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Recent advances in inhalable liposomes for treatment of pulmonary diseases: Concept to clinical stance
Journal Pre-proof Recent advances in inhalable liposomes for treatment of pulmonary diseases: Concept to clinical stance Piyush P. Mehta, Debjit Ghoshal, Atmaram P. Pawar, Shivajirao S. Kadam, Vividha S. Dhapte-Pawar PII:
S1773-2247(19)31859-3
DOI:
https://doi.org/10.1016/j.jddst.2020.101509
Reference:
JDDST 101509
To appear in:
Journal of Drug Delivery Science and Technology
Received Date: 1 December 2019 Revised Date:
28 December 2019
Accepted Date: 7 January 2020
Please cite this article as: P.P. Mehta, D. Ghoshal, A.P. Pawar, S.S. Kadam, V.S. Dhapte-Pawar, Recent advances in inhalable liposomes for treatment of pulmonary diseases: Concept to clinical stance, Journal of Drug Delivery Science and Technology (2020), doi: https://doi.org/10.1016/ j.jddst.2020.101509. This is a PDF file of an article that has undergone enhancements after acceptance, such as the addition of a cover page and metadata, and formatting for readability, but it is not yet the definitive version of record. This version will undergo additional copyediting, typesetting and review before it is published in its final form, but we are providing this version to give early visibility of the article. Please note that, during the production process, errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain. © 2020 Published by Elsevier B.V.
Recent advances in inhalable liposomes for treatment of pulmonary diseases: Concept to Clinical Stance
Piyush P. Mehta 1, Debjit Ghoshal 2, Atmaram P. Pawar 2, Shivajirao S. Kadam 3 and Vividha S Dhapte-Pawar 2*
1
Department of Quality Assurance, Poona College of Pharmacy, Bharati Vidyapeeth Deemed
University, Pune - 38, Maharashtra, India. 2
Department of Pharmaceutics, Poona College of Pharmacy, Bharati Vidyapeeth Deemed
University, Pune - 38, Maharashtra, India. 3
Bharati Vidyapeeth Bhavan, Bharati Vidyapeeth Deemed University, LBS Road, Pune- 30,
Maharashtra, India.
*Corresponding author Dr. Vividha S Dhapte-Pawar, Associate Professor, Department of Pharmaceutics, Bharati Vidyapeeth University, Poona College of Pharmacy, Erandwane, Kothrud, Pune 411038. Email: [email protected]; [email protected]
1
Recent advances in inhalable liposomes for treatment of pulmonary diseases: Concept
2
to Clinical Stance
3 4
Graphical Abstract
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6 7 8 9 10 11 12 13 14 15 16 17 1|Page
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Recent advances in inhalable liposomes for treatment of pulmonary diseases: Concept
19
to Clinical Stance
20 21
Abstract
22
Liposomes and liposomes-based drug delivery systems are promising carriers in the rapidly
23
evolving field of nanomedicine. Liposomes are receiving growing attention in the scientific
24
domain owing to their distinctive structural characteristics, physiological features and
25
biological properties. These versatile lipid vesicles represent unique platforms for drug
26
delivery, targeting, diagnostics, imaging as well as theranostics. The main objective of the
27
present article is to present insights into the pulmonary delivery of these versatile cargos for
28
nanomedicine. Inhalable liposomes for pulmonary delivery present unique advantages as they
29
are formulated from phospholipids similar to endogenous pulmonary surfactant. The article
30
begins by unfolding the important background information about liposomes and their key
31
advantages as nanomaterials. In the subsequent section, various liposomes and proliposome
32
formulations are summarized with a special emphasis on their physicochemical properties,
33
aerosolization performance and in-vivo aerodynamic behavior. Additionally, this article
34
includes a segment devoted to recent status on clinical trials of liposomal formulations for
35
treating pulmonary diseases. Moreover, the present article contains a section dedicated to the
36
market potential and scientific challenges related to inhalable liposomes. In summary, this
37
article is a comprehensive report of inhalable liposomes to appear in recent years.
38 39
Keywords
40
Liposomes, proliposome, nanomedicine, nanocarriers, dry powder inhalers, aerodynamic
41
behavior, drug delivery
42
2|Page
43
Abbreviations
44
Chronic obstructive pulmonary disease (COPD); world health organization (WHO);
45
pressurized metered-dose inhalers (pMDIs); soft mist inhalers (SMI); dry powder inhalers
46
(DPI); novel drug delivery systems (NDDS); small unilamellar vesicles (SUVs); large
47
unilamellar vesicles (LUVs); giant unilamellar vesicles (GUVs); multilamellar vesicles
48
(MLV); multi-vesicular vesicles (MVV); double emulsion templating (DET); microfluidic
49
hydrodynamic focusing (MHF); reticuloendothelial system (RES); polyethylene glycol
50
(PEG); enhanced permeability and retention (EPR); solid lipid nanoparticles (SLNs);
51
nanostructured lipid carriers (NLCs); nanoparticles embedded microparticles (NEMs);
52
encapsulation
53
bromide (MTT); half-maximal inhibitory concentration (IC50); folate receptor alpha
54
(SPCA1); hydrogenated soy phosphatidylcholine (HSPC); 1,2-Distearoyl-sn-glycero-3-
55
phosphoglycerol (DSPG); fine particle fraction (FPF); emitted dose (ED); mass median
56
aerodynamic diameter (MMAD); human lung adenocarcinoma (A549); maximum tolerated
57
dose (MTD); area under curve (AUC); mean residence time (MRT); 1,2-dioleoyl-sn-glycero-
58
3-[phospho-L-serine] (DOPS); twin stage impinger (TSI); maximum drug concentration
59
(Cmax); 4-aminophenyl-alpha-D-manno-pyranoside (PAM); next generation impactor (NGI);
60
3-methylcholanthrene (MCA); diethyl nitrosamine (DEN); tumor necrosis factor-α (TNF-α);
61
vascular endothelial growth factor (VEGF); malondialdehyde (MDA); caspase-3 and B-cell
62
lymphoma 2 protein (BCL-2); 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid (HEPES);
63
nuclear factor-κB (NF-κB); geometric standard deviation (GSD); idiopathic pulmonary
64
fibrosis (IPF); dipalmitoylphosphatidylcholine (DPPC); phosphatidylcholines (PC); the phase
65
transition temperature (Tm); normal human bronchial epithelial cells (NHBE); small airway
66
epithelial cells (SAEC); alveolar macrophages (AMs); enzyme linked immunosorbent assay
67
(ELISA); human lung cancer cell line cells (Calu-3); human alveolar basal epithelial cell
3|Page
efficiency
(EE);
3-(4,5-dimethylthiazole-2-yl)-2,5-diphenyltetrazolium
68
(NR8383); chemistry, manufacturing and control (CMC); forced vital capacity (FVC); forced
69
expiratory volume (FEV); Tiffeneau-Pinelli index (FEV1/FVC); maximal voluntary
70
ventilation (MVV).
71 72
1. Introduction
73
Even with the continuous efforts and progress made in pulmonary research, pulmonary
74
ailments still remain a major health issue across the globe. Pulmonary diseases are leading
75
causes of morbidity and mortality worldwide [1]. Some of the most prevalent pulmonary
76
ailments are asthma, chronic obstructive pulmonary disease (COPD), pulmonary tuberculosis,
77
lung cancer, pulmonary hypertension, chronic bronchitis, emphysema and occupational lung
78
diseases. COPD is not one single disease but an umbrella phrase commonly utilized to
79
express diverse chronic lung disorders that cause limitations in pulmonary airflow [2].
80
Almost 65 million individuals suffer from COPD and more than 3 million fatalities are
81
reported annually making it the third major cause of mortality worldwide. According to the
82
World Health Organization (WHO) statistics, approximately 235 million people suffer from
83
asthma and it is the most ordinary pulmonary disease among the paediatric population [3].
84
Moreover, the family, not just the individual, suffer the impact and crisis of pulmonary
85
diseases. In brief, pulmonary diseases are prevalent in every part of the world; this public
86
health problem is just not limited to 2nd and 3rd income countries but affects all countries
87
irrespective of their developmental levels. In view of the impact of pulmonary diseases on a
88
universal scale, it is apparent that these ailments have raised a significant public health
89
challenge [4].
90 91
Various pulmonary delivery systems such as pressurized metered-dose inhalers (pMDIs), soft
92
mist inhalers (SMI), dry powder inhalers (DPI) and nebulizers have been designed, developed
4|Page
93
and studied for the treatment of pulmonary diseases. Among them, DPIs are more preferred
94
dosage forms because of their better physicochemical stability and capability to deliver the
95
drug into deep lungs using the patient’s respiration [5,6]. Each type of delivery system has
96
distinct strengths and weaknesses considering the disease pathophysiology, severity and type
97
of prescribed formulation. Furthermore, it has been well reported that the therapeutic success
98
of inhalation therapy is highly dependent on particle size distribution, inhalation flow rate,
99
inhaler device resistance and dispersion capability [7]. Therefore, to maximize the clinical
100
outcome of inhalation therapy, a formulation with apt physicochemical properties is vital [8].
101 102
With the continuous growth of various scientific fields such as particle engineering, materials
103
science, biotechnology, molecular biology, green chemistry and biomaterials, rapid
104
advancement has been achieved in the domain of novel drug delivery systems (NDDS) [9].In
105
addition, recent progress in nanotechnology has opened avenues for augmenting the clinical
106
value of inhalation therapy for different pulmonary diseases [10]. The application of
107
nanotechnology to the design of NDDS such as polymeric systems (polymer-drug conjugates,
108
polymeric nanoparticles, polymeric micelles) [11], nanoparticles [12,13], nanoaggregates,
109
nanocomposites [14], dendrimers, quantum dots, fullerenes, novel lipid vesicles (liposomes,
110
pro-liposomes, solid lipid nanoparticles, nanostructured lipid carriers, cochleates and lipidic
111
spherulites), ethosomes, cubosomes, lipomers, polymeric microparticles and microspheres
112
[15] have illustrated numerous therapeutic advantages over traditional pulmonary
113
formulations [9,16]. The design and development of NDDS based DPIs have the potential to
114
surpass issues associated with the carrier as a critical formulation component. These novel
115
formulations help in improving physicochemical stability, tissue distribution, in-vivo drug
116
deposition and bioavailability [11,17]. To sum up, such novel formulations represent
117
promising alternatives to traditional inhaled formulations.
5|Page
118 119
Among various novel bioactive platforms, lipid vesicles i.e. liposomes have attracted the
120
focus of formulation scientists as they can be prepared from substances endogenous to the
121
pulmonary airways as components of the lung with surfactant and other unique
122
physicochemical and biopharmaceutical properties [18]. The application of liposomes in drug
123
delivery has been studied and investigated by numerous formulation scientists [19]. Several
124
active scientists and clinicians have studied the significance of liposomes systems for drug
125
delivery, drug targeting, tissue engineering, diagnostics, detection of genetic material and
126
imaging [20-22]. In brief, the superior therapeutic efficacy of liposomes has been evidenced
127
either in laboratory investigations or in clinical analysis, particularly in the therapy of
128
pulmonary diseases.
129 130
By knowing this continues development, in this article, we have analyzed inhalable liposomal
131
formulations for the treatment of numerous pulmonary ailments. In the opening section of
132
this article, we have discussed liposomes, generations of liposomes and the importance of
133
liposomes-based nanomaterials. In the next segment, various dry powdered liposomal
134
formulations are summarized with a special emphasis on their physicochemical properties,
135
aerosolization performance and in-vivo behavior. In the third part, the importance of
136
respirable pro-liposome formulations and their influence on pulmonary drug delivery are
137
thoroughly summarized. Additionally, this article includes a section devoted to recent efforts
138
taken in clinical trials of liposomal formulations for the treatment of pulmonary diseases.
139
Moreover, this article concisely explores the market potential of inhaled liposomal
140
formulations. Furthermore, the present article contains a segment dedicated to the practical
141
concerns, technical limitations and scientific challenges related to inhalable liposomes. This
6|Page
142
review can be of potential significance from the perspective of both academicians and
143
industrial scientists.
144 145
2. Methodology
146
We performed a broad literature search for liposomes and pro-liposomes for pulmonary drug
147
delivery. This literature search included scientific journals, books and book chapters from
148
various electronic sources such as ScienceDirect, PubMed, Google Scholar, Springer and
149
Web of Science. Present article investigates various key aspects associated to inhalable
150
liposomes and pro-liposomes with special importance on the prepration methods, in-vitro
151
aerodynamic performance and pulmokinetic parameters. References enlisted in present article
152
contains 130 articles, containing peered review articles, original research articles, clinical trial
153
reports and book chapters. All articles were examined, screened cautiously and then selected
154
for the review. ChemDraw software (Version 12.2; Perkin Elmer) was used to draw the
155
figures.
156 157
3. Liposomes
158
Liposomes perhaps are the most extensively explored and characterized lipid-based drug
159
delivery systems. Liposomes are the sub-micron spherical phospholipid vesicles consisting of
160
single or multiple concentric lipid bilayers enclosing aqueous interior [23]. Liposomes were
161
first discovered and explained in 1965 by Bangham and his co-workers. Since then, they are
162
extensively studied as a drug reservoir from the last five to six decades [24]. Accordingly,
163
liposomes have become one of the most explored drug reservoirs in diagnosis, treatment and
164
imaging of several diseases [25]. Generally, on basis of vesicular arrangement liposome can
165
be classified as unilamellar liposomes (single bilayer lipid membrane) i.e. small unilamellar
166
vesicles (SUVs; 20-100 nm); large unilamellar vesicles (LUVs; 100 nm -1 mm) and giant
7|Page
167
unilamellar vesicles (GUVs; > 1 mm) or multilamellar liposomes (numerous bilayer lipid
168
membranes)
169
MVV is also known as vesosomes (Fig. 1) [26]. Phospholipids and cholesterol are the main
170
two components of liposomal bilayers. Cholesterol plays a vital role in the physical stability
171
of liposome vesicles with its ability to increase the phospholipid phase transition temperature
172
(TM) [27,28]. Many techniques have been explored for the design and development of
173
liposomes (Table1). The most popular techniques that have been explored to develop
174
liposomes are the thin-film (Bangham method) and the ethanol injection method [29]. Each
175
type of technique has its own unique strengths and weaknesses considering the type of lipid
176
and the nature of drugs that can be utilized. On account of this, many formulation scientists
177
are now actively engaged in developing strategies to improve the physicochemical quality of
178
the liposomes. Microfluidic technology, membrane contactor, electrospray mechanism are a
179
helpful alternative for the fabrication of liposomes [29]. Moreover, new techniques derived
180
from microfluidic approaches for fabrication of liposome mainly include double emulsion
181
templating (DET) [30], electro formation and hydration [31], pulsed jetting [32], ice droplet
182
hydration [33], extrusion [34], transient membrane ejection [35], droplet emulsion transfer
183
[36] and microfluidic hydrodynamic focusing (MHF) [37]. Among these methods, templating
184
is a good option to fabricate liposomes of consistent size and high encapsulation efficiency
185
[29].
i.e. multilamellar vesicles (MLV) or multi-vesicular vesicles (MVV) [26].
186 187
Additionally, based on structural modifications, liposomes can be classified into the different
188
‘generations’. The first generation liposomes mainly consist of phospholipids and cholesterol
189
vesicles without any structural modifications. The mean particle size of first-generation
190
liposomes is in the range of 50 to 450 nm [48]. As a nanoscale drug reservoir, the liposomes
191
are biodegradable, biocompatible with low toxicity. As well, the liposomes are also
8|Page
192
convenient considering the scale-up and the quality control specifications [49]. Liposomes
193
are capable to hold both, lipophilic as well as hydrophilic molecules. Thus, a wide range of
194
therapeutic compounds can be easily incorporated into the liposomes [50]. Various synthetic
195
drugs, semi-synthetic compounds, herbs, herb extracts and phytoconstituents can be easily
196
incorporated into the liposomes. Ambisome® (Astellas Pharma), Amphotec® (Intermune), and
197
Abelcet® (Enzon) [amphotericin B liposomes] are few successful, commercial liposomal
198
products approved for the treatment of fungal infections across the globe [51]. Yet, these
199
first-generation liposomes displayed noticeable limitations such as leakage of drugs from
200
liposome vesicles, low loading capacity for hydrophilic compounds and rapid clearance of
201
vesicles by the reticuloendothelial system (RES) in the systemic circulation. Therefore, the
202
first generation liposomes were almost abandoned until the active drug loading and sterically
203
stable second-generation liposomes were discovered [52].
204 205
The second-generation liposomes are long-circulating, surface modified liposomes. Usually,
206
liposomes are coated with inert polymers i.e. polysaccharides, oligosaccharides,
207
glycoproteins and synthetic polymers [53]. Among these polymers, polyethylene glycol
208
(PEG) is most commonly explored to stabilize the liposomes. PEG-modified (PEG-coated)
209
liposomes are often denoted as sterically stabilized or ‘stealth liposomesʼ [54]. Several active
210
drug loading techniques have been explored to achieve higher drug encapsulation efficiency.
211
The active drug loading techniques are distant encapsulation methods that load the drug into
212
the liposomes by the differential chemical gradient across the membrane of the liposomes.
213
The chemical gradients mainly comprise the pH of the system, ammonium sulfate gradient or
214
calcium acetate gradient. As an effect, the liposomes exhibited favourable physicochemical
215
and biological properties such as reduced drug leakage, prolonged systemic circulation,
216
modified release pattern and altered bio-distribution. The prolonged systemic circulation is
9|Page
217
mainly ascribed to the steric hindrance effect presented by a hydrophilic polymer which
218
could avoid the surface-modified liposomes from being rapidly purged by the RES [53].
219
Consequently, a number of surface-modified liposomes were studied successfully and few of
220
them are under various phases of clinical trials. Few pegylated doxorubicin liposomes such as
221
Doxil® (PEG 2000 surface-modified liposome), Caelyx® (liposomes with surface-bound
222
methoxy-polyethylene glycol) and Myocet® (liposome-encapsulated doxorubicin citrate
223
complex) have been approved and marketed for the treatment of cancers [55]. Even though
224
great commercial achievements have been made, the second generation liposomes
225
demonstrated few downsides such as poor cancer cells selectivity and low cellular uptake
226
[56].
227 228
The targeting liposomes are known as the third generation liposomes wherein the targeting
229
ligand molecules or functional moieties are incorporated into liposomes to target a specific
230
site. The third generation liposomes may possess passive as well as active targeting
231
properties. Both of the first and second generation liposomes fit into the passive targeting
232
reservoirs with high lymphatic affinity attributed to the lipophilic casing of liposomes. These
233
first and second-generation liposomes displayed the enhanced accumulation at tumor site
234
owing to the enhanced permeability and retention (EPR) consequence [57]. Active targeting
235
liposomes can target cells and/or cellular organelles by a precise interaction between a
236
targeting ligand attached to the liposomes and a cell receptor [58]. Additionally, the diverse
237
class of chemical moieties such as carbohydrates, vitamins, monoclonal antibodies, peptides,
238
proteins and aptamers can be used as targeting ligands [59]. The enhanced therapeutic
239
efficacy of third-generation liposomes had been verified in the laboratory model particularly
240
in the treatment of cancer [56]. Nowadays, a novel dual-functional liposomes have appeared
241
as a promising drug delivery system. Dual-functional liposomes are typically formulated
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using a mixture of phospholipid and functional material by either chemical adaptation or
243
physicochemical inclusion of functional motifs against the membrane of liposomes. Current
244
investigations showed that the dual-functional liposomes are capable of improving the
245
therapeutic efficiency by modifying drug delivery as well as the mechanism of action [56].
246 247
For the pulmonary application, liposomes offer numerous benefits (Fig. 2) [60,61]. Superior
248
tolerability of liposomes in the pulmonary airways can be guaranteed, if lipids selected are
249
biodegradable resulting in non-toxic, endogenous degradation products [60,62]. Furthermore,
250
owing to their small (nm) size, they can be easily encapsulated into particles with suitable
251
aerosolization properties, which facilitates satisfactory deep lung deposition of a molecule.
252
Additionally, liposomes adhere to the mucosal surface of the airways for a longer time
253
compared to larger particles owing to the small size. Particle size, adhesion, accumulation
254
and retention in the pulmonary airways along with controlled release characteristics of
255
liposomes can lead to improved therapeutic outcomes leading to better patient compliance
256
[60,63]. This can assist an imperative function in the therapy for chronic diseases as many of
257
the existing inhalation formulations have to be applied at least twice a day because of the
258
comparatively less duration of the drug in the pulmonary airways [62,65]. With this
259
background, various liposomal DPIs are described in the following section with special
260
emphasis on their aerodynamic behavior and clinical outcomes.
261 262
4. Pulmonary drug delivery
263
It is fascinating to believe in pulmonary therapy as a modern strategy for drug delivery but
264
this practice has been well acknowledged in most of the ancient literature and it has a strong
265
history of more than 4000 years [66]. From the therapeutic perspective, pulmonary therapy
266
represents a few attractive benefits allied with the anatomical and physiological
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characteristics of the lung. Pulmonary therapy has a better potential of treating various
268
intrapulmonary and extrapulmonary diseases, such as asthma, COPD, cystic fibrosis,
269
pulmonary hypertension, pneumonia, cancer [67], diabetics and tuberculosis [68]. The
270
reduced invasiveness of pulmonary therapy may further improve clinical outcomes and
271
patient compliance with the treatment. Furthermore, owing to the scientific and technological
272
advances in formulations (e.g. Technosphere® particles, AerosphereTM delivery, iSPERSE™
273
platform and/or Capsugel® Zephyr™), particle replication in nonwetting templates (PRINT),
274
inkjet-printed aerogel particles and inhaler devices (e.g. Advair Diskus®, ProAir®
275
Digihaler™, Spiriva® Handihaler® and TwinCapsTM Inhaler), pulmonary therapy has become
276
more ‘patient-friendly’ and economically favourable [68,69].
277 278
Many different types of delivery systems such as nanoparticles (polymeric nanoparticles,
279
polymeric micelles), microparticles (microspheres), solid lipid nanoparticles (SLNs),
280
nanostructured lipid carriers (NLCs), polymer-drug conjugates, macromolecules (dendrimers)
281
and lipid vesicles (liposomes and proliposomes) have been designed and developed for
282
pulmonary delivery of several therapeutics. They fulfill many biopharmaceutical
283
requirements i.e. sufficient drug loading, shielding of the actives from degradation,
284
biocompatibility, biodegradability and stability during aerosolization. However, one major
285
limitation of these systems is that they can be readily exhaled from the lungs after pulmonary
286
delivery. Table 2 denotes various advantages and limitations of novel carrier-based
287
pulmonary drug delivery [70]. However, several formulation techniques, such as nano-
288
agglomeration processes, nanoparticle-rooted microparticles or nanoparticles embedded
289
microparticles (NEMs) can be utilized to form nanoparticles with appropriate aerodynamic
290
diameters for pulmonary delivery. Knowing these facts, lipid vesicles (i.e. liposomes and
291
proliposomes) for pulmonary delivery have been a popularized theme for the last two-three
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decades [8,11]. Accordingly, the present segment discusses and analyzes the available
293
literature on inhalable liposomes and proliposomes.
294 295
4.1 Liposomes based dry powder inhalers
296
Zhu et al (2019) developed folic-acid conjugated docetaxel (1:25) liposomes (LP-DTX-FA)
297
using phosphatidylcholine and cholesterol (6:1) by thin lipid film hydration method.
298
Furthermore, obtained LP-DTX-FA were spray-dried using mannitol (2 % w/v; carrier) and
299
leucine (anti-adherent) to obtain an inhalable dry powder. Spherical shaped, smooth surface
300
spray-dried LP-DTX-FA showed mean particle size, zeta potential and encapsulation
301
efficiency (EE) of 346.80 nm, -29.30 mV and 99.50 %, respectively. During the in-vitro
302
release study, LP-DTX-FA showed sustained release (>70 %) up to 50 h in phosphate buffer
303
saline (PBS; pH 7.4). In in-vitro cytotoxicity study using 3-(4,5-dimethylthiazole-2-yl)-2,5-
304
diphenyltetrazolium bromide (MTT) assay, LP-DTX-FA showed 3.3-fold superior half-
305
maximal inhibitory concentration (IC50) as compared to LP-DTX in high expression of folate
306
receptor alpha (SPCA1) cells. Additionally, during the in-vitro uptake study, LP-DTX-FA
307
showed significantly superior DTX uptake as compared to DTX alone due to conjugated
308
folic-acid. Moreover, during the in-vivo study after intratracheal administration in Sprague
309
Dawley rats, LP-DTX-FA showed 23.39-fold higher DTX deposition within the lung as
310
compared to intravenous LP-DTX-FA at the same dose of 1 mg/kg [71].
311 312
Gemcitabine intercalated liposomes (LP-GB) were formulated using a combination of lipids
313
i.e.
314
phosphoglycerol
315
emulsification solvent evaporation method. Gemcitabine liposomal powder (LP-GB-DPI)
316
was formulated using trehalose (1:2) as a cryoprotectant in the lyophilization technique.
hydrogenated
13 | P a g e
soy
(DSPG),
phosphatidylcholine cholesterol
and
(HSPC),
1,2-Distearoyl-sn-glycero-3-
mPEG2000-DSPE (5:2:2:0.8) by W/O
317
Spherical shape smooth surface LP-GB showed particle size and negative zeta potential of
318
339.49 nm and -53.1 mV, respectively while LP-GB-DPI demonstrated particle size of 8.79
319
µm with satisfactory flow properties. LP-GB-DPI showed satisfactory performance with fine
320
particle fraction (FPF), emitted dose (ED) and mass median aerodynamic diameter (MMAD)
321
of 56.12 %, 88.99 and 3.91 microns, respectively, measured at 60 L/min. LP-GB showed a
322
pH-dependent release profile over a period of 48 h. LP-GB displayed maximum (55.93 %)
323
drug release at pH 5.5 and pH 6.8 (41.45 %) whereas 7.4 (5.61 %) minimum drug release was
324
observed. Release kinetics showed that maximum gemcitabine release from liposome was
325
obtained at endosomal pH (5.5) or cancerous tissue pH (6.5). The reduced drug release at pH
326
7.4 confirmed the stability of liposomes within lung fluid. During the in-vitro toxicity study
327
using MTT assay on human lung adenocarcinoma (A549) cell 3.44-fold improvement in IC50
328
value was observed for LP-GB-DPI as compared to free drug. Moreover, maximum tolerated
329
dose (MTD) and edema index analysis showed retention of alveolar-capillary integrity and
330
negligible infiltration around bronchioles with LP-GB-DPI as compared to drug alone at the
331
same dose. Additionally, during the in-vivo study after intratracheal administration of
332
liposomes, a marked 8.31 and 5.69-fold improvement in area under curve (AUC) and mean
333
residence time (MRT) was observed as compared to drug alone at the same dose (2 mg/kg).
334
In brief, the formulated liposomes showed promising results for the management of lung
335
cancer [72].
336 337
In another study, a respirable lyophilized recombinant secretory leukocyte protease inhibitor
338
(RSLPI) loaded liposomes (RSLPI-DOPS-LP) was formulated using 1,2-dioleoyl-sn-glycero-
339
3-[phospho-L-serine] (DOPS)/ cholesterol (7:3) by thin-film hydration method. Additionally,
340
lyophilized RSLPI-DOPS-LP was micronized in the presence of mannitol (1:10). RSLPI-
341
DOPS-LP exhibited average particle size and EE of 153.60 nm and 74.10 %, respectively
14 | P a g e
342
while lyophilized RSLPI-DOPS-LP showed mean particle size and yield of 19.2 µm and
343
86.70 %, respectively. Micronization showed marked a 12-fold reduction in particle size of
344
lyophilized RSLPI-DOPS-LP. Micronized RSLPI-DOPS-LP powder retained the stability
345
and anti-neutrophil elastase activity of RSLPI without protein degradation as evident from
346
western blot analysis. During the in-vitro aerosolization study using low resistance
347
Spinhaler® device, micronized RSLPI-DOPS-LP powder showed FPF of 38.70 % by twin
348
stage impinger (TSI). In addition, micronized RSLPI-DOPS-LP powder showed FPF and
349
MMAD of 33.30 % and 2.44 µm, respectively, using cascade assembly at a flow rate of 90
350
L/min. This micronized RSLPI-DOPS-LP powder was more stable in terms of liposome size
351
for 5 months at room temperature [73]. In another study, Tang et al (2013) developed a pro-
352
drug oseltamivir phosphate (OP) loaded liposomes (1:10) using ovelecithin and cholesterol
353
by film dispersion method followed by spray drying (LP-OP-DPI). Smooth surface, spherical
354
OP liposomes displayed mean particle size, negative zeta potential and EE of 105.90 nm,-
355
13.65 mV and 60.43 %, respectively. Corrugated nonaggregating LP-OP-DPI showed
356
average particle size and yield of 3.5 µm and 55.37 % respectively, with acceptable flow
357
properties. During the in-vitro deposition study using TSI, LP-OP-DPI showed FPF of 35.4
358
%. In the in-vitro release study LP-OP-DPI displayed sustained release pattern for 20 hr in
359
PBS (pH 7.4) while OP solution showed more than 90 % drug release within the 2 h.
360
Moreover, during the in-vivo study after endotracheal administration, LP-OP-DPI showed a
361
marked 1.14 and 1.22-fold improvement in AUC and maximum drug concentration (Cmax), of
362
oseltamivir carboxylate (OSCA) respectively as compared to orally administered OP solution
363
at the same dose (12 mg/kg). The improved clinical outcome may be attributed to the
364
transformation of OP to OSCA in pulmonary airways by carboxylesterase enzyme [74].
365
15 | P a g e
366
Moxifloxacin loaded nanoliposomes (MFX-NL) were developed by a reversed-phase
367
evaporation method using L-α-phosphatidylcholine (type X-E) and cholesterol (7:3) lipids.
368
MFX-NL was decorated with targeting ligand 4-aminophenyl-alpha-D-manno-pyranoside
369
(PAM) to enhance the uptake of (NL) by alveolar macrophages. Additionally, MFX-NL was
370
embedded into microparticles using dextran as a carrier by spray drying. Corrugated surface
371
and dimple shaped MFX-NL microparticles showed particle size, zeta potential and EE of
372
277 nm, -12.31 mV and 66.25 %, respectively. During the in-vitro aerodynamic study,
373
microparticles showed higher respirable fraction (> 75 %) using the Aerolizer® device. In the
374
in-vitro release study, microparticles showed biphasic release pattern i.e. initial burst release
375
for first 2 h (~ 50 %) followed by controlled release (~ 100 %) up to 48 h in PBS (pH 7.4).
376
Moreover, during the in-vivo deposition study, after intrapulmonary administration (Penn-
377
century®) green fluorescence-labeled microparticles showed sufficiently higher MFX
378
deposition within the alveolar macrophages as compared to the upper respiratory tract.
379
Negatively charged MFX-NL provided higher anti-tubercular activity as compared to the
380
neutral NL whereas PAM decoration was capable enough to augment MFX alveolar delivery.
381
From a pharmacological viewpoint, higher alveolar deposition is important in the treatment
382
of pulmonary tuberculosis. Developed PAM decorated MFX-NL microparticles can be
383
suitable for the treatment of pulmonary tuberculosis and other pulmonary disorders [75].
384 385
Inhalable salbutamol sulphate (SS) loaded liposomal powder (SS-LP-DPI) was formulated
386
using soybean phosphatidylcholine by vesicular phospholipid gel technique followed by
387
lyophilization using lactose (1:5) as a cryoprotectant. In addition, the lyophilised powder was
388
lubricated with 0.5 % magnesium stearate and subjected to ball milling. SS-LP-DPI showed
389
80.71 and 44.35 % EE for SS before lyophilization and after rehydration, respectively.
390
Irregular shaped microparticles showed a marked 4-fold reduction in mean particle size after
16 | P a g e
391
ball milling. In an in-vitro drug release study using a dialysis bag technique SS-LP-DPI
392
displayed sustained release profile up to SS for 24 h in deionized water. During the in-vitro
393
aerodynamic study in TSI assembly using Spinhaler® device, ball-milled SS-LP-DPI
394
exhibited 2.53-fold superior FPF as compared to lyophilized powder. The present
395
investigation highlights the localized pulmonary delivery of liposomes containing anhydrous
396
SS [76]. Honmane et al (2018), prepared and optimized SS loaded liposomal DPI using
397
soybean lecithin and cholesterol (1:1) by thin-film hydration technique followed by spray
398
drying using lactose as a carrier (SS-LM-DPI). Optimized liposomes displayed mean particle
399
size, zeta potential and entrapment efficiency of 167.2 nm, 9.74 mV and 80.68 %,
400
respectively. While SS-LM-DPI showed a mean particle size of 6.35 µm suitable for
401
pulmonary drug delivery. During the in-vitro drug deposition study SS-LM-DPI
402
demonstrated marked 8.92-fold enhancement in FPF as compared to spray dried SS at a flow
403
rate of 60 L/min using the Rotahaler® inhaler device. Moreover, during the in-vitro
404
dissolution study, SS-LM-DPI displayed a controlled release profile for SS (~ 90 %) up to 14
405
hr in PBS (pH 7.4) [77]. Ye et al (2016), designed clarithromycin liposomes using a mixture
406
of soybean phosphatidylcholine: cholesterol: clarithromycin (4:1:2 w/w) by thin lipid film
407
hydration method. Inhalable powder formulation (CLA-LP-DPI) was formulated by
408
ultrasonic spray freeze drying (SFD) techniques using a combination of cryoprotectants i.e.
409
mannitol (15 % w/v) and sucrose (5 % w/v). Spherical shaped, porous CLA-LP-DPI showed
410
high drug recovery (85 %) with suitable particle size. During the in-vitro aerodynamic study
411
at a flow rate of 100 L/min, CLA-LP-DPI showed FPF and ED of 43.82 % and 53.78 %,
412
respectively. Additionally, during a three-month stability study at 25 °C and 60 % relative
413
humidity CLA-LP-DPI didn't show any major change in EE and mean particle size [78].
414
17 | P a g e
415
Curcumin (CUR) intercalated liposomal DPI (CUR-LP-DPI) were prepared using soybean
416
lecithin: cholesterol in the ratio of 5:1 by a film hydration method followed by freeze-drying
417
technique for the treatment of lung cancer. Irregular shaped microparticles showed a mean
418
particle size of 15.02 µm. During aerodynamic assessment CUR-LP-DPI showed FPF and
419
MMAD of 46.71 % and 5.81 µm using next-generation impactor (NGI). In in-vitro
420
cytotoxicity study using normal human bronchial epithelial cell (BEAS-2B) microparticles
421
showed marked 93-fold improvement in selection indices as compared to CUR alone at the
422
same dose (100 µmol/L) while microparticles showed satisfactory in-vitro anti-cancer cell
423
effect against A549 cells. Additionally, uptake of CUR from liposomes by A549 cells was
424
noticeably superior to that of CUR alone. For the in-vivo study, the primary lung cancer
425
model was developed using 3-methylcholanthrene (MCA) and diethylnitrosamine (DEN)
426
inducing agents. After pulmonary administration, CUR-LP-DPI displayed better anti-cancer
427
potential as compared to CUR alone and gemcitabine via regulating enzymatic markers such
428
as tumor necrosis factor-α (TNF-α), vascular endothelial growth factor (VEGF),
429
malondialdehyde (MDA), caspase-3 and B-cell lymphoma 2 protein (BCL-2) [79]. Dry
430
powders of liposomal encapsulated ciprofloxacin nanocrystals (CUR-LP-NC) were fabricated
431
using a freeze-thaw method followed by spray drying. CUR-LP-NC was formulated using
432
HSPC: cholesterol (7:3) and sucrose (2 % w/w) as a cryoprotectant. Corrugated dimple
433
shaped CUR-LP-NC microparticles showed particle size of 1.92 µm. In the in-vitro
434
dissolution study, CUR-LP-NC displayed a controlled release up to 12 h in HEPES (4-(2-
435
hydroxyethyl)-1-piperazineethanesulfonic acid) buffered saline (pH 7.4). During the in-vitro
436
aerodynamic study, CUR-LP-NC displayed higher FPF (69.70 %) as compared to CUR-LP-
437
NC formulated from 0.5 and 1 % w/w sucrose using Osmohaler device® at 105 L/min. The
438
developed formulation can be apt for a once-a-daily treatment schedule [80].
439
18 | P a g e
440
Li et al (2017) developed liposomal andrographolide powder (AG-LP-DPI) using soybean
441
lecithin and cholesterol (6:1) by the solvent injection method with subsequent by freeze-
442
drying using mannitol as a cryoprotectant. AG liposomes showed mean particle size and
443
negative zeta potential of 77.91 nm and -56.13 mV, respectively. While AG-LP-DPI showed
444
irregular and rough surface particles (7.44 µm) suitable for lung deposition. During the in-
445
vitro drug deposition study AG-LP-DPI showed a marked 2.75-fold improvement in FPF as
446
compared to conventional AG-DPI at 60 L/min flow rate using Twister® inhaler device.
447
Moreover, during in-vivo bacterial (S. aureus) study intratracheal administration of AG-LP-
448
DPI (1 mg) showed satisfactory anti-bacterial action at a 10-fold lower dose as compared to
449
AG (10 mg) alone. AG-LP-DPI considerably decreased pro-inflammatory cytokines such as
450
TNF-α, IL-1 and inhibited the phosphorylation of IκB-α in the nuclear factor-κB (NF-κB)
451
pathway (Fig. 3). In summary, phytoconstituent loaded inhaled liposomes are the potential
452
therapeutic agents for pulmonary therapy [81]. In another study, Viswanathan et al (2018),
453
fabricated inhalable liquorice acetone extract loaded liposomes (L-LP-DPI) using soybean
454
phosphatidylcholine by thin-film hydration method with further freeze-drying using trehalose
455
as cryoprotectant and carrier. Unilamellar spherical shaped liposomes showed a mean particle
456
size and EE of 210 nm and 75 %, respectively. During the multistage cascade analysis at 60
457
L/min flow rate using Lupihaler® device, L-LP-DPI showed the FPF, MMAD and geometric
458
standard deviation (GSD) of 54.68 %, 4.29 µm and 1.23 respectively. The in-vivo lung
459
deposition study using the nose-only apparatus in Swiss-albino mice showed more than 46 %
460
of drug deposited by L-LP-DPI in the lungs whereas only 16% of drug retained in the lungs
461
24 h of administration. Moreover, an in-vivo pharmacodynamic study in Mycobacterium
462
tuberculosis H37Rv infected Balb/c mice L-LP-DPI showed a satisfactory reduction in
463
bacterial count in lungs and spleen. In brief, liposomal containing liquorice extract found to
464
be a useful anti-tubercular medicine alone or as an adjunct to the existing standard drugs [82].
19 | P a g e
465 466
Chennakesavulu et al (2017) formulated colchicine (CL) and budesonide (BU) loaded
467
liposomal DPI using thin layer film hydration technique followed by lyophilization for
468
effective treatment of idiopathic pulmonary fibrosis (IPF). CL and BU liposomes were
469
prepared using a combination of various lipids i.e. DPPG: SPC: COL in a ration of 3:6:1 and
470
4:5:1, respectively. CL and BU liposomal DPI using mannitol as a carrier and glycine (10 %)
471
as an anti-adherent was developed. Spherical shaped, CL liposomes showed particle size, zeta
472
potential and drug entrapment of < 100 nm, -24.7 mV and 50.94 %, respectively while BU
473
liposomes showed particle size, zeta potential and drug entrapment of < 100 nm, -36.9 mV
474
and 74.22 %, respectively. During the in-vitro aerosolization study using Rotahaler® device,
475
CL and BU liposomal DPI showed FPF of 44.45and 48.62 %, respectively at a flow rate of
476
28.3 L/min. In-vitro diffusion study for both formulations showed sustained release up to 24
477
h and followed Higuchi’s diffusion-controlled kinetic model in PBS, pH 7.4 containing SLS
478
(1 % w/v). Additionally, during in-vivo studies, in bleomycin (2.5 U/kg) induced IPF rats, CL
479
and BU combined DPI showed 1.17 and 3.53-fold reduction in hydroxyproline content and
480
myeloperoxidase activity showing a positive effect against IPF. Moreover, six-month stability
481
studies at two different conditions i.e. long term (25 °C ± 2 °C, 60 % ± 5 % RH) and
482
refrigerated conditions (2-8 °C) confirmed the stability of lyophilized BU and CL liposomes.
483
Thus, liposomal based BU and CL DPI formulations can be potentially used for the treatment
484
of IPF [83]. Apart from these investigations, various, appealing case studies of inhalable
485
liposomal DPI are summarized in Table 3.
486 487
As summarized in Table 3, liposomes were thoroughly explored and studied as carriers for
488
the inhalation delivery of synthetic drugs [76,77], herbal extracts [82], phytoconstituents,
489
[79,81] vitamins, protein and peptides for the treatment of numerous pulmonary diseases and
20 | P a g e
490
pathological conditions. They were carefully explored for various pulmonary disorders
491
ranging from asthma, COPD to lung cancer. Several well-known and novel processing
492
techniques have been explored to deliver these as dry powder. In various scientific studies,
493
after judiciously selecting phospholipid composition and aerosolization technique, liposomes
494
retain their payload, mean size and do not aggregate after aerosolization. They reveal a
495
superior pulmonary deposition, satisfactory lung retention for a prolonged period after
496
inhalation and delivery of the payload inside the cells [10].
497 498
The drug distribution pattern within the pulmonary airways is highly depended on the size of
499
the aerosolized liposomes rather than vesicle size or type. Among the inhaled liposome
500
formulations, phosphatidylcholine was the most appropriate phospholipid to certify vesicle
501
stability and minimize lipid-drug interactions [91,92]. Normally, the addition of cholesterol in
502
liposomal vesicles enhances in-vivo stability with the decrease in drug release rate. The slow-
503
release pattern of liposomes extends the drug residence time in pulmonary airways and assists
504
the uptake by macrophages [92]. While the inclusion of phosphatidylglycerol assist the
505
spreading of liposomes at the alveolar interface and potentially improve the drug release [93].
506
Multilamellar liposomes are more suitable than unilamellar liposomes for sustained
507
pulmonary drug release. The fusion of lipid vesicles with the alveolar interface and/or lipid
508
exchange between lipid vesicles and pulmonary surfactants has a significant impact on the
509
drug release mechanism [93]. Moreover, liposomes appear to be more suitable for pulmonary
510
application if they are formulated from substances endogenous to the lung i.e. pulmonary
511
surfactant.
512
[dipalmitoylphosphatidylcholine (DPPC), phosphatidylglycerol, phosphatidylcholines (PC)]
513
and proteins where lipids account for >90 % of the surfactant by mass [94-96]. The fate of
514
liposomes deposited in the alveoli is mainly governed by clearance and re-utilization of lung
21 | P a g e
Pulmonary
surfactant
is
a
unique
mixture
of
lipids
515
surfactant. The rate and degree of pulmonary uptake of liposomes are a function of their
516
phospholipid
517
phospholipid showed superior pulmonary uptake [97,98] whereas, macrophage activity
518
played an important role in the clearance mechanism of liposomes [99]. The mean vesicle
519
size of liposome should be less than 400 nm as macrophage uptake of the formulation will
520
increase after 400 nm [100]. Furthermore, neutral or anionic and cationic liposomes showed
521
the different pharmacological outcome. No major adverse effects were found after the
522
delivery of neutral or anionic liposomes. But, cationic liposomes were found to be fatal to
523
human cells and can potentially initiate genetic abnormalities. Furthermore, side effects of
524
cationic liposomes considerably enhanced with an increase of positive charge of the carriers.
525
However, cationic liposomes usually form the almost neutral complexes with negatively
526
charged nucleic acids. Thus such structural modification of cationic carriers usually controls
527
adverse effects on the cells [101]. However, during storage liposomal formulation
528
experienced considerable loss of encapsulated drug and alteration in the physical integrity of
529
the liposomes [102]. To surpass the instability and delivery issues, liposomes can be dried
530
using spray drying, SFD or freeze-drying into proliposomes [63]. Proliposomes are free-
531
flowing particles that instantly form liposomes when in contact with aqueous media [92,103].
532
These delivery systems are thoroughly summarized in the following section.
composition.
In
general,
liposomes
containing
phosphatidylglycerol
533 534
4.2 Proliposomes
535
Proliposomes were initially explored by Payne and his co-workers in 1986 as dry
536
phospholipid formulations with better stability than liposomes [104]. The dry form of
537
proliposomes ease transportation and makes them a functional and efficient delivery system.
538
Proliposomes includes hydrophilic carriers that are layered with cholesterol and
539
phospholipid. They are superior for the entrapment/encapsulation of both hydrophilic as well
22 | P a g e
540
as lipophilic molecules. Later in 1991, the idea of proliposomes was extended to include
541
liquid phospholipid formulations that can produce liposomes upon addition of aqueous
542
medium. Proliposomes can be grouped into two types i.e. particulate-based proliposomes and
543
solvent-based proliposomes. The selection of an apt carrier is a key aspect for the formulation
544
of particulate-based proliposomes. Basically, the carrier is selected on account of its porosity
545
and capacity to hold phospholipids on its surface [105,106]. Whereas, solvent-based
546
proliposomes are fabricated using an organic solvent that dissolves lipids and all at once is
547
miscible with water. In this technique, high concentration of phospholipid in the organic
548
solvent is formulated and the resulting liposomes are collected by dispersion into the aqueous
549
phase. The company of phospholipid, ethanol and water initially produces a stacked bilayer
550
that transforms into liposomes by hydration. Ultimately, proliposomes can be transformed
551
into liposomes by the addition of aqueous phase above the phase transition temperature (Tm)
552
of the lipid followed by continuous shaking. In relation to traditional liposomal formulations,
553
proliposomes reveal more benefits in stability, solubility, drug entrapment and drug release.
554
In addition, production of proliposomes can be scaled up by routinely used techniques such
555
as spray drying, fluidized-bed coating and fluid-energy micronization (jet-milling). Various
556
inhalable proliposomes formulations have been fabricated and explored to augment the
557
aerosolization performance and physicochemical stability of various molecules [68,106].
558
Various studies on inhalable proliposomes formulations are described in the following
559
section.
560 561
Patil-Gadhe et al (2013), fabricated inhalable rifapentine loaded proliposomes for the
562
treatment of pulmonary TB using soy phosphatidylcholine (HSPC) and cholesterol by spray
563
drying and optimized using 32 experimental design. Developed proliposomes showed mean
564
particle size, entrapment efficacy and zeta potential of 578 nm, 72.08 % and 29.40 mV,
23 | P a g e
565
respectively. Smooth spherical, spray-dried rifapentine proliposomes showed satisfactory
566
flow properties. During the in-vitro aerosolization study using Rotahaler®, rifapentine loaded
567
proliposomes showed FPF and MMAD of 92.50 % and 2.62 microns, respectively at a flow
568
rate of 60 L/min. In in-vitro release study rifapentine proliposomes displayed controlled
569
release (~ 90 %) up to 24 h in PBS (pH 7.4). Moreover, during the in-vivo pulmonary
570
pharmacokinetic study after intratracheal administration rifapentine proliposomes displayed
571
13.99, 6.43, 6.40 and 2.35-fold improvement in AUC(0-∞), AUC(0-24), MRT and Cmax,
572
respectively as compared to drug alone at the same dose (250 µg) [107]. Rojanarat et al
573
(2011),
574
phosphatidylcholine and cholesterol (1:1) by spray drying technique with mannitol as an inert
575
carrier. Irregular shaped proliposomes demonstrated FPF and MMAD of 35 % and 2.99
576
microns, respectively, at a flow rate of 60 L/min. During the in-vitro toxicity study, in MTT
577
assay INH-proliposomes did not show any toxicity to pulmonary-associated cells i.e. growth
578
of normal human bronchial epithelial cells (NHBE) and small airway epithelial cells (SAEC).
579
In addition, INH-proliposomes did not activate the alveolar macrophages (AMs) to produce
580
inflammatory mediators i.e. interleukin-1β, tumor necrosis factor-α, and nitric oxide, at a
581
toxic level confirmed from enzyme-linked immunosorbent assay (ELISA) method. Most
582
significantly, INH-proliposomes showed superior anti-mycobacterial activity against M.
583
Bovis-infected AM as compared to INH alone [108].
formulated
inhalable
isoniazid
(INH)
proliposomes
using
soybean
584 585
Also, Rojanarat et al (2012), fabricated inhalable pyrazinamide (PAZ) proliposomes using
586
soybean phosphatidylcholine and cholesterol (1:1) by spray drying method with porous
587
mannitol as a carrier for releasing PAZ to AM infected with mycobacteria. Irregular shaped
588
pro-liposomes demonstrated FPF and MMAD of 29 % and 4.4 microns, respectively, at a
589
flow rate of 60 L/min. During in-vitro toxicity study, in MTT assay using A549, human lung
24 | P a g e
590
cancer cell line cells (Calu-3; as an upper airway cell) and human alveolar basal epithelial
591
cell (NR8383; as a lower airway cell), formulated pro-liposome did not show any toxicity up
592
to 250 µg/mL. Furthermore, formulated proliposome did not activate the AMs to produce
593
inflammatory mediators i.e. interleukin-1β, tumor necrosis factor-α, and nitric oxide, at a
594
toxic level confirmed from ELISA kits. Additionally, in-vivo studies in male Wistar rats after
595
intratracheal administration (4 mg/kg) proliposome did not show any liver or renal toxicity
596
[109]. The same research group also prepared fluoro-quinolone antibiotic (levofloxacin)
597
loaded proliposomes for the treatment of pulmonary TB. Irregular shaped proliposomes
598
showed FPF and ED of 38.10 % and 91.30 % respectively, at a flow rate of 60 L/min. During
599
the in-vitro toxicity study, in MTT assay, levofloxacin pro-liposome showed absences of
600
toxicity on Calu-3, NR8383 up to 5 µg/mL. Moreover, levofloxacin proliposomes could not
601
produce inflammatory mediators and hence, showed marked improvement in the minimum
602
inhibitory concentration value as compared to drug alone [110].
603 604
Proliposomes have illustrated better therapeutic outcomes in the treatment of pulmonary
605
diseases. As summarized in the above section, proliposomes are easily fabricated using a
606
combination of SPC or HSPC and cholesterol with mannitol, porous mannitol or lactose as an
607
inert
608
physicochemical, biological and aerosolization properties of various actives. In comparison to
609
all discussed proliposomes, the HSPC and lactose based proliposome have achieved a greater
610
improvement in FPF (>90%) for rifapentine. Therefore, the combination of a drug molecule
611
with carefully selected lipid moiety, carrier and method may be a perfect move to formulate
612
proliposomes with reasonable aerodynamic properties.
carrier
by
613 614
5. Clinical outcome
25 | P a g e
spray-drying
method.
Spray-dried
proliposomes
could
enhance
615
In the last few decades, formulation scientists have been dynamically exploring the field of
616
liposomes to strengthen the existing standard of therapeutics. This active growth and
617
development in liposomes call for sustained clinical translation and commercialization [111].
618
Even though the preclinical trials have revealed positive effects, there is uncertainty linked to
619
safety in human beings. Thus, it is necessary to conduct a clinical trial to realize the ways in
620
which liposomes interact with the pulmonary airways. Clinical trials are the key building
621
blocks of evidence-based medicine and therefore, nurturing the backbone of clinical practice
622
[112].
623 624
In the past few years, the regulatory bodies have approved some liposomes-based delivery
625
systems and more than 100 different liposomes are in various clinical phases. Thus, in the
626
present segment, we have discussed the liposome-based pulmonary formulation that has
627
reached various phases of clinical trials. Clinical trials related to ‘‘liposomes’’ have been
628
indexed
629
[https://clinicaltrials.gov/]) and European (EU Clinical Trials Register [https://www.clinical
630
trialsregister.eu/]). Clinical trials with inhaled liposomes have been executed for pulmonary
631
tuberculosis, cystic fibrosis, bronchiectasis, fungal infections, lung transplantation, cancer
632
(for osteosarcoma metastatic), analgesia (for post-operative pain), pulmonary moisturizer
633
along with the understanding of safety, efficacy and toxicity. Few remarkable clinical trials
634
are summarized in the following section while clinical efforts invested in inhaled liposomes
635
are listed in Table 4.
and
are
easily
accessible
in
the
US
FDA
(ClinicalTrials.gov
636 637
5.1 Liposomal amikacin (Arikayce®)
638
Insmed Incorporation (New Jersey, US) is a well-known global biopharmaceutical
639
organization with a focus on the design and development of various critical pharmaceutical
26 | P a g e
640
dosage forms. The lipid bilayer of Arikayce® consists of DPPC and cholesterol. The Insmed
641
studied inhaled liposomal amikacin(Arikayce®) to treat mycobacterium infections (phase 3;
642
NCT02344004) [113], cystic fibrosis (phase 3;NCT01316276) [114] and Pseudomonas
643
aeruginosa infection (phase 3; NCT01315678) [115]. The liposomal amikacin was expected
644
to modulate disease conditions. Similarly, Insmed also studied inhaled liposomal cisplatin for
645
the treatment of cancer i.e. osteosarcoma metastatic of lungs (phase 1; NCT00102531) [116].
646
All these studies are near completion and the Insmed is expecting positive outcomes.
647 648
5.2 Liposomal amphotericin B (Ambisome®)
649
Various medical institutes together with pharmaceutical industries investigated inhaled
650
liposomal amphotericin B (Ambisome®) for various pulmonary diseases. Ambisome® (~ 100
651
nm) is composed of amphotericin B, DSPG, hydrogenated soy phosphatidylcholine and
652
cholesterol in a 0.4:0.8:2:1 molar ratio. Interaction between amphotericin B and cholesterol
653
attributed to its sterol binding which is highly responsible for stabilization. Ambisome® was
654
studied for the treatment of allergic bronchopulmonary aspergillosis (phase 2;
655
NCT02273661) [117] and invasive pulmonary aspergillosis (phase 4; NCT00986713) [118].
656 657
5.3 Liposomal phospholipids (LipoAerosol©)
658
The weakening of airways surfaces fluid film (pulmonary surfactant) is one of the main
659
factors responsible for pulmonary disorders (e.g. asthma, COPD and pulmonary edema).
660
Thus, replenishment of pulmonary surfactants through a pharmacological treatment might be
661
an appropriate strategy in the treatment of pulmonary disorders. The lipid vesicles of
662
LipoAerosol® have composed of phospholipids i.e. phosphatidylcholine which is an integral
663
part of the natural pulmonary surfactant. LipoAerosol® provides moistening, warming and
664
cleaning of the upper and lower pulmonary airways as well as assists the natural moistening
27 | P a g e
665
of the film in airway diseases and irritations. Recently, Technische Universität München
666
(Germany)
667
complications (NCT02157129) [119] and hoping for positive results.
has
investigated
LipoAerosol® therapeutic
potential
for
tracheostomy
668 669
5.4 Aerosolized liposomal fentanyl (AeroLEF™)
670
AeroLEF™ (liposome loaded fentanyl) is a new aerosol that offers rapid, extended and
671
personalized analgesia for patients undergoing acute pain episodes. It is designed and
672
developed for the non-invasive route of administration. It can be used for the treatment of
673
moderate to severe pain. AeroLEF™ was assessed in a few clinical trials for controlling pain
674
and post-operative pain (NCT00791804) [120]. Moreover, YM BioSciences Inc. (Canadian
675
drug development company) has studied AeroLEF™ therapeutic potential with four different
676
inhaler devices for device characterization and qualification (NCT00794209) [121].
677 678
The clinical trial database of the US FDA reported no clinical trial on the inhaled liposomes
679
as dry powders. However, an extensive amount of time and effort has been devoted by active
680
scientists and clinicians over the years towards design, development and clinical assessment
681
of inhaled liposomes. All these active efforts signify a bridgehead for the clinical progress of
682
inhaled liposomes. Still, a number of human clinical trials of inhaled liposomes have yet to be
683
performed for a wide range of ailments and better pulmonary therapy.
684 685
6. Market overview
686
The excitement regarding liposomes and liposome-based products has accelerated gradually
687
over the past few years. Liposomes and liposome-based products have demonstrated a
688
beneficial impact on the pharmaceutical market. Inhaled liposomes have offered several key
689
advantages including local as well as systemic delivery, high drug loading capacity,
28 | P a g e
690
controlled release kinetics and good patient compliance. Furthermore, nanocarriers such as
691
liposomes or pro-liposomes also help to extend the patent life and consequently improves the
692
value of drug molecules. For example, the Doxil® (liposomal formulation of doxorubicin) has
693
shown a good impact on cancer management with great benefits for pharmaceutical
694
industries. While, DaunoXome® (liposomal daunorubicin, Galen, Craigavon, UK), Onivyde®
695
(liposomal irinotecan injection, Merrimack Pharmaceuticals, US), DepoCyt® (liposomal
696
cytarabine, Pacira Pharmaceuticals, US),Marqibo® (liposomal vincristine sulfate, Talon
697
Therapeutics, US), AmBisome® (liposomal Amphotericin B, NeXstar Pharmaceuticals, US),
698
Vyxeos® (daunorubicin and cytarabine encapsulated liposomes, Jazz Pharmaceutics, Ireland)
699
and Visudyne® (benzoporphyrin derivative liposomes, QLT Phototherapeutics, Canada) are
700
the known and FDA approved liposomal products [122]. Still, no inhaled liposomal as dry
701
powders were available in the market. However, bench-to-bedside translation of nanocarriers
702
and adopting the same into the mainstream level is often a critical task. There are several
703
chemistry, manufacturing and control (CMC) challenge along with regulatory issues that
704
need to be defeated before it can move on to extensive clinical applications and community
705
acceptance. Thus, by knowing this situation, in the upcoming segment of article authors
706
provide a view on the existing challenges and future directions for liposomes; especially for
707
pulmonary applications.
708 709
7. Discussion
710
Currently, nanocarriers such as liposomes as well as proliposomes are receiving growing
711
interest among formulation scientists for better pulmonary therapy. So far, several
712
formulation scientists and experts have invested their time and efforts in designing,
713
developing and analyzing respirable liposomes and proliposomes for better clinical outcomes.
714
However, investigation in this stage is still in the primitive phase. Many published articles
29 | P a g e
715
have evaluated and discussed particle size, surface charge, in-vitro aerosolization behavior
716
and pulmonary pharmacokinetics. Yet, various other challenges must be addressed when
717
formulating liposomes and proliposomes for inhalation. Several key benefits and challenges
718
of inhaled liposomes are listed in Fig. 4 [123].
719 720
Needless to say, but achieving adequate deep lung deposition and targeting the drug to
721
specific pulmonary airways, is still very tough [68]. Ahead of the complexity of respiratory
722
diseases and lung morphology, it should be kept in mind that disease severity, patient’s age,
723
breathing pattern, device configuration (single dose, multi-dose device) and design features
724
(capsule-based, pre-filed or disposable device) conclude the real fate of aerosol and the final
725
therapeutic outcome of inhaled therapy [68,124,125]. Normally, the successful aerosol
726
delivery depends on four mutually dependent factors: the formulation, the inhaler device
727
configuration, the metering system and lastly, the patient’s training/understanding [124,126].
728
In the above-reviewed articles, most scientists have addressed liposomes and proliposomes
729
manufacturing techniques, drug loading methods, drug targeting strategies and pulmonary
730
pharmacokinetic problems; however, they have merely discussed issues pertain to inhaler
731
devices. Beyond this, there is also a fascinating story to be told about inhaler devices.
732
Furthermore, the device dose metering system has great importance as the dose needed for
733
the effective therapeutic outcome of antimicrobial is too large (e.g. 500 µg) to be inhaled in a
734
single actuation [68]. In such cases, there is a strong need for disposable inhaler device for
735
safe, effective delivery and to avoid bacterial resistance. Basically, to attain satisfactory drug
736
deposition, the development of formulation and inhaler devices should be considered as a
737
whole medicinal product. As per our perceptive strong collaboration between formulation
738
experts and device, engineers is desired to accomplish complex devices/formulation
739
interfaces.
30 | P a g e
740 741
Similarly, to explore the complete therapeutic potential of these multifunctional nanocarriers,
742
exhaustive consideration is required for pulmokinetics, lung clearance rate and other
743
toxicological issues. These issues can be tackled by improving current animal models that can
744
simulate the pathology of the human pulmonary airways [126]. Further, the new imaging
745
tools and techniques can provide superior extrapolative pre-clinical models to understand
746
complex and difficult to treat pulmonary ailments [128-130]. In addition, regarding the
747
situation of respirable liposomes in human clinical trials, there are presently only a few
748
formulations. Hence, up to date data and knowledge of clinical measurements such as forced
749
vital capacity (FVC) and forced expiratory volume (FEV), Tiffeneau-Pinelli index
750
(FEV1/FVC) and maximal voluntary ventilation (MVV) will be crucial in understanding their
751
capability for treating respiratory ailments [68,11]. US FDA has recently published a
752
guidance document i.e. ʻLiposome Drug Productsʼ for a better understanding of the liposomal
753
drug. Likewise, to utilize the full potential of these multifunctional nanocarriers absolute
754
consideration must be paid to upcoming regulatory and CMC guidance. Simply, combined
755
research effort will be highly appreciated to surpass gaps in our understanding with the
756
intention that we can assemble and use all accessible tools and techniques across the pre-
757
clinical-translational-clinical-axis to deliver the best pulmonary dosage to the patient in the
758
most efficient manner.
759 760
8. Conclusion
761
With the growth of nanotechnology and other allied fields, a number of versatile liposomes
762
are designed, developed and investigated as drug delivery carriers. In the vicinity of
763
pulmonary drug delivery, many bioactive molecules have been effectively encapsulated into
764
different liposomal carriers. Additionally, numerous novel strategies have been adopted to
31 | P a g e
765
modify the liposomal drug release pattern, in-vivo drug targeting and biodistribution within
766
the pulmonary airways. The research carried until now proved a great potential for the use of
767
liposomes in the treatment of various intrapulmonary and extrapulmonary diseases. However,
768
it is apparent from the present article that there is a clear need to continue the efforts to
769
design, develop and systematically evaluate the inhaled liposomal formulations. The present
770
situation of inhaled liposomes highly demands formulation scientists to judge long term and
771
structured strategy to pave a way for successful clinical translation, regulatory clearance with
772
FDA approval.
773 774
9. Acknowledgments
775
The author is thankful to Bharati Deemed Vidyapeeth University, Poona College of
776
Pharmacy, Pune-38, India for support and institutional facilities.
777 778
10. Funding Information
779
No financial support and writing assistance was utilized in the production of this manuscript.
780 781
11. Conflicts of Interest
782
Author declare no conflict of interest.
783 784
12. References
785
1. Salvi S, Kumar GA, Dhaliwal RS, Paulson K, Agrawal A, Koul PA et al. The burden of
786
chronic respiratory diseases and their heterogeneity across the states of India: the Global
787
Burden of Disease Study 1990-2016. Lancet Glob Health. 2018;6(12):e1363-e1374.
788
32 | P a g e
789
2. World Health Organization, WHO, Chronic Respiratory Diseases (CRDs): Fact Sheets,
790
(2018) http://www.who.int/respiratory/en/.
791 792
3. Celli BR, Agusti A. COPD: time to improve its taxonomy?. ERJ Open Res. 2018;4(1). pii:
793
00132-2017.
794 795
4. Forum of International Respiratory Societies. The Global Impact of Respiratory Disease-
796
Second Edition. Sheffield, European Respiratory Society, 2017.
797 798
5. Mehta PP. Multi-dose dry powder inhaler: advance technology for drug delivery to
799
airways. Indian Drug, 2019;56 (11):59-62.
800 801
6. Mehta P, Bothiraja C, Kadam S, Pawar A. Effect of USP induction ports, glass sampling
802
apparatus, and inhaler device resistance on aerodynamic patterns of fluticasone propionate-
803
loaded engineered mannitol microparticles. AAPS PharmSciTech. 2019;20(5):197.
804 805
7. Behara SR, Larson I, Kippax P, Stewart P, Morton DA. Insight into pressure drop
806
dependent efficiencies of dry powder inhalers. Eur J Pharm Sci. 2012;46(3):142-148.
807 808
8. Mehta PP. Imagine the superiority of dry powder inhalers from carrier engineering. J Drug
809
Deliv. 2018;2018:5635010.
810 811
9. Mehta PP. Dry powder inhalers: a focus on advancements in novel drug delivery systems. J
812
Drug Deliv. 2016;2016:8290963.
33 | P a g e
813
10. Kuzmov A, Minko T. Nanotechnology approaches for inhalation treatment of lung
814
diseases. J Control Release. 2015;219:500-518.
815 816
11. Mehta P, Kadam S, Pawar A, Bothiraja C. Dendrimers for pulmonary delivery: current
817
perspectives and future challenges. New J. Chem. 2019;43(22):8396-8409.
818 819
12. Aguilera G, Berry C, West R, Gonzalez-Monterrubio E, Angulo-Molina A, Arias-
820
Carrión O et al. Carboxymethyl cellulose coated magnetic nanoparticles transport across a
821
human lung microvascular endothelial cell model of the blood-brain barrier. Nanoscale Adv.
822
2019;1:671-685.
823 824
13. Wongpinyochit T, Totten J, Johnstona B, Seib F. Microfluidic-assisted silk nanoparticle
825
tuning. Nanoscale Adv. 2019;1:873-883.
826 827
14. Abdelaziz HM, Gaber M, Abd-Elwakil MM, Mabrouk MT, Elgohary MM, Kamel NM, et
828
al. Inhalable particulate drug delivery systems for lung cancer therapy: nanoparticles,
829
microparticles, nanocomposites and nanoaggregates. J Control Release. 2018;269:374-392.
830 831
15. Muralidharan P, Malapit M, Mallory E, Hayes Jr D, Mansour HM. Inhalable
832
nanoparticulate powders for respiratory delivery. Nanomedicine. 2015;11(5):1189-1199.
833 834
16. Andrade F, Rafael D, Videira M, Ferreira D, Sosnik A, Sarmento B. Nanotechnology and
835
pulmonary delivery to overcome resistance in infectious diseases. Adv Drug Deliv
836
Rev. 2013;65(13-14):1816-1827.
837
34 | P a g e
838
17. Mehta P, Bothiraja C, Mahadik K, Kadam S, Pawar A. Phytoconstituent based dry
839
powder inhalers as biomedicine for the management of pulmonary diseases. Biomed
840
Pharmacother. 2018;108:828-837.
841 842
18. van Rijt SH, Bein T, Meiners S. Medical nanoparticles for next generation drug delivery
843
to the lungs. Eur Respir J. 2014;44(3):765-774.
844 845
19. Cipolla D, Gonda I, Chan HK. Liposomal formulations for inhalation. Ther
846
Deliv. 2013;4(8):1047-1072.
847 848
20. Jumeaux C, Kim E, Howes P, Kim H, Chandrawati R, Stevens M. Detection of
849
microRNA biomarkers via inhibition of DNA-mediated liposome fusion. Nanoscale Adv.
850
2019;1:532-536.
851 852
21. Willis L, Hayes D, Mansour HM. Therapeutic liposomal dry powder inhalation aerosols
853
for targeted lung delivery. Lung. 2012;190(3):251-262.
854 855
22. Khatib I, Khanal D, Ruan J, Cipolla D, Dayton F, Blanchard JD et al. Ciprofloxacin
856
nanocrystals liposomal powders for controlled drug release via inhalation. Int J
857
Pharm. 2019;566:641-651.
858 859
23. Laverman P, Boerman OC, Oyen WJ, Dams ET, Storm G, Corstens FH. Liposomes for
860
scintigraphic detection of infection and inflammation. Adv. Drug Deliv. Rev. 1999;37:225-
861
235.
862
35 | P a g e
863
24. Bangham AD, Standish MM, Watkins JC. Diffusion of univalent ions across the lamellae
864
of swollen phospholipids. J. Mol. Biol. 1965;13:238-252.
865 866
25. van der Meel R, Fens MH, Vader P, van Solinge WW, Eniola-Adefeso O, Schiffelers
867
RM. Extracellular vesicles as drug delivery systems: lessons from the liposome field. J.
868
Control. Release 2014;195:72-85.
869 870
26. Rideau E, Dimova R , Schwille P , Wurm FR, Landfester K. Liposomes and
871
polymersomes: a comparative review towards cell mimicking. Chem. Soc. Rev.
872
2018;47:8572-8610.
873 874
27. Chorilli M, Calixto G, Rimério TC, Scarpa MV. Caffeine encapsulated in small
875
unilamellar liposomes: characerization and in vitro release profile. J. Dispers. Sci. Technol.
876
2013;34:1465-1470.
877 878
28. Koynova R, Caffrey M. Phases and phase transitions of the phosphatidylcholines.
879
Biochim. Biophys. Acta. 1998;1376:91-145.
880 881
29. Van Tran V, Moon JY, Lee YC. Liposomes for delivery of antioxidants in
882
cosmeceuticals: Challenges and development strategies. J Control Release. 2019;300:114-
883
140.
884 885
30. Chu LY, Utada AS, Shah RK, Kim JW, Weitz DA. Controllable monodisperse multiple
886
emulsions. Angew. Chem. Int. Ed. 2007;46:8970-8974.
887
36 | P a g e
888
31. Aimon S, Manzi J, Schmidt D, Poveda Larrosa JA, Bassereau P, Toombes GE.
889
Functional reconstitution of a voltage-gated potassium channel in giant unilamellar vesicles.
890
PLoS One. 2011;6(10):e25529.
891 892
32. Stachowiak JC, Richmond DL, Li TH, Liu AP, Parekh SH, Fletcher DA. Unilamellar
893
vesicle formation and encapsulation by microfluidic jetting. Proc Natl Acad Sci U S
894
A. 2008;105(12):4697-4702.
895 896
33. Sugiura S, Kuroiwa T, Kagota T, Nakajima M, Sato S, Mukataka S et al. Novel method
897
for obtaining homogeneous giant vesicles from a monodisperse water-in-oil emulsion
898
prepared with a microfluidic device. Langmuir. 2008;24:4581-4588.
899 900
34. Mayer LD, Hope MJ, Cullis PR. Vesicles of variable sizes produced by a rapid extrusion
901
procedure. Biochim Biophys Acta. 1986;858(1):161-168.
902 903
35. Pautot S, Frisken BJ, Weitz DA. Engineering asymmetric vesicles. Proc Natl Acad Sci U
904
S A. 2003;100(19):10718-10721.
905 906
36. Ota S, Yoshizawa S, Takeuchi S. Microfluidic formation of monodisperse, cell‐sized, and
907
unilamellar vesicles. Angew Chem Int Ed Engl. 2009;48(35):6533-6537.
908 909
37. Mazzitelli S, Capretto L, Quinci F, Piva R, Nastruzzi C. Preparation of cell-encapsulation
910
devices in confined microenvironment. Adv Drug Deliv Rev. 2013;65(11-12):1533-1555.
911
37 | P a g e
912
38. Bangham AD, De Gier, Greville G. Osmotic properties and water permeability of
913
phospholipid liquid crystals. Chem. Phys. Lipids. 1967;1:225-246.
914 915
39. Grimaldi N, Andrade F, Segovia N, Ferrer-Tasies L, Sala S, Veciana J, et al. Lipid-based
916
nanovesicles for nanomedicine. ChemSoc Rev. 2016;45(23):6520-6545.
917 918
40. Grijalvo S, Mayr J, Eritja R, Díaz DD. Biodegradable liposome-encapsulated hydrogels
919
for biomedical applications: a marriage of convenience. Biomater Sci. 2016;4(4):555-574.
920 921
41. Szoka J, Papahadjopoulos D. Procedure for preparation of liposomes with large internal
922
aqueous space and high capture by reverse-phase evaporation. Proc. Natl. Acad. Sci. U. S. A.
923
1978;75:4194-4198.
924 925
42. Deamer DW. Preparation and properties of ether injection liposomes. Ann. N. Y. Acad.
926
Sci. 1978;308:250-258.
927 928
43. Jahn A, Vreeland W, Gaitan M, Locascio L. Controlled vesicle self-assembly in
929
microfluidic channels with hydrodynamic focusing. J. Am. Chem. Soc. 2004;126
930
:2674-2675.
931 932
44. van Swaay D, deMello AD. Microfluidic methods for forming liposomes. Lab
933
Chip. 2013;13(5):752-767.
934 935
45. Jaafar-Maalej C, Charcosset C, Fessi H. A new method for liposome preparation using a
936
membrane contactor. J. Liposome Res. 2011;21:213-220.
38 | P a g e
937 938
46. Duong AD, Collier MA, Bachelder EM, Wyslouzil BE, Ainslie KM. One Step
939
Encapsulation of Small Molecule Drugs in Liposomes via Electrospray-Remote Loading.
940
Mol Pharm. 2016;13(1):92-99.
941 942
47. Collier MA, Bachelder EM, Ainslie KM. Electrosprayed Myocet-like Liposomes: An
943
Alternative to Traditional Liposome Production. Pharm Res. 2017;34(2):419-426.
944 945
48. Etheridge ML, Campbell SA, Erdman AG, Haynes CL, Wolf SM, McCullough J. The big
946
picture on nanomedicine: the state of investigational and approved nanomedicine products.
947
Nanomedicine. 2013;9(1):1-14.
948 949
49. Simone EA, Dziubla TD, Muzykantov VR. Polymeric carriers: role of geometry in drug
950
delivery. Expert Opin Drug Deliv. 2008;5(12):1283-1300.
951 952
50. Coune A. Liposomes as drug delivery system in treatment of infectious diseases, potential
953
applications and clinical experience. Infection. 1988;16:141-147.
954 955
51. Plotnick AN. Lipid-based formulations of amphotericin B. J. Am. Vet. Med. Assoc.
956
2000;216:838-841.
957 958
52. de Pauw BE. Fungal infections: diagnostic problems and choice of therapy. Leuk. Suppl.
959
2012;1:S22-23.
960
39 | P a g e
961
53. Woodle MC. Sterically stabilized liposome therapeutics. Adv. Drug Deliver. Rev.
962
1995;16:249-265.
963 964
54. Papahadjopoulos D. Stealth liposomes: from steric stabilization to targeting. CRC Press,
965
Boca Raton FL USA; 1995.
966 967
55. Drummond DC, Meyer O, Hong K, Kirpotin DB, Papahadjopoulos D. Optimizing
968
liposomes for delivery of chemotherapeutic agents to solid tumors. Pharmacol Rev.
969
1999;51(4):691-743.
970 971
56. Mu LM, Ju RJ, Liu R, Bu YZ, Zhang JY, Li XQ, et al. Dual-functional drug liposomes in
972
treatment of resistant cancers. Adv Drug Deliv Rev. 2017;115:46-56.
973 974
57. Nogueira E, Gomes AC, Preto A, Cavaco-Paulo A. Design of liposomal formulations for
975
cell targeting. Colloids Surf B Biointerfaces. 20151;136:514-526.
976 977
58. Marqués-Gallego P, de Kroon AI. Ligation strategies for targeting liposomal
978
nanocarriers. Biomed Res Int. 2014;2014:129458.
979 980
59. Morshed M, Chowdhury E. Gene delivery and clinical applications, Reference module in
981
biomedical sciences, Encyclopedia of Biomedical Engineering. 2019:345-351.
982 983
60. Weber S, Zimmer A, Pardeike J. Solid Lipid Nanoparticles (SLN) and Nanostructured
984
Lipid Carriers (NLC) for pulmonary application: a review of the state of the art. Eur J Pharm
985
Biopharm. 2014;86(1):7-22.
40 | P a g e
986 987
61. Gaspar M, Bakowsky U, Ehrhardt C. Inhaled liposomes-current strategies and future
988
challenges. J. Biomed. Nanotechnol. 2008;4:1-13.
989 990
62. G. Pilcer, K. Amighi, Formulation strategy and use of excipients in pulmonary drug
991
delivery. Int. J. Pharm. 2010;392:1-19.
992 993
63. Misra A, Jinturkar K, Patel D, Lalani J, Chougule M. Recent advances in liposomal dry
994
powder formulations: preparation and evaluation. Expert Opin Drug Deliv. 2009;6(1):71-89.
995 996
65. Willis L, Hayes D, Mansour H. Therapeutic liposomal dry powder inhalation aerosols for
997
targeted lung delivery. Lung. 2012;190:251-262.
998 999
66. Sanders M. Inhalation therapy: an historical review. Prim Care Respir J. 2007;16:71-81.
1000 1001
67. Skupin Mrugalska P. Liposome-based drug delivery for lung cancer, in Kesharwani P
1002
(Eds.), Nanotechnology-based targeted drug delivery systems for lung cancer, Academic
1003
Press, 2019;123-160.
1004 1005
68. Mehta P, Bothiraja C, Kadam S, Pawar A. Potential of dry powder inhalers for
1006
tuberculosis therapy: facts, fidelity and future. Artif Cells Nanomed Biotechnol.
1007
2018;46(sup3):S791-S806.
1008 1009
69. Zhou Q, Tang P, Leung S, et al. Emerging inhalation aerosol devices and strategies:
1010
where are we headed? Adv Drug Deliv Rev. 2014;75:3-17.
41 | P a g e
1011 1012
70. Tagami T, Ozeki T. Recent Trends in Clinical Trials Related to Carrier-Based Drugs. J
1013
Pharm Sci. 2017;106(9):2219-2226.
1014 1015
71. Zhu X, Kong Y, Liu Q, Lu Y, Xing H, Lu X, et al. Inhalable dry powder prepared from
1016
folic acid-conjugated docetaxel liposomes alters pharmacodynamic and pharmacokinetic
1017
properties relevant to lung cancer chemotherapy. Pulm Pharmacol Ther. 2019;55:50-61.
1018 1019
72. Gandhi M, Pandya T, Gandhi R, Patel S, Mashru R, Misra A, Tandel H. Inhalable
1020
liposomal dry powder of gemcitabine-HCl: Formulation, in vitro characterization and in vivo
1021
studies. Int J Pharm. 2015;496(2):886-895.
1022 1023
73. Gibbons A, McElvaney NG, Cryan SA. A dry powder formulation of liposome-
1024
encapsulated recombinant secretory leukocyte protease inhibitor (rSLPI) for inhalation:
1025
preparation and characterisation. AAPS PharmSciTech. 2010;11(3):1411-1421.
1026 1027
74. Tang Y, Zhang H, Lu X, Jiang L, Xi X, Liu J, Zhu J. Development and evaluation of a
1028
dry powder formulation of liposome-encapsulated oseltamivir phosphate for inhalation. Drug
1029
Deliv. 2015;22(5):608-618.
1030 1031
75. Hamed A, Osman R, Al-Jamal KT, Holayel SM, Geneidi AS. Enhanced antitubercular
1032
activity, alveolar deposition and macrophages uptake of mannosylated stable nanoliposomes.
1033
J Drug Deliv Sci Technol. 2019;51:513-523.
1034
42 | P a g e
1035
76. Huang WH, Yang ZJ, Wu H, Wong YF, Zhao ZZ, Liu L. Development of liposomal
1036
salbutamol sulfate dry powder inhaler formulation. Biol Pharm Bull. 2010;33(3):512-517.
1037 1038
77. Honmane S, Hajare A, More H, Osmani RAM, Salunkhe S. Lung delivery of
1039
nanoliposomal salbutamol sulfate dry powder inhalation for facilitated asthma therapy. J
1040
Liposome Res. 2019;29(4):332-342.
1041 1042
78. Ye T, Yu J, Luo Q, Wang S, Chan H. Inhalable clarithromycin liposomal dry powders
1043
using ultrasonic spray freeze drying. Powder Technol. 2017;305:63-70.
1044 1045
79. Zhang T, Chen Y, Ge Y, Hu Y, Li M, Jin Y. Inhalation treatment of primary lung cancer
1046
using liposomal curcumin dry powder inhalers. Acta Pharm Sin B. 2018;8(3):440-448.
1047 1048
80. Khatib I, Khanal D, Ruan J, Cipolla D, Dayton F, Blanchard J, Chan H. Ciprofloxacin
1049
nanocrystals liposomal powders for controlled drug release via inhalation. Int J Pharm.
1050
2019;566:641-651.
1051 1052
81. Li M, Zhang T, Zhu L, Wang R, Jin Y. Liposomal andrographolide dry powder inhalers
1053
for treatment of bacterial pneumonia via anti-inflammatory pathway. Int J Pharm.
1054
2017;528(1-2):163-171.
1055 1056
82. Viswanathan V, Pharande R, Bannalikar A, Gupta P, Gupta U, Mukne A. Inhalable
1057
liposomes of Glycyrrhiza glabra extract for use in tuberculosis: formulation, in
1058
characterization, in vivo lung deposition, and in vivo pharmacodynamic studies. Drug Dev
1059
Ind Pharm. 2019;45(1):11-20.
43 | P a g e
vitro
1060 1061
83. Chennakesavulu S, Mishra A, Sudheer A, Sowmya C, Reddy C, Bhargav E. Pulmonary
1062
delivery of liposomal dry powderinhaler formulation for effective treatment ofidiopathic
1063
pulmonary fibrosis. Asian J. Pharm. Sci. 2018;13:91-100.
1064 1065
84. Ourique AF, Chaves Pdos S, Souto GD, Pohlmann AR, Guterres SS, Beck RC.
1066
Redispersible
1067
development, in vitro characterization and antioxidant activity. Eur J Pharm Sci.
1068
2014;65:174-182.
liposomal-N-acetylcysteine
powder
for
pulmonary
administration:
1069 1070
85. Khatib I, Khanal D, Ruan J, Cipolla D, Dayton F, Blanchard JD, Chan HK. Ciprofloxacin
1071
nanocrystals liposomal powders for controlled drug release via inhalation. Int J Pharm.
1072
2019;566:641-651.
1073 1074
86. Daman Z, Gilani K, Najafabadi A, Eftekhariand H, Barghi M. Formulation of inhalable
1075
lipid-based salbutamol sulfatemicroparticles by spray drying technique. Daru. 2014; 22(1):
1076
50.
1077 1078
87. Joshi M, Misra A. Dry powder inhalation of liposomal Ketotifen fumarate: formulation
1079
and characterization. Int J Pharm. 2001;223(1-2):15-27.
1080 1081
88. Chougule M, Padhi B, Misra A. Development of spray dried liposomal dry powder
1082
inhaler of Dapsone. AAPS PharmSciTech. 2008;9(1):47-53.
1083
44 | P a g e
1084
89. Chougule M, Padhi B, Misra A. Nano-liposomal dry powder inhaler of tacrolimus:
1085
preparation, characterization, and pulmonary pharmacokinetics. Int J Nanomedicine.
1086
2007;2(4):675-688.
1087 1088
90. Changsan N, Chan HK, Separovic F, Srichana T. Physicochemical characterization and
1089
stability of rifampicin liposome dry powder formulations for inhalation. J Pharm Sci.
1090
2009;98(2):628-639.
1091 1092
91. Giovagnoli S, Blasi P, Vescovi C, Fardella G, Chiappini I, Perioli L, Ricci M, Rossi C.
1093
Unilamellar
1094
physicochemical characterization. AAPS PharmSciTech. 2004;4(4):E69.
vesicles
as
potential
capreomycin
sulfate
carriers:
preparation
and
1095 1096
92. Parumasivam T, Chang RY, Abdelghany S, Ye TT, Britton WJ, Chan HK. Dry powder
1097
inhalable formulations for anti-tubercular therapy. Adv Drug Deliv Rev. 2016;102:83-101.
1098 1099
93. Taylor KM, Newton JM. Liposomes for controlled delivery of drugs to the lung. Thorax.
1100
1992 Apr;47(4):257-259.
1101 1102
94. King RJ. Isolation and chemical composition of pulmonary surfactant, in Robertson B,
1103
Van Golde LMG, Batenburg JJ (Eds.), Pulmonary surfactant, Amstedam, Elsevier, 1984:1-
1104
15.
1105 1106
95. Wright JR, Clements JA. Metabolism and turnover of lung surfactant. Am Rev Respir Dis
1107
1987;135:426-444.
1108
45 | P a g e
1109
96. Lopez-Rodriguez E, Pérez-Gil J. Structure-function relationships in pulmonary surfactant
1110
membranes: from biophysics to therapy. Biochim Biophys Acta. 2014;1838(6):1568-1585.
1111 1112
97. Oyarzun MJ, Clements JA, Baritussio A. Ventilation enhances the pulmonary alveolar
1113
clearance of radioactive dipalmitoylphosphatidylcholine in liposomes. Am Rev Respir Dis
1114
1980;121:709-721.
1115 1116
98. Morimoto Y, Adachi Y. Pulmonary uptake of liposomal phosphatidylcholine upon
1117
intratrachealadministation to rats. Chem Pharmaceut Bull 1982;30:2248-2251.
1118 1119
99. Martin WJ, Standing JE. Amiodarone pulmonary toxicity: biochemical evidence for a
1120
cellular phospholipidosis in the bronchoalveolar lavage of human subjects. J Pharmacol
1121
ExpTher 1988;244:774-779.
1122 1123
100. Geiser M, Casaulta M, Kupferschmid B, Schulz H, Semmler-Behnke M, Kreyling W.
1124
The role of macrophages in the clearance of inhaled ultrafine titanium dioxide particles. Am J
1125
Respir Cell Mol Biol. 2008;38(3):371-376.
1126 1127
101. Shah V, Taratula O, Garbuzenko OB, Patil ML, Savla R, Zhang M, Minko T.
1128
Genotoxicity of different nanocarriers: possible modifications for the delivery of nucleic
1129
acids. Curr Drug Discov Technol. 2013;10(1):8-15.
1130 1131
102. Cipolla D, Gonda I, Chan HK. Liposomal formulations for inhalation. Ther Deliv.
1132
2013;4(8):1047-1072.
1133
46 | P a g e
1134
103. Agnihotri SA, Soppimath KS, Betageri GV. Controlled release application of
1135
multilamellar vesicles: a novel drug delivery approach. Drug Deliv. 2010;17(2):92-101.
1136 1137
104. Payne NI, Timmins P, Ambrose CV, Ward MD, Ridgway F. Proliposomes: a novel
1138
solution to an old problem. J Pharm Sci 1986;75:325-329.
1139 1140
105. Elhissi A, Phoenix D, Ahmed W. Some approaches to large-scale manufacturing of
1141
liposomes, in Ahmed W and Jackson MJ (Eds.), Emerging Nanotechnologies for
1142
Manufacturing, 2nd Eds, William Andrew Publishing, 2015: 402-417.
1143 1144
106. Khan I, Elhissi A, Shah M, Alhnan MA, Ahmed W. Liposome-based carrier systems and
1145
devices used for pulmonary drug delivery, in Davim JP (Eds.), Biomaterials and Medical
1146
Tribology, Woodhead Publishing, 2013:395-443.
1147 1148
107. Patil-Gadhe A, Pokharkar V. Single step spray drying method to develop proliposomes
1149
for inhalation: a systematic study based on quality by design approach. Pulm Pharmacol
1150
Ther. 2014;27(2):197-207.
1151 1152
108. Rojanarat W, Changsan N, Tawithong E, Pinsuwan S, Chan HK, Srichana T. Isoniazid
1153
proliposome powders for inhalation-preparation, characterization and cell culture studies. Int
1154
J Mol Sci. 2011;12(7):4414-4434.
1155 1156
109.
Rojanarat
1157
Inhaled pyrazinamide proliposome for targeting alveolar macrophages. Drug Deliv. 2012;
1158
19(7):334-345.
47 | P a g e
W, Nakpheng
T, Thawithong
E, Yanyium
N, Srichana
T.
1159 1160
110. Rojanarat W, Nakpheng T, Thawithong E, Yanyium N, Srichana T. Levofloxacin-
1161
proliposomes: opportunities for use in lung tuberculosis. Pharmaceutics. 2012;4(3):385-412.
1162 1163
111. Hassan S, Prakash G, Ozturk AB, Saghazadeh S, Sohail MF, Seo J, et al. Evolution and
1164
clinical translation of drug delivery nanomaterials. Nano Today. 2017;15:91-106.
1165 1166
112. Masic I, Miokovic M, Muhamedagic B. Evidence based medicine–new approaches and
1167
challenges. Acta Informatica Medica. 2008;16(4):219.
1168 1169
113. Study to Evaluate Efficacy of LAI When Added to Multi-drug Regimen Compared to
1170
Multi-drug
1171
https://clinicaltrials.gov/ct2/show/NCT02344004?term=NCT02344004&rank=1.
Regimen
Alone
(CONVERT).
1172 1173
114. Extension Study of Liposomal Amikacin for Inhalation in Cystic Fibrosis (CF) Patients
1174
With
1175
https://clinicaltrials.gov/ct2/show/NCT01316276?term=NCT01316276&rank=1
Chronic
Pseudomonas
Aeruginosa
(Pa)
Infection.
1176 1177
115. Study to Evaluate Arikayce™ in CF Patients With Chronic Pseudomonas Aeruginosa
1178
Infections.
1179
https://clinicaltrials.gov/ct2/show/NCT01315678?term=NCT01315678&rank=1
1180 1181
116. Inhalation SLIT Cisplatin (Liposomal) for the Treatment of Osteosarcoma Metastatic to
1182
the Lung. https://clinicaltrials.gov/ct2/show/NCT00102531?term=NCT00102531&rank=1
1183
48 | P a g e
1184
117. Evaluation of a Therapeutic Strategy Including Nebulised Liposomal Amphotericin B
1185
(Ambisome®) in Maintenance Treatment of Allergic BronchopulmonaryAspergillosis
1186
(Cystic Fibrosis Excluded). (NEBULAMB)
1187
https://clinicaltrials.gov/ct2/show/NCT02273661?term=NCT02273661&rank=1
1188 1189
118. Value of Amphotericin B Inhalation for Prophylaxis of Invasive Pulmonary
1190
Aspergillosis
1191
Transplantation.https://clinicaltrials.gov/ct2/show/NCT00986713?term=NCT00986713&rank
1192
=1
After
Renal
1193 1194
119.LipoAerosol©
1195
Tracheostomy.https://clinicaltrials.gov/ct2/show/NCT02157129?term=NCT02157129&rank=
1196
1
Inhalation
After
1197 1198
120. Phase II Study of Inhaled AeroLEF for Analgesia After ACL Knee Surgery (Pain).
1199
https://clinicaltrials.gov/ct2/show/NCT00791804?term=NCT00791804&rank=1.
1200 1201
121. Study Evaluating Inhaled AeroLEF Delivered in 4 Aerosol Delivery Devices in Healthy
1202
Volunteers
1203
https://clinicaltrials.gov/ct2/show/NCT00794209?term=NCT00794209&rank=1.
(LEF-07).
1204 1205
122.
Farjadian
1206
Nanopharmaceuticals and
1207
opportunities. Nanomedicine (Lond). 2019;14(1):93-126.
1208
49 | P a g e
F, Ghasemi
A, Gohari
O, Roointan
nanomedicines currently on
A, Karimi
M, Hamblin
MR.
the market: challenges and
1209
123. Limeres MJ, Moretton MA, Bernabeu E, Chiappetta DA, Cuestas ML. Thinking small,
1210
doing big: Current success and future trends in drug delivery systems for improving cancer
1211
therapy with special focus on liver cancer. Mater Sci Eng C Mater Biol Appl. 2019;95:328-
1212
341.
1213 1214
124. Giovagnoli S, Schoubben A, Ricci M. The long and winding road to inhaled TB therapy:
1215
not only the bug’s fault. Drug Dev. Ind. Pharm. 2017;43(3):347-363.
1216 1217
125. Mehta PP, Kadam SS, Pawar AP. Influence of modified induction port, modified DUSA
1218
assembly and device air-inlet geometry on the aerosolization pattern of a dry powder inhaler.
1219
J Drug Deliv Sci Technol. 2020;55:101416.
1220 1221
126. Rogueda P, Traini D. The future of inhalers: how can we improve drug delivery in
1222
asthma and COPD? Expert Rev Respir Med. 2016; 10:1041-1044.
1223 1224
127. Guillon A, Sécher T, Dailey LA, Vecellio L, de Monte M, Si-Tahar M et al. Insights on
1225
animal models to investigate inhalation therapy: relevance for biotherapeutics. Int J
1226
Pharm. 2018;536(1):116-126.
1227 1228
128. Vulović A, Šušteršič T, Cvijić S, Ibrić S, Filipović N. Coupled in silico platform:
1229
Computational fluid dynamics (CFD) and physiologically-based pharmacokinetic (PBPK)
1230
modelling. Eur J Pharm Sci. 2018;113:171-184.
1231
50 | P a g e
1232
129. Backman P, Arora S, Couet W, Forbes B, de Kruijf W, Paudel A. Advances in
1233
experimental and mechanistic computational models to understand pulmonary exposure to
1234
inhaled drugs. Eur J Pharm Sci. 2018;113:41-52.
1235 1236
130. Mehta PP. Dry powder inhalers: upcoming platform technologies for formulation
1237
development. Ther Deliv. 2019 Sep;10(9):551-554.
1238 1239 1240 1241 1242 1243 1244 1245 1246 1247 1248 1249 1250 1251 1252 1253 1254 1255 1256
51 | P a g e
1257
Figure captions
1258 1259
Graphical Abstract - Applications and interaction of liposomes with the pulmonary
1260
membrane. Liposomes are synthesized from substances endogenous to pulmonary airways
1261
such as pulmonary surfactants and hence, explored in the pulmonary drug delivery field
1262
owing to their versatile attributes. By interacting with the pulmonary membrane,
1263
multifunctional liposomes can adhere to the mucosal surface for a longer time, open
1264
intercellular tight junctions in pulmonary epithelia along with modified drug release kinetics
1265
and reduced dosing frequency to improve therapeutic outcomes.
1266 1267
Figure 1 - Liposome types based on lamellarity and size. SUV: small unilamellar vesicles,
1268
LUV: large unilamellar vesicles, GUV: giant unilamellar vesicles, MLV: multilamellar
1269
vesicles and MVV: multivesicular vesicles (vesosomes). Moreover, a fraction of classic lipid
1270
bilayer with hydrophilic head groups and hydrophobic alkyl chains shown.
1271 1272
Figure 2 - Applications and interaction of liposomes with the pulmonary membrane.
1273
Liposomes are synthesized from substances endogenous to pulmonary airways such as
1274
pulmonary surfactants and hence, explored in the pulmonary drug delivery field owing to
1275
their versatile attributes. By interacting with the pulmonary membrane, multifunctional
1276
liposomes can adhere to the mucosal surface for a longer time, open intercellular tight
1277
junctions in pulmonary epithelia along with modified drug release kinetics and reduced
1278
dosing frequency to improve therapeutic outcomes. (MRT: mean resisdance time; Cmax: the
1279
maximum concentration of the drug achieved in the plasma and PEG: polyethylene glycol).
1280
52 | P a g e
1281
Figure 3 -Graphical representation depicting the mechanism of action for liposomal
1282
andrographolide DPI used in regulating immune responses (Lie et al., 2017) [81]. The figure
1283
showed that liposomal andrographolide DPI regulates immune responses by downregulating
1284
various inflammatory pathways such as inhibit phosphorylation of kappa B (IκB-α) inhibitor
1285
and reduced the expression of intercellular adhesion molecule (ICAM) family 1, Interferon-γ
1286
(IFN-γ) in immunocytes.
1287 1288
Figure 4 -Benefits and challenges of multifunctional inhaled liposomes. The figure showed
1289
crucial technical, biological and formulations benefits as well as challenges related to the
1290
design and development of multifunctional inhaled liposomes. This figure emphasis various
1291
key factors such as chemistry, manufacturing and control (CMC), center for devices and
1292
radiological health (CDRH), forced vital capacity (FVC) and forced expiratory volume
1293
(FEV). (IPR: intellectual property rights, ADME: absorption, distribution, metabolism, and
1294
excretion and MRT: mean resisdance time).
53 | P a g e
Table 1 - Advantages and disadvantages of liposome preparation methods Preparation
Particle size Advantages
method
(nm)
Disadvantages
References
Conventional method Thin-lipid
film 100-1000
hydration
A
simple,
conventional Low yield, difficult to scale up, low [38-40]
process
encapsulation
efficacy,
heterogeneous
particle size Reverse-phase
100-1000
High entrapment efficacy
Organic
evaporation Ethanol
solvent
traces,
difficult
to [39-41]
manufacture at large scale injection 70-200
Ability to control vesicle size
method
Reproducibility, difficult to remove ethanol [39,40,42] traces, heterogeneous particle size, require high temperature
Novel methods Microfluidic technology
100-300
High
entrapment
suitable homogenous
1|Page
for
efficacy, Prerequisites
of
equipment,
various [29,40,
scale-up, processing conditions; thus need more 43,44]
particle
size, optimization
appropriate
for
template-
based manufacturing Membrane
~ 100
contactor
Homogenous high
particle
entrapment
size, Encapsulation requires optimization
[40,45]
efficacy,
suitable for scale up Electrospray technology
100-500
Continuous
method,
encapsulation efficiency
high Prerequisites processing optimization
2|Page
of
equipment,
parameters;
thus
various [29,46,47] critical
Table 2 - Advantages and limitations of novel carrier-based pulmonary drug delivery system Advantages
Limitations
Protection from biological environment
Complex and multi-step process
(enzymes, hydration, pH variation) Masking of immunogenicity resulting from API
High cost
Both hydrophilic and hydrophobic API can be easily loaded
Complex control of reproducibility i.e. particle size distribution and/or surface charge
Controlled and modified drug release
Need to conduct toxicological studies
Triggered released in response to physical or chemical different Need to gather and understand ADME pattern stimuli (pH/temperature) API: active pharmaceutical ingredient; ADME: absorption, distribution, metabolism and elimination
3|Page
Table 3- Developed liposomal formulations for pulmonary delivery Drug
Method Key ingredient
Particle
size Device
FPF (%)
References
(nm) SS
FD
SPC, α-LM and MgSt (0.5 %)
137.00 (LUVs)
Spinhaler®
41.51
[76]
Docetaxel
SD
PC, CHOL, mannitol and leucine
346.80 (LUVs)
Reusable
10.10
[71]
device N-acetylcysteine
SD
SPC, CHOL and α-LM
117.00 (LUVs)
HandiHaler®
35.34
[84]
Ciprofloxacin
SD
HSPC, CHOL and sucrose
131.30 (LUVs)
Osmohaler®
69.70
[85]
Liquorice extract FD
SPC, CHOL and trehalose
212.60 (LUVs)
Lupihaler®
54.68
[82]
Andrographolide
FD
Soybean lecithin, CHOL and mannitol
77.91 (SUVs)
Twister®
23.03
[81]
SS
SD
SPC, CHOL and α-LM
167.20 (LUVs)
Rotahaler®
64.01
[77]
SS
SD
DPPC, CHOL and α-LM
-
Cyclohaler®
42.70
[86]
Ketotifen
FD
EPC, CHOL and sucrose
-
Rotahaler®
21.59
[87]
Dapsone
SD
DPPC, CHOL and α-LM
137.00 (LUVs)
-
75.60
[88]
Clarithromycin
FD
SPC, CHOL, mannitol (15 %) and 370.00 (LUVs)
-
53.78
[78]
fumarate
4|Page
sucrose (5 %) Gemcitabine
FD
HSPC, DSPG, CHOL mPEG2000- 331.42 (LUVs)
-
56.12
[72]
DSPE and trehalose Curcumin
FD
SPC, CHOL and mannitol
94.65 (SUVs)
-
46.71
[79]
Oseltamivir
SD
Ovelecithin, CHOL and leucine
105.90 (LUVs)
-
35.40
[74]
Tacrolimus
SD
HSPC, CHOL and trehalose
140.00 (LUVs)
-
71.10
[89]
Rifampicin
FD
SPC, CHOL and mannitol
255.00 (LUVs)
-
66.80
[90]
Moxifloxacin
SD
PC, CHOL and dextran
272.00 (LUVs)
-
72.08
[75]
phosphate
SS: salbutamol sulphate; SD: spray drying; FD: freeze drying; SPC: Soya phosphatidylcholine; CHOL: cholesterol; HSPC: hydrogenated soybean
phosphatidylcholine;
DSPG:
1,2-Distearoyl-sn-glycero-3-phosphoglycerol;
dipalmitoylphosphatidylcholine and EPC: egg phosphatidylcholine
5|Page
PC:
phosphatidylcholine;
DPPC:
Table 4 - A summary of ongoing clinical trials for liposomal formulations Product
Condition or disease
Intervention/ treatment
Phase
Identifier
Sponsor
Arikayce™
Cystic fibrosis
280 mg of matching placebo
I and II
NCT00777296
Insmed Incorporated., US
Arikayce™
Cystic fibrosis
70 mg; 140 mg and 560 mg
I and II
NCT00558844
Insmed Incorporated., US
Cisplatin
Osteosarcoma
24 mg/m2 once daily
I and II
NCT00102531
Insmed Incorporated., US
liposomes
metastatic
for 14 days
Cyclosporine
Bronchiolitis obliterans
2.5 mg/10 mg x 2/day
I and II
NCT01334892
Pari
liposomes
for 96 weeks
Cyclosporine
Lung
liposomes
and
transplantation -
Pharma
GmbH,
Germany I and II
NCT01650545
bronchiolitis
University of Maryland, US
obliterans Arikayce™
Cystic fibrosis
560 mg once daily dose for 6 II
NCT03905642
Insmed Incorporated., US
NCT03038178
Kevin Winthrop, Insmed
cycles over 18 months Arikayce™
MBI and NMBI
590 mg for 12 months
II
Incorporated and etc. Ambisome®
6|Page
Allergic
25 mg x 1/ week for 6 months
II
NCT02273661
Poitiers
University
bronchopulmonary
Hospital, France
aspergillosis Ambisome®
Lung transplantation
-
II
NCT01254708
University
Health
Network, Toronto Liposomal Nitro-20
9- Lung cancer (S)-
5 consecutive days per week II
NCT00250120
X 8 weeks, every 10 weeks
University
of
New
Mexico, US
camptothecin Amikacin
MBI and NMBI
590 mg/day for 12 months
II
NCT03038178
liposomes
Kevin Winthrop and Insmed Incorporated., US and etc.
Arikayce™
MBI and NMBI
590 mg administered
III
NCT02344004
Insmed Incorporated., US
III
NCT03270514
Kathirvel Subramaniam
III
NCT02628600
Insmed Incorporated., US
once daily Bupivacaine
Coronary artery disease
liposomes Liposomal amikacin
7|Page
Bupivacaine liposomes 226 mg
Non-tuberculous for infections
590 mg/day
inhalation Ambisome®
Lung
transplantation 1 mg/kg/day for 4 days
III
NCT00177710
and fungal infections
University of Pittsburgh and Astellas Pharma US, Inc.
Ciprofloxacin
Non-cystic
fibrosis -
liposomes
bronchiectasis
III
NCT01515007
Grifols
(ORBIT-3) Cyclosporine
Bronchiolitis obliterans
Twice daily inhalation for a III
NCT01439958
maximum of three years
Ambisome®
Chronic
with
aspergillosis
for
24
weeks)
Pari
Pharma
GmbH,
Germany
pulmonary Ambisome (25 mg x 2/week III
itraconazole
NCT03656081
and
Poitiers
University
Hospital, France
itraconazole (200 mg x 2/day)
Ciprofloxacin
Non-cystic
liposomes
bronchiectasis
fibrosis -
III
NCT02104245
Aradigm Corporation Grifols
(ORBIT-4)
8|Page
Therapeutics
LLC
liposomes
Liposomal
Aradigm Corporation and
Therapeutics
LLC Lung cancer
-
-
NCT00277082
University
of
New
camptothecin LipoAerosol®
Mexico, US Tracheostomy complications
5x/d for 30 min
-
NCT02157129
Technical University of Munich, Germany
ArikayceTM : amikacin liposome inhalation suspension; Ambisome ®: amphotericin B liposomal; MBI: mycobacterial infection and NMBI: nonmycobacterial infection.
9|Page
AUTHOR DECLARATION TEMPLATE
We wish to draw the attention of the Editor to the following facts which may be considered as potential conflicts of interest and to significant financial contributions to this work. We confirm that the manuscript has been read and approved by all named authors and that there are no other persons who satisfied the criteria for authorship but are not listed. We further confirm that the order of authors listed in the manuscript has been approved by all of us. We confirm that we have given due consideration to the protection of intellectual property associated with this work and that there are no impediments to publication, including the timing of publication, with respect to intellectual property. In so doing we confirm that we have followed the regulations of our institutions concerning intellectual property. We understand that the Corresponding Author is the sole contact for the Editorial process (including Editorial Manager and direct communications with the office). He/she is responsible for communicating with the other authors about progress, submissions of revisions and final approval of proofs. We confirm that we have provided a current, correct email address which is accessible by the Corresponding Author and which has been configured to accept email from [email protected]; [email protected]
Corresponding author; Dr. Vividha Dhapte-Pawar, Associate Professor, Department of Pharmaceutics, Bharati Vidyapeeth University, Poona College of Pharmacy, Pune -411038 Email: [email protected]; [email protected]
S1773-2247(19)31859-3
DOI:
https://doi.org/10.1016/j.jddst.2020.101509
Reference:
JDDST 101509
To appear in:
Journal of Drug Delivery Science and Technology
Received Date: 1 December 2019 Revised Date:
28 December 2019
Accepted Date: 7 January 2020
Please cite this article as: P.P. Mehta, D. Ghoshal, A.P. Pawar, S.S. Kadam, V.S. Dhapte-Pawar, Recent advances in inhalable liposomes for treatment of pulmonary diseases: Concept to clinical stance, Journal of Drug Delivery Science and Technology (2020), doi: https://doi.org/10.1016/ j.jddst.2020.101509. This is a PDF file of an article that has undergone enhancements after acceptance, such as the addition of a cover page and metadata, and formatting for readability, but it is not yet the definitive version of record. This version will undergo additional copyediting, typesetting and review before it is published in its final form, but we are providing this version to give early visibility of the article. Please note that, during the production process, errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain. © 2020 Published by Elsevier B.V.
Recent advances in inhalable liposomes for treatment of pulmonary diseases: Concept to Clinical Stance
Piyush P. Mehta 1, Debjit Ghoshal 2, Atmaram P. Pawar 2, Shivajirao S. Kadam 3 and Vividha S Dhapte-Pawar 2*
1
Department of Quality Assurance, Poona College of Pharmacy, Bharati Vidyapeeth Deemed
University, Pune - 38, Maharashtra, India. 2
Department of Pharmaceutics, Poona College of Pharmacy, Bharati Vidyapeeth Deemed
University, Pune - 38, Maharashtra, India. 3
Bharati Vidyapeeth Bhavan, Bharati Vidyapeeth Deemed University, LBS Road, Pune- 30,
Maharashtra, India.
*Corresponding author Dr. Vividha S Dhapte-Pawar, Associate Professor, Department of Pharmaceutics, Bharati Vidyapeeth University, Poona College of Pharmacy, Erandwane, Kothrud, Pune 411038. Email: [email protected]; [email protected]
1
Recent advances in inhalable liposomes for treatment of pulmonary diseases: Concept
2
to Clinical Stance
3 4
Graphical Abstract
5
6 7 8 9 10 11 12 13 14 15 16 17 1|Page
18
Recent advances in inhalable liposomes for treatment of pulmonary diseases: Concept
19
to Clinical Stance
20 21
Abstract
22
Liposomes and liposomes-based drug delivery systems are promising carriers in the rapidly
23
evolving field of nanomedicine. Liposomes are receiving growing attention in the scientific
24
domain owing to their distinctive structural characteristics, physiological features and
25
biological properties. These versatile lipid vesicles represent unique platforms for drug
26
delivery, targeting, diagnostics, imaging as well as theranostics. The main objective of the
27
present article is to present insights into the pulmonary delivery of these versatile cargos for
28
nanomedicine. Inhalable liposomes for pulmonary delivery present unique advantages as they
29
are formulated from phospholipids similar to endogenous pulmonary surfactant. The article
30
begins by unfolding the important background information about liposomes and their key
31
advantages as nanomaterials. In the subsequent section, various liposomes and proliposome
32
formulations are summarized with a special emphasis on their physicochemical properties,
33
aerosolization performance and in-vivo aerodynamic behavior. Additionally, this article
34
includes a segment devoted to recent status on clinical trials of liposomal formulations for
35
treating pulmonary diseases. Moreover, the present article contains a section dedicated to the
36
market potential and scientific challenges related to inhalable liposomes. In summary, this
37
article is a comprehensive report of inhalable liposomes to appear in recent years.
38 39
Keywords
40
Liposomes, proliposome, nanomedicine, nanocarriers, dry powder inhalers, aerodynamic
41
behavior, drug delivery
42
2|Page
43
Abbreviations
44
Chronic obstructive pulmonary disease (COPD); world health organization (WHO);
45
pressurized metered-dose inhalers (pMDIs); soft mist inhalers (SMI); dry powder inhalers
46
(DPI); novel drug delivery systems (NDDS); small unilamellar vesicles (SUVs); large
47
unilamellar vesicles (LUVs); giant unilamellar vesicles (GUVs); multilamellar vesicles
48
(MLV); multi-vesicular vesicles (MVV); double emulsion templating (DET); microfluidic
49
hydrodynamic focusing (MHF); reticuloendothelial system (RES); polyethylene glycol
50
(PEG); enhanced permeability and retention (EPR); solid lipid nanoparticles (SLNs);
51
nanostructured lipid carriers (NLCs); nanoparticles embedded microparticles (NEMs);
52
encapsulation
53
bromide (MTT); half-maximal inhibitory concentration (IC50); folate receptor alpha
54
(SPCA1); hydrogenated soy phosphatidylcholine (HSPC); 1,2-Distearoyl-sn-glycero-3-
55
phosphoglycerol (DSPG); fine particle fraction (FPF); emitted dose (ED); mass median
56
aerodynamic diameter (MMAD); human lung adenocarcinoma (A549); maximum tolerated
57
dose (MTD); area under curve (AUC); mean residence time (MRT); 1,2-dioleoyl-sn-glycero-
58
3-[phospho-L-serine] (DOPS); twin stage impinger (TSI); maximum drug concentration
59
(Cmax); 4-aminophenyl-alpha-D-manno-pyranoside (PAM); next generation impactor (NGI);
60
3-methylcholanthrene (MCA); diethyl nitrosamine (DEN); tumor necrosis factor-α (TNF-α);
61
vascular endothelial growth factor (VEGF); malondialdehyde (MDA); caspase-3 and B-cell
62
lymphoma 2 protein (BCL-2); 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid (HEPES);
63
nuclear factor-κB (NF-κB); geometric standard deviation (GSD); idiopathic pulmonary
64
fibrosis (IPF); dipalmitoylphosphatidylcholine (DPPC); phosphatidylcholines (PC); the phase
65
transition temperature (Tm); normal human bronchial epithelial cells (NHBE); small airway
66
epithelial cells (SAEC); alveolar macrophages (AMs); enzyme linked immunosorbent assay
67
(ELISA); human lung cancer cell line cells (Calu-3); human alveolar basal epithelial cell
3|Page
efficiency
(EE);
3-(4,5-dimethylthiazole-2-yl)-2,5-diphenyltetrazolium
68
(NR8383); chemistry, manufacturing and control (CMC); forced vital capacity (FVC); forced
69
expiratory volume (FEV); Tiffeneau-Pinelli index (FEV1/FVC); maximal voluntary
70
ventilation (MVV).
71 72
1. Introduction
73
Even with the continuous efforts and progress made in pulmonary research, pulmonary
74
ailments still remain a major health issue across the globe. Pulmonary diseases are leading
75
causes of morbidity and mortality worldwide [1]. Some of the most prevalent pulmonary
76
ailments are asthma, chronic obstructive pulmonary disease (COPD), pulmonary tuberculosis,
77
lung cancer, pulmonary hypertension, chronic bronchitis, emphysema and occupational lung
78
diseases. COPD is not one single disease but an umbrella phrase commonly utilized to
79
express diverse chronic lung disorders that cause limitations in pulmonary airflow [2].
80
Almost 65 million individuals suffer from COPD and more than 3 million fatalities are
81
reported annually making it the third major cause of mortality worldwide. According to the
82
World Health Organization (WHO) statistics, approximately 235 million people suffer from
83
asthma and it is the most ordinary pulmonary disease among the paediatric population [3].
84
Moreover, the family, not just the individual, suffer the impact and crisis of pulmonary
85
diseases. In brief, pulmonary diseases are prevalent in every part of the world; this public
86
health problem is just not limited to 2nd and 3rd income countries but affects all countries
87
irrespective of their developmental levels. In view of the impact of pulmonary diseases on a
88
universal scale, it is apparent that these ailments have raised a significant public health
89
challenge [4].
90 91
Various pulmonary delivery systems such as pressurized metered-dose inhalers (pMDIs), soft
92
mist inhalers (SMI), dry powder inhalers (DPI) and nebulizers have been designed, developed
4|Page
93
and studied for the treatment of pulmonary diseases. Among them, DPIs are more preferred
94
dosage forms because of their better physicochemical stability and capability to deliver the
95
drug into deep lungs using the patient’s respiration [5,6]. Each type of delivery system has
96
distinct strengths and weaknesses considering the disease pathophysiology, severity and type
97
of prescribed formulation. Furthermore, it has been well reported that the therapeutic success
98
of inhalation therapy is highly dependent on particle size distribution, inhalation flow rate,
99
inhaler device resistance and dispersion capability [7]. Therefore, to maximize the clinical
100
outcome of inhalation therapy, a formulation with apt physicochemical properties is vital [8].
101 102
With the continuous growth of various scientific fields such as particle engineering, materials
103
science, biotechnology, molecular biology, green chemistry and biomaterials, rapid
104
advancement has been achieved in the domain of novel drug delivery systems (NDDS) [9].In
105
addition, recent progress in nanotechnology has opened avenues for augmenting the clinical
106
value of inhalation therapy for different pulmonary diseases [10]. The application of
107
nanotechnology to the design of NDDS such as polymeric systems (polymer-drug conjugates,
108
polymeric nanoparticles, polymeric micelles) [11], nanoparticles [12,13], nanoaggregates,
109
nanocomposites [14], dendrimers, quantum dots, fullerenes, novel lipid vesicles (liposomes,
110
pro-liposomes, solid lipid nanoparticles, nanostructured lipid carriers, cochleates and lipidic
111
spherulites), ethosomes, cubosomes, lipomers, polymeric microparticles and microspheres
112
[15] have illustrated numerous therapeutic advantages over traditional pulmonary
113
formulations [9,16]. The design and development of NDDS based DPIs have the potential to
114
surpass issues associated with the carrier as a critical formulation component. These novel
115
formulations help in improving physicochemical stability, tissue distribution, in-vivo drug
116
deposition and bioavailability [11,17]. To sum up, such novel formulations represent
117
promising alternatives to traditional inhaled formulations.
5|Page
118 119
Among various novel bioactive platforms, lipid vesicles i.e. liposomes have attracted the
120
focus of formulation scientists as they can be prepared from substances endogenous to the
121
pulmonary airways as components of the lung with surfactant and other unique
122
physicochemical and biopharmaceutical properties [18]. The application of liposomes in drug
123
delivery has been studied and investigated by numerous formulation scientists [19]. Several
124
active scientists and clinicians have studied the significance of liposomes systems for drug
125
delivery, drug targeting, tissue engineering, diagnostics, detection of genetic material and
126
imaging [20-22]. In brief, the superior therapeutic efficacy of liposomes has been evidenced
127
either in laboratory investigations or in clinical analysis, particularly in the therapy of
128
pulmonary diseases.
129 130
By knowing this continues development, in this article, we have analyzed inhalable liposomal
131
formulations for the treatment of numerous pulmonary ailments. In the opening section of
132
this article, we have discussed liposomes, generations of liposomes and the importance of
133
liposomes-based nanomaterials. In the next segment, various dry powdered liposomal
134
formulations are summarized with a special emphasis on their physicochemical properties,
135
aerosolization performance and in-vivo behavior. In the third part, the importance of
136
respirable pro-liposome formulations and their influence on pulmonary drug delivery are
137
thoroughly summarized. Additionally, this article includes a section devoted to recent efforts
138
taken in clinical trials of liposomal formulations for the treatment of pulmonary diseases.
139
Moreover, this article concisely explores the market potential of inhaled liposomal
140
formulations. Furthermore, the present article contains a segment dedicated to the practical
141
concerns, technical limitations and scientific challenges related to inhalable liposomes. This
6|Page
142
review can be of potential significance from the perspective of both academicians and
143
industrial scientists.
144 145
2. Methodology
146
We performed a broad literature search for liposomes and pro-liposomes for pulmonary drug
147
delivery. This literature search included scientific journals, books and book chapters from
148
various electronic sources such as ScienceDirect, PubMed, Google Scholar, Springer and
149
Web of Science. Present article investigates various key aspects associated to inhalable
150
liposomes and pro-liposomes with special importance on the prepration methods, in-vitro
151
aerodynamic performance and pulmokinetic parameters. References enlisted in present article
152
contains 130 articles, containing peered review articles, original research articles, clinical trial
153
reports and book chapters. All articles were examined, screened cautiously and then selected
154
for the review. ChemDraw software (Version 12.2; Perkin Elmer) was used to draw the
155
figures.
156 157
3. Liposomes
158
Liposomes perhaps are the most extensively explored and characterized lipid-based drug
159
delivery systems. Liposomes are the sub-micron spherical phospholipid vesicles consisting of
160
single or multiple concentric lipid bilayers enclosing aqueous interior [23]. Liposomes were
161
first discovered and explained in 1965 by Bangham and his co-workers. Since then, they are
162
extensively studied as a drug reservoir from the last five to six decades [24]. Accordingly,
163
liposomes have become one of the most explored drug reservoirs in diagnosis, treatment and
164
imaging of several diseases [25]. Generally, on basis of vesicular arrangement liposome can
165
be classified as unilamellar liposomes (single bilayer lipid membrane) i.e. small unilamellar
166
vesicles (SUVs; 20-100 nm); large unilamellar vesicles (LUVs; 100 nm -1 mm) and giant
7|Page
167
unilamellar vesicles (GUVs; > 1 mm) or multilamellar liposomes (numerous bilayer lipid
168
membranes)
169
MVV is also known as vesosomes (Fig. 1) [26]. Phospholipids and cholesterol are the main
170
two components of liposomal bilayers. Cholesterol plays a vital role in the physical stability
171
of liposome vesicles with its ability to increase the phospholipid phase transition temperature
172
(TM) [27,28]. Many techniques have been explored for the design and development of
173
liposomes (Table1). The most popular techniques that have been explored to develop
174
liposomes are the thin-film (Bangham method) and the ethanol injection method [29]. Each
175
type of technique has its own unique strengths and weaknesses considering the type of lipid
176
and the nature of drugs that can be utilized. On account of this, many formulation scientists
177
are now actively engaged in developing strategies to improve the physicochemical quality of
178
the liposomes. Microfluidic technology, membrane contactor, electrospray mechanism are a
179
helpful alternative for the fabrication of liposomes [29]. Moreover, new techniques derived
180
from microfluidic approaches for fabrication of liposome mainly include double emulsion
181
templating (DET) [30], electro formation and hydration [31], pulsed jetting [32], ice droplet
182
hydration [33], extrusion [34], transient membrane ejection [35], droplet emulsion transfer
183
[36] and microfluidic hydrodynamic focusing (MHF) [37]. Among these methods, templating
184
is a good option to fabricate liposomes of consistent size and high encapsulation efficiency
185
[29].
i.e. multilamellar vesicles (MLV) or multi-vesicular vesicles (MVV) [26].
186 187
Additionally, based on structural modifications, liposomes can be classified into the different
188
‘generations’. The first generation liposomes mainly consist of phospholipids and cholesterol
189
vesicles without any structural modifications. The mean particle size of first-generation
190
liposomes is in the range of 50 to 450 nm [48]. As a nanoscale drug reservoir, the liposomes
191
are biodegradable, biocompatible with low toxicity. As well, the liposomes are also
8|Page
192
convenient considering the scale-up and the quality control specifications [49]. Liposomes
193
are capable to hold both, lipophilic as well as hydrophilic molecules. Thus, a wide range of
194
therapeutic compounds can be easily incorporated into the liposomes [50]. Various synthetic
195
drugs, semi-synthetic compounds, herbs, herb extracts and phytoconstituents can be easily
196
incorporated into the liposomes. Ambisome® (Astellas Pharma), Amphotec® (Intermune), and
197
Abelcet® (Enzon) [amphotericin B liposomes] are few successful, commercial liposomal
198
products approved for the treatment of fungal infections across the globe [51]. Yet, these
199
first-generation liposomes displayed noticeable limitations such as leakage of drugs from
200
liposome vesicles, low loading capacity for hydrophilic compounds and rapid clearance of
201
vesicles by the reticuloendothelial system (RES) in the systemic circulation. Therefore, the
202
first generation liposomes were almost abandoned until the active drug loading and sterically
203
stable second-generation liposomes were discovered [52].
204 205
The second-generation liposomes are long-circulating, surface modified liposomes. Usually,
206
liposomes are coated with inert polymers i.e. polysaccharides, oligosaccharides,
207
glycoproteins and synthetic polymers [53]. Among these polymers, polyethylene glycol
208
(PEG) is most commonly explored to stabilize the liposomes. PEG-modified (PEG-coated)
209
liposomes are often denoted as sterically stabilized or ‘stealth liposomesʼ [54]. Several active
210
drug loading techniques have been explored to achieve higher drug encapsulation efficiency.
211
The active drug loading techniques are distant encapsulation methods that load the drug into
212
the liposomes by the differential chemical gradient across the membrane of the liposomes.
213
The chemical gradients mainly comprise the pH of the system, ammonium sulfate gradient or
214
calcium acetate gradient. As an effect, the liposomes exhibited favourable physicochemical
215
and biological properties such as reduced drug leakage, prolonged systemic circulation,
216
modified release pattern and altered bio-distribution. The prolonged systemic circulation is
9|Page
217
mainly ascribed to the steric hindrance effect presented by a hydrophilic polymer which
218
could avoid the surface-modified liposomes from being rapidly purged by the RES [53].
219
Consequently, a number of surface-modified liposomes were studied successfully and few of
220
them are under various phases of clinical trials. Few pegylated doxorubicin liposomes such as
221
Doxil® (PEG 2000 surface-modified liposome), Caelyx® (liposomes with surface-bound
222
methoxy-polyethylene glycol) and Myocet® (liposome-encapsulated doxorubicin citrate
223
complex) have been approved and marketed for the treatment of cancers [55]. Even though
224
great commercial achievements have been made, the second generation liposomes
225
demonstrated few downsides such as poor cancer cells selectivity and low cellular uptake
226
[56].
227 228
The targeting liposomes are known as the third generation liposomes wherein the targeting
229
ligand molecules or functional moieties are incorporated into liposomes to target a specific
230
site. The third generation liposomes may possess passive as well as active targeting
231
properties. Both of the first and second generation liposomes fit into the passive targeting
232
reservoirs with high lymphatic affinity attributed to the lipophilic casing of liposomes. These
233
first and second-generation liposomes displayed the enhanced accumulation at tumor site
234
owing to the enhanced permeability and retention (EPR) consequence [57]. Active targeting
235
liposomes can target cells and/or cellular organelles by a precise interaction between a
236
targeting ligand attached to the liposomes and a cell receptor [58]. Additionally, the diverse
237
class of chemical moieties such as carbohydrates, vitamins, monoclonal antibodies, peptides,
238
proteins and aptamers can be used as targeting ligands [59]. The enhanced therapeutic
239
efficacy of third-generation liposomes had been verified in the laboratory model particularly
240
in the treatment of cancer [56]. Nowadays, a novel dual-functional liposomes have appeared
241
as a promising drug delivery system. Dual-functional liposomes are typically formulated
10 | P a g e
242
using a mixture of phospholipid and functional material by either chemical adaptation or
243
physicochemical inclusion of functional motifs against the membrane of liposomes. Current
244
investigations showed that the dual-functional liposomes are capable of improving the
245
therapeutic efficiency by modifying drug delivery as well as the mechanism of action [56].
246 247
For the pulmonary application, liposomes offer numerous benefits (Fig. 2) [60,61]. Superior
248
tolerability of liposomes in the pulmonary airways can be guaranteed, if lipids selected are
249
biodegradable resulting in non-toxic, endogenous degradation products [60,62]. Furthermore,
250
owing to their small (nm) size, they can be easily encapsulated into particles with suitable
251
aerosolization properties, which facilitates satisfactory deep lung deposition of a molecule.
252
Additionally, liposomes adhere to the mucosal surface of the airways for a longer time
253
compared to larger particles owing to the small size. Particle size, adhesion, accumulation
254
and retention in the pulmonary airways along with controlled release characteristics of
255
liposomes can lead to improved therapeutic outcomes leading to better patient compliance
256
[60,63]. This can assist an imperative function in the therapy for chronic diseases as many of
257
the existing inhalation formulations have to be applied at least twice a day because of the
258
comparatively less duration of the drug in the pulmonary airways [62,65]. With this
259
background, various liposomal DPIs are described in the following section with special
260
emphasis on their aerodynamic behavior and clinical outcomes.
261 262
4. Pulmonary drug delivery
263
It is fascinating to believe in pulmonary therapy as a modern strategy for drug delivery but
264
this practice has been well acknowledged in most of the ancient literature and it has a strong
265
history of more than 4000 years [66]. From the therapeutic perspective, pulmonary therapy
266
represents a few attractive benefits allied with the anatomical and physiological
11 | P a g e
267
characteristics of the lung. Pulmonary therapy has a better potential of treating various
268
intrapulmonary and extrapulmonary diseases, such as asthma, COPD, cystic fibrosis,
269
pulmonary hypertension, pneumonia, cancer [67], diabetics and tuberculosis [68]. The
270
reduced invasiveness of pulmonary therapy may further improve clinical outcomes and
271
patient compliance with the treatment. Furthermore, owing to the scientific and technological
272
advances in formulations (e.g. Technosphere® particles, AerosphereTM delivery, iSPERSE™
273
platform and/or Capsugel® Zephyr™), particle replication in nonwetting templates (PRINT),
274
inkjet-printed aerogel particles and inhaler devices (e.g. Advair Diskus®, ProAir®
275
Digihaler™, Spiriva® Handihaler® and TwinCapsTM Inhaler), pulmonary therapy has become
276
more ‘patient-friendly’ and economically favourable [68,69].
277 278
Many different types of delivery systems such as nanoparticles (polymeric nanoparticles,
279
polymeric micelles), microparticles (microspheres), solid lipid nanoparticles (SLNs),
280
nanostructured lipid carriers (NLCs), polymer-drug conjugates, macromolecules (dendrimers)
281
and lipid vesicles (liposomes and proliposomes) have been designed and developed for
282
pulmonary delivery of several therapeutics. They fulfill many biopharmaceutical
283
requirements i.e. sufficient drug loading, shielding of the actives from degradation,
284
biocompatibility, biodegradability and stability during aerosolization. However, one major
285
limitation of these systems is that they can be readily exhaled from the lungs after pulmonary
286
delivery. Table 2 denotes various advantages and limitations of novel carrier-based
287
pulmonary drug delivery [70]. However, several formulation techniques, such as nano-
288
agglomeration processes, nanoparticle-rooted microparticles or nanoparticles embedded
289
microparticles (NEMs) can be utilized to form nanoparticles with appropriate aerodynamic
290
diameters for pulmonary delivery. Knowing these facts, lipid vesicles (i.e. liposomes and
291
proliposomes) for pulmonary delivery have been a popularized theme for the last two-three
12 | P a g e
292
decades [8,11]. Accordingly, the present segment discusses and analyzes the available
293
literature on inhalable liposomes and proliposomes.
294 295
4.1 Liposomes based dry powder inhalers
296
Zhu et al (2019) developed folic-acid conjugated docetaxel (1:25) liposomes (LP-DTX-FA)
297
using phosphatidylcholine and cholesterol (6:1) by thin lipid film hydration method.
298
Furthermore, obtained LP-DTX-FA were spray-dried using mannitol (2 % w/v; carrier) and
299
leucine (anti-adherent) to obtain an inhalable dry powder. Spherical shaped, smooth surface
300
spray-dried LP-DTX-FA showed mean particle size, zeta potential and encapsulation
301
efficiency (EE) of 346.80 nm, -29.30 mV and 99.50 %, respectively. During the in-vitro
302
release study, LP-DTX-FA showed sustained release (>70 %) up to 50 h in phosphate buffer
303
saline (PBS; pH 7.4). In in-vitro cytotoxicity study using 3-(4,5-dimethylthiazole-2-yl)-2,5-
304
diphenyltetrazolium bromide (MTT) assay, LP-DTX-FA showed 3.3-fold superior half-
305
maximal inhibitory concentration (IC50) as compared to LP-DTX in high expression of folate
306
receptor alpha (SPCA1) cells. Additionally, during the in-vitro uptake study, LP-DTX-FA
307
showed significantly superior DTX uptake as compared to DTX alone due to conjugated
308
folic-acid. Moreover, during the in-vivo study after intratracheal administration in Sprague
309
Dawley rats, LP-DTX-FA showed 23.39-fold higher DTX deposition within the lung as
310
compared to intravenous LP-DTX-FA at the same dose of 1 mg/kg [71].
311 312
Gemcitabine intercalated liposomes (LP-GB) were formulated using a combination of lipids
313
i.e.
314
phosphoglycerol
315
emulsification solvent evaporation method. Gemcitabine liposomal powder (LP-GB-DPI)
316
was formulated using trehalose (1:2) as a cryoprotectant in the lyophilization technique.
hydrogenated
13 | P a g e
soy
(DSPG),
phosphatidylcholine cholesterol
and
(HSPC),
1,2-Distearoyl-sn-glycero-3-
mPEG2000-DSPE (5:2:2:0.8) by W/O
317
Spherical shape smooth surface LP-GB showed particle size and negative zeta potential of
318
339.49 nm and -53.1 mV, respectively while LP-GB-DPI demonstrated particle size of 8.79
319
µm with satisfactory flow properties. LP-GB-DPI showed satisfactory performance with fine
320
particle fraction (FPF), emitted dose (ED) and mass median aerodynamic diameter (MMAD)
321
of 56.12 %, 88.99 and 3.91 microns, respectively, measured at 60 L/min. LP-GB showed a
322
pH-dependent release profile over a period of 48 h. LP-GB displayed maximum (55.93 %)
323
drug release at pH 5.5 and pH 6.8 (41.45 %) whereas 7.4 (5.61 %) minimum drug release was
324
observed. Release kinetics showed that maximum gemcitabine release from liposome was
325
obtained at endosomal pH (5.5) or cancerous tissue pH (6.5). The reduced drug release at pH
326
7.4 confirmed the stability of liposomes within lung fluid. During the in-vitro toxicity study
327
using MTT assay on human lung adenocarcinoma (A549) cell 3.44-fold improvement in IC50
328
value was observed for LP-GB-DPI as compared to free drug. Moreover, maximum tolerated
329
dose (MTD) and edema index analysis showed retention of alveolar-capillary integrity and
330
negligible infiltration around bronchioles with LP-GB-DPI as compared to drug alone at the
331
same dose. Additionally, during the in-vivo study after intratracheal administration of
332
liposomes, a marked 8.31 and 5.69-fold improvement in area under curve (AUC) and mean
333
residence time (MRT) was observed as compared to drug alone at the same dose (2 mg/kg).
334
In brief, the formulated liposomes showed promising results for the management of lung
335
cancer [72].
336 337
In another study, a respirable lyophilized recombinant secretory leukocyte protease inhibitor
338
(RSLPI) loaded liposomes (RSLPI-DOPS-LP) was formulated using 1,2-dioleoyl-sn-glycero-
339
3-[phospho-L-serine] (DOPS)/ cholesterol (7:3) by thin-film hydration method. Additionally,
340
lyophilized RSLPI-DOPS-LP was micronized in the presence of mannitol (1:10). RSLPI-
341
DOPS-LP exhibited average particle size and EE of 153.60 nm and 74.10 %, respectively
14 | P a g e
342
while lyophilized RSLPI-DOPS-LP showed mean particle size and yield of 19.2 µm and
343
86.70 %, respectively. Micronization showed marked a 12-fold reduction in particle size of
344
lyophilized RSLPI-DOPS-LP. Micronized RSLPI-DOPS-LP powder retained the stability
345
and anti-neutrophil elastase activity of RSLPI without protein degradation as evident from
346
western blot analysis. During the in-vitro aerosolization study using low resistance
347
Spinhaler® device, micronized RSLPI-DOPS-LP powder showed FPF of 38.70 % by twin
348
stage impinger (TSI). In addition, micronized RSLPI-DOPS-LP powder showed FPF and
349
MMAD of 33.30 % and 2.44 µm, respectively, using cascade assembly at a flow rate of 90
350
L/min. This micronized RSLPI-DOPS-LP powder was more stable in terms of liposome size
351
for 5 months at room temperature [73]. In another study, Tang et al (2013) developed a pro-
352
drug oseltamivir phosphate (OP) loaded liposomes (1:10) using ovelecithin and cholesterol
353
by film dispersion method followed by spray drying (LP-OP-DPI). Smooth surface, spherical
354
OP liposomes displayed mean particle size, negative zeta potential and EE of 105.90 nm,-
355
13.65 mV and 60.43 %, respectively. Corrugated nonaggregating LP-OP-DPI showed
356
average particle size and yield of 3.5 µm and 55.37 % respectively, with acceptable flow
357
properties. During the in-vitro deposition study using TSI, LP-OP-DPI showed FPF of 35.4
358
%. In the in-vitro release study LP-OP-DPI displayed sustained release pattern for 20 hr in
359
PBS (pH 7.4) while OP solution showed more than 90 % drug release within the 2 h.
360
Moreover, during the in-vivo study after endotracheal administration, LP-OP-DPI showed a
361
marked 1.14 and 1.22-fold improvement in AUC and maximum drug concentration (Cmax), of
362
oseltamivir carboxylate (OSCA) respectively as compared to orally administered OP solution
363
at the same dose (12 mg/kg). The improved clinical outcome may be attributed to the
364
transformation of OP to OSCA in pulmonary airways by carboxylesterase enzyme [74].
365
15 | P a g e
366
Moxifloxacin loaded nanoliposomes (MFX-NL) were developed by a reversed-phase
367
evaporation method using L-α-phosphatidylcholine (type X-E) and cholesterol (7:3) lipids.
368
MFX-NL was decorated with targeting ligand 4-aminophenyl-alpha-D-manno-pyranoside
369
(PAM) to enhance the uptake of (NL) by alveolar macrophages. Additionally, MFX-NL was
370
embedded into microparticles using dextran as a carrier by spray drying. Corrugated surface
371
and dimple shaped MFX-NL microparticles showed particle size, zeta potential and EE of
372
277 nm, -12.31 mV and 66.25 %, respectively. During the in-vitro aerodynamic study,
373
microparticles showed higher respirable fraction (> 75 %) using the Aerolizer® device. In the
374
in-vitro release study, microparticles showed biphasic release pattern i.e. initial burst release
375
for first 2 h (~ 50 %) followed by controlled release (~ 100 %) up to 48 h in PBS (pH 7.4).
376
Moreover, during the in-vivo deposition study, after intrapulmonary administration (Penn-
377
century®) green fluorescence-labeled microparticles showed sufficiently higher MFX
378
deposition within the alveolar macrophages as compared to the upper respiratory tract.
379
Negatively charged MFX-NL provided higher anti-tubercular activity as compared to the
380
neutral NL whereas PAM decoration was capable enough to augment MFX alveolar delivery.
381
From a pharmacological viewpoint, higher alveolar deposition is important in the treatment
382
of pulmonary tuberculosis. Developed PAM decorated MFX-NL microparticles can be
383
suitable for the treatment of pulmonary tuberculosis and other pulmonary disorders [75].
384 385
Inhalable salbutamol sulphate (SS) loaded liposomal powder (SS-LP-DPI) was formulated
386
using soybean phosphatidylcholine by vesicular phospholipid gel technique followed by
387
lyophilization using lactose (1:5) as a cryoprotectant. In addition, the lyophilised powder was
388
lubricated with 0.5 % magnesium stearate and subjected to ball milling. SS-LP-DPI showed
389
80.71 and 44.35 % EE for SS before lyophilization and after rehydration, respectively.
390
Irregular shaped microparticles showed a marked 4-fold reduction in mean particle size after
16 | P a g e
391
ball milling. In an in-vitro drug release study using a dialysis bag technique SS-LP-DPI
392
displayed sustained release profile up to SS for 24 h in deionized water. During the in-vitro
393
aerodynamic study in TSI assembly using Spinhaler® device, ball-milled SS-LP-DPI
394
exhibited 2.53-fold superior FPF as compared to lyophilized powder. The present
395
investigation highlights the localized pulmonary delivery of liposomes containing anhydrous
396
SS [76]. Honmane et al (2018), prepared and optimized SS loaded liposomal DPI using
397
soybean lecithin and cholesterol (1:1) by thin-film hydration technique followed by spray
398
drying using lactose as a carrier (SS-LM-DPI). Optimized liposomes displayed mean particle
399
size, zeta potential and entrapment efficiency of 167.2 nm, 9.74 mV and 80.68 %,
400
respectively. While SS-LM-DPI showed a mean particle size of 6.35 µm suitable for
401
pulmonary drug delivery. During the in-vitro drug deposition study SS-LM-DPI
402
demonstrated marked 8.92-fold enhancement in FPF as compared to spray dried SS at a flow
403
rate of 60 L/min using the Rotahaler® inhaler device. Moreover, during the in-vitro
404
dissolution study, SS-LM-DPI displayed a controlled release profile for SS (~ 90 %) up to 14
405
hr in PBS (pH 7.4) [77]. Ye et al (2016), designed clarithromycin liposomes using a mixture
406
of soybean phosphatidylcholine: cholesterol: clarithromycin (4:1:2 w/w) by thin lipid film
407
hydration method. Inhalable powder formulation (CLA-LP-DPI) was formulated by
408
ultrasonic spray freeze drying (SFD) techniques using a combination of cryoprotectants i.e.
409
mannitol (15 % w/v) and sucrose (5 % w/v). Spherical shaped, porous CLA-LP-DPI showed
410
high drug recovery (85 %) with suitable particle size. During the in-vitro aerodynamic study
411
at a flow rate of 100 L/min, CLA-LP-DPI showed FPF and ED of 43.82 % and 53.78 %,
412
respectively. Additionally, during a three-month stability study at 25 °C and 60 % relative
413
humidity CLA-LP-DPI didn't show any major change in EE and mean particle size [78].
414
17 | P a g e
415
Curcumin (CUR) intercalated liposomal DPI (CUR-LP-DPI) were prepared using soybean
416
lecithin: cholesterol in the ratio of 5:1 by a film hydration method followed by freeze-drying
417
technique for the treatment of lung cancer. Irregular shaped microparticles showed a mean
418
particle size of 15.02 µm. During aerodynamic assessment CUR-LP-DPI showed FPF and
419
MMAD of 46.71 % and 5.81 µm using next-generation impactor (NGI). In in-vitro
420
cytotoxicity study using normal human bronchial epithelial cell (BEAS-2B) microparticles
421
showed marked 93-fold improvement in selection indices as compared to CUR alone at the
422
same dose (100 µmol/L) while microparticles showed satisfactory in-vitro anti-cancer cell
423
effect against A549 cells. Additionally, uptake of CUR from liposomes by A549 cells was
424
noticeably superior to that of CUR alone. For the in-vivo study, the primary lung cancer
425
model was developed using 3-methylcholanthrene (MCA) and diethylnitrosamine (DEN)
426
inducing agents. After pulmonary administration, CUR-LP-DPI displayed better anti-cancer
427
potential as compared to CUR alone and gemcitabine via regulating enzymatic markers such
428
as tumor necrosis factor-α (TNF-α), vascular endothelial growth factor (VEGF),
429
malondialdehyde (MDA), caspase-3 and B-cell lymphoma 2 protein (BCL-2) [79]. Dry
430
powders of liposomal encapsulated ciprofloxacin nanocrystals (CUR-LP-NC) were fabricated
431
using a freeze-thaw method followed by spray drying. CUR-LP-NC was formulated using
432
HSPC: cholesterol (7:3) and sucrose (2 % w/w) as a cryoprotectant. Corrugated dimple
433
shaped CUR-LP-NC microparticles showed particle size of 1.92 µm. In the in-vitro
434
dissolution study, CUR-LP-NC displayed a controlled release up to 12 h in HEPES (4-(2-
435
hydroxyethyl)-1-piperazineethanesulfonic acid) buffered saline (pH 7.4). During the in-vitro
436
aerodynamic study, CUR-LP-NC displayed higher FPF (69.70 %) as compared to CUR-LP-
437
NC formulated from 0.5 and 1 % w/w sucrose using Osmohaler device® at 105 L/min. The
438
developed formulation can be apt for a once-a-daily treatment schedule [80].
439
18 | P a g e
440
Li et al (2017) developed liposomal andrographolide powder (AG-LP-DPI) using soybean
441
lecithin and cholesterol (6:1) by the solvent injection method with subsequent by freeze-
442
drying using mannitol as a cryoprotectant. AG liposomes showed mean particle size and
443
negative zeta potential of 77.91 nm and -56.13 mV, respectively. While AG-LP-DPI showed
444
irregular and rough surface particles (7.44 µm) suitable for lung deposition. During the in-
445
vitro drug deposition study AG-LP-DPI showed a marked 2.75-fold improvement in FPF as
446
compared to conventional AG-DPI at 60 L/min flow rate using Twister® inhaler device.
447
Moreover, during in-vivo bacterial (S. aureus) study intratracheal administration of AG-LP-
448
DPI (1 mg) showed satisfactory anti-bacterial action at a 10-fold lower dose as compared to
449
AG (10 mg) alone. AG-LP-DPI considerably decreased pro-inflammatory cytokines such as
450
TNF-α, IL-1 and inhibited the phosphorylation of IκB-α in the nuclear factor-κB (NF-κB)
451
pathway (Fig. 3). In summary, phytoconstituent loaded inhaled liposomes are the potential
452
therapeutic agents for pulmonary therapy [81]. In another study, Viswanathan et al (2018),
453
fabricated inhalable liquorice acetone extract loaded liposomes (L-LP-DPI) using soybean
454
phosphatidylcholine by thin-film hydration method with further freeze-drying using trehalose
455
as cryoprotectant and carrier. Unilamellar spherical shaped liposomes showed a mean particle
456
size and EE of 210 nm and 75 %, respectively. During the multistage cascade analysis at 60
457
L/min flow rate using Lupihaler® device, L-LP-DPI showed the FPF, MMAD and geometric
458
standard deviation (GSD) of 54.68 %, 4.29 µm and 1.23 respectively. The in-vivo lung
459
deposition study using the nose-only apparatus in Swiss-albino mice showed more than 46 %
460
of drug deposited by L-LP-DPI in the lungs whereas only 16% of drug retained in the lungs
461
24 h of administration. Moreover, an in-vivo pharmacodynamic study in Mycobacterium
462
tuberculosis H37Rv infected Balb/c mice L-LP-DPI showed a satisfactory reduction in
463
bacterial count in lungs and spleen. In brief, liposomal containing liquorice extract found to
464
be a useful anti-tubercular medicine alone or as an adjunct to the existing standard drugs [82].
19 | P a g e
465 466
Chennakesavulu et al (2017) formulated colchicine (CL) and budesonide (BU) loaded
467
liposomal DPI using thin layer film hydration technique followed by lyophilization for
468
effective treatment of idiopathic pulmonary fibrosis (IPF). CL and BU liposomes were
469
prepared using a combination of various lipids i.e. DPPG: SPC: COL in a ration of 3:6:1 and
470
4:5:1, respectively. CL and BU liposomal DPI using mannitol as a carrier and glycine (10 %)
471
as an anti-adherent was developed. Spherical shaped, CL liposomes showed particle size, zeta
472
potential and drug entrapment of < 100 nm, -24.7 mV and 50.94 %, respectively while BU
473
liposomes showed particle size, zeta potential and drug entrapment of < 100 nm, -36.9 mV
474
and 74.22 %, respectively. During the in-vitro aerosolization study using Rotahaler® device,
475
CL and BU liposomal DPI showed FPF of 44.45and 48.62 %, respectively at a flow rate of
476
28.3 L/min. In-vitro diffusion study for both formulations showed sustained release up to 24
477
h and followed Higuchi’s diffusion-controlled kinetic model in PBS, pH 7.4 containing SLS
478
(1 % w/v). Additionally, during in-vivo studies, in bleomycin (2.5 U/kg) induced IPF rats, CL
479
and BU combined DPI showed 1.17 and 3.53-fold reduction in hydroxyproline content and
480
myeloperoxidase activity showing a positive effect against IPF. Moreover, six-month stability
481
studies at two different conditions i.e. long term (25 °C ± 2 °C, 60 % ± 5 % RH) and
482
refrigerated conditions (2-8 °C) confirmed the stability of lyophilized BU and CL liposomes.
483
Thus, liposomal based BU and CL DPI formulations can be potentially used for the treatment
484
of IPF [83]. Apart from these investigations, various, appealing case studies of inhalable
485
liposomal DPI are summarized in Table 3.
486 487
As summarized in Table 3, liposomes were thoroughly explored and studied as carriers for
488
the inhalation delivery of synthetic drugs [76,77], herbal extracts [82], phytoconstituents,
489
[79,81] vitamins, protein and peptides for the treatment of numerous pulmonary diseases and
20 | P a g e
490
pathological conditions. They were carefully explored for various pulmonary disorders
491
ranging from asthma, COPD to lung cancer. Several well-known and novel processing
492
techniques have been explored to deliver these as dry powder. In various scientific studies,
493
after judiciously selecting phospholipid composition and aerosolization technique, liposomes
494
retain their payload, mean size and do not aggregate after aerosolization. They reveal a
495
superior pulmonary deposition, satisfactory lung retention for a prolonged period after
496
inhalation and delivery of the payload inside the cells [10].
497 498
The drug distribution pattern within the pulmonary airways is highly depended on the size of
499
the aerosolized liposomes rather than vesicle size or type. Among the inhaled liposome
500
formulations, phosphatidylcholine was the most appropriate phospholipid to certify vesicle
501
stability and minimize lipid-drug interactions [91,92]. Normally, the addition of cholesterol in
502
liposomal vesicles enhances in-vivo stability with the decrease in drug release rate. The slow-
503
release pattern of liposomes extends the drug residence time in pulmonary airways and assists
504
the uptake by macrophages [92]. While the inclusion of phosphatidylglycerol assist the
505
spreading of liposomes at the alveolar interface and potentially improve the drug release [93].
506
Multilamellar liposomes are more suitable than unilamellar liposomes for sustained
507
pulmonary drug release. The fusion of lipid vesicles with the alveolar interface and/or lipid
508
exchange between lipid vesicles and pulmonary surfactants has a significant impact on the
509
drug release mechanism [93]. Moreover, liposomes appear to be more suitable for pulmonary
510
application if they are formulated from substances endogenous to the lung i.e. pulmonary
511
surfactant.
512
[dipalmitoylphosphatidylcholine (DPPC), phosphatidylglycerol, phosphatidylcholines (PC)]
513
and proteins where lipids account for >90 % of the surfactant by mass [94-96]. The fate of
514
liposomes deposited in the alveoli is mainly governed by clearance and re-utilization of lung
21 | P a g e
Pulmonary
surfactant
is
a
unique
mixture
of
lipids
515
surfactant. The rate and degree of pulmonary uptake of liposomes are a function of their
516
phospholipid
517
phospholipid showed superior pulmonary uptake [97,98] whereas, macrophage activity
518
played an important role in the clearance mechanism of liposomes [99]. The mean vesicle
519
size of liposome should be less than 400 nm as macrophage uptake of the formulation will
520
increase after 400 nm [100]. Furthermore, neutral or anionic and cationic liposomes showed
521
the different pharmacological outcome. No major adverse effects were found after the
522
delivery of neutral or anionic liposomes. But, cationic liposomes were found to be fatal to
523
human cells and can potentially initiate genetic abnormalities. Furthermore, side effects of
524
cationic liposomes considerably enhanced with an increase of positive charge of the carriers.
525
However, cationic liposomes usually form the almost neutral complexes with negatively
526
charged nucleic acids. Thus such structural modification of cationic carriers usually controls
527
adverse effects on the cells [101]. However, during storage liposomal formulation
528
experienced considerable loss of encapsulated drug and alteration in the physical integrity of
529
the liposomes [102]. To surpass the instability and delivery issues, liposomes can be dried
530
using spray drying, SFD or freeze-drying into proliposomes [63]. Proliposomes are free-
531
flowing particles that instantly form liposomes when in contact with aqueous media [92,103].
532
These delivery systems are thoroughly summarized in the following section.
composition.
In
general,
liposomes
containing
phosphatidylglycerol
533 534
4.2 Proliposomes
535
Proliposomes were initially explored by Payne and his co-workers in 1986 as dry
536
phospholipid formulations with better stability than liposomes [104]. The dry form of
537
proliposomes ease transportation and makes them a functional and efficient delivery system.
538
Proliposomes includes hydrophilic carriers that are layered with cholesterol and
539
phospholipid. They are superior for the entrapment/encapsulation of both hydrophilic as well
22 | P a g e
540
as lipophilic molecules. Later in 1991, the idea of proliposomes was extended to include
541
liquid phospholipid formulations that can produce liposomes upon addition of aqueous
542
medium. Proliposomes can be grouped into two types i.e. particulate-based proliposomes and
543
solvent-based proliposomes. The selection of an apt carrier is a key aspect for the formulation
544
of particulate-based proliposomes. Basically, the carrier is selected on account of its porosity
545
and capacity to hold phospholipids on its surface [105,106]. Whereas, solvent-based
546
proliposomes are fabricated using an organic solvent that dissolves lipids and all at once is
547
miscible with water. In this technique, high concentration of phospholipid in the organic
548
solvent is formulated and the resulting liposomes are collected by dispersion into the aqueous
549
phase. The company of phospholipid, ethanol and water initially produces a stacked bilayer
550
that transforms into liposomes by hydration. Ultimately, proliposomes can be transformed
551
into liposomes by the addition of aqueous phase above the phase transition temperature (Tm)
552
of the lipid followed by continuous shaking. In relation to traditional liposomal formulations,
553
proliposomes reveal more benefits in stability, solubility, drug entrapment and drug release.
554
In addition, production of proliposomes can be scaled up by routinely used techniques such
555
as spray drying, fluidized-bed coating and fluid-energy micronization (jet-milling). Various
556
inhalable proliposomes formulations have been fabricated and explored to augment the
557
aerosolization performance and physicochemical stability of various molecules [68,106].
558
Various studies on inhalable proliposomes formulations are described in the following
559
section.
560 561
Patil-Gadhe et al (2013), fabricated inhalable rifapentine loaded proliposomes for the
562
treatment of pulmonary TB using soy phosphatidylcholine (HSPC) and cholesterol by spray
563
drying and optimized using 32 experimental design. Developed proliposomes showed mean
564
particle size, entrapment efficacy and zeta potential of 578 nm, 72.08 % and 29.40 mV,
23 | P a g e
565
respectively. Smooth spherical, spray-dried rifapentine proliposomes showed satisfactory
566
flow properties. During the in-vitro aerosolization study using Rotahaler®, rifapentine loaded
567
proliposomes showed FPF and MMAD of 92.50 % and 2.62 microns, respectively at a flow
568
rate of 60 L/min. In in-vitro release study rifapentine proliposomes displayed controlled
569
release (~ 90 %) up to 24 h in PBS (pH 7.4). Moreover, during the in-vivo pulmonary
570
pharmacokinetic study after intratracheal administration rifapentine proliposomes displayed
571
13.99, 6.43, 6.40 and 2.35-fold improvement in AUC(0-∞), AUC(0-24), MRT and Cmax,
572
respectively as compared to drug alone at the same dose (250 µg) [107]. Rojanarat et al
573
(2011),
574
phosphatidylcholine and cholesterol (1:1) by spray drying technique with mannitol as an inert
575
carrier. Irregular shaped proliposomes demonstrated FPF and MMAD of 35 % and 2.99
576
microns, respectively, at a flow rate of 60 L/min. During the in-vitro toxicity study, in MTT
577
assay INH-proliposomes did not show any toxicity to pulmonary-associated cells i.e. growth
578
of normal human bronchial epithelial cells (NHBE) and small airway epithelial cells (SAEC).
579
In addition, INH-proliposomes did not activate the alveolar macrophages (AMs) to produce
580
inflammatory mediators i.e. interleukin-1β, tumor necrosis factor-α, and nitric oxide, at a
581
toxic level confirmed from enzyme-linked immunosorbent assay (ELISA) method. Most
582
significantly, INH-proliposomes showed superior anti-mycobacterial activity against M.
583
Bovis-infected AM as compared to INH alone [108].
formulated
inhalable
isoniazid
(INH)
proliposomes
using
soybean
584 585
Also, Rojanarat et al (2012), fabricated inhalable pyrazinamide (PAZ) proliposomes using
586
soybean phosphatidylcholine and cholesterol (1:1) by spray drying method with porous
587
mannitol as a carrier for releasing PAZ to AM infected with mycobacteria. Irregular shaped
588
pro-liposomes demonstrated FPF and MMAD of 29 % and 4.4 microns, respectively, at a
589
flow rate of 60 L/min. During in-vitro toxicity study, in MTT assay using A549, human lung
24 | P a g e
590
cancer cell line cells (Calu-3; as an upper airway cell) and human alveolar basal epithelial
591
cell (NR8383; as a lower airway cell), formulated pro-liposome did not show any toxicity up
592
to 250 µg/mL. Furthermore, formulated proliposome did not activate the AMs to produce
593
inflammatory mediators i.e. interleukin-1β, tumor necrosis factor-α, and nitric oxide, at a
594
toxic level confirmed from ELISA kits. Additionally, in-vivo studies in male Wistar rats after
595
intratracheal administration (4 mg/kg) proliposome did not show any liver or renal toxicity
596
[109]. The same research group also prepared fluoro-quinolone antibiotic (levofloxacin)
597
loaded proliposomes for the treatment of pulmonary TB. Irregular shaped proliposomes
598
showed FPF and ED of 38.10 % and 91.30 % respectively, at a flow rate of 60 L/min. During
599
the in-vitro toxicity study, in MTT assay, levofloxacin pro-liposome showed absences of
600
toxicity on Calu-3, NR8383 up to 5 µg/mL. Moreover, levofloxacin proliposomes could not
601
produce inflammatory mediators and hence, showed marked improvement in the minimum
602
inhibitory concentration value as compared to drug alone [110].
603 604
Proliposomes have illustrated better therapeutic outcomes in the treatment of pulmonary
605
diseases. As summarized in the above section, proliposomes are easily fabricated using a
606
combination of SPC or HSPC and cholesterol with mannitol, porous mannitol or lactose as an
607
inert
608
physicochemical, biological and aerosolization properties of various actives. In comparison to
609
all discussed proliposomes, the HSPC and lactose based proliposome have achieved a greater
610
improvement in FPF (>90%) for rifapentine. Therefore, the combination of a drug molecule
611
with carefully selected lipid moiety, carrier and method may be a perfect move to formulate
612
proliposomes with reasonable aerodynamic properties.
carrier
by
613 614
5. Clinical outcome
25 | P a g e
spray-drying
method.
Spray-dried
proliposomes
could
enhance
615
In the last few decades, formulation scientists have been dynamically exploring the field of
616
liposomes to strengthen the existing standard of therapeutics. This active growth and
617
development in liposomes call for sustained clinical translation and commercialization [111].
618
Even though the preclinical trials have revealed positive effects, there is uncertainty linked to
619
safety in human beings. Thus, it is necessary to conduct a clinical trial to realize the ways in
620
which liposomes interact with the pulmonary airways. Clinical trials are the key building
621
blocks of evidence-based medicine and therefore, nurturing the backbone of clinical practice
622
[112].
623 624
In the past few years, the regulatory bodies have approved some liposomes-based delivery
625
systems and more than 100 different liposomes are in various clinical phases. Thus, in the
626
present segment, we have discussed the liposome-based pulmonary formulation that has
627
reached various phases of clinical trials. Clinical trials related to ‘‘liposomes’’ have been
628
indexed
629
[https://clinicaltrials.gov/]) and European (EU Clinical Trials Register [https://www.clinical
630
trialsregister.eu/]). Clinical trials with inhaled liposomes have been executed for pulmonary
631
tuberculosis, cystic fibrosis, bronchiectasis, fungal infections, lung transplantation, cancer
632
(for osteosarcoma metastatic), analgesia (for post-operative pain), pulmonary moisturizer
633
along with the understanding of safety, efficacy and toxicity. Few remarkable clinical trials
634
are summarized in the following section while clinical efforts invested in inhaled liposomes
635
are listed in Table 4.
and
are
easily
accessible
in
the
US
FDA
(ClinicalTrials.gov
636 637
5.1 Liposomal amikacin (Arikayce®)
638
Insmed Incorporation (New Jersey, US) is a well-known global biopharmaceutical
639
organization with a focus on the design and development of various critical pharmaceutical
26 | P a g e
640
dosage forms. The lipid bilayer of Arikayce® consists of DPPC and cholesterol. The Insmed
641
studied inhaled liposomal amikacin(Arikayce®) to treat mycobacterium infections (phase 3;
642
NCT02344004) [113], cystic fibrosis (phase 3;NCT01316276) [114] and Pseudomonas
643
aeruginosa infection (phase 3; NCT01315678) [115]. The liposomal amikacin was expected
644
to modulate disease conditions. Similarly, Insmed also studied inhaled liposomal cisplatin for
645
the treatment of cancer i.e. osteosarcoma metastatic of lungs (phase 1; NCT00102531) [116].
646
All these studies are near completion and the Insmed is expecting positive outcomes.
647 648
5.2 Liposomal amphotericin B (Ambisome®)
649
Various medical institutes together with pharmaceutical industries investigated inhaled
650
liposomal amphotericin B (Ambisome®) for various pulmonary diseases. Ambisome® (~ 100
651
nm) is composed of amphotericin B, DSPG, hydrogenated soy phosphatidylcholine and
652
cholesterol in a 0.4:0.8:2:1 molar ratio. Interaction between amphotericin B and cholesterol
653
attributed to its sterol binding which is highly responsible for stabilization. Ambisome® was
654
studied for the treatment of allergic bronchopulmonary aspergillosis (phase 2;
655
NCT02273661) [117] and invasive pulmonary aspergillosis (phase 4; NCT00986713) [118].
656 657
5.3 Liposomal phospholipids (LipoAerosol©)
658
The weakening of airways surfaces fluid film (pulmonary surfactant) is one of the main
659
factors responsible for pulmonary disorders (e.g. asthma, COPD and pulmonary edema).
660
Thus, replenishment of pulmonary surfactants through a pharmacological treatment might be
661
an appropriate strategy in the treatment of pulmonary disorders. The lipid vesicles of
662
LipoAerosol® have composed of phospholipids i.e. phosphatidylcholine which is an integral
663
part of the natural pulmonary surfactant. LipoAerosol® provides moistening, warming and
664
cleaning of the upper and lower pulmonary airways as well as assists the natural moistening
27 | P a g e
665
of the film in airway diseases and irritations. Recently, Technische Universität München
666
(Germany)
667
complications (NCT02157129) [119] and hoping for positive results.
has
investigated
LipoAerosol® therapeutic
potential
for
tracheostomy
668 669
5.4 Aerosolized liposomal fentanyl (AeroLEF™)
670
AeroLEF™ (liposome loaded fentanyl) is a new aerosol that offers rapid, extended and
671
personalized analgesia for patients undergoing acute pain episodes. It is designed and
672
developed for the non-invasive route of administration. It can be used for the treatment of
673
moderate to severe pain. AeroLEF™ was assessed in a few clinical trials for controlling pain
674
and post-operative pain (NCT00791804) [120]. Moreover, YM BioSciences Inc. (Canadian
675
drug development company) has studied AeroLEF™ therapeutic potential with four different
676
inhaler devices for device characterization and qualification (NCT00794209) [121].
677 678
The clinical trial database of the US FDA reported no clinical trial on the inhaled liposomes
679
as dry powders. However, an extensive amount of time and effort has been devoted by active
680
scientists and clinicians over the years towards design, development and clinical assessment
681
of inhaled liposomes. All these active efforts signify a bridgehead for the clinical progress of
682
inhaled liposomes. Still, a number of human clinical trials of inhaled liposomes have yet to be
683
performed for a wide range of ailments and better pulmonary therapy.
684 685
6. Market overview
686
The excitement regarding liposomes and liposome-based products has accelerated gradually
687
over the past few years. Liposomes and liposome-based products have demonstrated a
688
beneficial impact on the pharmaceutical market. Inhaled liposomes have offered several key
689
advantages including local as well as systemic delivery, high drug loading capacity,
28 | P a g e
690
controlled release kinetics and good patient compliance. Furthermore, nanocarriers such as
691
liposomes or pro-liposomes also help to extend the patent life and consequently improves the
692
value of drug molecules. For example, the Doxil® (liposomal formulation of doxorubicin) has
693
shown a good impact on cancer management with great benefits for pharmaceutical
694
industries. While, DaunoXome® (liposomal daunorubicin, Galen, Craigavon, UK), Onivyde®
695
(liposomal irinotecan injection, Merrimack Pharmaceuticals, US), DepoCyt® (liposomal
696
cytarabine, Pacira Pharmaceuticals, US),Marqibo® (liposomal vincristine sulfate, Talon
697
Therapeutics, US), AmBisome® (liposomal Amphotericin B, NeXstar Pharmaceuticals, US),
698
Vyxeos® (daunorubicin and cytarabine encapsulated liposomes, Jazz Pharmaceutics, Ireland)
699
and Visudyne® (benzoporphyrin derivative liposomes, QLT Phototherapeutics, Canada) are
700
the known and FDA approved liposomal products [122]. Still, no inhaled liposomal as dry
701
powders were available in the market. However, bench-to-bedside translation of nanocarriers
702
and adopting the same into the mainstream level is often a critical task. There are several
703
chemistry, manufacturing and control (CMC) challenge along with regulatory issues that
704
need to be defeated before it can move on to extensive clinical applications and community
705
acceptance. Thus, by knowing this situation, in the upcoming segment of article authors
706
provide a view on the existing challenges and future directions for liposomes; especially for
707
pulmonary applications.
708 709
7. Discussion
710
Currently, nanocarriers such as liposomes as well as proliposomes are receiving growing
711
interest among formulation scientists for better pulmonary therapy. So far, several
712
formulation scientists and experts have invested their time and efforts in designing,
713
developing and analyzing respirable liposomes and proliposomes for better clinical outcomes.
714
However, investigation in this stage is still in the primitive phase. Many published articles
29 | P a g e
715
have evaluated and discussed particle size, surface charge, in-vitro aerosolization behavior
716
and pulmonary pharmacokinetics. Yet, various other challenges must be addressed when
717
formulating liposomes and proliposomes for inhalation. Several key benefits and challenges
718
of inhaled liposomes are listed in Fig. 4 [123].
719 720
Needless to say, but achieving adequate deep lung deposition and targeting the drug to
721
specific pulmonary airways, is still very tough [68]. Ahead of the complexity of respiratory
722
diseases and lung morphology, it should be kept in mind that disease severity, patient’s age,
723
breathing pattern, device configuration (single dose, multi-dose device) and design features
724
(capsule-based, pre-filed or disposable device) conclude the real fate of aerosol and the final
725
therapeutic outcome of inhaled therapy [68,124,125]. Normally, the successful aerosol
726
delivery depends on four mutually dependent factors: the formulation, the inhaler device
727
configuration, the metering system and lastly, the patient’s training/understanding [124,126].
728
In the above-reviewed articles, most scientists have addressed liposomes and proliposomes
729
manufacturing techniques, drug loading methods, drug targeting strategies and pulmonary
730
pharmacokinetic problems; however, they have merely discussed issues pertain to inhaler
731
devices. Beyond this, there is also a fascinating story to be told about inhaler devices.
732
Furthermore, the device dose metering system has great importance as the dose needed for
733
the effective therapeutic outcome of antimicrobial is too large (e.g. 500 µg) to be inhaled in a
734
single actuation [68]. In such cases, there is a strong need for disposable inhaler device for
735
safe, effective delivery and to avoid bacterial resistance. Basically, to attain satisfactory drug
736
deposition, the development of formulation and inhaler devices should be considered as a
737
whole medicinal product. As per our perceptive strong collaboration between formulation
738
experts and device, engineers is desired to accomplish complex devices/formulation
739
interfaces.
30 | P a g e
740 741
Similarly, to explore the complete therapeutic potential of these multifunctional nanocarriers,
742
exhaustive consideration is required for pulmokinetics, lung clearance rate and other
743
toxicological issues. These issues can be tackled by improving current animal models that can
744
simulate the pathology of the human pulmonary airways [126]. Further, the new imaging
745
tools and techniques can provide superior extrapolative pre-clinical models to understand
746
complex and difficult to treat pulmonary ailments [128-130]. In addition, regarding the
747
situation of respirable liposomes in human clinical trials, there are presently only a few
748
formulations. Hence, up to date data and knowledge of clinical measurements such as forced
749
vital capacity (FVC) and forced expiratory volume (FEV), Tiffeneau-Pinelli index
750
(FEV1/FVC) and maximal voluntary ventilation (MVV) will be crucial in understanding their
751
capability for treating respiratory ailments [68,11]. US FDA has recently published a
752
guidance document i.e. ʻLiposome Drug Productsʼ for a better understanding of the liposomal
753
drug. Likewise, to utilize the full potential of these multifunctional nanocarriers absolute
754
consideration must be paid to upcoming regulatory and CMC guidance. Simply, combined
755
research effort will be highly appreciated to surpass gaps in our understanding with the
756
intention that we can assemble and use all accessible tools and techniques across the pre-
757
clinical-translational-clinical-axis to deliver the best pulmonary dosage to the patient in the
758
most efficient manner.
759 760
8. Conclusion
761
With the growth of nanotechnology and other allied fields, a number of versatile liposomes
762
are designed, developed and investigated as drug delivery carriers. In the vicinity of
763
pulmonary drug delivery, many bioactive molecules have been effectively encapsulated into
764
different liposomal carriers. Additionally, numerous novel strategies have been adopted to
31 | P a g e
765
modify the liposomal drug release pattern, in-vivo drug targeting and biodistribution within
766
the pulmonary airways. The research carried until now proved a great potential for the use of
767
liposomes in the treatment of various intrapulmonary and extrapulmonary diseases. However,
768
it is apparent from the present article that there is a clear need to continue the efforts to
769
design, develop and systematically evaluate the inhaled liposomal formulations. The present
770
situation of inhaled liposomes highly demands formulation scientists to judge long term and
771
structured strategy to pave a way for successful clinical translation, regulatory clearance with
772
FDA approval.
773 774
9. Acknowledgments
775
The author is thankful to Bharati Deemed Vidyapeeth University, Poona College of
776
Pharmacy, Pune-38, India for support and institutional facilities.
777 778
10. Funding Information
779
No financial support and writing assistance was utilized in the production of this manuscript.
780 781
11. Conflicts of Interest
782
Author declare no conflict of interest.
783 784
12. References
785
1. Salvi S, Kumar GA, Dhaliwal RS, Paulson K, Agrawal A, Koul PA et al. The burden of
786
chronic respiratory diseases and their heterogeneity across the states of India: the Global
787
Burden of Disease Study 1990-2016. Lancet Glob Health. 2018;6(12):e1363-e1374.
788
32 | P a g e
789
2. World Health Organization, WHO, Chronic Respiratory Diseases (CRDs): Fact Sheets,
790
(2018) http://www.who.int/respiratory/en/.
791 792
3. Celli BR, Agusti A. COPD: time to improve its taxonomy?. ERJ Open Res. 2018;4(1). pii:
793
00132-2017.
794 795
4. Forum of International Respiratory Societies. The Global Impact of Respiratory Disease-
796
Second Edition. Sheffield, European Respiratory Society, 2017.
797 798
5. Mehta PP. Multi-dose dry powder inhaler: advance technology for drug delivery to
799
airways. Indian Drug, 2019;56 (11):59-62.
800 801
6. Mehta P, Bothiraja C, Kadam S, Pawar A. Effect of USP induction ports, glass sampling
802
apparatus, and inhaler device resistance on aerodynamic patterns of fluticasone propionate-
803
loaded engineered mannitol microparticles. AAPS PharmSciTech. 2019;20(5):197.
804 805
7. Behara SR, Larson I, Kippax P, Stewart P, Morton DA. Insight into pressure drop
806
dependent efficiencies of dry powder inhalers. Eur J Pharm Sci. 2012;46(3):142-148.
807 808
8. Mehta PP. Imagine the superiority of dry powder inhalers from carrier engineering. J Drug
809
Deliv. 2018;2018:5635010.
810 811
9. Mehta PP. Dry powder inhalers: a focus on advancements in novel drug delivery systems. J
812
Drug Deliv. 2016;2016:8290963.
33 | P a g e
813
10. Kuzmov A, Minko T. Nanotechnology approaches for inhalation treatment of lung
814
diseases. J Control Release. 2015;219:500-518.
815 816
11. Mehta P, Kadam S, Pawar A, Bothiraja C. Dendrimers for pulmonary delivery: current
817
perspectives and future challenges. New J. Chem. 2019;43(22):8396-8409.
818 819
12. Aguilera G, Berry C, West R, Gonzalez-Monterrubio E, Angulo-Molina A, Arias-
820
Carrión O et al. Carboxymethyl cellulose coated magnetic nanoparticles transport across a
821
human lung microvascular endothelial cell model of the blood-brain barrier. Nanoscale Adv.
822
2019;1:671-685.
823 824
13. Wongpinyochit T, Totten J, Johnstona B, Seib F. Microfluidic-assisted silk nanoparticle
825
tuning. Nanoscale Adv. 2019;1:873-883.
826 827
14. Abdelaziz HM, Gaber M, Abd-Elwakil MM, Mabrouk MT, Elgohary MM, Kamel NM, et
828
al. Inhalable particulate drug delivery systems for lung cancer therapy: nanoparticles,
829
microparticles, nanocomposites and nanoaggregates. J Control Release. 2018;269:374-392.
830 831
15. Muralidharan P, Malapit M, Mallory E, Hayes Jr D, Mansour HM. Inhalable
832
nanoparticulate powders for respiratory delivery. Nanomedicine. 2015;11(5):1189-1199.
833 834
16. Andrade F, Rafael D, Videira M, Ferreira D, Sosnik A, Sarmento B. Nanotechnology and
835
pulmonary delivery to overcome resistance in infectious diseases. Adv Drug Deliv
836
Rev. 2013;65(13-14):1816-1827.
837
34 | P a g e
838
17. Mehta P, Bothiraja C, Mahadik K, Kadam S, Pawar A. Phytoconstituent based dry
839
powder inhalers as biomedicine for the management of pulmonary diseases. Biomed
840
Pharmacother. 2018;108:828-837.
841 842
18. van Rijt SH, Bein T, Meiners S. Medical nanoparticles for next generation drug delivery
843
to the lungs. Eur Respir J. 2014;44(3):765-774.
844 845
19. Cipolla D, Gonda I, Chan HK. Liposomal formulations for inhalation. Ther
846
Deliv. 2013;4(8):1047-1072.
847 848
20. Jumeaux C, Kim E, Howes P, Kim H, Chandrawati R, Stevens M. Detection of
849
microRNA biomarkers via inhibition of DNA-mediated liposome fusion. Nanoscale Adv.
850
2019;1:532-536.
851 852
21. Willis L, Hayes D, Mansour HM. Therapeutic liposomal dry powder inhalation aerosols
853
for targeted lung delivery. Lung. 2012;190(3):251-262.
854 855
22. Khatib I, Khanal D, Ruan J, Cipolla D, Dayton F, Blanchard JD et al. Ciprofloxacin
856
nanocrystals liposomal powders for controlled drug release via inhalation. Int J
857
Pharm. 2019;566:641-651.
858 859
23. Laverman P, Boerman OC, Oyen WJ, Dams ET, Storm G, Corstens FH. Liposomes for
860
scintigraphic detection of infection and inflammation. Adv. Drug Deliv. Rev. 1999;37:225-
861
235.
862
35 | P a g e
863
24. Bangham AD, Standish MM, Watkins JC. Diffusion of univalent ions across the lamellae
864
of swollen phospholipids. J. Mol. Biol. 1965;13:238-252.
865 866
25. van der Meel R, Fens MH, Vader P, van Solinge WW, Eniola-Adefeso O, Schiffelers
867
RM. Extracellular vesicles as drug delivery systems: lessons from the liposome field. J.
868
Control. Release 2014;195:72-85.
869 870
26. Rideau E, Dimova R , Schwille P , Wurm FR, Landfester K. Liposomes and
871
polymersomes: a comparative review towards cell mimicking. Chem. Soc. Rev.
872
2018;47:8572-8610.
873 874
27. Chorilli M, Calixto G, Rimério TC, Scarpa MV. Caffeine encapsulated in small
875
unilamellar liposomes: characerization and in vitro release profile. J. Dispers. Sci. Technol.
876
2013;34:1465-1470.
877 878
28. Koynova R, Caffrey M. Phases and phase transitions of the phosphatidylcholines.
879
Biochim. Biophys. Acta. 1998;1376:91-145.
880 881
29. Van Tran V, Moon JY, Lee YC. Liposomes for delivery of antioxidants in
882
cosmeceuticals: Challenges and development strategies. J Control Release. 2019;300:114-
883
140.
884 885
30. Chu LY, Utada AS, Shah RK, Kim JW, Weitz DA. Controllable monodisperse multiple
886
emulsions. Angew. Chem. Int. Ed. 2007;46:8970-8974.
887
36 | P a g e
888
31. Aimon S, Manzi J, Schmidt D, Poveda Larrosa JA, Bassereau P, Toombes GE.
889
Functional reconstitution of a voltage-gated potassium channel in giant unilamellar vesicles.
890
PLoS One. 2011;6(10):e25529.
891 892
32. Stachowiak JC, Richmond DL, Li TH, Liu AP, Parekh SH, Fletcher DA. Unilamellar
893
vesicle formation and encapsulation by microfluidic jetting. Proc Natl Acad Sci U S
894
A. 2008;105(12):4697-4702.
895 896
33. Sugiura S, Kuroiwa T, Kagota T, Nakajima M, Sato S, Mukataka S et al. Novel method
897
for obtaining homogeneous giant vesicles from a monodisperse water-in-oil emulsion
898
prepared with a microfluidic device. Langmuir. 2008;24:4581-4588.
899 900
34. Mayer LD, Hope MJ, Cullis PR. Vesicles of variable sizes produced by a rapid extrusion
901
procedure. Biochim Biophys Acta. 1986;858(1):161-168.
902 903
35. Pautot S, Frisken BJ, Weitz DA. Engineering asymmetric vesicles. Proc Natl Acad Sci U
904
S A. 2003;100(19):10718-10721.
905 906
36. Ota S, Yoshizawa S, Takeuchi S. Microfluidic formation of monodisperse, cell‐sized, and
907
unilamellar vesicles. Angew Chem Int Ed Engl. 2009;48(35):6533-6537.
908 909
37. Mazzitelli S, Capretto L, Quinci F, Piva R, Nastruzzi C. Preparation of cell-encapsulation
910
devices in confined microenvironment. Adv Drug Deliv Rev. 2013;65(11-12):1533-1555.
911
37 | P a g e
912
38. Bangham AD, De Gier, Greville G. Osmotic properties and water permeability of
913
phospholipid liquid crystals. Chem. Phys. Lipids. 1967;1:225-246.
914 915
39. Grimaldi N, Andrade F, Segovia N, Ferrer-Tasies L, Sala S, Veciana J, et al. Lipid-based
916
nanovesicles for nanomedicine. ChemSoc Rev. 2016;45(23):6520-6545.
917 918
40. Grijalvo S, Mayr J, Eritja R, Díaz DD. Biodegradable liposome-encapsulated hydrogels
919
for biomedical applications: a marriage of convenience. Biomater Sci. 2016;4(4):555-574.
920 921
41. Szoka J, Papahadjopoulos D. Procedure for preparation of liposomes with large internal
922
aqueous space and high capture by reverse-phase evaporation. Proc. Natl. Acad. Sci. U. S. A.
923
1978;75:4194-4198.
924 925
42. Deamer DW. Preparation and properties of ether injection liposomes. Ann. N. Y. Acad.
926
Sci. 1978;308:250-258.
927 928
43. Jahn A, Vreeland W, Gaitan M, Locascio L. Controlled vesicle self-assembly in
929
microfluidic channels with hydrodynamic focusing. J. Am. Chem. Soc. 2004;126
930
:2674-2675.
931 932
44. van Swaay D, deMello AD. Microfluidic methods for forming liposomes. Lab
933
Chip. 2013;13(5):752-767.
934 935
45. Jaafar-Maalej C, Charcosset C, Fessi H. A new method for liposome preparation using a
936
membrane contactor. J. Liposome Res. 2011;21:213-220.
38 | P a g e
937 938
46. Duong AD, Collier MA, Bachelder EM, Wyslouzil BE, Ainslie KM. One Step
939
Encapsulation of Small Molecule Drugs in Liposomes via Electrospray-Remote Loading.
940
Mol Pharm. 2016;13(1):92-99.
941 942
47. Collier MA, Bachelder EM, Ainslie KM. Electrosprayed Myocet-like Liposomes: An
943
Alternative to Traditional Liposome Production. Pharm Res. 2017;34(2):419-426.
944 945
48. Etheridge ML, Campbell SA, Erdman AG, Haynes CL, Wolf SM, McCullough J. The big
946
picture on nanomedicine: the state of investigational and approved nanomedicine products.
947
Nanomedicine. 2013;9(1):1-14.
948 949
49. Simone EA, Dziubla TD, Muzykantov VR. Polymeric carriers: role of geometry in drug
950
delivery. Expert Opin Drug Deliv. 2008;5(12):1283-1300.
951 952
50. Coune A. Liposomes as drug delivery system in treatment of infectious diseases, potential
953
applications and clinical experience. Infection. 1988;16:141-147.
954 955
51. Plotnick AN. Lipid-based formulations of amphotericin B. J. Am. Vet. Med. Assoc.
956
2000;216:838-841.
957 958
52. de Pauw BE. Fungal infections: diagnostic problems and choice of therapy. Leuk. Suppl.
959
2012;1:S22-23.
960
39 | P a g e
961
53. Woodle MC. Sterically stabilized liposome therapeutics. Adv. Drug Deliver. Rev.
962
1995;16:249-265.
963 964
54. Papahadjopoulos D. Stealth liposomes: from steric stabilization to targeting. CRC Press,
965
Boca Raton FL USA; 1995.
966 967
55. Drummond DC, Meyer O, Hong K, Kirpotin DB, Papahadjopoulos D. Optimizing
968
liposomes for delivery of chemotherapeutic agents to solid tumors. Pharmacol Rev.
969
1999;51(4):691-743.
970 971
56. Mu LM, Ju RJ, Liu R, Bu YZ, Zhang JY, Li XQ, et al. Dual-functional drug liposomes in
972
treatment of resistant cancers. Adv Drug Deliv Rev. 2017;115:46-56.
973 974
57. Nogueira E, Gomes AC, Preto A, Cavaco-Paulo A. Design of liposomal formulations for
975
cell targeting. Colloids Surf B Biointerfaces. 20151;136:514-526.
976 977
58. Marqués-Gallego P, de Kroon AI. Ligation strategies for targeting liposomal
978
nanocarriers. Biomed Res Int. 2014;2014:129458.
979 980
59. Morshed M, Chowdhury E. Gene delivery and clinical applications, Reference module in
981
biomedical sciences, Encyclopedia of Biomedical Engineering. 2019:345-351.
982 983
60. Weber S, Zimmer A, Pardeike J. Solid Lipid Nanoparticles (SLN) and Nanostructured
984
Lipid Carriers (NLC) for pulmonary application: a review of the state of the art. Eur J Pharm
985
Biopharm. 2014;86(1):7-22.
40 | P a g e
986 987
61. Gaspar M, Bakowsky U, Ehrhardt C. Inhaled liposomes-current strategies and future
988
challenges. J. Biomed. Nanotechnol. 2008;4:1-13.
989 990
62. G. Pilcer, K. Amighi, Formulation strategy and use of excipients in pulmonary drug
991
delivery. Int. J. Pharm. 2010;392:1-19.
992 993
63. Misra A, Jinturkar K, Patel D, Lalani J, Chougule M. Recent advances in liposomal dry
994
powder formulations: preparation and evaluation. Expert Opin Drug Deliv. 2009;6(1):71-89.
995 996
65. Willis L, Hayes D, Mansour H. Therapeutic liposomal dry powder inhalation aerosols for
997
targeted lung delivery. Lung. 2012;190:251-262.
998 999
66. Sanders M. Inhalation therapy: an historical review. Prim Care Respir J. 2007;16:71-81.
1000 1001
67. Skupin Mrugalska P. Liposome-based drug delivery for lung cancer, in Kesharwani P
1002
(Eds.), Nanotechnology-based targeted drug delivery systems for lung cancer, Academic
1003
Press, 2019;123-160.
1004 1005
68. Mehta P, Bothiraja C, Kadam S, Pawar A. Potential of dry powder inhalers for
1006
tuberculosis therapy: facts, fidelity and future. Artif Cells Nanomed Biotechnol.
1007
2018;46(sup3):S791-S806.
1008 1009
69. Zhou Q, Tang P, Leung S, et al. Emerging inhalation aerosol devices and strategies:
1010
where are we headed? Adv Drug Deliv Rev. 2014;75:3-17.
41 | P a g e
1011 1012
70. Tagami T, Ozeki T. Recent Trends in Clinical Trials Related to Carrier-Based Drugs. J
1013
Pharm Sci. 2017;106(9):2219-2226.
1014 1015
71. Zhu X, Kong Y, Liu Q, Lu Y, Xing H, Lu X, et al. Inhalable dry powder prepared from
1016
folic acid-conjugated docetaxel liposomes alters pharmacodynamic and pharmacokinetic
1017
properties relevant to lung cancer chemotherapy. Pulm Pharmacol Ther. 2019;55:50-61.
1018 1019
72. Gandhi M, Pandya T, Gandhi R, Patel S, Mashru R, Misra A, Tandel H. Inhalable
1020
liposomal dry powder of gemcitabine-HCl: Formulation, in vitro characterization and in vivo
1021
studies. Int J Pharm. 2015;496(2):886-895.
1022 1023
73. Gibbons A, McElvaney NG, Cryan SA. A dry powder formulation of liposome-
1024
encapsulated recombinant secretory leukocyte protease inhibitor (rSLPI) for inhalation:
1025
preparation and characterisation. AAPS PharmSciTech. 2010;11(3):1411-1421.
1026 1027
74. Tang Y, Zhang H, Lu X, Jiang L, Xi X, Liu J, Zhu J. Development and evaluation of a
1028
dry powder formulation of liposome-encapsulated oseltamivir phosphate for inhalation. Drug
1029
Deliv. 2015;22(5):608-618.
1030 1031
75. Hamed A, Osman R, Al-Jamal KT, Holayel SM, Geneidi AS. Enhanced antitubercular
1032
activity, alveolar deposition and macrophages uptake of mannosylated stable nanoliposomes.
1033
J Drug Deliv Sci Technol. 2019;51:513-523.
1034
42 | P a g e
1035
76. Huang WH, Yang ZJ, Wu H, Wong YF, Zhao ZZ, Liu L. Development of liposomal
1036
salbutamol sulfate dry powder inhaler formulation. Biol Pharm Bull. 2010;33(3):512-517.
1037 1038
77. Honmane S, Hajare A, More H, Osmani RAM, Salunkhe S. Lung delivery of
1039
nanoliposomal salbutamol sulfate dry powder inhalation for facilitated asthma therapy. J
1040
Liposome Res. 2019;29(4):332-342.
1041 1042
78. Ye T, Yu J, Luo Q, Wang S, Chan H. Inhalable clarithromycin liposomal dry powders
1043
using ultrasonic spray freeze drying. Powder Technol. 2017;305:63-70.
1044 1045
79. Zhang T, Chen Y, Ge Y, Hu Y, Li M, Jin Y. Inhalation treatment of primary lung cancer
1046
using liposomal curcumin dry powder inhalers. Acta Pharm Sin B. 2018;8(3):440-448.
1047 1048
80. Khatib I, Khanal D, Ruan J, Cipolla D, Dayton F, Blanchard J, Chan H. Ciprofloxacin
1049
nanocrystals liposomal powders for controlled drug release via inhalation. Int J Pharm.
1050
2019;566:641-651.
1051 1052
81. Li M, Zhang T, Zhu L, Wang R, Jin Y. Liposomal andrographolide dry powder inhalers
1053
for treatment of bacterial pneumonia via anti-inflammatory pathway. Int J Pharm.
1054
2017;528(1-2):163-171.
1055 1056
82. Viswanathan V, Pharande R, Bannalikar A, Gupta P, Gupta U, Mukne A. Inhalable
1057
liposomes of Glycyrrhiza glabra extract for use in tuberculosis: formulation, in
1058
characterization, in vivo lung deposition, and in vivo pharmacodynamic studies. Drug Dev
1059
Ind Pharm. 2019;45(1):11-20.
43 | P a g e
vitro
1060 1061
83. Chennakesavulu S, Mishra A, Sudheer A, Sowmya C, Reddy C, Bhargav E. Pulmonary
1062
delivery of liposomal dry powderinhaler formulation for effective treatment ofidiopathic
1063
pulmonary fibrosis. Asian J. Pharm. Sci. 2018;13:91-100.
1064 1065
84. Ourique AF, Chaves Pdos S, Souto GD, Pohlmann AR, Guterres SS, Beck RC.
1066
Redispersible
1067
development, in vitro characterization and antioxidant activity. Eur J Pharm Sci.
1068
2014;65:174-182.
liposomal-N-acetylcysteine
powder
for
pulmonary
administration:
1069 1070
85. Khatib I, Khanal D, Ruan J, Cipolla D, Dayton F, Blanchard JD, Chan HK. Ciprofloxacin
1071
nanocrystals liposomal powders for controlled drug release via inhalation. Int J Pharm.
1072
2019;566:641-651.
1073 1074
86. Daman Z, Gilani K, Najafabadi A, Eftekhariand H, Barghi M. Formulation of inhalable
1075
lipid-based salbutamol sulfatemicroparticles by spray drying technique. Daru. 2014; 22(1):
1076
50.
1077 1078
87. Joshi M, Misra A. Dry powder inhalation of liposomal Ketotifen fumarate: formulation
1079
and characterization. Int J Pharm. 2001;223(1-2):15-27.
1080 1081
88. Chougule M, Padhi B, Misra A. Development of spray dried liposomal dry powder
1082
inhaler of Dapsone. AAPS PharmSciTech. 2008;9(1):47-53.
1083
44 | P a g e
1084
89. Chougule M, Padhi B, Misra A. Nano-liposomal dry powder inhaler of tacrolimus:
1085
preparation, characterization, and pulmonary pharmacokinetics. Int J Nanomedicine.
1086
2007;2(4):675-688.
1087 1088
90. Changsan N, Chan HK, Separovic F, Srichana T. Physicochemical characterization and
1089
stability of rifampicin liposome dry powder formulations for inhalation. J Pharm Sci.
1090
2009;98(2):628-639.
1091 1092
91. Giovagnoli S, Blasi P, Vescovi C, Fardella G, Chiappini I, Perioli L, Ricci M, Rossi C.
1093
Unilamellar
1094
physicochemical characterization. AAPS PharmSciTech. 2004;4(4):E69.
vesicles
as
potential
capreomycin
sulfate
carriers:
preparation
and
1095 1096
92. Parumasivam T, Chang RY, Abdelghany S, Ye TT, Britton WJ, Chan HK. Dry powder
1097
inhalable formulations for anti-tubercular therapy. Adv Drug Deliv Rev. 2016;102:83-101.
1098 1099
93. Taylor KM, Newton JM. Liposomes for controlled delivery of drugs to the lung. Thorax.
1100
1992 Apr;47(4):257-259.
1101 1102
94. King RJ. Isolation and chemical composition of pulmonary surfactant, in Robertson B,
1103
Van Golde LMG, Batenburg JJ (Eds.), Pulmonary surfactant, Amstedam, Elsevier, 1984:1-
1104
15.
1105 1106
95. Wright JR, Clements JA. Metabolism and turnover of lung surfactant. Am Rev Respir Dis
1107
1987;135:426-444.
1108
45 | P a g e
1109
96. Lopez-Rodriguez E, Pérez-Gil J. Structure-function relationships in pulmonary surfactant
1110
membranes: from biophysics to therapy. Biochim Biophys Acta. 2014;1838(6):1568-1585.
1111 1112
97. Oyarzun MJ, Clements JA, Baritussio A. Ventilation enhances the pulmonary alveolar
1113
clearance of radioactive dipalmitoylphosphatidylcholine in liposomes. Am Rev Respir Dis
1114
1980;121:709-721.
1115 1116
98. Morimoto Y, Adachi Y. Pulmonary uptake of liposomal phosphatidylcholine upon
1117
intratrachealadministation to rats. Chem Pharmaceut Bull 1982;30:2248-2251.
1118 1119
99. Martin WJ, Standing JE. Amiodarone pulmonary toxicity: biochemical evidence for a
1120
cellular phospholipidosis in the bronchoalveolar lavage of human subjects. J Pharmacol
1121
ExpTher 1988;244:774-779.
1122 1123
100. Geiser M, Casaulta M, Kupferschmid B, Schulz H, Semmler-Behnke M, Kreyling W.
1124
The role of macrophages in the clearance of inhaled ultrafine titanium dioxide particles. Am J
1125
Respir Cell Mol Biol. 2008;38(3):371-376.
1126 1127
101. Shah V, Taratula O, Garbuzenko OB, Patil ML, Savla R, Zhang M, Minko T.
1128
Genotoxicity of different nanocarriers: possible modifications for the delivery of nucleic
1129
acids. Curr Drug Discov Technol. 2013;10(1):8-15.
1130 1131
102. Cipolla D, Gonda I, Chan HK. Liposomal formulations for inhalation. Ther Deliv.
1132
2013;4(8):1047-1072.
1133
46 | P a g e
1134
103. Agnihotri SA, Soppimath KS, Betageri GV. Controlled release application of
1135
multilamellar vesicles: a novel drug delivery approach. Drug Deliv. 2010;17(2):92-101.
1136 1137
104. Payne NI, Timmins P, Ambrose CV, Ward MD, Ridgway F. Proliposomes: a novel
1138
solution to an old problem. J Pharm Sci 1986;75:325-329.
1139 1140
105. Elhissi A, Phoenix D, Ahmed W. Some approaches to large-scale manufacturing of
1141
liposomes, in Ahmed W and Jackson MJ (Eds.), Emerging Nanotechnologies for
1142
Manufacturing, 2nd Eds, William Andrew Publishing, 2015: 402-417.
1143 1144
106. Khan I, Elhissi A, Shah M, Alhnan MA, Ahmed W. Liposome-based carrier systems and
1145
devices used for pulmonary drug delivery, in Davim JP (Eds.), Biomaterials and Medical
1146
Tribology, Woodhead Publishing, 2013:395-443.
1147 1148
107. Patil-Gadhe A, Pokharkar V. Single step spray drying method to develop proliposomes
1149
for inhalation: a systematic study based on quality by design approach. Pulm Pharmacol
1150
Ther. 2014;27(2):197-207.
1151 1152
108. Rojanarat W, Changsan N, Tawithong E, Pinsuwan S, Chan HK, Srichana T. Isoniazid
1153
proliposome powders for inhalation-preparation, characterization and cell culture studies. Int
1154
J Mol Sci. 2011;12(7):4414-4434.
1155 1156
109.
Rojanarat
1157
Inhaled pyrazinamide proliposome for targeting alveolar macrophages. Drug Deliv. 2012;
1158
19(7):334-345.
47 | P a g e
W, Nakpheng
T, Thawithong
E, Yanyium
N, Srichana
T.
1159 1160
110. Rojanarat W, Nakpheng T, Thawithong E, Yanyium N, Srichana T. Levofloxacin-
1161
proliposomes: opportunities for use in lung tuberculosis. Pharmaceutics. 2012;4(3):385-412.
1162 1163
111. Hassan S, Prakash G, Ozturk AB, Saghazadeh S, Sohail MF, Seo J, et al. Evolution and
1164
clinical translation of drug delivery nanomaterials. Nano Today. 2017;15:91-106.
1165 1166
112. Masic I, Miokovic M, Muhamedagic B. Evidence based medicine–new approaches and
1167
challenges. Acta Informatica Medica. 2008;16(4):219.
1168 1169
113. Study to Evaluate Efficacy of LAI When Added to Multi-drug Regimen Compared to
1170
Multi-drug
1171
https://clinicaltrials.gov/ct2/show/NCT02344004?term=NCT02344004&rank=1.
Regimen
Alone
(CONVERT).
1172 1173
114. Extension Study of Liposomal Amikacin for Inhalation in Cystic Fibrosis (CF) Patients
1174
With
1175
https://clinicaltrials.gov/ct2/show/NCT01316276?term=NCT01316276&rank=1
Chronic
Pseudomonas
Aeruginosa
(Pa)
Infection.
1176 1177
115. Study to Evaluate Arikayce™ in CF Patients With Chronic Pseudomonas Aeruginosa
1178
Infections.
1179
https://clinicaltrials.gov/ct2/show/NCT01315678?term=NCT01315678&rank=1
1180 1181
116. Inhalation SLIT Cisplatin (Liposomal) for the Treatment of Osteosarcoma Metastatic to
1182
the Lung. https://clinicaltrials.gov/ct2/show/NCT00102531?term=NCT00102531&rank=1
1183
48 | P a g e
1184
117. Evaluation of a Therapeutic Strategy Including Nebulised Liposomal Amphotericin B
1185
(Ambisome®) in Maintenance Treatment of Allergic BronchopulmonaryAspergillosis
1186
(Cystic Fibrosis Excluded). (NEBULAMB)
1187
https://clinicaltrials.gov/ct2/show/NCT02273661?term=NCT02273661&rank=1
1188 1189
118. Value of Amphotericin B Inhalation for Prophylaxis of Invasive Pulmonary
1190
Aspergillosis
1191
Transplantation.https://clinicaltrials.gov/ct2/show/NCT00986713?term=NCT00986713&rank
1192
=1
After
Renal
1193 1194
119.LipoAerosol©
1195
Tracheostomy.https://clinicaltrials.gov/ct2/show/NCT02157129?term=NCT02157129&rank=
1196
1
Inhalation
After
1197 1198
120. Phase II Study of Inhaled AeroLEF for Analgesia After ACL Knee Surgery (Pain).
1199
https://clinicaltrials.gov/ct2/show/NCT00791804?term=NCT00791804&rank=1.
1200 1201
121. Study Evaluating Inhaled AeroLEF Delivered in 4 Aerosol Delivery Devices in Healthy
1202
Volunteers
1203
https://clinicaltrials.gov/ct2/show/NCT00794209?term=NCT00794209&rank=1.
(LEF-07).
1204 1205
122.
Farjadian
1206
Nanopharmaceuticals and
1207
opportunities. Nanomedicine (Lond). 2019;14(1):93-126.
1208
49 | P a g e
F, Ghasemi
A, Gohari
O, Roointan
nanomedicines currently on
A, Karimi
M, Hamblin
MR.
the market: challenges and
1209
123. Limeres MJ, Moretton MA, Bernabeu E, Chiappetta DA, Cuestas ML. Thinking small,
1210
doing big: Current success and future trends in drug delivery systems for improving cancer
1211
therapy with special focus on liver cancer. Mater Sci Eng C Mater Biol Appl. 2019;95:328-
1212
341.
1213 1214
124. Giovagnoli S, Schoubben A, Ricci M. The long and winding road to inhaled TB therapy:
1215
not only the bug’s fault. Drug Dev. Ind. Pharm. 2017;43(3):347-363.
1216 1217
125. Mehta PP, Kadam SS, Pawar AP. Influence of modified induction port, modified DUSA
1218
assembly and device air-inlet geometry on the aerosolization pattern of a dry powder inhaler.
1219
J Drug Deliv Sci Technol. 2020;55:101416.
1220 1221
126. Rogueda P, Traini D. The future of inhalers: how can we improve drug delivery in
1222
asthma and COPD? Expert Rev Respir Med. 2016; 10:1041-1044.
1223 1224
127. Guillon A, Sécher T, Dailey LA, Vecellio L, de Monte M, Si-Tahar M et al. Insights on
1225
animal models to investigate inhalation therapy: relevance for biotherapeutics. Int J
1226
Pharm. 2018;536(1):116-126.
1227 1228
128. Vulović A, Šušteršič T, Cvijić S, Ibrić S, Filipović N. Coupled in silico platform:
1229
Computational fluid dynamics (CFD) and physiologically-based pharmacokinetic (PBPK)
1230
modelling. Eur J Pharm Sci. 2018;113:171-184.
1231
50 | P a g e
1232
129. Backman P, Arora S, Couet W, Forbes B, de Kruijf W, Paudel A. Advances in
1233
experimental and mechanistic computational models to understand pulmonary exposure to
1234
inhaled drugs. Eur J Pharm Sci. 2018;113:41-52.
1235 1236
130. Mehta PP. Dry powder inhalers: upcoming platform technologies for formulation
1237
development. Ther Deliv. 2019 Sep;10(9):551-554.
1238 1239 1240 1241 1242 1243 1244 1245 1246 1247 1248 1249 1250 1251 1252 1253 1254 1255 1256
51 | P a g e
1257
Figure captions
1258 1259
Graphical Abstract - Applications and interaction of liposomes with the pulmonary
1260
membrane. Liposomes are synthesized from substances endogenous to pulmonary airways
1261
such as pulmonary surfactants and hence, explored in the pulmonary drug delivery field
1262
owing to their versatile attributes. By interacting with the pulmonary membrane,
1263
multifunctional liposomes can adhere to the mucosal surface for a longer time, open
1264
intercellular tight junctions in pulmonary epithelia along with modified drug release kinetics
1265
and reduced dosing frequency to improve therapeutic outcomes.
1266 1267
Figure 1 - Liposome types based on lamellarity and size. SUV: small unilamellar vesicles,
1268
LUV: large unilamellar vesicles, GUV: giant unilamellar vesicles, MLV: multilamellar
1269
vesicles and MVV: multivesicular vesicles (vesosomes). Moreover, a fraction of classic lipid
1270
bilayer with hydrophilic head groups and hydrophobic alkyl chains shown.
1271 1272
Figure 2 - Applications and interaction of liposomes with the pulmonary membrane.
1273
Liposomes are synthesized from substances endogenous to pulmonary airways such as
1274
pulmonary surfactants and hence, explored in the pulmonary drug delivery field owing to
1275
their versatile attributes. By interacting with the pulmonary membrane, multifunctional
1276
liposomes can adhere to the mucosal surface for a longer time, open intercellular tight
1277
junctions in pulmonary epithelia along with modified drug release kinetics and reduced
1278
dosing frequency to improve therapeutic outcomes. (MRT: mean resisdance time; Cmax: the
1279
maximum concentration of the drug achieved in the plasma and PEG: polyethylene glycol).
1280
52 | P a g e
1281
Figure 3 -Graphical representation depicting the mechanism of action for liposomal
1282
andrographolide DPI used in regulating immune responses (Lie et al., 2017) [81]. The figure
1283
showed that liposomal andrographolide DPI regulates immune responses by downregulating
1284
various inflammatory pathways such as inhibit phosphorylation of kappa B (IκB-α) inhibitor
1285
and reduced the expression of intercellular adhesion molecule (ICAM) family 1, Interferon-γ
1286
(IFN-γ) in immunocytes.
1287 1288
Figure 4 -Benefits and challenges of multifunctional inhaled liposomes. The figure showed
1289
crucial technical, biological and formulations benefits as well as challenges related to the
1290
design and development of multifunctional inhaled liposomes. This figure emphasis various
1291
key factors such as chemistry, manufacturing and control (CMC), center for devices and
1292
radiological health (CDRH), forced vital capacity (FVC) and forced expiratory volume
1293
(FEV). (IPR: intellectual property rights, ADME: absorption, distribution, metabolism, and
1294
excretion and MRT: mean resisdance time).
53 | P a g e
Table 1 - Advantages and disadvantages of liposome preparation methods Preparation
Particle size Advantages
method
(nm)
Disadvantages
References
Conventional method Thin-lipid
film 100-1000
hydration
A
simple,
conventional Low yield, difficult to scale up, low [38-40]
process
encapsulation
efficacy,
heterogeneous
particle size Reverse-phase
100-1000
High entrapment efficacy
Organic
evaporation Ethanol
solvent
traces,
difficult
to [39-41]
manufacture at large scale injection 70-200
Ability to control vesicle size
method
Reproducibility, difficult to remove ethanol [39,40,42] traces, heterogeneous particle size, require high temperature
Novel methods Microfluidic technology
100-300
High
entrapment
suitable homogenous
1|Page
for
efficacy, Prerequisites
of
equipment,
various [29,40,
scale-up, processing conditions; thus need more 43,44]
particle
size, optimization
appropriate
for
template-
based manufacturing Membrane
~ 100
contactor
Homogenous high
particle
entrapment
size, Encapsulation requires optimization
[40,45]
efficacy,
suitable for scale up Electrospray technology
100-500
Continuous
method,
encapsulation efficiency
high Prerequisites processing optimization
2|Page
of
equipment,
parameters;
thus
various [29,46,47] critical
Table 2 - Advantages and limitations of novel carrier-based pulmonary drug delivery system Advantages
Limitations
Protection from biological environment
Complex and multi-step process
(enzymes, hydration, pH variation) Masking of immunogenicity resulting from API
High cost
Both hydrophilic and hydrophobic API can be easily loaded
Complex control of reproducibility i.e. particle size distribution and/or surface charge
Controlled and modified drug release
Need to conduct toxicological studies
Triggered released in response to physical or chemical different Need to gather and understand ADME pattern stimuli (pH/temperature) API: active pharmaceutical ingredient; ADME: absorption, distribution, metabolism and elimination
3|Page
Table 3- Developed liposomal formulations for pulmonary delivery Drug
Method Key ingredient
Particle
size Device
FPF (%)
References
(nm) SS
FD
SPC, α-LM and MgSt (0.5 %)
137.00 (LUVs)
Spinhaler®
41.51
[76]
Docetaxel
SD
PC, CHOL, mannitol and leucine
346.80 (LUVs)
Reusable
10.10
[71]
device N-acetylcysteine
SD
SPC, CHOL and α-LM
117.00 (LUVs)
HandiHaler®
35.34
[84]
Ciprofloxacin
SD
HSPC, CHOL and sucrose
131.30 (LUVs)
Osmohaler®
69.70
[85]
Liquorice extract FD
SPC, CHOL and trehalose
212.60 (LUVs)
Lupihaler®
54.68
[82]
Andrographolide
FD
Soybean lecithin, CHOL and mannitol
77.91 (SUVs)
Twister®
23.03
[81]
SS
SD
SPC, CHOL and α-LM
167.20 (LUVs)
Rotahaler®
64.01
[77]
SS
SD
DPPC, CHOL and α-LM
-
Cyclohaler®
42.70
[86]
Ketotifen
FD
EPC, CHOL and sucrose
-
Rotahaler®
21.59
[87]
Dapsone
SD
DPPC, CHOL and α-LM
137.00 (LUVs)
-
75.60
[88]
Clarithromycin
FD
SPC, CHOL, mannitol (15 %) and 370.00 (LUVs)
-
53.78
[78]
fumarate
4|Page
sucrose (5 %) Gemcitabine
FD
HSPC, DSPG, CHOL mPEG2000- 331.42 (LUVs)
-
56.12
[72]
DSPE and trehalose Curcumin
FD
SPC, CHOL and mannitol
94.65 (SUVs)
-
46.71
[79]
Oseltamivir
SD
Ovelecithin, CHOL and leucine
105.90 (LUVs)
-
35.40
[74]
Tacrolimus
SD
HSPC, CHOL and trehalose
140.00 (LUVs)
-
71.10
[89]
Rifampicin
FD
SPC, CHOL and mannitol
255.00 (LUVs)
-
66.80
[90]
Moxifloxacin
SD
PC, CHOL and dextran
272.00 (LUVs)
-
72.08
[75]
phosphate
SS: salbutamol sulphate; SD: spray drying; FD: freeze drying; SPC: Soya phosphatidylcholine; CHOL: cholesterol; HSPC: hydrogenated soybean
phosphatidylcholine;
DSPG:
1,2-Distearoyl-sn-glycero-3-phosphoglycerol;
dipalmitoylphosphatidylcholine and EPC: egg phosphatidylcholine
5|Page
PC:
phosphatidylcholine;
DPPC:
Table 4 - A summary of ongoing clinical trials for liposomal formulations Product
Condition or disease
Intervention/ treatment
Phase
Identifier
Sponsor
Arikayce™
Cystic fibrosis
280 mg of matching placebo
I and II
NCT00777296
Insmed Incorporated., US
Arikayce™
Cystic fibrosis
70 mg; 140 mg and 560 mg
I and II
NCT00558844
Insmed Incorporated., US
Cisplatin
Osteosarcoma
24 mg/m2 once daily
I and II
NCT00102531
Insmed Incorporated., US
liposomes
metastatic
for 14 days
Cyclosporine
Bronchiolitis obliterans
2.5 mg/10 mg x 2/day
I and II
NCT01334892
Pari
liposomes
for 96 weeks
Cyclosporine
Lung
liposomes
and
transplantation -
Pharma
GmbH,
Germany I and II
NCT01650545
bronchiolitis
University of Maryland, US
obliterans Arikayce™
Cystic fibrosis
560 mg once daily dose for 6 II
NCT03905642
Insmed Incorporated., US
NCT03038178
Kevin Winthrop, Insmed
cycles over 18 months Arikayce™
MBI and NMBI
590 mg for 12 months
II
Incorporated and etc. Ambisome®
6|Page
Allergic
25 mg x 1/ week for 6 months
II
NCT02273661
Poitiers
University
bronchopulmonary
Hospital, France
aspergillosis Ambisome®
Lung transplantation
-
II
NCT01254708
University
Health
Network, Toronto Liposomal Nitro-20
9- Lung cancer (S)-
5 consecutive days per week II
NCT00250120
X 8 weeks, every 10 weeks
University
of
New
Mexico, US
camptothecin Amikacin
MBI and NMBI
590 mg/day for 12 months
II
NCT03038178
liposomes
Kevin Winthrop and Insmed Incorporated., US and etc.
Arikayce™
MBI and NMBI
590 mg administered
III
NCT02344004
Insmed Incorporated., US
III
NCT03270514
Kathirvel Subramaniam
III
NCT02628600
Insmed Incorporated., US
once daily Bupivacaine
Coronary artery disease
liposomes Liposomal amikacin
7|Page
Bupivacaine liposomes 226 mg
Non-tuberculous for infections
590 mg/day
inhalation Ambisome®
Lung
transplantation 1 mg/kg/day for 4 days
III
NCT00177710
and fungal infections
University of Pittsburgh and Astellas Pharma US, Inc.
Ciprofloxacin
Non-cystic
fibrosis -
liposomes
bronchiectasis
III
NCT01515007
Grifols
(ORBIT-3) Cyclosporine
Bronchiolitis obliterans
Twice daily inhalation for a III
NCT01439958
maximum of three years
Ambisome®
Chronic
with
aspergillosis
for
24
weeks)
Pari
Pharma
GmbH,
Germany
pulmonary Ambisome (25 mg x 2/week III
itraconazole
NCT03656081
and
Poitiers
University
Hospital, France
itraconazole (200 mg x 2/day)
Ciprofloxacin
Non-cystic
liposomes
bronchiectasis
fibrosis -
III
NCT02104245
Aradigm Corporation Grifols
(ORBIT-4)
8|Page
Therapeutics
LLC
liposomes
Liposomal
Aradigm Corporation and
Therapeutics
LLC Lung cancer
-
-
NCT00277082
University
of
New
camptothecin LipoAerosol®
Mexico, US Tracheostomy complications
5x/d for 30 min
-
NCT02157129
Technical University of Munich, Germany
ArikayceTM : amikacin liposome inhalation suspension; Ambisome ®: amphotericin B liposomal; MBI: mycobacterial infection and NMBI: nonmycobacterial infection.
9|Page
AUTHOR DECLARATION TEMPLATE
We wish to draw the attention of the Editor to the following facts which may be considered as potential conflicts of interest and to significant financial contributions to this work. We confirm that the manuscript has been read and approved by all named authors and that there are no other persons who satisfied the criteria for authorship but are not listed. We further confirm that the order of authors listed in the manuscript has been approved by all of us. We confirm that we have given due consideration to the protection of intellectual property associated with this work and that there are no impediments to publication, including the timing of publication, with respect to intellectual property. In so doing we confirm that we have followed the regulations of our institutions concerning intellectual property. We understand that the Corresponding Author is the sole contact for the Editorial process (including Editorial Manager and direct communications with the office). He/she is responsible for communicating with the other authors about progress, submissions of revisions and final approval of proofs. We confirm that we have provided a current, correct email address which is accessible by the Corresponding Author and which has been configured to accept email from [email protected]; [email protected]
Corresponding author; Dr. Vividha Dhapte-Pawar, Associate Professor, Department of Pharmaceutics, Bharati Vidyapeeth University, Poona College of Pharmacy, Pune -411038 Email: [email protected]; [email protected]