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Alternative formulations of paclitaxel
Cancer
Treatment
Alternative
Reviews
(1997)
23, 87-95
formulations
J. M. Meerum Terwogt*,t, J. H. Beijnen*,t,*
of paclitaxel
B. Nuijent,
W. W. Ten Bokkel Huinink*
and
* Department of Medical Oncology, The Netherlands Cancer Institute/ Antoni van Leeuwenhoek Hospital, Amsterdam, The Netherlands t Department of Pharmacy and Pharmacology, Slotervaatt Hospital/The Netherlands Cancer Institute, Amsterdam, The Netherlands $ Department of Pharmaceutical Analysis and Toxicology, Faculty of Pharmacy, State University of Utrecht, The Netherlands
Abstract Paclitaxel, a novel antitumour agent, is active clinically against advanced ovarian and breast cancer and under investigation for various other cancers. One of the problems associated with the intravenous administration of paclitaxel is its low solubility in water. The current pharmaceutical formulation consists of a 1:l (v/v) mixture of ethanol and Cremophor EL. This formulation, however, has been demonstrated to cause some severe hypersensitivity reactions. Therefore the development of a safer intravenous formulation devoid of Cremophor EL is an important investigational issue. This review deals with some of the most promising formulation alternatives.
Introduction Paclitaxel (TaxolB), a natural diterpene product isolated from the bark of Taxus brevifolia, has been shown to be highly cytotoxic and is clinically active against advanced ovarian and breast cancer. It has a unique mechanism of action, inducing the assembly of stable microtubules and inhibiting the depolymerization process (1). In spite of its promising antitumour activity, the drug has presented considerable difficulties related to its intravenous administration to patients.
Correspondence of Pharmacy and 0306-7372/97/020087
should be addressed to: J.M. Meerum Terwogt, Slotervaart Pharmacology, Louwesweg 6, 1066 EC, Amsterdam, The + 09 $12.00/O
0 1997 87
W.B.
Hospital, Netherlands. Saunders
Department
Company
Ltd
J. M. MEERUM
TERWOGT
HAL.
-N&Ho1; .&AH
Figure
Table
1.
Approximate
solubilii
1. Structural
of paclitaxel
formula
in various
Solubility paclitaxel Co-solvents/emulsions Soybean oil PEG 400 (75%) Triacetin Liposomes Nanocapsules Mixed micelles Prodrugs C-2’ esters C-7’ esters C-2’ carbonates Pro-prodrugs Cyclodextrins * Increasing
with
and
carbamates
increasing
total
lipid
of in medium
of paclitaxel.
media Reference
0.2-75 mg/ml 0.3 mglml 31 mg/ml 75 mg/ml 1 mg/ml 1.7-9.2 mol % 0.6 mg/ml 0.4-1.3 mg/ml*
Adams et a/. (3) Adams et a/. (3) Adams et a/. (3) Adams et al. (3) Bartoli et a/. (25) Sharma et a/. (27) Bartoli et al. (25) Alkan-Onyuksel et a/. (24)
up to 666 mg/ml N 10 mg/ml Not reported >I0 mglml 0.02-34.1 mg/ml
\Eyas(12) Was et Ueda et Was et Sharma
al. al. a/. et
(20) (19) (20) al. (32)
concentration.
One of these is its poor solubility in water, which has given rise to serious formulation problems. The solubility problem can be partly explained by the chemistry of paclitaxel. Paclitaxel is a complex diterpenoid product, having a bulky, extended fused ring system as well as a number of hydrophobic substituents (2) (Figure 1). It does not contain any functional ionizable group and, therefore, alteration of pH does not improve its solubility. For the same reason, the usual attempts to increase water solubility, such as salt formation or addition of charged complexing agents, are not usable in the case of paclitaxel. The current clinical dosage form of paclitaxel consists of a 5 ml size vial, with each millilitre of solution containing 6 mg of paclitaxel, 527 mg of Cremophor EL and 49.7% ethanol (?:I v/v) (3). This pharmaceutical formulation, however, is associated with a number of concerns including stability, filtering requirements and use of non-plasticized solution containers and administration sets. Moreover, some of the side-effects, such as severe hypersensitivity reactions, observed following paclitaxel
ALTERNATIVE
FORMULATIONS
OF PACLITAXEL
89
administration are considered to be formulation related. Studies have shown that the Cremophor EL vehicle induces histamine release and hypotension in dogs within 10 min after administration (4). It has also been demonstrated that Cremophor EL has a profound effect on the pharmacokinetics and biodistribution of paclitaxel (5, 6). Some investigators even observed that high levels of Cremophor EL antagonized the in vitro cytotoxicity of paclitaxel at certain concentrations (7). As the pharmaceutical vehicle is considered to be related with paclitaxel’s adverse effects, a lot of alternative formulations for the administration of paclitaxel have been, and still are, under investigation. Therefore, it seems worthwhile to study some of the approaches that have been made towards new formulations suitable for clinical evaluation.
Current
co-solvents
The addition of co-solvents is probably the most widely used solubilizing technique, because of their relatively low toxicity and their ability to increase the solubility of many non-polar drugs (8). An important disadvantage, however, is the possible precipitation upon the necessary dilution in the infusion fluid and possibly in the bloodstream. Paclitaxel has substantial solubility in several organic solvents such as ethanol and dimethylsulphoxide (DMSO). However, dilution with water of these solvents saturated with the poorly water-soluble drug paclitaxel, resulted in immediate precipitation of the drug. A 75% (v/v) polyethylene glycol400 (PEG 400) solution in water, containing 16mg/ml paclitaxel, was found to be chemically stable, but upon dilution for infusion, precipitation and cloudiness were noted (3). Furthermore, intraperitoneally (IP) administered paclitaxel in an aqueous PEG 400 solution was less active against an IP-implanted B16 melanoma tumour model than paclitaxel administered either as an aqueous suspension or as a solution containing Cremophor EL.
Emulsions An emulsion is a heterogenous mixture of two or more immiscible liquids, with a third component (emulsifier) used to stabilize the dispersed droplets (9). The use of fat emulsions can be a suitable approach to increase the solubility and stability of some antineoplastic drugs. However, many emulsifiers are toxic as a result of haemolytic reactions (10). The solubility of paclitaxel in lipids such as soybean oil is quite low, and precludes the use of simple oil-water emulsions for formulation considerations. Tarr et al. (II) developed an emulsion containing 50% triacetin, 2.0% ethyl oleate, 1.5% pluronic F68, 1.5% purified soybean oil and IOmg paclitaxel. Glycerol was added up to 10% to prevent creaming. This emulsion appeared stable enough for possible administration (11). However, triacetin (glyceryl triacetate) itself was toxic to mice when administered intravenously in concentrations required to deliver therapeutic doses of paclitaxel (12). Furthermore, no antitumour activity was reported with this formulation.
90
J. M. MEERUM TEFWOGT ETAL.
Prodrugs Several efforts to find a more convenient formulation for paclitaxel administration have involved the synthesis and evaluation of water-soluble prodrugs of paclitaxel. Prodrugs are inactive derivatives of the parent compound. Once within the body, the inactive prodrug is converted into the active form by spontaneous hydrolysis or by enzymatic degradation. Derivatization can increase the solubility of a drug with several orders of magnitude; however, it is a costly and difficult procedure, and the pharmacokinetic and pharmacodynamic properties may be altered undesirably (8). Mellado et al. found that the C-2’ hydroxyl group was the preferred functionality for future research, since derivatives at this position can be easily hydrolysed by enzymatic or chemical means (13). C-2’ esters are synthesized by the incorporation of ionizable groups such as amines, aminoacids and sulphonic acid groups. Many paclitaxel C-2’ esters have been investigated for their suitability as water-soluble prodrugs. These included succinate and glutarate derivatives, sulphonic acid derivatives and amino acid derivatives. Although these derivatives possess adequate water solubility and good biological activity in viva, most of them were found unsuitable as prodrugs of paclitaxel because of their chemical instability in an aqueous solution (12). One of the investigated esters, 2’-[3-(N,N-diethylamino)propionyllpaclitaxel, is relatively stable and therefore may be best suited for prodrug delivery of paclitaxel. It possessed adequate stability in aqueous solutions, although its half-life in human plasma was no longer than 5 min. Additionally, this prodrug exhibited in vivo activity against B16 melanoma cells similar to that of paclitaxel (14). Another series of C-2’ esters, the ‘protaxols’, was investigated by Nicolao et al. (15). These were mono-esters of dicarboxylic acids incorporating a heteroatom functionality (oxygen/sulphur) for the purpose of further enhancing water solubility. They displayed acceptable solubility and stability properties, and were capable of generating paclitaxel rapidly under basic or physiological conditions. In vitro, these ‘protaxols’ had greater potential as anticancer agents than the parent compound paclitaxel (15). However, in vivo antitumour activity of these ‘protaxols’ has not yet been reported. Greenwald et al. synthesized and evaluated polyethyleneglycolesters as potential prodrugs (16). The 2’ and -/-PEG esters of paclitaxel were very soluble in water and, when tested for in vitro antitumour activity, the 2’ ester exhibited similar activity to non-modified taxol, while the 7 ester showed reduced cytotoxic activity (16). Recently, Greenwald eta/. have successfully demonstrated the in vivo activity of some of these esters. Their ‘compound 9’ (a PEG ester with a molecular weight of 40 kDa) showed an enhanced antitumour activity and equivalent toxicity in comparison- with the paclitaxel-treated group of mice. Therefore, PEG esters appear to-offer an effective meansfor-paclitaxel delivery (17). Presenti et al. reported the synthesis. of ‘,hydroxypropylmethacrylamide polymers bearing an amino acid chW.that;is linked through an’esteric bond at the 2’ position of pat&axe+ (l8A ‘These compounds behave as paclitexel prodrugs; they are cle@‘able. by-, esterases releasing free paclitaxel in the .
ALTERNATIVE
FORMULATIONS
OF PACLITAXEL
91
CH3
CH3
‘CH,
CH2f”
- CH,
kT
NH&O \
O CH2
HAOH
NH
AH3
AH, H&OH
-
-
7
-NH-:H-
Figure
2. Structural
formula
of FCE 28161.
bloodstream. Furthermore, polymer-bound derivatives are easily solubilized in water and can be administered intravenously. Antitumour efficacy is dependent on the release profile in plasma, which is related to the composition of the amino acid chain. When tested against Ml09 murine lung carcinoma, one of these compounds [FCE 28161 (Figure 211showed increased antitumoural activity and lower toxicity in comparison with paclitaxel (18). Also, this compound was active against the M5076 murine reticulosarcoma whereas paclitaxel was not. To date, FCE 28161 is being evaluated in a clinical trial in the authors’ institute. In general the paclitaxel C-7 esters are not found suitable as water-soluble prodrugs, because of their enhanced in vivo stability towards esterases (12). For the same reason, paclitaxel-2’-carbonates and carbamates have found little utility as prodrugs thus far. Ueda et a/. found that some carbonates exhibited in viva antitumour activity comparable to that of paclitaxel, indicating that paclitaxel-2’ carbonates are probably converted to the parent drug under in vivo conditions (19). In spite of their efficacy, they were not found suitable for formulation, because of their extreme insolubility in water (12). Vyas et al. focused on the synthesis and evaluation of water soluble phosphate-cleavable prodrugs of paclitaxel (20). They selected the C-2’- and C7-phosphate derivatives as targets. In spite of their high water solubility, these derivatives were found unsuitable as prodrugs of paclitaxel because they lacked
92
J. M. MEERUM TERWOGT ETAL.
in vitro enzymatic cleavage and in vivo anti-tumour activity. However, Ueda et a/. (21) pursued this strategy with the synthesis of ‘pro-prodrugs’ of paclitaxel which can be activated by phosphatase. They designed paclitaxel derivatives having a phosphonoxyphenylpropionate ester group at the C-2’and C-7 position. These derivatives were able to generate paclitaxel in vivo, not by enzymatic hydrolysis, but by phosphate-initiated lactonization. When evaluated, the C-2 ester/phosphate of these pro-prodrugs was found to be marginally active in vivo against the murine solid tumour (Ml091 model, whereas the corresponding C-7 compound was as active as paclitaxel (21). In summary, a lot of studies focused on the paclitaxel prodrug design have yielded interesting results. Further investigations in this area seem to be justified. The clinical efficacy of either esterase- or phosphatase-activated prodrugs still needs to be established.
Liposomes
and micro-encapsulation
systems
Liposomes are microscopic vesicles (from 250 A to >20pm in diameter) composed of one or more lipid membranes surrounding discrete aqueous compartments (22). These vesicles can encapsulate lipid-soluble drugs within the lipid membrane, and thus offer the possibility of administering lipophilic drugs intravenously. Micro-encapsulation involves the application of a thin film of material around micronized solid or liquid to produce discrete units ranging in size from less than 1 pm to several millimetres (23). Nanoparticles and microspheres are examples of micro-encapsulation products. An important problem of all liposomal products is their long-term stability during storage (24). Another problem is often the large-scale manufacturing of sterile liposomes (8). An advantage of the use of liposomes is their relatively low toxicity, as a result of lower systemic exposure (8). In addition, liposomes and other encapsulation systems can be applied to target cytotoxic compounds more specifically to tumour cells (8). Bat-toli et al. studied the behaviour of paclitaxel encapsulated both in liposomes and in nanocapsules (vesicles with a diameter ranging from 200 to 500 nm). They concluded that paclitaxel in liposome form retains its properties in vitro as well as in vivo, at least with regard to the two tumour models used in this study-the P388 leukaemia model and the L1210 cells-whereas nanocapsules proved to be toxic, apparently due to their composition (25). Due to the fact that only these two tumour models were used, it is difficult to predict the possible clinical significance of these results (12). Alkan-Onyuksel et al. (24) reported the development of a mixed micellar formulation suitable for the administration of paclitaxel. Paclitaxel was solubilized in micelles formed by a mixture of bile salts and phospholipids. Precipitation was here avoided by the spontaneous formation of drug-loaded liposomes from these mixed micelles. In vitro, significant cytostatic activity was observed with the drug in this formulation. In addition, this mixed-micellar vehicle appeared to be less toxic than the current vehicle, Cremophor EL (24). This was partly due to the fact that a greater amount of Cremophor was needed
ALTERNATIVE
FORMULATIONS
OF PACLITAXEL
93
to solubilize an equivalent amount of paclitaxel, as compared to the mixed micelles. Straubinger et a/. developed over 300 liposome formulations and evaluated them for stability and activity (26-281. In vitro tests demonstrated that paclitaxel liposomes retained growth-inhibitory activity. Moreover, against Colon-26, a Taxol-resistant murine tumour, paclitaxel liposomes showed a delay of tumour progression (28). It seems rational to investigate these liposome-based formulations further on potential clinical activity. Recently, some studies have led to the development of microspheres (particles with diameter ranging in size from 1 to 100pm) as a new paclitaxel delivery system. Very high encapsulation efficiencies for paclitaxel in microspheres were reached. Studies using the chick chorioallantoic membrane (CAM) model showed that paclitaxel also possesses potent anti-angiogenic activity (29). This formulation may have potential for the targeted delivery of paclitaxel to a tumour via arterial chemo-embolization (30).
Complexes
with cyclodextrins
At present, complexation is a widely used method to increase the aqueous solubility and stability of a drug. Additionally, the complex of a cytotoxic drug with a site-specific carrier can deliver the drug more specifically to its tumour target, thereby increasing the therapeutic index. However, the dissociation of the complex in the body may lead to precipitation upon dilution. The solubility of the drug should therefore be studied at various concentrations and conditions (8). Cyclodextrins (CyDs) are molecular complexing agents that are produced by enzymatic starch degradation. Their capacity of enhancing solubility and stability derives from the formation of water-soluble inclusion complexes. Alpha(a)-, beta(p)- and gamma(y)-cyclodextrins are naturally occurring cyclic oligosaccharides containing 6, 7 and 8 glucopyranose rings, respectively (31). Sharma et a/. tested several /?- and y-cyclodextrins for their solubilizing properties (32). The p-cyclodextrins increased paclitaxel solubility considerably, but the solutions formed were highly viscous and removal of particulate matter remained difficult. The chemically modified cyclodextrin heptakis-2,6-di-Omethyl+CyD solubilized paclitaxel to the greatest content. Most of the investigated cyclodextrins have known toxicity. The relatively small solubilizing capacity of some of the cyclodextrins, however, made it impossible to administer the target dose of paclitaxel. Administration of paclitaxel doses near the maximum tolerated dose (of free drug in mice) required the injection of large quantities of a cyclodextrin, resulting in significant renal toxicity and haemolysis.
Conclusion It can be concluded that research into alternative formulations of paclitaxel is very active. To date, several promising alternatives have been developed. The
94
J. M. MEERUM
TERWOGT
ETAL.
suitability, advantages and disadvantages of each new formulation must be carefully assessed and compared to each other before progressing to clinical trials. While some approaches led to higher systemic toxicity, there are others that significantly reduced toxic reactions (e.g. the liposomal formulations). Furthermore, precipitation upon the necessary dilution during preparation of the infusion fluid still remains a major problem with many of the alternatives. However, with most of the prodrugs and the liposomes, this problem does not occur. Some of the prodrugs, particularly FCE 28161, as well as the liposomebased formulations seem to offer the possibility of safer intravenous administration of paclitaxel. Clinical testing is now required.
References 1. Huizing, M. T., Sewberath Misser, V. H., Pieters, R. C. et al. (1995) Taxanes: a new class of antitumour agents. Cancer invest. 13: 381-404. 2. Suffness, M. fed.) (1996) Biopharmaceutics of Pa&axe/ (Taxoll: Formulation, Activity and Pharmacokinetics-Taxok% Science and Applications. Straubinger, CRC Press, Boca Raton, pp. 237-258. 3. Adams, J. D., Flora, K. P., Goldspiel, B. R. et a/. (1993) Taxol: a history of pharmaceutical development and current pharmaceutical concerns. J. Nat/. Cancer lnsr. 15: 141-147. 4. Lorenz, W., Reimann, H. J., Schmal, A. et a/. (1977) Histamine release in dogs by Cremophor EL and its derivatives. Agents Actions 7: 63-67. 5. Sparreboom, A., van Tellingen, O., Nooijen, W. J. et a/. (1996) Nonlinear pharmacokinetics of paclitaxel in mice results from the pharmaceutical vehicle Cremophor EL. Cancer Res. 56: 2112-2115. 6. Sykes, E., Woodburn, K., Decker, D. et al. (1994) Effects of Cremophor EL on distribution of tax01 to serum lipoproteins. Br. J. Cancer 70: 401-404. 7. Liebmann, J., Cook, J., Lipschultz, C. et a/. (1994) The influence of Cremophor EL on the cell cycle effects of paclitaxel (Taxol) in human tumour cell lines. Cancer Chem. Pharm. 33: 331-339. 8. Jonkman-de Vries, J. D., Flora, K. P., Bult, A. et al. (1996) Pharmaceutical development of investigational anticancer agents for parenteral use-a review. Drug Dev. Ind. Pharm. 22: 475-494. 9. Davis, S. S., Washington, C., West, P. et a/. (1987) Lipid emulsions as drug delivery systems. Ann. NY Acad. Sci. 799: 75-88. 10. Hansrani, P. K., Davis, S. S. & Groves, M. J. (1983) The preparation and properties of sterile intravenous emulsions. J. Parent. Sci. Technol. 4: 145-150. 11. Tarr, B. D., Sambadan, T. G. & Yalkowski, S. H. (1987) A new parenteral emulsion for the administration of Taxol. fharm. Res. 4: 162-165. 12. Vyas, D. M. (1995) Paclitaxel (Taxol) formulation and prodrugs. In: Farina, V. ed., The Chemistry and Pharmacology of Taxol and its Derivatives. Amsterdam: Elsevier Science SK pp. 103-130. 13. Mellado, W., Magri, N. F., Kingston, D. G. I. et al. (1984) Preparation and biological of Taxol acetates. Biochem. Siophys. Res. Corn. 124: 329-336. 14. Mathew, A. E., Mejilano, M. R., Nath, J. P. et a/. (1992) Synthesis and evaluation of some water soluble prodrugs and derivatives of Taxol with antitumour activity. J. Med. Chem. 35: 141-151. 15. Nicolao, K. C., Dai, W. M. & Guy, R. K. (1993) Design, synthesis and biological activity of protaxols. Nature 364: 464-466. 16. Greenwald, R. B., Pendri, A., Bolikal, D. et a/. (1994) Highly water soluble taxol derivatives: 2 polyethylene glycol esters as potential prodrugs. Rioor& &led. Chem. Lett. 4: 2465-2470. 17. Greenwald, R. B., Gilbert, C. W., Pendri, A. et al. (1996) Drua deliverv svstems: water soluble taxol 2’ poty(ethyiene glycol) ester prodrugs--design and in‘;ivo effectiveness. J. Med. Chem. 39: 424-431. 18. Presenti, E., Franzetti, C., Biasoli, G. et a/. (1995) Synthesis and biological activity of water soluble polymer-bound taxol derivatives. Proc. Am. Ass. Cancer Res. 36: 307. 19. Ueda, Y., Wong, H., Matiskella, J. D. et a/. (1993) Synthesis and antitumour evaluation of 2’ oxycarbonylpaclitaxels (paclitaxel-2’-carbonates). Bioorg. Med. Chem. Lett. 4: 1861-1864. 20. Vyas, D. M., Ueda, Y., Wong, H. et a/. (1995) Phosphate-activated prodrugs of paclitaxel. Am. Chem. Sot. Symposium Series 9: 124-l 37.
ALTERNATIVE 21.
FORMULATIONS
OF PACLITAXEL
95
Ueda, Y., Mikkilineni, A. B., Knipe, J. 0. et al. (1993) Novel water soluble phosphate prodrugs of Taxol possessing in vivo antitumour activity. Bioorg. Med. Chem. Lert. 3: 1761-1766. 22. Ostro, M. J. & Cullis, P. R. (1989) Use of liposomes as injectable-drug-delivery systems. Am. J. Hosp. Pharm. 46: 1576-1587. 23. Chen. T.. Lausier. J. M.. Rhodes. C. T. (1986) Possible strateaies for the formulation of antineopiastic drubs. Drub Dev. Ink Phari. 12: 1041-1106. 24. Alkan-Onyuksel, H., Ramakrishnan, S., Chai, H. B. et a/. (1994) A mixed micellar formulation suitable for the parenteral administration of taxol. Pharm. Res. 11: 206-212. 25. Bartoli, M. H., Boitard, M., Fessi, H. et a/. (1990) In vitro and in vivo antitumoural activity of free and encapsulated taxol. J. Microencapsulation 7: 191-197. 26. Sharma, A., Mayhew, E. & Straubinger, R. M. (1993) Antitumour effect of taxol-containing liposomes in a taxol-resistant murine tumor model. Cancer Res. 53: 5877-5881. 27. Sharma, A. & Straubinger, R. M. (1994) Novel taxol formulations: preparation and characterization of taxol-containing liposomes. Pharm. Res. 11: 889-896. 28. Straubinger, R. M., Sharma, A., Murray, M. et a/. (1993) Novel taxol formulations: taxolcontaining liposomes. J. Nat. Cancer inst. Mon. 15: 69-78. 29. Dordunoo, S. K., Jackson, J. K., Arsenault, L. A. et al. (1995) Taxol encapsulation in poly(~caprolactone) microspheres. Cancer Chem. Pharm. 36: 279-282. 30. Burt, H. M., Jackson, J. K., Bains, S. K. eta/. (1995) Controlled delivery of taxol from microspheres composed of a blend of ethylene-vinyl acetate copolymer and poly(d,l-lactic) acid. Cancer Lett. 88: 73-79. 31. Bekers, O., Uijtendaal, E. V., Beijnen, J. H. et a/. (1991) Cyclodextrins in the pharmaceutical field. Drug Dev. Ind. Pharm. 17: 1503-1549. 32. Sharma, U. S., Balasubramanian, S. V. & Straubinger, R. M. (1995) Pharmaceutical and physical properties of paclitaxel (Taxol) complexes with cyclodextrins. Am. Chem. Sot. Am. Pharm. Ass. 84: 1223-1230.
Treatment
Alternative
Reviews
(1997)
23, 87-95
formulations
J. M. Meerum Terwogt*,t, J. H. Beijnen*,t,*
of paclitaxel
B. Nuijent,
W. W. Ten Bokkel Huinink*
and
* Department of Medical Oncology, The Netherlands Cancer Institute/ Antoni van Leeuwenhoek Hospital, Amsterdam, The Netherlands t Department of Pharmacy and Pharmacology, Slotervaatt Hospital/The Netherlands Cancer Institute, Amsterdam, The Netherlands $ Department of Pharmaceutical Analysis and Toxicology, Faculty of Pharmacy, State University of Utrecht, The Netherlands
Abstract Paclitaxel, a novel antitumour agent, is active clinically against advanced ovarian and breast cancer and under investigation for various other cancers. One of the problems associated with the intravenous administration of paclitaxel is its low solubility in water. The current pharmaceutical formulation consists of a 1:l (v/v) mixture of ethanol and Cremophor EL. This formulation, however, has been demonstrated to cause some severe hypersensitivity reactions. Therefore the development of a safer intravenous formulation devoid of Cremophor EL is an important investigational issue. This review deals with some of the most promising formulation alternatives.
Introduction Paclitaxel (TaxolB), a natural diterpene product isolated from the bark of Taxus brevifolia, has been shown to be highly cytotoxic and is clinically active against advanced ovarian and breast cancer. It has a unique mechanism of action, inducing the assembly of stable microtubules and inhibiting the depolymerization process (1). In spite of its promising antitumour activity, the drug has presented considerable difficulties related to its intravenous administration to patients.
Correspondence of Pharmacy and 0306-7372/97/020087
should be addressed to: J.M. Meerum Terwogt, Slotervaart Pharmacology, Louwesweg 6, 1066 EC, Amsterdam, The + 09 $12.00/O
0 1997 87
W.B.
Hospital, Netherlands. Saunders
Department
Company
Ltd
J. M. MEERUM
TERWOGT
HAL.
-N&Ho1; .&AH
Figure
Table
1.
Approximate
solubilii
1. Structural
of paclitaxel
formula
in various
Solubility paclitaxel Co-solvents/emulsions Soybean oil PEG 400 (75%) Triacetin Liposomes Nanocapsules Mixed micelles Prodrugs C-2’ esters C-7’ esters C-2’ carbonates Pro-prodrugs Cyclodextrins * Increasing
with
and
carbamates
increasing
total
lipid
of in medium
of paclitaxel.
media Reference
0.2-75 mg/ml 0.3 mglml 31 mg/ml 75 mg/ml 1 mg/ml 1.7-9.2 mol % 0.6 mg/ml 0.4-1.3 mg/ml*
Adams et a/. (3) Adams et a/. (3) Adams et a/. (3) Adams et al. (3) Bartoli et a/. (25) Sharma et a/. (27) Bartoli et al. (25) Alkan-Onyuksel et a/. (24)
up to 666 mg/ml N 10 mg/ml Not reported >I0 mglml 0.02-34.1 mg/ml
\Eyas(12) Was et Ueda et Was et Sharma
al. al. a/. et
(20) (19) (20) al. (32)
concentration.
One of these is its poor solubility in water, which has given rise to serious formulation problems. The solubility problem can be partly explained by the chemistry of paclitaxel. Paclitaxel is a complex diterpenoid product, having a bulky, extended fused ring system as well as a number of hydrophobic substituents (2) (Figure 1). It does not contain any functional ionizable group and, therefore, alteration of pH does not improve its solubility. For the same reason, the usual attempts to increase water solubility, such as salt formation or addition of charged complexing agents, are not usable in the case of paclitaxel. The current clinical dosage form of paclitaxel consists of a 5 ml size vial, with each millilitre of solution containing 6 mg of paclitaxel, 527 mg of Cremophor EL and 49.7% ethanol (?:I v/v) (3). This pharmaceutical formulation, however, is associated with a number of concerns including stability, filtering requirements and use of non-plasticized solution containers and administration sets. Moreover, some of the side-effects, such as severe hypersensitivity reactions, observed following paclitaxel
ALTERNATIVE
FORMULATIONS
OF PACLITAXEL
89
administration are considered to be formulation related. Studies have shown that the Cremophor EL vehicle induces histamine release and hypotension in dogs within 10 min after administration (4). It has also been demonstrated that Cremophor EL has a profound effect on the pharmacokinetics and biodistribution of paclitaxel (5, 6). Some investigators even observed that high levels of Cremophor EL antagonized the in vitro cytotoxicity of paclitaxel at certain concentrations (7). As the pharmaceutical vehicle is considered to be related with paclitaxel’s adverse effects, a lot of alternative formulations for the administration of paclitaxel have been, and still are, under investigation. Therefore, it seems worthwhile to study some of the approaches that have been made towards new formulations suitable for clinical evaluation.
Current
co-solvents
The addition of co-solvents is probably the most widely used solubilizing technique, because of their relatively low toxicity and their ability to increase the solubility of many non-polar drugs (8). An important disadvantage, however, is the possible precipitation upon the necessary dilution in the infusion fluid and possibly in the bloodstream. Paclitaxel has substantial solubility in several organic solvents such as ethanol and dimethylsulphoxide (DMSO). However, dilution with water of these solvents saturated with the poorly water-soluble drug paclitaxel, resulted in immediate precipitation of the drug. A 75% (v/v) polyethylene glycol400 (PEG 400) solution in water, containing 16mg/ml paclitaxel, was found to be chemically stable, but upon dilution for infusion, precipitation and cloudiness were noted (3). Furthermore, intraperitoneally (IP) administered paclitaxel in an aqueous PEG 400 solution was less active against an IP-implanted B16 melanoma tumour model than paclitaxel administered either as an aqueous suspension or as a solution containing Cremophor EL.
Emulsions An emulsion is a heterogenous mixture of two or more immiscible liquids, with a third component (emulsifier) used to stabilize the dispersed droplets (9). The use of fat emulsions can be a suitable approach to increase the solubility and stability of some antineoplastic drugs. However, many emulsifiers are toxic as a result of haemolytic reactions (10). The solubility of paclitaxel in lipids such as soybean oil is quite low, and precludes the use of simple oil-water emulsions for formulation considerations. Tarr et al. (II) developed an emulsion containing 50% triacetin, 2.0% ethyl oleate, 1.5% pluronic F68, 1.5% purified soybean oil and IOmg paclitaxel. Glycerol was added up to 10% to prevent creaming. This emulsion appeared stable enough for possible administration (11). However, triacetin (glyceryl triacetate) itself was toxic to mice when administered intravenously in concentrations required to deliver therapeutic doses of paclitaxel (12). Furthermore, no antitumour activity was reported with this formulation.
90
J. M. MEERUM TEFWOGT ETAL.
Prodrugs Several efforts to find a more convenient formulation for paclitaxel administration have involved the synthesis and evaluation of water-soluble prodrugs of paclitaxel. Prodrugs are inactive derivatives of the parent compound. Once within the body, the inactive prodrug is converted into the active form by spontaneous hydrolysis or by enzymatic degradation. Derivatization can increase the solubility of a drug with several orders of magnitude; however, it is a costly and difficult procedure, and the pharmacokinetic and pharmacodynamic properties may be altered undesirably (8). Mellado et al. found that the C-2’ hydroxyl group was the preferred functionality for future research, since derivatives at this position can be easily hydrolysed by enzymatic or chemical means (13). C-2’ esters are synthesized by the incorporation of ionizable groups such as amines, aminoacids and sulphonic acid groups. Many paclitaxel C-2’ esters have been investigated for their suitability as water-soluble prodrugs. These included succinate and glutarate derivatives, sulphonic acid derivatives and amino acid derivatives. Although these derivatives possess adequate water solubility and good biological activity in viva, most of them were found unsuitable as prodrugs of paclitaxel because of their chemical instability in an aqueous solution (12). One of the investigated esters, 2’-[3-(N,N-diethylamino)propionyllpaclitaxel, is relatively stable and therefore may be best suited for prodrug delivery of paclitaxel. It possessed adequate stability in aqueous solutions, although its half-life in human plasma was no longer than 5 min. Additionally, this prodrug exhibited in vivo activity against B16 melanoma cells similar to that of paclitaxel (14). Another series of C-2’ esters, the ‘protaxols’, was investigated by Nicolao et al. (15). These were mono-esters of dicarboxylic acids incorporating a heteroatom functionality (oxygen/sulphur) for the purpose of further enhancing water solubility. They displayed acceptable solubility and stability properties, and were capable of generating paclitaxel rapidly under basic or physiological conditions. In vitro, these ‘protaxols’ had greater potential as anticancer agents than the parent compound paclitaxel (15). However, in vivo antitumour activity of these ‘protaxols’ has not yet been reported. Greenwald et al. synthesized and evaluated polyethyleneglycolesters as potential prodrugs (16). The 2’ and -/-PEG esters of paclitaxel were very soluble in water and, when tested for in vitro antitumour activity, the 2’ ester exhibited similar activity to non-modified taxol, while the 7 ester showed reduced cytotoxic activity (16). Recently, Greenwald eta/. have successfully demonstrated the in vivo activity of some of these esters. Their ‘compound 9’ (a PEG ester with a molecular weight of 40 kDa) showed an enhanced antitumour activity and equivalent toxicity in comparison- with the paclitaxel-treated group of mice. Therefore, PEG esters appear to-offer an effective meansfor-paclitaxel delivery (17). Presenti et al. reported the synthesis. of ‘,hydroxypropylmethacrylamide polymers bearing an amino acid chW.that;is linked through an’esteric bond at the 2’ position of pat&axe+ (l8A ‘These compounds behave as paclitexel prodrugs; they are cle@‘able. by-, esterases releasing free paclitaxel in the .
ALTERNATIVE
FORMULATIONS
OF PACLITAXEL
91
CH3
CH3
‘CH,
CH2f”
- CH,
kT
NH&O \
O CH2
HAOH
NH
AH3
AH, H&OH
-
-
7
-NH-:H-
Figure
2. Structural
formula
of FCE 28161.
bloodstream. Furthermore, polymer-bound derivatives are easily solubilized in water and can be administered intravenously. Antitumour efficacy is dependent on the release profile in plasma, which is related to the composition of the amino acid chain. When tested against Ml09 murine lung carcinoma, one of these compounds [FCE 28161 (Figure 211showed increased antitumoural activity and lower toxicity in comparison with paclitaxel (18). Also, this compound was active against the M5076 murine reticulosarcoma whereas paclitaxel was not. To date, FCE 28161 is being evaluated in a clinical trial in the authors’ institute. In general the paclitaxel C-7 esters are not found suitable as water-soluble prodrugs, because of their enhanced in vivo stability towards esterases (12). For the same reason, paclitaxel-2’-carbonates and carbamates have found little utility as prodrugs thus far. Ueda et a/. found that some carbonates exhibited in viva antitumour activity comparable to that of paclitaxel, indicating that paclitaxel-2’ carbonates are probably converted to the parent drug under in vivo conditions (19). In spite of their efficacy, they were not found suitable for formulation, because of their extreme insolubility in water (12). Vyas et al. focused on the synthesis and evaluation of water soluble phosphate-cleavable prodrugs of paclitaxel (20). They selected the C-2’- and C7-phosphate derivatives as targets. In spite of their high water solubility, these derivatives were found unsuitable as prodrugs of paclitaxel because they lacked
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in vitro enzymatic cleavage and in vivo anti-tumour activity. However, Ueda et a/. (21) pursued this strategy with the synthesis of ‘pro-prodrugs’ of paclitaxel which can be activated by phosphatase. They designed paclitaxel derivatives having a phosphonoxyphenylpropionate ester group at the C-2’and C-7 position. These derivatives were able to generate paclitaxel in vivo, not by enzymatic hydrolysis, but by phosphate-initiated lactonization. When evaluated, the C-2 ester/phosphate of these pro-prodrugs was found to be marginally active in vivo against the murine solid tumour (Ml091 model, whereas the corresponding C-7 compound was as active as paclitaxel (21). In summary, a lot of studies focused on the paclitaxel prodrug design have yielded interesting results. Further investigations in this area seem to be justified. The clinical efficacy of either esterase- or phosphatase-activated prodrugs still needs to be established.
Liposomes
and micro-encapsulation
systems
Liposomes are microscopic vesicles (from 250 A to >20pm in diameter) composed of one or more lipid membranes surrounding discrete aqueous compartments (22). These vesicles can encapsulate lipid-soluble drugs within the lipid membrane, and thus offer the possibility of administering lipophilic drugs intravenously. Micro-encapsulation involves the application of a thin film of material around micronized solid or liquid to produce discrete units ranging in size from less than 1 pm to several millimetres (23). Nanoparticles and microspheres are examples of micro-encapsulation products. An important problem of all liposomal products is their long-term stability during storage (24). Another problem is often the large-scale manufacturing of sterile liposomes (8). An advantage of the use of liposomes is their relatively low toxicity, as a result of lower systemic exposure (8). In addition, liposomes and other encapsulation systems can be applied to target cytotoxic compounds more specifically to tumour cells (8). Bat-toli et al. studied the behaviour of paclitaxel encapsulated both in liposomes and in nanocapsules (vesicles with a diameter ranging from 200 to 500 nm). They concluded that paclitaxel in liposome form retains its properties in vitro as well as in vivo, at least with regard to the two tumour models used in this study-the P388 leukaemia model and the L1210 cells-whereas nanocapsules proved to be toxic, apparently due to their composition (25). Due to the fact that only these two tumour models were used, it is difficult to predict the possible clinical significance of these results (12). Alkan-Onyuksel et al. (24) reported the development of a mixed micellar formulation suitable for the administration of paclitaxel. Paclitaxel was solubilized in micelles formed by a mixture of bile salts and phospholipids. Precipitation was here avoided by the spontaneous formation of drug-loaded liposomes from these mixed micelles. In vitro, significant cytostatic activity was observed with the drug in this formulation. In addition, this mixed-micellar vehicle appeared to be less toxic than the current vehicle, Cremophor EL (24). This was partly due to the fact that a greater amount of Cremophor was needed
ALTERNATIVE
FORMULATIONS
OF PACLITAXEL
93
to solubilize an equivalent amount of paclitaxel, as compared to the mixed micelles. Straubinger et a/. developed over 300 liposome formulations and evaluated them for stability and activity (26-281. In vitro tests demonstrated that paclitaxel liposomes retained growth-inhibitory activity. Moreover, against Colon-26, a Taxol-resistant murine tumour, paclitaxel liposomes showed a delay of tumour progression (28). It seems rational to investigate these liposome-based formulations further on potential clinical activity. Recently, some studies have led to the development of microspheres (particles with diameter ranging in size from 1 to 100pm) as a new paclitaxel delivery system. Very high encapsulation efficiencies for paclitaxel in microspheres were reached. Studies using the chick chorioallantoic membrane (CAM) model showed that paclitaxel also possesses potent anti-angiogenic activity (29). This formulation may have potential for the targeted delivery of paclitaxel to a tumour via arterial chemo-embolization (30).
Complexes
with cyclodextrins
At present, complexation is a widely used method to increase the aqueous solubility and stability of a drug. Additionally, the complex of a cytotoxic drug with a site-specific carrier can deliver the drug more specifically to its tumour target, thereby increasing the therapeutic index. However, the dissociation of the complex in the body may lead to precipitation upon dilution. The solubility of the drug should therefore be studied at various concentrations and conditions (8). Cyclodextrins (CyDs) are molecular complexing agents that are produced by enzymatic starch degradation. Their capacity of enhancing solubility and stability derives from the formation of water-soluble inclusion complexes. Alpha(a)-, beta(p)- and gamma(y)-cyclodextrins are naturally occurring cyclic oligosaccharides containing 6, 7 and 8 glucopyranose rings, respectively (31). Sharma et a/. tested several /?- and y-cyclodextrins for their solubilizing properties (32). The p-cyclodextrins increased paclitaxel solubility considerably, but the solutions formed were highly viscous and removal of particulate matter remained difficult. The chemically modified cyclodextrin heptakis-2,6-di-Omethyl+CyD solubilized paclitaxel to the greatest content. Most of the investigated cyclodextrins have known toxicity. The relatively small solubilizing capacity of some of the cyclodextrins, however, made it impossible to administer the target dose of paclitaxel. Administration of paclitaxel doses near the maximum tolerated dose (of free drug in mice) required the injection of large quantities of a cyclodextrin, resulting in significant renal toxicity and haemolysis.
Conclusion It can be concluded that research into alternative formulations of paclitaxel is very active. To date, several promising alternatives have been developed. The
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TERWOGT
ETAL.
suitability, advantages and disadvantages of each new formulation must be carefully assessed and compared to each other before progressing to clinical trials. While some approaches led to higher systemic toxicity, there are others that significantly reduced toxic reactions (e.g. the liposomal formulations). Furthermore, precipitation upon the necessary dilution during preparation of the infusion fluid still remains a major problem with many of the alternatives. However, with most of the prodrugs and the liposomes, this problem does not occur. Some of the prodrugs, particularly FCE 28161, as well as the liposomebased formulations seem to offer the possibility of safer intravenous administration of paclitaxel. Clinical testing is now required.
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FORMULATIONS
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Ueda, Y., Mikkilineni, A. B., Knipe, J. 0. et al. (1993) Novel water soluble phosphate prodrugs of Taxol possessing in vivo antitumour activity. Bioorg. Med. Chem. Lert. 3: 1761-1766. 22. Ostro, M. J. & Cullis, P. R. (1989) Use of liposomes as injectable-drug-delivery systems. Am. J. Hosp. Pharm. 46: 1576-1587. 23. Chen. T.. Lausier. J. M.. Rhodes. C. T. (1986) Possible strateaies for the formulation of antineopiastic drubs. Drub Dev. Ink Phari. 12: 1041-1106. 24. Alkan-Onyuksel, H., Ramakrishnan, S., Chai, H. B. et a/. (1994) A mixed micellar formulation suitable for the parenteral administration of taxol. Pharm. Res. 11: 206-212. 25. Bartoli, M. H., Boitard, M., Fessi, H. et a/. (1990) In vitro and in vivo antitumoural activity of free and encapsulated taxol. J. Microencapsulation 7: 191-197. 26. Sharma, A., Mayhew, E. & Straubinger, R. M. (1993) Antitumour effect of taxol-containing liposomes in a taxol-resistant murine tumor model. Cancer Res. 53: 5877-5881. 27. Sharma, A. & Straubinger, R. M. (1994) Novel taxol formulations: preparation and characterization of taxol-containing liposomes. Pharm. Res. 11: 889-896. 28. Straubinger, R. M., Sharma, A., Murray, M. et a/. (1993) Novel taxol formulations: taxolcontaining liposomes. J. Nat. Cancer inst. Mon. 15: 69-78. 29. Dordunoo, S. K., Jackson, J. K., Arsenault, L. A. et al. (1995) Taxol encapsulation in poly(~caprolactone) microspheres. Cancer Chem. Pharm. 36: 279-282. 30. Burt, H. M., Jackson, J. K., Bains, S. K. eta/. (1995) Controlled delivery of taxol from microspheres composed of a blend of ethylene-vinyl acetate copolymer and poly(d,l-lactic) acid. Cancer Lett. 88: 73-79. 31. Bekers, O., Uijtendaal, E. V., Beijnen, J. H. et a/. (1991) Cyclodextrins in the pharmaceutical field. Drug Dev. Ind. Pharm. 17: 1503-1549. 32. Sharma, U. S., Balasubramanian, S. V. & Straubinger, R. M. (1995) Pharmaceutical and physical properties of paclitaxel (Taxol) complexes with cyclodextrins. Am. Chem. Sot. Am. Pharm. Ass. 84: 1223-1230.