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Vasoconstriction and anti-inflammatory properties of the selective α-adrenergic receptor agonist brimonidine
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Vasoconstriction and anti-inflammatory properties of the selective a-adrenergic receptor agonist brimonidine David Piwnica *, Carine Rosignoli, Se´verine Thibaut de Me´nonville, Thierry Alvarez, Marlene Schuppli Nollet, Olivier Roye, Andre´ Jomard, Je´roˆme Aubert Research, Galderma, Sophia Antipolis, France
A R T I C L E I N F O
A B S T R A C T
Article history: Received 23 January 2014 Received in revised form 1 April 2014 Accepted 3 April 2014
Background: The facial erythema of rosacea is recognized as the most prevalent and most difficult manifestation of rosacea to treat. A recent approach in patients with rosacea has been to reduce this erythema through vasoconstriction of cutaneous blood vessels by selectively targeting a2-adrenergic receptors with brimonidine. Objective: To further investigate the pharmacodynamic profile of brimonidine, its vasoconstrictive effects and its anti-inflammatory properties. Methods: The potency for the a1A, a1B, a2A, a2B and a2C receptors of brimonidine was measured, as well as performing a large target profiling study in order to determine the target selectivity profile of brimonidine. The vasoconstrictive effects of brimonidine were measured using ex vivo wire myography and human skin biopsy neuroinflammation models. The anti-inflammatory properties of brimonidine were measured using two in vivo mice ear inflammation models. Results: Brimonidine was found to be highly selective for the a2A adrenoreceptor (EC50 0.45 nM) over the other a-adrenoreceptors. Additionally, the large target profiling study demonstrated the high selectivity of brimonidine with minimal off-target effects. The ex vivo wire myography model showed that brimonidine is a potent vasoconstrictor of human subcutaneous vessels with a diameter of less than 200 mm (EC50 0.4 nM). The ex vivo human skin biopsy neuroinflammation model demonstrated that brimonidine completely inhibited vasodilation induced by capsaicin. Both in vivo mouse ear inflammation models highlighted that brimonidine inhibited ear edema (up to 76%) when compared to vehicle. Conclusion: The selectivity, vasoconstrictive and anti-inflammatory properties of brimonidine that have been described in these studies are in agreement with the benefits observed with this compound in the treatment of facial erythema in rosacea. ß 2014 Published by Elsevier Ireland Ltd on behalf of Japanese Society for Investigative Dermatology.
Keywords: Rosacea Vascular Inflammation Brimonidine Mirvaso Pharmacology
1. Introduction Rosacea is a chronic skin disorder with an unknown etiology, characterized by transient or persistent erythema, telangiectasia, papules and pustules, that primarily affects the central facial region (cheeks, nose, chin and forehead) [1–3]. Persistent facial erythema is most commonly associated with subtypes I and II of rosacea, namely erythematotelangiectatic (ETR) and papulopustular rosacea (PPR) [2,3]. Rosacea, in particular facial erythema, is often triggered by a wide range of environmental or lifestyle factors including, but not limited to: sun exposure, emotional
* Corresponding author. Tel.: +33 1492383026. E-mail address: [email protected] (D. Piwnica).
stress, temperature, exercise and alcohol consumption [4]. The facial erythema of rosacea is recognized to be the most prevalent and also the most difficult manifestation to treat [4–6]. The pathophysiology of facial erythema of rosacea is relatively unknown. It is suggested to result from dysregulation of cutaneous vasomotor responses, which causes abnormal, involuntary and persistent dilation of facial blood vessels [7–10]. The severity of rosacea has also been linked to prolonged vasodilation of small blood vessels (<200 mm), with evidence to suggest that vasodilation may result from inflammatory mediators released during the early phase of rosacea [7,9,11,12]. These characteristics of rosacea are similar in nature to the main features of neurogenic inflammation, a condition caused by the local release of inflammatory mediators from sensory neurons [13]. The link between rosacea and neurogenic inflammation is further strengthened by a
http://dx.doi.org/10.1016/j.jdermsci.2014.04.002 0923-1811/ß 2014 Published by Elsevier Ireland Ltd on behalf of Japanese Society for Investigative Dermatology.
Please cite this article in press as: Piwnica D, et al. Vasoconstriction and anti-inflammatory properties of the selective a-adrenergic receptor agonist brimonidine. J Dermatol Sci (2014), http://dx.doi.org/10.1016/j.jdermsci.2014.04.002
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proteases and 5 phosphodiesterases). The full assay list can be found in the supplementary information. Brimonidine response was normalized to the maximum signal detected in each assay.
number of recent studies that demonstrate the release of vasoactive peptides by sensory neurones in skin upon exposure to known triggers [7,11,14], and co-localization between facial sensory nerve endings and blood vessels in patients with rosacea [7,11,13]. One recent study has shown an increase of markers, which are also present in neurogenic inflammation such as mast cells, neuropeptides (substance P and CGRPa) as well as matrix remodeling, in patients with rosacea [15]. In addition, patients with rosacea have been associated with a higher prevalence of migraine, a condition also thought to involve neurogenic inflammation [16,17]. Neurovascular and neuroimmune interactions in rosacea have revealed the involvement of a number of genes including adrenergic receptors [13]. Novel approaches are being assessed for the treatment of facial erythema of rosacea. One recent approach has been to reduce erythema through vasoconstriction of cutaneous blood vessels by targeting both a-adrenergic receptors with oxymetazoline [18,19] and brimonidine [10,20]. Oxymetazoline is an agonist of the a1Aadrenoceptor and a partial agonist of the a2A-receptor agonist; it is currently in clinical development (Phase IIb) for the treatment of facial erythema in rosacea [18,19]. Brimonidine is a highly selective a2-adrenergic receptor agonist with potent vasoconstrictive activity that has been approved for the treatment of openangle glaucoma for almost 20 years [21–24]. Brimonidine has also recently been approved as a promising, new therapy for the topical treatment of persistent (nontransient) facial erythema of rosacea in adults 18 years of age or older (Mirvaso1, Galderma R&D, Sophia Antipolis, France) [25]. Recent Phase II and III studies with topical brimonidine 0.33% gel (once daily) demonstrated its efficacy and safety [10,20] for periods of up to one year [26]. The aim of this study was to further investigate the pharmacodynamic profile of brimonidine, its vasoconstrictive effects on subcutaneous arteries and veins and its anti-inflammatory properties in vivo, in comparison with oxymetazoline.
Subcutaneous blood vessels were isolated from human abdominal plastic surgery. Healthy human skin was placed in cold physiological saline solution (PSS) and stored at 4 8C until required. The subcutaneous blood vessels were removed from the skin using forceps and scissors under stereomicroscope and maintained in PSS at pH = 7.4. Blood vessels were cut into ring segments, roughly 2 mm in length, and stored in cold PSS. The 2 mm-segments were then mounted using two fine tungsten wires (40 mm in diameter) into a four chamber myograph (EMKABath4, EMKA Technologies). Data were recorded using the Iox 2.8 software. The vessel segments were then stretched to the tension corresponding to a physiological pressure of 90 mmHg for arteries and 18 mmHg for veins, and lumen diameter was measured using Normalize software from EMKA Technologies. Vessel segments were maintained in a preheated PSS solution gassed with a mixture of 5% CO2, 20% O2 and 75% N2 (pH = 7.4) for 1 h before being used. During this period, the bath solution was changed every 20 min. The viability of blood vessel segments was assessed by measuring the contraction obtained after two successive stimulations with the PSS solution containing 80 mM of KCl. Then, the smooth muscle vascular wall function was evaluated using a vasoconstrictor compound such as prostaglandin F2a at 10 mM, whereas the endothelium function was assessed using acetylcholine at 1 mM. Oxymetazoline and brimonidine were dissolved in DMSO. Both compounds were added in a range of concentrations directly to the chamber and the modulation of vascular tones was recorded.
2. Materials and methods
2.3. Ex vivo human skin model
2.1. Target-based assays
Healthy human skin was obtained from 10 different donors who underwent abdominoplasty. Skin biopsies (1 cm2) were obtained and placed in culture with the epithelium on top using an insert with 0.45 mm polycarbonate membrane (Fig. 1). Inserts were then set in 12-well plates pre-filled with culture medium (DMEM supplemented with FCS and antibiotics). Thus, this ex vivo organ culture system maintained the skin at the air–liquid interface and fed the dermis and epidermis by nutrient diffusion across the insert. Biopsies were then placed in a humidified atmosphere of 95% air and 5% CO2 at 37 8C. 20 mL/cm2 of 0.33% of brimonidine (Mirvaso1, Galderma R&D, Sophia Antipolis, France, 0.33 g brimonidine in 100 mL of gel equivalent to 11.3 mM) or vehicle gel (corresponding vehicle gel to Mirvaso1, Galderma R&D, Sophia Antipolis, France), was added topically to the skin biopsies 2 hours before addition of capsaicin and capsaicin was added at 10 mM to the culture medium for 4 h. Skin
An adrenoreceptor assay selectivity profile study was conducted at Cerep (Poitiers, France) with human a1A (Cerep Cat 1500), a1B (Cerep Cat 1901), a2A (Cerep Cat 2A2558), a2B (Cerep Cat 1813) and a2C (Cerep Cat 1736) receptors selected to evaluate the EC50 potency of oxymetazoline (Sigma–Aldrich, France) and brimonidine (Sigma–Aldrich, France). Both compounds were dissolved in DMSO then diluted in HEPES sodium salt buffer. Both compounds tested over a dose range of 1 pM to 10 mM. A large target profiling study of brimonidine (Bioprint1 profile) was performed at Cerep (Poitiers, France) with the panel mainly based on target diversity. The study included 104 binding assays (non-peptide, peptide and nuclear receptors, ion channels and amine transporters) and 32 enzyme assays (including 10 kinases, 9
2.2. In vitro wire myography
Fig. 1. Schematic diagram of the ex vivo human skin model used to measure the vasoconstrictor activity of brimonidine. Black arrowheads show vasodilation induced by capsaicin application.
Please cite this article in press as: Piwnica D, et al. Vasoconstriction and anti-inflammatory properties of the selective a-adrenergic receptor agonist brimonidine. J Dermatol Sci (2014), http://dx.doi.org/10.1016/j.jdermsci.2014.04.002
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biopsies were then fixed in formol liquid and embedded in paraffin for histological analyses. After staining the histological section with hematoxylin and eosin, the surface area covered by the lumen of the dilated vessels (mm2) and the percentage of vessels dilated were calculated. Using the image analysis software Image-Pro Plus (Media Cybernetics, Inc., MD, USA), 16 areas were observed on the histological section. An average of 50 vessels were analyzed across all the 16 sections. For statistical analysis, this experiment was repeated on biopsies from 10 donors and the statistical analysis was conducted using a reduced deviation Student’s t-test or the paired ttest with a risk of 5%. 2.4. In vivo inflammation model Female BALB/cByJ Rj mice were purchased from JANVIER (France). All experiments were conducted at the Unite´ d’Evaluation de Produits Pharmacologiques (UDEPP, France) under specific pathogen-free conditions with mice between 8 and 9 weeks of age. All experiments were conducted with the approval of and in accordance with the guidelines for animal experiments of the local ethics committee (CEEA, France). Arachidonic acid (Sigma–Aldrich, France) was dissolved in tetrahydrofuran/methanol (1:1) at 4% (AA-induced skin inflammation) and 12-O-Tetradecanoylphorbol-13-acetate (Sigma– Aldrich, France) was dissolved in ethanol at 0.01% (TPA-induced skin inflammation). Brimonidine (Sigma–Aldrich, France) was added to the AA or TPA solution to achieve a final concentration of 0.2% (0.2 g in 100 mL of solution equivalent to 6.9 mM), which was the highest concentration possible due to solubility reasons. 20 ml of AA or TPA solution, with or without brimonidine 0.2%, was applied topically on the internal side of the mice ear. Ear thickness was measured using a micrometer 1 h or 6 h after brimonidine application for the AA-induced skin inflammation and TPAinduced skin inflammation models respectively.
Fig. 2. Target profile of brimonidine at 10 mM on a broad panel of 141 biological targets confirmed the high selectivity for a-adrenergic receptors and especially a2 subtypes. For full size graph see supplementary information.
3. Results 3.1. Brimonidine has high selectivity for a-adrenergic receptors with minimal off-target activity The selectivity of both brimonidine and oxymetazoline for the adrenoreceptors was determined by measuring the EC50 values for the a1A, a1B, a2A, a2B and a2C receptors. Brimonidine was found to be mainly selective for the a2A receptor, with an EC50 of 0.45 nM. In comparison, oxymetazoline was found to be less selective with an EC50 of 1.1 nM and 0.33 nM on a2A and a1A receptors respectively (Table 1). A large target profiling study was performed, including binding assays (non-peptide, peptide and nuclear receptors, ion channels and amine transporters) and enzyme assays (kinases, proteases and phosphodiesterases) to determine the selectivity of brimonidine. A compound with an effect greater than 30% at 10 mM is considered as active against the target. Brimonidine was found to be highly selective for a-adrenergic receptors, in particular a2 subtypes, with virtually no off-target activity seen, as shown by the percentage of response demonstrated at 10 mM on a broad panel of targets (Fig. 2 and supplementary information). Table 1 Agonist activity of brimonidine and oxymetazoline on human a-adrenergic receptors.
Oxymetazoline EC50 (nM) Brimonidine EC50 (nM)
a1A
a1B
a2A
a2B
a2C
0.33 56
N.C.a N.C.a
1.1 0.45
240 1900
5.9 6.5
a N.C., not calculable (concentration–response curve shows less than 25% effect at the highest validated testing concentration).
3.2. Wire myography demonstrates that brimonidine is a potent vasoconstrictor of sub 200 mm human subcutaneous blood vessels The wire myography technique was used to evaluate the functional responses and vascular reactivity of various sized isolated subcutaneous arteries when subjected to either brimonidine or oxymetazoline. Oxymetazoline was found to have EC50 values of 2 nM, 31 nM, 5.1 nM and 4.5 nM for the 184 mm, 433 mm, 513 mm and 630 mm diameter vessels, respectively (Fig. 3A–D). Brimonidine was found to be 5 times more potent than oxymetaxoline as a vasoconstrictor on human subcutaneous arteries smaller than 200 mm diameter, with an EC50 of 0.4 nM for the 184 mm vessels (Fig. 3A). The potency and efficacy of brimonidine was found to decrease as the vessel diameter increased, with EC50 values reaching 490 nM and 50% of relative efficacy (compared to oxymetazoline) for vessels of 630 mm in diameter, displaying its selectivity for the sub 200 mm vessels only (Fig. 3D). In addition, the activity of brimonidine was compared on similar sized arteries and veins (222 mm and 210 mm, respectively). Brimonidine was shown to be active on both types of subcutaneous blood vessels with EC50 values of 2.9 nM and 24 nM on arteries and veins respectively (Fig. 4). 3.3. Topical application of brimonidine significantly reduces vasodilation on ex vivo human skin biopsies after capsaicin application An ex vivo human skin model was used to assess the activity of brimonidine on subcutaneous vessel dilation induced by capsaicin,
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Fig. 3. Effect of brimonidine on human subcutaneous vessels using wire myography technique. Arteries of 184 mm had EC50 values of 0.4 nM and 2 nM for brimonidine and oxymetazoline respectively (A). Arteries of 433 mm had EC50 values of 62 nM and 31 nM for brimonidine and oxymetazoline respectively (B). Arteries of 513 mm had EC50 values of 190 nM and 5.1 nM for brimonidine and oxymetazoline respectively (C). Arteries of 630 mm had EC50 values of 490 nM and 4.5 nM for brimonidine and oxymetazoline respectively (D).
a known trigger of neurogenic inflammation [7]. Topical treatment with brimonidine 0.33% (Mirvaso1, Galderma R&D, Sophia Antipolis, France) resulted in a statistically significant almost complete reduction, compared to the gel vehicle, in the surface area occupied by the blood vessels following capsaicin-induced
vasodilation, (66.7 mm2 vs. 180.5 mm2; p < 0.001) (Fig. 5A). This finding was mirrored in the percentage of dilated vessels, with a significantly reduced percentage of dilated vessels in the brimonidine treated group, when compared to vehicle (Fig. 5B). 3.4. In vivo inflammation is reduced with brimonidine application Two mouse models of inflammation were used to measure the anti-inflammatory effect of brimonidine in an in vivo setting. Arachidonic acid (AA)-induced ear edema was inhibited by 76% with topical administration of brimonidine 0.2% (Fig. 6A) and TPAinduced ear edema was inhibited by 50% by topical administration of brimonidine 0.2% (Fig. 6B). 4. Discussion
Fig. 4. Dose–response effect of brimonidine on the vascular tone of human subcutaneous veins and arteries. Veins of 210 mm and arteries of 222 mm were found to have EC50 values of 24 nM and 2.9 nM, respectively.
Facial erythema in rosacea is thought to be the result of vasodilation of the cutaneous vasculature in the central facial region [7–10], with vessel diameters lower than 200 mm in rosacea patients [12]. The similarities between rosacea and inflammation have recently been demonstrated in numerous studies including the observed cytokine, chemokine and cellular infiltration during intermittent flares [9,11,12]; the marked inflammatory infiltrate of CD4+ T lymphocytes, macrophages and mast cells seen in ETR [7]; the release of vasoactive peptides [7,11,14]; and the increased expression of vasoactive peptides calcitonin gene-related peptide [CGRP-a] and substance P [15]. Further supporting evidence of a
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Fig. 5. Effect of brimonidine 0.33% on surface area (A) and percentage (B) of dilated vessels in the capsaicin-induced vasodilation ex vivo human skin model. ***p < 0.001, for capsaicin with vehicle compared to: control, vehicle and capsaicin.
link between rosacea and neuroinflammation is highlighted by the co-localization between facial sensory nerve endings and blood vessels in patients with rosacea [11,13] and the higher prevalence of migraine in patients with rosacea [16,17]. These studies combined with a recent study displaying the involvement of a number of adrenergic receptors [13] have suggested the potential of using a highly specific a-adrenergic receptor agonist with vasoconstrictive and anti-inflammatory properties to treat the facial erythema of rosacea. Brimonidine 0.33% (Mirvaso1, Galderma R&D, Sophia Antipolis, France) [25] has recently received FDA approval for the treatment of facial erythema in rosacea [6]. In this paper, we have described the pharmacodynamic, ex vivo vasoconstrictor activity and in vivo anti-inflammatory effects of brimonidine. The profiling of brimonidine in more than 200 targets confirmed that it has very high selectivity toward the a2 adrenergic receptors. Moreover, the profiling data revealed that brimonidine was inactive against the 5-hydroxytryptamine 2B receptor (5HT2b), reported to be involved in valvular heart disease after long-term treatment, opposed to other vasoconstrictors such as oxymetazoline [27]. In addition, we demonstrated that brimonidine was more potent and selective than oxymetazoline on small subcutaneous arteries (<200 mm) using the wire myography technique. This is of particular importance in facial erythema as the higher selectivity exhibited by brimonidine for targeting the smaller vessels is of notable benefit as one previous study showed blood vessel
diameters lower than 200 mm are involved in rosacea [12]. Studies of the a2 receptor have suggested that a2A, a2B and a2D subtypes are of particular importance in arteries, and that a2A, a2C and a2D subtypes are of more importance on the venous side [18]. The importance of the a2A receptor subtype on both arteries and veins was demonstrated in this study by the ability of brimonidine to initiate vasoconstriction of both arteries and veins. Targeting of the venous side is of real potential in the treatment of facial erythema as it has previously been shown that one of the first abnormalities to manifest in patients with rosacea is a microcirculatory disturbance of facial angular veins [12]. In the human skin biopsy model, a model of neurogenic inflammation, brimonidine 0.33% (Mirvaso1, Galderma R&D, Sophia Antipolis, France) was shown to have vasoconstrictive properties that negated the effect of capsaicin application on the skin. These data are in agreement with other studies that have shown that brimonidine inhibited temperature-induced cutaneous vasodilation [28], demonstrating that the vasoconstrictor activity of brimonidine in human skin vessels is of potential direct relevance to the treatment of erythema in rosacea. Perivascular infiltration was observed on some of the histological sections (such as the one shown in Fig. 1), which could be due to the surgical operation (abdominoplasty) or an existing skin inflammation. However, this was not observed in all of the sections and importantly was not restricted to the capsaicin treatment only and was seen across all of the groups.
Fig. 6. Effect of brimonidine 0.2% in the AA-induced inflammatory mouse model (A) and the TPA-induced inflammatory mouse model (B). ** p < 0.01.
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In addition to the vasoconstrictive properties demonstrated, we also sought to determine the anti-inflammatory properties of brimonidine. In both of the in vivo inflammation models used, the AA-induced and TPA-induced ear inflammation models, brimonidine 0.2% displayed significantly reduced levels of edema when compared to the control group. The reduction of edema in rosacea is a critical requirement for any new treatment due to the presence of edema in almost all histology of ETR [29] and due to the close association of inflammation with edema [7]. In a recent study, brimonidine was also shown to suppress vasodilation in the UVB-induced mouse ear inflammation model, which suggests the anti-inflammatory effect may result from vasoconstriction [30]. In summary, these studies suggest that brimonidine would be beneficial in the treatment of facial erythema in rosacea due to the demonstrated vasoconstrictor activity and potential anti-inflammatory effect, both of which are thought to have direct relevance to the pathophysiology of facial erythema of rosacea. The high selectivity of brimonidine demonstrated here, in targeting of small vessels and absence of off-target activity, is also highly desirable from a safety aspect in the context of daily chronic treatment of the disease. Acknowledgements The authors would like to thank Fabrice Duprat for his scientific assistance. We thank Dr S. Boisnic, MD and Marie-Christine Branchet (GREDECO SA, la salpetrie`re, Paris) for evaluation of Mirvaso1 (Galderma R&D, Sophia Antipolis, France) on their organoculture model. Funding for the study was provided by Galderma R&D, Sophia Antipolis, France. Medical writing support was provided by Gavin Kenny, PhD and Natasha Singh Kent, PhD at Havas Life Medicom and was funded by Galderma R&D, Sophia Antipolis, France. Appendix A. Supplementary data Supplementary data associated with this article can be found, in the online version, at http://dx.doi.org/10.1016/j.jdermsci.2014.04.002. References [1] Crawford GH, Pelle MT, James WD, Rosacea: I. Etiology, pathogenesis, and subtype classification. J Am Acad Dermatol 2004;51:327–41. [2] Wilkin J, Dahl M, Detmar M, Drake L, Feinstein A, Odom R, et al. Standard classification of rosacea: report of the National Rosacea Society expert committee on the classification and staging of rosacea. J Am Acad Dermatol 2002;46:584–7. [3] Wilkin J, Dahl M, Detmar M, Drake L, Liang MH, Odom R, et al. Standard grading system for rosacea: report of the National Rosacea Society expert committee on the classification and staging of rosacea. J Am Acad Dermatol 2004;50:907–12. [4] Odom R, Dahl M, Dover J, Draelos Z, Drake L, Macsai M, et al. Standard management options for rosacea, Part I: overview and broad spectrum of care. Cutis 2009;84:43–7. [5] Gupta AK, Chaudhry MM. Rosacea and its management: an overview. J Eur Acad Dermatol Venereol 2005;19:273–85. [6] Del Rosso JQ. Advances in understanding and managing rosacea: part 2. The central role, evaluation, and medical management of diffuse and persistent facial erythema of rosacea. J Clin Aesthet Dermatol 2012;5(3):26–36. [7] Steinhoff M, Buddenkotte J, Aubert J, Sulk M, Novak P, Scwab VD, et al. Clinical, cellular and molecular aspects in the pathophysiology of rosacea. J Investig Dermatol Symp Proc 2011;15:2–11.
[8] Wilkin JK, Rosacea:. Pathophysiology and treatment. Arch Dermatol 1994; 130:359–62. [9] Del Rosso JQ. Advances in understanding and managing rosacea: part 1: connecting the dots between pathophysiological mechanisms and common clinical features of rosacea with emphasis on vascular changes and facial erythema. J Clin Aesthet Dermatol 2012;5:16–25. [10] Fowler J, Jackson JM, Moore A, Jarratt M, Jones T, Meadows K, et al. Efficacy and safety of once-daily topical brimonidine tartrate gel 0.5% for the treatment of moderate to severe facial erythema of rosacea: results of two randomized, double-blind, vehicle-controlled pivotal studies. J Drugs Dermatol 2013; 12(6):650–6. [11] Del Rosso JQ, Gallo RL, Tanghetti E, Webster G, Thiboutot. An evaluation of potential correlations between pathophysiologic mechanisms, clinical manifestations, and management of rosacea. Cutis 2013;91(3S):1–8. [12] Rosina P, Zamperetti MR, Giovannini A, Chieregato C, Girolomoni G. Videocapillaroscopic alterations in erythematotelangiectatic rosacea. J Am Acad Dermatol 2006;54:100–4. [13] Schwab VD, Sulk M, Seeliger S, Nowak P, Aubert J, Mess C, et al. Neurovascular and neuroimmune aspects in the pathophysiology of rosacea. J Investig Dermatol Symp Proc 2011;15:53–62. [14] Yamasaki K, Di Nardo A, Bardan A, Murakami M, Ohtake T, Coda A, et al. Increased serine protease activity and cathelicidin promotes skin inflammation in rosacea. Nat Med 2007;13(August):975–80. [15] Helfrich YR, Varani J, Fisher GJ, Chubb H, Hammerberg C, Bruce A, et al. Mast cell and neuropeptide expression are increased in erythematotelangiectatic rosacea in comparison to telangiectatic photoaging. In: Poster presented at: Society for Investigative Dermatology Annual Meeting; 2012. p. P093. [16] Tan SG, Cunliffe WJ. Rosacea and migraine. Br Med J 1976;3(January):21. [17] Spoendlin J, Voegel JJ, Jick SS, Meier CR. Migraine, triptans, and the risk of developing rosacea: a population-based study within the United Kingdom. J Am Acad Dermatol 2013;69(September (3)):399–406. [18] Shanler SD, Ondo AL. Successful treatment of the erythema and flushing of rosacea using a topically applied selective alpha1-adrenergic receptor agonist, oxymetazoline. Arch Dermatol 2007;143(11):1369–71. [19] Bikowski J. Rosacea therapy: current approaches and future directions. Pract Dermatol 2012;(July):31–2. [20] Fowler J, Jarratt M, Moore A, Meadows K, Pollack A, Steinhoff M, et al. Oncedaily topical brimonidine tartrate gel 0.5% is a novel treatment for moderate to severe facial erythema of rosacea: results of two multicentre, randomized and vehicle-controlled studies. Br J Dermatol 2012;166:633–41. [21] Rahman MQ, Ramaesh K, Montgomery DNI. Brimonidine for glaucoma. Expert Opin Drug Saf 2010;9:483–91. [22] Serle JB. A comparison of the safety and efficacy of twice daily brimonidine 0.2% versus betaxolol 0.25% in subjects with elevated intraocular pressure. The Brimonidine Study Group III. Surv Ophthalmol 1996;41(Suppl 1): S39–47. [23] Katz LJ. Brimonidine tartrate 0.2% twice daily vs timolol 0.5% twice daily: 1year results in glaucoma patients. Brimonidine study group. Am J Ophthalmol 1999;127:20–6. [24] Cantor LB. The evolving pharmacotherapeutic profile of brimonidine, an alpha 2-adrenergic agonist, after four years of continuous use. Expert Opin Pharmacother 2000;1(4):815–34. [25] Mirvaso1 Prescribing Information. [26] Moore A, Kempers S, Murakawa G, et al. Long-term safety and efficacy of oncedaily topical brimonidine tartrate gel 0.5% for the treatment of moderate to severe facial erythema of rosacea: results of a 1-year open-labeled study. J Drugs Dermatol 2014;13(1):56–61. [27] Huang XP, Setola V, Yadav PN, Allen JA, Rogan SC, Hanson BJ, et al. Parallel functional activity profiling reveals valvulopathogens are potent 5-hydroxytryptamine2B receptor agonists: implications for drug safety assessment. Mol Pharmacol 2009;76:710–22. [28] Hsia E, Tian M, Santori E, Mistry D, Gil D. The alpha adrenergic receptor agonist oxymetazoline inhibits temperature-induced vasodilation and LL-37 induced skin inflammation. J Investig Dermatol 2012;132:S1–8. [29] Cribier B. Rosacea under the microscope: characteristic histological findings. J Eur Acad Dermatol Venereol 2013;27(November (11)):1336–43. [30] Tian M, Hsia E, Andrews-Jones L, Manley K, Azartash K, Cheevers C, et al. The alpha adrenergic receptor agonist oxymetazoline decreases erythema and inflammation in a UVB-induced sunburn model. J Investig Dermatol 2013; 133:S209–21.
Please cite this article in press as: Piwnica D, et al. Vasoconstriction and anti-inflammatory properties of the selective a-adrenergic receptor agonist brimonidine. J Dermatol Sci (2014), http://dx.doi.org/10.1016/j.jdermsci.2014.04.002
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Vasoconstriction and anti-inflammatory properties of the selective a-adrenergic receptor agonist brimonidine David Piwnica *, Carine Rosignoli, Se´verine Thibaut de Me´nonville, Thierry Alvarez, Marlene Schuppli Nollet, Olivier Roye, Andre´ Jomard, Je´roˆme Aubert Research, Galderma, Sophia Antipolis, France
A R T I C L E I N F O
A B S T R A C T
Article history: Received 23 January 2014 Received in revised form 1 April 2014 Accepted 3 April 2014
Background: The facial erythema of rosacea is recognized as the most prevalent and most difficult manifestation of rosacea to treat. A recent approach in patients with rosacea has been to reduce this erythema through vasoconstriction of cutaneous blood vessels by selectively targeting a2-adrenergic receptors with brimonidine. Objective: To further investigate the pharmacodynamic profile of brimonidine, its vasoconstrictive effects and its anti-inflammatory properties. Methods: The potency for the a1A, a1B, a2A, a2B and a2C receptors of brimonidine was measured, as well as performing a large target profiling study in order to determine the target selectivity profile of brimonidine. The vasoconstrictive effects of brimonidine were measured using ex vivo wire myography and human skin biopsy neuroinflammation models. The anti-inflammatory properties of brimonidine were measured using two in vivo mice ear inflammation models. Results: Brimonidine was found to be highly selective for the a2A adrenoreceptor (EC50 0.45 nM) over the other a-adrenoreceptors. Additionally, the large target profiling study demonstrated the high selectivity of brimonidine with minimal off-target effects. The ex vivo wire myography model showed that brimonidine is a potent vasoconstrictor of human subcutaneous vessels with a diameter of less than 200 mm (EC50 0.4 nM). The ex vivo human skin biopsy neuroinflammation model demonstrated that brimonidine completely inhibited vasodilation induced by capsaicin. Both in vivo mouse ear inflammation models highlighted that brimonidine inhibited ear edema (up to 76%) when compared to vehicle. Conclusion: The selectivity, vasoconstrictive and anti-inflammatory properties of brimonidine that have been described in these studies are in agreement with the benefits observed with this compound in the treatment of facial erythema in rosacea. ß 2014 Published by Elsevier Ireland Ltd on behalf of Japanese Society for Investigative Dermatology.
Keywords: Rosacea Vascular Inflammation Brimonidine Mirvaso Pharmacology
1. Introduction Rosacea is a chronic skin disorder with an unknown etiology, characterized by transient or persistent erythema, telangiectasia, papules and pustules, that primarily affects the central facial region (cheeks, nose, chin and forehead) [1–3]. Persistent facial erythema is most commonly associated with subtypes I and II of rosacea, namely erythematotelangiectatic (ETR) and papulopustular rosacea (PPR) [2,3]. Rosacea, in particular facial erythema, is often triggered by a wide range of environmental or lifestyle factors including, but not limited to: sun exposure, emotional
* Corresponding author. Tel.: +33 1492383026. E-mail address: [email protected] (D. Piwnica).
stress, temperature, exercise and alcohol consumption [4]. The facial erythema of rosacea is recognized to be the most prevalent and also the most difficult manifestation to treat [4–6]. The pathophysiology of facial erythema of rosacea is relatively unknown. It is suggested to result from dysregulation of cutaneous vasomotor responses, which causes abnormal, involuntary and persistent dilation of facial blood vessels [7–10]. The severity of rosacea has also been linked to prolonged vasodilation of small blood vessels (<200 mm), with evidence to suggest that vasodilation may result from inflammatory mediators released during the early phase of rosacea [7,9,11,12]. These characteristics of rosacea are similar in nature to the main features of neurogenic inflammation, a condition caused by the local release of inflammatory mediators from sensory neurons [13]. The link between rosacea and neurogenic inflammation is further strengthened by a
http://dx.doi.org/10.1016/j.jdermsci.2014.04.002 0923-1811/ß 2014 Published by Elsevier Ireland Ltd on behalf of Japanese Society for Investigative Dermatology.
Please cite this article in press as: Piwnica D, et al. Vasoconstriction and anti-inflammatory properties of the selective a-adrenergic receptor agonist brimonidine. J Dermatol Sci (2014), http://dx.doi.org/10.1016/j.jdermsci.2014.04.002
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proteases and 5 phosphodiesterases). The full assay list can be found in the supplementary information. Brimonidine response was normalized to the maximum signal detected in each assay.
number of recent studies that demonstrate the release of vasoactive peptides by sensory neurones in skin upon exposure to known triggers [7,11,14], and co-localization between facial sensory nerve endings and blood vessels in patients with rosacea [7,11,13]. One recent study has shown an increase of markers, which are also present in neurogenic inflammation such as mast cells, neuropeptides (substance P and CGRPa) as well as matrix remodeling, in patients with rosacea [15]. In addition, patients with rosacea have been associated with a higher prevalence of migraine, a condition also thought to involve neurogenic inflammation [16,17]. Neurovascular and neuroimmune interactions in rosacea have revealed the involvement of a number of genes including adrenergic receptors [13]. Novel approaches are being assessed for the treatment of facial erythema of rosacea. One recent approach has been to reduce erythema through vasoconstriction of cutaneous blood vessels by targeting both a-adrenergic receptors with oxymetazoline [18,19] and brimonidine [10,20]. Oxymetazoline is an agonist of the a1Aadrenoceptor and a partial agonist of the a2A-receptor agonist; it is currently in clinical development (Phase IIb) for the treatment of facial erythema in rosacea [18,19]. Brimonidine is a highly selective a2-adrenergic receptor agonist with potent vasoconstrictive activity that has been approved for the treatment of openangle glaucoma for almost 20 years [21–24]. Brimonidine has also recently been approved as a promising, new therapy for the topical treatment of persistent (nontransient) facial erythema of rosacea in adults 18 years of age or older (Mirvaso1, Galderma R&D, Sophia Antipolis, France) [25]. Recent Phase II and III studies with topical brimonidine 0.33% gel (once daily) demonstrated its efficacy and safety [10,20] for periods of up to one year [26]. The aim of this study was to further investigate the pharmacodynamic profile of brimonidine, its vasoconstrictive effects on subcutaneous arteries and veins and its anti-inflammatory properties in vivo, in comparison with oxymetazoline.
Subcutaneous blood vessels were isolated from human abdominal plastic surgery. Healthy human skin was placed in cold physiological saline solution (PSS) and stored at 4 8C until required. The subcutaneous blood vessels were removed from the skin using forceps and scissors under stereomicroscope and maintained in PSS at pH = 7.4. Blood vessels were cut into ring segments, roughly 2 mm in length, and stored in cold PSS. The 2 mm-segments were then mounted using two fine tungsten wires (40 mm in diameter) into a four chamber myograph (EMKABath4, EMKA Technologies). Data were recorded using the Iox 2.8 software. The vessel segments were then stretched to the tension corresponding to a physiological pressure of 90 mmHg for arteries and 18 mmHg for veins, and lumen diameter was measured using Normalize software from EMKA Technologies. Vessel segments were maintained in a preheated PSS solution gassed with a mixture of 5% CO2, 20% O2 and 75% N2 (pH = 7.4) for 1 h before being used. During this period, the bath solution was changed every 20 min. The viability of blood vessel segments was assessed by measuring the contraction obtained after two successive stimulations with the PSS solution containing 80 mM of KCl. Then, the smooth muscle vascular wall function was evaluated using a vasoconstrictor compound such as prostaglandin F2a at 10 mM, whereas the endothelium function was assessed using acetylcholine at 1 mM. Oxymetazoline and brimonidine were dissolved in DMSO. Both compounds were added in a range of concentrations directly to the chamber and the modulation of vascular tones was recorded.
2. Materials and methods
2.3. Ex vivo human skin model
2.1. Target-based assays
Healthy human skin was obtained from 10 different donors who underwent abdominoplasty. Skin biopsies (1 cm2) were obtained and placed in culture with the epithelium on top using an insert with 0.45 mm polycarbonate membrane (Fig. 1). Inserts were then set in 12-well plates pre-filled with culture medium (DMEM supplemented with FCS and antibiotics). Thus, this ex vivo organ culture system maintained the skin at the air–liquid interface and fed the dermis and epidermis by nutrient diffusion across the insert. Biopsies were then placed in a humidified atmosphere of 95% air and 5% CO2 at 37 8C. 20 mL/cm2 of 0.33% of brimonidine (Mirvaso1, Galderma R&D, Sophia Antipolis, France, 0.33 g brimonidine in 100 mL of gel equivalent to 11.3 mM) or vehicle gel (corresponding vehicle gel to Mirvaso1, Galderma R&D, Sophia Antipolis, France), was added topically to the skin biopsies 2 hours before addition of capsaicin and capsaicin was added at 10 mM to the culture medium for 4 h. Skin
An adrenoreceptor assay selectivity profile study was conducted at Cerep (Poitiers, France) with human a1A (Cerep Cat 1500), a1B (Cerep Cat 1901), a2A (Cerep Cat 2A2558), a2B (Cerep Cat 1813) and a2C (Cerep Cat 1736) receptors selected to evaluate the EC50 potency of oxymetazoline (Sigma–Aldrich, France) and brimonidine (Sigma–Aldrich, France). Both compounds were dissolved in DMSO then diluted in HEPES sodium salt buffer. Both compounds tested over a dose range of 1 pM to 10 mM. A large target profiling study of brimonidine (Bioprint1 profile) was performed at Cerep (Poitiers, France) with the panel mainly based on target diversity. The study included 104 binding assays (non-peptide, peptide and nuclear receptors, ion channels and amine transporters) and 32 enzyme assays (including 10 kinases, 9
2.2. In vitro wire myography
Fig. 1. Schematic diagram of the ex vivo human skin model used to measure the vasoconstrictor activity of brimonidine. Black arrowheads show vasodilation induced by capsaicin application.
Please cite this article in press as: Piwnica D, et al. Vasoconstriction and anti-inflammatory properties of the selective a-adrenergic receptor agonist brimonidine. J Dermatol Sci (2014), http://dx.doi.org/10.1016/j.jdermsci.2014.04.002
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biopsies were then fixed in formol liquid and embedded in paraffin for histological analyses. After staining the histological section with hematoxylin and eosin, the surface area covered by the lumen of the dilated vessels (mm2) and the percentage of vessels dilated were calculated. Using the image analysis software Image-Pro Plus (Media Cybernetics, Inc., MD, USA), 16 areas were observed on the histological section. An average of 50 vessels were analyzed across all the 16 sections. For statistical analysis, this experiment was repeated on biopsies from 10 donors and the statistical analysis was conducted using a reduced deviation Student’s t-test or the paired ttest with a risk of 5%. 2.4. In vivo inflammation model Female BALB/cByJ Rj mice were purchased from JANVIER (France). All experiments were conducted at the Unite´ d’Evaluation de Produits Pharmacologiques (UDEPP, France) under specific pathogen-free conditions with mice between 8 and 9 weeks of age. All experiments were conducted with the approval of and in accordance with the guidelines for animal experiments of the local ethics committee (CEEA, France). Arachidonic acid (Sigma–Aldrich, France) was dissolved in tetrahydrofuran/methanol (1:1) at 4% (AA-induced skin inflammation) and 12-O-Tetradecanoylphorbol-13-acetate (Sigma– Aldrich, France) was dissolved in ethanol at 0.01% (TPA-induced skin inflammation). Brimonidine (Sigma–Aldrich, France) was added to the AA or TPA solution to achieve a final concentration of 0.2% (0.2 g in 100 mL of solution equivalent to 6.9 mM), which was the highest concentration possible due to solubility reasons. 20 ml of AA or TPA solution, with or without brimonidine 0.2%, was applied topically on the internal side of the mice ear. Ear thickness was measured using a micrometer 1 h or 6 h after brimonidine application for the AA-induced skin inflammation and TPAinduced skin inflammation models respectively.
Fig. 2. Target profile of brimonidine at 10 mM on a broad panel of 141 biological targets confirmed the high selectivity for a-adrenergic receptors and especially a2 subtypes. For full size graph see supplementary information.
3. Results 3.1. Brimonidine has high selectivity for a-adrenergic receptors with minimal off-target activity The selectivity of both brimonidine and oxymetazoline for the adrenoreceptors was determined by measuring the EC50 values for the a1A, a1B, a2A, a2B and a2C receptors. Brimonidine was found to be mainly selective for the a2A receptor, with an EC50 of 0.45 nM. In comparison, oxymetazoline was found to be less selective with an EC50 of 1.1 nM and 0.33 nM on a2A and a1A receptors respectively (Table 1). A large target profiling study was performed, including binding assays (non-peptide, peptide and nuclear receptors, ion channels and amine transporters) and enzyme assays (kinases, proteases and phosphodiesterases) to determine the selectivity of brimonidine. A compound with an effect greater than 30% at 10 mM is considered as active against the target. Brimonidine was found to be highly selective for a-adrenergic receptors, in particular a2 subtypes, with virtually no off-target activity seen, as shown by the percentage of response demonstrated at 10 mM on a broad panel of targets (Fig. 2 and supplementary information). Table 1 Agonist activity of brimonidine and oxymetazoline on human a-adrenergic receptors.
Oxymetazoline EC50 (nM) Brimonidine EC50 (nM)
a1A
a1B
a2A
a2B
a2C
0.33 56
N.C.a N.C.a
1.1 0.45
240 1900
5.9 6.5
a N.C., not calculable (concentration–response curve shows less than 25% effect at the highest validated testing concentration).
3.2. Wire myography demonstrates that brimonidine is a potent vasoconstrictor of sub 200 mm human subcutaneous blood vessels The wire myography technique was used to evaluate the functional responses and vascular reactivity of various sized isolated subcutaneous arteries when subjected to either brimonidine or oxymetazoline. Oxymetazoline was found to have EC50 values of 2 nM, 31 nM, 5.1 nM and 4.5 nM for the 184 mm, 433 mm, 513 mm and 630 mm diameter vessels, respectively (Fig. 3A–D). Brimonidine was found to be 5 times more potent than oxymetaxoline as a vasoconstrictor on human subcutaneous arteries smaller than 200 mm diameter, with an EC50 of 0.4 nM for the 184 mm vessels (Fig. 3A). The potency and efficacy of brimonidine was found to decrease as the vessel diameter increased, with EC50 values reaching 490 nM and 50% of relative efficacy (compared to oxymetazoline) for vessels of 630 mm in diameter, displaying its selectivity for the sub 200 mm vessels only (Fig. 3D). In addition, the activity of brimonidine was compared on similar sized arteries and veins (222 mm and 210 mm, respectively). Brimonidine was shown to be active on both types of subcutaneous blood vessels with EC50 values of 2.9 nM and 24 nM on arteries and veins respectively (Fig. 4). 3.3. Topical application of brimonidine significantly reduces vasodilation on ex vivo human skin biopsies after capsaicin application An ex vivo human skin model was used to assess the activity of brimonidine on subcutaneous vessel dilation induced by capsaicin,
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Fig. 3. Effect of brimonidine on human subcutaneous vessels using wire myography technique. Arteries of 184 mm had EC50 values of 0.4 nM and 2 nM for brimonidine and oxymetazoline respectively (A). Arteries of 433 mm had EC50 values of 62 nM and 31 nM for brimonidine and oxymetazoline respectively (B). Arteries of 513 mm had EC50 values of 190 nM and 5.1 nM for brimonidine and oxymetazoline respectively (C). Arteries of 630 mm had EC50 values of 490 nM and 4.5 nM for brimonidine and oxymetazoline respectively (D).
a known trigger of neurogenic inflammation [7]. Topical treatment with brimonidine 0.33% (Mirvaso1, Galderma R&D, Sophia Antipolis, France) resulted in a statistically significant almost complete reduction, compared to the gel vehicle, in the surface area occupied by the blood vessels following capsaicin-induced
vasodilation, (66.7 mm2 vs. 180.5 mm2; p < 0.001) (Fig. 5A). This finding was mirrored in the percentage of dilated vessels, with a significantly reduced percentage of dilated vessels in the brimonidine treated group, when compared to vehicle (Fig. 5B). 3.4. In vivo inflammation is reduced with brimonidine application Two mouse models of inflammation were used to measure the anti-inflammatory effect of brimonidine in an in vivo setting. Arachidonic acid (AA)-induced ear edema was inhibited by 76% with topical administration of brimonidine 0.2% (Fig. 6A) and TPAinduced ear edema was inhibited by 50% by topical administration of brimonidine 0.2% (Fig. 6B). 4. Discussion
Fig. 4. Dose–response effect of brimonidine on the vascular tone of human subcutaneous veins and arteries. Veins of 210 mm and arteries of 222 mm were found to have EC50 values of 24 nM and 2.9 nM, respectively.
Facial erythema in rosacea is thought to be the result of vasodilation of the cutaneous vasculature in the central facial region [7–10], with vessel diameters lower than 200 mm in rosacea patients [12]. The similarities between rosacea and inflammation have recently been demonstrated in numerous studies including the observed cytokine, chemokine and cellular infiltration during intermittent flares [9,11,12]; the marked inflammatory infiltrate of CD4+ T lymphocytes, macrophages and mast cells seen in ETR [7]; the release of vasoactive peptides [7,11,14]; and the increased expression of vasoactive peptides calcitonin gene-related peptide [CGRP-a] and substance P [15]. Further supporting evidence of a
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Fig. 5. Effect of brimonidine 0.33% on surface area (A) and percentage (B) of dilated vessels in the capsaicin-induced vasodilation ex vivo human skin model. ***p < 0.001, for capsaicin with vehicle compared to: control, vehicle and capsaicin.
link between rosacea and neuroinflammation is highlighted by the co-localization between facial sensory nerve endings and blood vessels in patients with rosacea [11,13] and the higher prevalence of migraine in patients with rosacea [16,17]. These studies combined with a recent study displaying the involvement of a number of adrenergic receptors [13] have suggested the potential of using a highly specific a-adrenergic receptor agonist with vasoconstrictive and anti-inflammatory properties to treat the facial erythema of rosacea. Brimonidine 0.33% (Mirvaso1, Galderma R&D, Sophia Antipolis, France) [25] has recently received FDA approval for the treatment of facial erythema in rosacea [6]. In this paper, we have described the pharmacodynamic, ex vivo vasoconstrictor activity and in vivo anti-inflammatory effects of brimonidine. The profiling of brimonidine in more than 200 targets confirmed that it has very high selectivity toward the a2 adrenergic receptors. Moreover, the profiling data revealed that brimonidine was inactive against the 5-hydroxytryptamine 2B receptor (5HT2b), reported to be involved in valvular heart disease after long-term treatment, opposed to other vasoconstrictors such as oxymetazoline [27]. In addition, we demonstrated that brimonidine was more potent and selective than oxymetazoline on small subcutaneous arteries (<200 mm) using the wire myography technique. This is of particular importance in facial erythema as the higher selectivity exhibited by brimonidine for targeting the smaller vessels is of notable benefit as one previous study showed blood vessel
diameters lower than 200 mm are involved in rosacea [12]. Studies of the a2 receptor have suggested that a2A, a2B and a2D subtypes are of particular importance in arteries, and that a2A, a2C and a2D subtypes are of more importance on the venous side [18]. The importance of the a2A receptor subtype on both arteries and veins was demonstrated in this study by the ability of brimonidine to initiate vasoconstriction of both arteries and veins. Targeting of the venous side is of real potential in the treatment of facial erythema as it has previously been shown that one of the first abnormalities to manifest in patients with rosacea is a microcirculatory disturbance of facial angular veins [12]. In the human skin biopsy model, a model of neurogenic inflammation, brimonidine 0.33% (Mirvaso1, Galderma R&D, Sophia Antipolis, France) was shown to have vasoconstrictive properties that negated the effect of capsaicin application on the skin. These data are in agreement with other studies that have shown that brimonidine inhibited temperature-induced cutaneous vasodilation [28], demonstrating that the vasoconstrictor activity of brimonidine in human skin vessels is of potential direct relevance to the treatment of erythema in rosacea. Perivascular infiltration was observed on some of the histological sections (such as the one shown in Fig. 1), which could be due to the surgical operation (abdominoplasty) or an existing skin inflammation. However, this was not observed in all of the sections and importantly was not restricted to the capsaicin treatment only and was seen across all of the groups.
Fig. 6. Effect of brimonidine 0.2% in the AA-induced inflammatory mouse model (A) and the TPA-induced inflammatory mouse model (B). ** p < 0.01.
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In addition to the vasoconstrictive properties demonstrated, we also sought to determine the anti-inflammatory properties of brimonidine. In both of the in vivo inflammation models used, the AA-induced and TPA-induced ear inflammation models, brimonidine 0.2% displayed significantly reduced levels of edema when compared to the control group. The reduction of edema in rosacea is a critical requirement for any new treatment due to the presence of edema in almost all histology of ETR [29] and due to the close association of inflammation with edema [7]. In a recent study, brimonidine was also shown to suppress vasodilation in the UVB-induced mouse ear inflammation model, which suggests the anti-inflammatory effect may result from vasoconstriction [30]. In summary, these studies suggest that brimonidine would be beneficial in the treatment of facial erythema in rosacea due to the demonstrated vasoconstrictor activity and potential anti-inflammatory effect, both of which are thought to have direct relevance to the pathophysiology of facial erythema of rosacea. The high selectivity of brimonidine demonstrated here, in targeting of small vessels and absence of off-target activity, is also highly desirable from a safety aspect in the context of daily chronic treatment of the disease. Acknowledgements The authors would like to thank Fabrice Duprat for his scientific assistance. We thank Dr S. Boisnic, MD and Marie-Christine Branchet (GREDECO SA, la salpetrie`re, Paris) for evaluation of Mirvaso1 (Galderma R&D, Sophia Antipolis, France) on their organoculture model. Funding for the study was provided by Galderma R&D, Sophia Antipolis, France. Medical writing support was provided by Gavin Kenny, PhD and Natasha Singh Kent, PhD at Havas Life Medicom and was funded by Galderma R&D, Sophia Antipolis, France. Appendix A. Supplementary data Supplementary data associated with this article can be found, in the online version, at http://dx.doi.org/10.1016/j.jdermsci.2014.04.002. References [1] Crawford GH, Pelle MT, James WD, Rosacea: I. Etiology, pathogenesis, and subtype classification. J Am Acad Dermatol 2004;51:327–41. [2] Wilkin J, Dahl M, Detmar M, Drake L, Feinstein A, Odom R, et al. Standard classification of rosacea: report of the National Rosacea Society expert committee on the classification and staging of rosacea. J Am Acad Dermatol 2002;46:584–7. [3] Wilkin J, Dahl M, Detmar M, Drake L, Liang MH, Odom R, et al. Standard grading system for rosacea: report of the National Rosacea Society expert committee on the classification and staging of rosacea. J Am Acad Dermatol 2004;50:907–12. [4] Odom R, Dahl M, Dover J, Draelos Z, Drake L, Macsai M, et al. Standard management options for rosacea, Part I: overview and broad spectrum of care. Cutis 2009;84:43–7. [5] Gupta AK, Chaudhry MM. Rosacea and its management: an overview. J Eur Acad Dermatol Venereol 2005;19:273–85. [6] Del Rosso JQ. Advances in understanding and managing rosacea: part 2. The central role, evaluation, and medical management of diffuse and persistent facial erythema of rosacea. J Clin Aesthet Dermatol 2012;5(3):26–36. [7] Steinhoff M, Buddenkotte J, Aubert J, Sulk M, Novak P, Scwab VD, et al. Clinical, cellular and molecular aspects in the pathophysiology of rosacea. J Investig Dermatol Symp Proc 2011;15:2–11.
[8] Wilkin JK, Rosacea:. Pathophysiology and treatment. Arch Dermatol 1994; 130:359–62. [9] Del Rosso JQ. Advances in understanding and managing rosacea: part 1: connecting the dots between pathophysiological mechanisms and common clinical features of rosacea with emphasis on vascular changes and facial erythema. J Clin Aesthet Dermatol 2012;5:16–25. [10] Fowler J, Jackson JM, Moore A, Jarratt M, Jones T, Meadows K, et al. Efficacy and safety of once-daily topical brimonidine tartrate gel 0.5% for the treatment of moderate to severe facial erythema of rosacea: results of two randomized, double-blind, vehicle-controlled pivotal studies. J Drugs Dermatol 2013; 12(6):650–6. [11] Del Rosso JQ, Gallo RL, Tanghetti E, Webster G, Thiboutot. An evaluation of potential correlations between pathophysiologic mechanisms, clinical manifestations, and management of rosacea. Cutis 2013;91(3S):1–8. [12] Rosina P, Zamperetti MR, Giovannini A, Chieregato C, Girolomoni G. Videocapillaroscopic alterations in erythematotelangiectatic rosacea. J Am Acad Dermatol 2006;54:100–4. [13] Schwab VD, Sulk M, Seeliger S, Nowak P, Aubert J, Mess C, et al. Neurovascular and neuroimmune aspects in the pathophysiology of rosacea. J Investig Dermatol Symp Proc 2011;15:53–62. [14] Yamasaki K, Di Nardo A, Bardan A, Murakami M, Ohtake T, Coda A, et al. Increased serine protease activity and cathelicidin promotes skin inflammation in rosacea. Nat Med 2007;13(August):975–80. [15] Helfrich YR, Varani J, Fisher GJ, Chubb H, Hammerberg C, Bruce A, et al. Mast cell and neuropeptide expression are increased in erythematotelangiectatic rosacea in comparison to telangiectatic photoaging. In: Poster presented at: Society for Investigative Dermatology Annual Meeting; 2012. p. P093. [16] Tan SG, Cunliffe WJ. Rosacea and migraine. Br Med J 1976;3(January):21. [17] Spoendlin J, Voegel JJ, Jick SS, Meier CR. Migraine, triptans, and the risk of developing rosacea: a population-based study within the United Kingdom. J Am Acad Dermatol 2013;69(September (3)):399–406. [18] Shanler SD, Ondo AL. Successful treatment of the erythema and flushing of rosacea using a topically applied selective alpha1-adrenergic receptor agonist, oxymetazoline. Arch Dermatol 2007;143(11):1369–71. [19] Bikowski J. Rosacea therapy: current approaches and future directions. Pract Dermatol 2012;(July):31–2. [20] Fowler J, Jarratt M, Moore A, Meadows K, Pollack A, Steinhoff M, et al. Oncedaily topical brimonidine tartrate gel 0.5% is a novel treatment for moderate to severe facial erythema of rosacea: results of two multicentre, randomized and vehicle-controlled studies. Br J Dermatol 2012;166:633–41. [21] Rahman MQ, Ramaesh K, Montgomery DNI. Brimonidine for glaucoma. Expert Opin Drug Saf 2010;9:483–91. [22] Serle JB. A comparison of the safety and efficacy of twice daily brimonidine 0.2% versus betaxolol 0.25% in subjects with elevated intraocular pressure. The Brimonidine Study Group III. Surv Ophthalmol 1996;41(Suppl 1): S39–47. [23] Katz LJ. Brimonidine tartrate 0.2% twice daily vs timolol 0.5% twice daily: 1year results in glaucoma patients. Brimonidine study group. Am J Ophthalmol 1999;127:20–6. [24] Cantor LB. The evolving pharmacotherapeutic profile of brimonidine, an alpha 2-adrenergic agonist, after four years of continuous use. Expert Opin Pharmacother 2000;1(4):815–34. [25] Mirvaso1 Prescribing Information. [26] Moore A, Kempers S, Murakawa G, et al. Long-term safety and efficacy of oncedaily topical brimonidine tartrate gel 0.5% for the treatment of moderate to severe facial erythema of rosacea: results of a 1-year open-labeled study. J Drugs Dermatol 2014;13(1):56–61. [27] Huang XP, Setola V, Yadav PN, Allen JA, Rogan SC, Hanson BJ, et al. Parallel functional activity profiling reveals valvulopathogens are potent 5-hydroxytryptamine2B receptor agonists: implications for drug safety assessment. Mol Pharmacol 2009;76:710–22. [28] Hsia E, Tian M, Santori E, Mistry D, Gil D. The alpha adrenergic receptor agonist oxymetazoline inhibits temperature-induced vasodilation and LL-37 induced skin inflammation. J Investig Dermatol 2012;132:S1–8. [29] Cribier B. Rosacea under the microscope: characteristic histological findings. J Eur Acad Dermatol Venereol 2013;27(November (11)):1336–43. [30] Tian M, Hsia E, Andrews-Jones L, Manley K, Azartash K, Cheevers C, et al. The alpha adrenergic receptor agonist oxymetazoline decreases erythema and inflammation in a UVB-induced sunburn model. J Investig Dermatol 2013; 133:S209–21.
Please cite this article in press as: Piwnica D, et al. Vasoconstriction and anti-inflammatory properties of the selective a-adrenergic receptor agonist brimonidine. J Dermatol Sci (2014), http://dx.doi.org/10.1016/j.jdermsci.2014.04.002