Tacrolimus: Title and subTitle BreakThe Drug for the Turn of the Millennium? FREE

IN DERMATOPHARMACOLOGY, there is a large demand for alternative noncorticosteroidal anti-inflammatory drugs. Although glucocorticosteroids represent the mainstay of therapy in inflammatory skin disorders, their clinical use is limited by systemic and local side effects, such as diabetes mellitus, osteoporosis, skin atrophy, telangiectasia, and striae cutis distensae. In the race to develop potent alternative immunomodulatory agents, considerable emphasis has been directed toward more selective drugs, such as macrolide lactones. Lead compounds of this class are sirolimus and tacrolimus, which inhibit the interleukin (IL) 2 gene expression in T cells.¹

Tacrolimus is the prototype of a class of topical immunosuppressive agents with a great potential in the treatment of inflammatory skin diseases, atopic dermatitis above all.² Almost 5 decades after the introduction of glucocorticosteroids, this drug is the first one with proved topical efficacy in atopic dermatitis, and successors and competitors are likely to follow at a rapid pace. Furthermore, atopic dermatitis is unlikely to remain the only area of use. Other inflammatory and autoimmune dermatoses are awaiting results of clinical trials, and the list of indications is likely to grow in the near future. The introduction of oral cyclosporine and topical tacrolimus has marked the beginning of the era of immunosuppressive drugs that is likely to dominate dermatology in the years to come.

Although cyclosporine has revolutionized transplantation and the treatment of many other diseases, eg, psoriasis and autoimmune uveitis, its use is limited by side effects, such as nephrotoxic effects, hypertension, and neurologic dysfunction. However, cyclosporine lacks topical activity, whereas tacrolimus has been shown to penetrate the epidermal barrier and to suppress experimental contact hypersensitivity reactions in humans and animals.^{3 - 5} Furthermore, a randomized, double-blind, multicenter study recently demonstrated that tacrolimus ointment is highly effective in the treatment of atopic dermatitis.²

The aim of this review is to summarize the current knowledge regarding the therapeutic action of tacrolimus in dermatological diseases and to give an outlook for putative indications in the future.

In 1984, a tacrolimus-producing strain was isolated from the fermentation broth of a soil sample from Tsukuba, Japan, and defined as Streptomyces tsukubaensis.⁶ Chemically classified as a hydrophobic macrolide lactone, tacrolimus has a molecular weight of 822 daltons and exerts, in vitro, 10 to 100 times higher immunosuppressive activity than cyclosporine (Figure 1).¹

Tacrolimus has been shown to act at a point in activation of T lymphocytes that lies between T-cell receptor ligation and the transcription of early genes. The action of tacrolimus on signal transduction pathways has been studied extensively in vitro in T cells. During T-cell activation, antigen binds to its specific T-cell receptor, which results in an increase in intracellular Ca²⁺. Subsequently, Ca²⁺ binds to calmodulin and activates the phosphatase calcineurin. The activated calcineurin, in turn, may dephosphorylate the cytoplasmic (or preexisting) subunit of the nuclear factor of activated T cells (NF-AT). Only the dephosphorylated form of cytoplasmic NF-AT can translocate from the cytosol into the nucleus. Within the nucleus, cytoplasmic NF-AT forms a complex with the nuclear subunit of NF-AT, which then can bind to the promoter region of several cytokine genes (IL-2, IL-3, IL-4, granulocyte-macrophage colony-stimulating factor, tumor necrosis factor α) and induce cytokine gene transcription. After penetrating the cell membrane, tacrolimus binds to its intracellular receptor, the FK-binding proteins. This complex subsequently blocks the function of the Ca²⁺- and calmodulin-dependent phosphatase, calcineurin, which results in a suppression of NF-AT–dependent cytokine gene transcription and immunosuppression in vivo (Figure 2).^{1 ,7 - 9}

Mast cells are present in abundance in human skin, often in close anatomical relation to blood vessels and nerves. They play a role in the pathogenesis of various skin disorders, such as acute and chronic urticaria,¹⁰ psoriasis,¹¹ contact dermatitis,¹² and atopic dermatitis.¹³ Studies by de Paulis et al¹⁴ demonstrated that tacrolimus inhibited histamine release from skin mast cells activated by anti-IgE in a concentration-dependent manner, and also impaired the de novo synthesis of prostaglandin D₂. Moreover, Eberlein-König et al¹⁵ showed that tacrolimus diminished the enhancing effect of IL-3 on anti-IgE–induced histamine release from basophils. These observations may provide a clue to the understanding of the antipruritic action of tacrolimus.

Regarding the mechanisms of action of topical tacrolimus in atopic dermatitis, there is increasing evidence in vitro and in vivo that tacrolimus may interfere with epidermal cytokine networks (tumor necrosis factor α, IL-1, IL-8), T_H1/T_H2 imbalance (interferon γ, IL-4), costimulatory molecule (B7 family), and Fcϵ-receptor expression.^{16 - 19}

In addition to the suppression of T-cell activation, tacrolimus has been suggested to interfere with the immunopathogenesis of psoriasis by down-regulation of the proinflammatory mediator IL-8 and its specific receptor on keratinocytes.^{20 - 21} Furthermore, tacrolimus has been shown to increase the negative regulator of the cell cycle, p53, in vitro, which has been shown to be decreased in psoriatic skin when compared with skin from uninvolved areas.²² Induction of a cell-cycle block may thus represent a novel approach to treating hyperplastic skin diseases.

Earlier studies suggested that topical tacrolimus penetrated human skin in amounts exceeding those of topical cyclosporine.^{5 ,23 - 24} These findings prompted further clinical studies on the efficacy and safety of topical tacrolimus.³ Recently, in vitro dermal penetration studies on intact human skin have shown a relatively low percutaneous absorption of tacrolimus from ointments. The average rate of percutaneous penetration from 0.03%, 0.1%, and 0.3% tacrolimus ointments was 3.1, 4.9, and 6.8 ng/cm² per hour, respectively. However, the average rate of percutaneous penetration in damaged skin was 40 ng/cm² per hour, which is approximately 7-fold higher than on intact skin. Interestingly, occlusion did not affect percutaneous penetration of tacrolimus (N. Undre, PhD, oral communication, October 1997).

After systemic administration of tacrolimus, bile represents the principal pathway of elimination, with a total-body clearance of 2.25 L/h and a systemic elimination half-time of approximately 40 hours. Cytochrome P450 3A4 isoenzyme is responsible for the metabolism of tacrolimus.^{25 - 26} In vitro, 8 metabolites have been characterized so far, and a 13-O-demethylated metabolite has been shown to dominate in vitro and in vivo; however, it reveals a relative pharmacological activity of 6.4% compared with tacrolimus. In vitro studies on viable human skin report no evident cutaneous metabolism of tacrolimus.²⁷

The generic name tacrolimus is a neologism composed of Tsukuba macrolide immunosuppressive. Tacrolimus was first introduced for prevention of allograft rejection and is now in use in kidney, liver, and heart transplantation. Furthermore, tacrolimus exerts good therapeutic efficacy after systemic administration in pulmonary disorders such as severe asthma, and inflammatory bowel diseases such as Crohn disease. Recently, tacrolimus was shown to exert potent therapeutic effects in various dermatological diseases after both systemic and topical application (Table 1).

Psoriasis. During recent years, a variety of studies provided evidence that T cells play a crucial role in the immunopathogenesis of psoriasis. The disease shows association with particular HLA types.^{39 - 40} Moreover, histopathological findings demonstrate that the invasion of mononuclear cells, particularly activated T cells, precedes epidermal hyperplasia.⁴¹ Remission of psoriasis after bone marrow transplantation and anti-CD4 antibodies supports the concept that T cells play a pivotal role in its pathogenesis.^{42 - 43} Recently, Schön et al⁴⁴ published a murine psoriasis model based on the transfer of naive T cells, demonstrating for the first time that T cells are capable of generating a psoriatic phenotype.

Initial clinical experience with systemic tacrolimus therapy showed a dramatic resolution of severe recalcitrant psoriasis in 7 patients.²⁸ Subsequently, a placebo-controlled, double-blind, prospective, multicenter study was initiated. Fifty patients with severe recalcitrant plaque-type psoriasis were enrolled by the European FK 506 Multicentre Psoriasis Study Group.²⁹ After 9 weeks of a daily oral tacrolimus application of 0.10 to 0.15 mg/kg, tacrolimus-treated patients had a significantly greater reduction in the Psoriasis Area and Severity Index than did the placebo-treated ones. The mean blood trough level of tacrolimus throughout the entire study period was 10 ng/mL. In comparison, mean blood trough levels during topical therapy for atopic dermatitis ranged from 0.05 to 0.25 ng/mL in the majority of all treated patients.² Diarrhea, paresthesia, and insomnia were the most frequently reported side effects related to tacrolimus. Thus, systemic tacrolimus appears to be highly effective and well tolerated in the treatment of severe recalcitrant psoriasis.

Behçet Disease. Cyclosporine has been shown to be a potent drug for the treatment of Behçet disease.^{45 - 46} So far, 3 clinical studies demonstrated the systemic efficacy of tacrolimus in Behçet disease. This multisystem chronic disease is characterized by recurrent oral ulcerations and 2 of the following symptoms: aphthous recurrent genital ulceration, eye lesions (uveitis and vascular retinitis), and skin lesions (erythema nodosum, papulopustular lesions, and acneiform nodules).⁴⁷ Doses of 0.1 to 0.15 mg/kg have been shown to be effective in the resolution of mucocutaneous symptoms, ophthalmologic manifestations, and pulmonary complications.^{30 - 31}

Pyoderma Gangrenosum. A report about 4 male patients with recalcitrant pyoderma gangrenosum unresponsive to conventional therapy showed efficacy of systemic tacrolimus administration. Complete clinical remission and healing of disfiguring ulcers on the trunk and lower extremities was achieved in all 3 patients who continued to receive the drug at an oral dosage of 0.15 mg/kg twice a day.^{32 ,48}

Regarding the association of pyoderma gangrenosum with inflammatory bowel disease, it is of interest that oral tacrolimus was effective in 3 patients suffering from complicated Crohn disease. The treatment was started at doses of 0.15 to 0.20 mg/kg per day and adjusted to a whole-blood tacrolimus concentration range of 10 to 20 ng/mL. The drug was well absorbed in all patients and provided rapid clinical improvement.³³

Atopic Dermatitis. Atopic dermatitis is characterized by pruritic eczematous skin lesions in typical distribution, affecting about 10% of all children and 4.7% of all young adults living in industrialized countries.^{2 ,49} The underlying immunopathogenesis consists of activation of T cells in skin and an imbalance of T_H1/T_H2 cytokine production, hyperstimulating Langerhans cells, and B-cell IgE overproduction.^{50 - 52} Bruynzeel-Koomen et al⁵³ and Bieber et al⁵⁴ identified the presence of IgE on Langerhans cells in patients with atopic dermatitis, and subsequently it has been shown that this binding was mediated by Fcϵ receptors. In vitro studies showed that IgE receptor–allergen complexes on Langerhans cells aided in antigen processing and resulted in an up to 1000-fold increased antigen presentation to T cells.⁵⁵ Therefore, the induction of atopic dermatitis lesions may depend on the expression of Fcϵ receptors on Langerhans cells, the serum concentration of allergen-specific IgE, the number of skin-infiltrating allergen-specific T cells, and the exposure to allergen.⁵⁶

Recently, 1% tacrolimus ointment showed dramatic and quick therapeutic effects in the treatment of face and neck lesions in patients with atopic dermatitis. In keeping with this observation, the efficacy of 0.03%, 0.1%, and 0.3% tacrolimus ointment was assessed in an open trial in 50 patients with recalcitrant facial atopic dermatitis.^{34 - 35} In agreement with animal studies, all 3 concentrations showed substantial, dose-dependent improvement. Lichenified lesions, however, needed longer to clear up. Tacrolimus markedly improved pruritus within 3 days, and histopathologically T-cell and eosinophil infiltration was significantly reduced. Tacrolimus blood levels ranged from 0.09 to 0.70 ng/mL, and no significant changes in laboratory results were seen. The only adverse event was a mild transient skin irritation, which receded as the clinical symptoms improved.

The European Tacrolimus Multicenter Atopic Dermatitis Study Group reported topical tacrolimus to improve skin lesions in patients with atopic dermatitis in a randomized, double-blind, short-term trial of tacrolimus ointment.² Patients with moderate to severe atopic dermatitis according to the criteria of Rajka and Langeland⁵⁷ received topical application of 0.03%, 0.1%, or 0.3% tacrolimus ointment or vehicle twice a day for 3 weeks (Figure 3). Furthermore, a follow-up period of 2 weeks was included. The study end points were primarily the change in erythema, edema, and pruritus. In this controlled study, each of the 3 tacrolimus ointments was significantly more effective than the vehicle alone. There were no statistically significant differences among the active treatments, although there was a trend favoring 0.1% over 0.03% tacrolimus. A sensation of burning at the site of application has been reported to be the only adverse event in tacrolimus-compared with vehicle-treated skin sites.³ Thus, tacrolimus ointment seems to be highly effective in the treatment of atopic dermatitis with a potency comparable with that of potent glucocorticosteroids. Studies currently in progress in Europe, the United States, and Japan address the issues of long-term safety, use in children, and comparison with glucocorticosteroids. Registration of the drug for use in atopic dermatitis is expected during 1999 or 2000 in all of these countries.

Regarding allergic contact dermatitis, topical tacrolimus was reported to suppress challenge-induced contact hypersensitivity responses in 5 male human volunteers sensitized to dinitrochlorobenzene. Healthy skin was exposed to the immunosuppressant at doses of 0.01%, 0.1%, and 1% for 48 hours, and after removal of the patches, 0.1% dinitrochlorobenzene was placed on the same skin sites for another 24 hours. Topical tacrolimus suppressed dinitrochlorobenzene reactions in a dose-dependent manner. Histopathological examination of the vehicle-treated reaction site disclosed intense dermatitis, whereas an adjacent dinitrochlorobenzene-challenged skin site pretreated with 1% tacrolimus did not show inflammatory reactions. Furthermore, there were no systemic adverse effects.^{5 ,23}

Alopecia Areata and Hair Growth. Alopecia areata is believed to be an autoimmune disease in which a mononuclear cell infiltrate develops in and around anagen hair follicles and causes circumscript hair loss.⁵⁸ In the Dundee experimental bald rat model of alopecia areata, promising results were seen with topical immunotherapy. Although oral tacrolimus was ineffective in restoring hair, topical use of the drug led to hair growth at the site of application in all animals.³⁶ Moreover, in vitro studies showed that tacrolimus directly stimulates hair follicles.³⁷ Effects of topical tacrolimus on the normal hair cycle of C57BL/6J mice demonstrated the induction of anagen hair in telogen mouse skin in a dose-dependent manner. These findings suggest the possibility of clinical efficacy of topically applied tacrolimus in humans with alopecia areata.³⁸

The immunosuppressive drugs most commonly used for transplantation and treatment of autoimmune diseases are cyclosporine and tacrolimus. Both have similar nephrotoxic and vasoconstrictive properties that may limit their clinical use,^{59 - 60} although hypertension has been reported less frequently with tacrolimus during the first year after liver transplantation.⁶¹ Renal toxic effects of tacrolimus are clinically characterized by an increase in serum creatinine level, hyperkalemia, hypomagnesemia, reduction of the glomerular filtration rate, and tubular injury.⁶² So far, the underlying molecular mechanism has not been elucidated. The stimulating effect of tacrolimus on endothelin 1 release^{63 - 64} and the influence of transforming growth factor β1 on the pathogenesis of interstitial fibrosis⁶⁵ are investigated as putative mechanisms of tacrolimus-induced nephrotoxic effects. Katari et al⁶⁶ investigated acute reversible tacrolimus nephrotoxic effects by needle biopsy in 22 (17%) of 128 kidney transplant recipients receiving tacrolimus in a daily oral maintenance dose of 0.3 mg/kg. The diagnosis of tacrolimus nephrotoxic reaction was based on rigorous criteria, namely an increase in serum creatinine level requiring biopsy, absence of histological changes of acute rejection, and clinical response to reduction in the dose of tacrolimus. The highest whole-blood tacrolimus levels ranged from 14.5 to 50.5 ng/mL during the toxic episodes. A stepwise reduction in tacrolimus dosage led to an improved serum creatinine level within 1 to 14 days.⁶⁶

Initial experience with tacrolimus in pediatric renal transplant recipients who received the drug by oral administration at a daily dose of 0.3 mg/kg for a mean 12-month follow-up period showed hypertension as the most frequent side effect, in 85% of the young patients.⁶⁷ The therapeutic doses used in this study were twice as high as those used for the treatment of skin diseases.

Other side effects reported for systemic tacrolimus therapy include neuropsychiatric side effects, such as anxiety, nightmares, transient apraxia of speech, generalized tonic-clonic seizures, agitation, and acute delirium and tremor.^{68 - 69}

Furthermore, tacrolimus treatment is not related to impaired glucose tolerance or new-onset diabetes; however, serum levels of cholesterol and triglycerides increase significantly under tacrolimus and cyclosporine treatment.⁷⁰

Cutaneous adverse effects of topical tacrolimus have been examined in animals and humans. In animal studies (rats, rabbits, guinea pigs, albino rats, and dark Yucatan miniature swine), tacrolimus ointment was used at concentrations of 0.03% to 1% and for treatment periods of 28 days to 26 weeks. There were neither reports about cutaneous toxic effects, phototoxic effects, and irritation, nor about sensitization and contact dermatitis. The Draize test disclosed no injuries to corneal epithelium, and depigmentation could not be seen (I. Lawrence, MD, PhD, oral communication, October 1997).

Cumulative irritation studies and repeated insult patch testing on human skin with tacrolimus vs marketed dermatology products demonstrated nonsensitizing but slightly irritating potency of tacrolimus ointment. Theoretical concerns include nephrotoxic reactions, hypertension, and neurotoxic effects resulting from systemic absorption. However, for topical tacrolimus therapy in atopic dermatitis, the mean serum creatinine values remained at a constant level of 53 µmol/L (0.6 mg/dL). The ointment was applied to a defined, symptomatic area of 200 to 1000 cm² of skin for 3 weeks. The highest blood concentration of tacrolimus was 1.0 ng/mL for patients receiving 0.03%, 2.4 ng/mL for patients receiving 0.1%, and 4.9 ng/mL for patients receiving 0.3% tacrolimus ointment. A large proportion of the patients had values below the detection limit of the assay (0.05 ng/mL), and only a few individuals had tacrolimus concentrations higher than 1 ng/mL. The mean tacrolimus blood levels with the use of 0.3% ointment were 0.32 ng/mL on day 4 and 0.18 ng/mL on day 22. Considering that the therapeutic range for patients who received a transplant is 5 to 20 ng/mL, these findings indicate the relatively low risk of systemic toxic effects for topically applied tacrolimus.² Of major importance is the finding that topical tacrolimus induces no atrophogenic effects in contrast to glucocorticosteroids. In vitro tacrolimus did not affect keratinocyte proliferation,^{20 ,71} and recent studies showed that tacrolimus does not interfere with collagen synthesis.⁷²

In general, cyclosporine-responsive dermatoses may represent the most promising targets for topical tacrolimus therapy.⁷³

T-cell–mediated skin diseases, such as lichen planus, represent a potential area of use. Animal studies in mice with lupus hold promise that the drug might be beneficial in cutaneous autoimmune diseases.⁷⁴ Mast cell–mediated diseases represent another potential target for tacrolimus because of its potent effects on mast cell cytokine expression and mediator release.^{14 - 15} Cyclosporine and tacrolimus are lead substances; however, a number of other immunosuppressive agents, such as sirolimus, ascomycin, SDZ IMM 128, leflunomide, mycophenolate mofetil, and fumaric acid esters,^{75 - 76} are currently clinically evaluated for their use in dermatology.

In conclusion, experience in human studies has shown significant efficacy vs placebo and an excellent safety profile with few notable adverse effects and minimal risk of systemic toxic effects. Further trials including greater duration and extent of exposure, greater experience in children, and comparison of potencies between topical tacrolimus and the criterion standards of anti-inflammatory therapy, glucocorticosteroids, are needed to fulfill the promises of topical tacrolimus in dermatology.

Available evidence suggests that topical tacrolimus may open a new era in dermatopharmacology. However, only time will tell whether, at the beginning of the new millennium, atopic dermatitis will be redefined as a tacrolimus-deficiency state.

Accepted for publication July 30, 1998.

Reprints: Thomas Ruzicka, MD, Department of Dermatology, University of Düsseldorf, Moorenstr 5, D-40225 Düsseldorf, Germany.