1 11/8/00 Pharmacology of Tacrolimus Ointment William Fitzsimmons, Pharm.D. Vice President Drug Development Project Management Fujisawa Healthcare, Inc.

2 Pharmacology of Tacrolimus Ointment Topical macrolide (non-steroidal) immunomodulator Topical macrolide (non-steroidal) immunomodulator Inhibits calcineurin phosphatase activity in T-lymphocytes leading to decreased synthesis of cytokines associated with atopic dermatitis Inhibits calcineurin phosphatase activity in T-lymphocytes leading to decreased synthesis of cytokines associated with atopic dermatitis Decreases inflammatory mediator release from skin mast cells and basophils Decreases inflammatory mediator release from skin mast cells and basophils

3 Nonclinical Studies of Tacrolimus Ointment 27 studies conducted 27 studies conducted 3 chronic studies 3 chronic studies – 104-week topical carcinogenicity in B6C3F 1 mice – 52-week photocarcinogenicity in hairless mice – 52-week topical toxicity in micropigs

4 Topical Carcinogenicity Study in Mice 104 weeks (lifetime of animal) 104 weeks (lifetime of animal) No increase in skin tumors No increase in skin tumors Mice had 89 times higher systemic tacrolimus exposure than typical atopic dermatitis patients Mice had 89 times higher systemic tacrolimus exposure than typical atopic dermatitis patients Increased incidence of non- cutaneous lymphomas in 0.1% tacrolimus group Increased incidence of non- cutaneous lymphomas in 0.1% tacrolimus group

5 Photocarcinogenicity Study in Hairless Mice 52 week study with controlled UVR exposure 52 week study with controlled UVR exposure All animals develop skin tumors All animals develop skin tumors Median time to onset of tumor is the primary endpoint Median time to onset of tumor is the primary endpoint

6 Photocarcinogencity in Hairless Mice (combined male & female) MTO: Median Tumor Onset: mortality adjusted time at which one-half of the group has one or more tumors > 1 mm 2. Tumor Amplification Factor: relative influence of a test article on the response of skin to UVR exposure. Tacrolimus(%) MTO (Weeks) Tumor Amplification Factor UntreatedControl421.0Vehicle

7 Topical Toxicity Study in Micropigs 52 weeks 52 weeks Micropig (juvenile to adult) Micropig (juvenile to adult) Up to 3% tacrolimus ointment Up to 3% tacrolimus ointment Blood levels similar to humans Blood levels similar to humans 40% Body Surface Area treated 40% Body Surface Area treated No noteworthy topical or systemic findings No noteworthy topical or systemic findings

8 Nonclinical Studies Conclusions Tacrolimus is not a mutagen or carcinogen Tacrolimus is not a mutagen or carcinogen In a 2 year mouse study, there was no increase in skin tumors In a 2 year mouse study, there was no increase in skin tumors In mice, high skin permeability and high blood levels resulted in immunosuppression and lymphoma In mice, high skin permeability and high blood levels resulted in immunosuppression and lymphoma In a mouse model, median time to onset of photo- induced skin tumors is reduced with vehicle and active treatment In a mouse model, median time to onset of photo- induced skin tumors is reduced with vehicle and active treatment No noteworthy topical or systemic toxicity in micropigs No noteworthy topical or systemic toxicity in micropigs

9 Early Patch Testing in Humans Tacrolimus Ointment No evidence for: No evidence for: – Contact hypersensitivity – Phototoxicity – Photosensitization – Reduction of collagen synthesis in the skin

10 Pharmacokinetic Study -008 in Atopic Dermatitis Patients 0.3% tacrolimus ointment 0.3% tacrolimus ointment – Single application on days 1 and 8 – Twice daily application days 2 through 7 Protocol-defined area of application Protocol-defined area of application – Pediatric: 50 or 100 cm 2 – Adult: 100 to 5000 cm 2 Minimal systemic absorption (bioavailability < 0.5%) Minimal systemic absorption (bioavailability < 0.5%) No systemic accumulation No systemic accumulation

11 Phase III US Studies Distribution of Tacrolimus Blood Concentration 0.03% Tacrolimus Ointment (n=195)(n=25)

12 Phase II/III US Studies Blood Concentrations-Pediatric Patients (n=78) 0.03% Tacrolimus Ointment 87% 12% 1% 0%

13 Phase III US Studies Distribution of Tacrolimus Blood Concentration 0.1% Tacrolimus Ointment (n=193)(n=30)

14 Mean Blood Levels (ng/mL) 0.1% Tacrolimus Ointment Visit Week 1 Week 12 Month 6 Month 12 AdultMean N PediatricMean N2618--AdultMean N USEurope

15 Population Pharmacokinetic Analysis 462 patients in US clinical trials 462 patients in US clinical trials Average % BSA affected at baseline = 43% Average % BSA affected at baseline = 43% Estimation of “typical” exposure Estimation of “typical” exposure Nonlinear Mixed Effect Modeling (NONMEM) Nonlinear Mixed Effect Modeling (NONMEM) Steady state concentration0.25 ng/mL AUC 0-24 hours 6 nghr/mL Similar in pediatrics Similar in pediatrics

16 Hypothetical Worst Case Systemic Exposure in Atopic Dermatitis Patients European adult pharmacokinetic study European adult pharmacokinetic study – Application of 0.1% tacrolimus ointment – AUC 0-24 hours = 20 nghr/mL Model assumptions (contrary to clinical evidence) Model assumptions (contrary to clinical evidence) – lesions don’t heal – % affected BSA doesn’t decrease – quantifiable blood concentration for prolonged periods

17 Typical AD Patient Hypothetical Worst Case AD Patient Transplant Patient Transplant Patient with PTLD Mice with Lymphoma Cumulative AUC 122 days 732 1,410 55,144 79,208 65,392 Comparative AUC for Tacrolimus Blood Levels

18 Relative to AD Patients: Transplant Recipients with PTLD Mice with Lymphoma Hypothetical “Worst” Case 39x 56x 46x “Typical” Case 75x 108x 89x Relative Cumulative Systemic Exposure of Tacrolimus Based on Blood Levels

19 Clinical Pharmacology Conclusion Minimal systemic absorption Minimal systemic absorption – Most patients do not have quantifiable levels No systemic accumulation No systemic accumulation – Levels are transient Pediatric patients have a lower frequency of detectable blood levels and lower average levels than adults Pediatric patients have a lower frequency of detectable blood levels and lower average levels than adults Large safety margin in comparison to immunosuppressed transplant patients or mice Large safety margin in comparison to immunosuppressed transplant patients or mice