1. Azithromycin Pharmacokinetics in Pregnancy James H. Fischer, Pharm.D., FCCP May 17, 2011

2. Azithromycin: Background Macrolide Commonly administered for community acquired respiratory, skin and gynecological infections Among 15 most frequently prescribed drugs to pregnant women (2004) Dose derived from non-pregnant women & men Practice disregards how dose requirements affected by pregnancy-related differences: –Pharmacokinetics –Functioning of immune system Piscitelli et al 1992; Andrade et al 2004; Jamieson 2006

3. Azithromycin: Pharmacokinetics Distinct profile Incomplete oral absorption (34%) Extensively distributes into tissues Eliminated by hepatobiliary excretion – P-glycoprotein – Multidrug resistance protein-2 (MRP-2) Only one previous PK study in pregnancy – Women from Papua-New Guinea receiving malarial prophylaxis Ballow et al 1998; Luke & Foulds 1997; Sugie et al 2004; Salmon et al 2010

4. Objectives Compare population pharmacokinetics of azithromycin between pregnant women and nonpregnant women of child bearing age Identify factors contributing to the inter-individual variability in azithromycin pharmacokinetics

5. Research Team Co-Principal Investigator Gloria Sarto, M.D., Ph.D. Brigham & Women’s Hospital, Harvard –Ruth Tuomala, M.D. –Karen McCarthy, R.N. University of Illinois at Chicago Patricia Fischer, R.N. Mitra Habibi, Pharm.D. Sarah Kilpatrick, M.D. Keith Rodvold, Pharm.D. University of Wisconsin Thomas Jenkins, M.D. Lori Wollett, R.N. University of Michigan –Janet Shier, Pharm.D., M.D. –Zan Daley, R.N. FDA Margaret Miller, Ph.D.

6. Methods: Study Design Pilot Study –Goals: assist in establishing structural model provide initial PK parameter estimates –12 healthy adult women of child bearing age –Traditional Multi-Dose Pharmacokinetic Study 500 mg *1, then 250 mg Q.D. for 4 days Extensive sampling performed for 96 h following last dose –Typical eligibility criteria (except allowed OC use)

7. Methods: Study Design Population Pharmacokinetic Study – Prospective, open-label multicenter design – Participants Women at least 18 years of age, Capable of bearing children, Receiving azithromycin for treatment of infection, and Either – Pregnant and at least 12 weeks gestational age, or – Nonpregnant and at least 3 months postpartum.

8. Methods: Study Design Sparse sampling strategy –5 samples collected within 4 sampling windows –First 3 with any dose, last 2 with final dose Drug intake assessed by diary and interviews

9. Methods: Laboratory Analysis Azithromycin Plasma Concentrations –HPLC with electrochemical detection –linear: 10 – 505 ng/ml –LLOQ: 10 ng/ml –inter-assay precision: 3.2% to 5.8%

10. Methods: Pharmacokinetic Analysis Nonlinear mixed effects modeling (NONMEM) with FOCE method Subjects with at least one evaluable azithromycin plasma concentration Data from healthy women included in population PK database Step 1: Identify structural (base) model Step 2: Covariate Analysis Step 3: Model validation

11. Covariates Body Size Measures total body weight lean body weight body mass index body surface area Continuous Variables age gestational age creatinine clearance azithromycin dose Categorical Variables pregnancy status Ethnicity study site type of infection concurrent medications oral contraceptives renal/hepatic disease administration with food healthy volunteer

12. Demographics Number53 pregnant (12-40 weeks GA) 25 non-pregnant Age (years)28 (18-49) Total Body Weight (kg)76 (45-178) Ethnicity African American19 Caucasian42 Hispanic9 Other8 Creatinine Clearance (ml/min)110 (37-229) Indication for Azithromycin Respiratory tract infection41 Premature rupture of membranes14 Chlamydia 8 Other 3 Healthy volunteer12

13. Structural Model Three-Compartment PK model Inter-individual variability (IIV) – exponential error Residual variability – proportional error Vc ka V-p1 CL D-P1 CL V-p2 [Lag time] CL D-P2

14. Two-Compartment Three-Compartment

15. Covariates: Oral Clearance (CL/F) Body Size Measures total body weight lean body weight body mass index body surface area Continuous Variables age gestational age creatinine clearance azithromycin dose Categorical Variables pregnancy status ethnicity pregnancy-ethnicity (pregnant non-African Americans) study site type of infection concurrent medications oral contraceptives renal/hepatic disease administration with food healthy volunteer

16. Covariate Models Oral Clearance (CL/F) CL/F = 134 L/h + (Ethn × Preg × (-51)) + (OC × (-51)) × LBW/50

17. Model Predicted CL/F for 50-kg Lean Body Weight (LBW) Woman SubjectsCL/F (L/h) Nonpregnant women of any race not receiving oral contraceptives 134 Pregnant African American women134 Pregnant Non-African American women83 Nonpregnant women receiving oral contraceptives (OC)83

18. Population PK Parameters: Model vs. Bootstrap Estimates ParameterFinal ModelBootstrap (n=1000) EstimateMedian2.5 th - 97.5 th Percentiles k a (h -1 )0.8--- Lag time (h)1.3 1.0 – 1.6 CL/F (l/h/50 kg LBW)13413385 – 176 Pregnancy, non-African Americans-51-44-78 – -4 Oral Contraceptive Use-51-44-78 – -4 CL D-P1 /F (l/h)401398235 - 609 CL D-P2 /F (l/h)12011535 - 208 V c /F (l)456436189 - 716 V P-1 /F (l)15601630925 - 3629 V P-2 /F (l)16100 174006124 - 31837 Inter-Individual Variability (CV, %) CL/F363416 - 49 CL D-P2 /F (l/h)86 3 - 133 V c /F11411675 - 161 V P-1 /F60 0.5 - 110 Residual error (%)32 18 - 42

19. Visual Predictive Check Nonpregnant women Pregnant African American women Pregnant Non-African American women Women receiving oral contraceptives

20. Influence of Pregnancy and Ethnicity on CL/F * * p<0.05, compared to non-pregnant women who were not receiving oral contraceptives

21. Influence of Oral Contraceptives on Cl/F * * p<0.05, compared to non-pregnant women who were not receiving oral contraceptives

22. Conclusions Ethnicity influences the effect of pregnancy on azithromycin CL/F Compared to non-pregnant women, azithromycin CL/F during pregnancy is unchanged in African American women and 40% lower in non-African Americans Concurrent administration of oral contraceptives (OCs) also reduced azithromycin CL oral

23. Conclusions Whether ethnicity also impacts the effect of OCs on CL/F requires further study as no African American women were in the OC cohort. These findings suggest that estrogen or progesterone mediate the effects of pregnancy and OCs on azithromycin CL/F.

24. Research in Pregnant Women Lessons Learned

25. IRB Challenges Vulnerable Population Options to Participation (other drug options) Coercion/Undue Influence Inclusions of subjects < 18 y/o Risks of Drug Therapy Restrictions on concurrent drug therapy Data Safety Monitoring Committee

26. IRB Challenges Frequency of Blood Sampling Translated consent forms Certificate of Confidentiality Follow up Data Collection on Newborn – Consent – Blood sampling

27. Overcoming Challenges Request Pre-review with IRB staff Study design: Population PK – missed clinic visit or time limitations have no impact on study integrity Be prepared for additional scrutiny

28. Reasons for not participating Time commitment Illness in addition to pregnancy – did not want additional burden Aversion to needle sticks Discomfort with 24 hr blood pressure monitor – Staff wore first to develop strategies

29. Subject Recruitment Strategies Involve Primary Physician – Assure physician that research is independent of clinical care – Provide overview to potential subject Experienced Study Coordinator Coordination with clinical care services – Other labs – Ultrasounds – Prolonged monitoring

30. Subject Recruitment Strategies Multiple sites – Adequate number of subjects – Diversity of population Age Ethnicity Diagnosis Liver/renal function Body size