1. Food and Drug Administration Food and Drug Administration Center for Biologics Evaluation and Research Biological Response Modifiers Advisory Committee

2. Preclinical Considerations for Gene Transfer Clinical Trials: Vector Biodistribution Mercedes A. Serabian, M.S., DABT Anne M. Pilaro, Ph.D. FDA/CBER FDA/CBER BRMAC Meeting November 17, 2000 Bethesda, MD

3. Biodistribution Studies Are... n Preclinical animal studies designed to determine the distribution of vector to sites other than the intended therapeutic site

4. Goals of Biodistribution Studies n Evaluation of potential dissemination of vector to the germline u Assay of gonadal tissue n Evaluation of potential distribution of vector to non-target tissues u Additional data on potential target organs for toxicity studies u Determine kinetics of vector transduction & persistence

5. Addressing These Goals n Presence of vector sequence by DNA-PCR n Both issues may be addressed in the same preclinical study u Use of normal, intact animals u Use of animal model of disease

6. Results of the March 12, 1999 RAC Discussion on Gonadal Distribution n Risk of foreign gene transfer to germ cells, future progeny perceived to be low u Acceptance by those present in context of somatic cell gene therapies

7. Results of the March 12, 1999 RAC Discussion on Gonadal Distribution n Evaluation of biodistribution to the gonads may not be needed prior to all Phase 1 clinical trials n Consent form should address: u The lack of data u The unknown risk(s)

8. Sample Informed Consent Form n Risks associated with treatment in this study could cause permanent genetic changes in some of your sperm (men) or eggs (women). These changes could be neutral or may eventually cause abnormalities. Some of these changes could lead to miscarriage or abnormalities in your future children. Other changes may have no apparent effects but could still be passed on to future generations.

9. Sample Informed Consent Form (Cont’d) n The likelihood of such outcomes is currently unknown…. u Studies to estimate the likelihood of such effects have not been done in animals or humans. u Some studies (animals/humans) have been performed, but the information available does not allow estimation of the likelihood of such effects. u Conclusive data regarding these potential effects in animals and humans are not yet available.

10. When Can You “Postpone” Biodistribution Studies? n “Previously defined” vector u Previous experience with similar vector, route of administration, formulation, and schedule F e.g. adenovirus type 5 vectors n Transgene product “innocuous” if expressed ectopically n Size of vector (i.e. pDNA) not excessively different

11. And When They May Not… n New class of vector [no/little experience] u e.g. AAV, lentivirus, others n Change in formulation (i.e. lipid carrier) n Change to intentional systemic route of administration with established vector n Transgene has the potential to induce toxicity if aberrantly expressed in non-target organ

12. Sample Regulatory Letter for Studies Lacking Biodistribution Data "The present submission does not contain data that demonstrate the extent to which this vector is able to disseminate out of the injected site and distribute to gonadal tissues. These data are necessary to determine the risk of inadvertent gene transfer to the germ cells, which may result in genetic changes in subsequent progeny….

13. Sample Regulatory Letter (Cont’d)...In the course of development of your product, you will be required to obtain these data and provide them to the Agency for review and comment. Data may be obtained either from biodistribution studies in animals, analysis of clinical samples, or from a combination of preclinical and clinical sample analyses. Clinical data should be derived from peripheral blood cells and semen samples during the treatment and follow-up periods for the clinical trial, and from gonadal tissues (primarily ova) obtained at autopsy from consenting patients. We will require that these data be provided in a timely fashion, so that the results may be used to guide further development and optimization of your product as a therapeutic agent. Please update the Agency on the status of these studies at the time of each annual report."

14. Design Considerations for Vector Biodistribution Studies n Species selection u Non-human primates not always needed n Animal gender u Male &/or female (reflects patient population) n Animal numbers u 3-5/sex/group minimum F Use of smaller animals (i.e. rodents) allows for inclusion of larger numbers

15. Design Considerations for Vector Biodistribution Studies n Dose selection u Control u Maximally feasible/clinically relevant dose level u Lower dose for establishment of NOAEL n ROA u Should mimic intended clinical ROA to the greatest extent possible u “Worst-case” scenario may not adequately represent risk

16. Design Considerations for Vector Biodistribution Studies n Sacrifice/sampling timepoints [kinetics] F Early - at time of peak vector transduction/ expression F Later - to be determined by intended clinical use F Later still - to determine clearance of signal from gonads & non-target organs

17. Design Considerations for Vector Biodistribution Studies n Tissue panel u Recommended list - F Peripheral blood, gonads, injection site F Highly-perfused organs (determination of toxicity) Brain, liver, lung, kidneys, heart, spleen, etc...Brain, liver, lung, kidneys, heart, spleen, etc... u Other tissues F Based on toxicity/pathology [determined by transgene] (i.e., bone marrow) F Based on ROA (i.e., draining, contralateral LNs)

18. Design Considerations for Vector Biodistribution Studies n Detection assay u Methodology should detect sequence of vector DNA (or RNA) unique to that product u Methodology should be appropriate to adequately detect vector sequence: F In tissue samples from preclinical animal studies F In clinical samples obtained during the initial trial(s)

19. In Summary…. n Biodistribution studies are designed to evaluate vector dissemination out of the injected site u In both gonadal & other non-target tissues u Current method - DNA-PCR analysis n These studies not always required prior to initiation of Phase 1 trials; depends on: u “Previously defined” vectors u Clinical context u Will require data during course of product development