1. Laboratory of Immunobiochemistry site visit Jay E. Slater, MD FDA/CBER/OVRR/DBPAP June 29, 2006

2. 2/19 Laboratory of Immunobiochemistry (LIB) LIB supports the regulatory mission of CBER and FDA in assuring the safety and efficacy of allergenic products in the US by original research directed research projects expert advice lot release data BLA and IND review

3. 3/19 The last dedicated site visit for LIB was in January 2002 LIB was “functioning at one of the best levels in recent memory” With the “limited resources available,” LIB needed to be “well focused to achieve worthwhile results” LIB encouraged to “direct future efforts and resources toward continued standardization of allergenic extracts”

4. 4/19 Scientific goals Allergen structure/function Product quality/safety/efficacy Characterization of allergen extracts Product quality/safety/efficacy Modulation of T cell function Review of novel agents/formulations

5. 5/19 Staffing Principal Investigators Jay E. Slater, MD, Lab Chief Ronald Rabin, MD - Senior Staff Fellow Research Fellows Bo Chi, MD, PhD Nicolette deVore, PhD (Shoba Amarnath, PhD) (Srikant Bykadi, MS) (Ming Zhang, MD) Research Technicians Mona Febus Cherry Valerio Katia Dobrovolskaia

6. 6/19 LIB research program PI: Ronald Rabin, MD Modification of multidrug resistant protein (MRP-1) activity as a potential mechanism of immunomodulation Characterization of responses to respiratory syncytial virus (RSV) by T-cells PI: Jay E. Slater, MD Endotoxins in allergen extracts Biological potency of German cockroach allergen extracts Antibody microarray methods to determine allergen potency and composition

7. German cockroach allergen standardization Katia DobrovolskaiaICAC Mona FebusDAIT/NIAID Cherry ValerioJA Woodfolk, PhD University of Virginia

8. 8/19 The problem Cockroach allergy has been associated with asthma in the inner city Rosenstreich et al. N Engl J Med 1997; 336:1356-63. Eggleston et al. J Allergy Clin Immunol 1998; 102:563-70. Cockroach allergen extracts are non- standardized Standardized extracts are needed to Increase the safety and efficacy of extracts used for allergen immunotherapy Perform valid scientific studies of this important public health problem

9. 9/19 Aims Establish biological potency for German cockroach allergenic extracts Establish surrogate in vitro method for estimating biological potency

10. 10/19 IND 11319: German Cockroach Allergen Standardization Evaluation (CASE) Sponsor: DAIT/NIAID Four sites Baltimore Washington DC Chicago Denver

11. 11/19 Purpose Determination of the biological potency of three (3) commercially available German cockroach allergen extracts and test of their bioequivalence. Patient Population Adults with a history of allergic disease or asthma and a demonstrated sensitivity to German cockroach allergen tested. IND 11319: German Cockroach Allergen Standardization Evaluation (CASE)

12. 12/19 Conclusions/next steps Biological potencies of three German cockroach extracts have been determined These potencies appear to be low, but successful IT dosing should be achievable No single allergen assay will provide an adequate measure of overall potency allergens other than those tested may be significant surrogate potency test will have to take this uncertainty into account

13. Antibody microarrays for allergen standardization (see Briefing Document, pp.56-62) Nicolette deVore, PhD

14. 14/19 How do we measure potency? Total protein (hymenoptera) Overall allergen (grasses, mites) Pooled human antibody Specific allergen (cat, ragweed) Sheep antibody

15. 15/19 The dilemma of these potency measures: In order to measure specific allergens, we need to know which allergens are relevant If we measure overall allergenicity, we are unable to detect the absence of specific (and potentially important) allergens

16. 16/19 Nitrocellulose coated chip Antibody microarray plan Apply clonal scFvs Allergens bind to specific scFvs Incubate w/allergens

17. 17/19 Aims To develop a recombinant antibody microarray method for identifying individual allergens in complex mixtures potency profile Test this method using known simpler extracts (cat and short ragweed) Apply this technique to complex extracts, such as German cockroach allergen extracts

18. 18/19 Summary In our studies so far, we have applied phagemid library screening techniques to raising specific scFv antibodies to allergens developed appropriate antibody-screening methods to assess scFv function in the antibody microarray platform validated the use of the antibody microarray to measure the potency of allergens

19. 19/19 Microarray development plan Develop a quantifiable “fingerprint” of complex allergen mixtures using clonal allergen-specific scFvs and polyvalent sera Advance to more complex allergen extracts Yellow jacket venom German cockroach American cockroach