# Presentation: 521084

## Design and Use of HIV-1 Reference Strains for Standardized Assessments of Vaccine-Elicited Neutralizing Antibody Responses

**David C. Montefiori, Ph.D.**
**Laboratory for AIDS Vaccine Research & Development**
**Duke University Medical Center**
**Durham, NC**
**monte@duke.edu**

## Standardization of HIV-1 Reference Strains

**Standardization of HIV-1 Reference Strains**

**HIV-1 exhibits an extraordinary degree of genetic variability that has given rise to multiple genetic subtypes and circulating recombinant forms (CRFs) of the virus.  Information on the neutralization properties of each major genetic subtype and CRFs will inform vaccine design and will facilitate standardized assessments of vaccine-elicited neutralizing antibody responses.**

## The Need for HIV-1 Reference Strains

**The choice of target viruses has a significant influence on the outcome and interpretation of assay results**

**The Need for HIV-1 Reference Strains**

## SPECTRUM OF NEUTRALIZATION-SENSITIVITIES

**Overall neutralization-sensitivity**

**Average**

**High**

**Low**

**V3 & CD4i sensitive**

**Where most vaccine-elicited responses are now**

**TCLA & some 1****o**** isolates**

**Relative abundance**

**SPECTRUM OF NEUTRALIZATION-SENSITIVITIES**

**Desired reference strains**

## How to Measure HIV-1 Vaccine-Elicited Neutralizing Antibody Responses

**How to Measure HIV-1 Vaccine-Elicited Neutralizing Antibody Responses**

**  ****An effective neutralizing antibody response will need to overcome ****	****the genetic variability, structural complexity and many ****	****neutralization escape mechanisms of the HIV-1 envelope ****	****glycoproteins.**
**  ****These same properties of the virus need to be part of the plan ****	****when assessing vaccine-elicited neutralizing antibody responses.**

## Need for Standard Panels of HIV-1 Reference Strains

**Prioritize the advancement of candidate vaccine**
**Identify small but significant improvements in immunogen design**
**Facilitate assay standardization and GCLP assay validation for clinical trials**
**Inform new vaccine designs**

**Need for Standard Panels of HIV-1 Reference Strains**

## Composition of Standard Virus Panels

**12 viruses from acute/early sexually acquired infections per panel**
**Not unusually sensitive or resistant to neutralization**
**Genetically and antigenically diverse**
**R5 biologic phenotype**
**Molecularly cloned Env pseudotyped viruses are required for stability, reproducibility, improved GCLP and subsequent epitope analysis**
**Grouping by clade – rational for this purpose:**
**6 major genetic subtypes: over 90% viruses**
**Clades A, B, C, D, E (CRF01), A/G (CRF02)**
**Need additional panels corresponding to vaccine trial sites**
**Address key scientific questions relating to optimal panel composition and the possible need to modify initial panels to improve their correlative value. **

## Multi-Tiered Approach to Assessing Vaccine-Elicited NAb Responses

## Scientific Issues to Address

**Effect of possible genetic drift – compare recent vs. older isolates**
**Early/acute “transmitted” isolates vs chronic isolates**
**Clonal viruses vs. virus swarms**
**Plasma viruses vs PBMC coculture viruses**
**Mother-to-child transmission (vaginal, breast milk viruses)**
**Env pseudoviruses vs full-length molecular clones**
**Effect of virus produced in non-lymphoid cells such as 293T compared to primary T- cells**
**Impact of genetically engineered cell lines as target cells**
**What is threshold bar for meaningful neutralization?**

## Full-length functional Env plasmids – recently donated to the NIH AIDS Research & Reference Reagent Program (B panel cat. #11227; C panel cat. #11326) 

Sexually acquired, acute/early infections

R5 biologic phenotype

Neutralization phenotypes represent primary isolates

Genetically and antigenically diverse – sequences deposited in Genbank

Well-characterized e.g., N-glycans, V-regions, MAb epitopes

Transmissions M-M, F-M, M-F

**Sexually acquired, acute/early infections**
**R5 biologic phenotype**
**Neutralization phenotypes represent primary isolates**
**Genetically and antigenically diverse – ****sequences deposited in Genbank**
**Well-characterized e.g., N-glycans, V-regions, MAb epitopes**
**Transmissions M-M, F-M, M-F**

**Initial Panels of 12 Clade B and 12 Clade C HIV-1 Reference Strains**

***J. Virol. 79:10108-10125 (2005) – Clade B panel***
***J. Virol., in press (2006) – Clade C panel***

##   Emphasis on assays with Tier 2/Tier 3 viruses.

  Consider the magnitude (e.g., ID50 titer, % neutralization) and 	breadth (coverage) of the response.

  Exceed the “minimum bar” in the Vax004 phase III trial (~10% 	coverage for 50-80% neutralization; 0% coverage for >80% 	neutralization).

**  ****Consider the magnitude (e.g., ID50 titer, % neutralization) and ****	****breadth (coverage) of the response.**
**  ****Exceed the “minimum bar” in the Vax004 phase III trial (~10% ****	****coverage for 50-80% neutralization; 0% coverage for >80% ****	****neutralization).**

***These parameters are only part of a decision pathway that includes other immune responses. ***

**Decision Pathway for Prioritizing NAb-Based Vaccine Candidates**

## REFERENCES

_**COMENTARY**_**:**
**Mascola, J. R.,  P. D'Souza, P. Gilbert, B. Hahn, N. L. Haigwood, L. Morris, C. J. Petropoulos, V. R. Polonis, M. Sarzotti-Kelsoe, and D. C. Montefiori.  2005. Recommendations for the design and use of standard virus panels to assess the neutralizing antibody response elicited by candidate human immunodeficiency virus type 1 vaccines.  **_**J. Virol**_**. 79:10103-10107.**
_**CLADE B REFERENCE PANEL**_**:**
**Li, M., F. Gao, J.R. Mascola, L. Stamatatos, V.R. Polonis, M. Koutsoukos, G. Voss, P. Goepfert, P. Gilbert, K.M. Greene, M. Bilska, D.L. Kothe, J.F. Salazar-Gonzalez, X. Wei, J.M. Decker, B.H. Hahn, and D.C. Montefiori.  2005. Human immunodeficiency virus type 1 *****env***** clones from acute and early subtype B infections for standardized assessments of vaccine-elicited neutralizing antibodies.  **_**J. Virol**_**., 79:10108-10125.**
_**CLADE C REFERENCE PANEL**_**:**
**Li, M,.  J.F. Salazar-Gonzalez, C.A. Derdeyn, L. Morris, C. Williamson, J.E. Robinson, J.M. Decker, Y. Li, M.G. Salazar,****  ****V.R. Polonis, K. Mlisana, S.A. Karim, K. Hong, K.M. Greene, M. Bilska, J.T. Zhou, S. Allen, E. Chomba, J. Mulenga, C. Vwalika, F. Gao, M. Zhang, B.T.M. Korber, E. Hunter, B.H. Hahn, and D.C. Montefiori.  2006.  Genetic and neutralization properties of acute and early subtype C human immunodeficiency virus type 1 molecular *****env***** clones from heterosexually acquired infections in southern Africa.  **_**J. Virol**_**., in press.**
_**ADDITIONAL PANELS**_**:**
**Brown, B. K., J. M. Darden, S. Tovanabutra, T. Oblander, J. Frost, E. Sanders-Buell, M. S. DeSouza, D. L. Birx, F. E. McCutchan, and V. R. Polonis.  2005. Biologic and genetic characterization of a panel of 60 human immunodeficiency virus type 1 (HIV-1) isolates, representing clades A, B, C, D, CRF01_AE, and CRF02_AG, for the development and assessment of candidate vaccines.  **_**J. Virol**_**. 79:6089-6101.**

**REFERENCES**

## Dr. Montefiori’s laboratory is funded by:

_**Division of AIDS/NIAID/NIH**_**: **
** ****Primate Core Immunology Laboratory for AIDS Vaccine Research and Development (PCIL)**
** ****HIV Vaccine Trials Network (HVTN)**
** ****Center for HIV/AIDS Vaccine Immunology (CHAVI)**
_**Bill & Melinda Gates Foundation**_**:**
** ****Collaboration for AIDS Vaccine Discovery (CAVD)**