Antibody therapy of HIV infection and AIDS has been limited by the ability of the virus to escape antibody neutralization. There is a need for antibodies that can potently neutralize clinical isolates of HIV-1 across several diverse clades. The immediate objective of this project is to identify and characterize catalytic anti-gp120 scFv by further screening our hybrid phage-display library (SLE VL -S1-1-VH ) consisting of VL genes from SLE patients paired with a single VH from an anti-gp120 antibody (S1-1) that neutralizes several isolates of HIV-1. Since individuals with SLE have been shown to possess light chains capable of protein catalysis, the hypothesis for this objective is that selected human catalytic light chains paired with a human anti-gp120 heavy chain will more effectively neutralize HIV than the parent IgG, by proteolysis of gp120. Basic science long-term objectives: Understand the proteolytic mechanism (i.e. serine protease-like activity?) for selected catalytic antibodies and improve their turnover rates while maintaining specificity; Clinical long-term objectives: Passive administration of a catalytic antibody would be therapeutically beneficial compared to conventional antibodies because conventional antibodies bind reversibly to a single antigenic determinant, but may also be removed from circulation by the RES after binding, while catalytic antibodies can bind and inactivate by cleaving target antigen (gp120) prior to removal from circulation.
Specific Aims are to (i) identify and sequence VL-VH pairs that bind and cleave HIV-1 gp120 as scFv-phage and soluble scFv (ii) test the VL-VH pair for neutralization of diverse HIV-1 isolates, and (iii) determine the turnover rate (kcat) of the VL-VH pair for gp120 substrate to position us for generation of a clinically useful IgG. Methods to achieve the specific aims include screening the SLE VL -SI-I-VH library and """"""""lead"""""""" VL-VH pairs for catalysis using biotinylated gp120 and gp120-CRA, HIV neutralization assays with diverse clinical isolates using fresh PBMC, and calculation of the rate of proteolysis in collaboration with Dr. Sudhir Paul. In this novel approach we take advantage of one disease state (SLE) to develop a treatment for another (HIV).
