The major goal of this program is to derive human monoclonal antibodies against human immunodeficiency virus 1 (HIV-1) and to characterize the specificities and biological activities of such monoclonals. The source of B cells for production of these monoclonals will be peripheral blood mononuclear cells (PBMC) from HIV-1 seropositive individuals. Information obtained from these studies will be used to dissect the human antibody response to HIV-1 in both healthy seropositives and ill (ARC and AIDS) individuals. It is important to conduct these studies for a variety of reasons. First of all, though anti-HIV-1 antibodies in ARC and AIDS patients do not protect these individuals from the disease, this may be because the antibody response is developed too late to be effective and/or because the specificities and biological activities of the antibodies produced by these individuals are not as protective as those of healthy seropositives. Thus, it is important to compare the anti-HIV-1 antibodies of healthy seropositives to those of ARC and AIDS patients. Secondly, it is likely that both humoral immunity and cell-mediated immunity will have to be elicited by any vaccine which is effective against AIDS. Yet the biologically important anti-HIV-1 antibody specificities and activities have not been clearly identified. Finally, the production and characterization of human monoclonals against HIV-1 is likely to have additional practical and theoretical implications relevant to AIDS. Among these are: 1) the potential use of human monoclonals with neutralizing activity for passive immunization against AIDS, 2) use of such monoclonals to define viral epitopes critical for elicitation of neutralizing antibody; these epitopes could be prepared biosynthetically for use as vaccines, 3) the possibility of developing an anti-idiotypic vaccine for AIDS by first developing human monoclonal(s) with viral neutralizing activity and then using these, in later experiments, to derive anti-idiotypic antibodies, and 4) use of human monoclonals against HIV-1 to probe the function of ill-defined viral components, such as the integrase and the tat gene product. The derivation of human monoclonal antibodies from ARC and AIDS patients should be facilitated by the recent discovery by ourselves and other investigators that these patients have an increased number of spontaneously transformed B cells in in vitro cultures of PBMC as compared to normals; the evidence points toward Epstein-Barr virus (EBV) as the transforming agent. We have also recently demonstrated that the long-lived PBMC cultures derived from ARC and AIDS patients contain oligoclonal B cells producing anti- HIV-1 antibodies. If we are able to enrich the population of transformed, anti-HIV-1 antibody-producing B cells via sib- selection of long-lived cultures, as our preliminary results indicate, we shall clone this population on soft agarose directly in order to derive human monoclonals. We shall also attempt to enrich for this population via exogenous EBV infection and HIV-I antigen stimulation. Fusion of the EBV-transformed B cells with myeloma/hybridoma cell lines and/or infection of these B cells with a retroviral vector containing a c-myc gene will be utilized to stabilize the cells either just before or just after cloning the cells. The monoclonals obtained will be characterized for viral component and epitope specificity binding to virus-infected cells, neutralizing activity, and ability to effect antibody-dependent cellular cytotoxicity and to effect antibody- and complement- dependent lysis. Additional aims of our project are: 1) to probe the basis of the spontaneous B cell transformation in PBMC cultures of ARC and AIDS patients, and 2) to develop and apply an ELISA assay for anti-HIV-2 antibodies toward identification of any HIV-2 seropositive individuals in our patient population.
