Current HIV Research, 2004, 2, 193-204 193 1570-162X/04 $45.00+.00 © 2004 Bentham Science Publishers Ltd. Progress Towards the Development of a HIV-1 gp41-Directed Vaccine Georgia B. McGaughey 1* , Gaetano Barbato 6 , Elisabetta Bianchi 6 , Roger M. Freidinger 2 , Victor M. Garsky 2 , William M. Hurni 3 , Joseph G. Joyce 3 , Xiaoping Liang 4 , Michael D. Miller 5 , Antonello Pessi 6 , John W. Shiver 4 and Michael J. Bogusky 2* Departments of 1 Molecular Systems, 2 Medicinal Chemistry, 3 Virus and Cell Biology, 4 Viral Vaccine Research, and 5 Biological Chemistry, Merck Research Laboratories, P.O. Box 4, West Point, PA 19486, USA, 6 Istituto di Ricerche di Biologia Molecolare P. Angeletti (IRBM), Via Pontina Km 30.600, 00040 Pomezia (Roma) Italy Abstract: The HIV-1 gp41 envelope glycoprotein mediates fusion of the viral and cellular membranes. The core of the gp41 ectodomain undergoes a receptor-triggered conformational transition forming a trimeric, α-helical coiled-coil structure. This trimer-of-hairpins species facilitates insertion of the viral envelope protein into the host cell membrane promoting viral entry. The prefusogenic conformation of gp41 is capable of stimulating a neutralizing antibody immune response and is therefore an attractive therapeutic target. Several broadly neutralizing HIV-1 monoclonal antibodies which bind to gp41 have been characterized and include 4E10, Z13 and 2F5. A conserved segment of gp41 (residues 661-684) has been identified as the epitope for the HIV-1 neutralizing antibody 2F5 (MAb 2F5). MAb 2F5 has attracted considerable attention because of the highly conserved recognition epitope and the ability to neutralize both laboratory-adapted and primary viral isolates. Antibodies which recognize the immunodominant regions of gp41 may provide protection against HIV infection if elicited at appropriate concentrations. Here we review the rational design, structure-activity relationships and conformational features of both linear and constrained peptide immunogens incorporating variants of both the 2F5 epitope and the gp41 ectodomain. This review describes a rational design approach combining structural characterization with traditional SAR to optimize MAb 2F5 antibody affinities of gp41- based peptide immunogens. The immunogens are shown to stimulate a high titer, peptide-specific immune response; however, the resulting antisera were incapable of viral neutralization. The implication of these findings with regard to structural and immunological considerations is discussed. Keywords: 2F5, HIV Vaccine, gp41, antibody INTRODUCTION The HIV/AIDS pandemic represents a major world health problem. Currently, there are more than 42 million people infected worldwide and it has been estimated that 45 million new infections will occur by the year 2010 if measures are not taken to control the spread of the virus [90]. There is an urgent need to develop a globally accessible, cost-effective therapy to address the crisis. The only realistic therapeutic solution would come in the form of an HIV-1 vaccine [50]. Ideally an HIV vaccine would prevent infection (sterilizing immunity), although a vaccine that limits HIV replication could be effective in preserving the immune system, thereby protecting against disease and reducing viral transmission rates. Developing a vaccine capable of preventing HIV infection presents a formidable challenge. HIV has a very high mutation rate, efficiently escapes the immune system by a variety of mechanisms and integrates into the host DNA where it remains dormant. The existence of individuals who, despite exposure to the virus remain uninfected, suggests that a vaccine eliciting sterilizing immunity may be *Address correspondence to these authors at the Departments of Molecular Systems, Merck Research Laboratories, P.O. Box 4, West Point, PA 19486, USA; Tel: 215.652.7183; Fax: 215.652.4625, P.O. Box 4, WP53F-301, West Point, PA 19486; E-mail: georgia_mcgaughey@merck.com Departments of Medicinal Chemistry, Merck Research Laboratories, P.O. Box 4, West Point, PA 19486, USA; Tel: 215.652.3801, Fax: 215.652.4674, P.O. Box 4, WP14-1, West Point, PA 19486; E-mail: michael_bogusky@merck.com possible [78, 91]. Considerable attention has been given to the development of viral envelope glycoprotein-based vaccines [8, 13, 26, 36, 42, 44, 50, 51, 67, 70, 87] since these proteins have been shown to be the major antigens for anti-HIV antibodies present in HIV seropositive individuals. Infection of cells by HIV-1 requires membrane attachment of the virion and subsequent fusion of the viral and cellular membranes. The fusion process is mediated by the viral outer envelope glycoprotein complex gp120/gp41 and target cell receptors. The envelope glycoprotein is synthesized as the precursor gp160 which is proteolytically cleaved into two non-covalently associated protein subunits, a surface subunit (gp120) and a transmembrane subunit (gp41) (Figure 1) [17, 39, 62] . The gp120 envelope protein is responsible for binding to the CD4 cell-surface receptor and a chemokine co-receptor, CCR5 or CXCR4 [10, 28, 100]. Following receptor binding, the membrane-anchored gp41 mediates fusion of the viral and target cell membranes. The gp41 ectodomain contains a hydrophobic, glycine-rich fusion peptide (amino acids 512-527) at the amino terminus which is essential for membrane fusion (numbering based on HXB2 gp160 variant as described in [16]). Two 4,3 hydrophobic repeat regions following the fusion peptide are defined by a heptad repeat (abcdefg) n , where the residues occupying the a and d positions are predominantly hydrophobic. The two heptad repeat regions are referred to as the N36 (residues 546-581) and C34 (residues 628-661) peptides. A loop region containing a disulfide linkage separates the two heptad repeat regions. The region of the