In this study, we have produced soluble versions of Env glycoproteins trimers heterolgous trimerization motifs (GCN4 or foldon) Breadth of neutralization indicate that these glycoproteins significantly increase neutralization breadth and are interesting platforms for further modificatFor trimer immunogenicity, experiments are in progress with the current generation of trimeric immunogens to better map the neutralizing responses with the tools under development describe above. Using the current YU2 gp140-foldon trimers, we have compared several adjuvant formulations and found that the trimers emulsified in the GlaxoSmithKline (GSK) adjuvant AS01B generated improved neutralization. This adjuvant is a liposome formulation that contains the TLR-4 ligand monophosphoryl lipid A (MPL) and QS21 and currently is our preferred adjuvant of choice. Accordingly, an active MCRADA is in effect with GSK to cover many of our preclinical immunogenicty trials. We are currently testing the gp140-foldon trimers emulsified in the GSK AS01b adjuvant in monkeys with collaborators at the Karolinska. Very ELISA titers were elicited and neutralizing antibodies that could inhibit the entry of multiple HIV-1 clade B strains were generated after several inoculations. Further analysis of these sera are ongoing here at the VRC. As mentioned previously, we will design trimers with alternative heterologous trimerization motifs for structural pursuits in collaboration with the Kwong lab. A selected subset of trimers that reflect the antigenic profile of the native spike (that is, preferential recognition by neutralizing ligands only) will be tested for immunogenicity. We will also integrate other clade B, C and A primary Envs into the process of developing improved soluble, stable trimers, as well as into the solid phase proteoliposome (EnvPLs) technology published previously. The EnvPLs will be analyzed for their ability to elicit antibodies to conserved determinants by sequential immunization in preclinical trials as another aspect of an immunofocusing strategy on shared envelope glycoprotein elementsion to better elicit broadly neutralizing antibodies. We are also using these trimers to identify the resting B cell memory pool of anti-Env antibodies in HIV-1 infected individuals who possess broad neutralization activity in their serum.
