SUMMARY – OVERALL
While there is an urgent need for vaccines eliciting broadly neutralizing antibodies (bnAbs) against the highly
mutable HIV-1 retrovirus to stem its global spread, this goal remains elusive. Antibodies produced against
trimeric gp160 sites of vulnerability during natural infection drive retroviral mutation further, diversifying quasi-
species in individuals. One apparent exception is the conserved membrane-proximal external region (MPER)
site, critical for hemifusion/fusion and which is stealth, largely immersed in lipid and only transiently revealed
during spontaneous ectodomain tilting. Liposome-arrayed MPER segments induce specific antibodies in mice
but without neutralizing activity, explained by the restricted antibody access to the native MPER residing in a
narrow “crawlspace” between the viral membrane below and the base of the gp120 and gp41 protomers of the
trimeric gp160 envelop. Biomaterial formulation of polymer “steric clouds” on liposomes or alternative origami-
based approaches on nanodiscs to limit access to the MPER, akin to that mandated by the trimer three-fold axis,
were technically cumbersome and confounded by the uncertainty of MPER topology. Our recent analysis of the
structure and dynamics of nanodisc-embedded trimeric HIV-1 spike protein provides compelling data for
alternative design. In addition, synthetic RNA lipid nanoparticle (LNP) encapsulation technology obviates vaccine
challenges with hydrophobic proteins such as the MPER and adjacent transmembrane segment (MPER-TM).
This P01 comprises two projects. Project 1 shall determine the immunogenicity of RNA vaccines encoding
MPER-TM trimers fused to structurally suitable self-proteins to focus antibody responses against the conjoint
membrane arrayed viral MPER-TM, with the self-proteins imposing steric restriction comparable to that exerted
by the gp160 ectodomain on the native MPER. Serological, single B cell, recombinant mAb methods,
neutralization assays and Ig bioinformatic analyses are applied. Automated computational searches of the pdb
informed by single-particle cryo-electron microscopy (cryo-EM) structures and using RNA technology reveal
antigenicity and immunogenicity of construct design for planned immunization studies in normal as well as knock-
in mice harboring human Ig D3-3 and JH6 gene segments for long CDRH3 loop generation. Project 2 will perform
cryo-EM and X-ray crystallographic studies to compare existing and additional HIV-1 patient-derived bnAbs
obtained from biobanked serial samples (Osier, IAVI) with those that are vaccine elicited, in the context of
nanodisc-arrayed MPER immunogens and native spike protein. We shall investigate Ig requirements of somatic
hypermutation (SHM), Fab molecular dynamics (MD) and co-evolution of antibody approach angles and affinities
of bnAbs of IgG1 and IgG3 isotypes. Projects 1 & 2 shall be performed by Kim/Weissman (DFCI/UPenn) and
Reinherz (DFCI), respectively, with four Technology Components: mRNA-LNP vaccine synthesis by Weissman
(U Penn); cryo-EM by Walz (Rockefeller), X-ray crystallography by Tan (APS) and Kwong (NIH); and MD by
Huang (Texas A&M). Administrative (Reinherz) and Antibody (Seaman, BIDMC) Cores serve all.
