P10.06CD8 T-cell Based HIV Vaccines - Is TargetingConserved Epitopes the Answer?
Shelby L. O’Connor, Dane Gellerup, Max Harris, ErickaBecker
University of Wisconsin-Madison, Madison, WI, United States
Background: Immunodominant CD8 T cell responses emergein the first few weeks after HIV/SIV infection, suppress virusreplication, and select for escape variants. Although T-cell basedHIV vaccines have not successfully provided sterilizing immu-nity, vaccine-elicited CD8 T cells may contribute to the controlof replication of breakthrough viruses. It is critical that a T-cellbased vaccine generates an effective pool of memory CD8 Tcells that are available to respond during acute HIV infection andeffectively control both acute and chronic virus replication.Designing a vaccine to elicit these potent CD8 T cells in allvaccinated individuals, however, is a daunting challenge. Con-ceptually, a vaccine that elicits CD8 T cell responses targetinghighly conserved virus peptide sequences would have the mostwidespread impact, but the efficacy of T cells targeting theseconserved epitopes is unknown.Methods: We employ a model of SIVmac239Dnef-infectedMHC-identical Mauritian cynomolgus macaques to test thehypothesis that CD8 T cells targeting conserved epitopes areunable to detect and destroy virally infected cells. Accordingly,we expect that CD8 T cells targeting epitopes that accumulatevariants are more effective at controlling virus replication. Totest this hypothesis, we are creating variants of live attenuatedSIVmac239Dnef designed to elicit CD8 T cells targeting epi-topes that do and do not accumulate variants.Results: We will determine whether the mutant viruses are ‘fit’and whether the included variant epitope sequences are no lon-ger detected by CD8 T cells. In the future, we will determine ifacute CD8 T cells targeting these different categories of epitopesare able to control virus replication, in vivo.Conclusions: The conclusions from this study will help identifywhether CD8 T cells that develop during acute HIV infectioncan be specific for highly conserved epitopes and whether theycan control virus replication.
P10.07Bivalent NYVAC-based Vaccine Candidates againstHIV/AIDS Expressing Clade C Trimeric Solublegp140(ZM96) and Gag(ZM96)-Pol-Nef(CN54) as VLPs
Beatriz Perdiguero1, Carmen Elena Gomez1, Marıa VictoriaCepeda1, Lucas Sanchez-Sampedro1, Juan Garcıa-Arriaza1,Ernesto Mejıas-Perez1, Victoria Jimenez1, Cristina Sanchez1,Carlos Oscar S. Sorzano1, Julie Delaloye2, Thierry Roger2, ThierryCalandra2, Ralf Wagner3, Benedikt Asbach3, Karen V. Kibler4,Bertram L Jacobs4, Giuseppe Pantaleo2, Mariano Esteban1
1Centro Nacional de Biotecnologıa, CNB-CSIC, Madrid, Spain,2Centre Hospitalier Universitaire Vaudois and University ofLausanne, Lausanne, Switzerland, 3University of Regensburg,Regensburg, Germany, 4The Biodesign Institute at ArizonaState University, Tempe, AZ, United States
Background: The generation of vaccine candidates againstHIV/AIDS able to induce long-lasting protective immunity re-mains a major goal in the HIV field. The reduced efficacy(31.2%) against HIV infection observed in the RV144 Thai
clinical trial highlighted the need to develop novel improvedpoxvirus-based recombinants.Methods: In the present study, we have generated two novelNYVAC vectors expressing HIV-1 clade C gp140(ZM96)(NYVAC-gp140) or Gag(ZM96)-Pol-Nef(CN54) (NYVAC-Gag-Pol-Nef) and defined their biological characteristics incultured cells and in mice.Results: Insertion of the HIV-1 genes in the viral genome does notaffect the replication capacity of the NYVAC recombinants in pri-mary chick cells and the HIV-1 antigens are correctly expressed andreleased from the cells, with Env protein as a trimer (NYVAC-gp140), while in cells infected with NYVAC-Gag-Pol-Nef, Gag-induced VLPs are abundant. Electron microscopy revealed thatVLPs are accumulated with time at the cell surface, with no inter-ference with NYVAC morphogenesis. Gag-Pol-Nef expression inhuman primary monocytes markedly increases the levels of immu-nomodulatory molecules, such as cytokines and chemokines. Bothrecombinant viruses show an attenuation profile in immunocom-promised BALB/c mice after intracranial inoculation. Analysis ofthe immune responses elicited in mice after homologous NYVACprime/NYVAC boost immunization shows that the two recombinantviruses induced polyfunctional Env-specific CD4 or Gag-specificCD8 T cell immune responses. Antibody responses against gp140and p17/p24 were also elicited by both NYVAC vectors.Conclusions: Our findings showed that bivalent NYVAC vec-tors can be considered as candidate vaccines against HIV/AIDS.
P10.08Characterization of the Binding Affinity of Siglec-1to gp120, gp145, and V2 Loop via Sialic AcidBinding Motif
Hung V. Trinh1, Ousman Jobe1, Guofen Gao2, Carl R. Alving3,Venigalla Rao2, Mangala Rao3
1U.S. Military HIV Research Program (MHRP)/HJF, Labora-tory of Adjuvant and Antigen Research, Silver Spring, MD,United States, 2Catholic University of America, Department ofBiology, Washington, DC, United States, 3U.S. Military HIVResearch Program (MHRP), Walter Reed Army Institute ofResearch, Laboratory of Adjuvant and Antigen Research, SilverSpring, MD, United States
Background: Although HIV-1 primarily infects T-cells throughthe interaction of viral envelope with CD4 and a co-receptorCCR5/CXCR4, CD4 expresses in low level in macrophages.Recently, sialic acid-binding Ig-like lectin 1 (Siglec-1, CD169),which is highly expressed on the cell surface of macrophages,has been identified as a major receptor for HIV-1 infection. It isknown that Siglec-1 binds to glycans containing sialic acid. Tofurther understand the mechanism; we determined the bindingaffinity/kinetics of monomeric gp120 and trimeric gp145 to as-sess if there were differences in their ability to bind to Siglec-1.We further narrowed down the env-binding motif to the V2 loopof gp120. Finally, we conducted an inhibition assay in thepresence/absence of lactose (LA), 2, 6¢-Sialyllactose (6¢-SL) andsialic acid (SA) to identify the nature of the interactions.Methods: Siglec-1 was immobilized on a CM5 chip on a Bia-core T200. JRFL and SF162 (clade B) gp120, gp145, and V2loop proteins were captured. In additional experiments, theseenv proteins were immobilized, while Siglec-1 was injected overthat surface. For the inhibition assays, LA, SL and SA weremixed with the env proteins or with Siglec-1 and then injectedonto the immobilized Siglec-1 or env proteins, respectively.
A119
