HUMORAL IMMUNE RESPONSES HAVE LITTLE EFFECT ON CONTROLLING VIREMIA DURING SIVagm INFECTION OF AFRICAN GREEN MONKEYS Thaidra Gaufin Division of Microbiology Tulane National Primate Research Center

BACKGROUND African non-human primate species rarely progress to AIDS even though the SIV viral load is the same or higher than the SIV viral load in pathogenic infections Titers of anti-SIV antibodies are lower in African non- human primates than in pathogenic rhesus macaque infections Recent study demonstrated in AGMs that there is an early activation of B cells in the LNs in comparison to SIVmac251-infected Rh (Cumont et. al, J. Virol, in press) Postulated that antibodies may be important in controlling SIVagm viral loads

CD20 DEPLETIONS IN RHESUS MACAQUES Previous in vivo CD20 depletion studies in Rh reported that antibodies may be involved in controlling post-peak viral load and in controlling disease progression These studies used SIV strains that are partially (SIVmac251) or highly (SIVmac239) resistant to neutralization

MATERIALS AND METHODS 9 AGMs were inoculated with 300 TCID50 of SIVagm.sab –Four AGMs treated intravenously with 50 mg/kg Rituxan at day -7 and every 21 days thereafter up to 180 days –Five AGMs were controls and received virus only Dynamics of major lymphocyte subsets (CD20, CD79A, CD4 and CD8) were investigated in peripheral blood, lymph nodes, and intestine by flow-cytometry and IHC Anti SIVagm antibodies (anti V3 and anti gp-41) were detected by ELISA Titers of neutralizing antibodies were measured using neutralizing antibody assay performed by Dr. Montefiori’s lab FCGR3A polymorphisms determined by PCR and sequencing Viral loads were measured by real-time PCR Levels of T cell immune activation, proliferation and apoptosis were compared in depleted vs non-depleted SIVagm-infected AGMs by flow-cytometry

CD20 DEPLETION IN PERIPHERAL BLOOD (%) CD20-depleted AGMsControl group AGMs SIVagm infection Rituxan administration SIVagm infection

CD20 DEPLETION IN PERIPHERAL BLOOD (#) CD20-depleted AGMsControl group AGMs

DEPLETION OF B CELL LINEAGE (CD20+ CD79a+ B cells) Percentage of CD20+ CD79a+ cells # of CD20+ CD79a+ cells/  l

CD20 DEPLETION IN LNs CD20-depleted AGMsControl group AGMs

CD20 DEPLETION IN THE INTESTINE CD20-depleted AGMsControl group AGMs

TISSUE DEPLETION OF CD20 IN AGMs (IHC FOR CD20) Lymph nodes Lamina propria D-7 D28

NO ASSOCIATION BETWEEN FCGRIIIA POLYMORPHISMS AND EFFICACY OF RITUXAN- INDUCED CD20 DEPLETION V229 was reported to be associated with effective depletion while I233 was reported to be associated with incomplete response

DYNAMICS OF ANTI-SIVagm ANTIBODIES Anti-GP41 antibodies Anti-V3 antibodies CD20-depleted AGMsControl group AGMs

ABLATION OF NEUTRALIZING ANTIBODY RESPONSE AGAINST AUTOLOGOUS SIVagm.sab STRAIN FOLLOWING B-CELL DEPLETION CD20-depleted AGMsControl group AGMs

DEPLETION OF CD20+ CELLS HAVE LITTLE IMPACT ON SIVagm.sab REPLICATION Comparison with undepleted SIVagm-infected AGMs No significant difference in viral loads between depleted and undepleted monkeys

CD4+ T CELL DYNAMICS: BLOOD CD20-depleted AGMsControl group AGMs

CD4+ T CELL DYNAMICS IN THE INTESTINE CD20-depleted AGMsControl group AGMs

EFFECT OF CD20 DEPLETION ON CD4+ T CELL PROLIFERATION AND ACTIVATION CD20-depleted AGMsControl group AGMs Ki67+ CD4+ T-CELLS Ki67+ CD8+ T-CELLS

EFFECT OF CD20 DEPLETION on CD4+ T CELL PROLIFERATION AND ACTIVATION CD20-depleted AGMsControl group AGMs

CONCLUSION Rituxan administration successfully depleted CD20+CD79a+ in blood, LN, and INT for all animals Depletion of CD20+ cells during SIVagm.sab infection had no effect on controlling viremia during the acute and chronic phases. Ablation of CD20+ cells did not modify the immunological parameters in SIVagm.sab infection

CONCLUSION Our results corroborate with our group’s previous data in CD20-depleted Rh infected with a neutralizable SIVsm strain (SIVsmD215) Rh: CD20 Depleted Group Rh: Control Group

CONCLUSION Humoral responses have little control in SIV replication in both pathogenic and non-pathogenic models

ACKNOWLEDGEMENTS Ivona Pandrea’s lab Jamie Cherry A. Chase Carter Crystal Stoulig Clint Coleman Cristian Apetrei’s lab Rajeev Gautam Mary Barnes Melissa Pattison Christopher Monjure Jeanne MacFarland Veterinary Medicine at the TNPRC Jason Dufour Ronald Veazey Preston Marx Andrew Lackner David Montefiori, Duke University Dina Washington, Genentech Supported by R01 AI (IP) R21 AI (IP) R01 AI (CA)