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Your logo. Long-Term Reduction in Peripheral Blood HIV-1 Reservoirs Following Reduced-Intensity Allogeneic Stem Cell Transplantation in Two HIV-Infected Individuals.

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  1. Your logo Long-Term Reduction in Peripheral Blood HIV-1 Reservoirs Following Reduced-Intensity Allogeneic Stem Cell Transplantation in Two HIV-Infected Individuals Timothy J. Henrich1,2, Gaia Sciaranghella3, Jonathan Z. Li1,2, Sebastien Gallien4, Vincent Ho5,2, Ann S. LaCasce5,2, and Daniel R. Kuritzkes1,2 1Brigham and Women's Hospital, Boston, MA; 2Harvard Medical School, Boston, MA; 3Ragon Institute of MGH, MIT, and Harvard, Boston, MA; 4 Hopital Saint-Louis, Paris, France; 5Dana-Farber Cancer Institute, Boston, MA.

  2. Background • One reported “functional cure” of HIV-1 infection: myeloablativeallogeneic HSCT from a homozygous ccr5Δ32donor1,2 • Several factors may have contributed to functional cure including pre-transplant myeloablative chemotherapy, GVHD, full engraftment of CCR5- donor cells • Cytotoxic chemotherapy alone insufficient to eliminate reservoirs as HIV-1 DNA persists after autologousHSCT3,4 • The long-term effects of allogeneic HSCT using CCR5+ stem cells have not been studied in detail 1Hutter et al. 2009; 2Allers et al. 2010; 3Simonelli et al. 2010; 4Cillo et al. 2011;

  3. Study Aims Study Aims: • Examine long-term changes in the peripheral HIV-1 reservoir following allogeneic HSCT in the setting of cART • Explore HIV-1 coreceptor usage, PBMC coreceptor expression and HIV-specific antibody responses pre- and post-HSCT Patients: 2 HIV-1 infected patients on combination ART who underwent reduced-intensity conditioning (RIC) allogeneic HSCT RIC = non-myeloablativechemotherapy, no total body irradiation or anti-thymocyte globulin

  4. Methods • Studied stored blood samples collected pre- and post-HSCT and prospectively collected samples (5 time points) • Quantified proviral HIV-1 DNA from peripheral blood mononuclear cells (PBMCs) and purified CD4+ T cells by real-time PCR • Quantified 2-LTR circles from PBMC episomal DNA • Quantified plasma viremia by a single-copy assay • Viral outgrowth assays using ~107 patient-derived CD4+ T cells and CD8 T cell-depleted lymphoblasts from an HIV-negative donor • CCR5 genotyping/flow cytometric quantification of CCR5 expression on CD3+ T lymphocytes • Genotypic and phenotypic determination of HIV-1 coreceptorusage • Quantified HIV-1-specific Ab levels & avidity

  5. Study Patients Patient A:Male with perinatally acquired HIV-1 on long-term ART 2006: Stage IV Hodgkin disease  standard treatment Disease recurrence  salvage therapy 2007: Autologous HSCT 2008: Relapse  RIC partially mismatched unrelated-donor HSCTcART: TDF/FTC/EFV 3-4 years pre-HSCT with undetectable VL Clinical course post-allogeneic HSCT

  6. Study Patients Patient B:Male with sexually acquired HIV-1 in mid-1980’s 2003: Large B-cell lymphoma  chemotherapy and cART started 2006: New stage IV Hodgkin lymphoma Disease recurrence  salvage therapy 2007: Autologous HSCT 2010: MDS (Tx-related)  RIC matched related-donor HSCTcART: TDF/FTC/RAL peri-transplant with undetectable VL Clinical course post-allogeneic HSCT

  7. Patient A Viral outgrowth assay negative day +1266 No 2-LTRs detected

  8. Patient B Viral outgrowth assay negative day +652 No 2-LTRs detected DLI= donor lymphocyte infusion

  9. CCR5 / Coreceptor Usage • Both patients heterozygous for ccr5Δ32mutation • PBMC homozygous wild-type for CCR5 after engraftment • Percentage of CCR5-expressing lymphocytes nearly doubled after full donor engraftment in Patient A (sufficient sample) • Full-length HIV-1 env amplified from proviral DNA at pre- and 1st post-HSCT PBMC samples (later timepoints negative) • V3-loop genotyping predicted CCR5 usage pre- and post-HSCT • R5 phenotype confirmed by tropism assay of pseudotyped viruses expressing PBMC-derived env

  10. Anti-HIV Ab Quantification Limited Sensitivity VITROS Assay Limiting-Antigen Avidiy Assay Patient A Patient B • HIV-specific Ab detected by VITROS assay pre- & post-HSCT • Decrease in Ab levels post-HSCT from diluted and undiluted plasma • Similar decrease in antigen avidity by limiting-antigen assay

  11. Summary & Conclusions • Allogeneic HSCT with RIC in the setting of suppressive ART led to a substantial and sustained reduction in the HIV-1 reservoir in PBMC - Reduction in proviral HIV-1 DNA correlated temporally with full donor engraftment • Engraftment of susceptible donor cells without infection adds supportive evidence that HIV-1 replication is fully suppressed by effective cART • Declining HIV-specific Ablevels/avidity provide further evidence for minimal persistence of HIV-1 antigen • Tissue sampling and analytic treatment interruption are necessary to fully assess the extent of HIV-1 reservoir reduction after allogeneic HSCT

  12. Acknowledgements BWH:Funding Sources: Members of the Kuritzkes Lab: NIH/NIAID 1K23AI098480-01A1 to TJH Zixin Hu UM1 AI068636 to DRK; P30 AI060354 Nina Lin U19 AI096109 to SGD Françoise Giguel Laura Lavoie AtheTsibris Members of the ID Division Blood Systems Research Institute/UCSF: Michael Busch ShealaKeeting Mila Lebedeva UCSF: Steven Deeks Harvard School of Public Health: Ronald Bosch (SDAC)

  13. Viral DNA Diversity Pre-HSCT Post-HSCT Patient A Single genome sequencing of full-length HIV-1 Env from PBMC DNA from pre- and 1st post-HSCT samples No major evolutionary changes observed following HSCT

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