1 / 17

2014 “ Towards an HIV Cure ” symposium Melbourne

2014 “ Towards an HIV Cure ” symposium Melbourne. Following in vitro culture with myeloid dendritic cells, negative regulators of T cell activation are expressed preferentially on latently infected CD4 + T cells

maeve
Télécharger la présentation

2014 “ Towards an HIV Cure ” symposium Melbourne

An Image/Link below is provided (as is) to download presentation Download Policy: Content on the Website is provided to you AS IS for your information and personal use and may not be sold / licensed / shared on other websites without getting consent from its author. Content is provided to you AS IS for your information and personal use only. Download presentation by click this link. While downloading, if for some reason you are not able to download a presentation, the publisher may have deleted the file from their server. During download, if you can't get a presentation, the file might be deleted by the publisher.

E N D

Presentation Transcript

  1. 2014 “Towards an HIV Cure” symposiumMelbourne Following in vitro culture with myeloid dendritic cells, negative regulators of T cell activation are expressed preferentially on latently infected CD4+ T cells VanessaA. Evans, Renée M. van der Sluis, Nitasha A. Kumar, Rafick-Pierre Sekaly, Remi Fromentin, Nicolas Chomont, Paul U. Cameron, Sharon R. Lewin

  2. Resting CD4+T cell Infection of resting CD4+ T cells: a role for cell-cell interactions Unactivated resting cells Ex vivo tissue blocks Eckstein et al. J Virology 2001 Endothelial Cells Shenet al. J Virology 2013 Dendritic Cells Evans et al. PLoSPathogens 2013

  3. Myeloid DC induce latency in resting memory CD4+ T cells Non-proliferating CD4+ T cells • Not mediated by soluble factors: CCL19, CCL21, CXCL10, IL-10, IL-6 • Close DC-T cell proximity required for induction of latency Evans et al. PLoS Pathogens 2013

  4. Negative regulators and latency • Negative regulators dampen the immune response and can be found on exhausted T cells Ahmed et al. J Immunol 2010; Day et al. Nature 2006 • Latently infected T cells express negative regulators of T cell activation eg. PD-1, Tim-3 and TIGIT Chomont et al. Nat Med 2010; Fromentin et al. CROI 2014 • Blocking PD-L1/PD-1 in vivo restores the SIV-specific cellular and humoral immune responses, improves viral control and reduces immune activation Velu et al. Nature 2009; Dyavar Shetty et al. J Clin Invest 2012; Finnefrock et al. J Immunol 2009

  5. Hypothesis Expression of negative regulators during DC-T cell interactions may actively suppress viral replication and maintain latency

  6. Resting CD4+T cells Magnetic Bead Depletion Resting CD4+ T cells >98% PBMC CD4 + mouse anti-human CD8, CD11b, CD14, CD16, CD19, CD69, HLA-DR CD69 CD25 HLA-DR CD3 Dendritic Cells Magnetic Bead Depletion Plasmacytoid DC Bulk DC PBMC CD123 + mouse anti-human CD3, CD11b, CD19 HLA-DR Myeloid DC Lineage Cocktail CD11c

  7. DC added 1:10 AND 1 day Not productively infected EGFP- Non-proliferating eFluor670hi R5-EGFP-HIV-1 (2h pulse) Latent infection Resting CD4+ T cells eFluor670 Day 5 p.i. eFluor670hiEGFP- CD4+ T cells + integrase inhibitor L8 aCD3/aCD28 eFluor670 Activation >99% 3 Days EGFP Productive infection

  8. OR + mDC 1 day R5-EGFP-HIV-1 (2h pulse) Expression of negative regulators following DC-T cell co-culture eFluor-resting CD4+ T cells Baseline PD-1/Tim-3/CTLA-4 Phenotyping on days 1, 2, 3, 4, 5 post-infection eFluorhiEGFP- Viable Single cells T cells PD-1, Tim-3 and CTLA-4 SSC-W eFluor SSC-A CD3 FSC-A SSC-H HLA-DR EGFP

  9. Expression of negative regulators following DC-T cell co-culture PD-1 Tim-3 ( ) Mean fold change 24 Mean fold change 32 % Positive cells n=3 Days post-infection Days post-infection CTLA-4 No change % Positive cells Days post infection

  10. Differential expression of negative regulator ligands on mDC and pDC Leitner PLoS Pathog 2013

  11. Latent infection PD-1 lo/- cells Are PD-1hi cells enriched for latency? PD-1 hi cells eFluorhiEGFP- CD4+ T cells Anti-PD-1 aCD3/28 +L8 eFluor670 EGFP

  12. HIV latency is enriched in PD-1hi cells

  13. Tim-3 lo/- cells HIV latency is enriched in Tim3hi cells Tim-3 hi cells eFluorhiEGFP- CD4+ T cells Anti-Tim-3

  14. Summary • Myeloid DC facilitate establishment of HIV latency in resting memory CD4+ T cells via direct infection • Negative regulators PD-1 and Tim-3 but not CTLA-4 are up-regulated on resting CD4+ T cells upon co-culture with mDC • Negative regulator ligands are differentially expressed by mDC and pDC • mDC-induced HIV latency is enriched in PD-1hi and Tim-3hi cells

  15. EGFP+ productively infected CD4+ T cells Latent Infection in sorted eFluorhiEGFP- CD4+ T cells Blocking interactions between negative regulators and their ligands Cell death virus production immune clearance PD-1/-L1blockade PD-1 PD-L1

  16. Implications • mDC may facilitate ongoing latent infection of resting CD4+ T cells leading to replenishment of the reservoir • Disrupting the function of PD-1 and/or Tim-3 could potentially be exploited to inhibit replenishment of the reservoir and/or reverse latency

  17. Acknowledgements Monash University • Sharon Lewin • Paul Cameron • Renée van der Sluis • Nitasha Kumar • Suha Saleh • Candida da Fonseca AMREP Flow Cytometry • Geza Paukovicks • Michael Thompson • Jeanne Le Masurier • Phil Donaldson VGTI Florida • Rafick Sekaly • Nicolas Chomont • Remi Fromentin University of Melbourne • Damian Purcell

More Related