System Biology analysis of innate and adaptive immune responses during HIV infection

Transcription

1 System Biology analysis of innate and adaptive immune responses during HIV infection

2 Model of T cell memory persistence and exhaustion Naive Ag+APC Effector TEM (Pfp, Gr.B, FasL, TNF) Ag stim. IL-2, IL-7, TCR (pfoxo3a) Self renewal persistence Memory (TCM &TEM) TCM Hyperimmune activation Chronic persistent Ag (HIV) Increased inflammation Decrease in helper cytokines Pre-mature death of Memory B cells Dysfunctional Memory T cells Pre-mature death of TCM What are the mechanisms?

3 Model of T cell memory persistence and exhaustion Naive Ag+APC Effector TEM (Pfp, Gr.B, FasL, TNF) Ag stim. IL-2, IL-7, TCR (pfoxo3a) Self renewal persistence Memory (TCM &TEM) TCM Hyperimmune activation Chronic persistent Ag (HIV) Increased inflammation Decrease in helper cytokines Pre-mature death of Memory B cells? Dysfunctional Memory T cells (PD-1) and? Pre-mature death of TCM (pfoxo3a) What are the mechanisms?

4 System Biology It is the systematic use of genomic, proteomic, and metabolomic technologies for the construction of network-based models of biological processes. It is the study and identification of targets in an integrated network rather than in isolation. It is the deciphering the effect of a given molecule or pathway in immune system in the context of these many networks.

5 Global system analysis of innate immunity Ex vivo purified innate immune cells (monocytes, DCs, NK cells macrophages) Healthy, HIV chronic, Resistant to HIV infection, Susceptible to HIV infection, Elite controllers TLR triggered innate cells Virally infected innate cells Yellow Fever, Pox virus, Adeno viruses, HIV and their mutants

6 We are looking for biological correlates of resistance to HIV infection We compare gene expression of CMCs collected from HIV resistant women (R) against new negatives (NN). Sample Count New Negatives 13 Resistant 22 In Collaboration with Blake Ball, Frank Plummer and Rafick Sekaly

7 Pathways Analysis Using GSEA Canonical pathways and gene sets from literature Enrichment in NN samples Pathway pfdr Interferon inducible < Inflammation < TLR < IFNA < LPS stimulation induced genes < TOLL Pathway <0.0001

8 Pathways Analysis Using GSEA Transcription Factor Targets Enrichment in NN samples Pathway pfdr IRF IRF ISRE 0.018

9 Increased expression in IRF7 signaling in patients susceptible to infection NN Resistant

10 Fold change Susceptible/Resistant Increase expression in interferon induced genes in Cervical mononuclear cells (CMC) from susceptible sex workers IFI44L IFI6 IFIH1 IFIT1 IFIT2 IFIT3 IFIT5 N=9

11 Pathways Analysis Using GSEA Canonical pathways and gene sets from literature Enrichment in R samples Pathway pfdr OXIDATIVE PHOSPHORYLATION 0.001

12 Global System analysis of innate immune cells Identify specific innate transcriptional signatures for multiple viruses and vaccines Vaccine vectors Ad5, poxvirus, YF, HIV TLR triggering TLR-2, TLR-3, TLR-4, TLR-5, TLR-6, TLR-7 Generation of cell specific response data base Monocytes, mdc, pdc, T cells Generation of tissue specific data base PBMC, vaginal lavages, gut, lymph nodes

13 Strategy of DC infection and maturation PBMC Isolation mdc Mature DC pdc Infection with viruses, viral vaccines Triggering with TLR Transcriptional profiling

14 Distinct transcriptional signatures in different Pox viruses and their mutants in infected mdcs Count Color Key Row Z-Score

15 Fold change over Mock chemokines and cytokine expression levels in MVAC- C, NYVAC-C and NYVAC mutants CXCL10 CXCL13 CXCL16 CXCL9 IL12A IL15 LITAF TNF

16 Fold change over Mock IFN expression levels in MVAC-C, NYVAC-C and NYVAC mutants IFNB1 IFNA10 IFNA16 IFNA2 IFNA21

17 Fold change over Mock Enhanced expression of pathogen sensing molecules RIG-1 TLR7 TLR9 IRF1 IRF7 IRF8 MYD88

18 Fold change over control Induction of IRF genes by macrophages was enhanced by mutant NYVAC-C IRAK2 IRF1 IRF2BP2 IRF2BP2 IRF5 IRF7 IRF7 IRF8-2 -3

19 Summary Differential expression in Early, and late chemokine genes Gens associated with trafficking of T cells, B cells, NK cells and neutrophils cytokines which activate T cells (IL- 15) Genes of the IFN-a and IFN-b machinery

20 GSEA analysis shows the increased enrichment of immune pathways in NYVACC-KC DM compared to NYVACC

21 YF specific innate immune signature

22 YF vaccine induces the production of IL-7 in mdc and pdc

23 Distinct transcriptional signatures in mdc from healthy and HIV infected subjects HIV Healthy

24 Deliverables Generation of innate immune response data base Vaccine vectors Ad5, poxvirus, YF, HIV TLR triggering TLR-2, TLR-3, TLR-4, TLR-5, TLR-6, TLR-7 Generation of adaptive immune response data base HIV, YF, CMV, EBV Generation of cell specific response data base Monocytes, mdc, pdc, T cells Generation of tissue specific data base PBMC, vaginal lavages, gut, lymph nodes

25 Global system analysis of Adaptive Immunity: Dysfunction during HIV infection

26 Determine the global transcriptional signatures of HIV and CMV specific CD8+ T cells in HIV infected subjects Primary HIV Chronic HIV Elite controllers HIV HIV HIV CMV CMV CMV Gene array analysis N3-12

27 Cluster analysis of HIV- and CMV-specific tetramers in HIV infection CMV HIV

28 Cluster analysis of HIV- and CMV-specific tetramers in chronic infection HIV CMV

29 Cluster analysis of HIV- and CMV-specific tetramers in elite controllers CMV HIV

30 Common and unique gene expression levels during HIV infection up-regulated down-regulated

31 HIV specific CD8+ T cells in chronic stages express negative regulators and are exhausted Fold change of HIV vs CMV P<0.05

32 CD160 CD160 is expressed on NK, NKT, intraepthelial T cells, and low frequency of peripheral CD4+ and CD8+ T cells CD160 has been shown to significantly enhances TCR induced IL-2 production, proliferation and cytotoxic activities Recently, CD160 has been shown to inhibit TCR induced proliferation and cytokine production The ligand for CD160 is HVEM

33 n = 63 Untreated (except for STs) Acute: Infected < 6 months Chronics: Infected > 6 months ST: undetectable viremia EC: Undetectable viremia LTNP: low detectable viremia Patient population Time from infection n CD4 CD8 Log VL copies Acute 2.8 ( ) ( ) 728 ( ) 5.2 ( ) ( ) Chronic 11.7 ( ) ( ) 717 ( ) 4.9 ( ) ( ) ST 86.2 ( ) ( ) 732 ( ) EC ( ) ( ) 776 ( ) 1.7 ( ) 50 (50-987) LTNP ( ) ( ) 941 ( ) 3.7 ( ) 4228 ( ) Healthy 9

34 PD-1 CD160 is upregulated during chronic HIV infection on both total and HIV-specific CD8 T cells CD8 PD-1 A Tetramer CD160 CD160 MFI 1320 MFI 5466

35 CD160 is upregulated during chronic HIV infection on total CD8 T cells

36 CD160 and PD-1 levels are upregulated during chronic HIV infection on total CD8 T cells

37 CD160 and PD-1 expression levels are upregulated during chronic HIV infection on HIV-specific CD8 T cells

38 PD-1+ CD160+ double compartment defines a new exhausted subset during chronic HIV on antigen specific CD8 T cells

39 PD-1+ CD160+ double compartment defines a new exhausted subset during chronic HIV on antigen specific CD8 T cells

40 PD-1+ CD160+ double compartment defines a new exhausted subset during chronic HIV on antigen specific CD8 T cells

41 CD160 Unstimulated SEB CMV peptide HIV peptide IFN TNF CD107a Cytokines

42 PD-1+ CD160+ CD8+ T cells are less polyfunctional than SP PD-1 CD160- CD160+

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44 Tetramer A*022 CMV B*07 CMV B*08 NEF B*08 p Isotype control 6.1% 5.26% 5.64% 0.5% HVEM 0.7% 0.51% 0.56% 0.02% PD-L1 4.28% 6.29% 2.62% 0.15% HVEM + PD-L1 CFSE

45 Blocking CD160 interaction with its ligands increases HIV-specific T cell proliferation

46 Blocking CD160 interaction with its ligands increases CMV-specific T cell proliferation

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48 PD-1+ CD160+ double compartment is a distinct subset and is transcriptionally different than PD-1+ single subset

49 PD-1+ CD160+ double compartment is a distinct subset and is transcriptionally different than PD-1+ single subset

50 Summary Novel compartment More dysfunctional Specific phenotype Identifies chronic and not acute

51 Acknowledgements R.P. Sekaly Y. Peretz J. Van Grevenhyngue Lydie Trautmann Y. Shi Collaborators Blake Ball Frank Plummer Gepi Pantaleo Rick Koup Bruce Walker Ali Filali JP Gaulet J.P. Routy M.R. Boulassel McGill University Health Center, Montreal Réseau SIDA et MI du FRSQ Montreal