PMED Opening Workshop - Beyond ART: Towards Eradication or Drug-Free Control of HIV Infection - David Margolis, August 13, 2018
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HIV persists within the body despite successful suppression of virus replication with antiretroviral therapy (ART). HIV lurks in latent and active reservoirs, leading to rebound of virus spread if ART is interrupted. The latent HIV reservoir is a natural consequence of the lifecycle of HIV, with integration of HIV into the genomes of cells that are or later enter the resting state, resulting in transcriptionally quiescent provirus. Resting CD4 T cells comprise the majority of the latent reservoir. Multiple factors such as the degree of virus exposure, timing of ART, and host factors can influence the size and characteristics of the HIV reservoir. Constructing and testing effective strategies for HIV eradication and measuring their impact will require a sophisticated knowledge of the HIV reservoir, detailed understanding of the antiviral immune response, and of the diversity and kinetics of the latent viral reservoir.
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PMED Opening Workshop - Beyond ART: Towards Eradication or Drug-Free Control of HIV Infection - David Margolis, August 13, 2018
- 1. David Margolis MD UNC HIV Cure Center Beyond ART: towards eradication or drug-free control of HIV infection
- 2. Predicted Survival if HIV+ at age 25 years Before modern therapy: 50% dead 7 years after infection
- 3. Treatment allows people to live with HIV “A 20-year old HIV-positive adult on [HIV medications] … is expected to live into their early 70s, a life expectancy approaching that of the general population.” PLOS ONE 2013 Dec (8)12:e81355
- 4. An Indian woman with H.I.V. shows her anti- retroviral drugs at her home in New Delhi. Andrew Caballero-Reynolds/Agence France-Presse – Getty Images
- 5. Generation of latently infected cells is probably the result of transcriptional silencing during memory cell differentiation Antige n Naïve T-cell Activated T-cell Resting memory T-cell Proliferating effector cells Resting memory T-cell with latent HIV provirusHIV infection
- 6. • Persistent resting cell infection occurs during acute infection, but the kinetics of the establishment of latency are unclear as is the stability of the early latent pool. • Here using mathematical modeling we studied the kinetics of latency establishment and the effects of early initiation of ART. • Hypothesis: viral latency is established in proportion to viral infection events
- 7. Model For Generation of Latently Infected Cells death death activation Perelson Ho et al Nature 1997 f << 1
- 8. Correlation between model prediction and experimental data: frequency of latent infection proportional to exposure to viremia Arbitrary units Normalize scales and estimate f; 1 in a few hundred thousand to a million Infections generate latency
- 9. Reservoirs of HIV M. Stevenson, Scientific American 2008
- 10. 1 2 3 4 5 6 7 Time on HAART (years) Persistent HIV infection despite ART (2003) - Time to eradication > 73.4 years 0.0001 0.001 0.01 0.1 1 10 100 1000 10000 0 Frequency (per106cells) 0.00001 Siliciano JD, et al. Nature Med. 2003
- 11. - 1 2 3 4 5 6 7 Time on HAART (years) Persistent HIV infection despite ART (2015) - Time to eradication > 73.4 years 0.0001 0.001 0.01 0.1 1 10 100 1000 10000 Frequency (per106cells) 0.00001 ! 0 5 1 0 1 5 0 .0 0 0 1 0 .0 1 1 1 0 0 1 0 0 0 0 2 0 2 5 3 0 IU P M o v e r tim e fro m A R T T im e o n H A A R T (y e a rs) IUPM Crooks et al. JID 2015 -
- 12. Margolis, Garcia, Hazuda, Haynes; Science 2016
- 13. Persistent, latent infection of memory CD4 cells decays slowly over time Margolis et al. Science 2016 • Given assay variance, a more than 6-fold RCI decrease would have likelihood 0.023 (2.3%) • Therefore a measurable goal is therapy that can reduce the latent reservoir by half a log
- 14. A first step to eliminate latent HIV infection Latency Reversal A second step to eliminate latent HIV infection
- 15. HIV RNA Cell-associated HIV DNA, RNA, antigen & viruses Ho, Cell 2013 Ericksson, PLoS Path 2013 The “Real” Reservoir HIV DNA QVOA Replication- competent virus
- 18. Pt. 1 Pt. 2 Pt. 3 Pt. 4 Pt. 5 Pt. 6 Pt. 7 Pt. 8 0 20 40 60 200 400 600 800 100 RelativeHIV-1gagRNAcopies Baseline ART VOR 400 mg Vorinostat: Lewin group, Melbourne Panobinistat, Romidepsin: Arrhus group, Denmark
- 20. HIV+ Resting CD4 culture Untreated VOR PHA 42 hrs 6 hour pulse 6 hour pulse N o-VO R InitialD ose PH A 0 200 400 600 1000 1500 2000 RelativeHIV-1gagRNAcopies No VOR Initial Dose PHA 6 hour VOR p = 0.0001 N o-VO R Pre-second dose Post-seconddose 0 200 400 600 1000 1500 2000 RelativeHIV-1gagRNAcopies No VOR Pre-Second Dose Post-Second Dose 24 hour VOR p = 0.0626 VORUntreated Washed 18 hrs 66 hrs 48 hour VOR N o-VO R Pre-Second dose Post-Second D ose 0 500 1000 1500 2000 RelativeHIV-1gagRNAcopies No VOR Pre-Second Dose Post-Second Dose p = 0.0029 N o VO R Pre-Second D ose Pre-Second D ose 0 200 400 600 1000 1500 2000 RelativeHIV-1gagRNAcopies No VOR Pre-Second Dose Post-Second Dose 72 hour VOR p = 0.0118 Archin et al., JCI 2017
- 21. Baseline Multidose 0 500 1000 1500 2000 2500 p<0.0001 HIV-1gagRNAcopiesperwell Participant 720 Participant 674 Multiple Doses: 10 x 72hr Baseline Multidose 0 1000 2000 3000 4000 5000 p<0.0001 HIV-1gagRNAcopiesperwell Mean + SD; Mann-Whitney Test Multiple doses of VOR administered every 72hrs resulted in sustained increased in HIV RNA in Participants 767 and 674 Archin et al., JCI 2017
- 22. Searching for new Latency Reversal approaches Histone Crotonylation New Targets Polycomb Complex Targeting Histone Methylation Targeting Key RNA/Protein Complexes: p-TEFb NFkB signaling
- 23. Challenges to clearing persistent infection after latency reversal • Recent absence of antigen – low frequency of HIV-specific antiviral responses • Immune dysfunction, deletion, or exhaustion • Archived viral diversity, including immune escape • Latency Reversing Agents (LRAs) are host- targeted, and alone or in combination may alter antiviral immune response • Viral antigen is rare, anatomically dispersed, and may be transiently expressed
- 24. Augmenting Antiviral Clearance Expanded & augmented NK cell approaches Enhancing antibody mediated HIV clearance mechanismsExpanded & augmented T cell approaches Immune Checkpoint Inhibitors and Innate Signaling HIV-1 VACCINE
- 25. Latency Reversing Agent & ART PBMCs oror Remove ART, add Effectors Limiting Dilution Co-culture Measure replication- competent HIV production at 2 weeks Culture Resting CD4+ cells Resting CD4+ cells by negative selection Remove Effectors Augmented Effectors Latency Clearance Assay (LCA) Autologous Effectors No Effectors
- 26. N o stim N o C D 8s E:T 1:10 E:T 1 :1 0 1 0 2 0 3 0 4 0 %ofplatedwellsp24+ * +V O R A . * Vorinostat renders the Replication-Competent Latent Reservoir of HIV Vulnerable to Clearance by CD8 T cells 335 nM x 6 hours *indicates p<0.01 by Kruskal-Wallis with Dunn’s posttest correction for multiple comparisons Sung et al., 2017 Latency Clearance Assay
- 28. HIV specific Ex-vivo Expanded T cells (HXTCs) HIV Specific CTL IL-2 IL-12 IL-15 IL-15IL-7 CD80/86 4-IBBL CD32 + K562 Irradiated PHA blast Irradiated Mature DC + gag/ pol/ nef peptides Immature DC PBMC T cell ARVs Cath Bollard Clio Rooney Participant 425 Participant 532 Participant 250 0 20 40 60 80 100 120 %Viralrecovery †† †† † †HXTC p<0.05 compared to No CD8s ††HXTC p<0.05 compared to CD8s HXTCs Reduce HIV Recovery from autologous resting CD4+ T cells pulsed with VOR Sung et al. JID 2015 HXTCs CD8s
- 29. Dual Affinity ReTargeting (DARTs) Molecules for HIV Dual Affinity Re-Targeting proteins direct T cell – mediated cytolysis of latently HIV-infected cells. Sung, JA, Pickeral, J, Liu, L, Stanfield-Oakley, SA, Lam, CY, Garrido, C, Pollara, J, LaBranche C. Bonsignori, M. Moody, MA, …..Haynes, BF, Nordstrom JL, Margolis, DM, Ferrari, G. JCI 2015 • Screened ADCC, non-neutralizing Abs that bound HIV-infected CD4 T cells for optimal ADCC • Constructed DARTs with non- Neutralizing mAbs A32XCD3 and 7B2XCD3 • Showed DARTs + CD8 CTL eliminated HIV-infected CD4 T cells in vitro by CD8 T cell-mediated cytolysis.
- 30. *p < 0.05 Sung unpublished data
- 31. Combination Latency Reversal and Clearance Trial Step 4 AGS EOS Term ~Week 15 - Week 24 ~Week 73 - Week 89 Visit 8 27 AGS Manufacturing EOS visits - no dosing Leukapheresis & rc-RNA IMand Proviral Viral inhibition and latency clearance assays visits - Vorinistat doses Safety labs SCA Acetylation visits - AGS-004 injections PK Samples Exploratory Immunology rc-RNA measurement determines progress Single Dose Step 7 AGS Dosing Step 8 Multiple VOR Dosing Step 1 Screen Step 2 Enrollment Step 3 Interval Step 5 AGS Dosing Step 6 Multiple VOR Dosing ~Week 39 -Week 54 Visits 14 - 17 Visits 18 - 22 ~Week 52 -Week 73 Visits 23 - 26 ~Week 64 -Week 81 Paired Doses Approximately Weeks0 -8 Injections X4 Interval dosing X 10 doses Injections X4 Interval dosing X 10 doses ~Week 27 -Week 46 Visit 9 - 13 BASELINE ~Week 6 -Week 13 Visits 1&2 Visit 3 & 4 ~Week 10 -Week 18 Vists 5 - 7 Vaccine Vaccine LRA x 10 Vaccine = Argos dendritic cell therapy LRA = vorinostat LRA x 1 LRA x 2 LRA x 10
- 32. Study endpoints Primary: • Safety • Decline in frequency of resting CD4+T cell infection (IUPM) via a quantitative viral outgrowth assay (QVOA) Secondary: • Resting CD4 cell-associated HIV RNA and DNA • Ultrasensitive measures of cell-associated viral antigen • Viral species and RNA genotypes recovered from resting CD4 T cells • Anti-HIV CD8 cell frequencies and specificities • Single-copy viremia
- 33. Summary • We are learning how to develop and test latency reversal agents in the clinic • A wide array of immunotherapies are under study to assist in the clearance of persistent HIV infection • At this stage we cannot be sure if the lack of early success is due to inadequate Latency Reversal, inefficient Viral Clearance, or both
- 36. Acknowledgements Nancie Archin Julia Sung Carolina Garrido Anne Marie Turner Nilu Goonetilleke Victor Garcia-Martinez Ed Browne Cynthia Gay, JoAnn Kuruc & trials team Joe Eron, Susan Pedersen & ACTU Yara Park & UNC Blood Bank Ron Swanstrom, Sarah Josephs, Shuntai Zhu, & UNC CFAR Mary Napier Charles Nicolette Mark DeBenedette Irina Tcherepanova University of North Carolina Cath Bollard Clio Rooney Cath Bollard, S Patel, CR Cruz, S Lam Rick Dunham Heather Madsen Brian Johns Thanks to participants for their altruism
- 37. • Find patients earlier, bring treatment to them • Develop ways to use ART as prevention • Develop vaccines that are protective • Test curative therapy
Editor's Notes
- (CD3+CD4+CD45RA-CD27+CCR7+)
- HIV latency. Potential obstacles to HIV eradication. (A) True virological and transcriptional latency, with little HIV RNA expression, and no detectable HIV antigen presentation. (B) So-called “active latency” with ongoing production of HIV RNA and antigen. (C) Proliferation of latently infected cells, driven by homeostatic forces, or by dysregulation of the host gene program by a viral integrant, without viral production. (D) The possibility that de novo infection occurs despite effective ART. (E) Failure of immune clearance owing to viral epitope escape or host immune exhaustion.
- Persistent, latent infection of memory CD4 cells decays slowly over time. Residual HIV replication and proliferation of latently infected cells might increase the frequency of latent infection, but these forces must be slightly outweighed by those that naturally deplete latent infection because a slow decay of latent infection is uniformly seen in stably treated patients. The goal of antilatency therapy is to effectively accelerate the clearance of persistent infection across all reservoirs of persistent infection. This data, collected over 10 years ago (35), has recently been precisely reproduced in a contemporary patient cohort using improved ART (36). [Adapted by permission from Macmillan Publishers, Nature Med. 2003.]
- Current paradigm that is beginning to be tested
- The first combination LRA and immunotherapy study underway is depicted here, with participants receiving multiple vorinostat doses along with the Argos dendritic cell therapy. EOS?
- As well as the industry and academic partners that comprise this collaboratory. Thank you for your attention.
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