1. HIV: Science, Sex and Society. Lecture 1. Mechanisms of viral persistence in the face of antiviral therapy. Mario Stevenson, Ph.D Department of Medicine.

2. Main topics: Implications of plasma viremia Use of antivirals and viremia to model reservoir dynamics The concept of viral latency Homeostatic maintenance of viral DNA Cryptic replication and viral sanctuaries

4. months on antiviral cocktails (ART) virus particles in plasma (log10) Antiviral treatment: impact on viremia 3210 0 10 7 10 6 10 5 10 4 10 3 10 2 10 1

5. months on antiviral cocktails (ART) Antiviral treatment: impact on viremia 3210 0 10 7 10 6 10 5 10 4 10 3 10 2 10 1 Antiretroviral cocktails cause a 5-6 log reduction in levels of virus in blood within several months of treatment initiation! virus particles in plasma (log10)

7. What sustains HIV-1 in the face of HAART? Modeling the dynamics of the viral reservoirs!

8. Faucet = virus source or reservoir Water = virus Drain = virus clearance processes

9. If there are different faucets or reservoirs, water in each sink empties at different rates

10. Cellular Reservoirs of HIV-1 Replication and Persistence M. Stevenson Nature Med. 2003

11. What sustains HIV in the face of therapy? Latent infection of CD4 T-cells or macrophages Homeostatic maintenance of viral DNA Cryptic replication in CD4 T-cells or macrophages Latent infection of CD4 T-cells or macrophages Homeostatic maintenance of viral DNA Cryptic replication in CD4 T-cells or macrophages

12. What sustains HIV in the face of therapy? Latent infection of CD4 T-cells or macrophages Homeostatic maintenance of viral DNA Cryptic replication in CD4 T-cells or macrophages Latent infection of CD4 T-cells or macrophages Homeostatic maintenance of viral DNA Cryptic replication in CD4 T-cells or macrophages

13. What is latency? Latency is a reversibly non-productive state of infection. This term describes an infected cell that is not producing virus but that has the capacity to do so under the appropriate conditions.

14. Courtesy Tae-Wook Chun

15. The problem with latency! In a latent infection, the virus is inactive- in other words, the only thing that distinguishes the latently infected cell from an uninfected cell is a viral chromosome. Unfortunately, since there is no viral RNA or protein being made, the latently infected cell is invisible to the immune system or to the antiviral drugs. In addition, the CD4 T-cell is long-lived!

16. Time on HAART (years) Latent Cell Frequency Slow decay of latently infected CD4 + T cells Time to eradication > 73.4 years Courtesy Bob Siliciano 45 30 15 0

17. What sustains HIV in the face of therapy? Latent infection of CD4 T-cells or macrophages Homeostatic maintenance of viral DNA Cryptic replication in CD4 T-cells or macrophages Latent infection of CD4 T-cells or macrophages Homeostatic maintenance of viral DNA Cryptic replication in CD4 T-cells or macrophages

18. Mechanisms of HIV persistence T cell survival Homeostatic proliferation Finzi et al. Science 1997 ; Wong et al. Science 1997 ; Chun et al. PNAS 1997 ; Palmer et al. PNAS 2008 ; Chomont et al. Nat Med 2009 Latent reservoir

19. Mechanisms of HIV persistence T cell survival Homeostatic proliferation Finzi et al. Science 1997 ; Wong et al. Science 1997 ; Chun et al. PNAS 1997 ; Palmer et al. PNAS 2008 ; Chomont et al. Nat Med 2009 Latent reservoir Homeostatic proliferation may maintain dead viruses! 6% of human DNA comprises dead ancestral viruses! This poses a problem when trying to determine if an individual has been cured! Homeostatic proliferation may maintain dead viruses! 6% of human DNA comprises dead ancestral viruses! This poses a problem when trying to determine if an individual has been cured!

20. Size of latent reservoir VOA Intact HIV DNA Scale=100/10 6 62 fold Ho et al, Cell, 2013

21. What sustains HIV in the face of therapy? Latent infection of CD4 T-cells or macrophages Homeostatic maintenance of viral DNA Cryptic replication in CD4 T-cells or macrophages Latent infection of CD4 T-cells or macrophages Homeostatic maintenance of viral DNA Cryptic replication in CD4 T-cells or macrophages

22. Multiple viral RNA species produced in infected cells

23. Multiple forms of viral RNA – prematurely terminated, spliced, unspliced – produced in infected cells. CA-RNA present in infected cells from individuals on suppressive ART (Furtado NEJM, 2009; Yerly JID, 2009; Hockett J. Exp. Med, 1999) CA-RNA predicts time to rebound post-treatment interruption Multiple forms of viral RNA – prematurely terminated, spliced, unspliced – produced in infected cells. CA-RNA present in infected cells from individuals on suppressive ART (Furtado NEJM, 2009; Yerly JID, 2009; Hockett J. Exp. Med, 1999) CA-RNA predicts time to rebound post-treatment interruption

24. ART de-intensification studies Rothenberger et al., PNAS. March 2015

25. Baseline reservoir size and timing of viral rebound Jonathan Li

26. If there is some residual viral activity, how could this occur in the face of such powerful antiviral forces? There may be anatomic sites that act as sanctuaries for the virus

27. Reservoirs of HIV-1 persistence in HAART M. Stevenson, Scientific American 2008

28. Reservoirs of HIV-1 persistence in HAART M. Stevenson, Scientific American 2008 Drugs may not access all tissues with equal efficiency. This could create sanctuaries!

29. Sanctuaries for viral replication?

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31. ART intensification studies

32. In infected individuals on potent therapy, are the drugs completely shutting off the faucet?