Novel Approaches for the Assessment of the In Vivo Residual Virus Pool and Viral Eradication Strategies in SIV-infected Rhesus Macaques
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1 Towards an HIV Cure Pre-Conference Symposium 20 & 21 July 2012 Novel Approaches for the Assessment of the In Vivo Residual Virus Pool and Viral Eradication Strategies in SIV-infected Rhesus Macaques Gregory Q. Del Prete, Ph.D. AIDS and Cancer Virus Program, SAIC-Frederick, Inc. Frederick National Laboratory for Cancer Research Frederick, MD, USA Frederick National Laboratory AIDS and Cancer Virus Program
2 Advantages of Non-Human Primate Models in studies of AIDS virus latency/eradication strategies Experimental Control Virus/Infection: Route, dose, timing, identity (clones) Specimen sampling Precise timing Sampling options more extensive than in humans, including scheduled necropsy Interventional Latitude Given nature and potential safety issues of proposed interventions, animal models critical
3 Challenges of Working with NHP Models Monkeys are not people MHC, NKs, etc. Blood volume limitations, fewer cells Drugs: PK, PD, bioavailability differences SIV is not HIV Drugs: ARV susceptibility differences (e.g. NNRTIs) Biology of reservoirs
4 Presentation Topics Novel 3 class, sustainable ART regimen to achieve and maintain plasma SIV RNA < 30 copy Eq/mL in unselected, SIVmac239 infected Indian rhesus macaques Novel ex vivo SIV RNA induction assay to evaluate the frequency of cells containing inducible SIV RNA from in vivo ART treated macaques
5 cart Regimen Specific goal: Suppress viral replication to clinicallyrelevant levels (< 50 RNA copy Eq/ml) in unselected SIVmac239-infected Indian rhesus macaques Feasible, safe, sustainable 5 Drug cart Regimen (3 Drug Classes) 2 NRTIs: Tenofovir (PMPA), FTC 1 INi: Dolutegravir Triple Coformulation Once Daily SubQ Injectable 1 boosted PI: Ritonavir boosted Darunavir Twice Daily Oral
6 5 Drug cart Regimen in SIVmac239- infected Indian Rhesus Macaques 1.E+08 cart 1.E+07 Plasma VL (RNA Copy Eq./ml) 1.E+06 1.E+05 1.E+04 1.E+03 1.E+02 DCW9 DCEG DCT3 DCJB DCCN DCHV 1.E RNA copy eq/ml 1.E Weeks Postinfection
7 Histone Acetylation Changes with HDACi treatment Peripheral blood CD4+ T cells from ART-suppressed SIVmac239-infected Indian rhesus macaques, treated DCT3 ex vivo with HDACi (SAHA, Romidepsin): Modulate histone acetylation? Induce virus expression? DCT3 DCT3_DMSO_ DCT3_PMA_I_ DCT3_RomHi_ DCT3_RomLo_ DCT3_SAHAHi_ DCT3_SAHALo_ Count Ac-Histone H4 Vehicle Ctrl PMA/Ionomycin SAHA 5 μm SAHA 1 μm Romidepsin 40 nm Romidepsin 5 nm
8 Ex Vivo Virus Induction Assay Evaluate frequency of CD4s harboring inducible SIV RNA Enriched blood CD4s treated and plated in replicate wells containing equivalent number cells/well (1x10 5, 2x10 4, 4x10 3, etc.) Neg.Ctrl (Vehicle) SAHA (5 um) SAHA (1 um) Romidepsin (40 nm) Romidepsin (5 nm) Pos. Ctrl (PMA/Iono.) A B C D E F G H /10 5/10 3/10 6/10 3/10 7/10
9 Ex Vivo Induction Assay % Wells With Induced SIV RNA Input Cells/Well 1 x 10^5 2 x 10^4 4 x 10^3 0 Vehicle Ctrl (Neg) Positive Ctrl SAHA (5 um) SAHA (1 um) Romidepsin (40 nm) Romidepsin (5 nm)
10 Summary Safe and effective sustainable ART regimen for durable suppression of SIVmac239 to clinically relevant levels in Indian rhesus macaques. HDACi modulate histone acetylation and induce SIV RNA production ex vivo in CD4+ T cells from ART-suppressed animals Establishes methods to evaluate strategies directed toward HIV (functional) cure in primary SIV/macaque cells; enables testing for impact on the inducible viral reservoirs in vivo
11 Future Directions Further assay optimization and application Determine how inducible reservoir in LNs and other tissues compare with blood Correlate ex vivo induction assay results with co-culture (IUPM) assays
12 Acknowledgments ACVP, SAIC-F Jeffrey Lifson Doug Schneider Rebecca Kiser Vicky Coalter Adam Wiles Rodney Wiles LASP, SAIC-F Jeremy Smedley Rhonda Macallister Mercy Gathuka Gilead Romas Geleziunas Joseph Hesselgesser Bing Lu Jim Zheng Bei Li Jillian Hattersley Merck Daria Hazuda Christopher Tan John Wai Rosa Sanchez Mike Piatak Kelli Oswald Rebecca Shoemaker Randy Fast C. Mac Trubey Abigail Lara Session 6: Abstract # July 14:00-15:30 Funding NCI/NIH Contract HHSN E
13 cart (started 4 wpi) SubQ: Tenofovir (20 mg/kg/day) FTC (40 mg/kg/day) Dolutegravir (2.5 mg/kg/day) P.O.: Darunavir (600 mg, b.i.d.) Ritonavir (100 mg b.i.d.)
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