Abstract #: WEAA0104

1. Abstract #: WEAA0104 Probiotic and IL-21 Treatment Promotes Th17 Cell Recovery in ARV-Treatment of Pigtail MacaquesAlexandra M. OrtizLaboratory of Jason BrenchleyJuly 23, 2014

2. Th17 Cells and Probiotics in HIV-associated Intestinal Immune Dysfunction • HIV is characterized by persistent GI immune dysfunction. • Preferential Th17 loss • Th17 are antimicrobial CD4+ T-cells essential for intestinal immunity. • Generated in the context of commensal signaling • Immune dysfunction correlates with microbial translocation and dysbiosis • Probiotic supplementation of chronically-infected, ARV-treated macaques: • Increased colonic CD4+ T-cell function and frequency • Increased function but not frequency of colonic Th17 cells • IL21 therapy in acutely SIV-infected macaques promotes Th17 retention during treatment (PallikkuthPLoSPathog2013) (KlattJCI 2013)

3. Experimental Design • Animals: 11 Pigtail Macaques (Maccacanemestrina) • 6 animals treated with ARV + Probiotics (Pb) + IL-21, 5ARV-only controls • Infection: 3000 TCID50, SIVmac239i.v. • ARVs: 20-30 mg/kg PMPA i.v., 30mg/kg FTC i.v., 240 mg L’812 – daily • Probiotics: 1011 live bacteria (VSL#3) oral - daily • Bifidobacterium: breve, longum and infantis • Lactobacillus: acidophilus, plantarum, paracasei, bulgaricus • Streptococcus thermophilus • IL-21: 50 ug/kg IL-21, s.c. – 5 weekly doses, early and late chronic • RM IL-21 fused to RM IgG2 Fc, produced in S-2 cells

4. IL-21 and Probiotic Therapy do not Interfere with Viral Suppression Treatment initiation Viral Load (log copies/mL) PTA2P031 (early) IL21 (late) IL21 Day (relative to infection)

5. IL-21and Probiotic Therapy Improve Intestinal CD4+ T-Cell Frequency Rectal Biopsy Jejunum ARV only ARV+Pb+IL21 PTA2P031 Percent CD4+ of CD3+ T-cells PTA2P031 Day (relative to infection)

6. IL-21and Probiotic Therapy do not Induce CD4+ T-cell Activation PBMC LN ARV only ARV+Pb+IL21 Percent Ki67+ of CD4+ TM Jej RB BAL Day (relative to infection)

7. IL-21and Probiotic Therapy Improve Intestinal Th17 Frequency Rectal Biopsy Jejunum ARV only ARV+Pb+IL21 Percent IL17+ of memory CD4+T-Cells Day (relative to infection)

8. IL-21and Probiotic Therapy Improve Polyfunctional Th17 Recovery Rectal Biopsy Jejunum ARV only ARV+Pb+IL21 Percent Polyfunctional of CD3+ T-Cells Day (relative to infection)

9. IL-21 and Probiotic Therapy Moderately Reduced Colonic IDO Expression Mesenteric LN Colon ARV only ARV+Pb+IL21 Percent IDO+ Tissue

10. IL-21and Probiotic Therapy Reduce Comorbidities Percent Incidence-Free Animals Day (relative to infection)

11. Conclusions and Future Directions • Therapeutic treatment with probiotics and IL21 improved disease progression in ARV-treated macaques as measured by: • Intestinal CD4+ T-cell recovery • Improved intestinal Th17 frequency and functionality • Decreased incidence of health complications • By what mechanism are Th17 cells expanded? • Would treatment promote enhanced protection against microbial challenge? • Does probiotic and IL21 treatment reduce microbial translocation and dysbiosis?

12. Acknowledgements Brenchley Lab • Jason Brenchley • Nina Calantone • Kirby Carmack • Sarah DiNapoli • Zachary Klase • Molly Perkins • Carol Vinton Veterinary Staff Heather Cronise Richard Herbert Joanna Swerczek LMM Core NCI-NIH • Jake Estes Emory University – IL21 • MirkoPaiardini • Francois Villinger Gilead – PMPA/FTC • Michael Miller Merck– L’812 • DariaHazuda Sigma-Tau – VSL#3 • Claudio De Simone