1. Randomized Therapeutic Vaccine Trial of Canarypox-HIV (CP-HIV) + Dendritic Cells vs. CP-HIV Alone in HIV- infected Patients on Antiretroviral Therapy: ACTG A5130 ALVAC-HIV (vCP1452): CP vector that contains HIV env and gag genes and CTL epitopes from nef and pol. HIV+ subjects vaccinated with CP-HIV alone have small and transient increases in HIV-specific immunity. However, vaccination with CP-HIV in patients treated with ART during acute or chronic HIV infection does not lead to virologic control during treatment interruption Jacobson JID 194:623; Kinloch-de Loes JID 192:607; Autran AIDS 22:1313 Dendritic cells infected with CP-HIV elicit HIV-specific CD4 & CD8 immune responses in vitro Engelmayer. J Virol. 75:2142 Hypothesis: Immunization of HIV+ patients on ART with DCs plus CP-HIV will be more immunogenic and lead to better control of viremia during a treatment interruption than vaccination with CP-HIV alone

2. = vaccination with DCs + CP-HIV or CP-HIV alone. Study Schema Wk 15Wk 7Wk 3 Arm B CP HIV N=15 29 HIV+ Subjects On ART VL<50 CD4 > 400 12-week ATI. VL setpoint at end of ATI At wks 3 and 7, subjects also received keyhole limpet hemocyanin (KLH). KLH was a control to determine whether priming of new immune responses occurred. Primary Endpoints: 1) VL setpoint at the end of a 12- week ATI; 2) Safety: ≥ grade 3 adverse event that is at least possibly related to study treatment. 1:1 Arm A DC+CP HIV N=14

3. Inject CP-HIV-infected DCs at wk 3, 7 & 15 (1.5 to 6 million DCs, SQ) Based on slide from Nina Bhardwaj, M.D., Ph.D. Infect with CP- HIV (ALVAC vCP1452) Mature DCs using cocktail of IL-1 , IL-6, TNF- , PGE2 Expand monocyte-derived DCs using IL-4 and-GM CSF (7 days in a cell processing lab at MGH) Isolate PBMC from leukapheresis product (1-3 billion PBMC) HIV+ pt on antiretroviral therapy

4. Effect of Vaccination on LPA Subjects who received KLH-pulsed DCs had a significantly greater increase in KLH LPA response prior to ATI than subjects who received KLH alone –Median SI fold change 5.7 in arm A vs. 1.8 in arm B, p=0.007 No change in LPA response to tetanus, candida or 5 pools of HIV peptides A: KLH-pulsed DCs. B: KLH alone

5. 5/12 (42%) subjects in arm A and 5/12 (42%) subjects in arm B had a >3-fold increase in summed ELISPOT response over baseline at 1 or more time points after vaccination and before ATI Fold-increase in ELISPOT Responses During Vaccination Responder: 3-fold increase in ELISPOT response that is also >30 SFC/million PBMC Arm A (n=12) DC+ CP-HIV Arm B (n=12) CP-HIV alone

6. Viral Load during ATI: Summary No difference in VL setpoint between subjects in Arm A and B (medians 4.1 and 4.5 log 10 c/mL) 4/13 (31%) in DC+CP-HIV arm had VL setpoint <5,000 compared with 0/13 (0%) in CP-HIV alone arm (p=0.096) –Of the 4 subjects who achieved VL setpoint 5,000 on follow-up (7 wks later) VL=5000 Setpoint VL <5000 c/mL Arm A (n=14) DC+CP-HIV Arm B (n=15) CP-HIV alone

7. ELISPOT response and VL Setpoint during ATI Summed ELISPOT level after vaccination negatively but weakly correlated with VL setpoint (not sig.) Greater fold-increases in gag, nef or summed ELISPOT response after vaccination are associated with lower VL setpoint (all p-values <0.05)

8. Conclusions CP-HIV & DC+CP-HIV therapeutic vaccines are safe and immunogenic in HIV-infected patients DCs effective as an adjuvant to prime responses to KLH, but did not clearly enhance HIV-specific ELISPOT responses Suggestion that subjects vaccinated with CP-HIV+DC had a higher rate of VL setpoint <5,000 c/mL during ATI than subjects immunized with CP-HIV alone; however, control was transient Subjects who had higher fold-increases in HIV-specific ELISPOT responses after vaccination had lower VL setpoints Further studies of the correlates of antiviral immunity and methods to more effectively boost antiviral immunity are warranted

9. Acknowledgments ACTG 5130 team Jeff Jacobson, protocol chair; Rajesh Gandhi and Nina Bhardwaj, vice-chairs David O’Neill, Ellen Chan, Ron Bosch Pat Bucy, Mary Marovich, Lynn Baglyos, Reena Masih, Barbara Schock, Lynette Purdue and the 5130 team Massachusetts General Hospital Teri Flynn, Amy Sbrolla, Janet Shopis, Kathy Habeeb, Nicole Burgett, Gil Roy, Daniel Kavanagh, Doug Kwon, Bruce Walker NYU Cancer Institute Vaccine Center Crystal M. Cruz, Angelica Angiulli and Francesca Angiulli Burroughs Wellcome, Elizabeth Glaser, Doris Duke Beth Israel, NY Ann Marshak, Gwen Constantini, Sondra Middleton, Donald Garmon, Scott Barnett, Donna Mildvan Study subjects at MGH and Beth Israel, NY