Control of Viremia and Prevention of AIDS following Immunotherapy of SIV-Infected Macaques with Peptide- Pulsed Blood Robert De Rose 1 , Caroline S. Fernandez 1 , Miranda Z. Smith 1 , C. Jane Batten 1 , Sheilajen Alca ˆ ntara 1 , Vivienne Peut 1 , Erik Rollman 1 , Liyen Loh 1 , Rosemarie D. Mason 1 , Kim Wilson 2 , Matthew G. Law 3 , Amanda J. Handley 1,4¤ , Stephen J. Kent 1 * 1 Department of Microbiology and Immunology, University of Melbourne, Parkville, Victoria, Australia, 2 National Serology Reference Laboratory, Fitzroy, Victoria, Australia, 3 National Centre for HIV Epidemiology and Clinical Research, University of New South Wales, Sydney, New South Wales, Australia, 4 Opal Therapeutics Pty Ltd, Melbourne, Victoria, Australia Abstract Effective immunotherapies for HIV are needed. Drug therapies are life-long with significant toxicities. Dendritic-cell based immunotherapy approaches are promising but impractical for widespread use. A simple immunotherapy, reinfusing fresh autologous blood cells exposed to overlapping SIV peptides for 1 hour ex vivo, was assessed for the control of SIV mac251 replication in 36 pigtail macaques. An initial set of four immunizations was administered under antiretroviral cover and a booster set of three immunizations administered 6 months later. Vaccinated animals were randomized to receive Gag peptides alone or peptides spanning all nine SIV proteins. High-level, SIV-specific CD4 and CD8 T-cell immunity was induced following immunization, both during antiretroviral cover and without. Virus levels were durably ,10-fold lower for 1 year in immunized animals compared to controls, and a significant delay in AIDS-related mortality resulted. Broader immunity resulted following immunizations with peptides spanning all nine SIV proteins, but the responses to Gag were weaker in comparison to animals only immunized with Gag. No difference in viral outcome occurred in animals immunized with all SIV proteins compared to animals immunized against Gag alone. Peptide-pulsed blood cells are an immunogenic and effective immunotherapy in SIV-infected macaques. Our results suggest Gag alone is an effective antigen for T-cell immunotherapy. Fresh blood cells pulsed with overlapping Gag peptides is proceeding into trials in HIV-infected humans. Citation: De Rose R, Fernandez CS, Smith MZ, Batten CJ, Alca ˆntara S, et al. (2008) Control of Viremia and Prevention of AIDS following Immunotherapy of SIV- Infected Macaques with Peptide-Pulsed Blood. PLoS Pathog 4(5): e1000055. doi:10.1371/journal.ppat.1000055 Editor: Richard A. Koup, National Institutes of Health-NIAID, United States of America Received January 24, 2008; Accepted April 1, 2008; Published May 2, 2008 Copyright: ß 2008 De Rose et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Funding: Supported by OPAL Therapeutics Pty Ltd, the Australian Centre for HIV and Hepatitis Research, and Australian National Health and Medical Research Council grants 299907, 251653, 454553, 359281. The funders played no role in the design and conduct of the study, nor in the collection, analysis and interpretation of the data, nor in the preparation, review or approval of the manuscript. Competing Interests: SJK spun out a venture capital–backed biotechnology start-up company, OPAL Therapeutics Pty Ltd, to pursue this technology towards clinical trials. SJK, CJB and the University of Melbourne hold shares in this company. AJH has been an employee of OPAL Therapeutics. * E-mail: skent@unimelb.edu.au ¤ Current address: Medicines Development Ltd, Melbourne, Victoria, Australia Introduction Several attempts at immunotherapy of HIV using more conventional vaccines have thus far been poorly immunogenic and weakly efficacious in human trials [1,2,3,4]. The use of professional antigen-presenting cells such as dendritic cells to deliver HIV immunotherapies has shown strong immunogenicity efficacy in macaques and pilot humans studies but is limited to highly specialized facilities [5,6,7]. A simple intermittent immu- notherapy that reduces the need for long-term antiretroviral therapy (ART) would be a quantum advance in treating HIV. We recently reported the robust T-cell immunogenicity of treating unfractionated whole blood or peripheral blood mono- nuclear cells (PBMC) with overlapping peptides of SIV, HIV-1 or hepatitis C virus in outbred pigtail monkeys [8,9]. We termed this simple immunotherapy OPAL (Overlapping Peptide-pulsed Autologous ceLls). This technique is attractive since there is no prolonged ex vivo culture of antigen-presenting cells, robust CD4 and CD8 T-cell responses to both structural and regulatory proteins can be induced, and peptide antigens are simple to manufacture to high purity. This study assessed whether OPAL vaccination improves the outcome of SIV-infected monkeys. Considerable debate exists regarding the most effective antigens to target for T-cell based therapeutic HIV vaccination. It has been widely believed that broader immunity to multiple proteins would be more efficacious [10,11]. In contrast, recent studies highlight the effectiveness of Gag-specific T cell immunity in comparison to T cell immunity to other antigens. We therefore also assessed whether narrower responses induced only to SIV Gag are as effective as more broadly targeting all 9 SIV proteins. Materials and Methods Animals Juvenile pigtail macaques (Macaca nemestrina) free from Simian retrovirus type D were studied in protocols approved by PLoS Pathogens | www.plospathogens.org 1 May 2008 | Volume 4 | Issue 5 | e1000055