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Abstract 1831 Strong epidemiological association of Epstein-Barr Virus (EBV) with various human lymphoid malignancies and in vitro studies demonstrating tumorigenic activity of many EBV latent gene products suggest a causal relationship between EBV and these diseases. However, as EBV maintains a latent state of infection in these lymphomas, typical anti-herpesviral drugs, such as the nucleoside analogs ganciclovir (GCV) or acyclovir, are ineffective as these pro-drugs require expression of a lytic phase EBV protein, thymidine kinase (TK), for their activity. Therefore, selective induction of EBV lytic-phase gene expression in lymphoma cells that harbor latent EBV, coupled with simultaneous exposure to anti-herpesviral drugs, has been advanced as promising targeted therapy, because of resulting targeting of cytotoxicity to the EBV-infected tumor cells. A variety of agents including short-chain fatty acids and chemotherapeutic drugs, have been used to induce EBV lytic-phase infection in cultured cells, but these in vitro studies have generally not translated into clinical application. We have successfully used arginine butyrate and GCV to treat EBV-positive lymphoid malignancies in a recent Phase I/II clinical trial. In this study of 15 patients with relapsed or refractory EBV-positive lymphoid tumors, 4 patients achieved complete tumor remissions and 6 patients partial tumor remissions. However, the rapid metabolism of butyrate requires continuous IV administration of high doses. Butyrate has pan-HDAC inhibitory activity, and we have established that this activity is responsible for the induction of the EBV-TK protein. In recent years, several potent HDAC inhibitors (HDACi) have been tested in the clinic as anti-cancer agents. In the current study, we have investigated a number of HDACi, including some new, highly-potent compounds, for their potential to induce EBV lytic phase gene expression and to kill EBV-infected cells in combination with anti-herpesviral drugs. Our study included short-chain fatty acids (sodium butyrate and valproic acid); hydroxamic acids [Oxamflatin, Scriptaid, Suberoyl anilide hydroxamic acid (SAHA), Panobinostat (LBH589) and Belinostat (PXD101)]; the benzamide MS275; cyclic tetrapeptide Apicidin, and newly-identified HDAC inhibitor Largazole, which was originally isolated from a marine cyanobacterium. We assayed the induction of lytic phase in EBV-positive lymphoma cell lines exposed to different HDACi for 24–48 hrs, then quantitated the expression of EBV TK and other EBV transcripts by RT-PCR analysis. To determine tumor cytotoxic activity of the combination of HDACi and GCV, EBV+ lymphoma cells were exposed to a range of concentrations of HDACi and GCV for 3 days and then to GCV alone for another 3 days. Efficacy of a particular HDACi in the combination treatment approach was then determined by enumerating living cells. With the exception of SAHA and PXD101, the other HDACi had synergistic activity with anti-viral agents in killing EBV+ lymphoma cells. The hydroxamic acid LBH589, the benzamide MS275, and synthetic largazole derivatives 234a and 234b were 104 to 105-times more potent in killing EBV+ lymphoma cells in presence of GCV, compared to sodium butyrate. The effective concentration of LBH589 was in the range of 50–100 nM, MS275 at 200–500 nM and Largazole 234a and 234b at 100–200 nM. Of note, at these concentrations, the drugs as single agents produced no growth inhibitory activity in the tumor cells. LBH589, MS275 and Largazole 234a and 234b also strongly induced EBV-TK expression in the tumor cells. The effectiveness of these HDACi compounds at such low concentrations makes them potentially applicable as sensitizers to anti-viral therapeutics for the treatment of EBV-associated lymphomas. Our finding therefore provides an intriguing possibility that these novel HDACi may be used as an alternative therapeutic option, in combination with nucleoside antivirals, for the treatment of EBV-associated tumors. Disclosures: Faller: HemaQuest Pharmaceuticals: Consultancy, Equity Ownership. Perrine:HemaQuest Pharmaceuticals, Inc: Employment, Equity Ownership, Membership on an entity's Board of Directors or advisory committees, Patents & Royalties, Research Funding. Williams:HemaQuest Pharmaceuticals: Consultancy, Equity Ownership. Berenson:HemaQuest Pharmaceuticals, Inc: Employment, Equity Ownership, Membership on an entity's Board of Directors or advisory committees, Patents & Royalties.

Abstract 4754 Antiviral drugs alone have been unsuccessful in the treatment of Epstein-Barr virus (EBV)-associated malignancies because the virus maintains a latent state of replication in these tumors. In recent years, we developed a novel therapeutic approach wherein the early lytic phase of the virus is induced prior to the use of cytotoxic antiviral drugs. Butyrate derivatives induce an early lytic pattern of EBV gene expression in patient-derived EBV-positive lymphoblastoid cell lines and, together with the nucleoside analog ganciclovir (GCV), effectively reduce or eliminate tumor growth in humans. In a completed Phase 1/2 trial in which patients with relapsed or refractory EBV-associated lymphoid malignancies were treated with 3 weeks of Arginine Butyrate combined with GCV, tumor responses (CR and PR) occurred in 10 of 15 patients. Butyrate requires administration by prolonged intravenous infusion, which is challenging over long time periods. In the Phase 1/2 clinical trial, tumor responses were observed within a few days of starting therapy. We therefore investigated whether brief or discontinuous exposure to Butyrate is also capable of initiating early lytic-phase gene expression and thymidine kinase induction, and sensitizing EBV-positive lymphoma cells to ganciclovir-mediated cell growth arrest and apoptosis. Multiple daily 6-hr exposures of the EBV-positive Burkitt's lymphoma cell line P3HR1 to butyrate induced sustained expression of the EBV TK and lytic-phase protein BMRF. Discontinuous exposure to butyrate in combination with ganciclovir also induced a similar level of tumor cell death as did continuous treatment, as measured by serial enumeration of viable cells, MTT cell proliferation assays, and measurement of cellular DNA content. Based on these observations, we have initiated a new clinical trial utilizing a 5-day infusion of Arginine Butyrate and 21 days of GCV/valganciclovir for treatment of patients with EBV-positive lymphoid malignancies. The first patient enrolled, with Rituximab-refractory EBV-positive PTLD following a cord stem cell transplant for Hodgkin's Disease, has been treated on this protocol. The therapy was well-tolerated and resulted in rapid resolution of fever and cough of several weeks duration, and a rapid decrease of markedly elevated LDH levels to the normal range. At the end of the first cycle, 4 of 6 target lesions resolved completely, and two additional lesions decreased in size. High EBV, CMV and HHV6 viral loads became undetectable. This response has been durable for 2 months. These findings together suggest that a shorter, more patient-accessible regimen of this virus-targeted therapeutic strategy may be efficacious, and the clinical trial is continuing. Disclosures: Faller: HemaQuest Pharmaceuticals: Consultancy, Equity Ownership, Patents & Royalties, Research Funding. Off Label Use: The use of Ganciclovir, administered in combination with Arginine Butyrate as an inducer of viral TK,to induce apoptosis in EBV-lymphoma and EBV lymphoproliferative disease. Ghosh:HemaQuest Pharmaceuticals: Research Funding. Lerner:HemaQuest Pharmaceuticals: Consultancy. Berenson:HemaQuest Pharmaceuticals: Employment, Equity Ownership, Membership on an entity's Board of Directors or advisory committees. Perrine:HemaQuest Pharmaceuticals: Employment, Equity Ownership, Patents & Royalties, Research Funding.