II. Clinical and Laboratory Studies of the Histone Deacetylase Inhibitor Depsipe

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We have studied the histone deacetylase inhibitor depsipeptide in both the clinic and in the laboratory. We originally became interested in depsipeptide in the context of a clinical trial strategy seeking to identify agents that could overcome or circumvent multidrug resistance. My laboratory identified the histone deacetylase inhibitor depsipeptide as an agent in preclinical development and a substrate for Pgp-mediated efflux. Because depsipeptide is avidly transported by Pgp, and because it induces MDR-1 in the constellation of genes altered by histone acetylation, we planned to eventually develop depsipeptide in combination with a Pgp modulator. However, in the Phase I setting, we made the serendipitous discovery that depsipeptide was highly effective in subsets of T cell lymphoma. While we have continued to be interested in our original strategy of preventing the emergence of resistance to this agent, we have pursued the use of depsipeptide as an orphan drug in T cell lymphoma, using both laboratory and clinical strategies. Our multi-institutional clinical trial for cutaneous and peripheral T cell lymphoma has enrolled 120 patients to date, divided into 6 cohorts. Cohort 1, patients with cutaneous T cell lymphoma with fewer than 2 systemic chemotherapy regimens, is complete and a manuscript is in preparation. The responses to depsipeptide are at times dramatic and have been very durable. As examples, one patient has received therapy continuously for over 5 years, remaining in a partial remission. Another patient remains in complete remission off of therapy for over 3 years. Depending upon how heavily pretreated the patient population being evaluated, the response rate for depsipeptide, now termed romidepsin, in cutaneous T cell lymphoma ranges from 30 - 50%. Gloucester Pharmaceuticals obtained Fast Track and Orphan Drug status from the FDA for development of this therapy for CTCL. A registration trial has completed accrual, and responses from our trial will be included in the NDA package. Our data are being readied for the NDA submission. NCI CTEP and our Cancer Therapeutics Branch (now Medical Oncology Branch) largely pushed the development of this agent alone during a period in which Fujisawa Pharmaceuticals debated the relative merits of becoming involved in an oncology development platform. Responses with PTCL are also durable and Gloucester has developed a registration strategy for that indication as well. A registration-directed Phase II clinical trial in PTCL has been launched by the company, and with Dr. Richard Piekarz as principal investigator at our site, we will join that trial as well. The trial has been submitted to the IRB for review. Our NCI Phase II trial has a major second objective in addition to proving efficacy in the various histologies. That is confirmation of the safety of the agent. EKG abnormalities have been noted following treatment and a great deal of effort has gone into demonstrating the lack of myocardial damage associated with administration of this agent. We reported in June of 2006 in Clinical Cancer Research, our review of 2,051 ECGs obtained in 42 patients treated with depsipeptide. These ECGs demonstrate the previously documented reversible ST and T wave changes in the majority of patients, unassociated with any abnormality in cardiac function or change in troponin level. In addition, a median increase in the corrected QT interval of 14 msec. This study concluded that there was no evidence of cardiac damage resulting from depsipeptide exposure. One final concern noted in the 2006 annual report -- in clinical trials sponsored by CTEP across the nation there were 5 unexpected deaths associated with depsipeptide among over 500 patients treated. Generally, the patients had predisposing factors for sudden death. Implemented in 2006, all depsipeptide protocols now exclude patients with known cardiac disease or pre-existing risks for sudden death. Our ability to intensively characterize the cardiac effects of depsipeptide, an effective agent in a rare disease, is an excellent example of the critical role that the intramural program can play in drug development. This role assumes greater significance when one recognizes that these cardiac findings represent a class effect observed with other histone deacetylase inhibitors as well. The trial has a significant translational component that has consumed a major fraction of my laboratory resources. We have developed a quantitative immunoblot assay for detecting and quantitating histone acetylation in patient samples, principally peripheral mononuclear cells as a surrogate. Results from these assays are currently being compared to pharmacokinetic data. We have also evaluated gene expression including CD25, p21, and MDR1 by RT-PCR, finding that only MDR1 expression is induced sufficiently following depsipeptide for routine assay in patient mononuclear cells. MDR1 is also analyzed in tumor samples before therapy is initiated and then at the time of disease progression. Additional studies include a Phase I trial of depsipeptide on a day 1, 3, and 5 schedule in hopes of achieving a more continuous drug effect. Dr. Richard Piekarz is PI on this study. This study has a focus in thyroid cancer in order to translate the observation made in Dr. Tito Fojos laboratory that among the genes induced by depsipeptide were the genes encoding the sodium iodide symporter and thyroglobulin. Induction of these genes in thyroid cancer cells enhanced the accumulation of radioiodine in vitro. Such an effect in patients with thyroid cancer could lead to increased radioactive iodine uptake. The Phase I trial is ongoing, with samples collected for pharmacokinetic and pharmacodynamic analysis. PBMCs for histone acetylation determination and pre/post treatment tumor samples have been obtained for gene expression analysis. In patients with thyroid cancer (one patient per dose level and then expanding at the MTD), radioiodine imaging is allowed, along with subsequent dosing with radioiodine in the event that increased radioiodine accumulation is demonstrated. Since only patients with thyroid cancer without visible radioiodine uptake are enrolled on the study, this will probably be difficult to achieve. It is our hope that pre/post treatment biopsies will at a minimum show induction of the Na+/I- symporter at the gene expression level, in cells assayed by RT-PCR, if radioiodine uptake cannot be documented. Finally, we have been interested for some time in mechanisms of depsipeptide sensitivity and resistance. This led us to the generation of cell lines with non-Pgp mediated depsipeptide resistance and we have begun to ask whether other mechanisms of resistance can be identified. Preliminary studies suggest that there is a drug accumulation defect in these cells and a mechanism underlying that is being sought. We continue to be interested in the mechanism of action of depsipeptide. At least 5 mechanisms have been cited for histone deacetylase inhibitors: induction of gene expression, acetylation of cytoplasmic proteins and altered function, increased degradation of cytoplasmic proteins due to impaired Hsp90 activity, altered angiogenesis, and mitotic effects. We contributed to this last mechanism where we reported, in collaboration with Drs. April Robbins and Dan Sackett, changes in the centromere following [summary truncated at 7800 characters]