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Schedule-Dependent Synergy between the Heat Shock Protein 90 Inhibitor 17-(Dimethylaminoethylamino)-17-Demethoxygeldanamycin and Doxorubicin Restores Apoptosis to p53-Mutant Lymphoma Cell Lines

Authors' Affiliations: ¹ Laboratory of Human Carcinogenesis, Center for Cancer Research, National Cancer Institute and ² Hematology Branch, National Heart, Lung and Blood Institute, NIH, Bethesda, Maryland

Requests for reprints: Ana I. Robles, Laboratory of Human Carcinogenesis, National Cancer Institute/NIH, MSC 4258, Room 3060, Building 37, 37 Convent Drive, Bethesda, MD 20892. Phone: 301-496-1729; Fax: 301-496-0497; E-mail: roblesa{at}mail.nih.gov.

Purpose: Loss of p53 function impairs apoptosis induced by DNA-damaging agents used for cancer therapy. Here, we examined the effect of the heat shock protein 90 (HSP90) inhibitor 17-(dimethylaminoethylamino)-17-demethoxygeldanamycin (DMAG) on doxorubicin-induced apoptosis in lymphoma. We aimed to establish the optimal schedule for administration of both drugs in combination and the molecular basis for their interaction.

Experimental Design: Isogenic lymphoblastoid and nonisogenic lymphoma cell lines differing in p53 status were exposed to each drug or combination. Drug effects were examined using Annexin V, active caspase-3, cell cycle, and cytotoxicity assays. Synergy was evaluated by median effect/combination index. Protein expression and kinase inhibition provided insight into the molecular mechanisms of drug interaction.

Results: Presence of mutant p53 conferred increased survival to single agents. Nevertheless, DMAG showed synergistic toxicity with doxorubicin independently of p53 status. Synergy required exposure to doxorubicin before DMAG. DMAG-mediated down-regulation of CHK1, a known HSP90 client, forced doxorubicin-treated cells into premature mitosis followed by apoptosis. A CHK1 inhibitor, SB-218078, reproduced the effect of DMAG. Administration of DMAG before doxorubicin resulted in G₁-S arrest and protection from apoptosis, leading to additive or antagonistic interactions that were exacerbated by p53 mutation.

Conclusions: Administration of DMAG to doxorubicin-primed cells induced premature mitosis and had a synergistic effect on apoptosis regardless of p53 status. These observations provide a rationale for prospective clinical trials and stress the need to consider schedule of exposure as a critical determinant of the overall response when DMAG is combined with chemotherapeutic agents for the treatment of patients with relapsed/refractory disease.

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				T. T. Koll, S. S. Feis, M. H. Wright, M. M. Teniola, M. M. Richardson, A. I. Robles, J. Bradsher, J. Capala, and L. Varticovski HSP90 inhibitor, DMAG, synergizes with radiation of lung cancer cells by interfering with base excision and ATM-mediated DNA repair Mol. Cancer Ther., July 1, 2008; 7(7): 1985 - 1992. [Abstract] [Full Text] [PDF]