Manipulation of base excision repair to sensitize ovarian cancer cells to alkylating agent temozolomide

Melissa Fishel, Ying He, Martin L. Smith, Mark Kelley

Research output: Contribution to journalArticle

100 Citations (Scopus)

Abstract

Purpose: To improve the treatment of women with ovarian cancer, we are investigating the modulation of a prominent DNA-dnmaging agent, temozolomide, by manipulating the DNA base excision repair (BER) pathway via BER inhibitor, methoxyamine, and overexpression of N-methylpurine DNA glycosylase (MPG). Experimental Design: Enhancement of temozolomide via methoxyamine and MPG overexpression was analyzed using in vitro assays, including 3-(4-5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl) -2H-tetrazolium salt (MTS) assay, apoptosis via Annexin staining, and Western blotting for H2AX phosphorylation to quantitate DNA damage. Results: Our data show that we can effectively modulate the activity of the chemotherapeutic agent, temozolomide, via modulator methoxyamine, in three ovarian cancer cell lines, SKOV-3x, Ovcar-3, and IGROV-1. This enhancement of temozolomide-induced cytotoxicity is not dependent on p53 status as we transfected an ovarian cancer cell line with a dominant-negative p53-expressing plasmid (IGROV-1mp53) and obtained similar results. Our results show that MPG-overexpressing IGROV-1 and IGROV-1mp53 cells are significantly more sensitive to the clinical chemotherapeutic temozolomide in combination with methoxyamine as assayed by cytotoxicity, apoptosis, and levels of DNA damage than either agent alone. Conclusions: These studies show that although clinical trials in ovarian cancer to determine temozolomide single-agent efficacy are in development, through manipulation of the BER pathway, an increase in response to temozolomide is achieved. The combination of temozolomide plus methoxyamine has potential for second-line therapy for patients who have failed standard platinum plus paclitaxel chemotherapy.

Original languageEnglish
Pages (from-to)260-267
Number of pages8
JournalClinical Cancer Research
Volume13
Issue number1
DOIs
StatePublished - Jan 1 2007

Fingerprint

temozolomide
Alkylating Agents
DNA Repair
Ovarian Neoplasms
DNA Glycosylases
DNA Damage
Tetrazolium Salts
Apoptosis
Annexins
Cell Line
DNA
Paclitaxel
Platinum

ASJC Scopus subject areas

  • Cancer Research
  • Oncology

Cite this

Manipulation of base excision repair to sensitize ovarian cancer cells to alkylating agent temozolomide. / Fishel, Melissa; He, Ying; Smith, Martin L.; Kelley, Mark.

In: Clinical Cancer Research, Vol. 13, No. 1, 01.01.2007, p. 260-267.

Research output: Contribution to journalArticle

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abstract = "Purpose: To improve the treatment of women with ovarian cancer, we are investigating the modulation of a prominent DNA-dnmaging agent, temozolomide, by manipulating the DNA base excision repair (BER) pathway via BER inhibitor, methoxyamine, and overexpression of N-methylpurine DNA glycosylase (MPG). Experimental Design: Enhancement of temozolomide via methoxyamine and MPG overexpression was analyzed using in vitro assays, including 3-(4-5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl) -2H-tetrazolium salt (MTS) assay, apoptosis via Annexin staining, and Western blotting for H2AX phosphorylation to quantitate DNA damage. Results: Our data show that we can effectively modulate the activity of the chemotherapeutic agent, temozolomide, via modulator methoxyamine, in three ovarian cancer cell lines, SKOV-3x, Ovcar-3, and IGROV-1. This enhancement of temozolomide-induced cytotoxicity is not dependent on p53 status as we transfected an ovarian cancer cell line with a dominant-negative p53-expressing plasmid (IGROV-1mp53) and obtained similar results. Our results show that MPG-overexpressing IGROV-1 and IGROV-1mp53 cells are significantly more sensitive to the clinical chemotherapeutic temozolomide in combination with methoxyamine as assayed by cytotoxicity, apoptosis, and levels of DNA damage than either agent alone. Conclusions: These studies show that although clinical trials in ovarian cancer to determine temozolomide single-agent efficacy are in development, through manipulation of the BER pathway, an increase in response to temozolomide is achieved. The combination of temozolomide plus methoxyamine has potential for second-line therapy for patients who have failed standard platinum plus paclitaxel chemotherapy.",
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