Abstract 5454: Drug effect analysis of methoxyamine through quantifying the drug on its therapeutic target

Abstract

Abstract Methoxyamine (MX) is the first DNA base-excision-repair (BER) inhibitor evaluated in humans. MX potentiates the cytotoxicity of alkylating and antimetabolic agents by condensing with DNA apurinic/pymidinic (AP) sites which are produced through the removal of alkylated or abnormal nucleobases from DNA backbone by DNA glycosylases. The formation of AP-MX complex blocks the BER pathway, and hence reverses tumor resistance to the chemotherapeutic agents. The further repair of MX-bound DNA by topoisomerase II results in ds-DNA breaks, and induces tumor cell death. In this work, we developed and validated a liquid chromatography-tandem mass spectrometry (LC-MS/MS) method for the quantitative analysis of AP-MX complex released from MX-bound DNA by a multi-enzyme system. The enzyme-released AP-MX (i.e., deoxyribose-MX, or dR-MX) was first separated from the enzyme digestion matrix on a Hypercarb® column, and then quantitated by a triple quadrupole mass analyzer using multiple-reaction-monitoring (MRM) mode with a mass transition of m/z 164 > 117. The calibration curve which corresponds to 3.55 to 227 AP-MX complexes/107 bases was established using AP-MX calibrators obtained by serial dilution of DNA oligomer containing known amount of AP-MX complex in blank calf thymus DNA. The method developed has been applied to AP-MX analysis of DNA extracts from T98G cells treated with temozolomide with MX, and blood samples from an ongoing Phase I study of methoxyamine and temozolomide in patient with advanced solid tumors. The dose, AP-MX complex, and time profiles have been obtained. This method provides a quantitative measure of methoxyamine at its proposed target sites. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 5454. doi:10.1158/1538-7445.AM2011-5454