Formation of active products of benzaldehyde dimethane sulfonate (NSC 281612, DMS612) in human blood and plasma and their activity against renal cell carcinoma lines

Abstract

Benzaldehyde dimethane sulfonate (BEN, DMS612, NSC281612) is an alkylating agent with activity against renal cell carcinoma and is being evaluated clinically. To support clinical trials, we developed an LC–MS/MS assay to detect and quantitate BEN and its metabolites/decomposition products. We tested the stability and products of BEN and benzoic acid dimethane sulfonate (BA) in plasma, blood and five renal carcinoma cell lines in vitro. Further, we determined the IC50 of BEN, BA and four of their products in these cell lines. Low temperature and pH stabilized the analytes, and utilizing this resulted in an accurate, precise and reproducible assay. The half-lives of BEN and BA added to plasma in vitro were 220 and 5 min, while the half-life of BEN in whole blood was 18 min. The generation and degradation of up to 12 analytes were monitored, and structures confirmed with available authentic standards. The IC50 for BEN was 5- to 500-fold lower than that of any of its products, while the cellular metabolic activity toward BEN correlated with ALDH activity and IC50 values. We detected six of the in vitro products and their respective glucuronides in murine plasma after dosing BEN. The information gained from these experiments will be instrumental in the evaluation of the pharmacology of BEN in ongoing human trials.

Characterization of the metabolism of benzaldehyde dimethane sulfonate (NSC 281612, DMS612)

“…We studied the metabolism of BEN in five different RCC lines and found that the ALDH activity correlated with the amount of BA-OH 2 generated [12]. This further supported for the idea that the conversion of BEN to BA is enzymatic.…”

“…BEN has structural similarities to both busulfan and melphalan. As with related alkylators, we have previously shown that BEN undergoes hydrolysis in blood and plasma [12]. In whole blood, but not in plasma, BEN is rapidly converted to its carboxylic acid analogue BA, suggesting conversion by an enzyme present in red blood cells.…”

“…For the blood metabolism experiments we used a previously developed assay to quantitate the analytes in plasma [13,12]. For the purified enzyme experiments we developed an assay to quantitate the analytes in PBS (Assay 2).…”

“…Later, we found that in whole blood, but not in plasma, BEN was rapidly converted to its carboxylic acid analogue (BA) [12]. Furthermore, BEN decomposed into at least eleven different products that were structural analogs of either BEN or BA.…”

“…Furthermore, BEN decomposed into at least eleven different products that were structural analogs of either BEN or BA. This led us to hypothesize that the conversion of BEN to BA in whole blood is an enzymatic process [12]. …”