Dependence of Aldehyde Dehydrogenase-mediated Oxazaphosphorine Resistance on Soluble Thiols: Importance of Thiol Interactions with the Secondary Metabolite Acrolein

• Published in: Biochemical pharmacology 1998-07, Vol.56 (1), p.31-39
• Main Authors: Bunting, Kevin D, Townsend, Alan J
• Format: Article
• Language: English
• Subjects: 4-hydroperoxycyclophosphamide; acrolein; Acrolein - metabolism; Acrolein - pharmacology; aldehyde dehydrogenase; Aldehyde Dehydrogenase - antagonists & inhibitors; Aldehyde Dehydrogenase - metabolism; Animals; Antineoplastic agents; Antineoplastic Agents, Alkylating - pharmacology; Biological and medical sciences; Cell Line; Chemotherapy; Cricetinae; Cricetulus; drug resistance; glutathione; Glutathione - metabolism; Humans; Isoenzymes - antagonists & inhibitors; Isoenzymes - metabolism; mafosfamide; Medical sciences; Pharmacology. Drug treatments; Sulfhydryl Compounds - metabolism
• ISSN: 0006-2952; 1873-2968
• DOI: 10.1016/S0006-2952(98)00111-7

Acrolein is a highly reactive and cytotoxic by-product released during activation of oxazaphosphorine (OAP) anticancer alkylating agents. Previously, we demonstrated that transfected human aldehyde dehydrogenase (ALDH, EC 1.2.1.3) isozymes (class 1 or 3) protect V79/SD1 cells from mafosfamide (MAF) cytotoxicity, but protection from 4-hydroperoxy-cyclophosphamide (4-hpCPA) was weaker. Acrolein, an ALDH inhibitor, may be detoxified by conjugation with the nucleophilic thiol 2-mercaptoethanesulfonate (MESNA), which is released from MAF but not from 4-hpCPA. We examined the effect of acrolein or acrolein/thiol conjugates on ALDH activity in vitro. We found that both ALDH isozymes were inhibited by acrolein, with ic 50 values of 35 and 144 μM for ALDH-1 or ALDH-3, respectively. Both isozymes were partially protected by NAD + cofactor, being at least five-fold more sensitive to acrolein if added before cofactor. In contrast, thiol conjugates of acrolein did not inhibit ALDH-3 activity, but were substrates only for ALDH-1. Further, acrolein was shown to be oxidized by ALDH-3, but not by ALDH-1. The effect of acrolein on ALDH-mediated resistance to OAP agents in intact cells was also examined. In control cells (without ALDH expression), acrolein and 4-hpCPA rapidly depleted intracellular GSH levels, whereas the effect of MAF was much less. Depletion of GSH by preincubation of V79/SD1 cells with a low concentration of acrolein (2 μM) before MAF exposure caused a two-fold reduction in ALDH-mediated resistance. Conversely, protection from 4-hpCPA cytotoxicity was enhanced by the addition of thiols (GSH, 2-mercaptoethanesulfonate, or N-acetylcysteine) during the drug exposure. These results suggest 1) that thiol content is an important determinant of the OAP resistance conferred by ALDH isoenzymes; and 2) a new mechanism whereby thiol modulation could increase the therapeutic index of OAP chemotherapy.