Role of O⁶-alkylguanine-DNA alkyltransferase in protecting against 1,3-bis(2-chloroethyl)-1-nitrosourea (BCNU)-induced long-term toxicities

O⁶-alkylguanine-DNA alkyltransferase (AGT) protects from the mutagenic and toxic lesions induced by 1,3-bis(2-chloroethyl)-1-nitrosourea (BCNU), and in many tumors, AGT overexpression provides a means of resistance. To circumvent this, O⁶-benzylguanine, an inactivator of AGT, has been developed and is currently in clinical development with BCNU; however, the potential long-term toxicities associated with this treatment are unknown. With the inactivation of AGT by O⁶-benzylguanine, a higher number of toxic and mutagenic O⁶-alkylguanine lesions introduced by methylating or chloroethylating agents would be expected. In this study, cohorts of mice were treated with vehicle, O⁶-benzylguanine (30 mg/kg), BCNU alone (low dose of 15 mg/kg or high dose of 50 mg/kg), or O⁶-benzylguanine (30 mg/kg) plus BCNU (15 mg/kg) and followed for 12 months post-treatment. Mice treated with O⁶-benzylguanine plus BCNU or high-dose BCNU died significantly earlier (p < 0.0001) than mice in the other three cohorts with a median survival of 8.3 (O⁶-benzylguanine plus BCNU) and 7.9 months (high-dose BCNU). Histopathologic sections of tissues revealed that the most common morphological diagnosis in animals treated with O⁶- benzylguanine plus BCNU (15 mg/kg) or BCNU (50 mg/kg) was cytomegaly in the lung with greater severity observed in mice receiving the combination O ⁶-benzylguanine plus BCNU. Four of five mice analyzed in this cohort had alveolar histiocytosis, with one also having alveolar edema. In contrast, liver and kidney toxicity was only observed in mice treated with BCNU (50 mg/kg). These results suggest that O⁶-benzylguanine enhances long-term pulmonary toxicity associated with BCNU in mice.