Differential inactivation of polymorphic variants of human O6-alkylguanine-DNA alkyltransferase.

The human DNA repair protein O(6)-alkylguanine-DNA alkyltransferase (hAGT) is an important source of resistance to some therapeutic alkylating agents and attempts to circumvent this resistance by the use of hAGT inhibitors have reached clinical trials. Several human polymorphisms in the MGMT gene that encodes hAGT have been described including L84F and the linked double alteration I143V/K178R. We have investigated the inactivation of these variants and the much rarer variant W65C by O(6)-benzylguanine, which is currently in clinical trials, and a number of other second generation hAGT inhibitors that contain folate derivatives (O(4)-benzylfolic acid, the 3' and 5' folate esters of O(6)-benzyl-2'-deoxyguanosine and the folic acid gamma ester of O(6)-(p-hydroxymethyl)benzylguanine). The I143V/K178R variant was resistant to all of these compounds. The resistance was due solely to the I143V change. These results suggest that the frequency of the I143V/K178R variant among patients in the clinical trials with hAGT inhibitors and the correlation with response should be considered.

Inactivation of O(6)-alkylguanine-DNA alkyltransferase by folate esters of O(6)-benzyl-2'-deoxyguanosine and of O(6)-[4-(hydroxymethyl)benzyl]guanine.

O6-Alkylguanine-DNA alkyltransferase (alkyltransferase) provides an important source of resistance to some cancer chemotherapeutic alkylating agents. Folate ester derivatives of O6-benzyl-2'-deoxyguanosine and of O6-[4-(hydroxymethyl)benzyl]guanine were synthesized and tested for their ability to inactivate human alkyltransferase. Inactivation of alkyltransferase by the gamma-folate ester of O6-[4-(hydroxymethyl)benzyl]guanine was similar to that of the parent base. The gamma-folate esters of O6-benzyl-2'-deoxyguanosine were more potent alkyltransferase inactivators than the parent nucleoside. The 3'-ester was considerably more potent than the 5'-ester and was more than an order of magnitude more active than O6-benzylguanine, which is currently in clinical trials to enhance therapy with alkylating agents. They were also able to sensitize human tumor cells to killing by 1,3-bis(2-chloroethyl)-1-nitrosourea, with O6-benzyl-3'-O-(gamma-folyl)-2'-deoxyguanosine being most active. These compounds provide a new class of highly water-soluble alkyltransferase inactivators and form the basis to construct more tumor-specific and potent compounds targeting this DNA repair protein.