Cytotoxic Mechanism of 6-Thioguanine:  hMutSα, the Human Mismatch Binding Heterodimer, Binds to DNA Containing S⁶-Methylthioguanine

Abstract

It has been suggested that the cytotoxicity of 6-thioguanine depends upon (1) incorporation of 6-thioguanine into DNA, (2) methylation by S-adenosylmethionine (SAM) of the thio group to give S6-methylthioguanine, (3) miscoding during DNA replication to give [SMeG] x T base pairs, and (4) recognition of these base pairs by proteins of the postreplicative mismatch repair system. Here we have investigated systematically the ability of proteins present in human cell extracts to bind to DNA containing S6-methylthioguanine. We found that [SMeG] x T base mismatches were bound by the mismatch binding complex, hMutS alpha, and that the level of binding was dependent upon the base 5' to the S6-methylthioguanine in the order G > C = A > T. Extracts from cells that lack either hMSH2 (LoVo cells) or GTBP (HCT-15 cells), two components of the hMutS alpha complex, were unable to bind the [SMeG] x T base pair. We also found that hMutS alpha was able to bind to [SMeG] x C base pairs when the S6-methylthioguanine was in the sequence 5'-Cp[SMeG]. This suggests that miscoding by S6-methylthioguanine residues in DNA during DNA synthesis may not be an absolutely required step in the mechanism of cytotoxicity. Also, since CpG sequences are so important in gene regulation, this result may be of considerable significance.

Alkyltransferase-like protein (eATL) prevents mismatch repair-mediated toxicity induced by O 6 -alkylguanine adducts in Escherichia coli

“…O 6 -alkylguanine adducts are induced by a variety of SN1-alkylating agents, including several drugs used for cancer chemotherapy (34). Another drug used in chemotherapy, 6-thioguanine, also produces G adducts that signal apoptosis via MMR (9,35). Consequently, many tumors deficient in MMR become resistant to these drugs, presumably because they no longer signal apoptosis following therapy-induced DNA damage.…”

“…Indeed, cytotoxicity involves the mismatch repair (MMR) system that recognizes the mutagenic O 6 -methylguanine: T (mG:T) replication intermediates. There is strong evidence from in vitro experiments that MutS proteins recognize mG:T mispairs more efficiently than mG:C pairs (5)(6)(7)(8)(9). In vivo data also show that mG:T mispairs are processed by MMR more efficiently than mG: C pairs (10).…”

Dose reduction of coadministered 6-mercaptopurine decreases myelotoxicity following high-dose methotrexate in childhood leukemia

“…[4][5][6][7][8] In hemopoietic cells, 6MP is anabolized to 6-thioguanine nucleotides (6TGNs), which are the most important mediators of the cytotoxic effect of 6MP through incorporation into DNA. 9 Erythrocytic accumulation of 6TGN correlate with the degree of myelotoxicity and remission duration in childhood ALL. 3,10,11 Another important metabolic pathway of 6MP is the methylation of 6MP and some of its metabolites by thiopurine methyltransferase (TPMT).…”

6-Mercaptopurine dosage and pharmacokinetics influence the degree of bone marrow toxicity following high-dose methotrexate in children with acute lymphoblastic leukemia

“…1 are the primary mediators of the cytotoxic effect of 6MP through their incorporation into DNA. 3 Another important metabolic pathway is the methylation by the enzyme thiopurine methyltransferase (TPMT, E.C. 2.1.1.67) of 6MP and some of its metabolites such as 6-thioinosine 5Ј-monophosphate.…”