Dosage and cycle effects of dacarbazine (DTIC) and fotemustine on O6-alkylguanine-DNA alkyltransferase in human peripheral blood mononuclear cells

Abstract

Summary There is increasing experimental evidence to suggest that endogenous expression of o6-alkylguanine-DNA-alkyltransferase (ATase) is a major factor in cellular resistance to certain chemotherapeutic agents including dacarbazine (DTIC). We al., 1989;Comis, 1976). It undergoes metabolic N-demethylation to give the cytotoxic monomethyl triazene, 5 -(3 -methyl -1 -triazeno) imidazole -4-carboxamide (MTIC) which methylates DNA, producing among 12 other lesions, 06-methylguanine (Meer et al., 1986). There is increasing evidence to suggest that 06-methylguanine is the principal cytotoxic event following DTIC and that 06-alkylguanine-DNA alkyltransferase (ATase) gene expression may be a major factor in cellular resistance to such agents. ATase is able to transfer the methyl group from the 06 position of guanine to an internal cysteine residue in an auto-inactivating stoichiometric reaction. Experimental models using ATasedeficient cell lines or xenografts show them to be more sensitive to DTIC than lines or xenografts with high activity (Catapano et al., 1987;D'Incalci et al., 1988;Foster et al., 1990;Gibson et al., 1986a;Hayward and Parsons, 1984;Lunn & Harris, 1988). The strongest evidence for the cytotoxic effects of 06-alkylguanine in DNA comes from ATase cDNA transfection experiments which show that expression of prokaryotic or eukaryotic ATase cDNA in mammalian cells protects them against the toxic effects to these agents (Brennand and Margison, 1986;Jelinek et al., 1988;Kataoka et al., 1986;Samson et al., 1986;Kaina et al., 1991 therapeutic implications especially when DTIC is combined with the subsequent administration of a chloroethylating nitrosourea. In this case, drug resistance appears to involve the same ATase DNA repair enzymes which remove the chloroethyl lesions from the 06-position of guanine, thereby preventing the formation of a cytotoxic DNA interstrand cross-link (see Lee et al., 1991a). Theoretically, enhanced therapeutic effects might be obtained when the nitrosourea is administered at the nadir of ATase activity following DTIC treatment assuming that the effect in peripheral mononuclear cells reflects that of tumour tissues. Materials and methodsPatients and blood samples Blood samples were collected from 30 treatment cycles of 25 pts with metastatic melanoma treated with sequential DTIC and fotemustine chemotherapy. Approval was obtained from the local ethical committee and all pts gave informed consent for the study. Pts received DTIC at fixed doses (for each pt) of 400, 500 or 800 mg m-2 by i.v. infusion followed by fotemustine lOOmg m2, 4h later. Treatment was repeated every 28 days and the number of cycles given depended on the individual pts response. Blood samples were collected just before chemotherapy and at 1, 2, 3, 4, 6 and 18 h after DTIC infusion; in addition, 5 h samples were collected for the 400 mg m-2 patients. Blood was drawn into a 20 ml univeral container containing 0.5% EDTA and stored at 4°C before isolation of PMCs. Fourteen sets of samples from were also collecte...

O6-(4-bromothenyl)guanine reverses temozolomide resistance in human breast tumour MCF-7 cells and xenografts

“…While the O 6 -alkylating agents are currently not as widely used in the management of breast cancer as they are in glioma, malignant melanoma and lymphoma, the possibility of overcoming alkylating agent resistance would make these agents useful additions to the chemotherapeutic management of this much more common malignancy. Attempts to improve the effectiveness of O 6 -alkylating agents therapy by prior inactivation of MGMT using methylating agents have foundered owing to the similar, and hence cumulative, toxicities of the inactivating and treatment drugs (Lee et al, 1993;Smith et al, 1996;Clemons et al, 2002;Hammond et al, 2004).…”

“…Clemons et al, 2003). These have been hampered by the similar toxicities of the two agents, and no useful increase in therapeutic index has been demonstrated (Micetich et al, 1992;Lee et al, 1993;Smith et al, 1996;Hammond et al, 2004). Because of this, interest has turned to inactivation of MGMT using inherently nontoxic pseudosubstrates for the protein, such as O 6 -benzylguanine (O 6 -BeG; reviewed in Pegg et al, 1995;Dolan and Pegg, 1997) and O 6 -(4-bromothenyl)guanine (PaTrin-2, Lomeguatrib;McElhinney et al, 1998;Middleton et al, 2002;Middleton and Margison, 2003).…”

Formation and loss of O6-methyldeoxyguanosine in human leucocyte DNA following sequential DTIC and fotemustine chemotherapy

“…ATase activity was depleted and the rate and extent of ATase depletion was patient, dosage and cycle dependent, with the nadir of ATase activity occurring 4-5 h after DTIC administration (Lee et al, 1991(Lee et al, , 1993a. These findings have been attributed to autoinactivation of ATase during the repair of 06-MedG formed in lymphocyte DNA.…”

“…Results 06-Alkylguanine-DNA alkyltransferase activity The data regarding the changes in ATase activity during combination chemotherapy with DTIC and fotemustine have been reported previously as mean values of a group of patients (Lee et al, 1993a). Here, we present the pretreatment and nadir (i.e.…”

“…These findings have been attributed to autoinactivation of ATase during the repair of 06-MedG formed in lymphocyte DNA. It was also shown that fotemustine administration was not associated with ATase depletion in peripheral lymphocytes (Lee et al, 1993a).…”