Hypothetical Mechanism of Therapeutic Synergism Induced by the Combination of 6-Thioguanine and 3-[(4-Amino-2-methyl-5- pyrimidinyl)methyl]-1 -(2-chloroethyl)-1 -nitrosourea Hydrochloride1

Comparative studies were carried out using various antimetabolites in order to determine whether the synergistic cellkilling effect on L1210 murine leukemia cells induced by a treatment combining 6-thioguanine (6-TG) and 3-[(4-amino2-methyl-5-pyrimidinyl)methyl]-1-(2-chloroethyl)-1-nitrosourea hydrochloride (ACNU) resulted from "complementary inhibi tion." The combinations of ACNU plus 5-fluorouracil and ACNU plus 5-bromo-2'-deoxyuridine induced 31 and 18-fold greater cell killing of L1210 cells in vitro over the expected additive survival (the product of the observed survival for each drug), respectively, whereas the cytocidal activity of the combinations of ACNU plus methotrexate and ACNU plus 6-methylmercaptopurine riboside was less than 2-fold greater. Moreover, ther apeutic synergism defined by two parameters (survival time and/or cure incidence) against iv. L1210 leukemia was clearly elicited only by the combinations of ACNU plus 5-fluorouracil, 5-bromo-2'-deoxyuridine, or 6-mercaptopurine. As a result, the following possibility that the 6-TG incorporated into the DNA in place of guanine increases the amount of the DNA interstrand cross-linking by ACNU was considered and exam ined using the DNAs isolated from 6-day-old L1210 cells in the peritoneal cavity untreated or treated with 30, 3, and 0.3 mg of 6-TG per kg for 5 hr. Significant increases in the amount of binding of [efrty/ene-14C]ACNU to the DNA and in the amount of the DNA interstrand cross-iinking determined by the hydroxylapatite technique were observed with the increase in the 6TG dose and, maximally, approximately 6-fold greater binding and 6to 12-fold increases in the double-stranded DNA fraction over those of control DNA were obtained by 4and 2-hr incubations at 37°,respectively.