Combination antifol therapy: observations on the development of resistant L1210 leukemic cells in vivo

L1210 murine leukemic cells were serially passaged in BDF1 mice and were treated in vivo with methotrexate (MTX) and 2,4-diamino-5-(3',4'-dichlorophenyl)-6-methylpyrimidine (DDMP) alone or in combination. The time course of emergence of resistance of the treated cell lines was studied both in vivo and in vitro. When used as a single drug, in vivo resistance to MTX developed gradually and was considerable at eight passages and complete by 11 passages. Complete in vivo resistance to DDMP used alone occurred by the fifth passage. Complete in vivo resistance to the drugs in combination was seen by the eighth passage. In cells demonstrating complete DDMP resistance, as determined in vivo, there was no evidence of cross-resistance to MTX measured either in vivo or in vitro, while MTX resistance was associated with incomplete cross-resistance to DDMP. The greatest degree of resistance, as determined in vitro, occurred in the cell line treated with the drug combination. In vitro tests of drug resistance correlated well with in vivo survival data. An important observation was that major in vivo drug resistance was accompanied by only a small measurable effect using standard in vitro screening techniques. The implications of this finding are discussed.