Natural resistance to MTX in three different types of malignant cells is discussed. Two causes of resistance found were impaired drug uptake and impaired polyglutamylation. These resistance mechanisms have also been described to occur in cell lines with acquired resistance to this drug. In cells with acquired drug resistance, these phenotypic changes may reflect mutations in genes associated with transport of MTX or the enzyme DHFR, or in some circumstances reversion of the cell to a different phenotype without a mutational event. In cells with intrinsic resistance, the genetic mechanism responsible for these phenotypes is not known. Alternate treatment programs, e.g., with TMTX, or with TMTX and CPG2, a folate depleting enzyme, are under evaluation for these MTX resistant neoplasms.