MELPHALAN UPTAKE, HYPERTHERMIC SYNERGISM AND DRUG-RESISTANCE IN A HUMAN CELL-CULTURE MODEL FOR THE ISOLATED LIMB PERFUSION OF MELANOMA

Isolated limb perfusion with melphalan is a long-standing treatment fo r melanoma but the clinical conditions have not been subjected to a sy stematic evaluation. In order to establish optimal conditions for perf usion, three human melanoma cell lines were cultured with melphalan in vitro under conditions comparable to in vivo therapy. The most import ant findings were that: (a) 41.5-degrees-C was synergistic for melphal an killing of three human melanoma cell lines; (b) prolonging the trea tment time beyond 1 h had little additional toxicity; and (c) varying the initial pH of the culture medium had no effect. After 1 h of treat ment, cells accumulated more melphalan at 41.5-degrees-C than at 37-de grees-C, relative to the extracellular concentration. A cell line (MM4 18) derived from a primary tumour was the most resistant of the three lines; pigmented or non-pigmented sublines were equally resistant. The A2058 line showed the lowest level of synergism with hyperthermia, an d displayed a marked plateau at 10% of controls in the dose-response f or survival, yet no melphalan-resistant subpopulation could be isolate d. The implications of this work are that (a) enhanced cellular uptake of melphalan may account for hyperthermic synergism of melphalan; (b) varying conditions other than treatment time will be necessary to dea l with the variation in resistance between tumours; and (c) repeated c ycles of treatment may be needed for phenotypes such as A2058 where me lphalan resistance appears to be based on an epigenetic mechanism.