Therapeutic resistance: characterization and inactivation by specific antiserum of a putative protein family produced by tumour cells

A multitherapy resistance (MTR) factor produced by Cloudman S91 mouse melan oma cells rescues a responsive cell line after gamma-irradiation, short wav elength ultraviolet light, mitomycin C, vinblastine and actinomycin D. A si milar activity with respect to ionizing radiation is now shown to be produc ed by human melanoma cells and by both human and mouse breast cancer cells but not by five normal cell lines. In these studies, the factor produced in serum-free conditioned medium (SFCM) by Cloudman S91/13 cells is further c haracterized. Its activity in a clonogenic assay using related Cloudman S91 /amel cells is destroyed by trypsin but nor by DNase and is stable for at l east 8 days at a variety of temperatures including 37 degrees C. Molecules greater than 30 kDa from SFCM collected from S91/13 cells were concentrated and separated by preparative zonal electrophoresis (PZE). Bioactivity was present in both the cathode- and the anode-running fractions. The active ac idic (anode) fractions were analysed by preparative isoelectric focusing. B ioactivity was present between pi 3.5 and 4.2. These PZE fractions were als o used to immunize two rabbits, both of which produced antiserum that abrog ated the bioactivity of SFCM and of the PZE cathode fractions. Antiserum al so decreased the survival of irradiated S91/13 producer cells that do not r espond to SFCM but nonetheless must require MTR proteins for the expression of radiation resistance. These studies present a model for the production of rescue factors by non-clonogenic tumour cells that may persist in some t umours for considerable periods of time. (C) 1999 Lippincott Williams & Wil kins.