In most self-renewing tissues, one control point in the regulation of cell growth is availability of hormones, and/or growth factors produced by other tissues. One method by which cancer cells may achieve autonomous growth is through production of autostimulatory growth factors. We have found that culture medium conditioned by the established human malignant melanoma cell line Hs0294 contains an activity which stimulates 3H-thymidine incorporation into DNA and increases cell number over time in the same cells which produce it. The activity of this melanoma growth stimulatory activity (MGSA) is acid stable, nondialyzable (greater than 3500 d.), stable to boiling, trypsin sensitive, sensitive to dithiotreitol reduction and there is no proteolytic activity associated with MGSA. The activity elutes from a Bio-gel P-30 column over a broad molecular weight range, from 6,000-22,000 d. Analysis of Bio-gel MGSA revealed the active components can be further resolved into two major subsets. Acetic acid extracts of the melanoma-conditioned medium also contain an activity which stimulates nonmalignant NRK cells to form colonies in soft agar. This "transforming activity" elutes from the Bio-gel P-30 column differently than MGSA but will coelute in the RP-HPLC with the 35% acetonitrile MGSA unless the TGFs are first removed from the Bio-gel MGSA preparation. The Hs0294 cells do not respond to the "transforming activity," suggesting that the serum-independent proliferation of Hs0294 cells is due to MGSA. Monoclonal antibodies have been raised to a heterogenous preparation of MGSA and antibodies produced by five of the hybridoma clones will inhibit the growth of Hs0294 cells. The specificity of these antibodies is being evaluated. Our studies will now be directed toward three goals: (1)\to further characterize and purify MGSA; (2)\to determine the cellular specificity of the response to MGSA; and (3)\to determine the effect of the MGSA and antibodies to MGSA on the growth of human melanoma tumors.