THE 92-KDA GELATINASE-B IS EXPRESSED BY ADVANCED-STAGE MELANOMA-CELLS- SUPPRESSION BY SOMATIC-CELL HYBRIDIZATION WITH EARLY-STAGE MELANOMA-CELLS

The production and local release of various proteolytic enzymes, eithe r by tumor cells or tumor-associated stromal cells, is thought to faci litate the malignant behavior of solid tumors. Human cutaneous melanom a offers an excellent clinical model to study the possible contributio n of such proteases to solid tumor progression because melanoma goes t hrough a series of well defined stages in its pathogenesis; moreover, permanent cell lines have been established from these various stages. As a first step to analyzing the gelatinolytic enzymes in melanoma pat hology, we examined cell lines derived from early stage primary melano mas in which patients were cured of their disease and compared the res ults to those obtained with cell lines established from advanced stage primary lesions or metastases (ie., from patients who eventually succ umbed to the disease), We found that 80% of cell lines examined from e arly stage lesions constitutively produced only the 72-kDa gelatinase A but never the 92-kDa gelatinase B. In contrast, the majority of adva nced stage cell lines examined produced both the 72-kDa gelatinase A a nd the 92-kDa gelatinase B. Advanced stage cell lines that did not con stitutively produce the 92-kDa gelatinase B could be induced to do so with transforming growth factor beta, interleukin 1 beta or 12-O-tetra decanoyl-phorbol-13-acetate. In total, 0 of 5 early stage cell lines c onstitutively expressed the 92-kDa gelatinase B, and only 2 of 5 could be induced to produce this activity. In contrast all advanced stage c ell lines that were evaluated either constitutively or inducibly produ ced the 92-kDa gelatinase B. To analyze the mechanism by which 92-kDa gelatinase B production is switched on in the advanced stage melanoma cell lines, somatic cell hybrids were constructed using an advanced st age melanoma cell line as one partner and either one of two early stag e cell lines as the other. Constitutive production of the 92-kDa gelat inase B in such hybrids was lost and could not be induced in such hybr ids. Coculture of the early and advanced stage cell lines failed to re capitulate what was seen after somatic hybridization, and zymographic analysis of lysates from hybrid cell lines demonstrated no 92-kDa gela tinase B activity. Reverse transcription-PCR analysis demonstrated tha t the loss of 92-kDa gelatinase B production occurred at the level of steady-state mRNA for the enzyme. The results indicate that malignant progression of human cutaneous melanoma is accompanied by the constitu tive or induced ability to produce the 92-kDa gelatinase B, and that t his switch in production is most likely related to the loss of a negat ive regulatory activity present in early stage nonmalignant melanomas.