Induction of the metastatic phenotype in a mouse tumor model by 5-azacytidine, and characterization of an antigen associated with metastatic activity

Abstract

The murine Lewis lung carcinoma is a long-term grafted tumor that, after subcutaneous inoculation, forms metastases to the lungs. Forty-two cell lines were established from a primary tumor site and 40 were established from lung metastatic foci. Cloned sublines were established from the original 82 lines, and 2 sublines among 405 were found to be tumorigenic but not metastatic (T+/M-), whereas the remaining 403 sublines were both tumorigenic and metastatic (T+/M+). The T+/M- phenotype was shown to be stable for >2 yr. However, treatment of the T+/M- cell lines for 3 days with 3 μM 5-azacytidine resulted in reexpression of the metastatic phenotype in otherwise stable T+/M- lines. Also, 5-aza-cytidine treatment could result in loss of the metastatic phenotype in lines that had been stable T+/M+. The changes in tumorigenic and metastatic phenotypes were not associated with altered immunogenicity of the cells. Monoclonal antibodies were generated against T+/M+ cells, and one antibody (M36D3) was found to bind only to T+/M+ cells. Reactivity of the antibody was found to co-vary with expression of the metastatic phenotype. The antigen recognized by M36D3 antibody thus seems to be associated with metastatic capability. The antigen was found by two-dimensional gel electrophoretic analysis to be a cellular protein of M(r) ~ 45,000 and pI ~ 6.7.