A SMALL-MOLECULE E2F INHIBITOR BLOCKS GROWTH IN A MELANOMA CULTURE MODEL.

This study by researchers at the H. Lee Moffitt Cancer Center and Research Institute demonstrated how MatTek’s Melanoma in vitro human skin tissue equivalent can be used to investigate both the efficacy of potential melanoma therapeutic agents and their potential toxicity on non-melanoma skin cells. HLM006474 was identified using a computer-based virtual screen and the known crystal structure of the DNA-bound E2F4/DP2 heterodimer. Treatment of multiple cell lines with HLM006474 resulted in the loss of intracellular E2F4 DNA binding activity as measured by electrophoretic mobility shift assay within hours. Overnight exposure to HLM006474 resulted in down-regulation of total E2F4 protein as well as known E2F targets. The effects of HLM006474 treatment on different cell lines varied, but included a reduction in cell proliferation and an increase in apoptosis. HLM006474 induced apoptosis in a manner distinct from cisplatin and doxorubicin. E2F4-null mouse embryonic fibroblasts were less sensitive than wild-type counterparts to the apoptosis-inducing activity of the compound, revealing its biological specificity. A375 cells were extremely sensitive to the apoptosis-inducing activity of the compound in two-dimensional culture. HLM006474 was a potent inhibitor of melanocytes proliferation and subsequent invasion in a three-dimensional tissue model system. Over time, the metastatic melanocytes in the untreated three dimensional tissue model proliferate and form nodes, which grow and invade the underlying collagen substrate. HLM006474 significantly inhibited the proliferation and subsequent invasion of the melanocytes into the collagen layer. These results suggest that HLM006474 is a highly effective inhibitor of malignant growth in this model system. The compound had no obvious deleterious effects on the other cells (fibroblasts and keratinocytes) making up the three-dimensional tissue. Together, these results suggest that interference with E2F activity using small molecules may have clinical application in cancer therapy.

Apoptosis, Cancer therapy, Cell proliferation, E2F4 protein, H&E staining, HLM006474, Ki-67, MLNM-FT-A375, Melanoma, S100

HLM006474