Molecular studies on the therapeutic implications and regulation of connexin 43.

摘要

Gap junctional intercellular communication (GJIC) allows for the transport of small signaling molecules between adjacent cells. Gap junctions and their constituents, connexins, are often impaired in cancer. Restoration of connexins and GJIC in cancer cells leads to reversal of the transformed phenotype, reduction of the rate of cell growth, and inhibition of in vivo tumorigenicity. In addition, restoration of connexin43 (Cx43), one of the most abundant connexins that is often reduced in cancer, allows for the transfer of chemotherapeutic agents among neighboring cells, a phenomenon known as the "Bystander Effect". In chapter 2, I'm reporting evidence that induction of Cx43 and GJIC with cyclic-AMP (cAMP) can enhance the cytotoxic effect of the prodrug ganciclovir following infection of breast cancer cells with an adenovirus expressing the ganciclovir-activating enzyme viral thymidine kinase (vTK). This induction may provide a therapeutic advantage in suicide gene therapy due to the bystander effect when a small proportion of the cell population is expressing vTK. In chapter 3, I'm reporting that breast tumor tissue samples from patients had low to undetectable Cx43 levels, in contrast to their matched normal tissues. In addition, Cx43 protein and RNA levels were undetectable in a panel of human breast cancer lines and in rat breast tumors induced by the carcinogen dimethylbenz[a]anthracene. Together, these observations indicate that the loss of Cx43 is a common marker of cancer cells. In chapter 4, I examined the hypothesis that Cx43 downregulation occurs primarily at the transcriptional level. To determine the mechanisms behind the transcriptional regulation of Cx43, the human Cx43 promoter was investigated. Overexpression of the H-Ras oncogene in NIH3T3 cells leads to the induction of the human Cx43 promoter activity, as well as Cx43 RNA and protein levels. This stimulation involves the MEK-ERK pathway downstream of H-Ras. The promoter sequence responsible for this effect is localized downstream of the transcription start site, and is not homologous to any known cis-elements. It is recognized by one main nuclear protein complex, which binds to the Cx43 promoter at a greater extent in H-Ras-overexpressing than wild-type cells, and contains c-Myc and HSP90. This complex may serve as a therapeutic target for the stimulation of Cx43 in cancer.