The research program of the Molecular Cytogenetics Section of the Laboratory of Experimental Carcinogenesis is aimed at the identification and characterization of genomic modifications associated with initiation and progression of the neoplastic phenotype. Using a combined approach based on integrated use of molecular cytogenetics and molecular biology, our group identified and examined a number of recurrent chromosome alterations that led to the discovery of several new cancer-relevant genes, and to the detection of alterations in a number of known ones. Analysis of these alterations may provide markers useful for the prognosis and diagnosis of cancer and potential targets for therapy. In the past year significant progress has been made molecular cytogenetics and molecular biology of cancer cells. New recurrent chromosome alterations and evidence regarding the role of DLC-1 gene in embryonic development and human breast cancer metastasis. were identifid.. The mouse DLC-1 gene was isolated, and the exon-intron organization was characterized. An intragenic polymorphic microsatellite marker was identified that was useful for linkage mapping and LOH analysis.To provide an animal model system for investigating the biological functions of DLC-1 in vivo, we successfully used homologous recombination in embryonic stem cells to generate mice with a disrupted DLC-1 gene. Mice heterozygous for the disrupted allele were viable and phenotypically normal, although with reduced levels of DLC-1 mRNA. No homozygous mutant progeny was obtained from mating of heterozygous animals, indicating that DLC-1 deficiency resulted in embryonic lethality. Analysis of timed pregnancies showed that DLC-1-/- embryos did not survive beyond 10.5 days gestation , and histological examination revealed defects in the neural tube, brain, heart, and placenta. In situ hybridization localization of DLC-1 mRNA in the wild type mouse embryo showed a widespread expression in embryonic and extraembryonic tissues as at the time when abnormalities were found in mutant embryos. This distribution is consistent with a role for DLC-1 in normal development. Cultured fibroblasts from homozygous mutant embryos displayed alterations in the organization of actin filaments and focal adhesions. These results suggest that the DLC-1 protein plays an important role in the assembly of the cytoskeleton and cell adhesion complexes and loss of DLC-1 may interfere with development by adversely affecting cell adhesion and migration Identification of molecular signatures characteristic of the biological mechanisms involved in the metastasis spread of cancer is required for the development of therapeutic interventions able to abrogate the process. Previously , we demonstrated that restoration of DLC-1 expression in cell lines derived from metastasic breast adenocarcinomas lacking endogenous gene expression caused significant growth inhibition and prevented the development of tumors in athymic nude mice.In collaboaration with Dr. Steve Goodison from University of Florida and his colleagues the role of DLC-1 in metastasis. was examined. Monoclonal cell lines M4A4 and NM2C5 are spontaneously occurring sublines of the MDA-MB235 which exhibit many phenotypic differences in growth, invasion, dissemination and spontaneous metastatic efficiency from an orthotopic site. The common origin of these cell lines enables the comparative investigation of cellular and molecular events in the metastatic process in a stable and isogeneic model. A 171-gene expression signature that correlated with metastatic phenotype highlighted several GTPase signaling components. One of these components, DLC 1 gene, was found down-regulated in the metastatic relative to the non-metastatic cells.
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