Deletion of chromosome 3 has been implicated in a number of human cancers including small cell lung cancer (SCLC). Chromosomal deletion in solid tumors is one of the hallmarks of a tumor suppressor gene. Microcell transfer experiments from our laboratory support the contention that there is a tumor suppressor gene on the short arm of chromosome 3. The goals of this research are to clone the putative tumor suppressor gene in the region chromosome 3p21 and to determine its role in tumorigenesis and its function in normal cells. A 2Mb fragment from the p2l-p22 region of chromosome 3 which suppresses the growth of mouse A9 cells will be the starting material for cloning this gene. This human DNA fragment contains sequences which are homozygously deleted in SCLC. Both genomic and cDNA strategies will be pursued in the cloning of the gene. For the genomic approach, YAC, P1 and cosmid libraries will be screened with markers which are known to be in this fragment and are also deleted in SCLC. cDNAs corresponding to this region will be isolated by hybrid selection, exon trapping, and direct screening methods. Tumor cell lines either from revertants of the somatic cell hybrid or from SCLC will be analyzed for altered gene structure or expression for each candidate locus. Mutation analysis of tumor samples will indicate the frequency of aberration of candidate genes in SCLC. To prove the tumor suppressor gene- lies on a clone, genomic or cDNA clones will be transfected into mouse A9 cells. The stable transfectants will be analyzed for tumor growth. Once the tumor suppressor is identified, this gene will be examined in tumors which have shown deletions of chromosome 3 including breast and ovarian cancer. Characterization of this gene and gene product will include sequencing the cDNA and production of antibodies to define the expression pattern of this protein. These experiments will define a tumor suppressor gene on chromosome 3 and initiate new research on determining the role of this gene in many types of human cancer.
