The objective of the proposed research is to identify and characterize a gene involved in lung cancer development and progression on the centromeric portion of chromosome 11p15.5. The appliant s previous studies have provided evidence for the existence of two regions with frequent allelic loss on chromosome 11p in lung cancer which suggests the presence of tumor suppressor genes. A region in the centromeric part of chromosome band 11p15.5, which was identified by the polymorphic marker D11S12, had loss of heterozygosity (LOH) in 57% of all 156 lung cancer cases analyzed. This region is approximately 0.5 megabases (Mb) in size and is located between the markers D11S1758 and D11S860. The objective of this research proposal is to clone the tumor suppressor gene/s in this region using a positional cloning approach. The applicant proposes 1) to complement the partial Pl phage and cosmid contig of the target region (D11S1758 to D11S860) with P1 phage and cosmid clones using the """"""""DuPont"""""""" and """"""""PAC"""""""" human genomic Pl libraries and the National Laboratory and ICRF chromosome 11 specific cosmid libraries. He will assemble the contig by """"""""chromosome walking"""""""" using Pl phage and cosmid-specific end-labeled probes generated by PCR with SP6, T3, and T7 primers and with Pl phage-specific primers generated by direct sequencing. Contig gaps will be filled with bacterial artificial chromosome (BAC) clones by screening the BAC human genomic library available from GenomeSystems, Inc.; 2) to refine the presently known region of allele loss to less than 500 Kb with new microsatellite markers. New markers will be generated by hybridization of a poly(CA/TG) probe to contig clones and by direct sequencing. Allele frequencies of these new markers will be assessed in an unrelated population survey. 3) to positionally clone genes within this region with a solution hybridization and magnetic bead capture method and by screening human cDNA libraries with contig subclones. For solution hybridization and magnetic bead capture cosmid and Pl clones from the contig will serve as """"""""selector"""""""" templates. """"""""Driver"""""""" templates will be synthesized from normal lung polyA RNA. Selected sequences will be analyzed for an open reading frame, and they will be compared with published sequences in data bases. Genes identified (""""""""candidate"""""""" genes) will then be screened for evolutionary conservation, expression in normal lung, and mutations.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Research Project (R01)
Project #
5R01CA070317-02
Application #
2633925
Study Section
Pathology B Study Section (PTHB)
Program Officer
Lively, Tracy (LUGO)
Project Start
1997-01-22
Project End
1999-12-31
Budget Start
1998-02-25
Budget End
1998-12-31
Support Year
2
Fiscal Year
1998
Total Cost
Indirect Cost
Name
Duke University
Department
Internal Medicine/Medicine
Type
Schools of Medicine
DUNS #
071723621
City
Durham
State
NC
Country
United States
Zip Code
27705
Bepler, Gerold; Zheng, Zhong; Gautam, Ashish et al. (2005) Ribonucleotide reductase M1 gene promoter activity, polymorphisms, population frequencies, and clinical relevance. Lung Cancer 47:183-92
Lipkus, Isaac M; McBride, Colleen M; Pollak, Kathryn I et al. (2004) Interpretation of genetic risk feedback among African American smokers with low socioeconomic status. Health Psychol 23:178-88
McBride, Colleen M; Pollak, Kathryn I; Garst, Jennifer et al. (2003) Distress and motivation for smoking cessation among lung cancer patients' relatives who smoke. J Cancer Educ 18:150-6
Gautam, Ashish; Li, Zhan-Rong; Bepler, Gerold (2003) RRM1-induced metastasis suppression through PTEN-regulated pathways. Oncogene 22:2135-42
Bepler, Gerold; Gautam, Ashish; McIntyre, Lauren M et al. (2002) Prognostic significance of molecular genetic aberrations on chromosome segment 11p15.5 in non-small-cell lung cancer. J Clin Oncol 20:1353-60
McBride, Colleen M; Bepler, Gerold; Lipkus, Isaac M et al. (2002) Incorporating genetic susceptibility feedback into a smoking cessation program for African-American smokers with low income. Cancer Epidemiol Biomarkers Prev 11:521-8
Zhao, B; Bepler, G (2001) Transcript map and complete genomic sequence for the 310 kb region of minimal allele loss on chromosome segment 11p15.5 in non-small-cell lung cancer. Oncogene 20:8154-64
McIntyre, L M; O'Briant, K C; McBride, C M et al. (2001) Rater agreement and utility of the mutagen-induced chromosome damage assay. Anticancer Res 21:605-9
McBride, C M; Pollak, K I; Bepler, G et al. (2001) Reasons for quitting smoking among low-income African American smokers. Health Psychol 20:334-40
McBride, C M; Halabi, S; Bepler, G et al. (2000) Maximizing the motivational impact of feedback of lung cancer susceptibility on smokers' desire to quit. J Health Commun 5:229-41

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