The goal of this work is to develop a detailed understanding of the structure and function of the c-myc gene product. The working hypothesis is that the c-myc protein is involved in the control of transcription and functions as part of a complex of proteins which bind to DNA. The intention is to use a combination of monoclonal antibody and recombinant DNA techniques to analyze the structure and function of this oncogene protein. A panel of epitope specific antibodies will be produced and characterized. These antibodies will be used to define epitopes on the surface of the protein, and regions of the molecule involved in interactions with DNA and with other proteins. Coprecipitation will be used in an attempt to detect proteins associated with c-myc. The antibodies will also be used to determine if there is structural homology between the c-myc protein and other proteins particularly those with known functions or others that bind DNA. In parallel with this work an assay for the biological activity of the c-myc will be developed based on either the transformation of primary lymphocytes or fibroblasts. This assay will then be used to evaluate the effects of site-directed mutagenesis on specific c- myc epitopes. The epitope specific antibodies will be assayed for their effects on the growth of cells into which they are incorporated by microinjection or electroporation. A long term goal is to understand the detailed structure and function of the c- myc protein. This may possibly lead to the ability to specifically modulate the activity of this protein that apparently plays an important role in the control of cell proliferation.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Research Project (R01)
Project #
5R01CA024263-08
Application #
3166387
Study Section
Molecular Cytology Study Section (CTY)
Project Start
1978-09-01
Project End
1989-11-30
Budget Start
1987-12-01
Budget End
1988-11-30
Support Year
8
Fiscal Year
1988
Total Cost
Indirect Cost
Name
University of Pennsylvania
Department
Type
Schools of Medicine
DUNS #
042250712
City
Philadelphia
State
PA
Country
United States
Zip Code
19104
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Ikegaki, N; Kennett, R H (1990) Molecular genetic characterization of epitope-specific monoclonal antibodies against the myc family proteins. Oncogene 5:397-403
Ikegaki, N; Minna, J; Kennett, R H (1989) The human L-myc gene is expressed as two forms of protein in small cell lung carcinoma cell lines: detection by monoclonal antibodies specific to two myc homology box sequences. EMBO J 8:1793-9
Ikegaki, N; Kennett, R H (1989) Glutaraldehyde fixation of the primary antibody-antigen complex on nitrocellulose paper increases the overall sensitivity of immunoblot assay. J Immunol Methods 124:205-10
Ikegaki, N; Polakova, K; Prince, L et al. (1988) Expression and localization of the NMYC protein in human neuroblastoma cells: analysis of the effect of gamma interferon treatment and distribution of the NMYC protein in the nucleus. Prog Clin Biol Res 271:133-44
Bukovsky, J; Kennett, R H (1987) Simple and rapid purification of monoclonal antibodies from cell culture supernatants and ascites fluids by hydroxylapatite chromatography on analytical and preparative scales. Hybridoma 6:219-28
Ikegaki, N; Bukovsky, J; Kennett, R H (1986) Identification and characterization of the NMYC gene product in human neuroblastoma cells by monoclonal antibodies with defined specificities. Proc Natl Acad Sci U S A 83:5929-33
Bukovsky, J; Evans, A; Tartaglione, M et al. (1985) Selection of variant neuroblastoma cell line which has lost cell surface expression of antigen detected by monoclonal antibody PI153/3. Somat Cell Mol Genet 11:517-22
Evans, A E; Griffin, G C; Tartaglione, M et al. (1985) A method of detecting neuroblastoma in human bone marrow by means of two monoclonal antibodies PI 153/3 and KE2. Hybridoma 4:289-96
Kennett, R H; Leunk, R; Meyer, B et al. (1985) Detection of E. coli colonies expressing the v-sis oncogene product with monoclonal antibodies made against synthetic peptides. J Immunol Methods 85:169-82