Variation in susceptibility to environmental cancer may, in part, be due to inherent differences between individuals to repair damaged site induced within their genetic material by radiation and chemical carcinogens. In xeroderma pigmentosum, a prototype genetic disorder, defective cellular repair of UV- induced DNA damage is etiologically linked with the marked predisposition of patients to develop UV-associated skin cancers. Epidemiologic studies of DNA repair and cancer susceptibility may provide important insights into the etiology of various forms of environmental cancer within the general population, and offer possible modes of prevention. The extent to which such studies may be initiated, however, is limited by the lack of validated laboratory procedures for measuring cellular repair proficiency within populations of interest. The objective of this proposal is to develop a highly sensitive and specific laboratory procedure for screening cellular repair proficiency on a population basis. With the future intent of field- validating, then applying this methodology within epidemiologic studies of human cancer, the following specific aims are proposed: - To develop a quantitative host cell reactivation (HCR) assay of DNA repair in peripheral blood lymphocytes utilizing UV-damaged recombinant DNA plasmids. - To investigate the use of transformed xeroderma pigmentosum and presumed normal lymphocyte cell lines as internal standards of repair proficiency. - To determine the feasibility of storing human lymphocytes over time by cryopreservation for subsequent simultaneous assay of repair proficiency. - To determine the laboratory precision associated with the established plasmid HCR repair assay protocol. The proposed HCR assay will employ a recombinant DNA plasmid containing a bacterial gene which is transiently expressed in cultured human lymphocytes, and whose respective gene product assay is amenable to screening. Gene expression will be inactivated by inducing a quantified amount of UV damage within the coding region of the gene. Repair proficiency will be expressed in terms of the percent gene expression reactivated in cell cultures receiving UV-irradiated plasmid, relative to parallel control cultures receiving non-damaged plasmid DNA. Lymphocyte cell lines at varying levels of repair proficiency will be used to standardize inter-individual variation in absolute repair.
