Chromosomal instabilities in human tumor and irradiated normal cells

Abstract

Radiation-induced genomic instability has been proposed as an initiating step in radiation-induced carcinogenesis. Numerous studies have established the occurrence of this phenomenon in various cells of both human and rodent origin. Genomic instability is considered to have been induced if new mutations or chromosomal aberrations occur in the progeny of cells surviving the radiation dose at a higher frequency than that for unirradiated cells. Such induced instability has been reported both in vivo and in vitro and for such genetic changes as delayed chromatid-type and chromosome-type aberrations, delayed mutations and delayed ceil lethality. In many of these studies, however, the cells were not "normal" initially and in many cases involved tumor-derived cell lines. If radiation-induced genomic instability is indeed the initiating step involved in radiation-induced carcinogenesis, the phenomenon is clearly of much greater interest if it occurs especially in cells that are apparently normal at the outset. The induction of an "initiating" or early step in carcinogenesis would be less interesting in tumor cells that are already initiated by definition. To this end, I studied a phenotypically normal human fibroblast cell line, (AG1521A) to determine whether they exhibit chromosomal instability in the progeny of surviving ceils after exposure to low and high LET radiation. Radiation exposures were administered while cells were in a non-cycling G0 state and assays for instability were performed on both mixed populations of cells and clones of cells surviving the exposure. Several cytogenetic techniques were used to determine induced chromosomal instabilities. These included solid Giemsa staining and classical cytogenetic analysis, whole chromosome painting by fluorescence in situ hybridization (FISH) and multiplex fluorescence in situ hybridization (M-FISH). I found no evidence for the induction of chromosomal instability in the phenotypically normal AG1521A human fibroblast cells after exposure to either high or low LET radiation following exposure of the cells in a G0 state. Based on review of published literature and our own data, I conclude that radiation-induced chromosomal instability does not occur universally, at least in Go irradiated normal cells. I therefore suggest that induced instability may not be the initiating step in radiation carcinogenesis in all cases. This does not rule out the occurrence of the phenomenon, nor does it suggest a possible lack of importance for it in systems with some predisposing factor, such as alterations in normal DNA damage processing. Nijmegen Breakage Syndrome results from a recessively inherited genetic disorder and among other things is characterized by hypersensitivity to ionizing radiation and chromosomal instability. NBS1 (nibrin or p95), the product of the NBS1 gene, complexes with Rad50 and Mre11 in response to DNA damage, especially double-strand breaks and is required for cell cycle control. The defect in DNA repair and the resulting hypersensitivity to ionizing radiation in cells from NBS patients led us to hypothesize that exposure to ionizing radiation may lead to elevated levels of complex exchange aberrations at doses considerably lower than would produce a similar effect in repair competent cells. Elevated frequencies of spontaneous and radiation-induced chromatid-type and chromosome-type aberrations were detected in the NBS fibroblast cell line GM7166A compared to repair competent AG1521A fibroblasts. Furthermore, the proportion of radiation-induced aberrations that were complex was higher in the NBS fibroblasts than previously reported for normal human lymphoblasts as determined by mFISH. These results suggest that the major defect in this NBS cell line concerns the ability to correctly repair DNA damage and leads to elevated levels of both simple and complex exchange aberrations after exposure to ionizing radiation.