Individual genetic variation in cellular DNA damage response pathways can influence DNA damage signaling thresholds, rates of DNA repair, and in vitro
radiosensitivity after exposure to low-dose and low-dose-rate ionizing radiation (1
). Such genetic variants presumably play an important role in determining an individual's predisposition to spontaneous and DNA-damaging agent-induced cancers (2
), and sensitivity to the genotoxic effects of mutagens has been shown to be highly heritable in studies examining monozygotic and dizygotic twins and first-degree relatives (6
). Significantly reduced DNA damage signaling and repair capacity has been documented in vitro
for several cancer predisposition and chromosomal instability syndromes, including ataxia telangiectasia (AT), Nijmegen breakage syndrome (NBS), and LIG4 syndrome (11
), while a broad spectrum of radiation responses has also been observed for cells derived from apparently normal individuals (1
The induction of chromosomal aberrations after exposure to radiation or other genotoxic agents (both exogenous and endogenous) is mediated through the misrepair or lack of repair of DNA double-strand breaks (DSBs), which subsequently determines (to a great extent) the proliferative and carcinogenic potential of surviving cells (18
). Several cytogenetic assays have been developed to measure radiation-induced chromosomal aberrations during different phases of the cell cycle for use as potential radiation and cancer biomarkers (22
). One assay, the G2
chromosomal radiosensitivity assay, has been developed to measure the response of cells (typically lymphocytes or fibroblasts) to irradiation in G2
). This G2
assay measures the yield of chromatid-type aberrations (mostly chromatid gaps and breaks) at the first postirradiation mitosis in cells irradiated hours previously during the G2
phase. The repair of radiation-induced DSBs in G2
-phase mammalian cells occurs primarily through non-homologous end joining (NHEJ) and homologous recombinational repair (HRR) (20
). The relative contribution of NHEJ and HRR to the repair of radiation-induced DSBs in G2
-phase human cells has not been fully established, although it is apparent that both repair pathways contribute to the repair of double-stranded DNA damage induced by radiation (19
assay has been applied to a number of radiation sensitivity and cancer predisposition syndromes, including AT, Bloom syndrome, dysplastic nevus syndrome, familial polyposis, Fanconi anemia, Gardner syndrome, Li-Fraumeni syndrome, Wilms tumor and xeroderma pigmentosum, as well as to patients with prostate, head and neck, breast and other types of cancer (27
). The high frequencies of chromatid-type aberrations observed in cells derived from these patients have been attributed to deficiencies in DNA repair or related signaling pathways (27
). However, the G2
assay has demonstrated only moderate correlation to the degree of in vivo
radiosensitivity observed in the clinic (40
hypersensitivity has been studied as a potential marker of heritable low-penetrance predisposition to cancer. Roberts et al.
-phase hypersensitivity in 23 of 37 first-degree relatives of radiosensitive breast cancer patients compared to only one of 15 first-degree relatives of breast cancer patients with normal radiosensitivity (9
). A recent large-scale examination of G2
chromosomal radiosensitivity and postirradiation apoptotic responses of peripheral blood lymphocytes from 211 untreated breast cancer patients and 170 matched controls did not reveal any significant differences between the two groups for either end point, although it was suggested that both cases and controls with high familial risk of breast cancer were more radiosensitive (40
Measurements of G2
-phase chromosomal radiosensitivity in retinoblastoma (RB) patients and first-degree family members were reported by Chaum et al.
) and in two reports by Sanford et al.
) using the radiomimetic agent bleomycin and radiation, respectively. The spontaneous and bleomycin-induced aberration frequencies in bilateral (hereditary-type) and unilateral (sporadic) RB lymphocytes reported in the study of Chaum et al.
) did not differ significantly from those of normal control lymphocytes. Similarly, a retinoblastoma tumor cell line examined by Darroudi et al.
) did not demonstrate an increased aberration induction after X irradiation in G2
phase compared to normal untransformed fibroblasts. On the other hand, the first study of Sanford et al.
reported a mean frequency of 3.2 chromatid breaks/cell (range 1.0–5.1) for eight bilateral (including one familial unilateral) RB fibroblast strains and 4.1 breaks/cell for a sporadic unilateral RB fibroblast strain compared to 0.4 breaks/cell (range 0.2–2.0) for 29 normal fibroblast strains after an X-ray dose of 53 cGy (26
). In the second study of Sanford et al.
, the authors reported mean frequencies of 2.2 breaks/cell (range 1.3–3.2) for 13 additional bilateral RB fibroblast strains, 0.7 breaks/cell (range 0.4–2.1) for six additional unilateral RB fibroblast strains, and 0.4 breaks/cell (range 0.3–0.5) for eight additional normal fibroblast strains after 53 cGy (46
). Some of the unaffected first-degree relatives displayed high aberration frequencies (≥1.1 breaks per cell) similar to those of the bilateral RB patients. In a later study by Scott et al.
, evaluation of the same collection of fibroblast strains demonstrated higher aberration frequencies in the normal strains (average of ~2.2 breaks/cell, range 1.1–3.3), different kinetics of repair, and more interexperimental variability, with 12 of the 53 normal strains (23%) being designated as moderately radiosensitive (33
We previously described an enhanced sensitivity for cell killing and γ-H2AX focus induction after both high-dose-rate and continuous low-dose-rate γ irradiation in primary fibroblast strains derived from 14 hereditary-type (bilateral) RB family members for both the affected RB1+/−
probands and the unaffected RB1+/+
). In the present study, we present data on the G2
-phase chromosomal radiosensitivity of fibroblasts derived from these RB family members and five Coriell controls (four normal strains and one RB1+/−
strain) after acute doses of 50 cGy and 1 Gy high-dose-rate 137
Cs γ radiation. The relative radiosensitivity of the Coriell control strains examined in this study was established by previous survival and cytogenetic assays performed in this laboratory2