Radiosensitivity Of Lymphocytes Research Articles

Radiation-induced damage to normal tissues after radiotherapy in patients treated for gynecologic tumors: Association with single nucleotide polymorphisms in XRCC1, XRCC3, and OGG1 genes and in vitro chromosomal radiosensitivity in lymphocytes

To examine the association of polymorphisms in XRCC1 (194Arg/Trp, 280Arg/His, 399Arg/Gln, 632Gln/Gln), XRCC3 (5' UTR 4.541A>G, IVS5-14 17.893A>G, 241Thr/Met), and OGG1 (326Ser/Cys) with the development of late radiotherapy (RT) reactions and to assess the correlation between in vitro chromosomal radiosensitivity and clinical radiosensitivity. Sixty-two women with cervical or endometrial cancer treated with RT were included in the study. According to the Common Terminology Criteria for Adverse Events, version 3.0, scale, 22 patients showed late adverse RT reactions. Polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) assays were performed to examine polymorphic sites, the G2 assay was used to measure chromosomal radiosensitivity, and patient groups were compared using actuarial methods. The XRCC3 IVS5-14 polymorphic allele was significantly associated with the risk of developing late RT reactions (odds ratio 3.98, p = 0.025), and the XRCC1 codon 194 variant showed a significant protective effect (p = 0.028). Patients with three or more risk alleles in XRCC1 and XRCC3 had a significantly increased risk of developing normal tissue reactions (odds ratio 10.10, p = 0.001). The mean number of chromatid breaks per cell was significantly greater in patients with normal tissue reactions than in patients with no reactions (1.16 and 1.34, respectively; p = 0.002). Patients with high chromosomal radiosensitivity showed a 9.2-fold greater annual risk of complications than patients with intermediate chromosomal radiosensitivity. Combining the G2 analysis with the risk allele model allowed us to identify 23% of the patients with late normal tissue reactions, without false-positive results. The results of the present study showed that clinical radiosensitivity is associated with an enhanced G2 chromosomal radiosensitivity and is significantly associated with a combination of different polymorphisms in DNA repair genes.

Cytogenetic Radiosensitivity of G0-Lymphocytes of Breast and Esophageal Cancer Patients as Determined by Micronucleus Assay

Enhanced chromosomal radiosensitivity is a feature of many cancer predisposition conditions, indicative of the important role of chromosomal alterations in carcinogenesis. In this study the cytokinesis-blocked micronucleous assay was used to compare the radiosensitivity of blood lymphocytes obtained from Iranian breast or esophageal cancer patients (n = 50, n = 16; respectively) with that of control individuals (n = 40). For each sample, one thousand binucleate lymphocytes were analyzed before and after in vitro exposure to 3 Gy of gamma rays. The radiation-induced frequency of micronucleus was significantly higher in the breast cancer group (261/1,000 binucleated cells) than in esophageal cancer group (241/1,000 binucleated cells, P < 0.01) or in the control group (240/1,000 binucleated cells, P < 0.01). The results indicate that breast cancer patients are more radiosensitive compared to normal healthy individuals or esophageal cancer patients. Increased radiosensitivity could be due to defects in DNA repair genes involved in breast cancer formation. Since patients with esophageal cancer did not show elevated radiosensitivity, it is assumed that the contribution of radiosensitivity-related genes to the development of esophageal cancer may be smaller than the contribution of those genes to breast cancer.

The use of IL-2 cultures to measure chromosomal radiosensitivity in breast cancer patients

Enhanced chromosomal radiosensitivity in breast cancer patients has been demonstrated in several studies. To investigate the chromosomal radiosensitivity of lymphocytes in breast cancer patients the G2 and micronucleus (MN) assays are often used. In these assays blood samples are exposed to ionizing radiation and the number of radiation-induced micronuclei or chromatid breaks are scored. In most studies investigating the in vitro chromosomal radiosensitivity of breast cancer patients the G2 and MN assays were performed on freshly drawn blood. The disadvantage of working with fresh blood samples is that in most cases only one blood sample can be obtained and that the assay cannot be easily repeated without further blood sampling. To allow repeated testing we propose the use of long-term cultures of T lymphocytes (IL-2 cultures). In this study we therefore investigated whether the radiation-induced MN response in IL-2 cultures was the same as in concordant whole blood cultures. For this study the MN assay (2 Gy) was performed on IL-2 cultures of 11 sensitive breast cancer patients and 20 healthy women. The results demonstrate that the enhanced chromosomal radiosensitivity observed in whole blood cultures of breast cancer patients is not present in IL-2 cultures derived from the same blood samples. Therefore, care has to be taken when IL-2 cultures are used to assess chromosomal radiosensitivity in breast cancer patients.

The use of EBV-transformed cell lines of breast cancer patients to measure chromosomal radiosensitivity.

To investigate the chromosomal radiosensitivity of lymphocytes in cancer patients the micronucleus (MN) assay is often used and performed on freshly drawn peripheral blood lymphocytes. The use of Epstein-Barr virus (EBV)-transformed lymphoblastoid cell lines may have a lot of advantages (e.g. large pool of cells) compared with fresh blood samples. In this study we have investigated whether the response of EBV-transformed lymphoblastoid cell lines to irradiation in the G1/S/G2 phases of the cell cycle is the same as in concordant whole blood cultures where primary lymphocytes were irradiated in the G0 phase of the cell cycle. For this study the MN assay (2 Gy) was performed on EBV-transformed cell lines of breast cancer patients and a group of healthy women. Those breast cancer patients were selected who showed an elevated chromosomal radiosensitivity in fresh blood samples in a previous study. The results demonstrated that the enhanced chromosomal radiosensitivity observed in fresh blood cultures of breast cancer patients is not present in EBV-transformed cell lines derived from the same blood samples. Therefore, care must be taken when EBV cell lines are used to assess chromosomal radiosensitivity in breast cancer patients.

Radiosensitivity of Peripheral Blood Lymphocytes Obtained from Patients with Cancers of the Breast, Head and Neck or Cervix as Determined with a Micronucleus Assay

The in vitro radiation sensitivities of peripheral blood lymphocytes obtained from 48 normal females and 168 female cancer patients were measured with the cytokinesis-blocking micronucleus assay. Cancer patients group had significantly higher mean baseline micronucleus frequency than normal healthy controls. Breast cancer patients were more radiosensitive than normal individuals. Cervical cancer cases were less radiation sensitive than normal subjects. The relative lack of radiation sensitivity in cervical-cancer cases could be due to modification of the radiosensitivity of patients' immune-responsible cells by human papillomaviruses infection. Normal individuals and cancer patients were classified according to their radiation sensitivity which was evaluated with the radiation-induced micronucleus frequencies. Such a classification will be an important initial step to characterize the radiosensitive, radioresistant, or cancer-prone individuals using specific SNP typing.

Lymphocyte radiosensitivity correlated with pelvic radiotherapy morbidity

Purpose To test the hypothesis that, before treatment, prostate cancer patients who demonstrate a high yield of ex vivo radiation-induced micronucleus (MN) in G 0 lymphocytes represent a patient population with a greater-than-average risk of developing radiotherapy (RT)-related morbidity. Methods and materials We prospectively conducted the cytokinesis-block MN assay of peripheral blood lymphocytes (PBLs) in 38 prostate cancer patients. Before the initiation of RT, PBLs from each patient were irradiated (1–4 Gy). The mean patient age ± SEM was 68.7 ± 1.0 years. The clinical stage was T1 in 17, T2 in 15, and T3 in 6. The preoperative prostate-specific antigen level was ≤4 ng/mL in 5, 4–10 ng/mL in 18, and >10 ng/mL in 15. All patients underwent standardized pelvic external beam radiotherapy (range 41.4–50.4 Gy) and boost (range 16–26 Gy). The mean follow-up ± SEM was 32.8 ± 4.6 months. At the end of follow-up, a radiation oncologist scored the GI or GU morbidity according to the Radiation Therapy Oncology Group criteria without knowledge of the MN data. Results We found that between the average reactors ( n = 25; i.e., patients who had Grade 1 or less RT-related morbidity) and over reactors ( n = 13; i.e., patients who developed Grade 2 or greater RT-related morbidity), the differences in the ex vivo radiation dose–response relationship of MN yield in PBLs were highly significant, especially at doses of ≥2 Gy. Also, the development of RT-related morbidity correlated with the radiation dose–response relationship of MN yield in PBLs before treatment, but did not correlate with any of the patients’ clinical variables. Conclusion Our findings suggest that the pre-RT ex vivo radiation dose–response relationship of MN yield in PBLs may be a significant predictive factor for the development of GI or GU morbidity in prostate cancer patients after pelvic RT.

Lymphocyte radiosensitivity is correlated with pelvic radiotherapy morbidity

Lymphocyte radiosensitivity is correlated with pelvic radiotherapy morbidity

Trends in biological dosimetry: an overview

Biological dosimetry (biodosimetry) is based on investigations of induced biological effects (biomarkers) in order to correlate them with radiation dose. Among the indicators employed in biodosimetry, scoring of chromosome aberrations is the most reliable method to quantify individual exposure to ionizing radiation. The technique, applied to circulating lymphocytes, has been developed into a routine procedure to evaluate the dose in the case of real or suspected accidental exposure. Considering the radiosensitivity of lymphocytes in vitro and in vivo as being the same, the dose effect relationship obtained after in vitro irradiation of blood has been widely used, with medico-legal value, for evaluating individual radiation exposure. This report presents an overview of strengths, limitations and perspectives on biodosimetry.

Chromosomal radiosensitivity in G2-phase lymphocytes identifies breast cancer patients with distinctive tumour characteristics.

A substantial proportion of women with breast cancer exhibit an abnormally high radiosensitivity as measured by the frequency of chromatid breaks induced in G2-phase, PHA stimulated lymphocytes. Chromatid break frequencies were compared for a cohort of previously untreated sporadic breast cancer patients and hospital outpatient controls. In the breast cancer group 46% showed high radiosensitivity compared to 14% of controls (P< 0.001). Comparison of those breast cancer patients with a high G2radiosensitivity (G2RS) versus those with a low G2RS showed no difference in menopausal status or age but the high G2RS group had on average a lower score on the Nottingham Prognostic Index. Predicted survival in the high G2RS group at 15 years was 55% compared to 36% for the low G2RS group. Furthermore, 81% of tumours from the high G2RS were oestrogen receptor positive compared to 45% from the low G2RS group. Thus high G2RS identifies a sub-population of patients with distinctive tumour characteristics and with a predicted improved prognosis as compared with those in the low G2RS group. Our findings imply that besides influencing risk of breast cancer the genetic factors determining G2radiosensitivity also influence the tumour characteristics and prognosis in these patients. © 2001 Cancer Research Campaign http://www.bjcancer.com

Lymphocyte radiosensitivity is a significant prognostic factor for morbidity in carcinoma of the cervix

Purpose: To study the relationship between pretreatment peripheral blood lymphocyte radiosensitivity and morbidity following radiation therapy. Methods and Materials: A prospective study was carried out in which patients with carcinoma of the cervix underwent radiation therapy. Intrinsic radiosensitivity was measured on pretreatment peripheral blood lymphocytes, using a limiting dilution clonogenic assay. Late morbidity was assessed using the Franco-Italian glossary. Results were correlated in an actuarial analysis. Results: There were no correlations between the measured lymphocyte radiosensitivity (SF2) and colony-forming efficiency, patient age, tumor grade, or disease stage. For 83 patients, lymphocyte SF2 was a significant prognostic factor for the probability of developing both any ( p = 0.002) and Grade 3 ( p = 0.026) morbidity. In 174 patients, stage showed borderline significance as a prognostic factor for morbidity ( p = 0.056). However, the type of treatment (intracavitary alone, intracavitary plus parametrial irradiation, single insertion plus whole-pelvis irradiation) was significantly associated with the probability of developing late complications ( p = 0.013). There was a weak significant inverse correlation between lymphocyte SF2 and grade of morbidity ( r = −0.34, p = 0.002). Conclusion: These data highlight the importance of normal cell radiosensitivity as a factor determining radiation therapy response. They also show that peripheral blood lymphocyte SF2 is a highly significant prognostic factor for the probability of developing late radiation morbidity, and that carcinoma of the cervix is a good model for testing radiobiologic principles in the clinic.

Pretreatment plasma TGFβ1 levels are prognostic for survival but not morbidity following radiation therapy of carcinoma of the cervix

Purpose: To determine whether pretreatment plasma-transforming growth factor beta 1 (TGFβ1) levels are prognostic for tumor control and late morbidity following radiation therapy in carcinoma of the cervix. Methods and Materials: The study was comprised of 79 patients undergoing radiotherapy with curative intent for Stage I-III carcinoma of the cervix. TGFβ1 levels were analyzed using ELISA. Late morbidity was measured using the Franco-Italian glossary. Data were available for the pretreatment levels of circulating tumor markers that represent disease burden, and for peripheral blood lymphocyte radiosensitivity measured as SF2. Results: Pretreatment TGFβ1 levels were a significant prognostic factor for survival and local control. There were weak significant correlations of TGFβ1 levels with disease stage and the levels of circulating tumor markers (CA125, TPA). There was a weak significant correlation between TGFβ1 levels and normal cell radiosensitivity (lymphocyte SF2). There was no relationship between TGFβ1 levels and grade of morbidity and pretreatment TGFβ1 levels were not a significant prognostic factor for the probability of developing late morbidity. Conclusion: In carcinoma of the cervix, pretreatment TGFβ1 levels reflect tumor burden and are a significant prognostic factor for survival. Despite an underlying weak relationship of TGFβ1 levels with intrinsic normal cell radiosensitivity, pretreatment levels are not prognostic for the probability of developing late complications. This finding does not rule out the possible usefulness of measurements toward the end of treatment once tumor burden has been reduced.

Gender difference in smoking effect on chromosome sensitivity to gamma radiation in a healthy population.

In the general population, there is variation in radiosensitivity associated with cancer risk. However, data on the role of epigenetic factors in the variation of radiosensitivity are scarce. Thus we investigated the effects of smoking and age on the radiosensitivity of human lymphocytes by measuring the frequency of chromosome aberrations after in vitro exposure to gamma rays in peripheral lymphocytes from 441 healthy subjects (18-95 years old). We analyzed the frequency of both spontaneous (baseline) and in vitro gamma-ray-induced (1.5 Gy) chromatid breaks in 50 well-spread metaphases per subject. The overall mean frequencies of spontaneous and induced breaks were 0.02 and 0.45 per cell, respectively. The mean frequency of induced breaks was significantly higher in men than in women (P = 0.03) but did not differ by age or ethnicity. Donors who had ever smoked showed a small but significantly increased frequency of induced breaks (mean = 0.47) compared to nonsmokers (mean = 0.41; P = 0.005). Further stratification and multivariate analyses revealed that the smoking effect was more pronounced in men than in women. These findings support a smoking effect on radiosensitivity in a healthy population, particularly in men. Therefore, when evaluating the association between radiosensitivity and susceptibility to smoking-related cancers, the effect of smoking should be taken into account.

DNA Ligase IV Deficiency in Mice Leads to Defective Neurogenesis and Embryonic Lethality via the p53 Pathway

DNA ligase IV (LIG4) is a nonhomologous end-joining (NHEJ) protein used for V(D)J recombination and DNA repair. In mice, Lig4 deficiency causes embryonic lethality, massive neuronal apoptosis, arrested lymphogenesis, and various cellular defects. Herein, we assess potential roles in this phenotype for INK4a/ARF and p53, two proteins implicated in apoptosis and senescence. INK4a/ARF deficiency rescued proliferation/senescence defects of Lig4-deficient fibroblasts but not other phenotypic aspects. In contrast, p53 deficiency rescued embryonic lethality, neuronal apoptosis, and fibroblast proliferation/senescence defects but not lymphocyte development or radiosensitivity. Young Lig4/p53 double null mice routinely died from pro-B lymphomas. Thus, in the context of Lig4 deficiency, embryonic lethality and neuronal apoptosis likely result from a p53-dependent response to unrepaired DNA damage, and neuronal apoptosis and lymphocyte developmental defects can be mechanistically dissociated.

Relationship between in vitro chromosomal radiosensitivity of peripheral blood lymphocytes and the expression of normal tissue damage following radiotherapy for breast cancer

Background and purpose: There is a need for rapid and reliable tests for the prediction of normal tissue responses to radiotherapy, as this could lead to individualization of patient radiotherapy schedules and thus improvements in the therapeutic ratio. Because the use of cultured fibroblasts is too slow to be practicable in a clinical setting, we evaluated the predictive role of assays of lymphocyte chromosomal radiosensitivity in patients having radiotherapy for breast cancer.Materials and methods: Radiosensitivity was assessed using a micronucleus (MN) assay at high dose rate (HDR) and low dose rate (LDR) on lymphocytes irradiated in the G0 phase of the cell cycle (Scott D, Barber JB, Levine EL, Burril W, Roberts SA. Radiation-induced micronucleus induction in lymphocytes identifies a frequency of radiosensitive cases among breast cancer patients: a test for predispostion? Br. J. Cancer 1998;77;614-620) and an assay of G2 phase chromatid radiosensitivity (‘G2 assay’) (Scott D, Spreadborough A, Levine E, Roberts SA. Genetic predisposition in breast cancer. Lancet 1994; 344: 1444). In a study of acute reactions, blood samples were taken from breast cancer patients before the start of radiotherapy, and the skin reaction documented. 116 patients were tested with the HDR MN assay, 73 with the LDR MN assay and 123 with the G2 assay. In a study of late reactions, samples were taken from a series of breast cancer patients 8–14 years after radiotherapy and the patients assessed for the severity of late effects according to the'LENT SOMA’ scales. 47 were tested with the HDR assay, 26 with the LDR assay and 19 with the G2 assay. For each clinical endpoint, patients were classified as being normal reactors or ‘highly radiosensitive patients’ (HR patients (Burnet NG. Johansen J, Turesson I, Nyman J. Describing patients' normal tissue reactions: Concerning the possiblity of individualising radiotherapy dose presciptions based on potential predictive assays of normal tissue radiosensitivity. Int. J. Cancer 1998;79:606-613)).Results: The HR patients could be identified in some of the assays. For example, for acute skin reactions, 9/123 patients were judged as HR; they had significantly higher G2 scores than normal reactors (P=0.004). For the late reactions, the mean HDR MN scores were higher for the 4/47 patients who had severe telangiectasia (P=0.042) and the 8/47 patients had severe fibrosis (P=0.055). However, there were no trends towards increased chromosomal radiosensitivity with the micronucleus scores at HDR or LDR, or with G2 chromosomal radiosensitivity.Conclusions: While these results support the concept of using lymphocytes to detect elevated sensitivity to radiotherapy (as an alternative to fibroblasts), these assays are unlikely to be of assistance for the prediction of normal tissue effects in the clinic in their present form.

Comparison of the radiosensitivity of normal-tissue cells with normal-tissue reactions after radiotherapy

Purpose : To investigate whether the in vitro radiosensitivity of normal lymphocytes and fibroblasts evaluated by the micronucleus (MN) assay predicts acute and late reactions after radiotherapy in cancer patients. Materials and methods : Studies were performed on blood samples from 31 cervical and head and neck cancer patients and on skin fibroblasts from eight of the cancer patients. The radiosensitivity of lymphocytes and of fibroblasts was also assessed in 24 and five healthy donors, respectively. Radiosensitivity was measured after in vitro irradiation with doses ranging from 2 to 5 Gy using micronucleus frequency (the number of micronuclei per single binucleated (BN) cell) and the percentage of BN cells with micronuclei. The in vitro results were compared with the maximum grade of acute and late reactions. Results : There was no significant difference in cellular radiosensitivity between cancer patients and healthy donors. Although cancer patients differed considerably in normal-cell radiosensitivity, no correlation was found between radiosensitivity, either of lymphocytes or fibroblasts, and acute and late clinically observed side effects. In addition, no relationship was observed between the radiosensitivity of lymphocytes and fibroblasts derived from the same donors. Conclusion : The data do not support the usefulness of the MN assay in predicting normal-tissue response to radiotherapy in cancer patients.

Chromosome radiosensitivity in human G2 lymphocytes and cell-cycle progression

Purpose : To investigate the possibility that the differential G2- phase radiosensitivity of human peripheral blood lymphocytes, found in normal individuals using the 'G2-phase chromosome radiosensitivity assay', could be attributed to heterogeneity in cellular progression to mitosis rather than di fferences in radiosensitivity. Materials and methods : Human peripheral blood lymphocytes, from four different donors, were exposed to 50cGy X-rays and sampled at different times. The progression of cells into mitosis was monitored by 5-bromo 2-deoxyuridine (BrdUrd) incorporation. Results : The heterogeneous G2-phase chromosome radiosensitivity among different donors was abolished when homogeneous G2-phase cell populations were scored; they contained similar frequencies of cells in early or late G2-phase. Conclusions : The heterogeneous G2-phase chromosome radiosensitivity, usually found in different normal donors, is caused by the analysis of different cell populations rather than reflecting intrinsic differences in radiosensitivity.

Chromosomal in-vitro radiosensitivity of lymphocytes in radiotherapy patients and AT-homozygotes.

We investigated the in-vitro radiosensitivity of peripheral blood lymphocytes with a special FISH/CISS-technique. From October 1993 through April 1996, a total number of 52 cancer patients was enrolled in the study. The tumor sites in these patients were: breast (n = 41), lung (n = 4), head and neck (n = 3) as well as prostate, bladder, rectal cancer and Hodgkin's disease (each n = 1). Twenty-six of them were examined prior to planned radiotherapy (prospective group) and 26 after radiotherapy (retrospective group). Three additional individuals (without cancer or radiotherapy) with proven ataxia telangiectasia (Louis-Bar syndrome, AT-homozygotes) were also investigated and their blood samples served as positive control for radiosensitivity. The clinical radiation response of normal tissue in radiotherapy patients was scored according to the WHO grading system for acute and according to the RTOG grading system for late effects. For to estimate the intrinsic radiosensitivity, blood samples were taken and irradiated in vitro with 0 (control) or 0.7 or 2 Gy with a 6 MV-linear accelerator, standard 48-hour lymphocyte cultures were prepared, chromosomes #1, #2 and #4 were simultaneously labeled with a FISH/CISS-technique and 200 to 1,000 metaphase spreads were scored for chromosomal aberrations. The radiation sensitivity of lymphocytes was expressed as the number of radiation-induced chromosomal breaks per mitosis after 0.7 Gy or 2 Gy corrected for the 0-Gy control value. The frequency of chromosomal breaks/mitosis in the unirradiated control lymphocytes was 0.020 +/- 0.015 in prospective patients who had not yet received radiotherapy. It was significantly higher in retrospective patients (0.264 +/- 0.164 breaks/mitosis) as a result of the previous radiation exposure. The 3 AT-homozygotes showed also an increased number of spontaneous chromosomal breaks (0.084 +/- 0.016 breaks/mitosis), probably resulting from the chromosomal instability in this disease. This figure, however, was significantly lower than in retrospective patients. The number of radiation-induced breaks after in-vitro irradiation was comparable in lymphocytes of patients who showed no normal tissue reaction (n = 11) as compared to those with mild to moderate radiation reaction (n = 32, acute reactions Grade 1 to 2, late reactions Grade 0 to 2). In 9 patients with unexpected severe plus late Grade 3 to 4 reactions, however, a significantly higher number of radiation-induced chromosomal breaks was measured; the highest number was observed in a patient with a radiation myelitis. The 3 AT-homozygotes showed, as expected, an extreme radiosensitivity of their lymphocytes. The number of breaks/mitosis after 0.7 Gy in vitro irradiation of lymphocytes was 0.103 +/- 0.059 in patients with no normal tissue radiation reaction (n = 11), 0.122 +/- 0.146 in the group with mild to moderate radiation reactions Grade 1 to 2 (n = 32), 0.359 + 0.226 in patients with unexpected Grade 3 to 4 normal tissue reactions (n = 9) and 0.550 +/- 0.243 in the 3 AT-homozygotes (p < 0.01, t-test). The difference in lymphocyte radiosensitivity between these 4 groups was also detected after in-vitro irradiation with 2 Gy (0.484 +/- 0.132 vs. 0.535 +/- 0.228 vs. 0.926 +/- 0.349 vs. 1.423 +/- 0.072). We found a significantly higher number of chromosomal breaks in lymphocytes of patients with severe or extreme radiation reaction of normal tissues as compared to patients with no or only mild to moderate radiation reactions. The radiosensitivity of lymphocytes in these radiosensitive patients was in the range between normal radiosensitivity and the radiosensitivity of AT-homozygotes. Detection of patients with severely enhanced intrinsic radiosensitivity might be possible with this method.

Radiation-induced micronucleus induction in lymphocytes identifies a high frequency of radiosensitive cases among breast cancer patients: a test for predisposition?

Enhanced sensitivity to the chromosome-damaging effects of ionizing radiation is a feature of many cancer-predisposing conditions. We previously showed that 42% of an unselected series of breast cancer patients and 9% of healthy control subjects showed elevated chromosomal radiosensitivity of lymphocytes irradiated in the G2 phase of the cell cycle. We suggested that, in addition to the highly penetrant genes BRCA1 and BRCA2, which confer a very high risk of breast cancer and are carried by about 5% of all breast cancer patients, there are also low-penetrance predisposing genes carried by a much higher proportion of breast cancer patients, a view supported by recent epidemiological studies. Ideally, testing for the presence of these putative genes should involve the use of simpler methods than the G2 assay, which requires metaphase analysis of chromosome damage. Here we report on the use of a simple, rapid micronucleus assay in G0 lymphocytes exposed to high dose rate (HDR) or low dose rate gamma-irradiation, with delayed mitogenic stimulation. Good assay reproducibility was obtained, particularly with the HDR protocol, which identified 31% (12 out of 39) of breast cancer patients compared with 5% (2 out of 42) of healthy controls as having elevated radiation sensitivity. In the long term, such cytogenetic assays may have the potential for selecting women for intensive screening for breast cancer.

The intrinsic radiosensitivity of normal and tumour cells

Purpose: To examine whether in vitro measurements of normal and tumour cell radiosensitivity can be used as prognostic factors in clinical oncology. Materials and methods: Stage I-III cervix carcinoma patients were treated with radical radiotherapy with a minimum of 3 years' follow-up. Lymphocyte and tumour radiosensitivities were assayed using, respectively, a limiting dilution and soft agar clonogenic assay to obtain surviving fraction at 2 Gy (SF2). The results were related, in an actuarial analysis, to late morbidity assessed using the Franco-Italian glossary. Results: Patients with radiosensitive lymphocytes had a significantly increased risk of developing late complications (n 93, p 0.002). Increasing tumour radiosensitivity was associated with an increased risk of morbidity (n 113, p 0.032). A significant correlation was found between fibroblast and tumour cell radiosensitivity (r 0.57, p 0.03), but a weak inverse association was found between lymphocyte and tumour cell radiosensitivity (r 0.32, p 0.03). Patients with radiosensitive lymphocytes and tumour cells had higher levels of late complications than those whose cells were radioresistant. Conclusion: The work described highlights the importance of cellular radiosensitivity as a parameter determining the clinical response to radiotherapy.

Apoptosis induced by fast neutrons versus 60Co gamma -rays in human peripheral blood lymphocytes

Purpose: To investigate the effectiveness of high LET fast neutrons compared with low LET 60Co gamma -rays to induce apoptosis in resting lymphocytes. Materials and methods: Apoptosis induction was quantified by light microscopic analysis of irradiated lymphocyte samples from healthy donors after 24 h culture. For the dose-response analyses doses ranging from 0.05 to 5 Gy were applied at 1.5 Gy/min (gamma -rays) or 0.2 Gy/min (fast neutrons). To investigate the role of DNA repair in apoptosis induction a dose of 2 Gy was also given at low dose-rate (0.006 Gy/min). Results: Dose-response curves obtained with both radiation qualities were characterized by an initial steep increase in the number of apoptotic cells below 1 Gy, with a flattening of the curves at higher doses towards 5 Gy. The calculated relative biological effectiveness (RBE) values for fast neutrons were close to unity. When a 2 Gy dose was administered at low rather than high dose-rate no decrease in apoptotic cell yield was observed. Conclusion: The dose-response data confirm the high radiosensitivity of lymphocytes and demonstrate that their response to undergo early interphase cell death by apoptosis is largely independent of LET. The observations that apoptosis induction is independent of LET and dose-rate may suggest that initial DNA damage, as opposed to DNA repair, dominate the induction of apoptosis in resting lymphocytes.