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

Baeyens, Ans; Thierens, Hubert; Vandenbulcke, Katia; Ridder, Leo De; Vral, Anne

doi:10.1093/mutage/geh029

Cited by 24 publications

(13 citation statements)

References 24 publications

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“…b Multivariate linear regression included terms for age (years), BMI (kg m À2 ), current smoker (yes/no), alcohol intake (total drinks of per day), physical activity (total hours per week), and total daily caloric intake (kcal). radiation when compared to peripheral blood lymphocytes (Baeyens et al, 2004). Strengths of our study include the evaluation of multiple endpoints (before and after exposure to g-irradiation), a relatively large sample of carriers, the use of controls who were from a family with a previously identified mutation, and employing a single technician who was blinded to the mutation status of the samples.…”

Section: Discussionmentioning

confidence: 99%

DNA repair capacity as a possible biomarker of breast cancer risk in female BRCA1 mutation carriers

et al. 2007

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The BRCA1 gene product helps to maintain genomic integrity through its participation in the cellular response to DNA damage: specifically, the repair of double-stranded DNA breaks. An impaired cellular response to DNA damage is a plausible mechanism whereby BRCA1 mutation carriers are at increased risk of breast cancer. Hence, an individual's capacity to repair DNA may serve as a useful biomarker of breast cancer risk. The overall aim of the current study was to identify a biomarker of DNA repair capacity that could distinguish between BRCA1 mutation carriers and non-carriers. DNA repair capacity was assessed using three validated assays: the single-cell alkaline gel electrophoresis (comet) assay, the micronucleus test, and the enumeration of g-H2AX nuclear foci. DNA repair capacity of peripheral blood lymphocytes from 25 cancer-free female heterozygous BRCA1 mutation carriers and 25 noncarrier controls was assessed at baseline and following cell exposure to g -irradiation (2 Gy). We found no significant differences in the mean tail moment, in the number of micronuclei or in the number of g-H2AX nuclear foci between the carriers and non-carriers at baseline, and following g-irradiation. These data suggest that these assays are not likely to be useful in the identification of women at a high risk for breast cancer. British Journal of Cancer (2007) The inheritance of a deleterious mutation in the breast cancer susceptibility gene, BRCA1, has been associated with a lifetime risk of breast cancer of between 45 and 87% (Ford et al, 1998;Antoniou et al, 2003). Genetic, reproductive, and environmental factors have all been suggested to influence breast cancer risk in BRCA1 mutation carriers (reviewed in Narod and Offit (2005)). The use of breast cancer as an endpoint to evaluate the protective role of potential modifying factors is not always feasible. Thus, there is a need to ascertain biomarkers of cancer susceptibility in these women. In turn, the identification of a valid biomarker of breast cancer risk would allow us to identify mutation carriers and help improve our ability to target, and to evaluate the effect of both dietary and lifestyle alterations, as well as, medical diagnostic and therapeutic interventions on breast cancer risk.The BRCA1 protein plays a vital role in maintaining genomic stability because of its key role in the repair of double-stranded DNA breaks by homologous recombination (reviewed in Welcsh et al (2000)). If double-stranded DNA breaks are left unrepaired, or are repaired inaccurately, aberrant chromosome breaks, deletions, and translocations may accumulate. Thus, an impaired cellular response to DNA damage appears to be a plausible mechanism whereby BRCA1 mutation carriers are at an increased risk of breast cancer (Scott, 2004). Hence, the evaluation of an individual's capacity to repair DNA may serve as a biomarker of breast cancer risk in carriers of these mutations.Two previous studies have shown higher levels of chromosomal aberrations and chromosomal breaks among women with a BRCA...

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Section: Discussionmentioning

confidence: 99%

DNA repair capacity as a possible biomarker of breast cancer risk in female BRCA1 mutation carriers

et al. 2007

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“…However, these assays require cell activation or transformation, and the time required to produce the data varied from days in case of cytogenetic tests in lymphocytes (dysenteric chromosomes, micronuclei, translocations, DNA fragments) to months for clonogenic survival. It has been reported that transformation itself may disrupt the radiosensitivity of normal cells [111; 112]. Geara and Peters [109; 113] reported lack of correlation between radiosensitivity of transformed lymphocytes and primary fibroblasts cultured from skin biopsy samples.…”

Section: Individual Radiosensitivitymentioning

confidence: 99%

Use of the γ-H2AX assay to monitor DNA damage and repair in translational cancer research

et al. 2012

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Formation of γ-H2AX in response to DNA double stranded breaks (DSBs) provides the basis for a sensitive assay of DNA damage in human biopsies. The review focuses on the application of γ-H2AX-based methods to translational studies to monitor the clinical response to DNA targeted therapies such as some forms of chemotherapy, external beam radiotherapy, radionuclide therapy or combinations thereof. The escalating attention on radiation biodosimetry has also highlighted the potential of the assay including renewed efforts to assess the radiosensitivity of prospective radiotherapy patients. Finally the γ-H2AX response has been suggested as a basis for an in vivo imaging modality.

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“…1), heterozygous for the c.190T>C mutation in BRCA1. Immortalization was achieved by exposure of lymphocytes to the supernatants of the B95-8 EBV producer cell line, according to a standard protocol (20). EBV cell lines were used to amplify transcript sequences of the BRCA1 gene by RT-PCR, and for radiosensitivity testing with the cytokinesisblock micronucleus (MN) assay, as previously described (20).…”

Section: Detection Of Brca1 and Brca2 Mutations And Polymorphismsmentioning

confidence: 99%

“…Immortalization was achieved by exposure of lymphocytes to the supernatants of the B95-8 EBV producer cell line, according to a standard protocol (20). EBV cell lines were used to amplify transcript sequences of the BRCA1 gene by RT-PCR, and for radiosensitivity testing with the cytokinesisblock micronucleus (MN) assay, as previously described (20). The EBV cell lines were maintained in RPMI-1640 medium supplemented with 10% fetal bovine serum, 2 mM L-glutamine, 100 U/ml penicillin, 0.1 mg/ml streptomycin with 10% FBS and cultured at 37˚C, 5% CO 2 , in a humidified incubator.…”

Section: Detection Of Brca1 and Brca2 Mutations And Polymorphismsmentioning

confidence: 99%

Characterization of the c.190T>C missense mutation in BRCA1 codon 64 (Cys64Arg)

Willems

Magri²,

Čretnik³

et al. 2009

Int J Oncol

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Abstract. In the Milan area (Northern Italy), we identified a family characterized by a high prevalence of ovarian and breast cancer cases (5 out of 6 subjects, over 3 generations), and a predominant prevalence of ovarian lesions (4 out of 5 patients). Analysis of BRCA1 and BRCA2 genes allowed the identification of the missense c.190T>C mutation in codon 64 (Cys64Arg) of BRCA1. The aims of the present investigation were to characterize the functional implications of the c.190T>C mutation at the molecular level, and to search whether additional polymorphisms might be linked to the peculiar phenotypic features observed in the Italian pedigree. Molecular modelling studies suggested that substitution of the cysteine 64 with an arginine likely disrupts the architecture of the BRCA1 RING finger domain, responsible for the interaction with BARD1, essential for the tumor-suppressor activity of the BRCA1-BARD1 complex. By splicing site information analysis, exonic splicing enhancer site characterization, and analysis of transcript fragment length and sequence, we showed that the c.190T>C mutation was able to modulate the splicing of exon 5 in a fashion opposite to the c.190T>G transversion, responsible for the functionally-related Cys64Gly amino acid substitution. Genotyping of BRCA1 and BRCA2 in the Italian family revealed the presence of two significant polymorphisms: the cancer-associated c.2612C>T SNP in BRCA1, and the c.-26G>A SNP in the BRCA2 gene, acting as an ovarian cancer risk modifier in carriers of deleterious BRCA1 mutations. Analysis of these SNPs in a genotypically-unrelated Polish family, characterized by prevalent breast neoplasms in carriers of the c.190T>C mutation, revealed a genetic profile consistent with the hypothetic role of both polymorphisms. IntroductionHereditary breast cancer accounts for 5-10% of all breast cancers worldwide. Prevalence of BRCA1 and BRCA2 mutations among high-risk cancer patients may vary by ethnicity, study inclusion criteria and mutation detection techniques. Since their early characterization, inherited BRCA1 mutations were found to be responsible for ~45% of inherited early-onset breast cancers, and for >80% of inherited breast and ovarian cancers (1). In Europe, BRCA1 mutation frequencies in breast cancer populations show considerable variability and range between 0.4% in Finland and 7% in Sweden (2). Meta-analytic mean cumulative cancer risks for mutation carriers at age 70 years are 57% for BRCA1 and 49% for BRCA2 mutation carriers; ovarian cancer risk is 40% for BRCA1 and 18% for BRCA2 mutation carriers (3). In Italy, breast cancer penetrance, estimated in a series of 568 families, was found to be 27% at age 50 and 39% at age 70 in BRCA1 mutation carriers, and 26% at age 50 and 44% at age 70 in BRCA2 mutation carriers (4). Ovarian cancer penetrances in Italy were estimated to be 14% at age 50 and 43% at age 70 in BRCA1 mutation carriers, and 3% at age 50 and 15% at age 70 in BRCA2 mutation carriers (4).BRCA1 is a gene with a coding sequence spanning over 23 exons, i...

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

Cited by 24 publications

References 24 publications

DNA repair capacity as a possible biomarker of breast cancer risk in female BRCA1 mutation carriers

DNA repair capacity as a possible biomarker of breast cancer risk in female BRCA1 mutation carriers

Use of the γ-H2AX assay to monitor DNA damage and repair in translational cancer research

Characterization of the c.190T>C missense mutation in BRCA1 codon 64 (Cys64Arg)

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