Irmgard Helmbold

Association between Polymorphisms in the DNA Repair Genes, XRCC1, APE1, and XPD and Acute Side Effects of Radiotherapy in Breast Cancer Patients

Purpose: Several DNA repair gene polymorphisms have been described, which affect DNA repair capacity and modulate cancer susceptibility. We evaluated the association of six polymorphisms in the DNA repair genes: XRCC1 (Arg194Trp, Arg280His, and Arg399Gln), APE1 (Asp148Glu), and XPD (Lys751Gln and Asp312Asn), with the risk of acute skin reactions following radiotherapy. Design: We conducted a prospective study of 446 female patients with breast cancer who received radiotherapy after breast-conserving surgery. Individual genetic polymorphisms were determined using melting point analysis of sequence-specific hybridization probes. The development of acute skin reactions (moist desquamation) associated with DNA repair gene polymorphisms was modeled using Cox proportional hazards, accounting for cumulative biologically effective radiation dose. Results: Overall, the development of acute toxicity, which presented in 77 patients, was not associated with the genetic variants studied, although the hazard ratios (HR) were generally below 1. Risks were however differential by body mass index. Among normal-weight patients only, both carriers of the APE1 148Glu and the XRCC1 399Gln alleles had decreased risk of acute skin reactions after radiotherapy (HR, 0.49 and 0.51, respectively). The results for XRCC1 were confirmed by haplotype analysis. When considering joint effects, we observed that compared with homozygote carriers of the wild-type allele in both genes, the risk was most strongly reduced in carriers of both APE1 148Glu and XRCC1 399Gln alleles with normal weight [HR, 0.19; 95% confidence interval (95% CI), 0.06-0.56] but not in those with overweight (HR, 1.39; 95% CI, 0.56-3.45; Pinteraction = 0.009). Conclusion: The XRCC1 399Gln or APE1 148Glu alleles may be protective against the development of acute side effects after radiotherapy in patients with normal weight.

Predictive factors for late normal tissue complications following radiotherapy for breast cancer.

Background and purposeRadiotherapy after breast-conserving surgery is commonly applied to reduce recurrence of breast cancer but may cause acute and late side effects. To identify prognostic factors for the development of late toxicity after radiotherapy, we conducted a prospective study of breast cancer patients.Patients and methodsWe assessed late complications of radiotherapy and collected information on epidemiologic factors in a cohort of breast cancer patients who had received radiotherapy after breast-conserving surgery. Among 416 patients with complete follow-up data, the association between possible risk factors and development of late complications was evaluated using multivariate logistic regression analysis.ResultsAfter a median follow-up time of 51xa0months, 131 (31.4%) patients presented with telangiectasia and 28 (6.7%) patients with fibrosis. We observed a strong association between development of telangiectasia and fibrosis (pxa0<xa00.01). Increasing age of the patient was a risk factor for both telangiectasia and fibrosis (p-value for trend <0.01 and 0.03, respectively). Patients with acute skin toxicity (odds ratio (OR) 1.8, 95% confidence interval (CI) 1.0–3.1) were at higher risk to develop telangiectasia. Long-term smoking was associated with a significant increase in risk of telangiectasia compared to non-smokers (OR 2.3, 95% CI 1.2–4.6).ConclusionsOur study revealed several factors other than radiation dose that may predispose to late complications following radiotherapy. Further understanding of differences in response to irradiation may advance individualized treatment and improve cosmetic outcome.

Personal characteristics, therapy modalities and individual DNA repair capacity as predictive factors of acute skin toxicity in an unselected cohort of breast cancer patients receiving radiotherapy

BACKGROUND AND PURPOSEnIntrinsic and extrinsic factors can affect the occurrence of side effects of radiotherapy. The influence of therapy modalities, personal characteristics and individual DNA repair capacity on the risk of acute skin toxicity was thus evaluated.nnnMATERIALS AND METHODSnIn a prospective study of 478 female breast cancer patients receiving adjuvant radiotherapy of the breast after breast-conserving surgery, acute skin toxicity was documented systematically using a modified version of the common toxicity criteria. Prognostic personal and treatment characteristics were identified for the entire cohort. Individual DNA repair capacity was determined in a subgroup of 113 patients with alkaline comet assay using phytohemagglutinin stimulated lymphocytes. Using proportional hazards analysis to account for cumulative biologically effective radiation dose, the hazard for the development of acute skin reactions (moist desquamation) associated with DNA repair capacity was modeled.nnnRESULTSnOf the 478 participants, 84 presented with acute reactions by the end of treatment. Higher body mass index was significantly associated with an increased risk for acute reactions (hazard ratio=1.09 per 1 kg/m(2)), adjusted for treating hospital and photon beam quality. The comet assay parameters examined, including background DNA damage in non-irradiated cells, DNA damage induced by 5 Gy, and DNA repair capacity, were not significantly associated with risk of acute skin toxicity.nnnCONCLUSIONSnHigher BMI is predictive of acute skin toxicity, however, individual repair parameters as determined by the alkaline comet assay are not informative enough. More comprehensive analyses including late effects of radiotherapy and repair kinetics optimized for different radiation-induced DNA lesions are warranted.

Radiation-induced DNA damage and repair in lymphocytes from breast cancer patients and their correlation with acute skin reactions to radiotherapy

PURPOSEnRepair of radiation-induced DNA damage plays a critical role for both the susceptibility of patients to side effects after radiotherapy and their subsequent cancer risk. The study objective was to evaluate whether DNA repair data determined in vitro are correlated with the occurrence of acute side effects during radiotherapy.nnnMETHODS AND MATERIALSnBreast cancer patients receiving radiation therapy after a breast-conserving surgery were recruited in a prospective epidemiologic study. As an indicator for clinical radiosensitivity, adverse reactions of the skin were recorded. Cryo-preserved lymphocytes from 113 study participants were gamma-irradiated with 5 Gy in vitro and analyzed using the alkaline comet assay. Reproducibility of the assay was determined by repeated analysis (n = 26) of cells from a healthy donor. A coefficient of variation of 0.3 was calculated.nnnRESULTSnThe various parameters determined to characterize the individual DNA repair capacity showed large differences between patients. Eleven patients were identified with considerably enhanced DNA damage induction, and 7 patients exhibited severely reduced DNA repair capacity after 15 and 30 min. Six patients were considered as clinically radiosensitive, indicated by moist desquamation of the skin after a total radiation dose of about 50 Gy.nnnCONCLUSIONSnUsing the alkaline comet assay as described here, breast cancer patients were identified showing abnormal cellular radiation effects, but this repair deficiency corresponded only at a very limited extent to the acute radiation sensitivity of the skin. Because impaired DNA repair could be involved in the development of late irradiation effects, individuals exhibiting severely reduced DNA repair capacity should be followed for the development of late clinical symptoms.

Genetic polymorphisms in DNA repair and damage response genes and late normal tissue complications of radiotherapy for breast cancer

Breast-conserving surgery followed by radiotherapy is effective in reducing recurrence; however, telangiectasia and fibrosis can occur as late skin side effects. As radiotherapy acts through producing DNA damage, we investigated whether genetic variation in DNA repair and damage response confers increased susceptibility to develop late normal skin complications. Breast cancer patients who received radiotherapy after breast-conserving surgery were examined for late complications of radiotherapy after a median follow-up time of 51 months. Polymorphisms in genes involved in DNA repair (APEX1, XRCC1, XRCC2, XRCC3, XPD) and damage response (TP53, P21) were determined. Associations between telangiectasia and genotypes were assessed among 409 patients, using multivariate logistic regression. A total of 131 patients presented with telangiectasia and 28 patients with fibrosis. Patients with variant TP53 genotypes either for the Arg72Pro or the PIN3 polymorphism were at increased risk of telangiectasia. The odds ratios (OR) were 1.66 (95% confidence interval (CI): 1.02–2.72) for 72Pro carriers and 1.95 (95% CI: 1.13–3.35) for PIN3 A2 allele carriers compared with non-carriers. The TP53 haplotype containing both variant alleles was associated with almost a two-fold increase in risk (OR 1.97, 95% CI: 1.11–3.52) for telangiectasia. Variants in the TP53 gene may therefore modify the risk of late skin toxicity after radiotherapy.

Polymorphisms in genes related to oxidative stress (CAT, MnSOD, MPO, and eNOS) and acute toxicities from radiation therapy following lumpectomy for breast cancer.

Purpose: Because radiotherapy exerts cytotoxic effects via generation of massive oxidative stress, we hypothesized that catalase, manganese superoxide dismutase, myeloperoxidase (MPO), and endothelial nitric oxide synthase (eNOS) genotypes might result in greater risk of radiotoxicity. Experimental Design: Cases (n = 446) were Caucasian women with breast cancer who received radiotherapy following lumpectomy. Genotypes were determined by matrix-assisted laser desorption/ionization time-of-flight. The development of acute reactions (moist desquamation) associated with genotypes was modeled using the Cox proportional hazards model, accounting for cumulative biologically effective radiation dose. Results: Genotypes associated with higher levels of reactive oxygen species (ROS) were not associated with risk of radiotoxicity. However, relationships between overweight/obesity [body mass index (BMI), >25] and radiotoxicity risk seemed to be modified by eNOS and MPO genotypes associated with higher generation of nitric oxide and ROS, respectively. Women with high BMI (>25) and eNOS GG genotypes were at more than a 6-fold increase in risk (hazard ratio, 6.39; 95% confidence interval, 2.53-16.15) compared with those with BMI <25, and for MPO, those with high BMI (>25) and GG genotypes also had greater risk of radiotoxicity (hazard ratio, 3.61; 95% confidence interval, 1.78-7.35) compared with those with BMI <25. Overweight/obesity was not a strong risk factor among women with other eNOS and MPO genotypes. Exploratory analysis using classification and regression trees indicated that total number of risk alleles contributed, in part, to acute toxicity outcomes among a subgroup of women. Conclusions: Associations between BMI and radiotoxicity risk may be most apparent among women with genotypes related to higher levels of oxidative stress. Regression trees may be useful in future studies to examine the contributions of multiple factors to individual susceptibility to adverse effects of cancer treatment.

Association between TP53 and p21 genetic polymorphisms and acute side effects of radiotherapy in breast cancer patients

Summaryp53 and p21 play an important role in G1/S checkpoint control in response to ionizing radiation. Yet the genetic polymorphisms in these genes have not been investigated with respect to radiation toxicity in patients. We therefore assessed the association between TP53 Arg72Pro, p53PIN3 and p21 Ser31Arg polymorphisms and the risk of acute skin toxicity after radiotherapy in a prospective study of 446 female breast cancer patients (average age 60.3±9.0xa0years) receiving radiotherapy after breast conserving surgery. The p53PIN3 polymorphism was determined by standard PCR, and TP53 Arg72Pro and p21 Ser31Arg polymorphisms using melting point analysis of sequence-specific hybridization probes. The development of acute skin toxicity (moist desquamation) was modelled using Cox proportional hazards, accounting for cumulative biologically effective radiation dose. Overall, the development of acute skin toxicity, which presented in 77 patients, was not significantly associated with the polymorphisms studied. Risks were however differential by body mass index. Compared to non-carriers, TP53 72Pro carriers had a non-significantly decreased risk of acute skin toxicity in normal weight women (hazard ratio 0.46, 95% CI, 0.18–1.18) but not in overweight patients (hazard ratio 1.07, 95% CI, 0.61–1.89) (pinteraction =0.14). Haplotype analysis for the TP53 polymorphisms suggested that effect modification by TP53 72Pro may differ according to the p53PIN3 allele (pinteraction=0.06). Furthermore, in TP53 72Pro carriers with p21 Ser/Ser genotype, the occurrence of acute toxicity was reduced in normal weight but not overweight patients. In conclusion, the TP53 72Pro variant may be associated with the development of acute skin toxicity after radiotherapy in patients with normal weight. Large clinical studies are needed to clearly confirm this association.

Genetic predictors of acute toxicities related to radiation therapy following lumpectomy for breast cancer: a case-series study

IntroductionThe cytotoxic effects of radiation therapy are mediated primarily through increased formation of hydroxyl radicals and reactive oxygen species, which can damage cells, proteins and DNA; the glutathione S-transferases (GSTs) function to protect against oxidative stress. We hypothesized that polymorphisms encoding reduced or absent activity in the GSTs might result in greater risk for radiation-associated toxicity.MethodsWomen receiving therapy in radiation units in Germany following lumpectomy for breast cancer (1998–2001) provided a blood sample and completed an epidemiological questionnaire (n = 446). Genotypes were determined using Sequonom MALDI-TOF (GSTA1, GSTP1) and Masscode (GSTM1, GSTT1). Biologically effective radiotherapy dose (BED) was calculated, accounting for differences in fractionation and overall treatment time. Side effects considered were grade 2c and above, as classified using the modified Common Toxicity Criteria. Predictors of toxicity were modelled using Cox regression models in relation to BED, with adjustment for treating clinic, photon field, beam energy and boost method, and potential confounding variables.ResultsLow activity GSTP1 genotypes were associated with a greater than twofold increase in risk for acute skin toxicities (adjusted hazard ratio 2.28, 95% confidence interval 1.04–4.99). No associations were noted for the other GST genotypes.ConclusionThese data indicate that GSTP1 plays an important role in protecting normal cells from damage associated with radiation therapy. Studies examining the effects of GSTP1 polymorphisms on toxicity, recurrence and survival will further inform individualized therapeutics based on genotypes.

Genetic predictors of long‐term toxicities after radiation therapy for breast cancer

Telangiectasia and subcutaneous fibrosis are the most common late dermatologic side effects observed in response to radiation treatment. Radiotherapy acts on cancer cells largely due to the generation of reactive oxygen species (ROS). ROS also induce normal tissue toxicities. Therefore, we investigated if genetic variation in oxidative stress‐related enzymes confers increased susceptibility to late skin complications. Women who received radiotherapy following lumpectomy for breast cancer were followed prospectively for late tissue side effects after initial treatment. Final analysis included 390 patients. Polymorphisms in genes involved in oxidative stress‐related mechanisms (GSTA1, GSTM1, GSTT1, GSTP1, MPO, MnSOD, eNOS, CAT) were determined from blood samples by MALDI‐TOF. The associations between telangiectasia and genotypes were evaluated by multivariate unconditional logistic regression models. Patients with variant GSTA1 genotypes were at significantly increased risk of telangiectasia (OR 1.86, 95% CI 1.11–3.11). Reduced odds ratios of telangiectasia were noted for women with lower‐activity eNOS genotype (OR 0.58, 95% CI 0.36–0.93). Genotype effects were modified by follow‐up time, with the highest risk observed after 4 years of radiotherapy for gene polymorphisms in ROS‐neutralizing enzymes. Decreased risk with eNOS polymorphisms was significant only among women with less than 4 years of follow‐up. All other risk estimates were nonsignificant. Late effects of radiation therapy on skin appear to be modified by variants in genes related to protection from oxidative stress. The application of genomics to outcomes following radiation therapy holds the promise of radiation dose adjustment to improve both cosmetic outcomes and quality of life for breast cancer patients.

Genetic Polymorphisms in the DNA Double-Strand Break Repair Genes XRCC3, XRCC2, and NBS1 Are Not Associated with Acute Side Effects of Radiotherapy in Breast Cancer Patients

Clinical sensitivity to ionizing radiation varies considerably among patients, and radiation-induced adverse effects developing in normal tissue can be therapy limiting in >10% of patients ([1][1]). Ionizing radiation induces both DNA single-strand breaks and double-strand breaks (DSB), with the