Jennifer S. Wilkinson

Independent validation of genes and polymorphisms reported to be associated with radiation toxicity: a prospective analysis study

BACKGROUND Several studies have reported associations between radiation toxicity and single nucleotide polymorphisms (SNPs) in candidate genes. Few associations have been tested in independent validation studies. This prospective study aimed to validate reported associations between genotype and radiation toxicity in a large independent dataset. METHODS 92 (of 98 attempted) SNPs in 46 genes were successfully genotyped in 1613 patients: 976 received adjuvant breast radiotherapy in the Cambridge breast IMRT trial (ISRCTN21474421, n=942) or in a prospective study of breast toxicity at the Christie Hospital, Manchester, UK (n=34). A further 637 received radical prostate radiotherapy in the MRC RT01 multicentre trial (ISRCTN47772397, n=224) or in the Conventional or Hypofractionated High Dose Intensity Modulated Radiotherapy for Prostate Cancer (CHHiP) trial (ISRCTN97182923, n=413). Late toxicity was assessed 2 years after radiotherapy with a validated photographic technique (patients with breast cancer only), clinical assessment, and patient questionnaires. Association tests of genotype with overall radiation toxicity score and individual endpoints were undertaken in univariate and multivariable analyses. At a type I error rate adjusted for multiple testing, this study had 99% power to detect a SNP, with minor allele frequency of 0·35, associated with a per allele odds ratio of 2·2. FINDINGS None of the previously reported associations were confirmed by this study, after adjustment for multiple comparisons. The p value distribution of the SNPs tested against overall toxicity score was not different from that expected by chance. INTERPRETATION We did not replicate previously reported late toxicity associations, suggesting that we can essentially exclude the hypothesis that published SNPs individually exert a clinically relevant effect. Continued recruitment of patients into studies within the Radiogenomics Consortium is essential so that sufficiently powered studies can be done and methodological challenges addressed. FUNDING Cancer Research UK, The Royal College of Radiologists, Addenbrookes Charitable Trust, Breast Cancer Campaign, Cambridge National Institute of Health Research (NIHR) Biomedical Research Centre, Experimental Cancer Medicine Centre, East Midlands Innovation, the National Cancer Institute, Joseph Mitchell Trust, Royal Marsden NHS Foundation Trust, Institute of Cancer Research NIHR Biomedical Research Centre for Cancer.

Randomized Controlled Trial of Intensity-Modulated Radiotherapy for Early Breast Cancer: 5-Year Results Confirm Superior Overall Cosmesis

PURPOSE There are few randomized controlled trial data to confirm that improved homogeneity with simple intensity-modulated radiotherapy (IMRT) decreases late breast tissue toxicity. The Cambridge Breast IMRT trial investigated this hypothesis, and the 5-year results are reported. PATIENTS AND METHODS Standard tangential plans of 1,145 trial patients were analyzed; 815 patients had inhomogeneous plans (≥ 2 cm(3) receiving 107% of prescribed dose: 40 Gy in 15 fractions over 3 weeks) and were randomly assigned to standard radiotherapy (RT) or replanned with simple IMRT; 330 patients with satisfactory dose homogeneity were treated with standard RT and underwent the same follow-up as the randomly assigned patients. Breast tissue toxicities were assessed at 5 years using validated methods: photographic assessment (overall cosmesis and breast shrinkage compared with baseline pre-RT photographs) and clinical assessment (telangiectasia, induration, edema, and pigmentation). Comparisons between different groups were analyzed using polychotomous logistic regression. RESULTS On univariate analysis, compared with standard RT, fewer patients in the simple IMRT group developed suboptimal overall cosmesis (odds ratio [OR], 0.68; 95% CI, 0.48 to 0.96; P = .027) and skin telangiectasia (OR, 0.58; 95% CI, 0.36 to 0.92; P = .021). No evidence of difference was seen for breast shrinkage, breast edema, tumor bed induration, or pigmentation. The benefit of IMRT was maintained on multivariate analysis for both overall cosmesis (P = .038) and skin telangiectasia (P = .031). CONCLUSION Improved dose homogeneity with simple IMRT translates into superior overall cosmesis and reduces the risk of skin telangiectasia. These results are practice changing and should encourage centers still using two-dimensional RT to implement simple breast IMRT.

Randomized controlled trial of forward-planned intensity modulated radiotherapy for early breast cancer: interim results at 2 years.

PURPOSE This single-center randomized trial was designed to investigate whether intensity-modulated radiotherapy (IMRT) reduces late toxicity in patients with early-stage breast cancer. METHODS AND MATERIALS The standard tangential plans of 1,145 nonselected patients were analyzed. The patients with inhomogeneous plans were randomized to a simple method of forward-planned IMRT or standard radiotherapy (RT). The primary endpoint was serial photographic assessment of breast shrinkage. RESULTS At 2 years, no significant difference was found in the development of any photographically assessed breast shrinkage between the patients randomized to the interventional or control group (odds ratio, 1.51; 95% confidence interval, 0.83-1.58; p = .41). The patients in the control group were more likely to develop telangiectasia than those in the IMRT group (odds ratio, 1.68; 95% confidence interval 1.13-2.40; p = .009). Poor baseline surgical cosmesis resulted in poor overall cosmesis at 2 years after RT. In patients who had good surgical cosmesis, those randomized to IMRT were less likely to deteriorate to a moderate or poor overall cosmesis than those in the control group (odds ratio, 0.63; 95% confidence interval, 0.39-1.03, p = .061). CONCLUSIONS IMRT can lead to a significant reduction in telangiectasia at comparatively early follow-up of only 2 years after RT completion. An important component of breast induration and shrinkage will actually result from the surgery and not from the RT. Surgical cosmesis is an important determinant of overall cosmesis and could partially mask the longer term benefits of IMRT at this early stage.

A randomised controlled trial of forward-planned radiotherapy (IMRT) for early breast cancer: baseline characteristics and dosimetry results.

BACKGROUND AND PURPOSE This large trial was designed to investigate whether correction of dose inhomogeneities using intensity-modulated radiotherapy (IMRT) reduces late toxicity and improves quality of life in patients with early breast cancer. This paper reports baseline characteristics of trial participants and dosimetry results. MATERIALS AND METHODS Standard tangential plans of 1145 trials were analysed. Patients with inhomogeneous plans, defined by ICRU recommendations, were randomised to forward-planned IMRT or standard radiotherapy. RESULTS Twenty-nine percentage of patients had adequate dosimetry with standard 2D radiotherapy. In the randomised patients, the decreases in mean volumes receiving greater than 107% (Vol>107) and less than 95% (Vol<95) of the prescribed dose in the IMRT compared with the control group were 34.0 cm(3) (95% CI 26.4-41.6; P<0.0001) and 48.1 cm(3) (95% CI 34.4-61.9; P<0.0001), respectively. In this study, 90% of patients who had a breast separation greater > or = 21 cm had Vol>107>2 cm(3) on standard radiotherapy plans. CONCLUSION This large trial, in which patients with all breast sizes were eligible, confirmed that breast dosimetry can be significantly improved with a simple method of forward-planned IMRT and has little impact on radiotherapy resources. It is shown that patients with larger breasts are more likely to have dose inhomogeneities and breast separation gives some indication of this likelihood. Photographic assessment of patients at 2 years after radiotherapy, as the next part of this randomised controlled trial, will show whether these results for IMRT translate into improved cosmetic outcome in patients with early breast cancer. This would provide impetus for the widespread adoption of 3D planning and IMRT.

A genome wide association study (GWAS) providing evidence of an association between common genetic variants and late radiotherapy toxicity

BACKGROUND AND PURPOSE This study was designed to identify common single nucleotide polymorphisms (SNPs) associated with toxicity 2years after radiotherapy. MATERIALS AND METHODS A genome wide association study was performed in 1850 patients from the RAPPER study: 1217 received adjuvant breast radiotherapy and 633 had radical prostate radiotherapy. Genotype associations with both overall and individual endpoints of toxicity were tested via univariable and multivariable regression. Replication of potentially associated SNPs was carried out in three independent patient cohorts who had radiotherapy for prostate (516 RADIOGEN and 862 Gene-PARE) or breast (355 LeND) cancer. RESULTS Quantile-quantile plots show more associations at the P<5×10(-7) level than expected by chance (164 vs. 9 for the prostate cases and 29 vs. 4 for breast cases), providing evidence that common genetic variants are associated with risk of toxicity. Strongest associations were for individual endpoints rather than an overall measure of toxicity in all patients. However, in general, significant associations were not validated at a nominal 0.05 level in the replication cohorts. CONCLUSIONS This largest GWAS to date provides evidence of true association between common genetic variants and toxicity. Associations with toxicity appeared to be tumour site-specific. Future GWAS require higher statistical power, in particular in the validation stage, to test clinically relevant effect sizes of SNP associations with individual endpoints, but the required sample sizes are achievable.

No association between SNPs regulating TGF-β1 secretion and late radiotherapy toxicity to the breast: results from the RAPPER study.

BACKGROUND AND PURPOSE Several small studies have reported associations between TGFB1 single nucleotide polymorphisms (SNPs), considered to increase secretion of TGF-β1, and greater than 3-fold increases in incidence of fibrosis - an indicator of late toxicity after radiotherapy in breast cancer patients. MATERIALS AND METHODS Two SNPs in TGFB1, C-509T (rs1800469) and L10P (rs1800470), were genotyped in 778 breast cancer patients who had received radiotherapy to the breast. Late radiotherapy toxicity was assessed two years after radiotherapy using a validated photographic technique, clinical assessment and patient questionnaires. RESULTS On photographic assessment, 210 (27%) patients showed some degree of breast shrinkage, whilst 45 (6%) patients showed marked breast shrinkage. There was no significant association of genotype at either of the TGFB1 SNPs with any measure of late radiation toxicity. CONCLUSION This adequately powered trial failed to confirm previously reported increases in fibrosis with TGFB1 genotype - any increase greater than 1.36 can be excluded with 95% confidence. Similar frequent failures to replicate associations with candidate genes have been resolved using genome-wide association scans: this methodology detects common, low risk alleles but requires even larger patient numbers for adequate statistical power.

Individual patient data meta-analysis shows no association between the SNP rs1800469 in TGFB and late radiotherapy toxicity

BACKGROUND AND PURPOSE Reported associations between risk of radiation-induced normal tissue injury and single nucleotide polymorphisms (SNPs) in TGFB1, encoding the pro-fibrotic cytokine transforming growth factor-beta 1 (TGF-β1), remain controversial. To overcome publication bias, the international Radiogenomics Consortium collected and analysed individual patient level data from both published and unpublished studies. MATERIALS AND METHODS TGFB1 SNP rs1800469 c.-1347T>C (previously known as C-509T) genotype, treatment-related data, and clinically-assessed fibrosis (measured at least 2years after therapy) were available in 2782 participants from 11 cohorts. All received adjuvant breast radiotherapy. Associations between late fibrosis or overall toxicity, reported by STAT (Standardised Total Average Toxicity) score, and rs1800469 genotype were assessed. RESULTS No statistically significant associations between either fibrosis or overall toxicity and rs1800469 genotype were observed with univariate or multivariate regression analysis. The multivariate odds ratio (OR), obtained from meta-analysis, for an increase in late fibrosis grade with each additional rare allele of rs1800469 was 0.98 (95% Confidence Interval (CI) 0.85-1.11). This CI is sufficiently narrow to rule out any clinically relevant effect on toxicity risk in carriers vs. non-carriers with a high probability. CONCLUSION This meta-analysis has not confirmed previous reports of association between fibrosis or overall toxicity and rs1800469 genotype in breast cancer patients. It has demonstrated successful collaboration within the Radiogenomics Consortium.

The Cambridge Breast Intensity-modulated Radiotherapy Trial: patient- and treatment-related factors that influence late toxicity.

AIMS The effect of patient- and treatment-related factors in the development of late normal tissue toxicity after radiotherapy is not yet fully established. The aim of this study was to elucidate the relative importance of such factors in the development of late toxicity after breast-conserving surgery and adjuvant breast radiotherapy. MATERIALS AND METHODS Patient- and treatment-related factors were analysed in 1014 patients who had received adjuvant radiotherapy to the breast in the Cambridge Breast Intensity-modulated Radiotherapy (IMRT) Trial. Late toxicity data were collected using photographic and clinical assessments and patient-reported questionnaires at 2 years after radiotherapy. RESULTS On multivariate analysis, a larger breast volume was statistically significantly associated with the development of breast shrinkage assessed by serial photographs (odds ratio per litre increase in breast volume = 1.98, 95% confidence interval 1.41, 2.78; P < 0.0005), telangiectasia (odds ratio = 3.94, 95% confidence interval 2.49, 6.24; P < 0.0005), breast oedema (odds ratio = 3.65, 95% confidence interval 2.54, 5.24; P < 0.0005) and pigmentation (odds ratio = 1.75, 95% confidence interval 1.21, 2.51; P = 0.003). Current smokers had an increased risk of developing pigmentation (odds ratio = 2.09, 95% confidence interval 1.23, 3.54; P = 0.006). Patients with a moderate or poor post-surgical cosmesis had a greatly increased risk of moderate or poor overall cosmesis (odds ratio = 38.19; 95% confidence interval 21.9, 66.7; P < 0.0005). Postoperative infection requiring antibiotics was associated with increased risk of telangiectasia (odds ratio = 3.39, 95% confidence interval 1.94, 5.91; P < 0.0005) and breast oversensitivity (odds ratio = 1.78, 95% confidence interval 1.27, 2.49; P = 0.001). CONCLUSIONS In this study, the greatest risk factors for the development of late toxicity 2 years after breast-conserving surgery and adjuvant radiotherapy were larger breast volume, baseline pre-radiotherapy surgical cosmesis, postoperative infection and possibly smoking. These factors seem to be more important than relatively small differences in dose inhomogeneity and the addition of boost radiotherapy at 2 years after the completion of radiotherapy. The modification of potentially preventable risk factors, such as postoperative infection and smoking, may limit the development of late toxicity after breast radiotherapy.

Standardized total average toxicity score: a scale- and grade-independent measure of late radiotherapy toxicity to facilitate pooling of data from different studies.

PURPOSE The search for clinical and biologic biomarkers associated with late radiotherapy toxicity is hindered by the use of multiple and different endpoints from a variety of scoring systems, hampering comparisons across studies and pooling of data. We propose a novel metric, the Standardized Total Average Toxicity (STAT) score, to try to overcome these difficulties. METHODS AND MATERIALS STAT scores were derived for 1010 patients from the Cambridge breast intensity-modulated radiotherapy trial and 493 women from the University Hospitals of Leicester. The sensitivity of the STAT score to detect differences between patient groups, stratified by factors known to influence late toxicity, was compared with that of individual endpoints. Analysis of residuals was used to quantify the effect of these covariates. RESULTS In the Cambridge cohort, STAT scores detected differences (p < 0.00005) between patients attributable to breast volume, surgical specimen weight, dosimetry, acute toxicity, radiation boost to tumor bed, postoperative infection, and smoking (p < 0.0002), with no loss of sensitivity over individual toxicity endpoints. Diabetes (p = 0.017), poor postoperative surgical cosmesis (p = 0.0036), use of chemotherapy (p = 0.0054), and increasing age (p = 0.041) were also associated with increased STAT score. When the Cambridge and Leicester datasets were combined, STAT was associated with smoking status (p < 0.00005), diabetes (p = 0.041), chemotherapy (p = 0.0008), and radiotherapy boost (p = 0.0001). STAT was independent of the toxicity scale used and was able to deal with missing data. There were correlations between residuals of the STAT score obtained using different toxicity scales (r > 0.86, p < 0.00005 for both datasets). CONCLUSIONS The STAT score may be used to facilitate the analysis of overall late radiation toxicity, from multiple trials or centers, in studies of possible genetic and nongenetic determinants of radiotherapy toxicity.

Association of breast tumour bed seroma with post-operative complications and late normal tissue toxicity: Results from the Cambridge Breast IMRT trial

AIMS There are two main surgical techniques for managing the tumour bed after breast cancer excision. Firstly, closing the defect by suturing the cavity walls together and secondly leaving the tumour bed open thus allowing seroma fluid to collect. There is debate regarding which technique is preferable, as it has been reported that a post-operative seroma increase post-operative infection rates and late normal tissue side effects. METHODS Data from 648 patients who participated in the Cambridge Breast IMRT trial were used. Seromas were identified on axial CT images at the time of radiotherapy planning and graded as not visible/subtle or easily visible. An association was sought between the presence of seroma and the development of post-operative infection, post-operative haematoma and 2 and 5 years normal tissue toxicity (assessed using serial photographs, clinical assessment and self assessment questionnaire). RESULTS The presence of easily visible seroma was associated with increased risk of post-operative infection (OR = 1.80; p = 0.004) and post-operative haematoma (OR = 2.1; p = 0.02). Breast seroma was an independent risk factor for whole breast induration and tumour bed induration at 2 and 5 years. The presence of breast seroma was also associated with inferior overall cosmesis at 5 years. There was no significant association between the presence of seroma and the development of either breast shrinkage or breast pain. CONCLUSION The presence of seroma at the time of radiotherapy planning is associated with increased rates of post-operative infection and haematoma. It is also an independent risk factor for late normal tissue toxicity. This study suggests that full thickness surgical closure may be desirable for patients undergoing breast conservation and radiotherapy.