Clinical Outcomes of Patients with Metastatic Breast Cancer Treated with Hypo-Fractionated Liver Radiotherapy.

PURPOSE: To retrospectively review the clinical outcomes of patients with metastatic breast cancer (MBCa) following liver directed ablative intent radiotherapy (RT). METHODS: Demographics, disease and treatment characteristics of patients with MBCa who received liver metastasis (LM) directed ablative RT between 2004-2020 were analysed. The primary outcome was local control (LC), secondary outcomes included overall survival (OS) and progression-free survival (PFS) analyzed by univariate (UVA) and multi-variable analysis (MVA). RESULTS: Thirty MBCa patients with 50 LM treated with 5-10 fraction RT were identified. Median follow-up was 14.6 (range 0.9-156.2) months. Class of metastatic disease was described as induced (12 patients, 40%), repeat (15 patients, 50%) and de novo (three patients, 10%). Median size of treated LM was 3.1 cm (range 1-8.8 cm) and median biologically effective dose delivered was 122 (Q1-Q3; 98-174) Gy3. One-year LC rate was 100%. One year and two-year survival was 89% and 63%, respectively, with size of treated LM predictive of OS (HR 1.35, p = 0.023) on UVA. Patients with induced OMD had a significantly higher rate of progression (HR 4.77, p = 0.01) on UVA, trending to significance on MVA (HR 3.23, p = 0.051). CONCLUSIONS: Hypo-fractionated ablative liver RT in patients with MBCa provides safe, tolerable treatment with excellent LC.

Comprehensive Cardiovascular Magnetic Resonance Tissue Characterization and Cardiotoxicity in Women With Breast Cancer.

Importance: There is a growing interest in understanding whether cardiovascular magnetic resonance (CMR) myocardial tissue characterization helps identify risk of cancer therapy-related cardiac dysfunction (CTRCD). Objective: To describe changes in CMR tissue biomarkers during breast cancer therapy and their association with CTRCD. Design, Setting, and Participants: This was a prospective, multicenter, cohort study of women with ERBB2 (formerly HER2)-positive breast cancer (stages I-III) who were scheduled to receive anthracycline and trastuzumab therapy with/without adjuvant radiotherapy and surgery. From November 7, 2013, to January 16, 2019, participants were recruited from 3 University of Toronto-affiliated hospitals. Data were analyzed from July 2021 to June 2022. Exposures: Sequential therapy with anthracyclines, trastuzumab, and radiation. Main Outcomes and Measures: CMR, high-sensitivity cardiac troponin I (hs-cTnI), and B-type natriuretic peptide (BNP) measurements were performed before anthracycline treatment, after anthracycline and before trastuzumab treatment, and at 3-month intervals during trastuzumab therapy. CMR included left ventricular (LV) volumes, LV ejection fraction (EF), myocardial strain, early gadolinium enhancement imaging to assess hyperemia (inflammation marker), native/postcontrast T1 mapping (with extracellular volume fraction [ECV]) to assess edema and/or fibrosis, T2 mapping to assess edema, and late gadolinium enhancement (LGE) to assess replacement fibrosis. CTRCD was defined using the Cardiac Review and Evaluation Committee criteria. Fixed-effects models or generalized estimating equations were used in analyses. Results: Of 136 women (mean [SD] age, 51.1 [9.2] years) recruited from 2013 to 2019, 37 (27%) developed CTRCD. Compared with baseline, tissue biomarkers of myocardial hyperemia and edema peaked after anthracycline therapy or 3 months after trastuzumab initiation as demonstrated by an increase in mean (SD) relative myocardial enhancement (baseline, 46.3% [16.8%] to peak, 56.2% [18.6%]), native T1 (1012 [26] milliseconds to 1035 [28] milliseconds), T2 (51.4 [2.2] milliseconds to 52.6 [2.2] milliseconds), and ECV (25.2% [2.4%] to 26.8% [2.7%]), with P <.001 for the entire follow-up. The observed values were mostly within the normal range, and the changes were small and recovered during follow-up. No new replacement fibrosis developed. Increase in T1, T2, and/or ECV was associated with increased ventricular volumes and BNP but not hs-cTnI level. None of the CMR tissue biomarkers were associated with changes in LVEF or myocardial strain. Change in ECV was associated with concurrent and subsequent CTRCD, but there was significant overlap between patients with and without CTRCD. Conclusions and Relevance: In women with ERBB2-positive breast cancer receiving sequential anthracycline and trastuzumab therapy, CMR tissue biomarkers suggest inflammation and edema peaking early during therapy and were associated with ventricular remodeling and BNP elevation. However, the increases in CMR biomarkers were transient, were not associated with LVEF or myocardial strain, and were not useful in identifying traditional CTRCD risk.

A Combined Echocardiography Approach for the Diagnosis of Cancer Therapy-Related Cardiac Dysfunction in Women With Early-Stage Breast Cancer.

IMPORTANCE: Diagnosis of cancer therapy-related cardiac dysfunction (CTRCD) remains a challenge. Cardiovascular magnetic resonance (CMR) provides accurate measurement of left ventricular ejection fraction (LVEF), but access to repeated scans is limited. OBJECTIVE: To develop a diagnostic model for CTRCD using echocardiographic LVEF and strain and biomarkers, with CMR as the reference standard. DESIGN, SETTING, AND PARTICIPANTS: In this prospective cohort study, patients were recruited from University of Toronto-affiliated hospitals from November 2013 to January 2019 with all cardiac imaging performed at a single tertiary care center. Women with human epidermal growth factor receptor 2 (HER2)-positive early-stage breast cancer were included. The main exclusion criterion was contraindication to CMR. A total of 160 patients were recruited, 136 of whom completed the study. EXPOSURES: Sequential therapy with anthracyclines and trastuzumab. MAIN OUTCOMES AND MEASURES: Patients underwent echocardiography, high-sensitivity troponin I (hsTnI), B-type natriuretic peptide (BNP), and CMR studies preanthracycline and postanthracycline every 3 months during and after trastuzumab therapy. Echocardiographic measures included 2-dimensional (2-D) LVEF, 3-D LVEF, peak systolic global longitudinal strain (GLS), and global circumferential strain (GCS). LVEF CTRCD was defined using the Cardiac Review and Evaluation Committee Criteria, GLS or GCS CTRCD as a greater than 15% relative change, and abnormal hsTnI and BNP as greater than 26 pg/mL and ≥ 35 pg/mL, respectively, at any follow-up point. Combinations of echocardiographic measures and biomarkers were examined to diagnose CMR CTRCD using conditional inference tree models. RESULTS: Among 136 women (mean [SD] age, 51.1 [9.2] years), CMR-identified CTRCD occurred in 37 (27%), and among those with analyzable images, in 30 of 131 (23%) by 2-D LVEF, 27 of 124 (22%) by 3-D LVEF, 53 of 126 (42%) by GLS, 61 of 123 (50%) by GCS, 32 of 136 (24%) by BNP, and 14 of 136 (10%) by hsTnI. In isolation, 3-D LVEF had greater sensitivity and specificity than 2-D LVEF for CMR CTRCD while GLS had greater sensitivity than 2-D or 3-D LVEF. Regression tree analysis identified a sequential algorithm using 3-D LVEF, GLS, and GCS for the optimal diagnosis of CTRCD (area under the receiver operating characteristic curve, 89.3%). The probability of CTRCD when results for all 3 tests were negative was 1.0%. When 3-D LVEF was replaced by 2-D LVEF in the model, the algorithm still performed well; however, its primary value was to rule out CTRCD. Biomarkers did not improve the ability to diagnose CTRCD. CONCLUSIONS AND RELEVANCE: Using CMR CTRCD as the reference standard, these data suggest that a sequential approach combining echocardiographic 3-D LVEF with 2-D GLS and 2-D GCS may provide a timely diagnosis of CTRCD during routine CTRCD surveillance with greater accuracy than using these measures individually. TRIAL REGISTRATION: ClinicalTrials.gov Identifier: NCT02306538.

Are we missing the post-operative cavity in whole breast radiotherapy?

BACKGROUND/PURPOSE: Whole breast radiotherapy (RT) following breast-conserving surgery is a standard treatment option in early-stage breast cancer patients. The whole breast RT technique targets the entire breast, traditionally identified based on breast palpation and the lumpectomy scar. The aim of this study is to evaluate dosimetry of the tumour bed (cavity) and location of recurrence in women treated with breast radiotherapy without explicit cavity delineation. MATERIALS/METHODS: 50 consecutive women previously treated with whole breast RT were retrospectively contoured to define the post-operative cavity with a 1.0 cm expansion for planning target volume (cPTV). The cavity and cPTV dosimetric coverage [volume receiving 92%(V92%) and 95%(V95%) prescription] were calculated. Cavity and cPTV location were classified as inside, at edge or outside of previous treatment fields and recurrence rates were collected. RESULTS: Forty-five (90%) women had cavities located inside the previous treatment fields (CAVin) and 5 women (10%) had cavities located outside(4) or at edge(1) of previous fields (CAVout/edge). CAVout/edge were located in extreme aspects of the breast: lateral(3); medial(1); or superior(1). Mean cavity_V92% was 91.6% vs 98.5% for CAVout/edge vs CAVin (p = 0.042). Mean cPTV_V92% was 78.7% vs 97.2% for cPTVout/edge vs cPTVin (p<0.001). At 5-year follow-up, 20% (1/5) of the CAVout/edge had 1 in-breast recurrence near the cavity (at previous field edge). Within the CAVin cohort, 11 patients were lost to follow-up and 6% (2/34) patients had in-breast recurrence. CONCLUSIONS: In patients treated with whole breast RT without cavity delineation, 10% did not have ideal dosimetric coverage of the cavity. Cavity delineation in treatment planning provides optimal tumour bed coverage for patients undergoing whole breast RT, and is of particular importance for the coverage of cavities located in the extreme margins of the breast.

Strategic Training in Transdisciplinary Radiation Science for the 21st Century (STARS21): 15-Year Evaluation of an Innovative Research Training Program.

PURPOSE: To evaluate the 15-year impact of a transdisciplinary research training program for graduate students, postdoctoral fellows, and clinical trainees focused on radiation science, entitled Strategic Training in Transdisciplinary Radiation Science for the 21st Century (STARS21) with a primary objective to build capacity in radiation research. METHODS AND MATERIALS: Alumni (n = 128) and mentors (n = 41) who participated in STARS21 between 2003 and 2018 were sent an anonymized online survey designed to evaluate the program. Twelve alumni and 7 mentors also volunteered to participate in semistructured interviews. The transcribed interviews were coded and analyzed using NVivo12-Pro software. Alumni employment and publications were assessed from program records and by web-based search queries. RESULTS: Alumni are located in 11 countries, and nearly 90% are employed in a research-oriented career and continue to publish in radiation medicine- or cancer-related fields. Of those invited, 46 alumni (36%) and 12 mentors (29%) completed the online survey. Approximately 87% of alumni valued interdisciplinary collaboration, and 80% indicated that STARS21 had encouraged them to pursue such collaborations. Alumni emphasized that STARS21 assisted their career development, and the majority of alumni and mentors would recommend STARS21 to other trainees (4.48 and 4.58, respectively; 5 = strongly agree). The time invested in the program was perceived by mentors as worthwhile for the knowledge and skills gained by trainees (4.67; 5 = strongly agree), and 64% of mentors indicated that these benefits were associated with improved trainee research productivity. From the alumni and mentor perspectives, the valuable skills acquired from STARS21 included scientific communication (85% and 83%, respectively) and networking (83% and 92%, respectively). CONCLUSIONS: STARS21 is an innovative research training program that promotes interdisciplinary collaboration in radiation medicine research, which is valued by alumni and mentor respondents. Alumni can acquire important skill sets for career development, with a large proportion of alumni currently engaged in radiation research around the world.

Clinical, Echocardiographic, and Biomarker Associations With Impaired Cardiorespiratory Fitness Early After HER2-Targeted Breast Cancer Therapy.

BACKGROUND: Cardiorespiratory fitness (CRF) is reduced in cancer survivors and predicts cardiovascular disease (CVD)-related and all-cause mortality. However, routine measurement of CRF is not always feasible. OBJECTIVES: The purpose of this study was to identify clinical, cardiac biomarker, and imaging measures associated with reduced peak oxygen consumption (VO2peak) (measure of CRF) early post-breast cancer therapy to help inform CVD risk. METHODS: Consecutive women with early-stage HER2+ breast cancer receiving anthracyclines and trastuzumab were recruited prospectively. Within 6 ± 2 weeks of trastuzumab completion, we collected clinical information, systolic/diastolic echocardiographic measures, high-sensitivity troponin I, B-type natriuretic peptide, and VO2peak using a cycle ergometer. Regression models were used to examine the association between VO2peak and clinical, imaging, and cardiac biomarkers individually and in combination. RESULTS: Among 147 patients (age 52.2 ± 9.3 years), the mean VO2peak was 19.1 ± 5.0 mL O2·kg-1·min-1 (84.2% ± 18.7% of predicted); 44% had a VO2peak below threshold for functional independence (<18 mL O2·kg-1·min-1). In multivariable analysis, absolute global longitudinal strain (GLS) (ß = 0.58; P = 0.007), age per 10 years (ß: -1.61; P = 0.001), and E/e' (measure of diastolic filling pressures) (ß = -0.45; P = 0.038) were associated with VO2peak. GLS added incremental value in explaining the variability in VO2peak. The combination of age ≥50 years, E/e' ≥7.8, and GLS <18% identified a high probability (85.7%) of compromised functional independence, whereas age <50 years, E/e' <7.8, and GLS ≥18% identified a low probability (0%). High-sensitivity troponin I and B-type natriuretic peptide were not associated with VO2peak. CONCLUSIONS: Readily available clinical measures were associated with VO2peak early post-breast cancer therapy. A combination of these parameters had good discrimination to identify patients with compromised functional independence and potentially increased future CVD risk.

Serial Cardiovascular Magnetic Resonance Strain Measurements to Identify Cardiotoxicity in Breast Cancer: Comparison With Echocardiography.

OBJECTIVES: This study sought to compare the prognostic value of cardiovascular magnetic resonance (CMR) and 2-dimensional echocardiography (2DE) derived left ventricular (LV) strain, volumes, and ejection fraction for cancer therapy-related cardiac dysfunction (CTRCD) in women with early stage breast cancer. BACKGROUND: There are limited comparative data on the association of CMR and 2DE derived strain, volumes, and LVEF with CTRCD. METHODS: A total of 125 prospectively recruited women with HER2+ early stage breast cancer receiving sequential anthracycline/trastuzumab underwent 5 serial CMR and 6 of 2DE studies before and during treatment. CMR LV volumes, left ventricular ejection fraction tagged-CMR, and feature-tracking (FT) derived global systolic longitudinal (GLS) and global circumferential strain (GCS) and 2DE-based LV volumes, function, GLS, and GCS were measured. CTRCD was defined by the cardiac review and evaluation committee criteria. RESULTS: Twenty-eight percent of patients developed CTRCD by CMR and 22% by 2DE. A 15% relative reduction in 2DE-GLS increased the CTRCD odds by 133% at subsequent follow-up, compared with 47%/50% by tagged-CMR GLS/GCS and 87% by FT-GCS. CMR and 2DE-LVEF and indexed left ventricular end-systolic volume (LVESVi) were also associated with subsequent CTRCD. The prognostic threshold change in CMR-left ventricular ejection fraction and FT strain for subsequent CTRCD was similar to the known minimum-detectable difference for these measures, whereas for tagged-CMR strain it was lower than the minimum-detectable difference; for 2DE, only the prognostic threshold for GLS was greater than the minimum-detectable difference. Of all strain methods, 2DE-GLS provided the highest increase in discriminatory value over baseline clinical risk factors for subsequent CTRCD. The combination of 2DE-left ventricular ejection fraction or LVESVi and strain provided greater increase in the area under the curve for subsequent CTRCD over clinical risk factors than CMR left ventricular ejection fraction or LVESVi and strain (18% to 22% vs. 9% to 14%). CONCLUSIONS: In women with HER2+ early stage breast cancer, changes in CMR and 2DE strain, left ventricular ejection fraction, and LVESVi were prognostic for subsequent CTRCD. When LVEF can be measured precisely by CMR, FT strain may function as an additional confirmatory prognostic measure, but with 2DE, GLS is the optimal prognostic measure. (Evaluation of Myocardial Changes During BReast Adenocarcinoma Therapy to Detect Cardiotoxicity Earlier With MRI [EMBRACE-MRI]; NCT02306538).

Utilization patterns and temporal trends of internal mammary nodal irradiation at a tertiary cancer center.

PURPOSE: The role and uptake of internal mammary nodal irradiation (IMNI) is variable. This study was designed to quantify the rates and determinants of IMNI at a tertiary cancer center. METHODS: Consecutively treated breast cancer patients receiving adjuvant locoregional radiation therapy (RT) from January 1, 2012 to December 31, 2017 were sorted by IMNI receipt, disease risk and time period of RT delivery (2012-2015 vs 2016-2017). Differences between risk categories and groups were evaluated using χ2/Fisher's and Mann-Whitney test for categorical and continuous variables, respectively. Univariable and multivariable logistic regression analysis was done to determine factors associated with IMNI receipt. RESULTS: A total of 1566 patients were eligible, with 376 in Group 1 (IMNI), and 1190 in Group 2 (no IMNI). The proportion of patients receiving IMNI increased significantly each year (p < 0.0001), and 83% of patients receiving IMNI had pT1-2/pN1 disease. On univariable analysis, younger age, lymphovascular invasion, medial/central quadrant, higher stage, PR negative, mastectomy, axillary dissection, receipt of chemotherapy and nodal positivity had higher odds of IMNI. On multivariable analysis, younger age (p = < 0.001), medial/central quadrant (p = 0.0026), PR negative (p = 0.0011), mastectomy (p = 0.0055), increasing nodal positivity (p < 0.0001) and late cohort (p = 0.001) had increased likelihood of IMNI. The use of deep-inspiration breath hold was significantly higher in those receiving IMNI (45% vs 26%, p < 0.0001), and permitted achievement of acceptable mean heart and lung doses. CONCLUSIONS: There was a significant increase in IMNI utilization after 2015. Younger age, medial/central quadrant, PR-negative and node-positive disease predicted for receipt of IMNI. Modern RT techniques permit the safe delivery of IMNI.

Patterns of Recurrence and Predictors of Survival in Breast Cancer Patients Treated with Neoadjuvant Chemotherapy, Surgery, and Radiation.

PURPOSE: Neoadjuvant chemotherapy (NAC) is standard of care for locally advanced breast cancer. There is wide variation in radiation therapy (RT) practice and limited data describing locoregional relapse (LRR) after NAC. We hypothesized a low LRR risk with modern NAC, surgery, and RT and aimed to elucidate patterns of LRR and predictors of disease-free survival (DFS) and overall survival (OS) in these patients. METHODS AND MATERIALS: Data from 416 patients with stage II/III breast cancer treated between 2008 and 2015 with NAC, surgery, and adjuvant RT were reviewed retrospectively. DFS and OS rates were calculated using the Kaplan-Meier method. The LRR rate was estimated using the cumulative incidence function, treating death as a competing risk. Multivariable survival analysis was performed using Cox regression. RESULTS: Median follow-up was 4.7 years. Most patients had cT2/3 (74%) cN1 (61%) disease and underwent mastectomy (75%) and axillary dissection (84%). Pathologic complete response (pCR) was achieved in 22% of patients. There were 27 LRRs (including 4 isolated LRRs) and 89 distant failures. Two patients developed LRR 2 months after surgery, before adjuvant RT. LRR could be mapped in 23 patients: most (20) recurred within the RT field; 1 in- and out-of-field; and 2 out-of-field. Five-year LRR, DFS, and OS were 6.4%, 77%, and 90%, respectively. On multivariable analysis, triple-negative subtype (hazard ratio [HR] 2.82; 95% confidence interval [CI], 1.78-4.47; P < .001), stage III disease (HR 1.72; 95% CI, 1.11-2.69; P = .016), and non-pCR (HR 4.76; 95% CI 2.13-10.0; P < .001) were associated with poor DFS and OS (HR 4.13 [95% CI, 2.21-7.72; P < .001]; HR 1.94 [95% CI, 1.001-3.75; P = .049]; and HR 2.38 [95% CI, 0.98-5.88; P = .055], respectively). CONCLUSIONS: Patients with breast cancer treated with modern NAC, surgery, and RT have a low 5-year LRR risk, with the majority occurring in-field. Triple-negative subtype, stage III disease, and non-pCR were associated with inferior DFS and OS.

Local control in young women with early-stage breast cancer treated with hypofractionated whole breast irradiation.

OBJECTIVE: To compare local control (LC) in young women with early-stage breast cancer (BC) treated with hypofractionated (HF) whole breast irradiation (WBI) vs conventional fractionation (CF) following breast-conserving surgery (BCS). MATERIALS AND METHODS: Women <50 years with pT1-2N0 BC following BCS treated with WBI, CF (50Gy/25 fractions) or HF (42.4Gy/16 fractions) followed by a tumor bed boost (10-16Gy/5-8 fractions) from 2009 to 2013 were identified from an institutional database. Median follow-up was 5.2 years (range 0.3-8.4). Kaplan-Meier analysis was used to estimate 5-year LC. Logistic regression identified factors associated with receipt of CF vs HF WBI. RESULTS: Of 270 eligible women, 227 (84%) were treated with HF and 43 (16%) with CF WBI. A tumor bed boost of 10 Gy/5 fractions was given in 97% of patients, 53% received adjuvant chemotherapy and 94% (225/239) with estrogen-positive disease received endocrine therapy. Median age was 45 years (range 30-49) in HF and 40 years (range 19-49) in the CF group. The 5-year LC rate was 99.3% (95% CI 97.9-100%, p = 0.495) in the HF and 97.5% (95% CI 92.8-100%) in the CF group. On univariate analysis, age ≤ 40 years or triple negative BC was associated with a decreased likelihood of receiving HF WBI. Only age remained significant on multivariate analysis [OR 2.82 (95% CI 1.45-5.48, p = 0.002)]. CONCLUSIONS: HF WBI was associated with excellent LC rates in this study cohort, comparable to CF WBI. However, CF WBI was more likely to be recommended to women <40 years.

The impact of active breathing control on internal mammary lymph node coverage and normal tissue exposure in breast cancer patients planned for left-sided postmastectomy radiation therapy.

PURPOSE: The purpose of this study was to evaluate the impact of the active breathing control (ABC) technique on IMN coverage and organs at risk in patients planned for postmastectomy radiation therapy (PMRT), with the inclusion of the internal mammary lymph nodes (IMNs). The effect of body mass index (BMI) on recorded dosimetric parameters was examined in the same patient cohort. METHODS AND MATERIALS: Fifty left-sided postmastectomy patients with breast cancer who underwent free-breathing (FB) and ABC-Elekta CT simulation scans were selected at random from an institutional breast cancer database between 2008 and 2014. The ABC plans were directly compared with FB plans from the same patient. RESULTS: The IMN planning target volume coverage met dosimetric criteria for coverage of receiving more than 90% of the prescribed dose (V90) >90%, although it decreased with ABC compared with FB (94.5% vs 98%, P < .001). Overall, ABC significantly reduced doses to all measured heart and left anterior descending coronary artery parameters, ipsilateral lung V20, and mean lung dose compared with FB (P < .001). There was no difference seen between the ABC and FB plans with respect to the dose to contralateral lung or contralateral breast. There was no correlation identified between BMI and any of the dosimetric parameters recorded from the ABC and FB plans. CONCLUSIONS: Our results suggest that ABC reduces IMN coverage in left-sided breast cancer patients planned for PMRT; however, dosimetric criteria for IMN coverage were still met, suggesting that this is not likely to be clinically significant. ABC led to significant sparing of organs at risk compared with FB conditions and was not affected by BMI. Collectively, the results support the use of ABC for breast cancer patients undergoing left-sided PMRT requiring regional nodal irradiation that includes the IMNs. Further prospective clinical studies are required to determine the impact of these results on late normal tissue effects.

Post-mastectomy Radiation: Should Subtype Factor into the Decision?

BACKGROUND: Traditional indications do not factor molecular subtype into the decision making for post-mastectomy radiation (PMRT). We sought to determine whether constructed subtype was associated with receipt of PMRT in an academic cancer center and to assess differences in locoregional recurrence (LRR) by constructed subtype. METHODS: Patients treated with mastectomy as the primary surgical therapy were identified. Univariate and stepwise multivariate logistic regression analyses examined the association between covariates and PMRT. Kaplan-Meier estimates for the time to either the earlier of LRR or last follow-up were obtained for each subtype, and Cox proportional hazards regression examined the effect of covariates on time to LRR in both univariate analyses and stepwise multivariate analysis. RESULTS: Overall, 884 patients with invasive breast cancer who underwent a primary mastectomy between January 2002 and May 2012 were included in the study. A total of 359 patients (41.6 %) received PMRT. Compared with other subtypes, triple negative (TN; HR-/HER2-) cancers were more likely to be smaller (95 % T1/T2; p = 0.02) and have a lower nodal burden (N0 65 %; p < 0.0001). On multivariate analysis, age < 50 years, lymphovascular invasion (LVI), T stage, N stage, and close or positive margins remained significantly associated with PMRT, while constructed subtype was not associated with PMRT. Compared with all other subtypes, TN had the highest rate of LRR [hazard ratio (HR) 5.70; p < 0.0001]. On multivariable analysis, TN status, LVI, and not receiving chemotherapy were significantly associated with LRR. CONCLUSIONS: Despite significant differences in LRR by constructed subtype, receptor status does not appear to be associated with the receipt of PMRT. In our series, TN cancers had the highest risk of LRR despite their relatively smaller size and limited nodal disease.

XRCC1 Polymorphism Associated With Late Toxicity After Radiation Therapy in Breast Cancer Patients.

PURPOSE: To identify single-nucleotide polymorphisms (SNPs) in oxidative stress-related genes associated with risk of late toxicities in breast cancer patients receiving radiation therapy. METHODS AND MATERIALS: Using a 2-stage design, 305 SNPs in 59 candidate genes were investigated in the discovery phase in 753 breast cancer patients from 2 prospective cohorts from Germany. The 10 most promising SNPs in 4 genes were evaluated in the replication phase in up to 1883 breast cancer patients from 6 cohorts identified through the Radiogenomics Consortium. Outcomes of interest were late skin toxicity and fibrosis of the breast, as well as an overall toxicity score (Standardized Total Average Toxicity). Multivariable logistic and linear regression models were used to assess associations between SNPs and late toxicity. A meta-analysis approach was used to summarize evidence. RESULTS: The association of a genetic variant in the base excision repair gene XRCC1, rs2682585, with normal tissue late radiation toxicity was replicated in all tested studies. In the combined analysis of discovery and replication cohorts, carrying the rare allele was associated with a significantly lower risk of skin toxicities (multivariate odds ratio 0.77, 95% confidence interval 0.61-0.96, P=.02) and a decrease in Standardized Total Average Toxicity scores (-0.08, 95% confidence interval -0.15 to -0.02, P=.016). CONCLUSIONS: Using a stage design with replication, we identified a variant allele in the base excision repair gene XRCC1 that could be used in combination with additional variants for developing a test to predict late toxicities after radiation therapy in breast cancer patients.

Identification and functional characterization of a Ku-binding motif in aprataxin polynucleotide kinase/phosphatase-like factor (APLF).

Aprataxin polynucleotide kinase/phosphatase-like factor (APLF) facilitates nonhomologous end joining (NHEJ) and associates with the core NHEJ components XRCC4-DNA ligase IV and Ku. The APLF forkhead-associated (FHA) domain directs interactions with XRCC4, but the APLF-Ku interaction has not been well characterized. Here we describe an evolutionarily conserved amino acid motif within APLF that is required for mediating the physical interaction between APLF and Ku. This APLF Ku-binding motif possesses a similarity to regions identified in other NHEJ factors, WRN and XLF, which also direct interactions with Ku. Indeed, peptides derived from the Ku-binding region of APLF, WRN, or XLF were sufficient to reconstitute the interaction with Ku in vitro. Although APLF is localized predominantly to the nucleus, it does not possess a nuclear localization signal (NLS). Interestingly, the disruption of the APLF-Ku interaction by substituting key residues in the APLF Ku-binding motif was associated with increased relocalization of APLF to the cytoplasm and reduced association with XRCC4, which was rescued by the introduction of an NLS onto APLF. When human cells stably depleted of APLF were reconstituted with APLF Ku-binding mutants, or with an APLF FHA mutant that is known to disrupt interactions with XRCC4, APLF-dependent NHEJ and the retention of APLF at sites of laser-generated DNA damage were impaired. These data suggest functional requirements for Ku and XRCC4 in APLF-dependent NHEJ and a unique role for Ku as a factor required to facilitate the nuclear retention of APLF.

The PARP3- and ATM-dependent phosphorylation of APLF facilitates DNA double-strand break repair.

APLF is a forkhead associated-containing protein with poly(ADP-ribose)-binding zinc finger (PBZ) domains, which undergoes ionizing radiation (IR)-induced and Ataxia-Telangiectasia Mutated (ATM)-dependent phosphorylation at serine-116 (Ser(116)). Here, we demonstrate that the phosphorylation of APLF at Ser(116) in human U2OS cells by ATM is dependent on poly(ADP-ribose) polymerase 3 (PARP3) levels and the APLF PBZ domains. The interaction of APLF at sites of DNA damage was diminished by the single substitution of APLF Ser(116) to alanine, and the cellular depletion or chemical inhibition of ATM or PARP3 also altered the level of accumulation of APLF at sites of laser-induced DNA damage and impaired the accumulation of Ser(116)-phosphorylated APLF at IR-induced γH2AX foci in human cells. The data further suggest that ATM and PARP3 participate in a common signalling pathway to facilitate APLF-Ser(116) phosphorylation, which, in turn, appears to be required for efficient DNA double-strand break repair kinetics and cell survival following IR. Collectively, these findings provide a more detailed understanding of the molecular pathway that leads to the phosphorylation of APLF following DNA damage and suggest that Ser(116)-APLF phosphorylation facilitates APLF-dependent double-strand break repair.

Concomitant intensity modulated boost during whole breast hypofractionated radiotherapy--a feasibility and toxicity study.

BACKGROUND: Breast cancer sensitivity to large fraction size may be enhanced using hypofractionated concomitant boost radiotherapy (CBRT), thereby shortening overall treatment time. This ethics approved, prospective single cohort feasibility study was designed to evaluate the dosimetry and toxicity of CBRT using an intensity-modulated radiotherapy (IMRT) technique, compared with a standard sequential boost technique (SBT). METHODS: Fifteen women (11 right-sided; 4 left-sided) received 42.4 Gy to the whole breast and an additional 10.08 Gy to the tumor bed in 16 daily fractions, using IMRT and standard dose constraints. Each patient was replanned with the SBT, using mixed photon-electrons. Clinical target volume (CTV), dose evaluation volume (DEV), and organs at risk (OAR) dose distributions were compared with the SBT. Toxicity and treatment times were prospectively recorded. RESULTS: All 15 CBRT plans achieved the desired CTV (V(49.9Gy) ≥ 99%) and DEV (V(49.9Gy) ≥ 95%), coverage of the boost, compared with only 10 (66.7%, p=0.03), and 12 (80%, p=0.125) SBT plans, respectively. Ipsilateral lung (p<0.0001), and heart (right-sided, p=0.001; left-sided, p=0.13) doses were lower. Grade 3 acute toxicity occurred in 1 (6.7%) patient. At 1 year, two (13.3%) additional patients had overall grade 2 late toxicity, compared with baseline. No grade 3-4 late toxicity was observed. CONCLUSIONS: CBRT using IMRT improved boost coverage and lowered OAR doses, compared with SBT. Toxicities were acceptable using a daily boost of 3.28 Gy. While resource utilization was greater, overall treatment time was reduced.

Structure and identification of ADP-ribose recognition motifs of APLF and role in the DNA damage response.

Poly(ADP-ribosyl)ation by poly(ADP-ribose) polymerases regulates the interaction of many DNA damage and repair factors with sites of DNA strand lesions. The interaction of these factors with poly(ADP-ribose) (PAR) is mediated by specific domains, including the recently identified PAR-binding zinc finger (PBZ) domain. However, the mechanism governing these interactions is unclear. To better understand the PBZ-PAR interaction, we performed a detailed examination of the representative PBZ-containing protein involved in the DNA damage response, aprataxin polynucleotide-kinase-like factor (APLF), which possesses two tandem PBZ domains. Here we present structural and biochemical studies that identify Y381/Y386 and Y423/Y428 residues in the conserved C(M/P)Y and CYR motifs within each APLF PBZ domain that are critical for the interaction with the adenine ring of ADP-ribose. Basic residues (R387 and R429 in the first and second PBZ domains, respectively) coordinate additional interactions with the phosphate backbone of ADP-ribose, suggesting that APLF binds to multiple ADP-ribose residues along PAR polymers. These C(M/P)Y and CYR motifs form a basic/hydrophobic pocket within a variant zinc finger structure and are required for APLF recruitment to sites of DNA damage in vivo.

Differential regulation of mitogen- and stress-activated protein kinase-1 and -2 (MSK1 and MSK2) by CK2 following UV radiation.

Mitogen- and stress-activated protein kinases, MSK1 and the closely related isoform MSK2, are nuclear kinases that are activated following mitogen stimulation or cellular stress, including UV radiation, by the ERK1/2 and p38 MAPK signaling cascades, respectively. However, factors that differentially regulate MSK1 and MSK2 have not been well characterized. Here we report that the CK2 protein kinase, which contributes to NF-kappaB activation following UV radiation in a p38-dependent manner, physically interacts with MSK2 but not MSK1 and that CK2 inhibition specifically impairs UV-induced MSK2 kinase activation. A putative site of CK2 phosphorylation was mapped to MSK2 residue Ser(324) and when substituted to alanine (S324A) also compromised MSK2 activity. RNA interference-mediated depletion of MSK2 in human MDA-MB-231 cells, but not MSK1 depletion, resulted in impaired UV-induced phosphorylation of NF-kappaB p65 at Ser(276) in vivo, which was restored by the ectopic expression of MSK2 but not by MSK2-S324A. Furthermore, UV radiation led to the activation of NF-kappaB-responsive gene expression in MDA-MB-231 cells and induced p65 transactivation capacity that was dependent on MSK2, MSK2 residue Ser(324), and p65-Ser(276). These results suggest that MSK1 and MSK2 are differentially regulated by CK2 during the UV response and that MSK2 is the major protein kinase responsible for the UV-induced phosphorylation of p65 at Ser(276) that positively regulates NF-kappaB activity in MDA-MB-231 cells.