Early and late skin reactions to radiotherapy for breast cancer and their correlation with radiation-induced DNA damage in lymphocytes

INTRODUCTION Radiotherapy outcomes might be further improved by a greater understanding of the individual variations in normal tissue reactions that determine tolerance. Most published studies on radiation toxicity have been performed retrospectively. Our prospective study was launched in 1996 to measure the in vitro radiosensitivity of peripheral blood lymphocytes before treatment with radical radiotherapy in patients with breast cancer, and to assess the early and the late radiation skin side effects in the same group of patients. We prospectively recruited consecutive breast cancer patients receiving radiation therapy after breast surgery. To evaluate whether early and late side effects of radiotherapy can be predicted by the assay, a study was conducted of the association between the results of in vitro radiosensitivity tests and acute and late adverse radiation effects. METHODS Intrinsic molecular radiosensitivity was measured by using an initial radiation-induced DNA damage assay on lymphocytes obtained from breast cancer patients before radiotherapy. Acute reactions were assessed in 108 of these patients on the last treatment day. Late morbidity was assessed after 7 years of follow-up in some of these patients. The Radiation Therapy Oncology Group (RTOG) morbidity score system was used for both assessments. RESULTS Radiosensitivity values obtained using the in vitro test showed no relation with the acute or late adverse skin reactions observed. There was no evidence of a relation between acute and late normal tissue reactions assessed in the same patients. A positive relation was found between the treatment volume and both early and late side effects. CONCLUSION After radiation treatment, a number of cells containing major changes can have a long survival and disappear very slowly, becoming a chronic focus of immunological system stimulation. This stimulation can produce, in a stochastic manner, late radiation-related adverse effects of varying severity. Further research is warranted to identify the major determinants of normal tissue radiation response to make it possible to individualize treatments and improve the outcome of radiotherapy in cancer patients.

Automatic classification of breast density

A recent trend in digital mammography is computer-aided diagnosis systems, which are computerised tools designed to assist radiologists. Most of these systems are used for the automatic detection of abnormalities. However, recent studies have shown that their sensitivity is significantly decreased as the density of the breast increases. This dependence is method specific. In this paper we propose a new approach to the classification of mammographic images according to their breast parenchymal density. Our classification uses information extracted from segmentation results and is based on the underlying breast tissue texture. Classification performance was based on a large set of digitised mammograms. Evaluation involves different classifiers and uses a leave-one-out methodology. Results demonstrate the feasibility of estimating breast density using image processing and analysis techniques

Combined low initial DNA damage and high radiation-induced apoptosis confers clinical resistance to long-term toxicity in breast cancer patients treated with high-dose radiotherapy

BACKGROUND. Either higher levels of initial DNA damage or lower levels of radiation-induced apoptosis in peripheral blood lymphocytes have been associated to increased risk for develop late radiation-induced toxicity. It has been recently published that these two predictive tests are inversely related. The aim of the present study was to investigate the combined role of both tests in relation to clinical radiation-induced toxicity in a set of breast cancer patients treated with high dose hyperfractionated radical radiotherapy. METHODS. Peripheral blood lymphocytes were taken from 26 consecutive patients with locally advanced breast carcinoma treated with high-dose hyperfractioned radical radiotherapy. Acute and late cutaneous and subcutaneous toxicity was evaluated using the Radiation Therapy Oncology Group morbidity scoring schema. The mean follow-up of survivors (n = 13) was 197.23 months. Radiosensitivity of lymphocytes was quantified as the initial number of DNA double-strand breaks induced per Gy and per DNA unit (200 Mbp). Radiation-induced apoptosis (RIA) at 1, 2 and 8 Gy was measured by flow cytometry using annexin V/propidium iodide. RESULTS. Mean DSB/Gy/DNA unit obtained was 1.70 ± 0.83 (range 0.63-4.08; median, 1.46). Radiation-induced apoptosis increased with radiation dose (median 12.36, 17.79 and 24.83 for 1, 2, and 8 Gy respectively). We observed that those "expected resistant patients" (DSB values lower than 1.78 DSB/Gy per 200 Mbp and RIA values over 9.58, 14.40 or 24.83 for 1, 2 and 8 Gy respectively) were at low risk of suffer severe subcutaneous late toxicity (HR 0.223, 95%CI 0.073-0.678, P = 0.008; HR 0.206, 95%CI 0.063-0.677, P = 0.009; HR 0.239, 95%CI 0.062-0.929, P = 0.039, for RIA at 1, 2 and 8 Gy respectively) in multivariate analysis. CONCLUSIONS. A radiation-resistant profile is proposed, where those patients who presented lower levels of initial DNA damage and higher levels of radiation induced apoptosis were at low risk of suffer severe subcutaneous late toxicity after clinical treatment at high radiation doses in our series. However, due to the small sample size, other prospective studies with higher number of patients are needed to validate these results.

Cannabinoids reduce ErbB2-driven breast cancer progression through Akt inhibition.

BACKGROUND ErbB2-positive breast cancer is characterized by highly aggressive phenotypes and reduced responsiveness to standard therapies. Although specific ErbB2-targeted therapies have been designed, only a small percentage of patients respond to these treatments and most of them eventually relapse. The existence of this population of particularly aggressive and non-responding or relapsing patients urges the search for novel therapies. The purpose of this study was to determine whether cannabinoids might constitute a new therapeutic tool for the treatment of ErbB2-positive breast tumors. We analyzed their antitumor potential in a well established and clinically relevant model of ErbB2-driven metastatic breast cancer: the MMTV-neu mouse. We also analyzed the expression of cannabinoid targets in a series of 87 human breast tumors. RESULTS Our results show that both Delta9-tetrahydrocannabinol, the most abundant and potent cannabinoid in marijuana, and JWH-133, a non-psychotropic CB2 receptor-selective agonist, reduce tumor growth, tumor number, and the amount/severity of lung metastases in MMTV-neu mice. Histological analyses of the tumors revealed that cannabinoids inhibit cancer cell proliferation, induce cancer cell apoptosis, and impair tumor angiogenesis. Cannabinoid antitumoral action relies, at least partially, on the inhibition of the pro-tumorigenic Akt pathway. We also found that 91% of ErbB2-positive tumors express the non-psychotropic cannabinoid receptor CB2. CONCLUSIONS Taken together, these results provide a strong preclinical evidence for the use of cannabinoid-based therapies for the management of ErbB2-positive breast cancer.

Functional characterization of E- and P-cadherin in invasive breast cancer cells

BACKGROUND Alterations in the cadherin-catenin adhesion complexes are involved in tumor initiation, progression and metastasis. However, the functional implication of distinct cadherin types in breast cancer biology is still poorly understood. METHODS To compare the functional role of E-cadherin and P-cadherin in invasive breast cancer, we stably transfected these molecules into the MDA-MB-231 cell line, and investigated their effects on motility, invasion and gene expression regulation. RESULTS Expression of either E- and P-cadherin significantly increased cell aggregation and induced a switch from fibroblastic to epithelial morphology. Although expression of these cadherins did not completely reverse the mesenchymal phenotype of MDA-MB-231 cells, both E- and P-cadherin decreased fibroblast-like migration and invasion through extracellular matrix in a similar way. Moreover, microarray gene expression analysis of MDA-MB-231 cells after expression of E- and P-cadherins revealed that these molecules can activate signaling pathways leading to significant changes in gene expression. Although the expression patterns induced by E- and P-cadherin showed more similarities than differences, 40 genes were differentially modified by the expression of either cadherin type. CONCLUSION E- and P-cadherin have similar functional consequences on the phenotype and invasive behavior of MDA-MB-231 cells. Moreover, we demonstrate for the first time that these cadherins can induce both common and specific gene expression programs on invasive breast cancer cells. Importantly, these identified genes are potential targets for future studies on the functional consequences of altered cadherin expression in human breast cancer.

Caracterització i diagnosi de masses en imatges de mamografia

Els objectius del projecte es divideixen en tres blocs: Primerament, realitzar unasegmentació automàtica del contorn d'una imatge on hi ha una massa central. Tot seguit, a partir del contorn trobat, caracteritzar la massa. I finalment, utilitzant les característiques anteriors classificar la massa en benigne o maligne. En el projecte s'utilitza el Matlab com a eina de programació. Concretament les funcions enfocades al processat de imatges del toolbox de Image processing (propi de Matlab) i els classificadors de la PRTools de la Delft University of Technology

Retreatment with trastuzumab in Her2-positive metastatic breast cancer patients: a clinical study

Objective: To study the benefit of trastuzumab in monotherapy or combined with different chemotherapeutic agents in the treatment for Her2+ metastatic breast cancer (MBC) patients after progression on prior trastuzumab therapy.

Lack of evidence for KRAS oncogenic mutations in triple-negative breast cancer.

BACKGROUND Mutational analysis of the KRAS gene has recently been established as a complementary in vitro diagnostic tool for the identification of patients with colorectal cancer who will not benefit from anti-epidermal growth factor receptor (EGFR) therapies. Assessment of the mutation status of KRAS might also be of potential relevance in other EGFR-overexpressing tumors, such as those occurring in breast cancer. Although KRAS is mutated in only a minor fraction of breast tumors (5%), about 60% of the basal-like subtype express EGFR and, therefore could be targeted by EGFR inhibitors. We aimed to study the mutation frequency of KRAS in that subtype of breast tumors to provide a molecular basis for the evaluation of anti-EGFR therapies. METHODS Total, genomic DNA was obtained from a group of 35 formalin-fixed paraffin-embedded, triple-negative breast tumor samples. Among these, 77.1% (27/35) were defined as basal-like by immunostaining specific for the established surrogate markers cytokeratin (CK) 5/6 and/or EGFR. KRAS mutational status was determined in the purified DNA samples by Real Time (RT)-PCR using primers specific for the detection of wild-type KRAS or the following seven oncogenic somatic mutations: Gly12Ala, Gly12Asp, Gly12Arg, Gly12Cys, Gly12Ser, Gly12Val and Gly13Asp. RESULTS We found no evidence of KRAS oncogenic mutations in all analyzed tumors. CONCLUSIONS This study indicates that KRAS mutations are very infrequent in triple-negative breast tumors and that EGFR inhibitors may be of potential benefit in the treatment of basal-like breast tumors, which overexpress EGFR in about 60% of all cases.

Human pregnane X receptor is expressed in breast carcinomas, potential heterodimers formation between hPXR and RXR-alpha.

BACKGROUND The human pregnane X receptor (hPXR) is an orphan nuclear receptor that induces transcription of response elements present in steroid-inducible cytochrome P-450 gene promoters. This activation requires the participation of retinoid X receptors (RXRs), needed partners of hPXR to form heterodimers. We have investigated the expression of hPXR and RXRs in normal, premalignant, and malignant breast tissues, in order to determine whether their expression profile in localized infiltrative breast cancer is associated with an increased risk of recurrent disease. METHODS Breast samples from 99 patients including benign breast diseases, in situ and infiltrative carcinomas were processed for immunohistochemistry and Western-blot analysis. RESULTS Cancer cells from patients that developed recurrent disease showed a high cytoplasmic location of both hPXR isoforms. Only the infiltrative carcinomas that relapsed before 48 months showed nuclear location of hPXR isoform 2. This location was associated with the nuclear immunoexpression of RXR-alpha. CONCLUSION Breast cancer cells can express both variants 1 and 2 of hPXR. Infiltrative carcinomas that recurred showed a nuclear location of both hPXR and RXR-alpha; therefore, the overexpression and the subcellular location changes of hPXR could be considered as a potential new prognostic indicator.

Randomized phase 3 trial of fluorouracil, epirubicin, and cyclophosphamide alone or followed by Paclitaxel for early breast cancer.

BACKGROUND Taxanes are among the most active drugs for the treatment of metastatic breast cancer, and, as a consequence, they have also been studied in the adjuvant setting. METHODS After breast cancer surgery, women with lymph node-positive disease were randomly assigned to treatment with fluorouracil, epirubicin, and cyclophosphamide (FEC) or with FEC followed by weekly paclitaxel (FEC-P). The primary endpoint of study-5-year disease-free survival (DFS)-was assessed by Kaplan-Meier analysis. Secondary endpoints included overall survival and analysis of the prognostic and predictive value of clinical and molecular (hormone receptors by immunohistochemistry and HER2 by fluorescence in situ hybridization) markers. Associations and interactions were assessed with a multivariable Cox proportional hazards model for DFS for the following covariates: age, menopausal status, tumor size, lymph node status, type of chemotherapy, tumor size, positive lymph nodes, HER2 status, and hormone receptor status. All statistical tests were two-sided. RESULTS Among the 1246 eligible patients, estimated rates of DFS at 5 years were 78.5% in the FEC-P arm and 72.1% in the FEC arm (difference = 6.4%, 95% confidence interval [CI] = 1.6% to 11.2%; P = .006). FEC-P treatment was associated with a 23% reduction in the risk of relapse compared with FEC treatment (146 relapses in the 614 patients in the FEC-P arm vs 193 relapses in the 632 patients in the FEC arm, hazard ratio [HR] = 0.77, 95% CI = 0.62 to 0.95; P = .022) and a 22% reduction in the risk of death (73 and 95 deaths, respectively, HR = 0.78, 95% CI = 0.57 to 1.06; P = .110). Among the 928 patients for whom tumor samples were centrally analyzed, type of chemotherapy (FEC vs FEC-P) (P = .017), number of involved axillary lymph nodes (P < .001), tumor size (P = .020), hormone receptor status (P = .004), and HER2 status (P = .006) were all associated with DFS. We found no statistically significant interaction between HER2 status and paclitaxel treatment or between hormone receptor status and paclitaxel treatment. CONCLUSIONS Among patients with operable breast cancer, FEC-P treatment statistically significantly reduced the risk of relapse compared with FEC as adjuvant therapy.

Adiposity, hormone replacement therapy use and breast cancer risk by age and hormone receptor status: a large prospective cohort study.

INTRODUCTION Associations of hormone-receptor positive breast cancer with excess adiposity are reasonably well characterized; however, uncertainty remains regarding the association of body mass index (BMI) with hormone-receptor negative malignancies, and possible interactions by hormone replacement therapy (HRT) use. METHODS Within the European EPIC cohort, Cox proportional hazards models were used to describe the relationship of BMI, waist and hip circumferences with risk of estrogen-receptor (ER) negative and progesterone-receptor (PR) negative (n = 1,021) and ER+PR+ (n = 3,586) breast tumors within five-year age bands. Among postmenopausal women, the joint effects of BMI and HRT use were analyzed. RESULTS For risk of ER-PR- tumors, there was no association of BMI across the age bands. However, when analyses were restricted to postmenopausal HRT never users, a positive risk association with BMI (third versus first tertile HR = 1.47 (1.01 to 2.15)) was observed. BMI was inversely associated with ER+PR+ tumors among women aged ≤49 years (per 5 kg/m2 increase, HR = 0.79 (95%CI 0.68 to 0.91)), and positively associated with risk among women ≥65 years (HR = 1.25 (1.16 to 1.34)). Adjusting for BMI, waist and hip circumferences showed no further associations with risks of breast cancer subtypes. Current use of HRT was significantly associated with an increased risk of receptor-negative (HRT current use compared to HRT never use HR: 1.30 (1.05 to 1.62)) and positive tumors (HR: 1.74 (1.56 to 1.95)), although this risk increase was weaker for ER-PR- disease (Phet = 0.035). The association of HRT was significantly stronger in the leaner women (BMI ≤22.5 kg/m2) than for more overweight women (BMI ≥25.9 kg/m2) for, both, ER-PR- (HR: 1.74 (1.15 to 2.63)) and ER+PR+ (HR: 2.33 (1.84 to 2.92)) breast cancer and was not restricted to any particular HRT regime. CONCLUSIONS An elevated BMI may be positively associated with risk of ER-PR- tumors among postmenopausal women who never used HRT. Furthermore, postmenopausal HRT users were at an increased risk of ER-PR- as well as ER+PR+ tumors, especially among leaner women. For hormone-receptor positive tumors, but not for hormone-receptor negative tumors, our study confirms an inverse association of risk with BMI among young women of premenopausal age. Our data provide evidence for a possible role of sex hormones in the etiology of hormone-receptor negative tumors.

Effectiveness of core stability exercises and recovery myofascial release massage on fatigue in breast cancer survivors: a randomized controlled clinical trial.

The purpose of the present paper was to evaluate the effects of an 8-week multimodal program focused on core stability exercises and recovery massage with DVD support for a 6-month period in physical and psychological outcomes in breast cancer survivors. A randomized controlled clinical trial was performed. Seventy-eight (n = 78) breast cancer survivors were assigned to experimental (core stability exercises plus massage-myofascial release) and control (usual health care) groups. The intervention period was 8 weeks. Mood state, fatigue, trunk curl endurance, and leg strength were determined at baseline, after the last treatment session, and at 6 months of followup. Immediately after treatment and at 6 months, fatigue, mood state, trunk curl endurance, and leg strength exhibited greater improvement within the experimental group compared to placebo group. This paper showed that a multimodal program focused on core stability exercises and massage reduced fatigue, tension, depression, and improved vigor and muscle strength after intervention and 6 months after discharge.

A woman with reddish nodule on the skin of the breast.

A 54-year-old woman presented a peri-areolar nodule located in the skin of the right breast. Clinical examination showed a 6 x 5 cm exophytic, lobed, ulcerated, and bleeding nodule. The patient reported that the tumor had grown gradually over a period of 3 months. The patient had been diagnosed 8 years prior to presentation with infiltrating ductal carcinoma of the right breast (pT2NO). This tumor was treated with partial mastectomy (conservative surgery) and lymph node dissection, then subsequently received 30 tangent field radiotherapy sessions to the breast for a total dose of 45 Gy. The rest of her cutaneous exam was normal. There was no family history of any similar tumor.

MicroRNA-200 family modulation in distinct breast cancer phenotypes.

The epithelial to mesenchymal transition (EMT) contributes to tumor invasion and metastasis in a variety of cancer types. In human breast cancer, gene expression studies have determined that basal-B/claudin-low and metaplastic cancers exhibit EMT-related characteristics, but the molecular mechanisms underlying this observation are unknown. As the family of miR-200 microRNAs has been shown to regulate EMT in normal tissues and cancer, here we evaluated whether the expression of the miR-200 family (miR-200f) and their epigenetic state correlate with EMT features in human breast carcinomas. We analyzed by qRT-PCR the expression of miR-200f members and various EMT-transcriptional inducers in a series of 70 breast cancers comprising an array of phenotypic subtypes: estrogen receptor positive (ER+), HER2 positive (HER2+), and triple negative (TN), including a subset of metaplastic breast carcinomas (MBCs) with sarcomatous (homologous or heterologous) differentiation. No MBCs with squamous differentiation were included. The DNA methylation status of miR-200f loci in tumor samples were inspected using Sequenom MassArray® MALDI-TOF platform. We also used two non-tumorigenic breast basal cell lines that spontaneously undergo EMT to study the modulation of miR-200f expression during EMT in vitro. We demonstrate that miR-200f is strongly decreased in MBCs compared with other cancer types. TN and HER2+ breast cancers also exhibited lower miR-200f expression than ER+ tumors. Significantly, the decreased miR-200f expression found in MBCs is accompanied by an increase in the expression levels of EMT-transcriptional inducers, and hypermethylation of the miR-200c-141 locus. Similar to tumor samples, we demonstrated that downregulation of miR-200f and hypermethylation of the miR-200c-141 locus, together with upregulation of EMT-transcriptional inducers also occur in an in vitro cellular model of spontaneous EMT. Thus, the expression and methylation status of miR-200f could be used as hypothetical biomarkers to assess the occurrence of EMT in breast cancer.

Obesity and survival in operable breast cancer patients treated with adjuvant anthracyclines and taxanes according to pathological subtypes: a pooled analysis.

INTRODUCTION Obesity is an unfavorable prognostic factor in breast cancer (BC) patients regardless of menopausal status and treatment received. However, the association between obesity and survival outcome by pathological subtype requires further clarification. METHODS We performed a retrospective analysis including 5,683 operable BC patients enrolled in four randomized clinical trials (GEICAM/9906, GEICAM/9805, GEICAM/2003-02, and BCIRG 001) evaluating anthracyclines and taxanes as adjuvant treatments. Our primary aim was to assess the prognostic effect of body mass index (BMI) on disease recurrence, breast cancer mortality (BCM), and overall mortality (OM). A secondary aim was to detect differences of such prognostic effects by subtype. RESULTS Multivariate survival analyses adjusting for age, tumor size, nodal status, menopausal status, surgery type, histological grade, hormone receptor status, human epidermal growth factor receptor 2 (HER2) status, chemotherapy regimen, and under-treatment showed that obese patients (BMI 30.0 to 34.9) had similar prognoses to that of patients with a BMI < 25 (reference group) in terms of recurrence (Hazard Ratio [HR] = 1.08, 95% Confidence Interval [CI] = 0.90 to 1.30), BCM (HR = 1.02, 0.81 to 1.29), and OM (HR = 0.97, 0.78 to 1.19). Patients with severe obesity (BMI ≥ 35) had a significantly increased risk of recurrence (HR = 1.26, 1.00 to 1.59, P = 0.048), BCM (HR = 1.32, 1.00 to 1.74, P = 0.050), and OM (HR = 1.35, 1.06 to 1.71, P = 0.016) compared to our reference group. The prognostic effect of severe obesity did not vary by subtype. CONCLUSIONS Severely obese patients treated with anthracyclines and taxanes present a worse prognosis regarding recurrence, BCM, and OM than patients with BMI < 25. The magnitude of the harmful effect of BMI on survival-related outcomes was similar across subtypes.