Software BreastAnalyser for the semi-automatic analysis of breast cancer immunohistochemical images.

Breast cancer is the most diagnosed cancer worldwide and represents the fifth cause of cancer mortality globally. It is a highly heterogeneous disease, that comprises various molecular subtypes, often diagnosed by immunohistochemistry. This technique is widely employed in basic, translational and pathological anatomy research, where it can support the oncological diagnosis, therapeutic decisions and biomarker discovery. Nevertheless, its evaluation is often qualitative, raising the need for accurate quantitation methodologies. We present the software BreastAnalyser, a valuable and reliable tool to automatically measure the area of 3,3'-diaminobenzidine tetrahydrocholoride (DAB)-brown-stained proteins detected by immunohistochemistry. BreastAnalyser also automatically counts cell nuclei and classifies them according to their DAB-brown-staining level. This is performed using sophisticated segmentation algorithms that consider intrinsic image variability and save image normalization time. BreastAnalyser has a clean, friendly and intuitive interface that allows to supervise the quantitations performed by the user, to annotate images and to unify the experts' criteria. BreastAnalyser was validated in representative human breast cancer immunohistochemistry images detecting various antigens. According to the automatic processing, the DAB-brown area was almost perfectly recognized, being the average difference between true and computer DAB-brown percentage lower than 0.7 points for all sets. The detection of nuclei allowed proper cell density relativization of the brown signal for comparison purposes between the different patients. BreastAnalyser obtained a score of 85.5 using the system usability scale questionnaire, which means that the tool is perceived as excellent by the experts. In the biomedical context, the connexin43 (Cx43) protein was found to be significantly downregulated in human core needle invasive breast cancer samples when compared to normal breast, with a trend to decrease as the subtype malignancy increased. Higher Cx43 protein levels were significantly associated to lower cancer recurrence risk in Oncotype DX-tested luminal B HER2- breast cancer tissues. BreastAnalyser and the annotated images are publically available https://citius.usc.es/transferencia/software/breastanalyser for research purposes.

Evaluation of MRI accuracy after primary systemic therapy in breast cancer patients considering tumor biology: optimizing the surgical planning.

Background We analyzed the accuracy of magnetic resonance imaging (MRI) after primary systemic therapy (PST) according to tumor subtype. Patients and methods Two-hundred and four breast cancer patients treated with PST were studied. MRI findings after PST were compared with pathologic findings, and results were stratified based on tumor subtype. Results Of the two-hundred and four breast cancer patients, eighty-four (41.2%) achieved a pathologic complete response (pCR) in the breast. The MRI accuracy for predicting pCR was highest in triple-negative (TN) and HER2-positive (non-luminal) breast cancer (83.9 and 80.9%, respectively). The mean size discrepancy between MRI-measured and pathologic residual tumor size was lowest in TN breast cancer and highest in luminal B-like (HER2-negative) breast cancer (0.45cm vs. 0.98 cm, respectively; p = 0.003). After breast conserving surgery (BCS), we found a lower rate of positive margins in TN breast cancer and a higher rate of positive margins in luminal B-like (HER2-negative) breast cancer (2.4% vs. 23.6%, respectively). Conclusions If tumor response after PST is assessed by MRI, tumor subtype should be considered when BCS is planned. The accuracy of MRI is highest in TN breast cancer.

Primary systemic therapy in HER2-positive operable breast cancer using trastuzumab and chemotherapy: efficacy data, cardiotoxicity and long-term follow-up in 142 patients diagnosed from 2005 to 2016 at a single institution.

OBJECTIVE: The aim of this study was to evaluate the efficacy, cardiotoxicity profile and long-term benefits of neoadjuvant therapy in human epidermal growth factor receptor 2-positive operable breast cancer patients. PATIENTS AND METHODS: A total of 142 patients diagnosed from 2005 to 2016 were included in the study. The treatment consisted of a sequential regimen of taxanes and anthracyclines plus trastuzumab. The clinical and pathological responses were evaluated and correlated with clinical and biological factors. The cardiotoxicity profile and long-term benefits were analyzed. RESULTS: The median age was 49 years, and 4%, 69% and 27% of patients had stage I, II and III breast cancer, respectively, while 10% had inflammatory breast cancer at diagnosis. Hormone receptor (HR) status was negative in 43%, and 62% had grade III breast cancer. The clinical complete response rate was 49% and 63% as assessed using ultrasound and magnetic resonance imaging, respectively, and this allowed a high rate of conservative surgery (66%). The pathological complete response (pCR) rate was 52%, and it was higher in HR-negative (64%) patients than in HR-positive (41%) patients and in grade III breast cancer (53%) patients than in grade I-II breast cancer (45%) patients. Patients who achieved pCR had longer disease-free survival and a trend toward improved overall survival. A total of 2% of patients showed a 10% decrease in left ventricular ejection fraction to <50% during treatment. All patients except one recovered after discontinuation of trastuzumab. CONCLUSION: A sequential regimen of taxanes and anthracyclines plus trastuzumab was effective, with high pCR rates and long-term benefit, and had a very good cardiotoxicity profile.

The C-terminal domain of connexin43 modulates cartilage structure via chondrocyte phenotypic changes.

Chondrocytes in cartilage and bone cells population express connexin43 (Cx43) and gap junction intercellular communication (GJIC) is essential to synchronize cells for coordinated electrical, mechanical, metabolic and chemical communication in both tissues. Reduced Cx43 connectivity decreases chondrocyte differentiation and defective Cx43 causes skeletal defects. The carboxy terminal domain (CTD) of Cx43 is located in the cytoplasmic side and is key for protein functions. Here we demonstrated that chondrocytes from the CTD-deficient mice, K258stop/Cx43KO and K258stop/K258stop, have reduced GJIC, increased rates of proliferation and reduced expression of collagen type II and proteoglycans. We observed that CTD-truncated mice were significantly smaller in size. Together these results demonstrated that the deletion of the CTD negatively impacts cartilage structure and normal chondrocyte phenotype. These findings suggest that the proteolytic cleavage of the CTD under pathological conditions, such as under the activation of metalloproteinases during tissue injury or inflammation, may account for the deleterious effects of Cx43 in cartilage and bone disorders such as osteoarthritis.

Risk factors for positive margins in conservative surgery for breast cancer after neoadjuvant chemotherapy. / Factores de riesgo de afectación de los márgenes quirúrgicos en la cirugía conservadora del cáncer de mama tras quimioterapia neoadyuvante.

BACKGROUND: Breast conservative surgery after neoadjuvant chemotherapy intends to remove any residual tumor with negative margins. The purpose of this study was to analyze the preoperative clinical-pathological factors influencing the margin status after conservative surgery in breast cancer patients receiving neoadjuvant chemotherapy. METHODS: A retrospective study of 91 breast cancer patients undergoing neoadjuvant chemotherapy (92 breast lesions) during the period 2006 to 2013. A Cox regression analysis to identify baseline tumor characteristics associated with positive margins after breast conservative surgery was performed. RESULTS: Of all cases, 71 tumors were initially treated with conservative surgery after neoadjuvant chemotherapy. Pathologic exam revealed positive margins in 16 of the 71 cases (22.5%). The incidence of positive margins was significantly higher in cancers with initial size >5cm (P=.021), in cancers with low tumor grade (P=.031), and in patients with hormone receptor-positive cancer (P=.006). After a median follow-up of 45.2 months, 7 patients of the 71 treated with conservative surgery had disease recurrence (9.8%). There was no significant difference in terms of disease-free survival according to the margin status (P=.596). CONCLUSIONS: A baseline tumor size >5cm, low tumor grade and hormone receptor-positive status increase the risk for surgical margin involvement in breast conservative surgery after neoadjuvant chemotherapy.

Diagnostic accuracy of MRI to evaluate tumour response and residual tumour size after neoadjuvant chemotherapy in breast cancer patients.

BACKGROUND: The aim, of the study was to estimate the accuracy of magnetic resonance imaging (MRI) in assessing residual disease in breast cancer patients receiving neoadjuvant chemotherapy (NAC) and to identify the clinico-pathological factors that affect the diagnostic accuracy of breast MRI to determine residual tumour size following NAC. PATIENTS AND METHODS: 91 breast cancer patients undergoing NAC (92 breast lesions) were included in the study. Breast MRI was performed at baseline and after completion of NAC. Treatment response was evaluated by MRI and histopathological examination to investigate the ability of MRI to predict tumour response. Residual tumour size was measured on post-treatment MRI and compared with pathology in 89 lesions. Clinicopathological factors were analyzed to compare MRI-pathologic size differences. RESULTS: The overall sensitivity, specificity, positive predictive value, negative predictive value, and accuracy for diagnosing invasive residual disease by using MRI were 75.00%, 78.57%, 88.89%, 57.89%, and 76.09% respectively. The Pearson's correlation coefficient (r) between tumour sizes determined by MRI and pathology was r = 0.648 (p < 0.001). The size discrepancy was significantly lower in cancers with initial MRI size ≤ 5 cm (p = 0.050), in cancers with high tumour grade (p < 0.001), and in patients with hormonal receptor-negative cancer (p = 0.033). CONCLUSIONS: MRI is an accurate tool for evaluating tumour response after NAC. The accuracy of MRI in estimating residual tumour size varies with the baseline MRI tumour size, the tumour grade and the hormonal receptor status.

Proteomic Analysis of Connexin 43 Reveals Novel Interactors Related to Osteoarthritis.

We have previously reported that articular chondrocytes in tissue contain long cytoplasmic arms that physically connect two distant cells. Cell-to-cell communication occurs through connexin channels termed Gap Junction (GJ) channels, which achieve direct cellular communication by allowing the intercellular exchange of ions, small RNAs, nutrients, and second messengers. The Cx43 protein is overexpressed in several human diseases and inflammation processes and in articular cartilage from patients with osteoarthritis (OA). An increase in the level of Cx43 is known to alter gene expression, cell signaling, growth, and cell proliferation. The interaction of proteins with the C-terminal tail of connexin 43 (Cx43) directly modulates GJ-dependent and -independent functions. Here, we describe the isolation of Cx43 complexes using mild extraction conditions and immunoaffinity purification. Cx43 complexes were extracted from human primary articular chondrocytes isolated from healthy donors and patients with OA. The proteomic content of the native complexes was determined using LC-MS/MS, and protein associations with Cx43 were validated using Western blot and immunolocalization experiments. We identified >100 Cx43-associated proteins including previously uncharacterized proteins related to nucleolar functions, RNA transport, and translation. We also identified several proteins involved in human diseases, cartilage structure, and OA as novel functional Cx43 interactors, which emphasized the importance of Cx43 in the normal physiology and structural and functional integrity of chondrocytes and articular cartilage. Gene Ontology (GO) terms of the proteins identified in the OA samples showed an enrichment of Cx43-interactors related to cell adhesion, calmodulin binding, the nucleolus, and the cytoskeleton in OA samples compared with healthy samples. However, the mitochondrial proteins SOD2 and ATP5J2 were identified only in samples from healthy donors. The identification of Cx43 interactors will provide clues to the functions of Cx43 in human cells and its roles in the development of several diseases, including OA.

[Proposal for a new multidisciplinary therapeutic strategy in the breast cancer patient with sentinel lymph node metastasis]. / Propuesta de una nueva estrategia terapéutica multidisciplinar en la mujer con cáncer de mama y afectación del ganglio centinela.

Sentinel lymph node (SLN) biopsy is the standard of practice for assessing axillary spread in clinically node-negative breast cancer patients. On the other hand, axillary lymph node dissection (ALND) is the ideal procedure for patients with SLN metastasis. Different studies over the last few years have suggested that some patients with positive SLN can be treated without ALND. This article presents a literature review carried out by our multidisciplinary group and its strategy for avoiding routine ALND in women with SLN metastases. In this new strategy ALND should not be performed on women with T1 tumours, with 1-2 positive SLN and undergoing breast conservative surgery. On the other hand, ALND would be indicated in those patients with three or more positive SLN, presence of extracapsular invasion, mastectomised women and triple negative subtype or HER2+ tumours that have not received biological treatment with antibodies.

[Informing the surgical patient. Reflections on the basic law of patient autonomy]. / El proceso informativo en el paciente quirúrgico. Reflexiones sobre la ley básica reguladora de la autonomía de los pacientes.

The basic law of patient autonomy represents a new point of reference for surgeons in terms of their obligation to inform the patient and obtain consent. The main contribution of this new law is its explicit recognition of the patients autonomy in decision-making in medicine, leading to new obligations among health professionals in matters of information and consent. Nevertheless, because of the special characteristics of surgery, these legal principles need to be contextualized within the reality of surgical practice in order to harmonize rights, obligations and ethical values. In light of this new law, the present article aims to provide a reflection on the complexities of providing information in surgical practice with a view to identifying areas of conflict and making recommendations for their resolution.

[Informed consent in the surgical patient. Reflections on the basic law of patient autonomy]. / El consentimiento informado en el paciente quirúrgico. Reflexiones sobre la Ley Básica Reguladora de la Autonomía de los Pacientes.

Informed consent is the final step in informing the surgical patient. The basic law of patient autonomy specifies the conditions in which patient authorization should be given with the aim of guaranteeing that a free decision is made. However, surgeons are sometimes faced with clinical situations in which it is difficult to obtain informed consent, whether because the patient is unable to provide consent or because of the need to intervene in a life-threatening situation, or because the informed consent document must specify the risks and consequences of the intervention. The present article aims to analyze what happens when the patient is unable to provide consent ("consent by representation") and the characteristics of the informed consent document according to the stipulations of the basic law of patient autonomy.