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Biomedical subjects

Viviana Galimberti

Publications and source records attributed to Viviana Galimberti.

At least 19 recordsLinked to original sources

IART: intraoperative avidination for radionuclide treatment. A new way of partial breast irradiation.

A new procedure, known as Intraoperative Avidination for Radionuclide Therapy (IART), is described in breast cancer patients. In this paper, we provide proof of the principle that intraoperative injection of avidin in the tumour bed after quadrantectomy allows homing in of intravenously (IV) administered radioactive biotin to the target site. This approach of targeted therapy consists of two steps: (i) "avidination" of the anatomical area of the tumour with avidin injected by the surgeon, into and around the tumour bed; (ii) targeting the anatomical area of the tumour by IV injection of radiolabelled biotin. The scintigraphic images demonstrated fast and stable uptake of labelled biotin at the site of operated breast. The radiation dose released to the index quadrant was more than 5 Gy/GBq, consistent with a boost of 20 Gy for an activity of 3.7 GBq 90Y-biotin (100mCi). A further large clinical trial facing IART in combination with reduced external-beam radiotherapy is, in our opinion, fully justified.

Adult↗

Ipsilateral breast tumor reappearance in patients treated with conservative surgery after primary chemotherapy. The role of surgical margins on outcome.

BACKGROUND AND OBJECTIVES: To evaluate the ipsilateral breast tumor reappearance (IBTR) rate after breast conservative surgery (BCS) following primary chemotherapy (PC) and to assess whether positive margins affects IBTR rate and overall survival (OS). METHODS: Three hundred nine women candidates for mastectomy received PC before surgery. One hundred ninety-five patients (63.1%) underwent BCS and 114 patients (36.9%) a modified radical mastectomy. RESULTS: After a median follow-up of 41 months (range 7-90), 13 patients of the 195 treated with BCS had an IBTR (6.7%), 6 patients had a regional relapse (3.1%), 28 women had distant metastases (14.4%). Twenty-three patients died of breast cancer (11.8%). Twenty-four patients treated with BCS had positive margins (12.3%). At 3 years, the crude cumulative incidence of local recurrence was 4.7% in women with negative margins, and 13.3% in women with positive margins (P=0.05). Cumulative incidence of distant metastases was similar in patients with positive and negative margins (P=0.16) and there was no significant difference in terms of OS according to the margin status (P=0.577). CONCLUSIONS: BCS after PC has an acceptable rate of IBTR. After a short follow-up, the presence of positive margins does not affect OS.

Adult↗

Assessing the status of axillary sentinel lymph nodes of breast carcinoma patients by a real-time quantitative RT-PCR assay for mammaglobin 1 mRNA.

The aim of the study was to assess the accuracy of a real-time quantitative RT-PCR (qRT-PCR) assay for mammaglobin 1 mRNA in the detection of metastatic breast cancer in axillary sentinel lymph nodes (SLN), comparing the results with those of qualitative RT-PCR assays and of an extensive histopathological examination. A retrospective series of 81 SLN from 72 patients and a validation series of 61 SLN from 61 patients were evaluated. In the retrospective series, the qRT-PCR assay was positive for 23 (28.4%) of the 81 SLN. The overall concordance with histopathology was 93.8%, with a sensitivity of 90.9%, a specificity of 94.9%, a positive predictive value (PPV) of 87% and a negative predictive value (NPV) of 96.6%. In the same series, qualitative RT-PCR showed an overall concordance with histopathology of 86.4%, a sensitivity of 72.7%, a specificity of 91.5%, a PPV of 76.2% and a NPV of 90%. In the validation series, including 23 patients with pure in situ carcinoma, the real-time qRT-PCR assay showed an overall concordance with the histopathologic findings of 93.4%, with a sensitivity of 75.0%, a specificity of 94.7%, a PPV of 50.0% and a NPV of 98.2%. We conclude that real-time qRT-PCR assays for mammaglobin 1 are more sensitive and specific that qualitative RT-PCR assays for the detection of metastatic breast carcinoma in axillary SLN, but it should not be regarded as a possible substitute for an extensive histopathological scrutiny of the SLN in the clinical practice.

Axilla↗

Video-assisted management of malignant pleural effusion in breast carcinoma.

BACKGROUND: Advanced breast carcinoma almost always leads to a malignant pleural effusion, conditioning the performance status of patients and consequently quality of life. The treatment of malignant pleural effusion should be a priority in the management of such patients. The results of video-thoracoscopic approach (VATS) chemical pleurodesis was analyzed in patients with recurrent pleural effusion from breast carcinoma. METHODS: From October 1998 to June 2004, 71 consecutive patients with breast carcinoma-related pleural effusion were treated by the same thoracic-surgeon team with intracavitary nebulization of 8 g of asbestos-free sterilized talc via VATS. Multiple pleural biopsies were performed to determine biologic characteristics of recurrent disease. RESULTS: Talc pleurodesis was performed in all cases, with no intraoperative or postoperative complications. Median length of hospital stay was 5 days (range, 5-8). The overall success rate of the surgical procedure was 89% (confidence interval [CI], 79-95%) with a mean follow-up of 22 months (range, 2-81 mos). The overall survival time was 17 months (range, 2-80). Biopsies showed a switch on receptor status and c-erB-2 status from negative (primary tumor) to positive (pleural metastasis) in 11 (15%) patients. In another 7 (9.8%) patients, we obtained completely new information that was hitherto unknown. CONCLUSION: Talc pleurodesis via VATS is an effective and safe procedure that yields a high rate of success at the first attempt and achieves long-term control of malignant pleural effusion due to breast carcinoma. Concomitant biopsies performed during the VATS procedure were a determining factor in the subsequent decision-making process.

Adult↗

Sentinel-lymph-node biopsy as a staging procedure in breast cancer: update of a randomised controlled study.

BACKGROUND: In women with breast cancer, sentinel-lymph-node biopsy (SLNB) provides information that allows surgeons to avoid axillary-lymph-node dissection (ALND) if the SLN does not have metastasis, and has a favourable effect on quality of life. Results of our previous trial showed that SLNB accurately screens the ALN for metastasis in breast cancers of diameter 2 mm or less. We aimed to update this trial with results from longer follow-up. METHODS: Women with breast tumours of diameter 2 cm or less were randomly assigned after breast-conserving surgery either to SLNB and total ALND (ALND group), or to SLNB followed by ALND only if the SLN was involved (SLN group). Analysis was restricted to patients whose tumour characteristics met eligibility criteria after treatment. The main endpoints were the number of axillary metastases in women in the SLN group with negative SLNs, staging power of SLNB, and disease-free and overall survival. FINDINGS: Of the 257 patients in the ALND group, 83 (32%) had a positive SLN and 174 (68%) had a negative SLN; eight of those with negative SLNs were found to have false-negative SLNs. Of the 259 patients in the SLN group, 92 (36%) had a positive SLN, and 167 (65%) had a negative SLN. One case of overt clinical axillary metastasis was seen in the follow-up of the 167 women in the SLN group who did not receive ALND (ie, one false-negative). After a median follow-up of 79 months (range 15-97), 34 events associated with breast cancer occurred: 18 in the ALND group, and 16 in the SLN group (log-rank p=0.6). The overall 5-year survival of all patients was 96.4% (95% CI 94.1-98.7) in the ALND group and 98.4% (96.9-100) in the SLN group (log-rank p=0.1). INTERPRETATION: SLNB can allow total ALND to be avoided in patients with negative SLNs, while reducing postoperative morbidity and the costs of hospital stay. The finding that only one overt axillary metastasis occurred during follow-up of patients who did not receive ALND (whereas eight cases were expected) could be explained by various hypotheses, including those from cancer-stem-cell research.

Adult↗

Real-time in vivo dosimetry using micro-MOSFET detectors during intraoperative electron beam radiation therapy in early-stage breast cancer.

PURPOSE: In a previous paper we reported the results of off-line in vivo measurements using radiochromic films in IOERT. In the present study, a further step was made, aiming at the improvement of the effectiveness of in vivo dosimetry, based on a real-time check of the dose. MATERIALS AND METHODS: Entrance dose was determined using micro-MOSFET detectors placed inside a thin, sterile, transparent catheter. The epoxy side of the detector was faced towards the beam to minimize the anisotropy. Each detector was plugged into a bias supply (standard sensitivity) and calibrated at 5 Gy using 6 MeV electrons produced by a conventional linac. Detectors were characterized in terms of linearity, precision and dose per pulse dependence. No energy and temperature dependence was found. The sensitivity change of detectors was about 1% per 20 Gy accumulated dose. Correction factors to convert surface to entrance dose were determined for each combination of energy and applicator. From November 2004 to May 2005, in vivo dosimetry was performed on 45 patients affected by early-stage breast cancer, who underwent IOERT to the tumour bed. IOERT was delivered using electrons (4-10 MeV) at high dose per pulse, produced by either a Novac7 or a Liac mobile linac. RESULTS: The mean ratio between measured and expected dose was 1.006+/-0.035 (1 SD), in the range 0.92-1.1. The procedure uncertainty was 3.6%. Micro-MOSFETs appeared suitable for in vivo dosimetry in IOERT, although some unfavourable aspects, like the limited lifetime and the anisotropy with no build-up, were found. Prospectively, a real-time action level (+/-6%) on dose discrepancy was defined. CONCLUSIONS: Excellent agreement between measured and expected doses was found. Real-time in vivo dosimetry appeared feasible, reliable and more effective than the method previously published.

Breast Neoplasms↗

Second biopsy of axillary sentinel lymph node for reappearing breast cancer after previous sentinel lymph node biopsy.

BACKGROUND: Sentinel lymph node biopsy (SLNB) is a safe and accurate axillary staging procedure for patients with primary operable breast cancer. An increasing proportion of these patients undergo breast-conserving surgery, and 5% to 15% will develop local relapses that necessitate reoperation. Although a previous SLNB is often considered a contraindication for a subsequent SLNB, few data support this concern. METHODS: Between January 2000 and June 2004, 79 patients who were previously treated at our institution with breast-conserving surgery and who had a negative SLNB for early breast cancer developed, during follow-up, local recurrence that was amenable to reoperation. Eighteen of these patients were offered a second SLNB because of a clinically negative axillary status an average of 26.1 months after the primary event. RESULTS: In all 18 patients (7 with ductal carcinoma-in-situ and 11 with invasive recurrences), preoperative lymphoscintigraphy showed an axillary sentinel lymph node, with a preoperative identification rate of 100%, and 1 or more SLNs (an average of 1.3 per patient) were surgically removed. Sentinel lymph node metastases were detected in two patients with invasive recurrence, and a complete axillary dissection followed. At a median follow up of 12.7 months, no axillary recurrences have occurred in patients who did not undergo axillary dissection. CONCLUSIONS: Second SLNB after previous SLNB is technically feasible and likely effective in selected breast cancer patients. A larger population and longer follow-up are necessary to confirm these preliminary data.

Adult↗

Size of breast cancer metastases in axillary lymph nodes: clinical relevance of minimal lymph node involvement.

BACKGROUND: Overt ipsilateral axillary lymph node metastases of breast cancer are the most significant prognostic indicators for women who have undergone surgery, yet the clinical relevance of minimal involvement (isolated tumor cells and micrometastases) of these nodes is uncertain. PATIENTS AND METHODS: We evaluated biologic features, adjuvant treatment recommendations, and prognosis for 1,959 consecutive patients with pT1-3, pN0, minimal lymph node involvement (pN1mi or pN0i+), or pN1a (single positive node) and M0, who were operated on and counseled for medical therapy from April 1997 to December 2000. RESULTS: Patients with pN1a and pN1mi/pN0i+, when compared with patients with pN0 disease, were more often prescribed anthracycline-containing chemotherapy (39.1% v 33.2% v 6.1%, respectively; P < .0001) and were less likely to receive endocrine therapy alone (9.8% v 19.4% v 41.9%, respectively; P < .0001). At the multivariate analysis, a statistically significant difference in disease-free survival (DFS) and in the risk of distant metastases was observed for patients with pN1a versus pN0 disease (hazard ratio [HR] = 2.04; 95% CI, 1.46 to 2.86; P < .0001 for DFS; HR = 2.32; 95% CI, 1.42 to 3.80; P = .0007 for distant metastases) and for patients with pN1mi/pN0i+ versus pN0 disease (HR = 1.58; 95% CI, 1.01 to 2.47; P = .047 for DFS; HR = 1.94; 95% CI, 1.04 to 3.64; P = .037 for distant metastases). CONCLUSION: Even minimal involvement of a single axillary node in breast cancer significantly correlates with worse prognosis compared with no axillary node involvement. Further studies are required before widespread modification of clinical practice.

Adult↗

The sentinel lymph node biopsy under local anesthesia in breast carcinoma: experience of the European Institute of Oncology and impact on quality of life.

BACKGROUND: Sentinel lymph node biopsy performed under local anesthesia currently represents one of the main fields of investigation in breast cancer surgery because of its positive impact on the management and organisation of treatment. We previously published our preliminary experience with 115 patients; here we present an update of our experience with particular attention devoted to quality of life. PATIENTS AND METHODS: From September 2000 to April 2003 we treated 244 patients with this technique. We developed a 'questionnaire of impact on quality of life - LA' to be administered to these patients, to assess their reactions to the procedure: we administered the questionnaire to 243 patients. Our aim was to verify whether the advantages in terms of treatment planning are accompanied by an improvement in quality of life and a good degree of acceptance by the patients. RESULTS: The questionnaire was administered to the patients during follow-up and we obtained positive results; the impact of the procedure under local anesthesia was positive for 81% of patients, while the remaining 19% reported that it had 'no impact at all'. We can conclude that axillary sentinel node biopsy, when performed under local anesthesia, meets with a very high degree of acceptance by the patients and exerts an optimal impact on quality of life.

Adult↗

The sentinel node biopsy after previous breast surgery: preliminary results on 543 patients treated at the European Institute of Oncology.

BACKGROUND: Sentinel lymph node biopsy (SLNB) is an accurate alternative to complete axillary lymph node dissection (ALND) in clinically node-negative breast cancer patients. A previous breast biopsy has been considered a relative contraindication to SLNB. We examined the accuracy of SLNB by following the axillary relapses after the procedure in patients who had undergone a breast biopsy before SLNB. PATIENTS AND METHODS: Up to December 2003, 4351 patients with the diagnosis of invasive breast cancer underwent SLNB at the European Institute of Oncology. Already, 543 of these patients had undergone a breast biopsy; from June 1997 to January 2004, these patients received SLNB by lymphoscintigraphy performed on the biopsy area. We followed these patients with a clinical assessment every 6 months and instrumental examinations every year, particularly focusing on the research of axillary relapse of disease. RESULTS: In 70.4% of cases, the sentinel node was negative, and only three cases underwent further axillary dissection. The sentinel node was identified in 99% of cases and this was the only positive node in 61.5% of cases with positive axillary nodes. The median follow-up was 2 years; 4 nodal recurrences were observed: 3 axillary lymph node relapses and 1 loco-regional. CONCLUSIONS: SLNB accuracy after a previous breast biopsy is comparable with the results obtained in validation studies. SLNB after a previous breast biopsy can be considered a standard procedure. Lymphoscintigraphy identifies the sentinel node in 99% of patients.

Adult↗

Conservative treatment of breast cancer: its evolution.

BACKGROUND: Over recent decades, breast carcinoma surgery has witnessed a considerable evolution. The extent of surgery undertaken has progressively reduced, leading to less disfigurement and a significant improvement in quality of life, thereby offering women considerable motivation to seek early diagnosis. From an oncological perspective, outcome was found to be equally effective, and this key observation provided us with significant impetus to investigate the effects of reducing the radiotherapy field. PATIENTS AND METHODS: We present the achievements made so far in the conservative treatment of breast carcinoma, based on experiences at the European Institute of Oncology in Milan and, prior to 1994, at the Milan Cancer Institute. RESULTS AND DISCUSSION: Conservative surgery of breast carcinoma, both in the breast and in the axillary nodes, has yielded very good results in terms of overall survival and impact on quality of life. The reduction of the radiation field, conducted in our Institute by means of electron intraoperative radiotherapy (ELIOT), is currently the subject of evaluation via a randomised trial.

Breast Neoplasms↗

The pilot trial on intraoperative radiotherapy with electrons (ELIOT): update on the results.

BACKGROUND: Intraoperative radiotherapy with electrons (ELIOT) is under evaluation in the conservative treatment of breast carcinoma. At the European Institute of Oncology we started our study on ELIOT in 1999; the first step has been a phase I-II trial to test feasibility and tolerance of ELIOT in patients candidate to breast conserving surgery. PATIENTS AND METHODS: From July 1999 to September 2000, 101 patients affected by unifocal breast carcinoma up to a diameter of 2.5 cm received wide resection of the breast followed by ELIOT. The trial was based on a dose-escalation starting from 10 Gy: we tested the dose-levels of 10, 15, 17, 19 and 21 Gy. The dose-levels of 10 and 15 Gy were followed by a reduced course of external fractionated radiotherapy. Most patients received 21 Gy intraoperatively. Patients were evaluated 1, 3, 6 and 12 months after surgery, and thereafter every 6 months, to look for early, intermediate, late complications, and other events. RESULTS: After a mean follow-up of 42 months, 16 patients (16%) developed breast fibrosis, mild in 15, severe in one, which resolved within 24 months. Two patients suffered of postoperative infection, and four developed a lyponecrosis in the treated area. CONCLUSION: The first group of patients treated with ELIOT in the phase I-II trial are confirming the good impact of the procedure in terms of tolerance and quality of life.

Aged↗

Sentinel node biopsy in breast cancer: early results in 953 patients with negative sentinel node biopsy and no axillary dissection.

Sentinel node biopsy in patients with breast carcinoma accurately predicts the axillary nodal status. However, in some 6% of patients with negative sentinel nodes the remaining axillary nodes harbour metastases. Our purpose was to observe a large number of patients who did not undergo an axillary dissection after a negative sentinel node biopsy for the appearance of overt axillary metastases. 953 patients treated from 1996 to 2000, with negative sentinel nodes not submitted to axillary dissection, were followed-up to 7 years, with a median follow-up of 38 months. Fifty-five unfavourable events occurred among the 953 patients, 37 (4%) related to the primary breast carcinoma. Three cases of overt axillary metastases were found: they received total axillary dissection and are presently alive and well. The 5 year overall survival rate of the whole series was 98%. Patients with negative sentinel node biopsies not submitted to axillary dissection show during follow-up a rate of overt axillary metastases that is lower than that expected.

Adult↗

Predicting the risk for additional axillary metastases in patients with breast carcinoma and positive sentinel lymph node biopsy.

OBJECTIVE: To assess whether the risk for nonsentinel node metastases may be predicted, thus sparing a subgroup of patients with breast carcinoma and a positive sentinel lymph node (SLN) biopsy completion axillary lymph node dissection (ALND). SUMMARY BACKGROUND DATA: The SLN is the only involved axillary lymph node in the majority of the patients undergoing ALND for a positive SLN biopsy. A model to predict the status of nonsentinel axillary lymph nodes could help tailor surgical therapy to those patients most likely to benefit from completion ALND. METHODS: All the axillary sentinel and nonsentinel lymph nodes of 1228 patients were reviewed histologically and reclassified according to the current TNM classification of malignant tumors as bearing isolated tumor cells only, micrometastases, or (macro)metastases. The prevalence of metastases in nonsentinel lymph nodes was correlated to the type of SLN involvement and the size of the metastasis, the number of affected SLNs, and the prospectively collected clinicopathologic variables of the primary tumors. RESULTS: In multivariate analysis, further axillary involvement was significantly associated with the type and size of SLN metastases, the number of affected SLNs, and the occurrence of peritumoral vascular invasion in the primary tumor. A predictive model based on the characteristics most strongly associated with nonsentinel node metastases was able to identify subgroups of patients at significantly different risk for further axillary involvement. CONCLUSIONS: Patients with the most favorable combination of predictive factors still have no less than 13% risk for nonsentinel lymph node metastases and should be offered completion ALND outside of clinical trials of SLN biopsy without back-up axillary clearing.

Adult↗

Full-dose intraoperative radiotherapy with electrons during breast-conserving surgery: experience with 590 cases.

BACKGROUND: Previous studies show that local recurrences after breast-conserving treatment occur in the site of the primary tumor. The need for postoperative radiotherapy on the whole breast is challenged in favor of radiotherapy limited to the area of the breast at high risk of recurrence. The new mobile linear accelerators easily moved close to the operating table to allow the full-dose irradiation during surgery. PATIENTS AND METHODS: From July 1999 to December 2003, 590 patients affected by unifocal breast carcinoma up to a diameter of 2.5 cm received wide resection of the breast followed by intraoperative radiotherapy with electrons (ELIOT). Most patients received 21 Gy intraoperatively, biologically equivalent to 58 to 60 Gy in standard fractionation. Patients were evaluated 1, 3, 6, and 12 months after surgery, and thereafter every 6 months, to look for early, intermediate, late complications, and other events. RESULTS: After a follow-up from 4 to 57 months (mean, 24 months; median, 20 months), 19 patients (3.2%) developed breast fibrosis, mild in 18, severe in 1, which resolved within 24 months. Three patients (0.5%) developed local recurrences, 3 patients ipsilateral carcinomas in other quadrants and other 5 patients contralateral breast carcinoma. One patient (0.2%) died of distant metastases. CONCLUSIONS: ELIOT is a safe method for treating conservatively operated breasts, avoids the long period of postoperative radiotherapy, and reduces drastically the cost of radiotherapy. ELIOT reduces radiation to normal tissues and organs. Results on short-term and middle-term toxicity up to 5 years of follow-up are good. Data on local control are encouraging.

Adenocarcinoma↗

Loss of negative regulation by Numb over Notch is relevant to human breast carcinogenesis.

The biological antagonism between Notch and Numb controls the proliferative/differentiative balance in development and homeostasis. Although altered Notch signaling has been linked to human diseases, including cancer, evidence for a substantial involvement of Notch in human tumors has remained elusive. Here, we show that Numb-mediated control on Notch signaling is lost in approximately 50% of human mammary carcinomas, due to specific Numb ubiquitination and proteasomal degradation. Mechanistically, Numb operates as an oncosuppressor, as its ectopic expression in Numb-negative, but not in Numb-positive, tumor cells inhibits proliferation. Increased Notch signaling is observed in Numb-negative tumors, but reverts to basal levels after enforced expression of Numb. Conversely, Numb silencing increases Notch signaling in normal breast cells and in Numb-positive breast tumors. Finally, growth suppression of Numb-negative, but not Numb-positive, breast tumors can be achieved by pharmacological inhibition of Notch. Thus, the Numb/Notch biological antagonism is relevant to the homeostasis of the normal mammary parenchyma and its subversion contributes to human mammary carcinogenesis.

Breast Neoplasms↗

Chemotherapy is more effective in patients with breast cancer not expressing steroid hormone receptors: a study of preoperative treatment.

PURPOSE: The purpose of this research was to identify factors predicting response to preoperative chemotherapy. EXPERIMENTAL DESIGN: In a large volume laboratory using standard immunohistochemical methods, we reviewed the pretreatment biopsies and histologic specimens at final surgery of 399 patients with large or locally advanced breast cancer (cT2-T4, N0-2, M0) who were treated with preoperative chemotherapy. The incidence of pathological complete remission and the incidence of node-negative status at final surgery were assessed with respect to initial pathological and clinical findings. Menopausal status, estrogen receptor status, progesterone receptor status [absent (0% of the cells positive) versus expressed], clinical tumor size, histologic grade, Ki-67, Her-2/neu expression, and type and route of chemotherapy were considered. RESULTS: High rates of pathological complete remission were associated with absence of estrogen receptor and progesterone receptor expression (P < 0.0001), and grade 3 (P = 0.001). Significant predictors of node-negative status at surgery were absence of estrogen receptor and progesterone receptor expression (P < 0.0001), clinical tumor size <5 cm (P < 0.001), and use of infusional regimens (P = 0.003). The chance of obtaining pathological complete remission or node-negative status for patients with endocrine nonresponsive tumors compared with those having some estrogen receptor or progesterone receptor expression was 4.22 (95% confidence interval, 2.20-8.09, 33.3% versus 7.5%) and 3.47 (95% confidence interval, 2.09-5.76, 42.9% versus 21.7%), respectively. Despite the significantly higher incidence of pathological complete remission and node-negative status achieved by preoperative chemotherapy for patients with estrogen receptor and progesterone receptor absent disease, the disease-free survival was significantly worse for this cohort compared with the low/positive expression cohort (4-year disease-free survival %: 41% versus 74%; hazard ratio 3.22; 95% confidence interval, 2.28-4.54; P < 0.0001). CONCLUSIONS: Response to preoperative chemotherapy is significantly higher for patients with endocrine nonresponsive tumors. New chemotherapy regimens or combinations should be explored in this cohort of patients with poor outcome. For patients with endocrine responsive disease, the role of preoperative endocrine therapies should be studied.

Adolescent↗