Targeting Breast Cancer Using 177 Lu-Labeled Trastuzumab and Trastuzumab Fragment : First-in-Human Clinical Experience.

PURPOSE: A monoclonal antibody, trastuzumab, is used for immunotherapy for HER2-expressing breast cancers. Large-sized antibodies demonstrate hepatobiliary clearance and slower pharmacokinetics. A trastuzumab fragment (Fab; 45 kDa) has been generated for theranostic use. PATIENTS AND METHODS: Fab was generated by papain digestion. Trastuzumab and Fab have been radiolabelled with 177 Lu after being conjugated with a bifunctional chelating. The affinity and target specificity were studied in vitro. The first-in-human study was performed. RESULTS: The bifunctional chelating agent conjugation of 1-2 molecules with trastuzumab and Fab was detected at the molar ratio 1:10 in bicarbonate buffer (0.5 M, pH 8) at 37°-40°C. However, 2-3 molecules of bifunctional chelating agent were conjugated when DMSO in PBS (0.1 M, pH 7) was used as a conjugation buffer at a molar ratio of 1:10. The radiolabelling yield of DOTA-conjugated Fab and trastuzumab at pH 5, 45°C to 50°C, with incubation time 2.5-3 hours was 80% and 41.67%, respectively. However, with DOTAGA-conjugated trastuzumab and Fab, the maximum radiolabelling yield at pH 5.5, 37°C, and at 2.5-3 hours was 80.83% and 83%, respectively. The calculated K d of DOTAGA Fab and trastuzumab with HER2-positive SKBR3 cells was 6.85 ± 0.24 × 10 -8 M and 1.71 ± 0.10 × 10 -8 M, respectively. DOTAGA-Fab and trastuzumab showed better radiolabelling yield at mild reaction conditions.177 Lu-DOTAGA-Fab demonstrated higher lesion uptake and lower liver retention as compared with 177 Lu-DOTAGA-trastuzumab. However, 177 Lu-DOTAGA-Fab as compared with 177 Lu-DOTAGA-trastuzumab showed a relatively early washout (5 days) from the lesion. CONCLUSIONS: 177 Lu-DOTAGA-Fab and trastuzumab are suitable for targeting the HER2 receptors.

A prospective study evaluating patient reported outcome measures in patients who have undergone chest wall perforator flaps.

AIMS: To evaluate Patient Reported Outcome Measures (PROMs) and surgical outcomes in patients undergoing Chest Wall Perforator Flaps (CWPFs). METHODS: This was an observational single cohort study using an audit approach and a survey instrument. 84 patients who had undergone CWPFs in the last 5 years at the Department of Breast Surgery, City Hospital Birmingham, were identified from a pre-existing database. Surgical outcomes were recorded. Patients were contacted telephonically or in person at the time of follow-up and were asked to fill up a PROMs questionnaire. RESULTS: Out of 84 patients, 58 patients chose to respond. The average age of the patients was 51.3 years (±8.2 years). The average follow-up was 15.4 months (±9.9 months). The most common histological subtype was Infiltrating ductal carcinoma (IDC)-Not otherwise specified 34/58 (58.6%). Majority of the patients had T2 cancers-28/58 (48.3%). 26/58 (44.8%) were node negative. Eight patients (13.7%) had post-operative complications. No patient had total/partial flap loss. Nine patients (15.5%) had margin re-excision. One patient developed distant metastasis while 1 patient developed a second primary. Fifty-one patients (88%) were either satisfied or very satisfied with the post-operative appearance of the breasts. Thirty-six patients (62%) had no/little persistent pain or tenderness post-surgery. Eighty-six per cent (38/44) of the patients undergoing Lateral Intercostal Artery Perforator (LICAP) Flap and 16/18 (89%) of patients undergoing Anterior Intercostal Artery Perforator (AICAP) flap had no/little difficulty in carrying out normal activities at follow up. CONCLUSION: CWPFs are associated with a low complication rate and a high patient satisfaction rate.

HYPofractionated Adjuvant RadioTherapy in 1 versus 2 weeks in high-risk patients with breast cancer (HYPART): a non-inferiority, open-label, phase III randomised trial.

BACKGROUND: Breast cancer is the most common cancer in women. Radiotherapy is an important part of breast cancer treatment after surgery. Breast cancer radiotherapy is usually delivered in 3-5 weeks. This is a long duration for women with breast cancer to stay away from the family and work. We wanted to reduce this duration so that the wages loss and the logistics can be minimised for these patients. Hypofractionation, i.e. high dose per fraction, is delivered in a smaller number of days. In this study, we will compare a 1-week schedule of hypofractionated adjuvant whole breast/chest wall and/or regional nodal radiotherapy against 2 weeks for locoregional disease control, toxicities, quality of life (QoL), survival and second cancers after primary surgery in patients with breast cancer. METHODS: Eligible patients with breast cancer after mastectomy or breast conserving surgery (BCS) will be treated with a radiotherapy dose of 26 Gy in 5 fractions over 1 week in the study arm and 34 Gy in 10 fractions over 2 weeks in the control arm. The primary endpoint of this noninferiority study will be locoregional tumour control. Secondary endpoints will be early and late radiation toxicities, quality of life, contralateral primary tumours, regional and distant metastases, survival and second cancers. A total of 1018 patients will be randomised (1:1) to receive 1 week or 2 weeks of radiotherapy. An event-driven analysis will be performed after at least 94 patients have documented locoregional recurrences. Acute radiation toxicity will be assessed and scaled according to the RTOG grading system. Late radiation toxicity will be assessed with the Radiation Therapy Oncology Group and the European Organisation for Research and Treatment of Cancer late radiation morbidity scale. Cosmetic assessment will be done using Harvard/NSABP/RTOG breast cosmesis grading scale at baseline and 3 and 5 years. QoL will be assessed with EORTC QLQ-30 and EORTC QLQ-BR 23 at baseline and 3 and 5 years. DISCUSSION: Hypofractionation reduces treatment time to half while maintaining breast cosmesis and gives control rates equal to conventional fractionation. This is possible because breast tissue can tolerate high dose per fraction. In this study, we presume that 1-week radiotherapy will be non-inferior to 2 week radiotherapy, i.e. disease control will be similar with both the schedules without additional side effects, and QoL of these patients will be maintained. If we are able to achieve these outcomes, then patients will be able to complete their radiotherapy in less duration. There is not much data on regional nodal irradiation with hypofraction in breast cancer. We have used hypofraction for regional nodal irradiation in the past and not encountered any safety issue. If we are able to prove that late-term effects are comparable in the two schedules, it will make the radiation oncologist confident about hypofractionation in breast cancer. As breast cancer is a leading cancer in females and radiation therapy is an integral part of its local management, hypofractionation will help radiation centres worldwide to meet the growing need for radiation treatment in breast cancer, particularly in developing countries where resources are limited. It will also reduce the financial burden on the patient and family. Since we will treat these patients with both simple and complex radiotherapy techniques, it will also be possible for the low-income countries to follow this trial without needing a high-end or expensive radiotherapy equipment as the planning and treatment process will be very simple. TRIAL REGISTRATION: The trial is registered with ClinicalTrials.gov ID NCT04472845 and CTRI with REF/2020/09/037050.

Assessment of Pathological Complete Response Using Vacuum-Assisted Biopsy in Breast Cancer Patients Who Have Clinical and Radiological Complete Response After Neo-Adjuvant Chemotherapy.

Background: Any treatment protocol that leads to complete elimination of surgery may lead to a better patient acceptance of breast cancer treatments. Objectives: We conducted this study to assess the feasibility of preoperative vacuum-assisted biopsies in identifying pathological complete response (pCR) and its accuracy in correlation to final histopathology report (HPR), in an Indian setting. Methods: This was a prospective study conducted between October 1, 2019, and March 31, 2021. Patients with early breast cancer, estrogen and progesterone receptors negative and either Her2 positive or negative, and who were fit to undergo marker placement at the centre of the tumour and to receive third-generation chemotherapy (4 cycles of 3 weekly doxorubicin and cyclophosphamide followed by 4 cycles of 3 weekly docetaxel) were included in the study. Following the enrolment, a tissue marker was placed at the centre of the tumour and appropriate chemotherapy was started. Patients who achieved clinical complete response were subjected to ultrasound-guided vacuum-assisted biopsy (VAB) from the tumour bed before surgery. Pathology results of the VAB and resected specimen were then compared. Descriptive statistics were used in the study. Results: Eighteen patients were enrolled in the study, with a mean age of 43.6 ± 9.8 years. However, only 10 were eligible for VAB procedure, and sensitivity and specificity were calculated based on the results of these 10 patients only. Vacuum-assisted biopsy showed sensitivity of 50% and specificity of 100% in identifying pCR. Combination of mammography, ultrasonography, and VAB showed sensitivity of 77.8% and specificity of 66.7% in identifying pCR. Conclusion: Vacuum-assisted biopsy of tumour bed may not be sensitive enough to eliminate surgery even in patients who have had exceptional response to neo-adjuvant chemotherapy.

Pushing the Boundaries of Pedicled Chest Wall Perforator Flaps in Oncoplastic Breast Surgery.

Among 145 chest wall perforator flaps (CWPFs) performed at City Hospital, Birmingham (September 2017-February 2022), 11 were for novel indications, four were for whole breast reconstructions, two were for implant salvage procedures, three were CWPFs with skin paddle to replace excised skin/nipple-areola complex, and two were for upper inner quadrant tumours. Tumour characteristics and post-operative complications were noted. Patient-reported outcomes measures (PROMs) were measured using a questionnaire adapted from the National Mastectomy and Breast Reconstruction Audit (NMBRA) study. Among 11 patients, nine (81.81%) did not develop any complications. Ten patients responded to PROMs (median follow-up of eight months). The PROMs assessment showed that all patients (100%) were satisfied with the post-operative breast appearance. Of the patients, 90% (9/10) felt the results of their surgery to be good, very good, or excellent. Of the patients, 70% (7/10) said that they have no/little persistent pain. None of the patients had difficulty carrying out normal activities. Thus, the applications of CWPFs could be extended for whole breast reconstruction, implant salvage procedures, where skin paddle is needed, and for upper inner quadrant tumours.

Expression of Proteinase-activated Receptor 2 (PAR2) as a Correlate of Concern in Triple-negative Breast Cancer (TNBC).

PURPOSE: Triple-negative breast cancer (TNBC), a highly aggressive cancer with poor outcome and lacking specific diagnostic, prognostic, or targeted therapeutic strategies, constitutes roughly 20% of all breast cancer cases. TNBC cells lack receptors for estrogen, progesterone, and human epidermal growth factor. The effort continues to find a suitable correlate that could serve as a TNBC biomarker, or as therapeutic target, or both. MATERIALS AND METHODS: A retrospective study was performed with 88 TNBC and 74 non-TNBC patients who had undergone mastectomy/lumpectomy with axillary clearance for carcinoma breast. Immunohistochemical staining was carried out for levels of proteinase-activated receptor 2 (PAR2), encoded by F2RL1 gene, and staining scores were calculated, based on intensity and percentage positivity. RESULTS: PAR2 levels were markedly upregulated in TNBC patients, compared with patients with other breast cancer subtypes. Amongst different non-TNBC subtypes, higher expression was noted in luminal B (88.8%) and HER2+ (100%), compared with luminal A (52.5%). PAR2 levels were significantly high in TNBC patients with age more than 40 years than corresponding patients of non-TNBC group (P=0.0017). Furthermore, there was a statistically significant increase in levels of PAR2 expression in lymph node negative (P=0.0096) and early stage (P=0.005) of TNBC versus non-TNBC patients. PAR2 staining of ductal carcinoma in situ and invasive ductal carcinoma revealed lower expression in invasive component. CONCLUSIONS: Our data suggest that PAR2 levels constitute a correlate of concern for TNBC, tying in with a recent report that higher levels of F2RL1 gene expression correlate with poorer disease-free, as well as overall survival in TNBCs.

Tc-99m-tamoxifen: A novel diagnostic imaging agent for estrogen receptor-expressing breast cancer patients.

PURPOSE The aim of the study was to radiolabel, characterize, and perform in vitro and in vivo assessment of Technetium-99m (Tc-99m) tamoxifen for screening ER expressing lesions in breast cancer patients. METHODS In this study, tamoxifen has been radiolabeled with Tc-99m via Tc-99m-tricarbonyl core. The characterization and quality control tests of Tc-99m-tamoxifen were performed. In vitro recep- tor binding and blocking studies were performed in both positive control (MCF-7) and negative control cell lines (MDA-MB-231). Normal biodistribution studies were performed in female Wistar albino rats. The pilot clinical studies were performed in 4 ER-expressing breast cancer patients. Of the 4 patients, 1 was on tamoxifen therapy. All 4 patients had also undergone Fluorine-18 fluorodeoxyglucose (F-18-FDG) positron emission tomography/computed tomography. RESULTS Tamoxifen was radiolabeled with Tc-99m via Tc-99m-tricarbonyl core with more than 95% radio- chemical yield. Mass spectra showed a peak corresponding to the molecular weight of Tc-99m- tricarbonyl and Tc-99m-tamoxifen. The site of binding of Tc-99m-tricarbonyl with tamoxifen was determined by proton nuclear magnetic resonance. The Tc-99m-tamoxifen showed 30% binding with MCF-7 and only 1%-2% receptor binding with MDA-MB-231 cell lines. Also, the percentage of receptor binding was drastically reduced (up to 72%) when ER was saturated with 50 times the excess molar ratio of unlabeled tamoxifen. In a pilot patient study, Tc-99m-tamoxifen uptake was observed in primary and metastatic lesions. However, no uptake was observed in a patient who was on tamoxifen therapy. The uptake of F-18-FDG was noted in all the patients. CONCLUSION Tamoxifen was radiolabeled with an in-house-synthesized Tc-99m-tricarbonyl core. The radio- labeled complex has been characterized and evaluated for receptor specificity in in vitro and in vivo studies. Also, this is the first clinical study using Tc-99m-tamoxifen for imaging ER. More patients need to be evaluated to further explore the role of Tc-99m-tamoxifen in ER-expressing lesions.

Comparison of surgical and oncological outcomes between oncoplastic breast-conserving surgery versus conventional breast-conserving surgery for treatment of breast cancer: A systematic review and meta-analysis of 31 studies.

AIMS: To evaluate comparative outcomes of oncoplastic breast conserving surgery (OBCS) versus conventional breast conserving surgery (BCS) for breast cancer treatment. METHODS: A systematic search of multiple electronic data sources was conducted, and all eligible studies comparing OBCS and BCS were included. Characteristics of the tumour includes preoperative size of tumour on imaging and the weight of the specimen after resection. While positive margins rate, re-excision rate, completion mastectomy rate and loco-regional recurrence were considered as oncological outcome parameters. Post-operative complications include surgical site infection (SSI), seroma, haematoma and skin/nipple necrosis. RESULTS: Thirty-one studies reporting a total number of 115011 patients who underwent OBCS (n = 11978) or BCS (n = 103033) were included. OBCS group showed lower risk of positive margins rate [OR 0.76, P = 0.05], re-excision rate [OR 0.72, P = 0.02], and loco-regional recurrence [OR 0.62, P = 0.03] compared to BCS group. There was no significant difference between the two groups regarding post-operative complications. CONCLUSION: Although there is a lack of level 1 evidence, the available studies clearly demonstrate superior or at least equivalent outcomes when comparing OBCS with conventional BCS. The benefits of OBCS include dealing with larger tumours, wider surgical margins and better aesthetic results for patients.

Development 68Ga trastuzumab Fab and bioevaluation by PET imaging in HER2/neu expressing breast cancer patients.

INTRODUCTION: Receptors on breast cancer cells play a crucial role in the management of patients. Trastuzumab is a widely used drug for the treatment of HER2/neu expressing tumors. ImmunoPET with trastuzumab is not feasible due to slow pharmacokinetics. Fragment of antigen-binding (Fab) radiolabeled with positron emitters can be used for immunoPET. METHODS: Fab has been generated by papain digestion and conjugated with the bifunctional chelating agent NOTA. The SDS-PAGE and MALDI-TOF were used to see the integrity of Fab and conjugated Fab. In-vitro stability and target specificity for HER2/neu receptors were performed in plasma and receptor binding with bio-layer interferometry (BLI) techniques. Radiolabeling was standardized with 68GaCl3 and PET imaging was performed in seven patients showing 18F fluorodeoxyglucose (18F-FDG) uptake and correlated with HER2/neu expression by immunohistochemistry. RESULTS: Fab production was optimized at molar ratio 23:1 of trastuzumab and papain at 37 °C with a constant stirrer at 850 rpm for 22-24 h, at pH 8. Conjugation with NOTA was standardized at molar ratio 1:25 of trastuzumab Fab and NOTA. Molecular mass of trastuzumab Fab-NOTA was found approximately 46.3 kDa (~1/3 of intact antibody). Trastuzumab Fab-NOTA showed radiolabelling efficiency of 48-70% with incubation time 15 min at 37-40 °C and pH 4.5-5.0. BLI demonstrated the affinity of trastuzumab, trastuzumab Fab and trastuzumab Fab-NOTA towards HER2/neu receptor with KD of <1pM, ~0.5nM and ~20nM, respectively. All immunohistochemistry proven patients showed uptake in primary breast lesion and lymph nodes. CONCLUSION: Trastuzumab Fab-NOTA is suitable for radiolabelling with 68Ga and ImmunoPET imaging of HER2/neu receptor.

Impact of an antimicrobial stewardship and monitoring of infection control bundle in a surgical intensive care unit of a tertiary-care hospital in India.

OBJECTIVES: Antimicrobial stewardship (AMS) in resource-limited settings lacks models that can be readily adapted to their settings. Here we discuss the impact of a combined strategy of AMS and monitoring of infection control practices in a tertiary-care centre of a developing country. METHODS: This study was undertaken in the surgical unit of a tertiary-care hospital over an 8-month period. In the first 2 months (baseline phase), prospective audit and feedback alone was undertaken, while in the next 6 months (intervention phase) this was supplemented with strategies such as antimicrobial timeout, correction of doses and bundle approach for prevention of hospital-acquired infections. RESULTS: A total of 337 patients were included (94 in the baseline phase and 243 in the intervention phase). There was a decrease in days of therapy per 1000 patient-days (1000PD) (1112.3 days vs. 1048.6 days), length of therapy per 1000PD (956 days vs. 936.3 days) and defined daily doses (DDD) per 1000PD for most antimicrobials. A decrease in double cover for Gram-negative infections (9.6% vs. 2.9%) but an increase in double anaerobic cover (4.2% vs. 7.4%) was observed. There was a decrease in the incidence of ventilator-associated pneumonia per 1000 ventilator-days in the intervention phase (46.4 vs. 35.4), whereas central line-associated bloodstream infections per 1000 central line-days remained the same (14.7 vs. 14.8). CONCLUSION: This study shows that implementation of routine AMS activities with monitoring of infection control practices can help decrease overall antimicrobial use. With furtherance of measures to control infection, antimicrobial use may be further curtailed.

Molecular Subtyping of Triple Negative Breast Cancer by Surrogate Immunohistochemistry Markers.

Triple negative breast cancer (TNBC) is a heterogeneous disease and an attempt was made to classify TNBCs into surrogate molecular subtypes using immunohistochemical markers. Tissue microarrays were constructed for 245 cases of TNBCs. For classification of TNBCs immunohistochemistry was done on tissue microarrays for cytokeratin 5/6, 4/14 (CK5/6, CK4/14), epidermal growth factor receptor (EGFR), vimentin, E-cadherin, claudin 3 and 7, androgen receptor (AR) and aldehyde dehydrogenase1A. The TNBCs were classified into basal-like 1 (BL1) type (CK5/6+, CK4/14+, EGFR- n=32; 13.1%), basal-like 2 (BL2) type (EGFR+, n=4; 1.6%), mesenchymal type (Vimentin+, E-cadherin ̅, claudin 3-and 7-, n=70; 28.6%), luminal androgen type (AR+, n=41; 16.7%), mixed type (n=37; 15.1%), and unclassified type (n=61; 24.9%). Luminal androgen receptor subtype showed apocrine features, and was associated with older age group, lower proliferation index and high frequency of lymph node metastasis. Basal subtype was cellular with rich stromal lymphocytic infiltrate. Mesenchymal stem like subtype was associated with younger age group with metaplastic and mesenchymal features. Mesenchymal stem like and unclassified subtype had shorter overall survival with median of 68.2 and 69.2 months, respectively, and the BL2 had median disease-free survival of 35.4 months. On immunohistochemistry TNBC is a heterogeneous entity composed of 6 major subtypes. Immunohistochemical subtyping of TNBC can provide information on prognostication and selection of appropriate targeted therapy for these patients.

Wire- and Ultrasound-Guided Localization: A Novel Technique for Excision of Nonpalpable Breast Tumors.

BACKGROUND: Excision of nonpalpable breast lesions requires intraoperative guidance. Wire-guided localization and intraoperative ultrasounds have been used successfully but suffer from some disadvantages. We describe a new modification of the standard technique using a combination of preoperative ultrasound in conjunction with standard wire-guided localization. METHODS: Wire and ultrasound-guided localization (WUGL) technique was used for the excision of nonpalpable breast lesions. RESULTS: Sixty-nine patients with nonpalpable breast lesions were subjected to excision using WUGL, out of whom 63 patients had a preoperative diagnosis of invasive/noninvasive breast cancer. Six patients had a preoperative diagnosis of benign lesions, out of which 3 patients were converted to invasive breast cancer on final pathology. Only 1 patient had positive margin. CONCLUSIONS: WUGL is a technique that uses a combination of well-accepted and easily available techniques. It has given good results and has the potential for widespread acceptance in resource-constrained situations.

99mTc Tamoxifen for Imaging Estrogen Receptor Expression in Metastatic Breast Cancer Patient.

Estrogen receptor-expressing breast cancer exhibits better prognosis due to responsiveness to antiestrogen treatment. Therefore, knowledge of the estrogen receptor status of a tumor is an important prognostic and predictive indicator in breast cancer. We present noninvasive imaging of estrogen receptors with Tc tamoxifen that can identify the active tumor and approachable sites for biopsy. It may help in selection of patients for hormone replacement therapy and in assessment of receptor status in recurrent disease and also in response evaluation.