Clinical relevance of circulating ESR1 mutations during endocrine therapy for advanced hormone-dependent endometrial carcinoma.

OBJECTIVE: Endocrine therapy is frequently administered in patients with hormone dependent (HR+) metastatic endometrial cancer. ESR1 mutations have emerged as a key mechanism of aromatase inhibitor (AI) resistance in HR + metastatic breast cancer and can be monitored using circulating tumor DNA (ctDNA). The aim of this study was to explore the incidence and clinical relevance of circulating ESR1 mutations in patients treated by AI or megestrol acetate (M) for advanced endometrial carcinoma. METHODOLOGY: This single-center retrospective study was performed at the Henri Becquerel Center (Rouen) and looked for circulating ESR1 gene mutations by droplet digital PCR (E380Q, L536R, Y537S, Y537N, Y537C, D538G, S463P) in patients with advanced HR + endometrial carcinoma treated between 2008 and 2020 for at least 30 days by AI or M. Analyses were performed before exposure and at progression/during endocrine therapy. RESULTS: Twenty-two patients were included: 13 were treated with AI, 12 of whom progressed; 9 patients were treated with M, 8 of whom progressed. 68.1% of the patients had low-grade endometrial carcinoma and 54.5% had received chemotherapy in the metastatic setting. The median duration of treatment was 152 days (min 47 - max 629) with AI and 155 days (min 91-max 1297) with M. Under AI, there was no ESR1 mutation at baseline, and one Y537C mutation at progression with a variant allele frequency (VAF) of 0.14%. Under M, one patient had a Y537C (VAF 0.2%) at baseline that disappeared during treatment. Another patient had a Y537S mutation emergence at progression after 91 days of treatment (VAF 1.83%). There was no significant difference between the circulating DNA concentration before and after hormone therapy (p = 0.16). CONCLUSION: ESR1 mutations do not seem to be involved in the mechanisms of resistance to AI or M in HR+ endometrial cancer. The clinical relevance of their detection is not demonstrated.

Results of a worldwide survey on the currently used histopathological diagnostic criteria for invasive lobular breast cancer.

Invasive lobular carcinoma (ILC) represents the second most common subtype of breast cancer (BC), accounting for up to 15% of all invasive BC. Loss of cell adhesion due to functional inactivation of E-cadherin is the hallmark of ILC. Although the current world health organization (WHO) classification for diagnosing ILC requires the recognition of the dispersed or linear non-cohesive growth pattern, it is not mandatory to demonstrate E-cadherin loss by immunohistochemistry (IHC). Recent results of central pathology review of two large randomized clinical trials have demonstrated relative overdiagnosis of ILC, as only ~60% of the locally diagnosed ILCs were confirmed by central pathology. To understand the possible underlying reasons of this discrepancy, we undertook a worldwide survey on the current practice of diagnosing BC as ILC. A survey was drafted by a panel of pathologists and researchers from the European lobular breast cancer consortium (ELBCC) using the online tool SurveyMonkey®. Various parameters such as indications for IHC staining, IHC clones, and IHC staining procedures were questioned. Finally, systematic reporting of non-classical ILC variants were also interrogated. This survey was sent out to pathologists worldwide and circulated from December 14, 2020 until July, 1 2021. The results demonstrate that approximately half of the institutions use E-cadherin expression loss by IHC as an ancillary test to diagnose ILC and that there is a great variability in immunostaining protocols. This might cause different staining results and discordant interpretations. As ILC-specific therapeutic and diagnostic avenues are currently explored in the context of clinical trials, it is of importance to improve standardization of histopathologic diagnosis of ILC diagnosis.

Deep learning analysis of contrast-enhanced spectral mammography to determine histoprognostic factors of malignant breast tumours.

OBJECTIVE: To evaluate if a deep learning model can be used to characterise breast cancers on contrast-enhanced spectral mammography (CESM). METHODS: This retrospective mono-centric study included biopsy-proven invasive cancers with an enhancement on CESM. CESM images include low-energy images (LE) comparable to digital mammography and dual-energy subtracted images (DES) showing tumour angiogenesis. For each lesion, histologic type, tumour grade, estrogen receptor (ER) status, progesterone receptor (PR) status, HER-2 status, Ki-67 proliferation index, and the size of the invasive tumour were retrieved. The deep learning model used was a CheXNet-based model fine-tuned on CESM dataset. The area under the curve (AUC) of the receiver operating characteristic (ROC) curve was calculated for the different models: images by images and then by majority voting combining all the incidences for one tumour. RESULTS: In total, 447 invasive breast cancers detected on CESM with pathological evidence, in 389 patients, which represented 2460 images analysed, were included. Concerning the ER, the deep learning model on the DES images had an AUC of 0.83 with the image-by-image analysis and of 0.85 for the majority voting. For the triple-negative analysis, a high AUC was observable for all models, in particularity for the model on LE images with an AUC of 0.90 for the image-by-image analysis and 0.91 for the majority voting. The AUC for the other histoprognostic factors was lower. CONCLUSION: Deep learning analysis on CESM has the potential to determine histoprognostic tumours makers, notably estrogen receptor status, and triple-negative receptor status. KEY POINTS: • A deep learning model developed for chest radiography was adapted by fine-tuning to be used on contrast-enhanced spectral mammography. • The adapted models allowed to determine for invasive breast cancers the status of estrogen receptors and triple-negative receptors. • Such models applied to contrast-enhanced spectral mammography could provide rapid prognostic and predictive information.

[2021 update of the GEFPICS' recommendations for HER2 status assessment in invasive breast cancer in France]. / Mise à jour 2021 des recommandations du GEFPICS pour l'évaluation du statut HER2 dans les cancers infiltrants du sein en France.

The last international guidelines on HER2 determination in breast cancer have been updated in 2018 by the American Society of Clinical Oncology and College of American Pathologists, on the basis of a twenty-year practice and results of numerous clinical trials. Moreover, the emerging HER2-low concept for 1+ and 2+ non amplified breast cancers lead to refine French practices for HER2 status assessment. The GEFPICS group, composed of expert pathologists, herein presents the latest French recommendations for HER2 status evaluation in breast cancer, taking into account the ASCO/CAP guidelines and introducing the HER2-low concept. In the era of personalized medicine, HER2 status assessment remains one of the most important biomarkers in breast cancer and its quality guaranties the optimal patients' care. French pathologists' commitment in theranostic biomarker quality is more than ever required to provide the most efficient cares in oncology.

[GEFPICS' guidelines for management of breast cancer tissue samples in the neoadjuvant setting]. / Recommandations du GEFPICS pour la prise en charge des prélèvements dans le cadre du traitement néoadjuvant du cancer du sein.

Neoadjuvant therapy is an increasing treatment option in the management of breast cancer. The tumor response to neoadjuvant therapy, especially the pathological complete response, is a validated endpoint frequently used in clinical trials. However, there is still a lack of standardization for the surgical specimen management in the neoadjuvant setting. This leads to heterogeneity in the specimen handling and might lead to significant bias for the prognostic assessment of patients or in clinical trials. The GEFPICS group, composed of expert breast cancer pathologists, herein presents guidelines for the management of breast and axillary specimen before treatment (management of biopsy, items of the pathological report) and after neoadjuvant therapy (specimen handling, histological assessment of response, items of the pathological report and response grading systems).

Treatment Response, Tumor Infiltrating Lymphocytes and Clinical Outcomes in Inflammatory Breast Cancer-Treated with Neoadjuvant Systemic Therapy.

Inflammatory breast cancer (IBC) is a rare (1%-5%), aggressive form of breast cancer, accounting for approximately 10% of breast cancer mortality. In the localized setting, standard of care is neoadjuvant chemotherapy (NACT) ± anti-HER2 therapy, followed by surgery. Here we investigated associations between clinicopathologic variables, stromal tumor-infiltrating lymphocytes (sTIL), and pathologic complete response (pCR), and the prognostic value of pCR. We included 494 localized patients with IBC treated with NACT from October 1996 to October 2021 in eight European hospitals. Standard clinicopathologic variables were collected and central pathologic review was performed, including sTIL. Associations were assessed using Firth logistic regression models. Cox regressions were used to evaluate the role of pCR and residual cancer burden (RCB) on disease-free survival (DFS), distant recurrence-free survival (DRFS), and overall survival (OS). Distribution according to receptor status was as follows: 26.4% estrogen receptor negative (ER-)/HER2-; 22.0% ER-/HER2+; 37.4% ER+/HER2-, and 14.1% ER+/HER2+. Overall pCR rate was 26.3%, being highest in the HER2+ groups (45.9% for ER-/HER2+ and 42.9% for ER+/HER2+). sTILs were low (median: 5.3%), being highest in the ER-/HER2- group (median: 10%). High tumor grade, ER negativity, HER2 positivity, higher sTILs, and taxane-based NACT were significantly associated with pCR. pCR was associated with improved DFS, DRFS, and OS in multivariable analyses. RCB score in patients not achieving pCR was independently associated with survival. In conclusion, sTILs were low in IBC, but were predictive of pCR. Both pCR and RCB have an independent prognostic role in IBC treated with NACT. SIGNIFICANCE: IBC is a rare, but very aggressive type of breast cancer. The prognostic role of pCR after systemic therapy and the predictive value of sTILs for pCR are well established in the general breast cancer population; however, only limited information is available in IBC. We assembled the largest retrospective IBC series so far and demonstrated that sTIL is predictive of pCR. We emphasize that reaching pCR remains of utmost importance in IBC.

Impact of preoperative staging with contrast-enhanced mammography for localized breast cancer management.

OBJECTIVE: A precise evaluation of the disease extent is mandatory before surgery for early breast cancer (EBC). Contrast-enhanced mammography (CEDM) is a recent technique that may help define adequate surgery. METHODS: This retrospective study included consecutive patients referred to a cancer center between November 2016 and July 2017 for biopsy-confirmed invasive EBC management. The primary objective was to evaluate the rate of surgical changes after incorporating the results of the preoperative staging examination, including CEDM. RESULTS: A total of 231 patients were screened for inclusion, and 132 patients were included, corresponding to 134 lesions. The first surgical plan was modified for 33 patients (25%), which represented 34 lesions. For 8 patients (6%), the surgery was cancelled in preference for neoadjuvant chemotherapy; for 16 patients (12.1%), the primary tumor procedure was enlarged; and for 23 patients (17.4%) the lymph node management was modified. Surgery was changed only due to the CEDM results for 24 patients (18.5%) and consisted of a more invasive procedure due to a more extended, multifocal or multicentric lesion than seen on the standard imaging. Anatomopathological surgery piece findings were well correlated with contrast-enhanced mammography results. Overall, there was no increase in the delay between the planned date of surgery and the effective surgical procedure (median 0 days). CONCLUSION: CEDM added to preoperative staging helped define better surgical management without increasing delay in the surgical procedure. ADVANCES IN KNOWLEDGE: CEDM is a reliable technique that should be considered as part of preoperative staging for EBC.

Circulating PIK3CA mutation detection at diagnosis in non-metastatic inflammatory breast cancer patients.

Inflammatory breast cancer (IBC) is an aggressive BC subtype with poor outcomes. A targetable somatic PIK3CA mutation is reported in 30% of IBC, allowing for treatment by PI3Kα-specific inhibitors, such as alpelisib. The aim of this study was to evaluate the detection rate of circulating PIK3CA mutation in locally-advanced IBC (LAIBC) patients harbouring a PIK3CA mutation on initial biopsy. This monocentric retrospective study was based on available stored plasma samples and tumour biopsies at diagnosis from all LAIBC patients treated with neo-adjuvant chemotherapy (NCT) between 2008 and 2018 at the Centre Henri Becquerel. PIK3CA mutations (E542K, E545K, H1047R/L) were assessed by droplet digital PCR (ddPCR) in plasma samples and tumoral tissue at diagnosis. A total of 55 patients were included. Overall, 14/55 patients (25%) had a PIK3CA mutation identified on baseline biopsy (H1047R = 8; H1047L = 3; E545K = 2; E542K = 1). Among them, 11 (79%) patients had enough DNA for circulating DNA analyses, and corresponding circulating PIK3CA mutations were found in 6/11 (55%). Among the 41 patients without PIK3CA mutations on biopsy, 32 (78%) had enough DNA for circulating DNA analysis, and no circulating PIK3CA mutation was identified. Our results revealed no prognostic or predictive value of PIK3CA mutations at the diagnosis of non-metastatic IBC but highlighted the prognostic value of the cfDNA rate at diagnosis. Our study showed that a corresponding circulating PIK3CA mutation was identified in 55% of LAIBC patients with PIK3CA-mutated tumours, while no circulating mutation was found among patients with PI3KCA wild-type tumours.

Phenotypic discordance between primary and metastatic breast cancer in the large-scale real-life multicenter French ESME cohort.

Expression of hormone receptor (HR) for estrogens (ER) and progesterone (PR) and HER2 remains the cornerstone to define the therapeutic strategy for breast cancer patients. We aimed to compare phenotypic profiles between matched primary and metastatic breast cancer (MBC) in the ESME database, a National real-life multicenter cohort of MBC patients. Patients with results available on both primary tumour and metastatic disease within 6 months of MBC diagnosis and before any tumour progression were eligible for the main analysis. Among the 16,703 patients included in the database, 1677 (10.0%) had available biopsy results at MBC diagnosis and on matched primary tumour. The change rate of either HR or HER2 was 27.0%. Global HR status changed (from positive = either ER or PR positive, to negative = both negative; and reverse) in 14.2% of the cases (expression loss in 72.5% and gain in 27.5%). HER2 status changed in 7.8% (amplification loss in 45.2%). The discordance rate appeared similar across different biopsy sites. Metastasis to bone, HER2+ and RH+/HER2- subtypes and previous adjuvant endocrine therapy, but not relapse interval were associated with an HR discordance in multivariable analysis. Loss of HR status was significantly associated with a risk of death (HR adjusted = 1.51, p = 0.002) while gain of HR and HER2 discordance was not. In conclusion, discordance of HR and HER2 expression between primary and metastatic breast cancer cannot be neglected. In addition, HR loss is associated with worse survival. Sampling metastatic sites is essential for treatment adjustment.

Reliability of Prognostic and Predictive Factors Evaluated by Needle Core Biopsies of Large Breast Invasive Tumors.

OBJECTIVES: Preoperative biopsy of breast cancer allows for prognostic/predictive marker assessment. However, large tumors, which are the main candidates for preoperative chemotherapy, are potentially more heterogeneous than smaller ones, which questions the reliability of histologic analyses of needle core biopsy (NCB) specimens compared with whole surgical specimens (WSS). We studied the histologic concordance between NCB specimens and WSS in tumors larger than 2 cm. METHODS: Early pT2 or higher breast cancers diagnosed between 2008 and 2011 in our center, with no preoperative treatments, were retrospectively screened. We assessed the main prognostic and predictive validated parameters. Comparisons were performed using the κ test. RESULTS: In total, 163 matched NCB specimens and WSS were analyzed. The correlation was excellent for ER and HER2 (κ = 0.94 and 0.91, respectively), moderate for PR (κ = 0.79) and histologic type (κ = 0.74), weak for Ki-67 (κ = 0.55), and minimal for SBR grade (κ = 0.29). Three of the 21 HER2-positive cases (14% of HER2-positive patients or 1.8% of all patients), by WSS analysis, were initially negative on NCB specimens even after chromogenic in situ hybridization. CONCLUSIONS: NCB for large breast tumors allowed reliable determination of ER/PR expression. However, the SBR grade may be deeply underestimated, and false-negative evaluation of the HER2 status would have led to a detrimental lack of trastuzumab administration.

CXCL12 and CXCR4, but not CXCR7, are primarily expressed by the stroma in head and neck squamous cell carcinoma.

The CXCL12/CXCR4 axis is involved in numerous models of metastatic dissemination, including head and neck squamous cell carcinoma (HNSCC). We assessed the relative expressions of CXCL12, CXCR4 and CXCR7 in the stroma and the tumour of HNSCC, and evaluated the methylation status of the CXCL12 promoter.Snap-frozen, HPV negative HNSCC samples were micro-dissected to isolate the tumoural and stromal compartments. The expression levels of CXCL12, CXCR4 and CXCR7 were assessed by qRT-PCR, and the methylation level of the CXCL12 promoter was evaluated by pyrosequencing.In total, 23 matched tumour/stroma samples were analysed. Higher expressions of CXCR4 and CXCL12 were observed in the stroma (p = 0.012 and p < 0.0001, respectively). No significant difference in expression was observed for CXCR7. A high methylation level (>40%) of the CXCL12 promoter was observed in only a few tumoural samples (5/23) and was associated with a lower expression of the gene (p = 0.03).Stromal cells, rather than the tumour itself, are mainly responsible for the expression of both CXCL12 and CXCR4 expression in HNSCC. CXCR7 expression did not differ between the two compartments and was not related to CXCL12 or CXCR4 expression. Finally, the methylation of the CXCL12 promoter could only explain the low intra-tumoural expression of this gene in 20% of cases.

The gene expression profile of inflammatory, hypoxic and metabolic genes predicts the metastatic spread of human head and neck squamous cell carcinoma.

OBJECTIVES: To assess the prognostic value of the expression profile of the main genes implicated in hypoxia, glucose and lactate metabolism, inflammation, angiogenesis and extracellular matrix interactions for the metastatic spread of head and neck squamous cell carcinoma. PATIENTS AND METHODS: Using a high-throughput qRT-PCR, we performed an unsupervised clustering analysis based on the expression of 42 genes for 61 patients. Usual prognostic factors and clustering analysis results were related to metastasis free survival. RESULTS: With a median follow-up of 48months, 19 patients died from a metastatic evolution of their head and neck squamous cell carcinoma and one from a local recurrence. The unsupervised clustering analysis distinguished two groups of genes that were related to metastatic evolution. A capsular rupture (p=0.005) and the "cluster CXCL12 low" (p=0.002) were found to be independent prognostic factors for metastasis free survival. Using a Linear Predictive Score methodology, we established a 9-gene model (VHL, PTGER4, HK1, SLC16A4, DLL4, CXCL12, CXCR4, PTGER3 and CA9) that was capable of classifying the samples into the 2 clusters with 90% accuracy. CONCLUSION: In this cohort, our clustering analysis underlined the independent prognostic value of the expression of a panel of genes involved in hypoxia and tumor environment. It allowed us to define a 9-gene model which can be applied routinely to classify newly diagnosed head and neck squamous cell carcinoma. If confirmed by an independent prospective study, this approach may help future clinical management of these aggressive tumors.

Delineation of small mobile tumours with FDG-PET/CT in comparison to pathology in breast cancer patients.

PURPOSE: Various segmentation methods for 18F-fluoro-2-deoxy-d-glucose (FDG) positron emission tomography/computed tomography (PET/CT) images were correlated with pathological volume in breast cancer patients as a model of small mobile tumours. METHODS: Thirty women with T2-T3/M0 breast invasive ductal carcinoma (IDC) were included prospectively. A FDG-PET/CT was acquired 4 ± 3d before surgery in prone and supine positions, with/without respiratory gating. The segmentation methods were as follows: manual (Vm), relative (Vt%) and adaptive (Va) standard uptake value (SUV) threshold and semi-automatic on CT (Vct). Pathological volumes (Vpath) were measured for 26 lesions. RESULTS: The mean (±SD) Vpath was 4.1 ± 2.9 mL, and the lesion displacements were 3.9 ± 2.8 mm (median value: 3 mm). The delineated VOIs did not vary with the acquisition position nor with respiration, regardless of the segmentation method. The Vm, Va, Vct and Vt% methods, except Vt30%, were correlated with Vpath (0.5