Initial MRI findings predictive of a pathological complete response to neoadjuvant treatments in HER2-positive breast cancers.

PURPOSE: This study aimed to determine if initial MRI findings could predict a pathological complete response (pCR) following neoadjuvant systemic therapy (NST) in HER2-positive breast cancers. METHODS: The study retrospectively included 111 patients (Center 1, training set) and 71 patients (Center 2, validation set) with HER2-positive cancer who underwent NST. Initial clinicopathological data and MRI findings were recorded. Continuous variables were analyzed using the Mann-Whitney and Student's t-tests, while categorical variables were analyzed using the χ2 or Fisher's exact test. Univariate analysis was conducted to determine the associations between these variables and pathological complete response (pCR), defined as the absence of invasive malignant cells in the breast and lymph nodes. Interobserver reproducibility was assessed for associated non-mass enhancement (NME) parameter by analyzing 50 MR studies (intraclass correlation coefficient). RESULTS: pCR was achieved in 67 patients, 51 (46 %) from Center 1 and 16 (23%) from Center 2 (p = 0.003), with significant differences between Centers 1 and 2 in tumor-infiltrating lymphocyte levels and lymphovascular invasion (p < 0.001). The initial presence of suspicious associated NME was the only significant parameter predictive of pCR (p < 0.001 for Center 1 and 0.04 for Center 2). The inter-observer reproducibility for this MRI feature was good, with an intraclass correlation coefficient of 0.872 (95 % CI: 0.73-1.00). CONCLUSION: The presence of suspicious associated NME in HER2-positive cancers on the initial MRI study was predictive of achieving pCR after NST. This significant preliminary finding warrants confirmation through prospective multicenter studies.

E-Cadherin Mutational Landscape and Outcomes in Breast Invasive Lobular Carcinoma.

Invasive lobular carcinomas (ILC) are characterized by the loss of E-cadherin expression and CDH1 gene inactivation. Diagnostic reproducibility for this tumor type is currently suboptimal and could be improved by a better understanding of its histomolecular and clinical heterogeneity. We have analyzed the relationship between the presence, type, or position of CDH1 mutations, E-cadherin expression, and clinicopathological features (including outcome) in a retrospective series of 251 primary ILC with a long follow-up (median: 9.5 years). The mutational status of E-cadherin gene (CDH1) was determined by RNA sequencing from frozen tumor samples. E-cadherin immunohistochemistry (IHC) was performed with antibodies directed against the intracellular domain (clone 4A2C7) and the extracellular domain (clone NCH38). IHC expression of p120 and ß-catenin was also assessed in E-cadherin diffusely positive cases. Three major patterns of E-cadherin membrane expression were identified by IHC, with good agreement between the 2 clones (overall concordance: 83.8%, Kappa 0.67): null/focal expression (≤10%) (72.8% of cases for 4A2C7 and 83.8% for NCH38), heterogeneous expression (11%-89%) (19.2% of cases for 4A2C7 and 6.9% for NCH38), and diffuse expression (≥90%) (8% of cases for 4A2C7 and 9.3% for NCH38). E-cadherin membranous expression, when present, was abnormal (incomplete labeling and/or reduced intensity). ILC with diffuse E-cadherin expression showed abnormal ß-catenin or p120-catenin staining in 21% of cases. Interestingly, these cases with diffusely expressed E-cadherin had a CDH1 mutation rate as high as the E-cadherin null/focal cases (∼70%) but were enriched in nontruncating mutations. Regarding CDH1 mutation location, intracytoplasmic domain mutations correlated with a divergent E-cadherin IHC phenotype between the 2 antibodies (4A2C7 ≤ 10%/NCH38 ≥ 10%). Clinico-pathological correlation analyses found that stromal amount (inversely correlated with tumor cellularity) and tumor-infiltrating lymphocytes were less abundant in ILC with E-cadherin null/focal cases. In addition, CDH1 truncating mutations were associated with radiohistologic size discordance and were identified in multivariate survival analysis as an independent poor prognosis factor in terms of metastasis risk and breast cancer-related mortality. Overall, our study highlights the importance of the precise mutational status of CDH1 in the clinical, radiological, histologic, and phenotypic expression of lobular carcinomas. These findings should be taken into account in future attempts to improve diagnostic criteria or methods for ILC, as well as for clinicobiological studies dedicated to this tumor type.

Deciphering the spatial landscape and plasticity of immunosuppressive fibroblasts in breast cancer.

Although heterogeneity of FAP+ Cancer-Associated Fibroblasts (CAF) has been described in breast cancer, their plasticity and spatial distribution remain poorly understood. Here, we analyze trajectory inference, deconvolute spatial transcriptomics at single-cell level and perform functional assays to generate a high-resolution integrated map of breast cancer (BC), with a focus on inflammatory and myofibroblastic (iCAF/myCAF) FAP+ CAF clusters. We identify 10 spatially-organized FAP+ CAF-related cellular niches, called EcoCellTypes, which are differentially localized within tumors. Consistent with their spatial organization, cancer cells drive the transition of detoxification-associated iCAF (Detox-iCAF) towards immunosuppressive extracellular matrix (ECM)-producing myCAF (ECM-myCAF) via a DPP4- and YAP-dependent mechanism. In turn, ECM-myCAF polarize TREM2+ macrophages, regulatory NK and T cells to induce immunosuppressive EcoCellTypes, while Detox-iCAF are associated with FOLR2+ macrophages in an immuno-protective EcoCellType. FAP+ CAF subpopulations accumulate differently according to the invasive BC status and predict invasive recurrence of ductal carcinoma in situ (DCIS), which could help in identifying low-risk DCIS patients eligible for therapeutic de-escalation.

The prognosis of patients treated with everolimus for advanced ER-positive, HER2-negative breast cancer is driven by molecular features.

Everolimus is widely used in patients with advanced ER-positive, HER2-negative breast cancer. We looked at alterations in the PIK3CA/AKT/mTOR pathway in a multicenter cohort as potential biomarkers of efficacy. Patients with advanced ER-positive, HER2-negative breast cancer treated with everolimus and endocrine therapy between 2012 and 2014 in two cancer centers were included. Targeted sequencing examined mutations in PIK3CA, ESR1, and AKT1 genes. An immunochemical analysis was conducted to evaluate expression of PTEN, INPP4B, STK11, p4EBP1, and pS6. We analyzed 71 patients (44 primary tumors; 27 metastatic tissues). Median age was 63 years [58-69]. All patients had heavily pretreated advanced disease. A mutation in the PIK3CA pathway was observed in 32 samples (PIK3CA exons 10 and 21 and AKT1 exon 4 in 15.5%, 24.0%, and 5.6% of samples), and in ESR1 in 5 samples (7.0%), respectively. Most samples showed cytoplasmic expression of the PIK3CA pathway proteins. Progression-free survival was longer in patients with a pS6 or p4EBP1 histoscore ≥ median value (6.6 versus 3.7 months, p = 0.037), and in patients with a PTEN histoscore ≤ median value (7.1 versus 5.3 months, p = 0.02). Overall survival was longer in patients with pS6 ≥ 3rd quartile (27.6 versus 19.3 months, p = 0.038) and in patients with any mutation in the PIK3CA/AKT/mTOR pathway (27.6 versus 19.3 months, p = 0.011). The prognosis of patients treated with everolimus for advanced ER-positive, HER2-negative breast cancer appears primarily driven by molecular features associated with the activation of the PIK3CA/AKT/mTOR pathway.

A novel bioinformatic approach reveals cooperation between Cancer/Testis genes in basal-like breast tumors.

Breast cancer is the most prevalent type of cancer in women worldwide. Within breast tumors, the basal-like subtype has the worst prognosis, prompting the need for new tools to understand, detect, and treat these tumors. Certain germline-restricted genes show aberrant expression in tumors and are known as Cancer/Testis genes; their misexpression has diagnostic and therapeutic applications. Here we designed a new bioinformatic approach to examine Cancer/Testis gene misexpression in breast tumors. We identify several new markers in Luminal and HER-2 positive tumors, some of which predict response to chemotherapy. We then use machine learning to identify the two Cancer/Testis genes most associated with basal-like breast tumors: HORMAD1 and CT83. We show that these genes are expressed by tumor cells and not by the microenvironment, and that they are not expressed by normal breast progenitors; in other words, their activation occurs de novo. We find these genes are epigenetically repressed by DNA methylation, and that their activation upon DNA demethylation is irreversible, providing a memory of past epigenetic disturbances. Simultaneous expression of both genes in breast cells in vitro has a synergistic effect that increases stemness and activates a transcriptional profile also observed in double-positive tumors. Therefore, we reveal a functional cooperation between Cancer/Testis genes in basal breast tumors; these findings have consequences for the understanding, diagnosis, and therapy of the breast tumors with the worst outcomes.

Molecular and Clinical Portrait of HER2-low Invasive Lobular Carcinomas.

Invasive lobular carcinomas (ILCs) have a low frequency of ERBB2 amplification, therefore restricting the use of conventional anti-HER2 therapies for this histologic special type. Conversely, ILCs with low HER2 overexpression may represent a broader target for the use of emerging antibody drug conjugate therapies targeting HER2, since these treatments have proven effective in HER2-low breast cancers. Very scarce data about HER2-low ILCs have been so far published, although these tumors could have different prevalence and histomolecular specificities compared with invasive breast carcinoma of no special type (IBC-NST). Our aims in that context were to decipher the clinicopathological and molecular features of a large series of HER2-low ILCs. Comparative evaluation of HER2-low prevalence was done based on a retrospective series of 7970 patients from Institut Curie, with either primary invasive lobular (N = 1103) or no special type (N = 6867) invasive carcinoma. Clinicopathological and molecular analyses of HER2-zero, HER2-low, and HER2-positive ILCs were performed on a subgroup of 251 patients who underwent surgery for a primary ILC between 2005 and 2008. The mutational profile of these 251 cases was determined from RNAseq data. Compared with HER2-negative IBC-NSTs, the HER2-negative ILCs were found to display a higher frequency of HER2-zero cases (59.4% vs 53.7%) and a lower frequency of HER2-low (40.6% vs 46.3%) (P < .001). Clinicopathological features associated with HER2-low status (vs HER2-zero) in ILC were older age, postmenopausal status, nonclassic ILC histological types, higher grade, proliferation, and estrogen receptor expression levels. Survival curve analysis showed a significantly lower risk of local recurrence for HER2-low (vs HER2-zero) ILCs, but no association was found between HER2 status and either breast cancer-specific survival or distant metastasis-free interval. ERBB3 was the unique mutated gene exclusively associated with HER2-low ILCs yet being mutated at a low frequency (7.1%) (false discovery rate < 0.05). In conclusion, HER2-low ILCs exhibit their own particularities, both on clinical-pathological and molecular levels. Our findings call for larger multicenter validation studies.

Residual ANTXR1+ myofibroblasts after chemotherapy inhibit anti-tumor immunity via YAP1 signaling pathway.

Although cancer-associated fibroblast (CAF) heterogeneity is well-established, the impact of chemotherapy on CAF populations remains poorly understood. Here we address this question in high-grade serous ovarian cancer (HGSOC), in which we previously identified 4 CAF populations. While the global content in stroma increases in HGSOC after chemotherapy, the proportion of FAP+ CAF (also called CAF-S1) decreases. Still, maintenance of high residual CAF-S1 content after chemotherapy is associated with reduced CD8+ T lymphocyte density and poor patient prognosis, emphasizing the importance of CAF-S1 reduction upon treatment. Single cell analysis, spatial transcriptomics and immunohistochemistry reveal that the content in the ECM-producing ANTXR1+ CAF-S1 cluster (ECM-myCAF) is the most affected by chemotherapy. Moreover, functional assays demonstrate that ECM-myCAF isolated from HGSOC reduce CD8+ T-cell cytotoxicity through a Yes Associated Protein 1 (YAP1)-dependent mechanism. Thus, efficient inhibition after treatment of YAP1-signaling pathway in the ECM-myCAF cluster could enhance CD8+ T-cell cytotoxicity. Altogether, these data pave the way for therapy targeting YAP1 in ECM-myCAF in HGSOC.

Biopathologic Characterization and Grade Assessment of Breast Cancer With 3-D Multiparametric Ultrasound Combining Shear Wave Elastography and Backscatter Tensor Imaging.

OBJECTIVE: Despite recent improvements in medical imaging, the final diagnosis and biopathologic characterization of breast cancers currently still requires biopsies. Ultrasound is commonly used for clinical examination of breast masses. B-Mode and shear wave elastography (SWE) are already widely used to detect suspicious masses and differentiate benign lesions from cancers. But additional ultrasound modalities such as backscatter tensor imaging (BTI) could provide relevant biomarkers related to tissue organization. Here we describe a 3-D multiparametric ultrasound approach applied to breast carcinomas in the aims of (i) validating the ability of BTI to reveal the underlying organization of collagen fibers and (ii) assessing the complementarity of SWE and BTI to reveal biopathologic features of diagnostic interest. METHODS: Three-dimensional SWE and BTI were performed ex vivo on 64 human breast carcinoma samples using a linear ultrasound probe moved by a set of motors. Here we describe a 3-D multiparametric representation of the breast masses and quantitative measurements combining B-mode, SWE and BTI. RESULTS: Our results reveal for the first time that BTI can capture the orientation of the collagen fibers around tumors. BTI was found to be a relevant marker for assessing cancer stages, revealing a more tangent tissue orientation for in situ carcinomas than for invasive cancers. In invasive cases, the combination of BTI and SWE parameters allowed for classification of invasive tumors with respect to their grade with an accuracy of 95.7%. CONCLUSION: Our results highlight the potential of 3-D multiparametric ultrasound imaging for biopathologic characterization of breast tumors.