Clinicopathologic and genetic analysis of invasive breast carcinomas in women with germline CHEK2 variants.

PURPOSE: Germline pathogenic variants in checkpoint kinase 2 (CHEK2) are associated with a moderately increased risk of breast cancer (BC). The spectrum of clinicopathologic features and genetics of these tumors has not been fully established. METHODS: We characterized the histopathologic and clinicopathologic features of 44 CHEK2-associated BCs from 35 women, and assessed responses to neoadjuvant chemotherapy. A subset of cases (n = 23) was additionally analyzed using targeted next-generation DNA sequencing (NGS). RESULTS: Most (94%, 33/35) patients were heterozygous carriers for germline CHEK2 variants, and 40% had the c.1100delC allele. Two patients were homozygous, and five had additional germline pathogenic variants in ATM (2), PALB2 (1), RAD50 (1), or MUTYH (1). CHEK2-associated BCs occurred in younger women (median age 45 years, range 25-75) and were often multifocal (20%) or bilateral (11%). Most (86%, 38/44) were invasive ductal carcinomas of no special type (IDC-NST). Almost all (95%, 41/43) BCs were ER + (79% ER + HER2-, 16% ER + HER2 + , 5% ER-HER2 +), and most (69%) were luminal B. Nottingham grade, proliferation index, and results of multiparametric molecular testing were heterogeneous. Biallelic CHEK2 alteration with loss of heterozygosity was identified in most BCs (57%, 13/23) by NGS. Additional recurrent alterations included GATA3 (26%), PIK3CA (226%), CCND1 (22%), FGFR1 (22%), ERBB2 (17%), ZNF703 (17%), TP53 (9%), and PPM1D (9%), among others. Responses to neoadjuvant chemotherapy were variable, but few patients (21%, 3/14) achieved pathologic complete response. Most patients (85%) were without evidence of disease at time of study (n = 34). Five patients (15%) developed distant metastasis, and one (3%) died (mean follow-up 50 months). CONCLUSION: Almost all CHEK2-associated BCs were ER + IDC-NST, with most classified as luminal B with or without HER2 overexpression. NGS supported the luminal-like phenotype and confirmed CHEK2 as an oncogenic driver in the majority of cases. Responses to neoadjuvant chemotherapy were variable but mostly incomplete.

Targeted DNA Sequencing in Diagnosis of Malignant Phyllodes Tumors With Emphasis on Tumors With Keratin and p63 Expression.

The differential diagnosis of malignant spindle cell neoplasms in the breast most frequently rests between malignant phyllodes tumor (MPT) and metaplastic carcinoma (MBC). Diagnosis of MPT can be challenging due to diffuse stromal overgrowth, keratin (CK) and/or p63 immunopositivity, and absent CD34 expression, which can mimic MBC, especially in core biopsies. Distinction of MPT from MBC has clinical implications, with differences in surgical approach, chemotherapy, and radiation. In this study, we evaluated MPTs (78 tumors, 64 patients) for stromal CK, p63, and CD34 expression and profiled a subset (n = 31) by targeted next-generation DNA sequencing, with comparison to MBC (n = 44). Most MPTs (71%) were CK+ and/or p63+, including 32% CK+ (25/77 focal) and 65% p63+ (32/66 focal, 10/66 patchy, and 1/66 diffuse). Thirty percent of MPTs expressed both CK and p63 (20/66), compared with 95% of MBCs (40/42, P < .001). CK and/or p63 were positive in CD34+ and CD34- MPTs. Recurrent genetic aberrations in MPTs involved TERT, TP53, MED12, CDKN2A, chromatin modifiers, growth factor receptors/ligands, and phosphoinositide-3 kinase (PI-3K) and MAPK pathway genes. Only MED12 (39%, 12/31) and SETD2 (13%, 4/31) were exclusively mutated in MPTs and not MBCs (P < .001 and P = .044, respectively), whereas PIK3R1 mutations were only found in MBCs (37%, 13/35, P < .001). Comparative literature review additionally identified ARID1B, EGFR, FLNA, NRAS, PDGFRB, RAD50, and RARA alterations enriched or exclusively in MPTs vs MBCs. MED12 was mutated in MPTs with diffuse stromal overgrowth (53%, 9/17), CD34- MPTs (41%, 7/17), and CK+ and/or p63+ MPTs (39%, 9/23), including 36% of CD34- MPTs with CK and/or p63 expression. Overall, MED12 mutation and/or CD34 expression were observed in 68% (21/31) MPTs, including 61% (14/23) of CK+ and/or p63+ tumors. Our results emphasize the prevalence of CK and p63 expression in MPTs and demonstrate the diagnostic utility of next-generation DNA sequencing, especially in MPTs with confounding factors that can mimic MBC.

RB1 Genetic Alterations in Estrogen Receptor-Positive Breast Carcinomas: Correlation With Neuroendocrine Differentiation.

Genetic alterations in the retinoblastoma susceptibility gene (RB1) are present in up to 40% of triple-negative breast cancers (BCs) and frequent in tumors with neuroendocrine differentiation, including small cell neuroendocrine carcinoma. Data on RB1 genetic alterations in estrogen receptor (ER)-positive BCs are scarce. In this study, we sought to define the morphologic, immunohistochemical, and genetic features of ER-positive BCs harboring somatic alterations in RB1, with emphasis on neuroendocrine differentiation. ER-positive BCs with pathogenic RB1 genetic alterations were identified in <1% of cases (N = 55) from a cohort of 6026 BCs previously subjected to targeted next-generation sequencing, including 23 primary BCs (pBCs) and 32 recurrent/metastatic BCs (mBCs). In cases where loss of heterozygosity of the wild-type RB1 allele could be assessed (93%, 51/55), most pBCs (82%, 18/22) and mBCs (90%, 26/29) exhibited biallelic RB1 inactivation, primarily through loss-of-function mutation and loss of heterozygosity (98%, 43/44). Upon histologic review, a subset of RB1-altered tumors exhibited neuroendocrine morphology (13%, 7/55), which correlated with expression of neuroendocrine markers (39%, 9/23) in both pBCs (27%, 3/11) and mBCs (50%, 6/12). Loss of Rb protein expression was observed in BCs with biallelic RB1 loss only, with similar frequency in pBCs (82%, 9/11) and mBCs (75%, 9/12). All cases with neuroendocrine marker expression (n = 9) and/or neuroendocrine morphology (n = 7) harbored biallelic genetic inactivation of RB1 and exhibited Rb loss of expression. TP53 (53%, 29/55) and PIK3CA (45%, 25/55) were the most frequently comutated genes across the cohort. Overall, these findings suggest that ER-positive BCs with biallelic RB1 genetic alterations frequently exhibit Rb protein loss, which correlates with neuroendocrine differentiation in select BCs. This study provides insights into the molecular and phenotypic heterogeneity of BCs with RB1 genetic inactivation, underscoring the need for further research into the potential clinical implications associated with these tumors.

Malignant phyllodes tumour with lymph node metastasis: a diagnostic conundrum resolved by next generation DNA sequencing.

A malignant neoplasm with spindle cell and chondroid differentiation in the breast, metastatic to lymph node. In this context, a metaplastic carcinoma is typically favored given the exceptional nature of lymph node metastases in malignant phyllodes tumors (MPT). However, we demonstrate pathognomonic hotspot mutations in MED12 and the promoter of the TERT gene by targeted next-generation DNA sequencing, supporting a diagnosis of MPT.

Salivary gland-like tumors of the breast: Histopathologic and genetic features with clinical implications.

Salivary gland-like tumors of the breast are rare neoplasms that share morphologic, immunophenotypic, and/or genetic features with their salivary gland counterparts, highlighting a shared underlying histopathogenesis in most cases. Salivary gland-like carcinomas included in the World Health Organization classification of breast tumors are adenoid cystic carcinoma, secretory carcinoma, mucoepidermoid carcinoma, acinic cell carcinoma, and the exceedingly rare polymorphous adenocarcinoma. These carcinomas are usually triple negative for estrogen receptor and progesterone receptor expression and HER2 overexpression, yet generally have favorable prognosis, in contrast to high-grade triple negative carcinomas of no special type. On the other hand, a small subset, such as solid-basaloid adenoid cystic carcinoma, rare high-grade carcinomas, and those associated with transformation to other types of high-grade invasive carcinoma can behave more aggressively. Other salivary gland-like tumors of the breast, such as pleomorphic adenoma and adenomyoepithelioma, are usually benign but can rarely undergo malignant transformation. Although clinical experience with salivary gland-like breast tumors is overall limited, their recognition and accurate classification has important implications for prognosis and clinical management, especially to avoid overtreatment of salivary gland-like carcinomas. The identification of characteristic genetic alterations and/or immunohistochemical surrogates in many of these tumors has practical applications to establishing an accurate diagnosis and directing clinical management. This review highlights the histopathologic and genetic characteristics of salivary gland-like breast tumors and the implications of the diagnosis for current clinical management.

HER-2 low status in early-stage invasive lobular carcinoma of the breast: associated factors and outcomes in an institutional series.

PURPOSE: HER2 overexpression has a central role in breast cancer carcinogenesis and is associated with poor prognosis if untreated. Lately, identification of HER2-low breast cancer has been proposed to select patients for novel HER2-directed chemotherapy and includes cancers with immunohistochemistry 1 + or 2 + with negative FISH, encompassing approximately 55-60% of all breast carcinomas. In early-stage breast cancer, the prognostic significance of HER2 low-disease is less well understood, with a particular paucity of data evaluating the prevalence and implications of HER2-low status in invasive lobular carcinoma (ILC). METHODS: We evaluated 666 stage I-III ILC tumors from a prospectively maintained institutional database, comparing clinicopathologic features and disease-free survival (DFS) using a multivariable Cox proportional hazards model. RESULTS: HER2-low status was common in this cohort of patients with ILC, but most clinicopathologic features did not differ between HER2-low and HER2-negative cases. However, when adjusting for tumor size, number of positive nodes, ER/PR status, and local therapy received, patients with HER2-low status had worse disease-free survival (DFS) than those with HER2-negative tumors (hazard ratio 2.0, 95% confidence interval 1.0-4.1, p = 0.05). CONCLUSION: This difference in DFS supports the notion that HER2-low and HER2-negative early stage ILC may differ clinically, despite similar clinicopathologic features. Further investigation into the potential benefit of HER2 targeted therapy in HER2-low early-stage breast cancer, and specifically lobular cancer, is warranted to ensure optimal outcomes in this distinct tumor subtype.

Quantifying hormone receptor status in lobular breast cancer in an institutional series: the relationship between estrogen and progesterone receptor status and outcomes.

PURPOSE: Recent guidelines defined a new reporting category of ER-low-positive breast cancer based on immunohistochemistry (IHC). While low positivity of either hormone receptor is uncommon in invasive lobular carcinoma (ILC), we sought to investigate whether relatively low hormone receptor positivity was associated with tumor characteristics and patient outcomes in a single institutional cohort. METHODS: We searched an institutional database for cases of stage I-III ILC with available IHC reports. Based on prior published categories in ILC, ER was classified as low, medium, or high as defined by ER staining of 10-69%, 70-89%, and ≥ 90% respectively. PR low and high tumors were defined by < 20%, or ≥ 20% staining respectively. We used chi-squared tests, t-tests, and Cox proportional hazards models to evaluate associations between ER/PR categories and tumor characteristics or disease-free survival (DFS). RESULTS: The cohort consisted of 707 ILC cases, with 11% of cases categorized as ER low, 15.1% as medium, and 73.8% as high. The majority (67.6%) were PR high. Patients with ER low/medium expression were significantly younger, and more likely to also have PR low and/or HER2 positive tumors compared to those that were ER high. In a Cox proportional hazards model adjusting for age, stage, grade, pleomorphic histology, and treatment, ER category was not prognostic for DFS, but PR negative and PR low status each had significantly worse DFS compared to PR high status (HR 3.5, 95% CI 1.8-6.7, p < 0.001; and HR 2.0, 95% CI 1.1-3.5, p = 0.015, respectively). CONCLUSION: These findings highlight the relevance of quantifying ER and PR within ILC.

Solid-Basaloid Adenoid Cystic Carcinoma of the Breast: An Aggressive Subtype Enriched for Notch Pathway and Chromatin Modifier Mutations With MYB Overexpression.

Adenoid cystic carcinoma (AdCC) is a rare triple-negative breast cancer analogous to its extramammary counterparts. Diagnosis of the more aggressive solid-basaloid variant of AdCC (SB-AdCC) can be challenging due to poorly defined histopathologic and molecular features. We characterized 22 invasive and in situ basaloid carcinomas by morphology, immunohistochemistry, genetics, and MYB status using multiple platforms and assessed clinical behavior and neoadjuvant chemotherapy responses. After consensus review, 16/22 cases were classified as SB-AdCC. All SB-AdCC had predominantly solid growth and at least focal myxohyaline stroma and were immune-poor. Eosinophilic squamoid cells (69%, 11/16) and basement membrane-like secretions (69%, 11/16) were common, and intercalated ducts (31%, 5/16) were less frequent. SB-AdCC typically expressed SOX10 (100%, 16/16) and luminal markers (100%, 16/16 CK7; 88%, 14/16 CD117; 93%, 13/14 CAM5.2). SMA (40%, 6/15) expression was less common, and SMM (27%, 3/11), GATA3 (20%, 3/15), and p63 (25%, 4/16) were mostly negative. MYB protein and/or MYB RNA overexpression was universal in evaluable cases (13/13), with RNA in situ hybridization (10/10) more reliable than immunohistochemistry (10/11, plus 4 excisions inconclusive). Fluorescence in situ hybridization and/or next-generation sequencing identified MYB rearrangements (20%, 3/15) and amplifications/copy gains (60%, 9/15) but no MYB::NFIB fusions. SB-AdCC often had aberrations in Notch pathway (60%, including 40% NOTCH1 and 20% NOTCH2) and/or chromatin modifier (60%, including 33% CREBBP) genes, with relatively infrequent TP53 mutations (27%). Unclassified invasive basaloid carcinomas lacking described histologic features of SB-AdCC (n = 4) and basaloid ductal carcinoma in situ (n = 2) showed similar immunoprofiles and genetics as SB-AdCC, including Notch aberrations and MYB overexpression with MYB rearrangements/amplifications. Overall, nodal (22%) and distant (33%) metastases were common, and 23% of patients died of disease (mean follow-up, 35 months; n = 22). Responses were poor in all 7 neoadjuvant chemotherapy-treated patients, without any achieving pathologic complete response. The data highlight the histopathologic spectrum of basaloid carcinomas including SB-AdCC and reveal shared genetics and MYB activation, which can be diagnostically useful. Aggressive behavior and poor treatment responses emphasize a need for additional treatment approaches.

Triple-Negative Apocrine Carcinomas: Toward a Unified Group With Shared Molecular Features and Clinical Behavior.

Triple-negative apocrine carcinomas (TNACs) are rare breast tumors with limited studies evaluating their molecular characteristics and clinical behavior. We performed a histologic, immunohistochemical, genetic, and clinicopathologic assessment of 42 invasive TNACs (1 with a focal spindle cell component) from 41 patients, 2 pure apocrine ductal carcinomas in situ (A-DCIS), and 1 A-DCIS associated with spindle cell metaplastic carcinoma (SCMBC). All TNACs had characteristic apocrine morphology and expressed androgen receptor (42/42), gross cystic disease fluid protein 15 (24/24), and CK5/6 (16/16). GATA3 was positive in most cases (16/18, 89%), and SOX10 was negative (0/22). TRPS1 was weakly expressed in a minority of tumors (3/14, 21%). Most TNACs had low Ki67 proliferation (≤10% in 67%, 26/39), with a median index of 10%. Levels of tumor infiltrating lymphocytes were low (≤10% in 93%, 39/42, and 15% in 7%, 3/42). Eighteen percent of TNACs presented with axillary nodal metastasis (7/38). No patients treated with neoadjuvant chemotherapy achieved pathologic complete response (0%, 0/10). Nearly all patients with TNAC (97%, n = 32) were without evidence of disease at the time of study (mean follow-up of 62 months). Seventeen invasive TNACs and 10 A-DCIS (7 with paired invasive TNAC) were profiled by targeted capture-based next-generation DNA sequencing. Pathogenic mutations in phosphatidylinositol 3-kinase pathway genes PIK3CA (53%) and/or PIK3R1 (53%) were identified in all TNACs (100%), including 4 (24%) with comutated PTEN. Ras-MAPK pathway genes, including NF1 (24%), and TP53 were mutated in 6 tumors each (35%). All A-DCIS shared mutations, such as phosphatidylinositol 3-kinase aberrations and copy number alterations with paired invasive TNACs or SCMBC, and a subset of invasive carcinomas showed additional mutations in tumor suppressors (NF1, TP53, ARID2, and CDKN2A). Divergent genetic profiles between A-DCIS and invasive carcinoma were identified in 1 case. In summary, our findings support TNAC as a morphologically, immunohistochemically, and genetically homogeneous subgroup of triple-negative breast carcinomas and suggest overall favorable clinical behavior.

The clinical behavior and genomic features of the so-called adenoid cystic carcinomas of the solid variant with basaloid features.

Classic adenoid cystic carcinomas (C-AdCCs) of the breast are rare, relatively indolent forms of triple negative cancers, characterized by recurrent MYB or MYBL1 genetic alterations. Solid and basaloid adenoid cystic carcinoma (SB-AdCC) is considered a rare variant of AdCC yet to be fully characterized. Here, we sought to determine the clinical behavior and repertoire of genetic alterations of SB-AdCCs. Clinicopathologic data were collected on a cohort of 104 breast AdCCs (75 C-AdCCs and 29 SB-AdCCs). MYB expression was assessed by immunohistochemistry and MYB-NFIB and MYBL1 gene rearrangements were investigated by fluorescent in-situ hybridization. AdCCs lacking MYB/MYBL1 rearrangements were subjected to RNA-sequencing. Targeted sequencing data were available for 9 cases. The invasive disease-free survival (IDFS) and overall survival (OS) were assessed in C-AdCC and SB-AdCC. SB-AdCCs have higher histologic grade, and more frequent nodal and distant metastases than C-AdCCs. MYB/MYBL1 rearrangements were significantly less frequent in SB-AdCC than C-AdCC (3/14, 21% vs 17/20, 85% P < 0.05), despite the frequent MYB expression (9/14, 64%). In SB-AdCCs lacking MYB rearrangements, CREBBP, KMT2C, and NOTCH1 alterations were observed in 2 of 4 cases. SB-AdCCs displayed a shorter IDFS than C-AdCCs (46.5 vs 151.8 months, respectively, P < 0.001), independent of stage. In summary, SB-AdCCs are a molecularly heterogeneous but clinically aggressive group of tumors. Less than 25% of SB-AdCCs display the genomic features of C-AdCC. Defining whether these tumors represent a single entity or a collection of different cancer types with a similar basaloid histologic appearance is warranted.

Histologic and genomic features of breast cancers with alterations affecting the SWI/SNF (SMARC) genes.

The SWI/SNF family of proteins is a multisubunit ATPase complex frequently altered in human cancer. Inactivating mutations in SWI/SNF-related matrix-associated actin-dependent regulator of chromatin (SMARCs) underpin a subset of tumors such as the malignant rhabdoid tumor and small cell carcinoma of the ovary, hypercalcemic type. Here, we investigated the genotypic and phenotypic characteristics of breast cancers harboring somatic genetic alterations affecting genes of the SMARC family. We analyzed a series of 6026 primary and metastatic breast cancers subjected to targeted-capture sequencing. SMARC core subunit (SMARCA4, SMARCB1, and SMARCA2) alterations were identified in <1% of all breast cancers, consisting of 27 primary and 30 recurrent/metastatic tumors. The majority of SMARC alterations were monoallelic mutations (47/57, 82%) and thus categorized into two groups: Class 1 alterations consisting of potentially pathogenic mutations and rearrangements and Class 2 alterations consisting of missense mutations and small in-frame deletions of unknown significance. Biallelic events in a SMARC gene were present in a minority of cases (10/57, 18%). Histologic patterns in the form of rhabdoid, composite rhabdoid, sarcomatoid or anaplastic features were observed in a subset of Class 1 primary and metastatic tumors (7/57, 12%). SMARC protein was preserved in nearly all tumors analyzed with immunohistochemistry (26/30, 87%). Four Class 1 tumors demonstrated altered SMARC protein expression in the form of loss (1/30, 3%) or mosaic pattern (3/30, 10%). Complete loss of SMARCA2 (BRM) was observed in a sole tumor with composite rhabdoid morphology, and biallelic hits in the SMARCA2 gene. The genomic landscape of both primary Class 1 and 2 breast cancers did not reveal any characteristic findings. In summary, SMARC alterations likely contribute to the biology of a rare subset of breast cancers in the form of biallelic or pathogenic alterations in SMARC, as evidenced by SMARC-deficient phenotype or altered expression of SMARC protein.

Gynaecological perivascular epithelioid cell tumour (PEComa): comparative analysis of proposed algorithms for prediction of clinical outcome.

AIMS: Perivascular epithelioid cell tumours (PEComas) are rare mesenchymal tumours that coexpress smooth muscle and melanocytic markers. They have a predilection for gynaecological organs, where they present a unique diagnostic challenge, because of morphological and immunohistochemical overlap with more common smooth muscle and stromal tumours. Limited information regarding the natural history, owing to the rarity of this tumour, makes accurate risk stratification difficult. We aimed to review clinicopathological features of gynaecological PEComa and compare accuracy of five different classification systems for prediction of prognosis. METHODS AND RESULTS: We have described the clinicopathological features of 13 new cases and tested five prognostic algorithms in a total of 67 cases of gynaecological PEComa. Receiver operating characteristic curves were constructed and areas under the curve (AUCs) were calculated to evaluate predictive accuracy. The modified gynaecological-specific algorithm showed high sensitivity and specificity and yielded the highest AUC (0.864). It's earlier version, the gynaecological-specific algorithm, suffered from lower specificity (AUC = 0.843). The post-hoc McNemar test confirmed significant differences between the performances of the modified gynaecological-specific algorithm and the gynaecological-specific algorithm (P = 0.008). The original Folpe algorithm for PEComas of all sites showed low specificity, had a lower AUC (0.591), and was inapplicable in 18% of cases. Its two later versions (the revised Folpe algorithm and the modified Folpe algorithm) also yielded lower AUCs (0.690 and 0.591, respectively). CONCLUSION: We have shown that the modified gynaecological-specific algorithm predicts the clinical outcome of gynaecological PEComa with high accuracy, and have validated its use for prognostic stratification of gynaecological PEComa.

Interaction of photoperiod and vernalization determines flowering time of Brachypodium distachyon.

Timing of flowering is key to the reproductive success of many plants. In temperate climates, flowering is often coordinated with seasonal environmental cues such as temperature and photoperiod. Vernalization is an example of temperature influencing the timing of flowering and is defined as the process by which a prolonged exposure to the cold of winter results in competence to flower during the following spring. In cereals, three genes (VERNALIZATION1 [VRN1], VRN2, and FLOWERING LOCUS T [FT]) have been identified that influence the vernalization requirement and are thought to form a regulatory loop to control the timing of flowering. Here, we characterize natural variation in the vernalization and photoperiod responses in Brachypodium distachyon, a small temperate grass related to wheat (Triticum aestivum) and barley (Hordeum vulgare). Brachypodium spp. accessions display a wide range of flowering responses to different photoperiods and lengths of vernalization. In addition, we characterize the expression patterns of the closest homologs of VRN1, VRN2 (VRN2-like [BdVRN2L]), and FT before, during, and after cold exposure as well as in different photoperiods. FT messenger RNA levels generally correlate with flowering time among accessions grown in different photoperiods, and FT is more highly expressed in vernalized plants after cold. VRN1 is induced by cold in leaves and remains high following vernalization. Plants overexpressing VRN1 or FT flower rapidly in the absence of vernalization, and plants overexpressing VRN1 exhibit lower BdVRN2L levels. Interestingly, BdVRN2L is induced during cold, which is a difference in the behavior of BdVRN2L compared with wheat VRN2 during cold.

Morphologic and Genomic Characteristics of Breast Cancers Occurring in Individuals with Lynch Syndrome.

PURPOSE: Lynch syndrome is defined by germline pathogenic mutations involving DNA mismatch repair (MMR) genes and linked with the development of MMR-deficient colon and endometrial cancers. Whether breast cancers developing in the context of Lynch syndrome are causally related to MMR deficiency (MMRd), remains controversial. Thus, we explored the morphologic and genomic characteristics of breast cancers occurring in Lynch syndrome individuals. EXPERIMENTAL DESIGN: A retrospective analysis of 20,110 patients with cancer who underwent multigene panel genetic testing was performed to identify individuals with a likely pathogenic/pathogenic germline variant in MLH1, MSH2, MSH6, or PMS2 who developed breast cancers. The histologic characteristics and IHC assessment of breast cancers for MMR proteins and programmed death-ligand 1 (PD-L1) expression were assessed on cases with available materials. DNA samples from paired tumors and blood were sequenced with Memorial Sloan Kettering-Integrated Mutation Profiling of Actionable Cancer Targets (≥468 key cancer genes). Microsatellite instability (MSI) status was assessed utilizing MSISensor. Mutational signatures were defined using SigMA. RESULTS: A total of 272 individuals with Lynch syndrome were identified, 13 (5%) of whom had primary breast cancers. The majority of breast cancers (92%) were hormone receptor-positive tumors. Five (42%) of 12 breast cancers displayed loss of MMR proteins by IHC. Four (36%) of 11 breast cancers subjected to tumor-normal sequencing showed dominant MSI mutational signatures, high tumor mutational burden, and indeterminate (27%) or high MSISensor scores (9%). One patient with metastatic MMRd breast cancer received anti-PD1 therapy and achieved a robust and durable response. CONCLUSIONS: A subset of breast cancers developing in individuals with Lynch syndrome are etiologically linked to MMRd and may benefit from anti-PD1/PD-L1 immunotherapy.

Cancer-Causative Mutations Occurring in Early Embryogenesis.

Mosaic mutations in normal tissues can occur early in embryogenesis and be associated with hereditary cancer syndromes when affecting cancer susceptibility genes (CSG). Their contribution to apparently sporadic cancers is currently unknown. Analysis of paired tumor/blood sequencing data of 35,310 patients with cancer revealed 36 pathogenic mosaic variants affecting CSGs, most of which were not detected by prior clinical genetic testing. These CSG mosaic variants were consistently detected at varying variant allelic fractions in microdissected normal tissues (n = 48) from distinct embryonic lineages in all individuals tested, indicating their early embryonic origin, likely prior to gastrulation, and likely asymmetrical propagation. Tumor-specific biallelic inactivation of the CSG affected by a mosaic variant was observed in 91.7% (33/36) of cases, and tumors displayed the hallmark pathologic and/or genomic features of inactivation of the respective CSGs, establishing a causal link between CSG mosaic variants arising in early embryogenesis and the development of apparently sporadic cancers. SIGNIFICANCE: Here, we demonstrate that mosaic variants in CSGs arising in early embryogenesis contribute to the oncogenesis of seemingly sporadic cancers. These variants can be systematically detected through the analysis of tumor/normal sequencing data, and their detection may affect therapeutic decisions as well as prophylactic measures for patients and their offspring. See related commentary by Liggett and Sankaran, p. 889. This article is highlighted in the In This Issue feature, p. 873.

Molecular analysis of encapsulated papillary carcinoma of the breast with and without invasion.

Encapsulated papillary carcinomas (EPCs) of the breast are a unique variant of papillary carcinoma confined to a cystic space with absent or attenuated myoepithelial cell layer. Although staged as an in situ lesion, it can be associated with invasive ductal carcinoma (IDC). We sought to compare the genomic characteristics of pure EPC and EPC with associated invasive carcinoma (EPCi) at the genomic level. All cases of EPCi harbored recurrent hotspot mutations in PIK3CA. PIK3CA, KMT2A, and CREBBP deleterious somatic events were found across both tumor groups, irrespective of invasion status. At the whole transcriptomic level, EPCi cases displayed remarkably similar mRNA profiles when compared to EPC. When EPCi cases were compared with their corresponding IDC, despite significant overlap, we identified differential gene expression in 39 genes with enrichment of multiple pathways including extracellular matrix regulation, cell adhesion, and collagen fibril organization. Despite morphologic, genotypic, and transcriptomic overlap between pure EPC and EPCi, the latter tumors are likely advanced lesions with PIK3CA activating mutations and enrichment of stromal-related genes implicated in the switch to IDC.

Pathologic Evaluation of Breast Tissue From Transmasculine Individuals Undergoing Gender-Affirming Chest Masculinization.

CONTEXT.­: Bilateral mastectomy for chest masculinization is one of the gender-affirming procedures for transmasculine individuals. OBJECTIVE.­: To optimize gross handling protocols and assess histopathologic findings in transmasculine breast tissue specimens. DESIGN.­: We identified all gender-affirming mastectomies from 2015 to 2018. We sequentially identified reduction mammoplasty (RM) cases for macromastia from the same period as control. Significant findings were defined as atypical ductal or lobular hyperplasia (ADH, ALH), ductal or lobular carcinoma in situ (DCIS, LCIS), or invasive carcinoma. RESULTS.­: Significant findings were present in 6 of 211 gender-affirming mastectomies (2.8%) as follows: ADH (n = 5) and LCIS together with ALH (n = 1). By comparison, 19 of 273 RM specimens (7%) yielded significant findings as follows: ALH (n = 11), ADH (n = 4), LCIS (n = 2), DCIS (n = 1), and invasive lobular carcinoma (n = 1). In the gender-affirming group, 142 transmen underwent androgen therapy before surgery, of whom 2 had significant pathologic findings. Thirty and 41 individuals had a family history of breast cancer in the gender-affirming and RM group, of whom 1 and 3 individuals had significant pathologic findings, respectively. CONCLUSIONS.­: Our study demonstrates that we handle transmasculine mastectomy specimens by examining 2.8 times more slides on average than for RMs, with a 2.5 times lower rate of significant pathologic findings. Prior family history of breast cancer or the use of androgen therapy before surgery in gender-affirming individuals did not increase the risk of identifying significant breast lesions. We recommend submitting 4 tissue blocks per mastectomy for individuals undergoing gender-affirming breast surgery.

Primary Esophageal Mixed Sarcomatoid and Small Cell Neuroendocrine Carcinoma With Brain Metastasis: A Challenging Diagnosis on Biopsy.

Mixed carcinomas in the esophagus are highly uncommon neoplasms that represent a diagnostic challenge on small tissue biopsies. We present a case of a primary mixed sarcomatoid-small cell carcinoma of the esophagus that was diagnosed after repeat sampling of the lesion. The components were morphologically distinct and could be further classified by immunohistochemistry. Next-generation sequencing identified mutations in PIK3CA and CDKN2A. The small cell component morphology was also identified in brain metastasis.

Revisiting multifocal breast cancer: a clonality study of ductal carcinoma using whole exome sequencing.

Multifocal breast cancer (MFBC), ductal type, has been hypothesized to arise by one of two mechanisms: either through intramammary/intralymphatic spread from a single index tumor (MBC-1), or as multiple independent tumors with each focus carrying its corresponding ductal carcinoma in-situ (MBC-2). In order to improve our understanding of MFBC pathogenesis, we employed laser capture microdissection coupled with whole-exome sequencing to study clonal origin in MFBC. We selected three cases of MBC-1 (C1 to C3) and MBC-2 (C4 to C6) and analyzed three foci from each case. MBC-1 cases were histologically similar and showed a strong predilection for satellite foci, vascular invasion and nodal metastasis when compared to MBC-2. Our bioinformatics approach provided strong evidence for clonal relationships in MBC-1, as demonstrated by distinct clusters of genes conserved across all tumor foci. Conversely, no gene clusters were shared across all the foci in MBC-2, suggesting multiple independent tumors. These findings provide further support for the two distinct pathogenetic mechanisms in MFBC.

Microglandular adenosis is an advanced precursor breast lesion with evidence of molecular progression to matrix-producing metaplastic carcinoma.

Microglandular adenosis (MGA) is a rare breast lesion reported to be associated with invasive carcinoma in up to 20% to 30% of cases and has been proposed as a nonobligate precursor to basal-like breast cancers. We identified a case of matrix-producing metaplastic carcinoma with morphologic and immunohistochemical evidence of progression from MGA to atypical MGA, carcinoma in situ, and invasive carcinoma. We performed whole-exome sequencing of each component (MGA, atypical MGA, carcinoma in situ, and cancer) to characterize the mutational landscape of these foci. There was a significant copy number overlap between all foci, including a segmental amplification of the CCND1 locus (partial chromosome 11 trisomy) and MYC (8q24.12-13). Using a bioinformatics approach, we were able to identify 3 putative mutational clusters and recurrent, stop-gain nonsynonymous mutations in both ZNF862 and TP53 that were shared across all foci. Finally, we identified a novel deleterious splice-acceptor site mutation of chr5:5186164 G>T (chromosome 5p15) encoding the gene, ADAMTS16, in the invasive component.