SMARCB1 Expression Research Articles

Renal cell carcinoma, unclassified with medullary phenotype: poorly differentiated adenocarcinomas overlapping with renal medullary carcinoma

Renal medullary carcinoma (RMC) is a highly aggressive renal cell carcinoma arising in the collecting system and requiring careful correlation with status of sickle cell trait. A panel of international experts has recently proposed provisional diagnostic terminology, renal cell carcinoma, unclassified, with medullary phenotype, based on encountering an extraordinarily rare tumor with RMC morphology and immunophenotype in an individual proven not to have a hemoglobinopathy. Herein, we extend this observation to a cohort of 5 such tumors, morphologically similar to RMC, lacking SMARCB1 expression by immunohistochemistry, but each without evidence of a hemoglobinopathy. The tumors arose in 4 men and 1 woman with a mean age of 44 years, occurring in 3 left and 2 right kidneys. Clinically, aggression was apparent with involvement of perinephric adipose tissue in all 5 cases, nodal metastasis in 4 of 5 cases, and death of disease in 4 of 5 cases within 3-27 months. Histologic sections showed poorly differentiated adenocarcinoma, often with solid and nested growth patterns, as well as infiltrative glandular, tubulopapillary, cribriform, or reticular growth. Rhabdoid and sarcomatoid cytomorphology was seen in a subset. All tumors showed PAX8 nuclear positivity and SMARCB1 loss, with OCT3/4 expression in 4 of 5 cases. In summary, this first series of renal cell carcinoma, unclassified, with medullary phenotype documents tumors with morphologic, immunophenotypic, and prognostic features of RMC occurring in individuals without sickle cell trait. Although greater biologic and molecular understanding is needed, the available evidence points to these cases representing a sporadic counterpart to sickle cell trait-associated RMC.

Abstract 4875: Identification of the cellular origin and "stemness" phenotype of Malignant Rhabdoid Tumors (MRT) may represent a new therapeutic approach in paediatric oncology

Abstract Malignant Rhabdoid Tumors (MRT) are highly aggressive childhood malignancies characterized by a single mutation; biallelic inactivation of SMARCB1, a component of the SWI/SNF chromatin remodeling complex. These tumors may occur anywhere, most frequently in the brain (Atypical Teratoid/Rhabdoid Tumor, ATRT) and in the kidneys and soft tissues (Extra Cranial Rhabdoid tumor, ECRT). MRT presents immune markers from multiple lineages exhibiting a teratoid phenotype, characterized by cells with heterogeneous morphology within the same tumor leading to speculation that the MRT cell of origin is a type of multipotent stem cell. Despite recent advances in treating other solid tumors, treatment for MRT still remains ineffective and we hypothesized that MRT “stemness” characteristics are key to its aggressive clinical course and resistance to treatment. Here we analyze the expression profiles of primary MRT (n= 134) and catalog the expression of a program of “stemness” genes capable of driving aberrant self-renewal. Further, we show by re-expression of SMARCB1 in MRT cells that several of these key “stemness” genes are aberrantly activated by SMARCB1 mutation. One such gene, BMI1 was further demonstrated to be critical to MRT tumorigenesis by shRNA knockout, and a novel anti-BMI1 drug, PTC209 was tested to show efficacy in MRT. Knockdown of BMI1 in MRT cells reduced the self-renewal capability of cells as seen from the number of tumorspheres formed in limiting dilution assay (LDA). Interestingly, BMI1 loss triggers upregulation of p16(INK4a) expression and this mimics the expression profile when SMARCB1 was re-expressed into MRT cells. To identify a putative MRT cell of origin we performed a meta-analysis cross-referencing expression profiles from primary MRT (n= 20), and functional models in which SMARCB1 was re-expressed (n=5 lines) with expression profiles from multiple stem cell types including epithelial, embryonic, neural mesenchymal and neural crest (n= 446). Analysis by t-SNE and NMF consensus clustering suggested that MRT bore the closest resemblance to either early neural progenitors (ATRT) or early neural crest cells (ECRT). In MRT cells in which SMARCB1 was forcibly re-expressed, cells became gradually more differentiated and their expression profile altered from that of an early neural crest to a more differentiated mesenchymal pattern. For the first time, we show evidence that ECRTs and ATRTs appear to have different cells of origin, despite having the same mutation and tumor appearance. We further demonstrate that SMARCB1 expression is necessary for MRT cells to maintain a de-differentiated “stem-like” state and finally that the SMARCB1-dependent stemness gene BMI1 shows potential as a novel therapeutic target in MRT. Citation Format: Ras A. Ramli, Martina A. Finetti, Matthew P. Selby, Yura Grabovska, Stephen Crosier, Amanda J. Smith, Steven C. Clifford, Daniel Williamson. Identification of the cellular origin and "stemness" phenotype of Malignant Rhabdoid Tumors (MRT) may represent a new therapeutic approach in paediatric oncology [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 4875. doi:10.1158/1538-7445.AM2017-4875

Abstract 408: Investigating the biology of atypical teratoid/rhabdoid tumors by whole genome CRISPR/Cas9 screening

Abstract Atypical Teratoid/Rhabdoid Tumors are rare, highly aggressive, paediatric tumours with a dismal prognosis. Rhabdoid Tumours (RT) have one key genetic defect (biallelic inactivation of SMARCB1 in > 90% of patients) which, when re-expressed in RT cells, causes differentiation and growth arrest. We used whole genome CRISPR (clustered regularly interspaced short palindromic repeats)/Cas9 screening; a technique by which mutations are created systematically in each gene in the genome - to identify novel SMARCB1-dependent therapeutic targets and synthetic lethal relationships in RT. A panel of RT cell lines were transduced with a GeCKO (Genome Scale CRISPR Knock-Out) library which contains 122,441 guide constructs targeting 19,050 genes and 1,864 miRNAs; each cell contained a single CRISPR insert. This panel was further transduced with a SMARCB1 expression lentivirus or an empty vector control, cells were collected at 0, 7 and14 days post infection (300x library coverage, 3.7 x 107 cells per time point). CRISPR inserts were sequenced, counted, normalized and analyzed to identify those whose abundance was significantly altered, representing candidate gene targets which promote or inhibit RT cell growth/viability in a SMARCB1-dependent manner. CRISPR hits were further cross-validated with primary patient and functional model genomic data (RNA-sequencing/450k DNA methylation array) to identify several high confidence deleterious events with strong SMARCB1 dependency. Hundreds of significantly enriched (both positively and negatively) genes were identified which alter when SMARCB1 is present or absent including MYC family targets, cell cycle targets, SHH pathway and the Aurora kinase family. TGFβ was identified as a top upstream SMARCB1/methylation dependent regulator and when SMARCB1 re-expression data was overlaid onto the TGFβ canonical pathway showed a convincing enrichment in pathway regulation. Pathway members SMAD 2/3 showed a ‘synthetic lethal’ phenotype and represent a functional SMARCB1-dependent relationship which is also observed in SMARCB1 re-expression and methylation model data and is characteristic of primary RT. This phenotype was validated functionally by expression of a dominant negative TGFBR2. Combining RNA-sequencing and 450K DNA methylation microarray data from primary tumours and experimental models, coupled with whole-genome functional screening, provides a genome-wide view of key RT tumorigenic genes/pathways and their SMARCB1 dependence. These data suggest synthetic lethal phenotypes and enable prioritisation of true SMARCB1-dependent events and therapeutic targets, in this rare yet lethal paediatric tumour. Citation Format: Matthew P. Selby, Martina A. Finetti, Matthew Bashton, Ruth E. Cranston, Yura Grabovska, Alicia Del-Carpio-Pons, Amanda Smith, Simon Bailey, Steven C. Clifford, Daniel Williamson. Investigating the biology of atypical teratoid/rhabdoid tumors by whole genome CRISPR/Cas9 screening [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 408. doi:10.1158/1538-7445.AM2017-408

ATRT-05. PRECLINICAL EFFICACY OF RADIATION-FREE TREATMENT OF ATYPICAL TERATOID/RHABDOID TUMORS BY COMBINED MEK AND MELK INHIBITION

AbstractAtypical teratoid/rhabdoid tumors (AT/RT) are highly malignant tumors that occur both in- and outside the central nervous system, predominantly in young children. Although therapeutic interventions, combining surgery with intensive chemotherapy and conformal radiotherapy, have shown curative potential, the long-term consequences of these treatment modalities in young children are often severe. Therefore, there is a dire need for novel, targeted therapeutic strategies that potentially reduce or eliminate chemo- and radiotherapy, especially for children under three years of age. Genetically, AT/RT is characterized by a biallelic inactivation of SMARCB1 or SMARCA4, causing aberrant chromatin remodeling and thereby expression of a variety of oncogenes. Recently, the importance of the mitogen-activated protein kinase (MAPK) pathway in the pathogenesis of AT/RT has been demonstrated, with preclinical efficacy of MEK inhibition in AT/RT cells. Preclinical patient-derived AT/RT models are largely lacking. We developed a patient-derived primary cell line and xenograft model, VUMC-AT/RT-01, from a surgical specimen of an AT/RT patient that had not received prior therapy. Immunohistochemistry was performed on xenograft tumors, revealing typical loss of SMARCB1 expression as well as heterogeneous expression of glial, epithelial, mesenchymal and neuronal differentiation markers. In silico analysis of gene expression revealed strong upregulation of MELK in AT/RT tumors compared to normal brain tissues. These high levels of MELK were subsequently confirmed in our xenograft model. Inhibition of MELK with the small molecule OTSSP167 effectively reduced proliferation and induced cell death in primary AT/RT neurospheres. Combined treatment of primary AT/RT cells with OTSSP167 and the MEK inhibitor Trametinib showed strongly synergistic cytotoxicity at low nanomolar concentrations. Treatment of mice carrying VUMC-AT/RT-01-Fluc xenografts with OTSSP167 and Trametinib confirmed this synergy in vivo. We conclude that combined MEK and MELK inhibition represents a promising targeted therapy strategy for AT/RT that might ultimately reduce or eliminate the need for radiation and intensive chemotherapy.

Recurrent copy number alterations in low-grade and anaplastic pleomorphic xanthoastrocytoma with and without BRAF V600E mutation.

Pleomorphic xanthoastrocytoma (PXA) is a rare localized glioma characterized by frequent BRAF V600E mutation and CDKN2A/B deletion. We explored the association of copy-number variants (CNVs) with BRAF mutations, tumor grade, and patient survival in a cohort of 41 PXA patients using OncoScan chromosomal microarray. Primary resection specimens were available in 38 cases, including 24 PXA and 14 anaplastic PXA (A-PXA), 23 BRAF V600E mutant tumors (61%). CNVs were identified in all cases and most frequently involved chromosome 9 with homozygous CDKN2A/B deletion (n = 33, 87%), a higher proportion than previously detected by comparative genomic hybridization (50%-60%) (37). CDKN2A/B deletion was present in similar proportion of PXA (83%), A-PXA (93%), BRAF V600E (87%), and wild-type (87%) tumors. Whole chromosome gains/losses were frequent, including gains +7 (n = 15), +2 (n = 11), +5 (n = 10), +21 (n = 10), +20 (n = 9), +12 (n = 8), +15 (n = 8), and losses -22 (n = 11), -14 (n = 7), -13 (n = 5). Losses and copy-neutral loss of heterozygosity were significantly more common in A-PXA, involving chromosomes 22 (P = 0.009) and 14 (P = 0.03). Amplification of 8p and 12q was identified in a single tumor. Histologic grade was a robust predictor of overall survival (P = 0.003), while other copy-number changes, including CDKN2A/B deletion, did not show significant association with survival. Distinct histologic patterns of anaplasia included increased mitotic activity in an otherwise classic PXA or associated with small cell, fibrillary, or epithelioid morphology, with loss of SMARCB1 expression in one case. In 10 cases, matched specimens were compared, including A-PXA with areas of distinct low- and high-grade morphology (n = 2), matched primary/tumor recurrence (n = 7), or both (n = 1). Copy-number changes on recurrence/anaplastic transformation were complex and highly variable, from nearly identical profiles to numerous copy-number changes. Overall, we confirm CDKN2A/B deletion as key a feature of PXA not associated with tumor grade or BRAF mutation, but central to the underlying genetics of PXA.

SMARCB1 (INI-1)-deficient Sinonasal Carcinoma: A Series of 39 Cases Expanding the Morphologic and Clinicopathologic Spectrum of a Recently Described Entity.

To more fully characterize the clinical and pathologic spectrum of a recently described tumor entity of the sinonasal tract characterized by loss of nuclear expression of SMARCB1 (INI1), we analyzed 39 SMARCB1-deficient sinonasal carcinomas collected from multiple medical centers. The tumors affected 23 males and 16 females with an age range of 19 to 89 years (median, 52). All patients presented with locally advanced disease (T3, n=5; T4, n=27) involving the sinuses (mainly ethmoid) with variable involvement of the nasal cavity. Thirty patients received surgery and/or radiochemotherapy with curative intent. At last follow-up, 56% of patients died of disease 0 to 102 months after diagnosis (median, 15), 2 were alive with disease, and 1 died of an unrelated cause. Only 9 patients (30%) were alive without disease at last follow-up (range, 11 to 115 mo; median, 26). The original diagnosis of retrospectively identified cases was most often sinonasal undifferentiated carcinoma (n=14) and nonkeratinizing/basaloid squamous cell carcinoma (n=5). Histologically, most tumors displayed either a predominantly basaloid (61%) or plasmacytoid/rhabdoid morphology (36%). The plasmacytoid/rhabdoid form consisted of sheets of tumor cells with abundant, eccentrically placed eosinophilic cytoplasm, whereas similar cells were typically rare and singly distributed in the basaloid variant. Glandular differentiation was seen in a few tumors. None of the cases showed squamous differentiation or surface dysplasia. By immunohistochemistry, the tumors were positive for pancytokeratin (97%), CK5 (64%), p63 (55%), and CK7 (48%); and they were negative for NUT (0%). Epstein-Barr virus and high-risk human papillomavirus was not detected by in situ hybridization. Immunohistochemical loss of SMARCB1 (INI1) expression was confirmed for all 39 tumors. Investigation of other proteins in the SWI/SNF complex revealed co-loss of SMARCA2 in 4 cases, but none were SMARCA4 deficient or ARID1A deficient. Of 27 tumors with SMARCB1 fluorescence in situ hybridization analysis, 14 showed homozygous (biallelic) deletions and 7 showed heterozygous (monoallelic) deletions. SMARCB1-deficient sinonasal carcinoma represents an emerging poorly differentiated/undifferentiated sinonasal carcinoma that (1) cannot be better classified as another specific tumor type, (2) has consistent histopathologic findings (albeit with some variability) with varying proportions of plasmacytoid/rhabdoid cells, and (3) demonstrates an aggressive clinical course. This entity should be considered in any difficult-to-classify sinonasal carcinoma, as correct diagnosis will be mandatory for optimizing therapy and for further delineation of this likely underdiagnosed disease.

Embryonic signature distinguishes pediatric and adult rhabdoid tumors from other SMARCB1-deficient cancers.

Extra-cranial rhabdoid tumors (RT) are highly aggressive malignancies of infancy, characterized by undifferentiated histological features and loss of SMARCB1 expression. The diagnosis is all the more challenging that other poorly differentiated cancers lose SMARCB1 expression, such as epithelioid sarcomas (ES), renal medullary carcinomas (RMC) or undifferentiated chordomas (UC). Moreover, late cases occurring in adults are now increasingly reported, raising the question of differential diagnoses and emphasizing nosological issues. To address this issue, we have analyzed the expression profiles of a training set of 32 SMARCB1-deficient tumors (SDT), with ascertained diagnosis of RT (n = 16, all < 5 years of age), ES (n = 8, all > 10 years of age), UC (n = 3) and RMC (n = 5). As compared with other SDT, RT are characterized by an embryonic signature, and up-regulation of key-actors of de novo DNA methylation processes. Using this signature, we then analysed the expression profiling of 37 SDT to infer the appropriate diagnosis. Thirteen adult onset tumors showed strong similarity with pediatric RT, in spite of older age; by exome sequencing, these tumors also showed genomic features indistinguishable from pediatric RT. In contrary, 8 tumors were reclassified within carcinoma, ES or UC categories, while the remaining could not be related to any of those entities. Our results demonstrate that embryonic signature is shared by all RT, whatever the age at diagnosis; they also illustrate that many adult-onset SDT of ambiguous histological diagnosis are clearly different from RT. Finally, our study paves the way for the routine use of expression-based signatures to give accurate diagnosis of SDT.

Highlights from Recent Cancer Literature

Highlights from Recent Cancer Literature

Loss of expression of SMARCA4 (BRG1), SMARCA2 (BRM) and SMARCB1 (INI1) in undifferentiated carcinoma of the endometrium is not uncommon and is not always associated with rhabdoid morphology.

Abnormalities of SMARCB1 (INI1), which encodes a member of the SWI/SNF pathway, are found in neoplasms with rhabdoid morphology, such as malignant rhabdoid tumour of the kidney and atypical teratoid/rhabdoid tumour of the central nervous system. SMARCA4 (BRG1), which encodes another member of the SWI/SNF pathway, and which is mutated in almost all small-cell carcinomas of the ovary, hypercalcaemic type, has been investigated in endometrial carcinomas, and mutations with resultant loss of immunohistochemical staining have been demonstrated in some endometrial undifferentiated carcinomas/dedifferentiated carcinomas. The aim of this study was to evaluate immunohistochemical expression of SMARCA4, SMARCB1 and SMARCA2 in a cohort of undifferentiated endometrial carcinomas, and to correlate expression of these markers with rhabdoid morphology and clinical outcome. Forty undifferentiated endometrial carcinomas (18 pure and 22 dedifferentiated carcinomas) were stained with SMARCA4 (n = 40), SMARCB1 (n = 27), and SMARCA2 (n = 37). SMARCA4 expression was intact in 26 of 40 (65%) cases, lost in 13 of 40 (32.5%) cases, and unassessable in one case (2.5%). SMARCB1 expression was intact in 26 of 27 (96%) cases and lost in one of 27 (4%) cases. SMARCA2 expression was intact in 23 of 37 (62%) cases, lost in 10 of 37 (27%) cases, and unassessable in four cases. SMARCA2 expression showed corresponding loss in nine of the 13 (69%) SMARCA4-deficient cases. Rhabdoid morphology was present in three of 13 (23%) SMARCA4-deficient cases, in two of 10 (20%) SMARCA2-deficient cases, in four of 26 (15%) SMARCA4-intact cases, and in four of 23 (17%) SMARCA2-intact cases. There was no correlation between SMARCA4 or SMARCA2 expression and clinical outcome. Our study demonstrated that almost one-third of endometrial undifferentiated carcinomas show loss of SMARCA4 and SMARCA2 expression, and that a subset show rhabdoid morphology. The majority of the SMARCA4-deficient cases show concomitant loss of SMARCA2 expression. There is no correlation between SMARCA4 or SMARCA2 expression and outcome. Our results confirm that the SWI/SNF chromatin-remodelling complex is involved in the pathogenesis of endometrial undifferentiated carcinomas.

GENT-45. HETEROGENEITY OF SWI-SNF COMPONENTS IN MALIGNANT RHABDOID TUMORS

Malignant Rhabdoid Tumors (MRT’s) are highly aggressive and often fatal pediatric tumors that can occur in the central nervous system (CNS) or outside the CNS. When they occur in the CNS, they are called atypical teratoid rhabdoid tumors (AT/RT) and are most common in children younger than 3 years of age. The loss of the tumor suppressor SMARCB1 on chromosome 22 as a result of homozygous deletions, truncating non-sense mutations or frameshift mutations is the common driver event in all MRT’s. SMARCB1 is a core component of the SWI/SNF chromatin-remodeling complex that regulates gene expression by reorganizing chromatin in an ATP dependent manner. Histopathologically, MRT’s are characterized by polyphenotypic differentiation. Since the SWI/SNF complex closely regulates differentiation, we examined the composition of the SWI/SNF complex in MRT’s derived from different locations using immunohistochemistry in a cohort of 43 rhabdoid tumors. Further, the expression of these SWI/SNF components during brain development was analyzed using gene expression data from the Allen Brain Atlas. We found that the expression of SMARCB1 was highest in the cerebellum throughout development. Global levels of other members of the SWI/SNF complex did not vary significantly during brain development. In MRTs, the expression of the core SWI/SNF members (SMARCC1, SMARCC2) remained relatively stable across tumors from different locations. Surprisingly, other SWI/SNF components including ARID1A, ARID1B and BRD7 show heterogeneity in expression depending on the location of tumors within the CNS. Other SWI/SNF members such as ACTL6A, ACTL6B, SMARCD1 and PBRM1 showed differences between the CNS and non-CNS tumors. These data have implications to understand the complex biology of MRTs.

SMARCA4 and SMARCA2 deficiency in non-small cell lung cancer: immunohistochemical survey of 316 consecutive specimens.

The chromatin remodeling switch sucrose nonfermentable (SWI/SNF) complex has been increasingly implicated in the pathogenesis and dedifferentiation of neoplasms from several organs with prognostic and potential therapeutic implications. We herein investigated the expression of the SWI/SNF complex catalytic subunits SMARCA4 (BRG1) and SMARCA2 (BRM) in 316 consecutive non-small cell lung cancer (NSCLC) specimens on tissue microarrays (171 adenocarcinomas [ADCAs], 130 squamous cell carcinomas [SCCs], 9 adenosquamous carcinomas, and 6 large cell carcinomas) excluding undifferentiated/giant cell or rhabdoid carcinomas. Complete loss of SMARCA4 was observed in 8 (5.5%) of 146 evaluable pulmonary ADCAs and 6 (5.2%) of 115 evaluable pulmonary SCCs, whereas 9 (6.4%) of 140 ADCAs and 2 (1.7%) of 117 SCCs showed SMARCA2 loss. Two of 6 large cell carcinomas were SMARCA2 deficient. Concurrent loss of both markers was observed in 4 cases (2 ADCAs and 2 SCCs). Of 15 ADCAs with loss of either or both markers, 12 (80%) were TTF1 negative. In conclusion, SMARCA4 and SMARCA2 deficiency is observed in 5.1% and 4.8% of NSCLC, respectively. SMARCB1 expression was intact in all cases. The presence of differentiated histology (glandular or squamous) is a novel aspect among SWI/SNF-deficient carcinomas which in other organs generally are associated with undifferentiated/rhabdoid morphology. The predominance of TTF1 negativity among SWI/SNF-deficient pulmonary ADCA (80%) underlines the need to include these 2 markers in the evaluation of TTF1-negative ADCA of putative pulmonary origin. Given the recently documented potential of SMARCA4 loss as a predictor of chemosensitivity to platinum-based chemotherapy in NSCLC, recognition of the clinicopathological features of SMARCA4-deficient NSCLC in routine surgical pathology practice is recommended.

Dual Targeting of PDGFRα and FGFR1 Displays Synergistic Efficacy in Malignant Rhabdoid Tumors

SummarySubunits of the SWI/SNF chromatin remodeling complex are mutated in a significant proportion of human cancers. Malignant rhabdoid tumors (MRTs) are lethal pediatric cancers characterized by a deficiency in the SWI/SNF subunit SMARCB1. Here, we employ an integrated molecular profiling and chemical biology approach to demonstrate that the receptor tyrosine kinases (RTKs) PDGFRα and FGFR1 are coactivated in MRT cells and that dual blockade of these receptors has synergistic efficacy. Inhibitor combinations targeting both receptors and the dual inhibitor ponatinib suppress the AKT and ERK1/2 pathways leading to apoptosis. MRT cells that have acquired resistance to the PDGFRα inhibitor pazopanib are susceptible to FGFR inhibitors. We show that PDGFRα levels are regulated by SMARCB1 expression, and assessment of clinical specimens documents the expression of both PDGFRα and FGFR1 in rhabdoid tumor patients. Our findings support a therapeutic approach in cancers with SWI/SNF deficiencies by exploiting RTK coactivation dependencies.

Prognostic factors and survival in non-central nervous system rhabdoid tumors

IntroductionNon-central nervous system (non-CNS) rhabdoid tumors tend to present at a young age and have an extremely aggressive course, with dismal overall survival rates. Inactivation of the tumor suppressor gene SMARCB1 has been shown in rhabdoid tumors regardless of anatomic location, suggesting a common genetic basis. We retrospectively analyzed our institutional experience with non-CNS rhabdoid tumors to determine overall survival and prognostic variables. MethodsWe reviewed records of pediatric patients (age<22y) with non-CNS rhabdoid tumor at our institution between 1980 and 2014. Variables evaluated for correlation with survival included: age > or <1.5years (median) at diagnosis, M1 status, and radiation therapy. The log-rank test was used to compare Kaplan–Meier probability distributions with P values adjusted for multiple testing using the false discovery rate approach. ResultsNineteen consecutive patients (10 female) with histologically verified rhabdoid tumor were identified. Mean age at diagnosis was 3.2years (median 1.5y, range 1.3mo–21.8y). Primary tumors were located in the kidney (n=10), head and neck (n=5), and in the liver, thigh, mediastinum and retroperitoneum (n=1 each). SMARCB1 expression was absent in all 10 patients tested. Eight patients had distant metastases at diagnosis. Median overall survival was 1.2years. Age greater than the median and radiation therapy were associated with better outcome, with a median overall survival of 2.7years (P=0.049 and P=0.003, respectively). ConclusionSurvival rates for rhabdoid tumor remain poor, but prognosis is better in older children, regardless of primary tumor location. Because of its rarity, clinical trials with present agents are difficult to conduct. Further progress will require a focus on therapies targeted at tumor biology rather than anatomic location for non-CNS rhabdoid tumors.

Cribriform neuroepithelial tumor: molecular characterization of a SMARCB1-deficient non-rhabdoid tumor with favorable long-term outcome.

Rhabdoid phenotype and loss of SMARCB1 expression in a brain tumor are characteristic features of atypical teratoid/rhabdoid tumors (ATRT). Rare non-rhabdoid brain tumors showing cribriform growth pattern and SMARCB1 loss have been designated cribriform neuroepithelial tumor (CRINET). Small case series suggest that CRINETs may have a relatively favorable prognosis. However, the long-term outcome is unclear and it remains uncertain whether CRINET represents a distinct entity or a variant of ATRT. Therefore, 10 CRINETs were clinically and molecularly characterized and compared with 10 ATRTs of each of three recently described molecular subgroups (i.e. ATRT-TYR, ATRT-SHH and ATRT-MYC) using Illumina Infinium HumanMethylation450 arrays, FISH, MLPA, and sequencing. Furthermore, outcome was compared to a larger cohort of 27 children with ATRT-TYR. Median age of the 6 boys and 4 girls harboring a CRINET was 20 months. On histopathological examination, all CRINETs demonstrated a cribriform growth pattern and distinct tyrosinase staining. On unsupervised cluster analysis of methylation data, all CRINETs examined exclusively clustered within the ATRT-TYR molecular subgroup. As ATRT-TYR, CRINETs mainly showed large heterozygous 22q deletions (9/10) and SMARCB1 mutations of the other allele. In two patients, SMARCB1 mutations were also present in the germline. Estimated mean overall survival in patients with CRINETs was 125 months (95% confidence interval 100-151 months) as compared to only 53 (33-74) months in patients with ATRTs of the ATRT-TYR subgroup (Log-Rank P < 0.05). In conclusion, CRINET represents a SMARCB1-deficient non-rhabdoid tumor, which shares molecular similarities with the ATRT-TYR subgroup but has distinct histopathological features and favorable long-term outcome.

Loss of SMARCA4 (BRG1) protein expression as determined by immunohistochemistry in small-cell carcinoma of the ovary, hypercalcaemic type distinguishes these tumours from their mimics.

Molecular investigation of small-cell carcinoma of the ovary, hypercalcaemic type (SCCOHT) has revealed that it is a monogenetic tumour characterized by alteration of SMARCA4 (BRG1), encoding a member of the switch/sucrose non-fermentable (SWI/SNF) chromatin remodelling complex. A large majority of cases show loss of expression of the corresponding SMARCA4/BRG1 protein. Furthermore, three cases of SCCOHT with retained SMARCA4 protein expression showed loss of SMARCB1/INI1 expression. The aim of this study was to assess the sensitivity and specificity of loss of SMARCA4 expression as a diagnostic test for SCCOHT. We performed SMARCA4 and SMARCB1 staining in 245 tumours, many of which were potentially in the differential diagnosis of SCCOHT. We also stained 56 cases of SCCOHT for SMARCA4 and 37 of these for SMARCB1. Fifty-four of the SCCOHT cases showed complete absence of SMARCA4 expression. The two cases with retained expression showed molecular alteration of SMARCA4. Of the 217 other neoplasms with interpretable staining, all retained SMARCA4 expression. Although the majority showed diffuse, strong nuclear expression, a heterogeneous, typically weak staining pattern was present in 13% of cases. All 37 cases of SCCOHT tested and all other neoplasms, apart from three malignant rhabdoid tumours, showed retained nuclear SMARCB1 expression. Loss of SMARCA4 expression had a sensitivity of 96.55% and specificity of 100%. Loss of SMARCA4 expression is sensitive and specific for SCCOHT. Although some mimics show heterogeneous expression, there is retention of nuclear staining in at least a part of the tumour; therefore, only complete loss of staining should be regarded as being supportive of SCCOHT.

Medullary renal cell carcinoma (RCC): Genomics and treatment outcomes.

4556Background: Medullary RCC is a rare and aggressive type of kidney cancer that primarily affects young adults with sickle cell trait. Recent reports have identified loss of expression of SMARCB1...

Loss of switch/sucrose non-fermenting complex protein expression is associated with dedifferentiation in endometrial carcinomas

Dedifferentiated endometrial carcinoma is an aggressive type of endometrial cancer that contains a mix of low-grade endometrioid and undifferentiated carcinoma components. We performed targeted sequencing of eight dedifferentiated carcinomas and identified somatic frameshift/nonsense mutations in SMARCA4, a core ATPase of the switch/sucrose non-fermenting (SWI/SNF) complex, in the undifferentiated components of four tumors. Immunohistochemical analysis confirmed the loss of SMARCA4 in the undifferentiated component of these four SMARCA4-mutated cases, whereas the corresponding low-grade endometrioid component showed retained SMARCA4 expression. An expanded survey of other members of the SWI/SNF complex showed SMARCB1 loss in the undifferentiated component of two SMARCA4-intact tumors, and all SMARCA4- or SMARCB1-deficient tumors showed concomitant loss of expression of SMARCA2. We subsequently examined the expression of SMARCA2, SMARCA4, and SMARCB1 in an additional set of 22 centrally reviewed dedifferentiated carcinomas and 31 grade 3 endometrioid carcinomas. Combining the results from the index and the expansion set, 15 of 30 (50%) of the dedifferentiated carcinomas examined showed either concurrent SMARCA4 and SMARCA2 loss (37%) or concurrent SMARCB1 and SMARCA2 loss (13%) in the undifferentiated component. The loss of SMARCA4 or SMARCB1 was mutually exclusive. All 31 grade 3 endometrioid carcinomas showed intact expression of these core SWI/SNF proteins. The majority (73%) of the SMARCA4/SMARCA2-deficient and half of SMARCB1/SMARCA2-deficient undifferentiated component developed in a mismatch repair-deficient molecular context. The observed spatial association between SWI/SNF protein loss and histologic dedifferentiation suggests that inactivation of these core SWI/SNF proteins may contribute to the development of dedifferentiated endometrial carcinoma.

The diagnostic utility of reduced immunohistochemical expression of SMARCB1 in synovial sarcomas: a validation study

Synovial sarcoma is a malignant mesenchymal neoplasm of uncertain histogenesis, characterized by a specific SS18-SSX fusion. The diagnosis of synovial sarcoma can be challenging based on morphology and conventional immunohistochemistry alone, and identification of the fusion gene by molecular genetics may be necessary for diagnosis. Several recent studies have demonstrated the diagnostic utility of the reduced expression of SMARCB1 in synovial sarcomas as measured using immunohistochemistry. Therefore, we undertook a validation study using synovial sarcomas and other spindle or round cell tumors that could enter differential diagnosis of monophasic or poorly differentiated synovial sarcomas. Among 36 synovial sarcomas that were successfully evaluated, the expression of SMARCB1 was diffusely reduced in 33 cases (92%) at variable degrees. In contrast, the expression of SMARCB1 was not reduced in any of the 93 evaluable non-synovial sarcoma tumors (5 thymomas, 5 sarcomatoid mesotheliomas, 10 schwannomas, 9 mesenchymal chondrosarcomas, 20 solitary fibrous tumors, 19 Ewing sarcomas, and 25 malignant peripheral nerve sheath tumors). A few schwannomas and malignant peripheral nerve sheath tumors showed mosaic or complete loss of SMARCB1 expression. Reduced expression of SMARCB1 immunoreactivity was therefore found to be highly sensitive and specific for synovial sarcoma, and can be useful for rapidly and accurately confirming the diagnosis of synovial sarcoma. This reduction in SMARCB1 expression likely reflects the BAF47 ejection mechanism of the SS18-SSX fusion product and can therefore be viewed as an indirect visualization of this fusion product.

Reappraisal of sinonasal undifferentiated carcinoma: SMARCB1 (INI1)-deficient sinonasal carcinoma: a single-institution experience.

Sinonasal carcinomas are rare and of diverse histology, often involve critical anatomic structures, and are associated with an aggressive clinical course and poor prognosis. Differentiating these tumor types may have clinical impact as advances in entity-specific therapeutic intervention could increase survival and quality of life and occasionally result in a cure. Recently, a unique subset of sinonasal carcinomas characterized by basaloid/rhabdoid tumor morphology and loss of expression of SMARCB1 (INI1) was identified. We tested a total of 256 tumors including head and neck (n = 241) and thoracic (n = 15) tumors with basaloid/rhabdoid morphology for loss of expression of SMARCB1 (INI1) using full tissue sections and tissue microarrays. Among these, four tumors of the sinonasal tract were found to be SMARCB1 (INI1) deficient and were reclassified as SMARCB1 (INI1)-deficient sinonasal carcinomas. These tumors appear to be restricted to the sinonasal tract, and their unique clinical, morphological, and immunohistochemical features seem to warrant inclusion as a separate new entity among the existing high-grade sinonasal neoplasms. Separation from the other types of sinonasal malignancies is important as the identification of SMARCB1 (INI1) deficiency may provide a new target for novel treatment approaches and may ultimately lead to improved patient survival.

Myoepithelioma-like Tumors of the Vulvar Region: A Distinctive Group of SMARCB1-deficient Neoplasms.

We describe 9 tumors that resemble soft tissue myoepitheliomas but possess certain traits that do not fit perfectly into this category. These tumors, herein referred to as "myoepithelioma-like tumors of the vulvar region," occurred in the subcutis of the vulva and surrounding regions of adult women aged 24 to 65 years. Histologically, the tumors measured 2 to 7.7 cm and were well circumscribed, focally encapsulated, and lobulated. Tumor cells had an epithelioid to spindled shape, with fine amphophilic cytoplasm, and uniform nuclei with vesicular chromatin and nucleoli. The tumor stroma was relatively hypervascular, and comprised a mixture of myxoid and nonmyxoid components. Myxoid areas accounted for <5% to 95% of the tumor volume, wherein cells proliferated singly or in a loosely cohesive manner. In nonmyxoid areas, tumors cells grew in diffuse sheets or storiform arrangements. Immunohistochemically, all tested tumors were positive for vimentin, epithelial membrane antigen, and estrogen receptor; most tumors expressed actin. All tumors were negative for S100 protein, glial fibrillary acidic protein, and CD34. Cytokeratin expression was absent in all but 2 tumors, which showed rare positivity. SMARCB1 expression was deficient in all cases. EWSR1, FUS, and NR4A3 rearrangements were absent. All tumors were treated through surgery. Although 3 tumors regrew or recurred after intralesional excision, all 9 patients were alive without metastases at a mean follow-up of 66 months. Myoepithelioma-like tumors of the vulvar region constitute a distinct group of tumors, although future research is required to determine whether they are an unusual subtype of soft tissue myoepitheliomas or a separate disease.