APOBEC3 mutagenesis drives therapy resistance in breast cancer.

Acquired genetic alterations commonly drive resistance to endocrine and targeted therapies in metastatic breast cancer 1-7 , however the underlying processes engendering these diverse alterations are largely uncharacterized. To identify the mutational processes operant in breast cancer and their impact on clinical outcomes, we utilized a well-annotated cohort of 3,880 patient samples with paired tumor-normal sequencing data. The mutational signatures associated with apolipoprotein B mRNA-editing enzyme catalytic polypeptide-like 3 (APOBEC3) enzymes were highly prevalent and enriched in post-treatment compared to treatment-naïve hormone receptor-positive (HR+) cancers. APOBEC3 mutational signatures were independently associated with shorter progression-free survival on antiestrogen plus CDK4/6 inhibitor combination therapy in patients with HR+ metastatic breast cancer. Whole genome sequencing (WGS) of breast cancer models and selected paired primary-metastatic samples demonstrated that active APOBEC3 mutagenesis promoted resistance to both endocrine and targeted therapies through characteristic alterations such as RB1 loss-of-function mutations. Evidence of APOBEC3 activity in pre-treatment samples illustrated a pervasive role for this mutational process in breast cancer evolution. The study reveals APOBEC3 mutagenesis to be a frequent mediator of therapy resistance in breast cancer and highlights its potential as a biomarker and target for overcoming resistance.

Converging and evolving immuno-genomic routes toward immune escape in breast cancer.

The interactions between tumor and immune cells along the course of breast cancer progression remain largely unknown. Here, we extensively characterize multiple sequential and parallel multiregion tumor and blood specimens of an index patient and a cohort of metastatic triple-negative breast cancers. We demonstrate that a continuous increase in tumor genomic heterogeneity and distinct molecular clocks correlated with resistance to treatment, eventually allowing tumors to escape from immune control. TCR repertoire loses diversity over time, leading to convergent evolution as breast cancer progresses. Although mixed populations of effector memory and cytotoxic single T cells coexist in the peripheral blood, defects in the antigen presentation machinery coupled with subdued T cell recruitment into metastases are observed, indicating a potent immune avoidance microenvironment not compatible with an effective antitumor response in lethal metastatic disease. Our results demonstrate that the immune responses against cancer are not static, but rather follow dynamic processes that match cancer genomic progression, illustrating the complex nature of tumor and immune cell interactions.

Comparison of Immunohistochemistry, Next-generation Sequencing and Fluorescence In Situ Hybridization for Detection of MTAP Loss in Pleural Mesothelioma.

9p21 deletions involving MTAP/CDKN2A genes are detected in diffuse pleural mesotheliomas (DPM) but are absent in benign mesothelial proliferations. Loss of MTAP expression by immunohistochemistry (IHC) is well accepted as a surrogate for 9p21 deletion to support a diagnosis of DPM. Accurate interpretation can be critical in the diagnosis of DPM, but variations in antibody performance may impact interpretation. The objectives of this study were to compare the performance of MTAP monoclonal antibodies (mAbs) EPR6893 and 1813 and to compare MTAP expression by IHC with 9p21 copy number status in DPM. Cytoplasmic expression of MTAP IHC with mAbs EPR6893 (ab126770; Abcam) and 1813 (NBP2-75730, Novus Biologicals) was evaluated in 56 DPM (47 epithelioid, 7 biphasic, and 2 sarcomatoid) profiled by targeted next-generation sequencing. 9p21 Copy number status was assessed by Fraction and Allele-Specific Copy Number Estimates from Tumor Sequencing (FACETS) analysis and also by CDKN2A fluorescence in situ hybridization in discrepant cases when material was available. MTAP mAb 1813 showed stronger immunoreactivity, more specific staining, and no equivocal interpretations compared to mAb EPR6893 which showed equivocal staining in 19 (34%) of cases due to weak or heterogenous immunoreactivity, lack of definitive internal positive control, and/or nonspecific background staining. MTAP expression with mAb 1813 showed near perfect agreement with 9p21 copy number by combined FACETS/fluorescence in situ hybridization calls (κ = 0.85; 95% CI, 0.71-0.99; P < .001). MTAP IHC with mAb 1813 was 96% sensitive, 86% specific, and 93% accurate for 9p21 homozygous deletion. The findings of this study suggest that interpretation of MTAP IHC is improved with mAb 1813 because mAb EPR6893 was often limited by equivocal interpretations. We show that MTAP IHC and molecular assays are complementary in detecting 9p21 homozygous deletion. MTAP IHC may be particularly useful for low tumor purity samples and in low-resource settings.

Immunohistochemistry for PRAME in Dermatopathology.

ABSTRACT: Preferentially expressed antigen in melanoma (PRAME) is a tumor-associated antigen first identified in a melanoma patient and found to be expressed in most melanomas as well as in variable levels in other malignant neoplasms of epithelial, mesenchymal, or hematolymphoid lineage. Detection of PRAME expression in formalin-fixed paraffin-embedded tissue is possible by immunohistochemistry (IHC) with commercially available monoclonal antibodies. In situ and invasive melanoma frequently show a diffuse pattern of nuclear PRAME immunoreactivity which contrasts with the infrequent and typically nondiffuse staining seen in nevi. In many challenging melanocytic tumors, results of PRAME IHC and other ancillary tests correlate well, but not always: The tests are not interchangeable. Most metastatic melanomas are positive for PRAME, whereas nodal nevi are not. Numerous studies on PRAME IHC have become available in the past few years with results supporting the value of PRAME IHC as an ancillary tool in the evaluation of melanocytic lesions and providing insights into limitations in sensitivity and specificity as well as possible pitfalls that need to be kept in mind by practicing pathologists.

BCG-Induced Tumor Immunity Requires Tumor-Intrinsic CIITA Independent of MHC-II.

For decades, BCG immunotherapy has been the standard of care for non-muscle-invasive bladder cancer. Despite this clinical experience, the mechanism by which BCG stimulates tumor-eliminating immunity is unclear, and there is still a need for more accurate prediction of clinical outcomes in advance of treatment initiation. We have shown that BCG stimulates tumor-specific T-cell immunity that requires tumor cell expression of the IFNγ receptor (IFNGR); however, the downstream components of IFNGR signaling responsible for responsiveness to BCG are unknown. Here, we demonstrate that the IFNγ-driven, tumor cell intrinsic expression of the class II transactivator CIITA is required for activation of a tumor-specific CD4 T-cell response and BCG-induced tumor immunity. Despite the established role for CIITA in controlling MHC-II antigen presentation machinery, the requirement for CIITA is independent of MHC-II and associated genes. Rather, we find that CIITA is required for a broader tumor-intrinsic transcriptional program linked to critical pathways of tumor immunity via mechanisms that remain to be determined. Tumor cell intrinsic expression of CIITA is not required for a response to immunotherapy targeting programmed cell death protein 1 (PD-1), suggesting that different modalities of immunotherapy for bladder cancer could be employed based on tumor-intrinsic characteristics.

Expression of novel neuroendocrine markers in breast carcinomas: a study of INSM1, ASCL1, and POU2F3.

INSM1, ASCL1, and POU2F3 are novel transcription factors involved in neuroendocrine (NE) differentiation of neoplasms in several organs, but data on their expression in breast carcinomas (BCs) are limited. We retrospectively evaluated the expression of these markers in a series of 97 BCs (58 with NE morphology and 39 with otherwise uncommon morphology) tested prospectively using immunohistochemistry (IHC). Nuclear staining in >50% of the cells was used as the positive cut-off. Thirty-two of the 97 BCs (33%) were INSM1-positive. INSM1-positivity correlated significantly with histologic type and presence of stromal mucin. INSM1 also correlated with synaptophysin and chromogranin, established markers of NE differentiation (P < .0001 and P = .0023, respectively). In BC with NE morphology, the expression of INSM1 supported NE differentiation, and INSM1 was more specific than synaptophysin and more sensitive and specific than chromogranin. INSM1 was the most expressed NE marker in 17 BCs. INSM1-positive BCs included 56% of solid papillary BCs, 88% of BCs with solid papillary features, and 75% of high-grade NE carcinomas. Of 35 BCs tested for POU2F3 and ASCL1, only 1 and 4 cases were positive, respectively. Our results show that INSM1 is a sensitive marker of NE differentiation in BC and should be included with synaptophysin and chromogranin in the IHC panel used to evaluate NE differentiation in BC with NE morphology. ASCL1 and POU2F3 are uncommon in BC and their routine assessment is not warranted.

POU2F3 in SCLC: Clinicopathologic and Genomic Analysis With a Focus on Its Diagnostic Utility in Neuroendocrine-Low SCLC.

INTRODUCTION: POU2F3 is a recent marker of a small cell lung carcinoma (SCLC) subtype related to chemosensory tuft cells (SCLC-P). The characteristics of SCLC-P have not been fully defined, and the data on POU2F3 expression in other lung tumors are scarce. METHODS: We screened 254 SCLC for POU2F3 expression and comprehensively analyzed histopathologic, genomic, and clinical characteristics of POU2F3-positive tumors. We also explored POU2F3 expression in other major lung cancer types (n = 433) and a targeted set of potential diagnostic mimics of SCLC (n = 123). RESULTS: POU2F3 was expressed in 30 of 254 (12%) SCLC and was strongly associated with low expression of standard neuroendocrine markers (synaptophysin, chromogranin A, CD56, INSM1). Notably, POU2F3 was expressed in 75% of SCLC with entirely negative or minimal neuroendocrine marker expression (15/20) and was helpful in supporting the diagnosis of SCLC in such cases. Broad targeted next-generation sequencing revealed that SCLC-P (n = 12) exhibited enrichment in several alterations, including PTEN inactivation, MYC amplifications, and 20q13 amplifications, but similar rates of RB1 and TP53 alterations as other SCLC (n = 155). Beyond SCLC, POU2F3 expression was exclusively limited to large cell neuroendocrine carcinoma (12%) and basaloid squamous cell carcinoma (22%). CONCLUSIONS: This is the largest cohort of SCLC-P clinical samples to date, where we describe the diagnostic utility of POU2F3 in a challenging subset of SCLC with low or absent expression of standard neuroendocrine markers. The distinct genomic alterations in SCLC-P may offer a novel avenue for therapeutic targeting. The role of POU2F3 in a narrow subset of other lung cancer types warrants further study.

Cancer testis antigen (PRAME) as an independent marker for survival in oral squamous cell carcinoma (OSCC).

BACKGROUND: The objective was to assess the expression patterns of the cancer testis antigen PRAME, NY-ESO1, and SSX2 in oral squamous cell carcinoma (OSSC) and to correlate the expression with clinical and histopathological parameters including progression-free survival analysis. METHODS: The study variables of this retrospective cohort study (n = 83) included demographic data, histopathological data, and information on progression-free survival. PRAME expression patterns were rated based on immunohistochemistry on tissue microarrays (TMA). The survival rate was assessed by Kaplan-Meier method and Cox regression model. The primary predictor variable was defined as the expression of PRAME and the outcome variable was progression-free survival. RESULTS: Analysis of progression-free survival using Kaplan-Meier method showed that patients with positive expression of PRAME had lower probabilities of progression-free survival (p < 0.001). According to the Cox regression model, the level of PRAME expression had a considerable and significant independent influence on progression-free survival (positive PRAME expression increasing the hazards for a negative outcome by 285% in our sample; HR = 3.85, 95% CI: 1.45-10.2, p = 0.007). The expression of SSX2 (n = 1) and NY-ESO-1 (n = 5) in our samples was rare. CONCLUSION: PRAME is expressed in OSCC and appears to be a suitable marker of progression-free survival, correlates with severe course, and may allow identification of high-risk patients with aggressive progression.

Anatomic position determines oncogenic specificity in melanoma.

Oncogenic alterations to DNA are not transforming in all cellular contexts1,2. This may be due to pre-existing transcriptional programmes in the cell of origin. Here we define anatomic position as a major determinant of why cells respond to specific oncogenes. Cutaneous melanoma arises throughout the body, whereas the acral subtype arises on the palms of the hands, soles of the feet or under the nails3. We sequenced the DNA of cutaneous and acral melanomas from a large cohort of human patients and found a specific enrichment for BRAF mutations in cutaneous melanoma and enrichment for CRKL amplifications in acral melanoma. We modelled these changes in transgenic zebrafish models and found that CRKL-driven tumours formed predominantly in the fins of the fish. The fins are the evolutionary precursors to tetrapod limbs, indicating that melanocytes in these acral locations may be uniquely susceptible to CRKL. RNA profiling of these fin and limb melanocytes, when compared with body melanocytes, revealed a positional identity gene programme typified by posterior HOX13 genes. This positional gene programme synergized with CRKL to amplify insulin-like growth factor (IGF) signalling and drive tumours at acral sites. Abrogation of this CRKL-driven programme eliminated the anatomic specificity of acral melanoma. These data suggest that the anatomic position of the cell of origin endows it with a unique transcriptional state that makes it susceptible to only certain oncogenic insults.

VISTA in Soft Tissue Sarcomas: A Perspective for Immunotherapy?

(1) Background: V domain immunoglobulin suppressor of T cell activation (VISTA) plays a critical role in antitumor immunity and may be a valuable target in cancer immunotherapy. To date, it has never been studied in a large and well-characterised cohort of soft tissue sarcomas (STS). (2) Methods: Using immunohistochemistry, we examined VISTA expression in tumour tissues of 213 high-risk STS. We then analysed whether VISTA was associated with other clinicopathological parameters, including tumour-infiltrating lymphocyte (TIL) counts, programmed death receptor-1 (PD1), programmed death ligand-1 (PDL1), CD3, grading, and long-term survival. (3) Results: We observed VISTA expression in 96 (45%) of 213 specimens with distinct patterns ranging from 26 to 63% for histological subtypes. VISTA was associated with higher grade (G3 vs. G2, p = 0.019), higher TIL counts (p = 0.033), expression of PD1 (p = 0.046), PDL1 (p = 0.031), and CD3+ (p = 0.023). In patients without CD3+ TILs, 10-year survival was higher when VISTA was expressed compared to when there was no VISTA expression (p = 0.013). In a multivariate analysis, VISTA expression was independently associated with prolonged survival (p = 0.043). (4) Conclusions: VISTA is expressed in different STS subtypes and is associated with increased TILs, PD-1, PD-L1, and CD3 expression. Patients with VISTA+ tumours show improved survival. These results may help define future immunotherapeutic approaches in STS.

Genomic characterization of small cell carcinomas of the uterine cervix.

Small cell carcinoma (SCC) of the uterine cervix is a rare and aggressive form of neuroendocrine carcinoma, which resembles small cell lung cancer (SCLC) in its histology and poor survival rate. Here, we sought to define the genetic underpinning of SCCs of the uterine cervix and compare their mutational profiles with those of human papillomavirus (HPV)-positive head and neck squamous cell carcinomas, HPV-positive cervical carcinomas, and SCLCs using publicly available data. Using a combination of whole-exome and targeted massively parallel sequencing, we found that the nine uterine cervix SCCs, which were HPV18-positive (n = 8) or HPV16-positive (n = 1), harbored a low mutation burden, few copy number alterations, and other than TP53 in two cases no recurrently mutated genes. The majority of mutations were likely passenger missense mutations, and only few affected previously described cancer-related genes. Using RNA-sequencing, we identified putative viral integration sites on 18q12.3 and on 8p22 in two SCCs of the uterine cervix. The overall nonsilent mutation rate of uterine cervix SCCs was significantly lower than that of SCLCs, HPV-driven cervical adeno- and squamous cell carcinomas, or HPV-positive head and neck squamous cell carcinomas. Unlike SCLCs, which are reported to harbor almost universal TP53 and RB1 mutations and a dominant tobacco smoke-related signature 4, uterine cervix SCCs rarely harbored mutations affecting these genes (2/9, 22% TP53; 0% RB1) and displayed a dominant aging (67%) or APOBEC mutational signature (17%), akin to HPV-driven cancers, including cervical adeno- and squamous cell carcinomas and head and neck squamous cell carcinomas. Taken together, in contrast to SCLCs, which are characterized by highly recurrent TP53 and RB1 alterations, uterine cervix SCCs were positive for HPV leading to inactivation of the suppressors p53 and RB, suggesting that these SCCs are convergent phenotypes.

TSC2-mutant uterine sarcomas with JAZF1-SUZ12 fusions demonstrate hybrid features of endometrial stromal sarcoma and PEComa and are responsive to mTOR inhibition.

Uterine perivascular epithelioid cell tumor (PEComa) is a rare mesenchymal neoplasm that occasionally shares morphologic and immunohistochemical overlap with low- and high-grade endometrial stromal sarcoma (LGESS and HGESS). In this study, we sought to characterize the clinical, morphologic, genetic, and epigenetic features of five uterine sarcomas that display histologic features of LGESS, HGESS, and PEComa. All tumors demonstrated epithelioid cells often associated with a low-grade spindled component resembling LGESS, with both regions expressing CD10, ER, PR, variable HMB45, and Melan-A immunoreactivity, and strong cathepsin K and pS6 expression. Targeted massively parallel sequencing analysis revealed the presence of somatic TSC2 mutations in all five cases, of which four harbored concurrent or consecutive JAZF1-SUZ12 gene fusions. Unsupervised hierarchical clustering analysis of methylation profiles of TSC2-mutant uterine sarcomas (n = 4), LGESS (n = 10), and HGESS (n = 12) demonstrated two clusters consisting of (1) all LGESS and TSC2-mutant uterine sarcomas and (2) all HGESS. KEGG pathway analysis detected methylation differences in genes involved in PI3K/AKT, calcium, and Rap1 signaling. TSC2-mutant uterine sarcomas were responsive to hormone suppression, and mTOR inhibition demonstrated clinical benefit in four patients with these neoplasms. Our results suggest that these tumors represent histologically distinctive LGESS with TSC2 mutations. TSC2 mutations and JAZF1-SUZ12 fusion may help diagnose these tumors and possibly direct effective treatment.

Expression Analysis of GD2 by Immunohistochemistry in Invasive Breast Carcinoma: Clinical and Pathologic Correlation.

The glycosphingolipid disialoganglioside GD2 is a cell surface-associated antigen expressed on tumors of neuroectodermal origin that serves as a target of immunotherapy in select cancer types. Information about the expression of GD2 in breast cancer is limited. In the present study, we investigate the utility of GD2 as a potential biomarker for targeted treatment. The study cohort consists of 386 breast carcinomas of several histologic types. GD2 expression was assessed in both whole tumor sections and tissue microarrays with anti-GD2 3F8 monoclonal antibody immunohistochemistry and correlated with clinicopathologic features and survival outcomes. A total of 134 (35%) breast carcinomas were positive for GD2, with a median H-score of 100. 3F8 staining displayed granular and predominantly cytoplasmic or perinuclear patterns, which was confined to the neoplastic tissue in nearly all cases. GD2 positivity was significantly associated with tumor histologic type (P=0.0015), low grade (P<0.0001), estrogen receptor positivity (P<0.0001), low stage (P=0.0014), and multifocality (P=0.022). Event-free survival and overall survival of patients with GD2-positive and GD2-negative tumors were not significantly different. Our results support further assessment of GD2 using the 3F8 antibody as a predictive and prognostic biomarker in breast cancer.

IDH2 R172 Mutations Across Poorly Differentiated Sinonasal Tract Malignancies: Forty Molecularly Homogenous and Histologically Variable Cases With Favorable Outcome.

IDH2 R172 mutations occur in sinonasal undifferentiated carcinoma (SNUC), large-cell neuroendocrine carcinoma (LCNEC), sinonasal adenocarcinomas, and olfactory neuroblastoma (ONB). We performed a clinical, pathologic, and genetic/epigenetic analysis of a large IDH2-mutated sinonasal tumor cohort to explore their distinct features. A total 165 sinonasal/skull base tumors included 40 IDH2 mutants studied by light microscopy, immunohistochemistry, and genome-wide DNA methylation, and 125 IDH2 wild-type tumors used for comparison. Methylation profiles were analyzed by unsupervised hierarchical clustering, t-distributed stochastic neighbor embedding dimensionality reduction and assessed for copy number alterations (CNA). Thirty-nine histologically assessable cases included 25 (64.1%) SNUC, 8 (20.5%) LCNEC, 2 (5.1%) poorly differentiated adenocarcinomas, 1 (2.7%) ONB, and 3 (7.7%) IDH2-mutated tumors with ONB features. All cases were high-grade showing necrosis (82.4%), prominent nucleoli (88.9%), and median 21 mitoses/10 HPFs. AE1/AE3 and/or CAM 5.2 were positive in all and insulinoma-associated protein 1 (INSM1) in 80% cases. All IDH2 mutants formed one distinct group by t-distributed stochastic neighbor embedding dimensionality reduction separating from all IDH2 wild-type tumors. There was no correlation between methylation clusters and histopathologic diagnoses. Recurrent CNA included 1q gain (79.3%), 17p loss (75.9%), and 17q gain (58.6%). No CNA differences were observed between SNUC and LCNEC. IDH2 mutants showed better disease-specific survival than SMARCB1-deficient (P=0.027) and IDH2 wild-type carcinomas overall (P=0.042). IDH2-mutated sinonasal tumors are remarkably homogeneous at the molecular level and distinct from IDH2 wild-type sinonasal malignancies. Biology of IDH2-mutated sinonasal tumors might be primarily defined by their unique molecular fingerprint rather than by their respective histopathologic diagnoses.

PD-L1 Expression in Metaplastic Breast Carcinoma Using the PD-L1 SP142 Assay and Concordance Among PD-L1 Immunohistochemical Assays.

Immunotherapy for the treatment of programmed death-ligand 1 (PD-L1) positive locally advanced or metastatic triple negative breast cancer may benefit patients with metaplastic breast cancer (MpBC). Previous study of PD-L1 in MpBC scored tumor cells (TCs), different from Food and Drug Administration-approved scoring methods. We sought to define PD-L1 expression in MpBCs and to evaluate concordance of 3 PD-L1 assays. Primary, treatment naive MpBC treated at our Center from 1998 to 2019 were identified. PD-L1 expression was assessed using SP142, E1L3n, and 73-10. We evaluated PD-L1 expression on tumor infiltrating immune cells (IC) and also in TCs. For each assay, we scored PD-L1 expression using ≥1% IC expression according to the IMpassion130 trial criteria and using combined positive score (CPS) ≥10 according to the KEYNOTE-355 trial cutoff. A total of 42 MpBCs were identified. Most MpBC had PD-L1 positivity in ≥1% IC with all 3 assays (95%, 95%, 86%) in contrast to a maximum 71% with a CPS ≥10. PD-L1 IC expression was comparable between the SP142 and 73-10 assays and was lowest with E1L3n. PD-L1 TC expression was lowest using SP142. The overall concordance for IC scoring was 88% while 62% had concordant CPS. For each assay, the results of the 2 scoring algorithms were not interchangeable. The SP142 assay showed distinct expression patterns between IC (granular, dot-like) and TC (membranous) while 73-10 and E1L3n showed membranous and/or cytoplasmic expression in both IC and TC. Most MpBC in our cohort were positive for PD-L1 indicating eligibility for anti-PD-L1/programmed death-1 immunotherapy.

PRAME Immunohistochemistry as an Ancillary Test for the Assessment of Melanocytic Lesions.

PRAME (PReferentially expressed Antigen in MElanoma) is a melanoma-associated antigen expressed in cutaneous and ocular melanomas and some other malignant neoplasms, while its expression in normal tissue and benign tumors is limited. Detection of PRAME protein expression by immunohistochemistry in a cohort of 400 melanocytic tumors showed diffuse nuclear immunoreactivity for PRAME in most metastatic and primary melanomas. In contrast, most nevi were negative for PRAME or showed nondiffuse immunoreactivity. The difference in the extent of immunoreactivity for PRAME in unambiguous melanocytic tumors prompted the study of PRAME as an ancillary tool for evaluating melanocytic lesions in more challenging scenarios.

Immunohistochemical Detection of Cancer-Testis Antigen PRAME.

Cancer-testis (CT) antigens were identified by their ability to elicit T- or B-cell immune responses in the autologous host. They are typically expressed in a wide variety of neoplasms and in normal adult tissues are restricted to testicular germ cells. PReferentially expressed Antigen of Melanoma (PRAME) is a member of the family of nonclassical CT antigens being expressed in a few other normal tissues besides testis. Interestingly, knowledge about the protein expression of many CT antigens is still incomplete due to the limited availability of reagents for their immunohistochemical detection. Here, we tested several commercially available serological reagents and identified a monoclonal antibody suitable for the immunohistochemical detection of PRAME in formalin-fixed paraffin-embedded specimens. We also tested a wide array of normal and neoplastic tissues. PRAME protein expression in normal tissues is congruent with original molecular data being present in the testis, and at low levels in the endometrium, adrenal cortex, and adult as well as fetal ovary. In tumors, there is diffuse PRAME immunoreactivity in most metastatic melanomas, myxoid liposarcomas, and synovial sarcomas. Other neoplasms such as seminomas and carcinomas of various origins including endometrial, serous ovarian, mammary ductal, lung, and renal showed an intermediate proportion of cases and variable extent of tumor cells positive for PRAME protein expression. As seen with other CT antigens, hepatocellular and colorectal carcinoma, Leydig cell tumors, mesothelioma, and leiomyosarcoma are poor expressers of PRAME.

Clonal relationship and directionality of progression of synchronous endometrial and ovarian carcinomas in patients with DNA mismatch repair-deficiency associated syndromes.

Sporadic synchronous endometrial (ECs) and ovarian cancers (OCs), although clinically considered to be independent primaries, have been shown to be clonally related and likely constitute metastases from each other. We sought to define whether synchronous ECs/OCs in patients with DNA mismatch repair (MMR)-deficiency syndromes would be clonally related. We subjected synchronous ECs/OCs from four patients (LS3-LS6) with clinically confirmed Lynch syndrome (LS) and one patient with constitutional mismatch repair-deficiency syndrome (CMMRD) to massively parallel sequencing targeting 468 cancer-related genes. Somatic mutations, copy number alterations (CNAs), clonal relatedness and clonal decomposition analyses were performed using previously described bioinformatics methods. All synchronous ECs/OCs analyzed were considered independent primaries based on clinicopathologic criteria. Sequencing analysis revealed that the ECs/OCs of three cases (LS2-CMMRD, L3, L4) harbored similar repertoires of somatic mutations and CNAs and were clonally related. In these three cases, a subset of subclonal mutations in the EC became clonal in the OC, suggesting that the EC was likely the substrate from which the OC developed. LS5's EC/OC had distinct mutational profiles but shared specific CNAs. In contrast, LS6's EC/OC harbored distinct somatic mutations and lacked CNAs, consistent with each tumor constituting an independent primary lesion. In LS5 and LS6, PTEN mutations and PTEN loss of protein expression were found to be restricted to the EC. Finally, re-analysis of sequencing data of sporadic synchronous ECs/OCs supported the observations made in the current study that the directionality of progression is likely from the endometrium to the ovary. In conclusion, contrary to sporadic synchronous ECs/OCs, which are almost invariably clonally related, ECs/OCs simultaneously involving the uterus and ovary in LS patients may represent distinct primary tumors. A subset of MMR-deficiency syndrome-related synchronous ECs/OCs, however, may originate from a single primary tumor at variance with their clinical diagnosis, with the endometrium being the likeliest site of origin.

A phase II trial of durvalumab and tremelimumab in metastatic, non-urothelial carcinoma of the urinary tract.

BACKGROUND: Immune checkpoint blockade has made a significant impact on the clinical outcomes of patients with metastatic urothelial carcinoma (UC). However, evidence for this approach in patients with non-UC of the urinary tract is limited. METHODS: This was a phase II open-label study of durvalumab 1500 mg and tremelimumab 75 mg every 4 weeks for four cycles followed by durvalumab 1500 mg every 4 weeks. Eligible patients had metastatic non-UC with ECOG PS 0-1 regardless of prior therapy (except small cell carcinoma who were pretreated). The primary endpoint was overall response rate per RECIST v1.1. A Simon's minimax two-stage design was employed, with 13 patients planned for stage one. Pre-treatment tumors underwent PD-L1 staining and next-generation sequencing. RESULTS: Thirteen patients were treated, including seven small cell carcinoma, three squamous cell carcinoma, and three adenocarcinoma. Eleven patients had visceral metastases. No responses were observed; 11 patients had PD and 2 patients had SD. Median PFS was 1.8 months (95% CI, 1.25-not reached [NR]) with a median follow-up of 7.38 months (range, 5.23-21.99 months). Median OS was 6.97 months (95% CI, 4.34-NR). One patient's tumor was PD-L1 positive and all sequenced tumors (n = 8) were microsatellite stable. Grades 3-4 treatment-related adverse events occurred in 38.4% of patients. CONCLUSIONS: In a poor prognosis cohort of patients with non-UC, durvalumab and tremelimumab lacked clinical activity while demonstrating a manageable safety profile.

Genomic Profiling Aids Classification of Diagnostically Challenging Uterine Mesenchymal Tumors With Myomelanocytic Differentiation.

Although diagnosis of high-grade uterine mesenchymal tumors (UMTs) exhibiting classic morphologic features is straightforward, diagnosis is more challenging in tumors in which prototypical features are poorly developed, focal, and/or coexist with features seen in other neoplasms. Here, we sought to define the repertoire of somatic genetic alterations in diagnostically challenging UMTs with myomelanocytic differentiation, including some reported as perivascular epithelioid cell tumors (PEComas). In 17 samples from 15 women, the tumors were histologically heterogenous. Immunohistochemical expression of at least 1 melanocytic marker (HMB45, Melan-A, or MiTF) was identified in all tumors, and of myogenic markers (desmin or smooth muscle actin) in most tumors. Targeted massively parallel sequencing revealed several genetic alterations, most commonly in TP53 (41% mutation, 12% deletion), TSC2 (29% mutation, 6% deletion), RB1 (18% deletion), ATRX (24% mutation), MED12 (12% mutation), BRCA2 (12% deletion), CDKN2A (6% deletion) as well as FGFR3, NTRK1, and ERBB3 amplification (each 6%). Gene rearrangements (JAZF1-SUZ12; DNAJB6-PLAG1; and SFPQ-TFE3) were identified in 3 tumors. Integrating histopathologic, immunohistochemical, and genetic findings, tumors from 4 patients were consistent with malignant PEComa (1 TFE3-rearranged); 6 were classified as leiomyosarcomas; 3 showed overlapping features of PEComa and other sarcoma types (leiomyosarcoma or low-grade endometrial stromal sarcoma); and 2 were classified as sarcoma, not otherwise specified. Our findings suggest that diagnostically challenging UMTs with myomelanocytic differentiation represent a heterogenous group of neoplasms which harbor a diverse repertoire of somatic genetic alterations; these genetic alterations can aid classification.