Diffuse pleural mesotheliomas with genomic near-haploidization: a newly recognized subset with distinct clinical, histologic, and molecular features.

PURPOSE: Diffuse pleural mesotheliomas (DPMs) with genomic near-haploidization (GNH) represent a novel subtype first recognized by the TCGA project; however, its clinicopathologic and molecular features remain poorly defined. EXPERIMENTAL DESIGN: We analyzed clinical genomic profiling data from 290 patients with DPM using the MSK-IMPACT assay. Allele-specific copy number analysis was performed using the FACETS algorithm. RESULTS: 210 patients were evaluable for LOH analysis using FACETS. In this cohort, GNH was detected in 10 cases (4.8%). Compared to non-GNH tumors, GNH DPMs were associated with younger age and less frequent self-reported history of occupational asbestos exposure. Histologically, GNH DPMs were enriched in biphasic subtype (80% vs. 14.5%) and showed abundant tumor infiltrating lymphocytes (TILs). Genomic analysis revealed a higher frequency of TP53 alterations, while SETDB1 mutations were present in nearly all and only in this subset. The clinicopathologic and molecular findings were further validated in a separate cohort. Despite the younger age, patients with GNH DPMs had a shorter overall survival (10.9 vs. 25.4 months, p=0.004); the poor prognostic impact of GNH remained significant after controlling for biphasic histology. Out of three patients with GNH DPMs who received immune checkpoint blockade (ICB), two achieved a clinician assessed partial response. CONCLUSIONS: GNH defines an aggressive subtype of mainly biphasic DPMs in younger patients with recurrent alterations in SETDB1 and TP53. The enrichment in biphasic histology and TILs, together with our preliminary ICB response data and anecdotal clinical trial data, suggests that further evaluation of immunotherapy may be warranted in this subset.

Enhanced clinical assessment of hematologic malignancies through routine paired tumor and normal sequencing.

Genomic profiling of hematologic malignancies has augmented our understanding of variants that contribute to disease pathogenesis and supported development of prognostic models that inform disease management in the clinic. Tumor only sequencing assays are limited in their ability to identify definitive somatic variants, which can lead to ambiguity in clinical reporting and patient management. Here, we describe the MSK-IMPACT Heme cohort, a comprehensive data set of somatic alterations from paired tumor and normal DNA using a hybridization capture-based next generation sequencing platform. We highlight patterns of mutations, copy number alterations, and mutation signatures in a broad set of myeloid and lymphoid neoplasms. We also demonstrate the power of appropriate matching to make definitive somatic calls, including in patients who have undergone allogeneic stem cell transplant. We expect that this resource will further spur research into the pathobiology and clinical utility of clinical sequencing for patients with hematologic neoplasms.

Mutant-RB1 circulating tumor DNA in the blood of unilateral retinoblastoma patients: What happens during enucleation surgery: A pilot study.

Cell free DNA (cfDNA) and circulating tumor cell free DNA (ctDNA) from blood (plasma) are increasingly being used in oncology for diagnosis, monitoring response, identifying cancer causing mutations and detecting recurrences. Circulating tumor RB1 DNA (ctDNA) is found in the blood (plasma) of retinoblastoma patients at diagnosis before instituting treatment (naïve). We investigated ctDNA in naïve unilateral patients before enucleation and during enucleation (6 patients/ 8 mutations with specimens collected 5-40 minutes from severing the optic nerve) In our cohort, following transection the optic nerve, ctDNA RB1 VAF was measurably lower than pre-enucleation levels within five minutes, 50% less within 15 minutes and 90% less by 40 minutes.

Overall survival with circulating tumor DNA-guided therapy in advanced non-small-cell lung cancer.

Circulating tumor DNA (ctDNA) sequencing guides therapy decisions but has been studied mostly in small cohorts without sufficient follow-up to determine its influence on overall survival. We prospectively followed an international cohort of 1,127 patients with non-small-cell lung cancer and ctDNA-guided therapy. ctDNA detection was associated with shorter survival (hazard ratio (HR), 2.05; 95% confidence interval (CI), 1.74-2.42; P < 0.001) independently of clinicopathologic features and metabolic tumor volume. Among the 722 (64%) patients with detectable ctDNA, 255 (23%) matched to targeted therapy by ctDNA sequencing had longer survival than those not treated with targeted therapy (HR, 0.63; 95% CI, 0.52-0.76; P < 0.001). Genomic alterations in ctDNA not detected by time-matched tissue sequencing were found in 25% of the patients. These ctDNA-only alterations disproportionately featured subclonal drivers of resistance, including RICTOR and PIK3CA alterations, and were associated with short survival. Minimally invasive ctDNA profiling can identify heterogeneous drivers not captured in tissue sequencing and expand community access to life-prolonging therapy.

Diagnostic yield and clinical relevance of expanded genetic testing for cancer patients.

BACKGROUND: Genetic testing (GT) for hereditary cancer predisposition is traditionally performed on selected genes based on established guidelines for each cancer type. Recently, expanded GT (eGT) using large hereditary cancer gene panels uncovered hereditary predisposition in a greater proportion of patients than previously anticipated. We sought to define the diagnostic yield of eGT and its clinical relevance in a broad cancer patient population over a 5-year period. METHODS: A total of 17,523 cancer patients with a broad range of solid tumors, who received eGT at Memorial Sloan Kettering Cancer Center between July 2015 to April 2020, were included in the study. The patients were unselected for current GT criteria such as cancer type, age of onset, and/or family history of disease. The diagnostic yield of eGT was determined for each cancer type. For 9187 patients with five common cancer types frequently interrogated for hereditary predisposition (breast, colorectal, ovarian, pancreatic, and prostate cancer), the rate of pathogenic/likely pathogenic (P/LP) variants in genes that have been associated with each cancer type was analyzed. The clinical implications of additional findings in genes not known to be associated with a patients' cancer type were investigated. RESULTS: 16.7% of patients in a broad cancer cohort had P/LP variants in hereditary cancer predisposition genes identified by eGT. The diagnostic yield of eGT in patients with breast, colorectal, ovarian, pancreatic, and prostate cancer was 17.5%, 15.3%, 24.2%, 19.4%, and 15.9%, respectively. Additionally, 8% of the patients with five common cancers had P/LP variants in genes not known to be associated with the patient's current cancer type, with 0.8% of them having such a variant that confers a high risk for another cancer type. Analysis of clinical and family histories revealed that 74% of patients with variants in genes not associated with their current cancer type but which conferred a high risk for another cancer did not meet the current GT criteria for the genes harboring these variants. One or more variants of uncertain significance were identified in 57% of the patients. CONCLUSIONS: Compared to targeted testing approaches, eGT can increase the yield of detection of hereditary cancer predisposition in patients with a range of tumors, allowing opportunities for enhanced surveillance and intervention. The benefits of performing eGT should be weighed against the added number of VUSs identified with this approach.

Molecular Characterization of Peritoneal Mesotheliomas.

INTRODUCTION: Malignant peritoneal mesothelioma (MPeM) is clinically distinct and less studied than malignant pleural mesothelioma. We report the genomic and immunophenotypic features of a prospectively collected MPeM cohort. METHODS: Next-generation sequencing (NGS) was performed on MPeM tumors. Genomic near-haploidization (GNH) was assessed. WT1, BAP1, mesothelin, VISTA, and programmed death-ligand 1 were evaluated by immunohistochemistry (IHC) when tissue was available. Overall survival was stratified by selected genomic and IHC features. RESULTS: A total of 50 consented patients with MPeM (45 epithelioid, 5 nonepithelioid) were studied exhibiting common alterations in BAP1 (60%; 30 of 50), NF2 (24%; 12 of 50) SETD2 (22%; 11 of 50), and TP53 (16%; 8 of 50). A total of 76% (38 of 50) of specimens were assessable for allele-specific copy number analysis; 8% (3 of 38) had GNH. IHC positivity rates were 93% (37 of 40) for mesothelin, 96% (46 of 48) for WT1, 50% (19 of 38) for programmed death-ligand 1, and 89% (34 of 38) for VISTA. BAP1 loss by IHC was observed in 76% (29 of 38), including five wild-type on NGS. Combining NGS and IHC for BAP1, overall survival was worse with alteration or loss compared with wild-type or retained in all patients (n = 37 versus 13, 43.8 versus 117.3 mo, p = 0.04) Three of 30 patients had a pathogenic germline variant: POT1 I78T, MUTYH R109Y, and BAP1 E402∗. CONCLUSIONS: MPeM has distinct biology and genomic composition. CDKN2A/B alterations were rare in MPeM, whereas BAP1, NF2, TP53, SETD2, and LATS2 were common. BAP1 alteration/loss was associated with shorter survival when all patients were included. A notable minority of specimens had GNH associated with NF2, TP53, and SETDB1 mutations. Pathogenic germline mutations were found in 3 of 30 patients.

The context-specific role of germline pathogenicity in tumorigenesis.

Human cancers arise from environmental, heritable and somatic factors, but how these mechanisms interact in tumorigenesis is poorly understood. Studying 17,152 prospectively sequenced patients with cancer, we identified pathogenic germline variants in cancer predisposition genes, and assessed their zygosity and co-occurring somatic alterations in the concomitant tumors. Two major routes to tumorigenesis were apparent. In carriers of pathogenic germline variants in high-penetrance genes (5.1% overall), lineage-dependent patterns of biallelic inactivation led to tumors exhibiting mechanism-specific somatic phenotypes and fewer additional somatic oncogenic drivers. Nevertheless, 27% of cancers in these patients, and most tumors in patients with pathogenic germline variants in lower-penetrance genes, lacked particular hallmarks of tumorigenesis associated with the germline allele. The dependence of tumors on pathogenic germline variants is variable and often dictated by both penetrance and lineage, a finding with implications for clinical management.

Paired Tumor-Normal Sequencing Provides Insights Into the TP53-Related Cancer Spectrum in Patients With Li-Fraumeni Syndrome.

BACKGROUND: Genetic testing for Li-Fraumeni syndrome (LFS) is performed by using blood specimens from patients selected based on phenotype-dependent guidelines. This approach is problematic for understanding the LFS clinical spectrum because patients with nonclassical presentations are missed, clonal hematopoiesis-related somatic blood alterations cannot be distinguished from germline variants, and unrelated tumors cannot be differentiated from those driven by germline TP53 defects. METHODS: To provide insights into the LFS-related cancer spectrum, we analyzed paired tumor-blood DNA sequencing results in 17 922 patients with cancer and distinguished clonal hematopoiesis-related, mosaic, and germline TP53 variants. Loss of heterozygosity and TP53 mutational status were assessed in tumors, followed by immunohistochemistry for p53 expression on a subset to identify those lacking biallelic TP53 inactivation. RESULTS: Pathogenic/likely pathogenic TP53 variants were identified in 50 patients, 12 (24.0%) of which were clonal hematopoiesis related and 4 (8.0%) of which were mosaic. Twelve (35.3%) of 34 patients with germline TP53 variants did not meet LFS testing criteria. Loss of heterozygosity of germline TP53 variant was observed in 96.0% (95% confidence interval [CI] = 79.7% to 99.9%) of core LFS spectrum-type tumors vs 45.5% (95% CI = 16.8% to 76.6%) of other tumors and 91.3% (95% CI = 72.0% to 98.9%) of tumors from patients who met LFS testing criteria vs 61.5% (95% CI = 31.6% to 86.1%) of tumors from patients who did not. Tumors retaining the wild-type TP53 allele exhibited wild-type p53 expression. CONCLUSIONS: Our results indicate that some TP53 variants identified in blood-only sequencing are not germline and a substantial proportion of patients with LFS are missed based on current testing guidelines. Additionally, a subset of tumors from patients with LFS do not have biallelic TP53 inactivation and may represent cancers unrelated to their germline TP53 defect.

Prospective pan-cancer germline testing using MSK-IMPACT informs clinical translation in 751 patients with pediatric solid tumors.

The spectrum of germline predisposition in pediatric cancer continues to be realized. Here we report 751 solid tumor patients who underwent prospective matched tumor-normal DNA sequencing and downstream clinical use (clinicaltrials.gov NCT01775072). Germline pathogenic and likely pathogenic (P/LP) variants were reported. One or more P/LP variants were found in 18% (138/751) of individuals when including variants in low, moderate, and high penetrance dominant or recessive genes, or 13% (99/751) in moderate and high penetrance dominant genes. 34% of high or moderate penetrance variants were unexpected based on the patient's diagnosis and previous history. 76% of patients with positive results completed a clinical genetics visit, and 21% had at least one relative undergo cascade testing as a result of this testing. Clinical actionability additionally included screening, risk reduction in relatives, reproductive use, and use of targeted therapies. Germline testing should be considered for all children with cancer.

Therapeutic Implications of Germline Testing in Patients With Advanced Cancers.

PURPOSE: Tumor mutational profiling is increasingly performed in patients with advanced cancer. We determined the extent to which germline mutation profiling guides therapy selection in patients with advanced cancer. METHODS: Patients with cancer undergoing tumor genomic profiling were prospectively consented for germline cancer predisposition gene analysis (2015-2019). In patients harboring germline likely pathogenic or pathogenic (LP/P) alterations, therapeutic actionability was classified using a precision oncology knowledge base. Patients with metastatic or recurrent cancer receiving germline genotype-directed therapy were determined. RESULTS: Among 11,947 patients across > 50 malignancies, 17% (n = 2,037) harbored a germline LP/P variant. By oncology knowledge base classification, 9% (n = 1042) had an LP/P variant in a gene with therapeutic implications (4% level 1; 4% level 3B; < 1% level 4). BRCA1/2 variants accounted for 42% of therapeutically actionable findings, followed by CHEK2 (13%), ATM (12%), mismatch repair genes (11%), and PALB2 (5%). When limited to the 9,079 patients with metastatic or recurrent cancer, 8% (n = 710) harbored level 1 or 3B genetic findings and 3.2% (n = 289) received germline genotype-directed therapy. Germline genotype-directed therapy was received by 61% and 18% of metastatic cancer patients with level 1 and level 3B findings, respectively, and by 54% of BRCA1/2, 75% of mismatch repair, 43% of PALB2, 35% of RAD51C/D, 24% of BRIP1, and 19% of ATM carriers. Of BRCA1/2 patients receiving a poly(ADP-ribose) polymerase inhibitor, 45% (84 of 188) had tumors other than breast or ovarian cancer, wherein the drug, at time of delivery, was delivered in an investigational setting. CONCLUSION: In a pan-cancer analysis, 8% of patients with advanced cancer harbored a germline variant with therapeutic actionability with 40% of these patients receiving germline genotype-directed treatment. Germline sequence analysis is additive to tumor sequence analysis for therapy selection and should be considered for all patients with advanced cancer.

Enhanced specificity of clinical high-sensitivity tumor mutation profiling in cell-free DNA via paired normal sequencing using MSK-ACCESS.

Circulating cell-free DNA from blood plasma of cancer patients can be used to non-invasively interrogate somatic tumor alterations. Here we develop MSK-ACCESS (Memorial Sloan Kettering - Analysis of Circulating cfDNA to Examine Somatic Status), an NGS assay for detection of very low frequency somatic alterations in 129 genes. Analytical validation demonstrated 92% sensitivity in de-novo mutation calling down to 0.5% allele frequency and 99% for a priori mutation profiling. To evaluate the performance of MSK-ACCESS, we report results from 681 prospective blood samples that underwent clinical analysis to guide patient management. Somatic alterations are detected in 73% of the samples, 56% of which have clinically actionable alterations. The utilization of matched normal sequencing allows retention of somatic alterations while removing over 10,000 germline and clonal hematopoiesis variants. Our experience illustrates the importance of analyzing matched normal samples when interpreting cfDNA results and highlights the importance of cfDNA as a genomic profiling source for cancer patients.

Structure-function analysis of oncogenic EGFR Kinase Domain Duplication reveals insights into activation and a potential approach for therapeutic targeting.

Mechanistic understanding of oncogenic variants facilitates the development and optimization of treatment strategies. We recently identified in-frame, tandem duplication of EGFR exons 18 - 25, which causes EGFR Kinase Domain Duplication (EGFR-KDD). Here, we characterize the prevalence of ERBB family KDDs across multiple human cancers and evaluate the functional biochemistry of EGFR-KDD as it relates to pathogenesis and potential therapeutic intervention. We provide computational and experimental evidence that EGFR-KDD functions by forming asymmetric EGF-independent intra-molecular and EGF-dependent inter-molecular dimers. Time-resolved fluorescence microscopy and co-immunoprecipitation reveals EGFR-KDD can form ligand-dependent inter-molecular homo- and hetero-dimers/multimers. Furthermore, we show that inhibition of EGFR-KDD activity is maximally achieved by blocking both intra- and inter-molecular dimerization. Collectively, our findings define a previously unrecognized model of EGFR dimerization, providing important insights for the understanding of EGFR activation mechanisms and informing personalized treatment of patients with tumors harboring EGFR-KDD. Finally, we establish ERBB KDDs as recurrent oncogenic events in multiple cancers.

Retrospective Evaluation of Somatic Alterations in Cell-Free DNA from Blood in Retinoblastoma.

Purpose: Analysis of circulating tumor DNA (ctDNA) in the plasma of patients with retinoblastoma and simulating lesions. Design: Retrospective cross-sectional study of the association of plasma ctDNA from retinoblastoma and simulating lesions with disease course. Participants: Fifty-eight Memorial Sloan Kettering Cancer Center patients with retinoblastoma comprising 68 plasma ctDNA samples and 5 with retinoblastoma-simulating lesions. Methods: The ctDNA analyzed with hybridization capture and next-generation sequencing in blood (plasma) of patients who had retinoblastoma or simulating lesions were evaluated for association with clinical course of the disease. Main Outcome Measures: Presence or absence of molecular aberrations in the RB1 gene and correlations with clinical features. Results: RB1 cell-free DNA (cfDNA) was detected in 16 of 19 patients with newly diagnosed, untreated intraocular retinoblastoma and in 3 of 3 patients with newly diagnosed, untreated metastatic disease. It was also present in 3 patients with recurrent intraocular disease before therapy, but was not present in patients with recurrent disease who received intra-arterial chemotherapy, nor in 21 patients who had undergone enucleation for unilateral disease. In 1 patient who had delayed treatment (insurance reasons) and showed rapid growth of the intraocular tumor, the variant allele frequency increased in 1 month from 0.34% to 2.48%. No RB1 mutations were detected in the cfDNA from plasma of patients with simulating lesions (3 with Coats disease and 1 with persistent fetal vasculature [PFV]). In 2 patients, we identified 2 independent RB1 mutations in plasma. Conclusions: Mutations in RB1 were found in the cfDNA from blood of patients with newly diagnosed, untreated retinoblastoma and in patients who showed disease recurrence in the eye after prior treatment, but not in unilateral retinoblastoma after enucleation Levels of ctDNA increase in patients with progressive disease who did not receive any treatment. High plasma cfDNA levels were detected in patients with newly diagnosed metastatic disease, and these levels decreased after systemic chemotherapy was administered. Further validation is needed for measuring the somatic alterations in cfDNA from blood in retinoblastoma that could provide a promising method of monitoring patients in the future.

Adverse histology, homozygous loss of CDKN2A/B, and complex genomic alterations in locally advanced/metastatic renal mucinous tubular and spindle cell carcinoma.

Mucinous tubular and spindle cell carcinoma (MTSCC) is a rare subtype of renal cell carcinoma with characteristic histologic features and chromosomal alterations. Although typically indolent, a small subset of cases has been reported to exhibit aggressive clinical behavior. We retrospectively identified 33 patients with MTSCC, consisting of 10 cases of locally advanced/metastatic MTSCC (pT3 or N1 or M1) and 23 kidney-confined MTSCC (pT1/T2) without disease recurrence or progression. Utilizing a single-nucleotide polymorphism array and a targeted next-generation sequencing platform, we examined genome-wide molecular alterations in 24 cases, including 11 available samples from 8 patients with locally advanced/metastatic MTSCC. Ten patients with locally advanced/metastatic MTSCC were 8 females (80%) and 2 males (20%). At nephrectomy, 7 of these 10 cases (70%) were pT3 or pN1 while the remaining 3 (30%) were pT1/T2. Eight patients (80%) developed metastases and common sites included lymph node (4, 40%), bone (4, 40%), and retroperitoneum (3, 30%). Four patients died of disease (40%) during follow-up. Locally advanced/metastatic MTSCCs shared typical MTSCC genomic profiles with loss of chromosomes 1, 4, 6, 8, 9, 13, 14, 15, and 22, while some exhibited additional complex genomic alterations, most frequently a relative gain of 1q (7/8). Homozygous loss of CDKN2A/B was observed in 3 (38%) locally advanced/metastatic MTSCCs. Tumor necrosis, solid nested/sheet pattern, irregular trabecular/single-file infiltration in a desmoplastic stroma, lymphovascular space invasion, and increased mitotic activity were associated with locally advanced/metastatic MTSCCs (all p < 0.05). Our findings reveal that MTSCCs with aggressive clinical behavior have progressed through clonal evolution; CDKN2A/B deletion and additional complex genomic abnormalities may contribute to this process. Recognizing the morphologic presentation of high-grade MTSCC and evaluating adverse histologic features seen in these tumors can help establish a definitive diagnosis and stratify patients for treatment and prognostication.

A Performance Comparison of Commonly Used Assays to Detect RET Fusions.

PURPOSE: Selpercatinib and pralsetinib induce deep and durable responses in patients with advanced RET fusion-positive lung and thyroid cancer. RET fusion testing strategies with rapid and reliable results are critical given recent FDA approval. Here, we assess various clinical assays in a large pan-cancer cohort. EXPERIMENTAL DESIGN: Tumors underwent DNA-based next-generation sequencing (NGS) with reflex to RNA-based NGS if no mitogenic driver or if a RET structural variant of unknown significance (SVUS) were present. Canonical DNA-level RET fusions and RNA-confirmed RET fusions were considered true fusions. Break-apart FISH and IHC performance were assessed in subgroups. RESULTS: A total of 171 of 41,869 patients with DNA NGS harbored RET structural variants, including 139 canonical fusions and 32 SVUS. Twelve of 32 (37.5%) SVUS were transcribed into RNA-level fusions, resulting in 151 oncogenic RET fusions. The most common RET fusion-positive tumor types were lung (65.6%) and thyroid (23.2%). The most common partners were KIF5B (45%), CCDC6 (29.1%), and NCOA4 (13.3%). DNA NGS showed 100% (46/46) sensitivity and 99.6% (4,459/4,479) specificity. FISH showed 91.7% (44/48) sensitivity, with lower sensitivity for NCOA4-RET (66.7%, 8/12). A total of 87.5% (7/8) of RET SVUS negative for RNA-level fusions demonstrated rearrangement by FISH. The sensitivity of IHC varied by fusion partner: KIF5B sensitivity was highest (100%, 31/31), followed by CCDC6 (88.9%, 16/18) and NCOA4 (50%, 6/12). Specificity of RET IHC was 82% (73/89). CONCLUSIONS: Although DNA sequencing has high sensitivity and specificity, RNA sequencing of RET SVUS is necessary. Both FISH and IHC demonstrated lower sensitivity for NCOA4-RET fusions.

Genomic Alterations as Potential Therapeutic Targets in Extramammary Paget's Disease of the Vulva.

PURPOSE: To identify genomic alterations as potential therapeutic targets in extramammary Paget disease (EMPD) of the vulva. METHODS: We identified all patients with primary vulvar EMPD who were treated at our institution and underwent paired tumor-normal massively parallel sequencing of 410-468 cancer-related genes (MSK-IMPACT assay). EMPD of the vulva samples sequenced from 2014 to 2019 were reviewed and somatic mutations identified, with specific focus on mutations of potential therapeutic targets. Clinical data were abstracted from electronic medical records. Microsatellite instability (MSI) was assessed by MSIscore. RESULTS: Tumors of 26 patients with EMPD underwent genomic sequencing. At diagnosis, all patients had noninvasive or microinvasive (< 1 mm) disease; invasive disease eventually developed in 2 patients. Primary treatment was surgery for 19 patients (73%) and imiquimod topical therapy for 7 (27%). Seven patients had ≥ 2 surgeries as part of clinical course (1 patient had 5 vulvar resections). Samples had a median of 2 coding mutations in the genes analyzed (range, 0-29). The most common mutations were in PIK3CA (n = 9; 35%), ERBB2 (4 mutations and 3 copy number alterations; 27%), and TP53 (n = 7; 27%). MSIscore was available for 23 samples; all were microsatellite stable. After tumor genomic profiling, a patient who was initially treated with multiple resections and imiquimod was found to have a PIK3CA p.E542K mutation. She underwent PI3K-inhibitor treatment for 18 months before disease progression. CONCLUSION: EMPD of the vulva has a chronic and relapsing course, often requiring multiple surgical resections. Effective topical treatments are lacking. We identified targetable mutations (PIK3CA or ERBB2) in > 25% of a real-world clinical cohort. Additional prospective research implementing targetable therapies for EMPD treatment is warranted.

Fumarate hydratase FH c.1431_1433dupAAA (p.Lys477dup) variant is not associated with cancer including renal cell carcinoma.

Fumarate hydratase (FH) mutations underpin the autosomal recessive syndrome. FH deficiency and the autosomal dominant syndrome hereditary leiomyomatosis and renal cell carcinoma (HLRCC). The FH c.1431_1433dupAAA (p.Lys477dup) genomic alteration has been conclusively shown to contribute to FH deficiency when occurring with another FH germline alteration. However, a sufficiently large dataset has been lacking to conclusively determine its clinical significance to cancer predisposition in the heterozygous state. We reviewed a series of 7,571 patients with cancer who received germline results through MSK-IMPACT testing at the Memorial Sloan Kettering Cancer Center. The FH c.1431_1433dupAAA (p.Lys477dup) variant was detected in 24 individuals, none of whom was affected with renal cancer. Eleven of the 372 patients with renal cancer were identified to carried pathogenic FH variants associated with HLRCC. None of these 372 patients with renal cancer carried the FH c.1431_1433dupAAA variant. Our data indicate the FH c.1431_1433dupAAA is not associated with cancer including renal cell carcinoma.

Novel Germline Mutations in DNA Damage Repair in Patients with Malignant Pleural Mesotheliomas.

INTRODUCTION: Although next-generation sequencing (NGS) has brought insight into critical mutations or pathways (e.g., DNA damage sensing and repair) involved in the etiology of many cancers and has directed new screening, prevention, and therapeutic approaches for patients and families, it has only recently been used in malignant pleural mesotheliomas (MPMs). METHODS: We analyzed the blood samples from patients with MPM using the NGS platform MSK-IMPACT to explore cancer-predisposing genes. The loss-of-function variants or pathogenic entries were identified, and clinicopathologic information was collected. RESULTS: Of 84 patients with MPM, 12% (10 of 84) had pathogenic variants. Clinical characteristics were similar between cohorts, although patients with germline pathogenic variants were more likely to have more than two first-degree family members with cancer than those without germline mutations (40% versus 12%; Fisher's exact test, p < 0.05). Novel, deleterious variants in mesotheliomas included MutS homolog 3 (1% [one of 84]; 95% confidence interval [CI]: 0%-7%), breast cancer gene 1-associated ring domain 1 (1% [one of 84]; 95% CI: 0%-7%), and RecQ-like helicase 4 (2% [two of 84]; 95% CI: 0%-9%). Pathogenic variants previously reported on germline testing in patients with mesotheliomas were breast cancer gene 1-associated protein 1 (4% [three of 84]; 95% CI: 1%-10%), breast cancer gene 2 (1% [one of 84]; 95% CI: 0%-7%), and MRE11 homolog, double strand break repair nuclease (1% [one of 84]; 95% CI: 0%-7%). One patient (1% [one of 84]; 95% CI: 0%-7%) had a likely pathogenic alteration in SHQ1, H/ACA ribonucleoprotein assembly factor that has not been associated with a heritable susceptibility to cancer. CONCLUSIONS: Our study lends further support for the role of aberrations in DNA damage repair genes in the pathogenesis of MPMs and suggests that targeting the members of these pathways for screening and treatment warrants further study.

Genetic hallmarks of recurrent/metastatic adenoid cystic carcinoma.

BACKGROUNDAdenoid cystic carcinoma (ACC) is a rare malignancy arising in salivary glands and other sites, characterized by high rates of relapse and distant spread. Recurrent/metastatic (R/M) ACCs are generally incurable, due to a lack of active systemic therapies. To improve outcomes, deeper understanding of genetic alterations and vulnerabilities in R/M tumors is needed.METHODSAn integrated genomic analysis of 1,045 ACCs (177 primary, 868 R/M) was performed to identify alterations associated with advanced and metastatic tumors. Intratumoral genetic heterogeneity, germline mutations, and therapeutic actionability were assessed.RESULTSCompared with primary tumors, R/M tumors were enriched for alterations in key Notch (NOTCH1, 26.3% vs. 8.5%; NOTCH2, 4.6% vs. 2.3%; NOTCH3, 5.7% vs. 2.3%; NOTCH4, 3.6% vs. 0.6%) and chromatin-remodeling (KDM6A, 15.2% vs. 3.4%; KMT2C/MLL3, 14.3% vs. 4.0%; ARID1B, 14.1% vs. 4.0%) genes. TERT promoter mutations (13.1% of R/M cases) were mutually exclusive with both NOTCH1 mutations (q = 3.3 × 10-4) and MYB/MYBL1 fusions (q = 5.6 × 10-3), suggesting discrete, alternative mechanisms of tumorigenesis. This network of alterations defined 4 distinct ACC subgroups: MYB+NOTCH1+, MYB+/other, MYBWTNOTCH1+, and MYBWTTERT+. Despite low mutational load, we identified numerous samples with marked intratumoral genetic heterogeneity, including branching evolution across multiregion sequencing.CONCLUSIONThese observations collectively redefine the molecular underpinnings of ACC progression and identify further targets for precision therapies.FUNDINGAdenoid Cystic Carcinoma Research Foundation, Pershing Square Sohn Cancer Research grant, the PaineWebber Chair, Stand Up 2 Cancer, NIH R01 CA205426, the STARR Cancer Consortium, NCI R35 CA232097, the Frederick Adler Chair, Cycle for Survival, the Jayme Flowers Fund, The Sebastian Nativo Fund, NIH K08 DE024774 and R01 DE027738, and MSKCC through NIH/NCI Cancer Center Support Grant (P30 CA008748).

Prevalence and Preliminary Validation of Screening Criteria to Identify Carriers of Germline BAP1 Mutations.

INTRODUCTION: Inherited mutations are easily detected factors that influence the disease courses and optimal treatment strategies of some cancers. Germline mutations in BRCA1 associated protein 1 (BAP1) are associated with unique disease profiles in mesothelioma, atypical spitz nevi, and uveal melanoma, but the patient characteristics of an unselected population of BAP1 carriers identified by an ascertainment prevalence study are unknown. METHODS: We collected blood samples, cancer histories, and occupational exposures from 183 unselected patients with BAP1-related diseases. Clinical information for each patient was obtained from medical records. Germline DNA was extracted from blood samples and sequenced using a next-generation sequencing assay. We tested screening criteria developed to identify patients with a possible germline BAP1 mutation. RESULTS: Pathogenic or likely pathogenic germline BAP1 mutations were observed in 5 of 180 sequenced specimens and were exclusively found in patients identified by our screening criteria. Several patients with characteristics suspicious for a heritable deleterious mutation did not have a germline BAP1 mutation. The prevalence of pathogenic germline BAP1 mutations in patients with mesothelioma was 4.4% (95% confidence interval 1.1-11.1). CONCLUSIONS: Results from the first unselected prevalence ascertainment study of germline BAP1 alterations suggest that the frequency of this mutation is low among patients with mesothelioma. The proposed screening criteria successfully identified all patients with germline BAP1-mutant mesothelioma. These screening guidelines may assist physicians in selecting patients who would benefit from genetic testing. Future efforts should validate and refine these criteria and search for other germline mutations associated with mesothelioma and related diseases.