Cell-free DNA from nail clippings as source of normal control for genomic studies in hematologic malignancies.

Comprehensive genomic sequencing is becoming a critical component in the assessment of hematologic malignancies, with broad implications for patient management. In this context, unequivocally discriminating somatic from germline events is challenging but greatly facilitated by matched analysis of tumor:normal pairs. In contrast to solid tumors, conventional sources of normal control (peripheral blood, buccal swabs, saliva) could be highly involved by the neoplastic process, rendering them unsuitable. In this work we describe our real-world experience using cell free DNA (cfDNA) isolated from nail clippings as an alternate source of normal control, through the dedicated review of 2,610 tumor:nail pairs comprehensively sequenced by MSK-IMPACT-heme. Overall, we find nail cfDNA is a robust source of germline control for paired genomic studies. In a subset of patients, nail DNA may have tumor DNA contamination, reflecting unique attributes of the hematologic disease and transplant history. Contamination is generally low level, but significantly more common among patients with myeloid neoplasms (20.5%; 304/1482) compared to lymphoid diseases (5.4%; 61/1128) and particularly enriched in myeloproliferative neoplasms with marked myelofibrosis. When identified in patients with lymphoid and plasma-cell neoplasms, mutations commonly reflected a myeloid profile and correlated with a concurrent/evolving clonal myeloid neoplasm. For nails collected after allogeneic stem-cell transplantation, donor DNA was identified in 22% (11/50). In this cohort, an association with recent history of graft-vs-host disease was identified. These findings should be considered as a potential limitation for the use of nail as normal control but could also provide important diagnostic information regarding the disease process.

Effects of radioactive iodine on clonal hematopoiesis in patients with thyroid cancer: A prospective study.

OBJECTIVE: Exposure to therapeutic radioactive iodine (RAI) is associated with an increased relative risk of myeloid malignancies. Clonal hematopoiesis (CH) is a precursor state that can be detected in blood of healthy individuals decades before overt development of leukemia. We prospective studied the effects of RAI on CH. DESIGN: Prospective cohort study. PATIENTS AND MEASUREMENTS: We examined the effect of RAI on CH in 20 patients exposed to RAI for thyroid carcinoma and 20 age-matched unexposed controls. CH status was determined at baseline, 6, 12, 18 and 24 months. We also examined the effect of CH on structural progression of disease. RESULTS: No CH mutations were observed in the patient population that were not present at baseline. Using a variant allelic fraction (VAF) of 2% to define CH, 6/20 older patients (55-80 years old) had CH compared to 2/20 younger patients (20-40 years old) (p = 0.11). Six patients exposed to RAI had CH compared to two patients not exposed to RAI (30% vs. 10%, p = 0.11). There was no significant difference in CH VAF increase in patients treated with RAI compared to untreated age-matched controls (3.8% vs. 1.2%, p = 0.2). CH was significantly associated with somatic BRAFV600E mutations and with worse progression-free survival in the overall cohort as well as among BRAFV600E-mutant tumors. CONCLUSIONS: There was no increase in CH in patients treated with RAI over a 2-year follow-up period. Larger studies with longer follow-up periods are needed to investigate the association between RAI and clonal dynamics. The presence of CH is associated with worse structural progression in both BRAFV600E-mutant and wild-type thyroid cancers.

First-line pembrolizumab and trastuzumab in HER2-positive oesophageal, gastric, or gastro-oesophageal junction cancer: an open-label, single-arm, phase 2 trial.

BACKGROUND: Addition of trastuzumab to first-line chemotherapy improves overall survival in patients with HER2-positive metastatic gastric cancer. We assessed the safety and activity of pembrolizumab in combination with trastuzumab and chemotherapy in first-line HER2-positive metastatic oesophagogastric (gastric, oesophageal, or gastroesophageal junction) cancer. METHODS: This study was an investigator-initiated, open-label, non-randomised, single-arm, single centre, phase 2 trial in patients aged 18 years or older with HER2-positive metastatic oesophagogastric cancer. Eligible patients had measurable or evaluable non-measurable disease, Eastern Cooperative Oncology Group performance status of 0, 1, or 2, and left ventricular ejection fraction of at least 53%. Patients were eligible to receive an initial induction cycle of 200 mg flat dose of intravenous pembrolizumab and 8 mg/kg loading dose of intravenous trastuzumab. For subsequent cycles, patients received 130 mg/m2 of intravenous oxaliplatin or 80 mg/m2 of cisplatin on day 1, 850 mg/m2 of oral capecitabine twice a day for 2 weeks followed by 1 week off (or intravenous 5-fluorouracil, 800 mg/m2 per day on days 1-5), and a 200 mg flat dose of intravenous pembrolizumab, and 6 mg/kg of trastuzumab, administered on day 1 of each 3-week cycle. The primary endpoint was 6-month progression-free survival, defined as the proportion of patients alive and free of progression at 6 months, assessed in patients who received at least one dose of trastuzumab and pembrolizumab. The regimen would be considered worthy of further investigation if 26 or more of 37 patients were progression-free at 6 months. This trial is registered with ClinicalTrials.gov, NCT02954536, and is ongoing, but closed to enrolment. FINDINGS: Between Nov 11, 2016, and Jan 23, 2019, 37 patients were enrolled. At the time of data cutoff on Aug 6, 2019, median follow-up among survivors was 13·0 months (IQR 11·7-23·5). The primary endpoint was achieved; 26 (70%; 95% CI 54-83) of 37 patients were progression-free at 6 months. The most common treatment-related adverse event of any grade was neuropathy, which was reported in 36 (97%) of 37 patients. The most common grade 3 or 4 adverse events were lymphocytopenia (seven [19%] patients with grade 3 and two [5%] with grade 4), grade 3 decreased electrolytes (six [16%] patients), and grade 3 anaemia (four [11%] patients). Serious adverse events occurred in two patients patients (both grade 3 nephritis leading to treatment discontinuation). Four patients discontinued pembrolizumab because of immune-related adverse events. There were no treatment-related deaths. INTERPRETATION: Pembrolizumab can be safely combined with trastuzumab and chemotherapy and has promising activity in HER2-positive metastatic oesophagogastric cancer. A randomised phase 3 clinical trial assessing the efficacy and safety of pembrolizumab versus placebo in combination with trastuzumab and chemotherapy in first-line HER2-positive metastatic oesophagogastric cancer is underway. FUNDING: Merck & Co.

DNA methylation-based classification of sinonasal undifferentiated carcinoma.

Sinonasal undifferentiated carcinoma (SNUC) is an aggressive malignancy harboring IDH2 R172 mutations in >80% cases. We explored the potential of genome-wide DNA methylation profiling to elucidate tumor biology and improve the diagnosis of sinonasal undifferentiated carcinoma and its histologic mimics. Forty-two cases, including sinonasal undifferentiated, large cell neuroendocrine, small cell neuroendocrine, and SMARCB1-deficient carcinomas and olfactory neuroblastoma, were profiled by Illumina Infinium Methylation EPIC array interrogating >850,000 CpG sites. The data were analyzed using a custom bioinformatics pipeline. IDH2 mutation status was determined by the targeted exome sequencing (MSK-IMPACTTM) in most cases. H3K27 methylation level was assessed by the immunohistochemistry-based H-score. DNA methylation-based semi-supervised hierarchical clustering analysis segregated IDH2 mutants, mostly sinonasal undifferentiated (n = 10) and large cell neuroendocrine carcinomas (n = 4), from other sinonasal tumors, and formed a single cluster irrespective of the histologic type. t-distributed stochastic neighbor embedding dimensionality reduction analysis showed no overlap between IDH2 mutants, SMARCB1-deficient carcinoma and olfactory neuroblastoma. IDH2 mutants demonstrated a global methylation phenotype and an increase in repressive trimethylation of H3K27 in comparison to IDH2 wild-type tumors (p < 0.001). Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis showed no difference in pathway activation between IDH2-mutated sinonasal undifferentiated and large cell neuroendocrine carcinomas. In comparison to SMARCB1-deficient, IDH2-mutated carcinomas were associated with better disease-free survival (p = 0.034) and lower propensity for lung metastasis (p = 0.002). ARID1A mutations were common in small cell neuroendocrine carcinoma but not among IDH2 mutants (3/3 versus 0/18 and p < 0.001). IDH2 mutations in sinonasal carcinomas induce a hypermethylator phenotype and define a molecular subgroup of tumors arising in this location. IDH2-mutated sinonasal undifferentiated carcinoma and large cell neuroendocrine carcinoma likely represent a phenotypic spectrum of the same entity, which is distinct from small cell neuroendocrine and SMARCB1-deficient sinonasal carcinomas. DNA methylation-based analysis of the sinonasal tumors has potential to improve the diagnostic accuracy and classification of tumors arising in this location.

Frequent IDH2 R172 mutations in undifferentiated and poorly-differentiated sinonasal carcinomas.

Sinonasal undifferentiated carcinoma (SNUC) is a high-grade malignancy with limited treatment options and poor outcome. A morphological spectrum of 47 sinonasal tumours including 17 (36.2%) SNUCs was analysed at genomic level. Thirty carcinomas (cohort 1) were subjected to a hybridization exon-capture next-generation sequencing assay (MSK-IMPACTTM ) to interrogate somatic variants in 279 or 410 cancer-related genes. Seventeen sinonasal tumours (cohort 2) were examined only for the presence of IDH1/2 exon 4 mutations by Sanger sequencing. IDH2 R172 single nucleotide variants were overall detected in 14 (82.4%) SNUCs, in two (20%) poorly-differentiated carcinomas with glandular/acinar differentiation, and in one of two high-grade neuroendocrine carcinomas, large cell type (HGNECs). No IDH2 mutation was detected in any of five olfactory neuroblastomas or in any of five SMARCB1-deficient carcinomas. Among 12 IDH2-mutated cases in cohort 1, five (41.7%) harboured co-existing TP53 mutations, four (33.3%) CDKN2A/2B loss-of-function alterations, four (33.3%) MYC amplification, and three (25%) had concurrent SETD2 mutations. AKT1 E17K and KIT D816V hotspot variants were each detected in one IDH2-mutated SNUC. The vast majority of SNUCs and variable proportions of other poorly-differentiated sinonasal carcinomas may be amenable to IDH2-targeted therapy. Copyright © 2017 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

Mammary analog secretory carcinoma of the thyroid gland: A primary thyroid adenocarcinoma harboring ETV6-NTRK3 fusion.

ETV6-NTRK3 fusion was identified in several cancers including the recently described mammary analog secretory carcinoma (MASC) of the salivary glands and a minority of papillary thyroid carcinomas. We describe three cases of primary MASC of the thyroid gland and provide a detailed clinical and pathological characterization of the tumor morphology, immunoprofile, and genetic background. Immunohistochemistry for PAX8, TTF-1, thyroglobulin, mammaglobin, GCDFP-15, S-100 protein, and p63 was used to define the tumor immunophenotype. Fluorescence in situ hybridization for ETV6 rearrangement was performed in three, and the next-generation sequencing assay MSK-IMPACT™ (Memorial Sloan Kettering-Integrated Mutation Profiling of Actionable Cancer Targets) was performed in two cases. Primary MASC of the thyroid occurred in two women and one man, age 47-72 years. All patients presented with high T stage, infiltrative, locally aggressive tumors with extrathyroidal extension. Two cases were associated with well-differentiated papillary thyroid carcinoma. Histologically, they appeared as low-grade tumors, resembling MASC of the salivary glands and labeled positive for mammaglobin, GCDFP-15, S-100 protein, p63, weakly positive for PAX8, and negative for TTF-1 and thyroglobulin. Fluorescence in situ hybridization revealed ETV6 rearrangement in all cases. In two tested cases MSK-IMPACT™ confirmed the presence of ETV6-NTRK3 gene fusion. Two patients had at least two local recurrences, one was alive with disease, and one was alive and free of disease after 14 and 17 years, respectively. The third patient was alive and free of disease after 2 years. MASC of the thyroid is histologically, immunophenotypically, and genetically similar to its salivary gland counterpart. Thyroid MASC can be associated with a well-differentiated papillary thyroid carcinoma component, supporting follicular cell origin. Clinically, these carcinomas may show frequent recurrences but are associated with long-term survival. Patients with MASC of the thyroid may potentially benefit from Trk molecular-targeted therapy.

Respiratory Syncytial Virus Utilizes a tRNA Fragment to Suppress Antiviral Responses Through a Novel Targeting Mechanism.

Target identification is highly instructive in defining the biological roles of microRNAs. However, little is known about other small noncoding RNAs; for example, tRNA-derived RNA Fragments (tRFs). Some tRFs exhibit a gene-silencing mechanism distinctly different from that of typical microRNAs. We recently demonstrated that a respiratory syncytial virus (RSV)-induced tRF, called tRF5-GluCTC, promotes RSV replication. RSV is the single most important cause of lower respiratory tract infection in children. By using biochemical screening and bioinformatics analyses, we have identified apolipoprotein E receptor 2 (APOER2) as a target of tRF5-GluCTC. The 3'-portion of tRF5-GluCTC recognizes a target site in the 3'-untranslated region of APOER2 and suppresses its expression. We have also discovered that APOER2 is an anti-RSV protein whose suppression by tRF5-GluCTC promotes RSV replication. Our report represents the first identification of a natural target of a tRF and illustrates how a virus utilizes a host tRF to control a host gene to favor its replication.

High-risk and silent clonal hematopoietic genotypes in patients with nonhematologic cancer.

ABSTRACT: Clonal hematopoiesis (CH) identified by somatic gene variants with variant allele fraction (VAF) ≥ 2% is associated with an increased risk of hematologic malignancy. However, CH defined by a broader set of genotypes and lower VAFs is ubiquitous in older individuals. To improve our understanding of the relationship between CH genotype and risk of hematologic malignancy, we analyzed data from 42 714 patients who underwent blood sequencing as a normal comparator for nonhematologic tumor testing using a large cancer-related gene panel. We cataloged hematologic malignancies in this cohort using natural language processing and manual curation of medical records. We found that some CH genotypes including JAK2, RUNX1, and XPO1 variants were associated with high hematologic malignancy risk. Chronic disease was predicted better than acute disease suggesting the influence of length bias. To better understand the implications of hematopoietic clonality independent of mutational function, we evaluated a set of silent synonymous and noncoding mutations. We found that silent CH, particularly when multiple variants were present or VAF was high, was associated with increased risk of hematologic malignancy. We tracked expansion of CH mutations in 26 hematologic malignancies sequenced with the same platform. JAK2 and TP53 VAF consistently expanded at disease onset, whereas DNMT3A and silent CH VAFs mostly decreased. These data inform the clinical and biological interpretation of CH in the context of nonhematologic cancer.

Associations Between Cancer Predisposition Mutations and Clonal Hematopoiesis in Patients With Solid Tumors.

PURPOSE: Clonal hematopoiesis (CH), the expansion of clones in the hematopoietic system, has been linked to different internal and external features such as aging, genetic ancestry, smoking, and oncologic treatment. However, the interplay between mutations in known cancer predisposition genes and CH has not been thoroughly examined in patients with solid tumors. METHODS: We used prospective tumor-blood paired sequencing data from 46,906 patients who underwent Memorial Sloan Kettering-Integrated Mutation Profiling of Actionable Cancer Targets (MSK-IMPACT) testing to interrogate the associations between CH and rare pathogenic or likely pathogenic (P/LP) germline variants. RESULTS: We observed an enrichment of CH-positive patients among those carrying P/LP germline mutations and identified a significant association between P/LP germline variants in ATM and CH. Germline and CH comutation patterns in ATM, TP53, and CHEK2 suggested biallelic inactivation as a potential mediator of clonal expansion. Moreover, we observed that CH-PPM1D mutations, similar to somatic tumor-associated PPM1D mutations, were depleted in patients with P/LP germline mutations in the DNA damage response (DDR) genes ATM, CHEK2, and TP53. Patients with solid tumors and harboring P/LP germline mutations, CH mutations, and mosaicism chromosomal alterations might be at an increased risk of developing secondary leukemia while germline variants in TP53 were identified as an independent risk factor (hazard ratio, 36; P < .001) for secondary leukemias. CONCLUSION: Our results suggest a close relationship between inherited variants and CH mutations within the DDR genes in patients with solid tumors. Associations identified in this study might translate into enhanced clinical surveillance for CH and associated comorbidities in patients with cancer harboring these germline mutations.

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.

Bioinformatically Expanded Next-Generation Sequencing Analysis Optimizes Identification of Therapeutically Relevant MET Copy Number Alterations in >50,000 Tumors.

PURPOSE: Clinical relevance thresholds and laboratory methods are poorly defined for MET amplification, a targetable biomarker across malignancies. EXPERIMENTAL DESIGN: The utility of next-generation sequencing (NGS) in assessing MET copy number alterations was determined in >50,000 solid tumors. Using fluorescence in situ hybridization as reference, we validated and optimized NGS analysis. RESULTS: Incorporating read-depth and focality analyses achieved 91% concordance, 97% sensitivity, and 89% specificity. Tumor heterogeneity, neoplastic cell proportions, and genomic focality affected MET amplification assessment. NGS methodology showed superiority in capturing overall amplification status in heterogeneous tumors and defining amplification focality among other genomic alterations. MET copy gains and amplifications were found in 408 samples across 23 malignancies. Total MET copy number inversely correlated with amplified segment size. High-level/focal amplification was enriched in certain genomic subgroups and associated with targeted therapy response. CONCLUSIONS: Leveraging our integrated bioinformatic approach, targeted therapy benefit was observed across diverse MET amplification contexts.

Cancer-Causative Mutations Occurring in Early Embryogenesis.

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

Bone Marrow Surveillance of Pediatric Cancer Survivors Identifies Clones that Predict Therapy-Related Leukemia.

PURPOSE: Therapy-related myelodysplastic syndrome and acute leukemias (t-MDS/AL) are a major cause of nonrelapse mortality among pediatric cancer survivors. Although the presence of clonal hematopoiesis (CH) in adult patients at cancer diagnosis has been implicated in t-MDS/AL, there is limited published literature describing t-MDS/AL development in children. EXPERIMENTAL DESIGN: We performed molecular characterization of 199 serial bone marrow samples from 52 patients treated for high-risk neuroblastoma, including 17 with t-MDS/AL (transformation), 14 with transient cytogenetic abnormalities (transient), and 21 without t-MDS/AL or cytogenetic alterations (neuroblastoma-treated control). We also evaluated for CH in a cohort of 657 pediatric patients with solid tumor. RESULTS: We detected at least one disease-defining alteration in all cases at t-MDS/AL diagnosis, most commonly TP53 mutations and KMT2A rearrangements, including involving two novel partner genes (PRDM10 and DDX6). Backtracking studies identified at least one t-MDS/AL-associated mutation in 13 of 17 patients at a median of 15 months before t-MDS/AL diagnosis (range, 1.3-32.4). In comparison, acquired mutations were infrequent in the transient and control groups (4/14 and 1/21, respectively). The relative risk for development of t-MDS/AL in the presence of an oncogenic mutation was 8.8 for transformation patients compared with transient. Unlike CH in adult oncology patients, TP53 mutations were only detectable after initiation of cancer therapy. Last, only 1% of pediatric patients with solid tumor evaluated had CH involving myeloid genes. CONCLUSIONS: These findings demonstrate the clinical relevance of identifying molecular abnormalities in predicting development of t-MDS/AL and should guide the formation of intervention protocols to prevent this complication in high-risk pediatric patients.

Genetic basis of SMARCB1 protein loss in 22 sinonasal carcinomas.

SMARCB1-deficient sinonasal carcinoma (SNC) is an aggressive malignancy characterized by INI1 loss mostly owing to homozygous SMARCB1 deletion. With the exception of a few reported cases, these tumors have not been thoroughly studied by massive parallel sequencing (MPS). A retrospective cohort of 22 SMARCB1-deficient SNCs were studied by light microscopy, immunohistochemistry, fluorescence in situ hybridization (n = 9), targeted exome MPS (n = 12), and Fraction and Allele-Specific Copy Number Estimates from Tumor Sequencing (FACETS) (n = 10), a bioinformatics pipeline for copy number/zygosity assessment. SMARCB1-deficient SNC was found in 13 (59%) men and 9 (41%) women. Most common growth patterns were the basaloid pattern (59%), occurring mostly in men (77%), and plasmacytoid/eosinophilic/rhabdoid pattern (23%), arising mostly in women (80%). The former group was significantly younger (median age = 46 years, range = 24-54, vs 79 years, range = 66-95, p < 0.0001). Clear cell, pseudoglandular, glandular, spindle cell, and sarcomatoid features were variably present. SMARCB1-deficient SNC expressed cytokeratin (100%), p63 (72%), neuroendocrine markers (52%), CDX-2 (44%), S-100 (25%), CEA (4/4 cases), Hepatocyte (2/2 cases), and aberrant nuclear ß-catenin (1/1 case). SMARCB1 showed homozygous deletion (68%), hemizygous deletion (16%), or truncating mutations associated with copy neutral loss of heterozygosity (11%). Coexisting genetic alterations were 22q loss including loss of NF2 and CHEK2 (50%), chromosome 7 gain (25%), and TP53 V157F, CDKN2A W110∗, and CTNNB1 S45F mutations. At 2 years and 5 years, the disease-specific survival and disease-free survival were 70% and 35% and 13% and 0%, respectively. SMARCB1-deficient SNCs are phenotypically and genetically diverse, and these distinctions warrant further investigation for their biological and clinical significance.

The Clinical Management of Clonal Hematopoiesis: Creation of a Clonal Hematopoiesis Clinic.

The acquisition of mutations in hematologic stem cells (clonal hematopoiesis) is common with normal aging and can be identified as an incidental finding through clinical genetic testing. Clonal hematopoiesis is associated with a heightened risk of developing hematologic neoplasms (especially myeloid) and accelerated atherosclerotic cardiovascular disease. This article discusses a multidisciplinary clinical approach to the management of patients with clonal hematopoiesis. Key areas of research needed to establish evidence-based clinical care guidelines and intervention strategies for individuals with clonal hematopoiesis are discussed.

SMARCA4-Deficient Thoracic Sarcomatoid Tumors Represent Primarily Smoking-Related Undifferentiated Carcinomas Rather Than Primary Thoracic Sarcomas.

INTRODUCTION: Highly aggressive thoracic neoplasms characterized by SMARCA4 (BRG1) deficiency and undifferentiated round cell or rhabdoid morphology have been recently described and proposed to represent thoracic sarcomas. However, it remains unclear whether such tumors may instead represent sarcomatoid carcinomas, and how their clinicopathologic characteristics compare with those of nonsarcomatoid SMARCA4-deficient non-small cell lung carcinomas (SD-NSCC). METHODS: We identified 22 SMARCA4-deficient thoracic sarcomatoid tumors (SD-TSTs) with round cell and/or rhabdoid morphology and 45 SD-NSCCs, and comprehensively analyzed their clinicopathologic, immunohistochemical, and genomic characteristics using 341-468 gene next-generation sequencing and other molecular platforms. RESULTS: The relationship of SD-TSTs with NSCC was supported by (1) the presence of NSCC components juxtaposed with sarcomatoid areas in five cases, (2) focal expression of NSCC lineage markers TTF1 or p40 in four additional cases, (3) smoking history in all except one patient (mean = 51 pack-years), accompanied by genomic smoking signature, and (4) high tumor mutation burden (mean = 14.2 mutations per megabase) and mutations characteristic of NSCC in a subset. Compared with SD-NSCCs, SD-TSTs exhibited considerably larger primary tumor size (p < 0.0001), worse survival (p = 0.004), and more frequent presentation at younger age (30-50 years) despite heavier smoking history. Distinctive pathologic features of SD-TSTs included consistent lack of adhesion molecule claudin-4, SMARCA2 (BRM) codeficiency, and frequent expression of stem cell markers. CONCLUSIONS: SD-TSTs represent primarily smoking-associated undifferentiated/de-differentiated carcinomas rather than primary thoracic sarcomas. Despite their histogenetic relationship with NSCC, these tumors have unique clinicopathologic characteristics, supporting their recognition as a distinct entity. Further studies are warranted to determine therapeutic approaches to this novel class of exceptionally aggressive thoracic tumors.

Cancer therapy shapes the fitness landscape of clonal hematopoiesis.

Acquired mutations are pervasive across normal tissues. However, understanding of the processes that drive transformation of certain clones to cancer is limited. Here we study this phenomenon in the context of clonal hematopoiesis (CH) and the development of therapy-related myeloid neoplasms (tMNs). We find that mutations are selected differentially based on exposures. Mutations in ASXL1 are enriched in current or former smokers, whereas cancer therapy with radiation, platinum and topoisomerase II inhibitors preferentially selects for mutations in DNA damage response genes (TP53, PPM1D, CHEK2). Sequential sampling provides definitive evidence that DNA damage response clones outcompete other clones when exposed to certain therapies. Among cases in which CH was previously detected, the CH mutation was present at tMN diagnosis. We identify the molecular characteristics of CH that increase risk of tMN. The increasing implementation of clinical sequencing at diagnosis provides an opportunity to identify patients at risk of tMN for prevention strategies.

Clinicopathologic and genetic characterization of nonacute NPM1-mutated myeloid neoplasms.

NPM1-mutated myeloid neoplasms (NPM1 + MNs) with <20% blood or bone marrow blasts are rare and have been previously shown in limited case series to exhibit an aggressive clinical course. We assembled the largest cohort of NPM1 + MN cases to date (n = 45) and compared it with NPM1 - MN (n = 95) and NPM1 + de novo acute myeloid leukemia (AML; n = 119) patients. Compared with NPM1 - MN, NPM1 + MN were associated with younger age (P = .007), a normal karyotype (P < .0001), more frequent mutations involving DNMT3A (P = .01) and PTPN11 (P = .03), and fewer involving ASXL1 (P = .003), RUNX1 (P = .0004), and TP53 (P = .02). Mutations involving IDH1 or IDH2 (IDH1/2) (P = .007) and FLT3 (internal tandem duplication, P < .0001; noninternal tandem duplication, P = .01) were less frequent in NPM1 + MN than in NPM1 + AML. In multivariable analyses performed in patients with myelodysplastic syndrome only, total mutation count (hazard ratio [HR], 1.3; P = .05), NPM1 mutation (HR, 3.6; P = .02), TP53 mutation (HR, 5.2; P = .01), and higher International Prognostic Scoring System-R score (HR, 1.7; P = .0003) were independently associated with shorter overall survival, whereas stem cell transplant conferred a favorable effect (HR, 0.1; P < .0001). These data suggest that NPM1 + MN are biologically distinct from NPM1 - MN. Similar to NPM1 + AML, patients with NPM1-mutated myelodysplastic syndrome may benefit from more intensive therapeutic regimens.

Prevalence of Clonal Hematopoiesis Mutations in Tumor-Only Clinical Genomic Profiling of Solid Tumors.

Importance: Although clonal hematopoiesis (CH) is well described in aging healthy populations, few studies have addressed the practical clinical implications of these alterations in solid-tumor sequencing. Objective: To identify and quantify CH-related mutations in patients with solid tumors using matched tumor-blood sequencing, and to establish the proportion that would be misattributed to the tumor based on tumor-only sequencing (unmatched analysis). Design, Setting, and Participants: Retrospective analysis of samples from 17 469 patients with solid cancers who underwent prospective clinical sequencing of DNA isolated from tumor tissue and matched peripheral blood using the MSK-IMPACT assay between January 2014 and August 2017. Main Outcomes and Measures: We identified the presence of CH-related mutations in each patient's blood leukocytes and quantified the fraction of DNA molecules harboring the mutation in the corresponding matched tumor sample. Results: The mean age of the 17 469 patients with cancer at sample collection was 59.2 years (range, 0.3-98.9 years); 53.6% were female. We identified 7608 CH-associated mutations in the blood of 4628 (26.5%) patients. A total of 1075 (14.1%) CH-associated mutations were also detectable in the matched tumor above established thresholds for calling somatic mutations. Overall, 912 (5.2%) patients would have had at least 1 CH-associated mutation erroneously called as tumor derived in the absence of matched blood sequencing. A total of 1061 (98.7%) of these mutations were absent from population scale databases of germline polymorphisms and therefore would have been challenging to filter informatically. Annotating variants with OncoKB classified 534 (49.7%) as oncogenic or likely oncogenic. Conclusions and Relevance: This study demonstrates how CH-derived mutations could lead to erroneous reporting and treatment recommendations when tumor-only sequencing is used.

Chromosome 20q Amplification Defines a Subtype of Microsatellite Stable, Left-Sided Colon Cancers with Wild-type RAS/RAF and Better Overall Survival.

Here, comprehensive analysis was performed on the molecular and clinical features of colorectal carcinoma harboring chromosome 20q amplification. Tumor and normal DNA from patients with advanced colorectal carcinoma underwent next-generation sequencing via MSK-IMPACT, and a subset of case samples was subjected to high-resolution microarray (Oncoscan). Relationships between genomic copy number and transcript expression were assessed with The Cancer Genome Atlas (TCGA) colorectal carcinoma data. Of the colorectal carcinoma patients sequenced (n = 401) with MSK-IMPACT, 148 (37%) had 20q gain, and 30 (7%) had 20q amplification. In both the MSK-IMPACT and TCGA datasets, BCL2L1 was the most frequently amplified 20q oncogene. However, SRC was the only recognized 20q oncogene with a significant inverse relationship between mRNA upregulation and RAS/RAF mutation (OR, -0.4 ± 0.2, P = 0.02). In comparison with 20q diploid colorectal carcinoma, 20q gain/amplification was associated with wild-type KRAS (P < 0.001) and BRAF (P = 0.01), microsatellite stability (P < 0.001), distal primary tumors (P < 0.001), and mutant TP53 (P < 0.001), but not stage. On multivariate analysis, longer overall survival from the date of metastasis was observed with chromosome 20q gain (P = 0.02) or amplification (P = 0.04) compared with diploid 20q.Implications: 20q amplification defines a subset of colorectal cancer patients with better overall survival from the date of metastasis, and further studies are warranted to assess whether the inhibition of 20q oncogenes, such as SRC, may benefit this subset of patients. Mol Cancer Res; 15(6); 708-13. ©2017 AACR.