Validation of the VirCapSeq-VERT system for differential diagnosis, detection, and surveillance of viral infections.

IMPORTANCE: Broad range assay for accurate and sensitive diagnostics.

Clinical Utility and Reimbursement of Next-Generation Sequencing-Based Testing for Myeloid Malignancies.

Next-generation sequencing is becoming increasingly important for the diagnosis, risk stratification, and management of patients with established or suspected myeloid malignancies. These tests are being incorporated into clinical practice guidelines and many genetic alterations now constitute disease classification criteria. However, the reimbursement for these tests is uncertain. This study analyzed the clinical impact, ordering practices, prior authorization, and reimbursement outcomes of 505 samples from 477 patients sequenced with a 50-gene myeloid next-generation sequencing panel or a 15-gene myeloproliferative neoplasm subpanel. Overall, 98% (496 of 505) of tests provided clinically useful data. Eighty-nine percent of test results, including negative findings, informed or clarified potential diagnoses, 94% of results informed potential prognoses, and 19% of tests identified a potential therapeutic target. Sequencing results helped risk-stratify patients whose bone marrow biopsy specimens were inconclusive for dysplasia, monitor genetic evolution associated with disease progression, and delineate patients with mutation-defined diagnoses. Despite the clinical value, prior authorization from commercial payors or managed government payors was approved for less than half (45%) of requests. Only 51% of all cases were reimbursed, with lack of medical necessity frequently cited as a reason for denial. This study demonstrates the existence of a substantial gap between clinical utility and payor policies on test reimbursement.

Current clinical practices and challenges in molecular testing: a GOAL Consortium Hematopathology Working Group report.

While molecular testing of hematologic malignancies is now standard of care, there is variability in practice and testing capabilities between different academic laboratories, with common questions arising on how to best meet clinical expectations. A survey was sent to hematopathology subgroup members of the Genomics Organization for Academic Laboratories consortium to assess current and future practice and potentially establish a reference for peer institutions. Responses were received from 18 academic tertiary-care laboratories regarding next-generation sequencing (NGS) panel design, sequencing protocols and metrics, assay characteristics, laboratory operations, case reimbursement, and development plans. Differences in NGS panel size, use, and gene content were reported. Gene content for myeloid processes was reported to be generally excellent, while genes for lymphoid processes were less well covered. The turnaround time (TAT) for acute cases, including acute myeloid leukemia, was reported to range from 2 to 7 calendar days to 15 to 21 calendar days, with different approaches to achieving rapid TAT described. To help guide NGS panel design and standardize gene content, consensus gene lists based on current and future NGS panels in development were generated. Most survey respondents expected molecular testing at academic laboratories to continue to be viable in the future, with rapid TAT for acute cases likely to remain an important factor. Molecular testing reimbursement was reported to be a major concern. The results of this survey and subsequent discussions improve the shared understanding of differences in testing practices for hematologic malignancies between institutions and will help provide a more consistent level of patient care.

Papillary Intralymphatic Angioendothelioma in a Child With PIK3CA-Related Overgrowth Spectrum: Implication of PI3K Pathway in the Vascular Tumorigenesis.

Papillary intralymphatic angioendothelioma (PILA) is an extremely rare vascular tumor and its pathogenesis is unknown. Phosphatidylinositol-4,5-bisphosphate 3-kinase catalytic subunit alpha (PIK3CA)-related overgrowth spectrum (PROS) is a heterogeneous group of disorders caused by mosaicism for activating mutations of PIK3CA and characterized by asymmetric overgrowth, skeletal anomalies, skin lesions, and vascular malformations. An association between PILA and PROS has not been known. We report a case of PILA involving the spleen of a young girl with the clinical and molecular diagnosis of PROS. Sequencing of the patient's germ-line DNA detected a pathogenic PIK3CA variant c.1357G>A in 10.6% of alleles. Splenectomy revealed a 4-cm tumor composed of ectatic lymphatics with intraluminal papillary projections, consistent with PILA. The tumor cells showed immunohistochemical expression of CD31, CD34, ERG, FLI-1, PROX1, and caldesmon, while D2-40 was negative. The latter may suggest that the tumor derived from an endothelial precursor arrested in the final steps of lymphothelial differentiation, in keeping with the known role of the PIK3CA-governed molecular pathway in the progression of vascular progenitors to mature endothelial cells. The data implicates PIK3CA in the pathogenesis of PILA and broadens the spectrum of phenotypic expressions of PROS.

Pineal region ganglioglioma: A neoplasm with a bimodal age distribution.

Background: Gangliogliomas arise very rarely in the pineal region, where their natural histories and pathologic features are poorly understood. Case Description: In this report, we describe a 36-year-old woman who presented with a seizure followed by worsening headache, dizziness, confusion, and intermittent left facial numbness over the next few weeks. A head CT scan showed a partially calcified pineal region mass with hydrocephalus. After an endoscopic third ventriculostomy, the patient underwent a resection of the tumor that contained dysplastic ganglion cells and piloid glial cells. Molecular profiling of this CNS WHO Grade 1 ganglioglioma revealed polysomies of chromosomes 7 and 9, and a BUB1 variant of uncertain significance, without known MAP kinase pathway alterations. From a review of the literature, we found two distinct age distributions for pineal ganglioglioma, with modes at 1 and 36 years of age. Conclusion: Although very rare, this tumor should be considered in the differential diagnosis of pineal region tumors in children and young adults.

Pan-tumor screening for NTRK gene fusions using pan-TRK immunohistochemistry and RNA NGS fusion panel testing.

Targetable NTRK gene fusions can be detected across tumor types using methodologies such as pan-TRK IHC, DNA or RNA NGS testing, or FISH. Challenges for implementation of clinical testing for NTRK fusions may arise due to the range in NTRK fusion prevalence across tumors, endogenous levels of TRK expression in tissues, and the large number of potential fusion partners. In this study, we examined our experience evaluating driver mutation negative lung, urothelial or cholangiocarcinoma cases, in addition to cases with positive, equivocal, or weak staining by pan-TRK IHC for NTRK fusions. 63/127 (49.6%) of these cases were positive for pan-TRK IHC, of which 71.4% showed weak or focal staining, potentially due to physiologic or non-specific TRK expression. Of these 127 cases, 4 harbored a NTRK fusion (1 fusion was seen in two separate samples from the same patient) as confirmed by RNA fusion panel testing. Pan-TRK IHC was positive in 1 case with TPM3-NTRK1 fusion, equivocal in 1 case with GOLGA4-NTRK3 fusion, and negative in 2 samples with ADAM19-NTRK3 fusion. Our findings show that we were able to successfully identify NTRK fusions that resulted in targeted therapy. However, our results suggest limited sensitivity of pan-TRK IHC for NTRK3 fusions, and that the reduced specificity for pan-TRK IHC in tumors with physiologic or non-specific TRK expression, results in false positive samples that require confirmatory testing by RNA based NGS.

Phenogenomic heterogeneity of post-transplant plasmablastic lymphomas.

Plasmablastic lymphoma (PBL) is a rare and clinically aggressive neoplasm that typically occurs in immunocompromised individuals, including those with HIV infection and solid organ allograft recipients. Most prior studies have focused on delineating the clinicopathologic features and genetic attributes of HIV-related PBLs, where MYC deregulation and EBV infection, and more recently, mutations in JAK/STAT, MAP kinase, and NOTCH pathway genes have been implicated in disease pathogenesis. The phenotypic spectrum of post-transplant (PT)-PBLs is not well characterized and data on underlying genetic alterations are limited. Hence, we performed comprehensive histopathologic and immunophenotypic evaluation and targeted sequencing of 18 samples from 11 patients (8 males, 3 females, age range 12-76 years) with PT-PBL; 8 de novo and 3 preceded by other types of PTLDs. PT-PBLs displayed morphologic and immunophenotypic heterogeneity and some features overlapped those of plasmablastic myeloma. Six (55%) cases were EBV+ and 5 (45%) showed MYC rearrangement by fluorescence in situ hybridization. Recurrent mutations in epigenetic regulators (KMT2/MLL family, TET2) and DNA damage repair and response (TP53, mismatch repair genes, FANCA, ATRX), MAP kinase (KRAS, NRAS, HRAS, BRAF), JAK/STAT (STAT3, STAT6, SOCS1), NOTCH (NOTCH1, NOTCH3, SPEN), and immune surveillance (FAS, CD58) pathway genes were observed, with EBV+ and EBV- cases exhibiting similarities and differences in their mutational profiles. Clinical outcomes also varied, with survival ranging from 0-15.9 years postdiagnosis. Besides uncovering the biological heterogeneity of PT-PBL, our study highlights similarities and distinctions between PT-PBLs and PBLs occurring in other settings and reveals potentially targetable oncogenic pathways in disease subsets.

ALK Gene Rearrangements in Lung Adenocarcinomas: Concordance of Immunohistochemistry, Fluorescence In Situ Hybridization, RNA In Situ Hybridization, and RNA Next-Generation Sequencing Testing.

INTRODUCTION: The 2018 updated molecular testing guidelines for patients with advanced lung cancer incorporated ALK immunohistochemistry (IHC) analysis as an equivalent to fluorescence in situ hybridization (FISH) method recommended in 2013. Nevertheless, no specific recommendation for alternative methods was proposed owing to insufficient data. The aim of this study was to compare the results of ALK IHC, FISH, RNA next-generation sequencing (NGS), and RNA in situ hybridization (ISH) with available clinical data. METHODS: A search for lung carcinomas with ALK testing by greater than or equal to one modality (i.e., ALK IHC, FISH, NGS) was performed; a subset underwent RNA ISH. When available, clinical data were recorded. RESULTS: The results were concordant among all performed testing modalities in 86 of 90 cases (95.6%). Of the four discordant cases, two were ALK positive by FISH but negative by IHC, RNA NGS, and RNA ISH. The remaining two cases failed RNA NGS testing, one was IHC negative, FISH positive, RNA ISH negative and the second was IHC positive, FISH positive, RNA ISH equivocal. RNA NGS identified one rare and one novel ALK fusion. Sufficient therapy data were available in 10 cases treated with tyrosine kinase inhibitors; three had disease progression, including one with discordant results (FISH positive, RNA NGS negative, IHC negative, RNA ISH negative) and two with concordant ALK positivity among all modalities. CONCLUSIONS: Our results reveal high concordance among IHC, RNA NGS, and RNA ISH. In cases of discordance with available RNA NGS, FISH result was positive whereas IHC and ISH results were negative. On the basis of our data, multimodality testing is recommended to identify discrepant results and patients (un)likely to respond to tyrosine kinase inhibitors.

Rosette-Forming Glioneuronal Tumor in the Pineal Region: A Series of 6 Cases and Literature Review.

Resected lesions from the pineal region are rare specimens encountered by surgical pathologists, and their heterogeneity can pose significant diagnostic challenges. Here, we reviewed 221 pineal region lesions resected at New York-Presbyterian Hospital/Columbia University Irving Medical Center from 1994 to 2019 and found the most common entities to be pineal parenchymal tumors (25.3%), glial neoplasms (18.6%), and germ cell tumors (17.6%) in this predominantly adult cohort of patients. Six cases of a rare midline entity usually found exclusively in the fourth ventricle, the rosette-forming glioneuronal tumor, were identified. These tumors exhibit biphasic morphology, with a component resembling pilocytic astrocytoma admixed with variable numbers of small cells forming compact rosettes and perivascular pseudorosettes. Targeted sequencing revealed a 100% co-occurrence of novel and previously described genetic alterations in the mitogen-activated protein kinase (MAPK) and phosphoinositide 3-kinase (PI3K) signaling pathways, suggesting a synergistic role in tumor formation. The most common recurrent mutation, PIK3CA H1047R, was identified in tumor cells forming rosettes and perivascular pseudorosettes. A review of the literature revealed 16 additional cases of rosette-forming glioneuronal tumors in the pineal region. Although rare, this distinctive low-grade tumor warrants consideration in the differential diagnosis of pineal region lesions.

Immunophenotypic Spectrum and Genomic Landscape of Refractory Celiac Disease Type II.

Refractory celiac disease type II (RCD II), also referred to as "cryptic" enteropathy-associated T-cell lymphoma (EATL) or "intraepithelial T-cell lymphoma," is a rare clonal lymphoproliferative disorder that arises from innate intraepithelial lymphocytes. RCD II has a poor prognosis and frequently evolves to EATL. The pathogenesis of RCD II is not well understood and data regarding the immunophenotypic spectrum of this disease and underlying genetic alterations are limited. To gain further biological insights, we performed comprehensive immunophenotypic, targeted next-generation sequencing, and chromosome microarray analyses of 11 RCD II cases: CD4-/CD8- (n=6), CD8+ (n=4), and CD4+ (n=1), and 2 of 3 ensuing EATLs. Genetic alterations were identified in 9/11 (82%) of the RCD II cases. All 9 displayed mutations in members of the JAK-STAT signaling pathway, including frequent, recurrent STAT3 (7/9, 78%) and JAK1 (4/9, 44%) mutations, and 9/10 evaluable cases expressed phospho-STAT3. The mutated cases also harbored recurrent alterations in epigenetic regulators (TET2, n=5 and KMT2D, n=5), nuclear factor-κB (TNFAIP3, n=4), DNA damage repair (POT1, n=3), and immune evasion (CD58, n=2) pathway genes. The CD4-/CD8- and other immunophenotypic subtypes of RCD II exhibited similar molecular features. Longitudinal genetic analyses of 4 RCD II cases revealed stable mutation profiles, however, additional mutations were detected in the EATLs, which occurred at extraintestinal sites and were clonally related to antecedent RCD II. Chromosome microarray analysis demonstrated copy number changes in 3/6 RCD II cases, and 1 transformed EATL with sufficient neoplastic burden for informative analysis. Our findings provide novel information about the immunophenotypic and genomic characteristics of RCD II, elucidate early genetic events in EATL pathogenesis, and reveal potential therapeutic targets.

Compound heterozygous inheritance of two novel COQ2 variants results in familial coenzyme Q deficiency.

BACKGROUND: Primary coenzyme Q10 deficiency is a rare disease that results in diverse and variable clinical manifestations. Nephropathy, myopathy and neurologic involvement are commonly associated, however retinopathy has also been observed with certain pathogenic variants of genes in the coenzyme Q biosynthesis pathway. In this report, we describe a novel presentation of the disease that includes nephropathy and retinopathy without neurological involvement, and which is the result of a compound heterozygous state arising from the inheritance of two recessive potentially pathogenic variants, previously not described. MATERIALS AND METHODS: Retrospective report, with complete ophthalmic examination, multimodal imaging, electroretinography, and whole exome sequencing performed on a family with three affected siblings. RESULTS: We show that affected individuals in the described family inherited two heterozygous variants of the COQ2 gene, resulting in a frameshift variant in one allele, and a predicted deleterious missense variant in the second allele (c.288dupC,p.(Ala97Argfs*56) and c.376C > G,p.(Arg126Gly) respectively). Electroretinography results were consistent with rod-cone dystrophy in the affected individuals. All affected individuals in the family exhibited the characteristic retinopathy as well as end-stage nephropathy, without evidence of any neurological involvement. CONCLUSIONS: We identified two novel compound heterozygous variants of the COQ2 gene that result in primary coenzyme Q deficiency. Targeted sequencing of coenzyme Q biosynthetic pathway genes may be useful in diagnosing oculorenal clinical presentations syndromes not explained by more well known syndromes (e.g., Senior-Loken and Bardet-Biedl syndromes).

Predictive testing for neurodegenerative diseases in the age of next-generation sequencing.

The availability and cost of next-generation sequencing (NSG) now allow testing large numbers of genes simultaneously. However, the gold standard for predictive testing has been to test only for a known family mutation or confirmed family disease. The goal of this study was to investigate the psychological impact of predictive testing for autosomal dominant neurodegenerative diseases without a known family mutation using next-generation sequencing panels compared to single-gene testing of a known family mutation. Fourteen individuals from families with a known mutation and 10 individuals with unknown family mutations participated. Participants completed questionnaires on demographics, genetic knowledge, and psychological measures of anxiety, depression, perceived personal control, rumination, and intolerance to uncertainty at baseline and 1 and 6 months after receiving results. Decision regret was measured 1 and 6 months after receiving results. Participants completed a modified Huntington disease genetic testing protocol with genetic counseling and neurological and psychological evaluation. Genetic testing of either the known family mutation or an NGS panel of neurodegenerative disease genes was performed. Semi-structured interviews were performed at 6 months post-results about their experience. Two-sample t tests were performed on data collected at each time point to identify significant between-group differences in demographic variables, baseline psychological scores, and baseline genetic knowledge scores. Within-group change over time was assessed by a mixed-effects model. Results of this study indicate that NGS panels for predictive testing for neurodegenerative disease are safe and beneficial to participants when performed within a modified HD protocol. Though significant differences in psychological outcomes were found, these differences may have been driven by genetic results and baseline psychological differences between individuals within the groups. Participants did not regret their decision to test and were largely pleased with the testing protocol.

Diffuse midline glioma with novel, potentially targetable, FGFR2-VPS35 fusion.

We report a case of a slow-growing, diffuse, infiltrating glioma in the right brainstem of a 9-yr-old boy. The tumor was negative by immunohistochemical staining for histone H3 K27M, BRAF V600E, and IDH1 R132H mutations. Fluorescence in situ hybridization did not reveal a BRAF duplication. Genomic profiling of the tumor, by DNA methylation array and cancer whole-exome and transcriptome sequencing, was performed. This analysis showed copy-number alterations, including gains of several chromosomes. In addition, a novel fusion involving the first 17 exons of FGFR2 fused to exon 2 of VPS35 was identified. This novel fusion is predicted to result in activation of fibroblast growth factor receptor (FGFR) signaling and is potentially targetable using FGFR inhibitors. This tumor expands the spectrum of pediatric diffuse gliomas.

Refractory celiac disease type II: An atypical case highlighting limitations of the current classification system.

Refractory celiac disease (RCD) is a rare condition associated with high morbidity that develops in individuals with celiac disease. It is known to be biologically heterogeneous, and currently two types are recognized based on immunophenotypic and molecular features, type I (RCD I) and type II (RCD II). Differentiating between RCD I and RCD II is critical, as patients with RCD II have substantially worse outcomes and a high risk of developing enteropathy-associated T-cell lymphoma. However, the current RCD classification is limited in scope, and atypical presentations and immunophenotypes are not recognized at present. Herein, we describe a unique case of RCD II with atypical clinical (primarily neurologic manifestations and lack of significant gastrointestinal symptoms), histopathologic (no villous atrophy), immunophenotypic (virtual absence of cytoplasmic CD3 expression), and molecular features (absence of clonal TR rearrangement and identification of pathogenic STAT3 and KMT2D mutations). This case highlights limitations of the current RCD classification system and the utility of next generation sequencing (NGS) studies in the diagnostic workup of RCD. Future algorithms need to recognize extraintestinal manifestations and incorporate atypical histopathologic and immunophenotypic features, as well as results of NGS analysis for RCD II classification.

Clinical Utilization, Utility, and Reimbursement for Expanded Genomic Panel Testing in Adult Oncology.

PURPOSE: The routine use of large next-generation sequencing (NGS) pan-cancer panels is required to identify the increasing number of, but often uncommon, actionable alterations to guide therapy. Inconsistent coverage and variable payment is hindering NGS adoption into clinical practice. A review of test utilization, clinical utility, coverage, and reimbursement was conducted in a cohort of patients diagnosed with high-risk cancer who received pan-cancer panel testing as part of their clinical care. MATERIALS AND METHODS: The Columbia Combined Cancer Panel (CCCP), a 467-gene panel designed to detect DNA variations in solid and liquid tumors, was performed in the Laboratory of Personalized Genomic Medicine at Columbia University Irving Medical Center. Utilization was characterized at test order. Results were reviewed by a molecular pathologist, followed by a multidisciplinary molecular tumor board where clinical utility was classified by consensus. Reimbursement was reviewed after payers provided final coverage decisions. RESULTS: NGS was performed on 359 high-risk tumors from 349 patients. Reimbursement data were available for 246 cases. The most common reason providers ordered CCCP testing was for patients diagnosed with a treatment-resistant or recurrent tumor (n = 214; 61%). Findings were clinically impactful for 229 cases (64%). Molecular alterations that may inform future therapy in the event of progression or relapse were found in 42% of cases, and a targeted therapy was initiated in 23 cases (6.6%). The majority of tests were denied coverage by payers (n = 190; 77%). On average, insurers reimbursed 10.75% of the total NGS service charge. CONCLUSION: CCCP testing identified clinically impactful alterations in 64% of cases. Limited coverage and low reimbursement remain barriers, and broader reimbursement policies are needed to adopt pan-cancer NGS testing that benefits patients into clinical practice.

Genetic and phenotypic characterization of indolent T-cell lymphoproliferative disorders of the gastrointestinal tract.

Indolent T-cell lymphoproliferative disorders of the gastrointestinal tract are rare clonal T-cell diseases that more commonly occur in the intestines and have a protracted clinical course. Different immunophenotypic subsets have been described, but the molecular pathogenesis and cell of origin of these lymphocytic proliferations is poorly understood. Hence, we performed targeted next-generation sequencing and comprehensive immunophenotypic analysis of ten indolent T-cell lymphoproliferative disorders of the gastrointestinal tract, which comprised CD4+ (n=4), CD8+ (n=4), CD4+/CD8+ (n=1) and CD4-/CD8- (n=1) cases. Genetic alterations, including recurrent mutations and novel rearrangements, were identified in 8/10 (80%) of these lymphoproliferative disorders. The CD4+, CD4+/CD8+, and CD4-/CD8- cases harbored frequent alterations of JAK-STAT pathway genes (5/6, 82%); STAT3 mutations (n=3), SOCS1 deletion (n=1) and STAT3-JAK2 rearrangement (n=1), and 4/6 (67%) had concomitant mutations in epigenetic modifier genes (TET2, DNMT3A, KMT2D). Conversely, 2/4 (50%) of the CD8+ cases exhibited structural alterations involving the 3' untranslated region of the IL2 gene. Longitudinal genetic analysis revealed stable mutational profiles in 4/5 (80%) cases and acquisition of mutations in one case was a harbinger of disease transformation. The CD4+ and CD4+/CD8+ lymphoproliferative disorders displayed heterogeneous Th1 (T-bet+), Th2 (GATA3+) or hybrid Th1/Th2 (T-bet+/GATA3+) profiles, while the majority of CD8+ disorders and the CD4-/CD8- disease showed a type-2 polarized (GATA3+) effector T-cell (Tc2) phenotype. Additionally, CD103 expression was noted in 2/4 CD8+ cases. Our findings provide insights into the pathogenetic bases of indolent T-cell lymphoproliferative disorders of the gastrointestinal tract and confirm the heterogeneous nature of these diseases. Detection of shared and distinct genetic alterations of the JAK-STAT pathway in certain immunophenotypic subsets warrants further mechanistic studies to determine whether therapeutic targeting of this signaling cascade is efficacious for a proportion of patients with these recalcitrant diseases.

Cytogenetic analysis of 130 renal oncocytomas identify three distinct and mutually exclusive diagnostic classes of chromosome aberrations.

The cytogenetic alterations in renal oncocytoma (RO) are poorly understood. We analyzed 130 consecutive RO for karyotypic alterations. Clonal chromosome abnormalities were identified in 63 (49%) cases, which could be categorized into three classes of mutually exclusive cytogenetic categories. Class 1 (N = 20) RO had diploid karyotypes with characteristic 11q13 rearrangement in balanced translocations with 10 or more different chromosome partners in all cases. We identified recurrent translocation partners at 5q35, 6p21, 9p24, 11p13-14, and 11q23, and confirmed that CCND1 gene rearrangement at 11q13 utilizing fluorescence in situ hybridization (FISH). Class 2 RO (N = 25) exhibited hypodiploid karyotypes with loss of chromosome 1 and/or losses of Y in males and X in females in all cases. The class 3 tumors comprising of 18 cases showed diverse types of abnormalities with the involvement of two or more chromosomes exclusive of abnormalities seen in classes 1 and 2 tumors. Furthermore, karyotypically uninformative cases were subjected to FISH analysis to identify classes 1 and 2 abnormalities. In this group, we found similar frequencies of CCND1 rearrangement, loss of chromosome 1 or Y as with karyotypically abnormal cases. We validated our results against 91 tumors from the Mitelman database. Correlation of clinical data with all the three classes of ROs showed no clear evidence of overall patient survival. Our findings support the hypothesis that RO exhibit three principal cytogenetic categories, which may have different roles in initiation and/or progression. These cytogenetic markers provide a key tool in the diagnostic evaluation of RO.

Multi-Institutional Evaluation of Interrater Agreement of Variant Classification Based on the 2017 Association for Molecular Pathology, American Society of Clinical Oncology, and College of American Pathologists Standards and Guidelines for the Interpretation and Reporting of Sequence Variants in Cancer.

This multi-institutional study was undertaken to evaluate interrater reliability of the 2017 Association for Molecular Pathology/American Society of Clinical Oncology/College of American Pathologists guidelines for interpretation and reporting of oncology sequence variants and to assess current practices and perceptions surrounding these guidelines. Fifty-one variants were distributed to 20 participants from 10 institutions for classification using the new guidelines. Agreement was assessed using chance-corrected agreement (Cohen κ). κ was 0.35. To evaluate if data sharing could help resolve disagreements, a summary of variant classifications and additional information about each variant were distributed to all participants. κ improved to 0.7 after the original classifications were revised. Participants were invited to take a web-based survey regarding their perceptions of the guidelines. Only 20% (n = 3) of the survey respondents had prior experience with the guidelines in clinical practice. The main perceived barriers to guideline implementation included the complexity of the guidelines, discordance between clinical actionability and pathobiologic relevance, lack of familiarity with the new classifications, and uncertainty when applying criteria to potential germline variants. This study demonstrates noteworthy discordances between pathologists for variant classification in solid tumors when using the 2017 Association for Molecular Pathology/American Society of Clinical Oncology/College of American Pathologists guidelines. These findings highlight potential areas for clarification/refinement before mainstream clinical adoption.

Malignant Rhabdoid Tumor, an Aggressive Tumor Often Misclassified as Small Cell Variant of Hepatoblastoma.

The clinical management of pediatric liver tumors involves stratification into risk groups. One previously defined, high-risk group of hepatoblastomas is the small cell undifferentiated variant. In light of molecular studies showing SMARCB1 deletion in these tumors, it is now recognized that most small cell, undifferentiated liver tumors represent an aggressive unrelated tumor-the malignant rhabdoid tumor (MRT). SMARCB1 is a member of the chromatin remodeling SWI/SNF complex and encodes the INI1 protein. The histologic diagnosis of MRT is currently based on INI1 negative immunoreactivity and the presence of rhabdoid morphology. INI1-negative small cell liver tumors lacking classic rhabdoid morphology are often misclassified as small cell undifferentiated hepatoblastomas (SCUD-HB), according to the current classification. Pediatric liver tumors diagnosed between 2003-2017 as SCUD-HB (four cases) or MRT (two cases) were identified from the Columbia University Pathology Department Archives. All tumors were associated with normal or low serum alpha fetoprotein levels, and showed an absence of immunohistochemical staining of hepatocellular markers (Hep-par1, Arginase) and loss of INI1 staining. Two cases were initially diagnosed as MRT, one with prominent rhabdoid morphology, the other with predominant small cell morphology. The remaining four cases with small cell morphology were classified as SCUD-HB. Ancillary molecular studies confirmed the loss of SMARCB1, supporting the diagnosis of MRT in all cases, proving morphology an unreliable criterion. It is critical to eliminate the term INI1-negative hepatoblastoma from the current classification scheme, and classify INI1-negative tumors as MRT, particularly since high-risk HB-chemotherapy regimens are not effective for treating MRT.