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Increased adoption of personalized medicine has brought comprehensive genomic profiling (CGP) to the forefront. However, differences in assay, bioinformatics, and reporting systems and lack of understanding of their complex interplay are a challenge for implementation and achieving uniformity in CGP testing. Two commercially available, tissue-based, in-house CGP assays were compared, in combination with a tertiary analysis solution in a research use only (RUO) context: the AVENIO Tumor Tissue CGP RUO Kit paired with navify Mutation Profiler (RUO) software and the TruSight Oncology 500 RUO assay paired with PierianDx Clinical Genomics Workspace software. Agreements and differences between the assays were assessed for short variants, copy number alterations, rearrangements, tumor mutational burden, and microsatellite instability, including variant categorization and clinical trial-matching (CTM) recommendations. Results showed good overall agreement for short variant, known gene fusion, and microsatellite instability detection. Important differences were obtained in tumor mutational burden scoring, copy number alteration detection, and CTM. Differences in variant and biomarker detection could be explained by bioinformatic approaches to variant calling, filtering, tiering, and normalization; differences in CTM, by underlying reported variants and conceptual differences in system parameters. Thus, distinctions between different approaches may lead to inconsistent results. Complexities in calling, filtering, and interpreting variants illustrate key considerations for implementation of any high-quality CGP in the laboratory and bringing uniformity to genomic insight results., Competing Interests: Disclosure Statement H.-P.A., F.F., and U.S. are employees of Signature Diagnostics GmbH (H.-P.A. at the time of the analysis, but now retired). M.C.H., J.O., and J.M.A. are employees of Foundation Medicine, Inc. (J.O. at the time of the analysis, but is no longer an employee), and M.C., S.O., and T.W. are employees of Foundation Medicine GmbH; all hold stock in F. Hoffmann-La Roche Ltd. K.H. is an employee of Helios MVZ Pathologie Erfurt GmbH. M.T. is an employee of F. Hoffmann-La Roche AG and holds stock in F. Hoffmann-La Roche Ltd. D.G. holds stock in Univ8 Genomics Ltd.; has received honoraria (institution) from Roche, Illumina, Novartis, AstraZeneca, Eli Lilly, Amgen, and Janssen; has participated in a consulting or advisory role (institution) for AstraZeneca and Novartis; has participated in a speakers' bureau (institution) for AstraZeneca; and has received travel, accommodations, and expenses (institution) from Roche, AstraZeneca, and Illumina. E.P. received compensation for speaking engagements and advisory roles, with payments made directly to his affiliated university, from Roche, AstraZeneca, Novartis, and MSD. H.S. and M.D.V. are employees of Roche Diagnostics Solutions, Inc., and M.D.V. holds stock in F. Hoffmann-La Roche Ltd. E.S. reports lectures for Bio-Rad, Seracare, Novartis, Roche, Biocartis, Illumina, Eli Lilly, Janssen-Cilag, Pfizer, AstraZeneca, and Agena Bioscience; he is consultant in advisory boards for MSD/Merck, GSK, AstraZeneca, Astellas Pharma, Sysmex, Roche, Pfizer, Novartis, Bayer, Bristol Myers Squibb, Eli Lilly, Amgen, Biocartis, Illumina, Agena Bioscience, Janssen-Cilag (Johnson & Johnson), Sinnovisionlab, Diaceutics, and CC Diagnostics; and received research grants from Pfizer, Biocartis, Invitae-ArcherDX, AstraZeneca, Agena Bio-science, Bristol Myers Squibb, Bio-Rad, Roche, Boehringer Ingelheim, CC Diagnostics, and Abbott (all paid to University Medical Center Groningen account); and travel reimbursements from Bio-Rad, Abbott, Illumina, Agena Bioscience, and Roche. All authors received support in the form of third-party writing assistance for this manuscript, furnished by Nucleus Global, an Inizio company, and provided by F. Hoffmann-La Roche Ltd./Roche Diagnostics Solutions, Inc., (Copyright © 2024 Association for Molecular Pathology and American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.)

Microsatellite instability high (MSI-H) and mismatch repair deficient (dMMR) tumor status have been demonstrated to predict patient response to immunotherapies. We developed and validated a next-generation sequencing (NGS)-based companion diagnostic (CDx) to detect MSI-H solid tumors via a comprehensive genomic profiling (CGP) assay, FoundationOne®CDx (F1CDx). To determine MSI status, F1CDx calculates the fraction of unstable microsatellite loci across >2000 loci using a fraction-based (FB) analysis. Across solid tumor types, F1CDx demonstrated a high analytical concordance with both PCR (n = 264) and IHC (n = 279) with an overall percent agreement (OPA) of 97.7% and 97.8%, respectively. As part of a retrospective bridging clinical study from KEYNOTE-158 Cohort K and KEYNOTE-164, patients with MSI-H tumors as determined by F1CDx demonstrated an objective response rate (ORR) of 43.0% to pembrolizumab. In real-world cancer patients from a deidentified clinicogenomic database, F1CDx was at least equivalent in assessing clinical outcome following immunotherapy compared with MMR IHC. Demonstrated analytical and clinical performance of F1CDx led to the pan-tumor FDA approval in 2022 of F1CDx to identify MSI-H solid tumor patients for treatment with pembrolizumab. F1CDx is an accurate, reliable, and FDA-approved method for the identification of MSI-H tumors for treatment with pembrolizumab., (© 2024. The Author(s).)

For cancer clinical trials that require central confirmation of tumor genomic profiling, exhaustion of tissue from standard-of-care testing may prevent enrollment. For Lung-MAP, a master protocol that requires results from a defined centralized clinical trial assay to assign patients to a therapeutic substudy, we developed a process to repurpose existing commercial vendor raw genomic data for eligibility: genomic data reanalysis (GDR). Molecular results for substudy assignment were successfully generated for 369 of the first 374 patients (98.7%) using GDR for Lung-MAP, with a median time from request to result of 9 days. During the same period, 691 of 791 (87.4%) tissue samples received successfully yielded results, in a median of 14 days beyond sample acquisition. GDR is a scalable bioinformatic pipeline that expedites reanalysis of existing data for clinical trials in which validated integral biomarker testing is required for participation., (© The Author(s) 2024. Published by Oxford University Press.)

Myxofibrosarcoma (MFS) is a common soft tissue sarcoma of the elderly that typically shows low tumor mutational burden, with mutations in TP53 and in genes associated with cell cycle checkpoints ( RB1 , CDKN2A ). Unfortunately, no alterations or markers specific to MFS have been identified and, as a consequence, there are no effective targeted therapies. The receptor tyrosine kinase AXL, which drives cellular proliferation, is targetable by new antibody-based therapeutics. Expression of AXL messenger RNA is elevated in a variety of sarcoma types, with the highest levels reported in MFS, but the pathogenic significance of this finding remains unknown. To assess a role for AXL abnormalities in MFS, we undertook a search for AXL genomic alterations in a comprehensive genomic profiling database of 463,546 unique tumors (including 19,879 sarcomas, of which 315 were MFS) interrogated by targeted next-generation DNA and/or RNA sequencing. Notably, the only genomic alterations recurrent in a specific sarcoma subtype were AXL W451C (n = 8) and AXL W450C (n = 2) mutations. The tumors involved predominantly older adults (age: 44 to 81 [median: 72] y) and histologically showed epithelioid and spindle-shaped cells in a variably myxoid stroma, with 6 cases diagnosed as MFS, 3 as undifferentiated pleomorphic sarcoma (UPS), and 1 as low-grade sarcoma. The AXL W451C mutation was not identified in any non-sarcoma malignancy. A review of publicly available data sets revealed a single AXL W451C-mutant case of UPS that clustered with MFS/UPS by methylation profiling. Functional studies revealed a novel activation mechanism: the W451C mutation causes abnormal unregulated dimerization of the AXL receptor tyrosine kinase through disulfide bond formation between pairs of mutant proteins expressing ectopic cysteine residues. This dimerization triggers AXL autophosphorylation and activation of downstream ERK signaling. We further report sarcomas of diverse histologic subtypes with AXL gene amplifications, with the highest frequency of amplification identified in MFS cases without the W451C mutation. In summary, the activating AXL W451C mutation appears highly specific to MFS, with a novel mechanism to drive unregulated signaling. Moreover, AXL gene amplifications and messenger RNA overexpression are far more frequent in MFS than in other sarcoma subtypes. We conclude that these aberrations in AXL are distinct features of MFS and may aid diagnosis, as well as the selection of available targeted therapies., Competing Interests: Conflicts of Interest and Source of Funding: E.A.W., D.A.M., M.C.H., and J.S.R. are employees/consultants of Foundation Medicine, Inc., a wholly-owned subsidiary of Roche Holdings, Inc. and Roche Finance Ltd.; these employees have an equity interest in an affiliate of these Roche entities. For the remaining authors, none were declared., (Copyright © 2024 The Author(s). Published by Wolters Kluwer Health, Inc.)

Background: Schwannomas are common peripheral nerve sheath tumors that can cause severe morbidity given their stereotypic intracranial and paraspinal locations. Similar to many solid tumors, schwannomas and other nerve sheath tumors are primarily thought to arise due to aberrant hyperactivation of the RAS growth factor signaling pathway. Here, we sought to further define the molecular pathogenesis of schwannomas., Methods: We performed comprehensive genomic profiling on a cohort of 96 human schwannomas, as well as DNA methylation profiling on a subset. Functional studies including RNA sequencing, chromatin immunoprecipitation-DNA sequencing, electrophoretic mobility shift assay, and luciferase reporter assays were performed in a fetal glial cell model following transduction with wildtype and tumor-derived mutant isoforms of SOX10., Results: We identified that nearly one-third of sporadic schwannomas lack alterations in known nerve sheath tumor genes and instead harbor novel recurrent in-frame insertion/deletion mutations in SOX10, which encodes a transcription factor responsible for controlling Schwann cell differentiation and myelination. SOX10 indel mutations were highly enriched in schwannomas arising from nonvestibular cranial nerves (eg facial, trigeminal, vagus) and were absent from vestibular nerve schwannomas driven by NF2 mutation. Functional studies revealed these SOX10 indel mutations have retained DNA binding capacity but impaired transactivation of glial differentiation and myelination gene programs., Conclusions: We thus speculate that SOX10 indel mutations drive a unique subtype of schwannomas by impeding proper differentiation of immature Schwann cells., (© The Author(s) 2023. Published by Oxford University Press on behalf of the Society for Neuro-Oncology.)

Context.—: Gene editing-based therapies are currently in development in the areas of oncology, inherited disease, and infectious disease. These potentially life-altering therapies are derived from decades of research in both academic and industry settings that developed technologies rooted in principles and products of nature. However, with such technologic developments come many important considerations, including adverse risks, high cost, and ethical questions., Objective.—: To educate pathologists about gene editing technologies, inform them of potential indications and risks, outline regulatory and practical issues that could affect hospital-based practice and laboratory testing, and advocate that pathologists need to be present at discussions among industry and regulators pertaining to gene editing-based therapies., Design.—: A Gene Editing Workgroup, facilitated by the College of American Pathologists Personalized Health Care Committee and consisting of pathologists of various backgrounds, was convened to develop an educational paper to serve as a stimulus to increase pathologist involvement and inquiry in gene editing therapeutic and diagnostic implementation., Results.—: Through multiple discussions and literature review, the workgroup identified potential gaps in pathologists' knowledge of gene editing. Additional topics that could impact pathology and laboratory medicine were also identified and summarized in order to facilitate pathologists as stakeholders in gene editing therapy administration and monitoring and potential use in diagnostics., Conclusions.—: Gene editing therapy is a complex but potentially transformative area of medicine. This article serves as an introduction to pathologists to assist them in future discussions with colleagues and potentially identify and alter pathology practices that relate to gene editing., (© 2023 College of American Pathologists.)

Context—: Gene editing-based therapies are currently in development in the areas of oncology, inherited disease, and infectious disease. These potentially life-altering therapies are derived from decades of research in both academic and industry settings that developed technologies rooted in principles and products of nature. However, with such technologic developments come many important considerations, including adverse risks, high cost, and ethical questions., Objective—: To educate pathologists about gene editing technologies, inform them of potential indications and risks, outline regulatory and practical issues that could affect hospital-based practice and laboratory testing, and advocate that pathologists need to be present at discussions among industry and regulators pertaining to gene editing-based therapies., Design—: A Gene Editing Workgroup, facilitated by the College of American Pathologists Personalized Health Care Committee and consisting of pathologists of various backgrounds, was convened to develop an educational paper to serve as a stimulus to increase pathologist involvement and inquiry in gene editing therapeutic and diagnostic implementation., Results—: Through multiple discussions and literature review, the workgroup identified potential gaps in pathologists' knowledge of gene editing. Additional topics that could impact pathology and laboratory medicine were also identified and summarized in order to facilitate pathologists as stakeholders in gene editing therapy administration and monitoring and potential use in diagnostics., Conclusions—: Gene editing therapy is a complex but potentially transformative area of medicine. This article serves as an introduction to pathologists to assist them in future discussions with colleagues and potentially identify and alter pathology practices that relate to gene editing., (© 2023 College of American Pathologists.)

Gene fusions involving EWSR1 or FUS as the 5' partner have been reported in a diverse array of sarcomas. Here, we characterize the histopathology and genomics of six tumors harboring a gene fusion between EWSR1 or FUS and POU2AF3, an understudied, putative colorectal cancer predisposition gene. Striking morphologic features reminiscent of synovial sarcoma were observed including a biphasic appearance with variable fusiform to epithelioid cytomorphology and staghorn-type vasculature. RNA sequencing demonstrated variable breakpoints in EWSR1/FUS along with similar breakpoints in POU2AF3 that encompassed a 3' portion of this gene. For cases in which additional information was available, the behavior of these neoplasms was aggressive with local spread and/or distant metastases. Although further studies are needed to confirm the functional significance of our findings, POU2AF3 fusions to EWSR1 or FUS may define a novel type of POU2AF3-rearranged sarcomas with aggressive, malignant behavior., (© 2023 Wiley Periodicals LLC.)

Background: In 2020, pembrolizumab was approved as a therapy for triple-negative breast cancer (TNBC) with the companion diagnostic DAKO 22C3 programmed death ligand-1 (PD-L1) immunohistochemistry assay. The study aimed to determine the landscape of PD-L1 expression as detected by the DAKO 22C3 PD-L1 assay in breast cancer subtypes and compare the clinicopathologic and genomic characteristics of PD-L1 positive and negative TNBC., Methods: PD-L1 expression using the DAKO 22C3 antibody was scored using a combined positive score (CPS) and positive status was defined as CPS ≥10. Comprehensive genomic profiling was performed using the FoundationOne CDx assay., Results: Of the 396 BC patients stained with DAKO 22C3, the majority were HR+/HER2- and TNBC (42% and 36%, respectively). Median PD-L1 expression and frequency of CPS ≥10 was highest in TNBC cases (median: 7.5, 50% CPS ≥10) and lowest in the HR+/HER2- group (median: 1.0, 15.5% CPS ≥10) (P < .0001). A comparison of PD-L1 positive and PD-L1 negative TNBC demonstrated no significant differences in clinicopathologic or genomic characteristics. TNBC tissue samples from the breast did have an observed enrichment for PD-L1 positivity compared to TNBC tissue samples from a metastatic site (57% vs. 44%), but this was not statistically significant (P = .1766). In the HR+/HER2- group, genomic alterations in TP53, CREBBP, and CCNE1 were more prevalent and genomic loss of heterozygosity was higher in the PD-L1(+) group compared to the PD-L1(-) group., Conclusions: The subtypes of breast cancer have distinct patterns of PD-L1 expression, supporting that further research of immunotherapies may include specific evaluation of optimum cutoffs for non-TNBC patients. In TNBC, PD-L1 positivity is not associated with other clinicopathologic or genomic features and should be integrated into future studies of immunotherapy efficacy., (© The Author(s) 2023. Published by Oxford University Press.)

Treatment of non-small cell lung cancer is increasingly biomarker driven with multiple genomic alterations, including those in the epidermal growth factor receptor (EGFR) gene, that benefit from targeted therapies. We developed a set of algorithms to assess EGFR status and morphology using a real-world advanced lung adenocarcinoma cohort of 2099 patients with hematoxylin and eosin (H&E) images exhibiting high morphological diversity and low tumor content relative to public datasets. The best performing EGFR algorithm was attention-based and achieved an area under the curve (AUC) of 0.870, a negative predictive value (NPV) of 0.954 and a positive predictive value (PPV) of 0.410 in a validation cohort reflecting the 15% prevalence of EGFR mutations in lung adenocarcinoma. The attention model outperformed a heuristic-based model focused exclusively on tumor regions, and we show that although the attention model also extracts signal primarily from tumor morphology, it extracts additional signal from non-tumor tissue regions. Further analysis of high-attention regions by pathologists showed associations of predicted EGFR negativity with solid growth patterns and higher peritumoral immune presence. This algorithm highlights the potential of deep learning tools to provide instantaneous rule-out screening for biomarker alterations and may help prioritize the use of scarce tissue for biomarker testing., (© 2023. The Author(s).)

Context.—: Multiple procedural techniques can be used to obtain tissue to create a formalin-fixed, paraffin-embedded specimen for comprehensive genomic profiling (CGP) in lung cancer. The literature is mixed on whether the procedure affects CGP success., Objective.—: To examine whether biopsy procedure affects lung cancer CGP success., Design.—: This was a cross-sectional study of all patients with lung cancer whose specimens were submitted for CGP between January and February 2020. Multiple quality control metrics were used to determine whether cases were successfully profiled., Results.—: In all, 3312 samples were identified. Overall, 67.5% (2236 of 3312) of samples were obtained from biopsies, 13.0% (432 of 3312) from fine-needle aspirations (FNAs), 9.7% (321 of 3312) from resections, 5.3% (174 of 3312) from fluid cytology cell blocks, and 4.5% (149 of 3312) from bone biopsies. Overall, 70.1% (2321 of 3312) of cases passed CGP, 15.4% (510 of 3312) of cases were released as qualified reports, and 14.5% (481 of 3312) of cases failed CGP. Resection samples were the most likely to be successfully sequenced, failing in only 2.8% (9 of 321) of instances, while fluid cytology specimens were the least likely, failing in 23.0% (40 of 174) of instances. Biopsy (14.5% [324 of 2236]), FNA (18.5% [80 of 432]), and bone biopsy (18.8% [28 of 149]) specimens failed at intermediate frequencies. On multivariate logistic regression analysis of CGP success on specimen type, fluid cytology (odds ratio [OR], 0.08; 95% CI, 0.03-0.19), biopsy (OR, 0.25; 95% CI, 0.11-0.52), FNA (OR, 0.14; 95% CI, 0.06-0.32), and bone biopsy (OR, 0.07; 95% CI, 0.03-0.17) specimens had decreased odds of CGP success relative to resection samples. Among patients with successfully sequenced samples, 48.0% were eligible for at least 1 therapy, based on a companion diagnostic or National Comprehensive Cancer Network biomarker., Conclusions.—: The method of tissue acquisition was an important preanalytic factor that determined whether a sample would be successfully sequenced and whether a clinically actionable genomic alteration would be detected.

In this study, we report the clinicopathologic and genomic profiles of 891 patients with RET fusion driven advanced solid tumors. All patient samples were tested using a tissue-based DNA hybrid capture next generation sequencing (NGS) assay and a subset of the samples were liquid biopsies tested using a liquid-based hybrid capture NGS assay. RET fusions were found in 523 patients with NSCLC and in 368 patients with other solid tumors. The two tumor types with the highest number of RET fusion were lung adenocarcinoma and thyroid papillary carcinoma, and they had a prevalence rate 1.14% (455/39,922) and 9.09% (109/1199), respectively. A total of 61 novel fusions were discovered in this pan-tumor cohort. The concordance of RET fusion detection across tumor types among tissue and liquid-based NGS was 100% (8/8) in patients with greater than 1% composite tumor fraction (cTF). Herein, we present the clinicopathologic and genomic landscape of a large cohort of RET fusion positive tumors and we observed that liquid biopsy-based NGS is highly sensitive for RET fusions at cTF ≥1%., (© 2023. The Author(s).)

Challenges with sequencing tissue samples from patients with prostate cancer have been reported in clinical trials. To assess the success rate of comprehensive genomic profiling (CGP) for prostate cancer patients, we analyzed a real-world cohort who underwent sequencing of their prostate tissue sample as well as a subset of patients with a reflex liquid biopsy. Overall, a significant majority (82%) of tissue prostate carcinoma samples yielded reportable CGP results. Of those samples that were unsuccessful, most (75%) were inadequate samples that did not meet pre-established criteria to advance into sequencing. For cases where liquid CGP was performed if tissue CGP was unsuccessful, mutations that were likely attributable to prostate carcinoma were observed in most cases and all cases were successful in generating a report. These results suggest that, for CGP testing, prostate cancer tissue is a reasonable matrix type and that liquid samples can be effectively used as an alternative to tissue., (© The Author(s) 2022. Published by Oxford University Press.)

Activation of the tyrosine kinase receptor IGF1R is targetable with existing tyrosine kinase inhibitors (TKIs) and monoclonal antibodies, but mutations in IGF1R have not been systematically characterized. Pan-cancer analysis of 326,911 tumors identified two distinct, activating non-frameshift insertion hotspots in IGF1R, which were significantly enriched in adenoid cystic carcinomas (ACCs). IGF1R alterations from 326,911 subjects were analyzed by variant effect prediction class, position within the gene, and cancer type. 6502 (2.0%) samples harbored one or more alterations in IGF1R. Two regions were enriched for non-frameshift insertions: codons 663-666 at the hinge region of the fibronectin type 3 domain and codons 1034-1049 in the tyrosine kinase domain. Hotspot insertions were highly enriched in ACCs (27.3-fold higher than in the remainder of the pan-cancer dataset; P = 2.3 × 10 -17 ). Among salivary gland tumors, IGF1R hotspot insertions were entirely specific to ACCs. IGF1R alterations were most often mutually exclusive with other ACC drivers (9/15, 60%). Tumors with non-frameshift hotspot IGF1R insertions represent a novel, potentially targetable subtype of ACC. Additional studies are needed to determine whether these patients respond to existing IGF1R inhibitors., (© 2022. The Author(s), under exclusive licence to United States & Canadian Academy of Pathology.)

Background: In patients with non-small cell lung cancer (NSCLC), 10%-40% will eventually develop brain metastases. We present the clinicopathologic, genomic, and biomarker landscape of a large cohort of NSCLC brain metastases (NSCLC-BM) samples., Materials and Methods: We retrospectively analyzed 3035 NSCLC-BM tested with comprehensive genomic profiling (CGP) during routine clinical care. In addition, we compared the NSCLC-BM to a separate cohort of 7277 primary NSCLC (pNSCLC) specimens. Finally, we present data on 67 paired patients with NSCLC-BM and pNSCLC., Results: Comprehensive genomic profiling analysis of the 3035 NSCLC-BMs found that the most frequent genomic alterations (GAs) were in the TP53, KRAS, CDKN2A, STK11, CDKN2B, EGFR, NKX2-1, RB1, MYC, and KEAP1 genes. In the NSCLC-BM cohort, there were significantly higher rates of several targetable GAs compared with pNSCLC, including ALK fusions, KRAS G12C mutations, and MET amplifications; and decreased frequency of MET exon14 skipping mutations (all P < .05). In the subset of NSCLC-BM (n = 1063) where concurrent PD-L1 immunohistochemistry (IHC) was performed, 54.7% of the patients with NSCLC-BM were eligible for pembrolizumab based on PD-L1 IHC (TPS ≥ 1), and 56.9% were eligible for pembrolizumab based on TMB-High status. In addition, in a series 67 paired pNSCLC and NSCLC-BM samples, 85.1% (57/67) had at least one additional GA discovered in the NSCLC-BM sample when compared with the pNSCLC sample., Conclusions: Herein, we defined the clinicopathologic, genomic, and biomarker landscape of a large cohort of patients with NSCLC-BM which can help inform study design of future clinical studies for patients with NSCLC with BM. In certain clinical situations, metastatic NSCLC brain tissue or cerebral spinal fluid specimens may be needed to fully optimize personalized treatment., (© The Author(s) 2022. Published by Oxford University Press.)

Background: In the current study, we examined the real-world prevalence of highly pigmented advanced melanomas (HPMel) and the clinicopathologic, genomic, and ICPI biomarker signatures of this class of tumors., Materials and Methods: Our case archive of clinical melanoma samples for which the ordering physician requested testing for both PD-L1 immunohistochemistry (IHC) and comprehensive genomic profiling (CGP) was screened for HPMel cases, as well as for non-pigmented or lightly pigmented advanced melanoma cases (LPMel)., Results: Of the 1268 consecutive melanoma biopsies in our archive that had been submitted for PD-L1 IHC, 13.0% (165/1268) were HPMel and 87.0% (1103/1268) were LPMel. In the HPMel cohort, we saw a significantly lower tumor mutational burden (TMB, median 8.8 mutations/Mb) than in the LPMel group (11.4 mut/Mb), although there was substantial overlap. In examining characteristic secondary genomic alterations (GA), we found that the frequencies of GA in TERTp, CDKN2A, TP53, and PTEN were significantly lower in the HPMel cases than in LPMel. A higher rate of GA in CTNNB1, APC, PRKAR1A, and KIT was identified in the HPMel cohort compared with LPMel., Conclusions: In this study, we quantified the failure rates of melanoma samples for PD-L1 testing due to high melanin pigmentation and showed that CGP can be used in these patients to identify biomarkers that can guide treatment decisions for HPMel patients. Using this practical clinical definition for tumor pigmentation, our results indicate that HPMel are frequent at 13% of melanoma samples, and in general appear molecularly less developed, with a lower TMB and less frequent secondary GA of melanoma progression., (© The Author(s) 2022. Published by Oxford University Press.)

P53 immunohistochemical staining with antibodies targeted to epitopes at or near the N-terminus are commonly used in diagnostic pathology practice as a surrogate for TP53 mutations. The abnormal staining patterns indicating TP53 mutations include nuclear overexpression, null, and the recently described cytoplasmic staining. The latter staining pattern occurs with the less common TP53 mutations affecting its nuclear localization and/or tetramerization domains that are located toward the C-terminus. Here we describe the first two cases of pediatric sarcomas with cytoplasmic staining with P53 antibody against N-terminus epitope and the absence of staining with P53 antibody against C-terminus epitope. We propose that a more precise description of P53 immunohistochemical staining patterns should include the nature of the antibody used.

Inactivating mutations in tumor suppressor genes TP53 and RB1 are considered central drivers in leiomyosarcomas (LMSs). In high-risk human papillomavirus (HPV)-related tumors, a similar functional outcome is achieved through oncoproteins E6 and E7, which inactivate the p53 and RB1 proteins, respectively. Here, we hypothesized that HPV infection could provide an alternative mechanism for tumorigenesis in a subset of TP53/RB1-wildtype LMS. We evaluated tumor samples from 2585 consecutive unique patients carrying a diagnosis of gynecologic or soft tissue LMS. Tumor DNA and available RNA were analyzed by hybrid-capture-based next-generation sequencing/comprehensive genomic profiling of 406 genes and transcripts (FoundationOneHeme). Of the initial 2585 cases, we excluded 16 based on the presence of molecular alterations that are considered defining for sarcomas other than LMS. In the remaining 2569 cases, we searched for LMS that were TP53/RB1-wildtype (n=486 of 2569; 18.9%). We also searched LMS tumors for HPV sequences that we then classified into genotypes by de novo assembly of nonhuman sequencing reads followed by alignment to the RefSeq database. Among TP53/RB1-wildtype LMS, we identified 18 unique cases harboring HPV sequences. Surprisingly, most (n=11) were HPV51-positive, and these 11 represented all HPV51-positive tumors in our entire LMS database (n=11 of 2569; 0.4%). The absence of genomic alterations in TP53 or RB1 in HPV51-positive LMS represented a marked difference from HPV51-negative LMS (n=2558; 0% vs. 72% [P<0.00001], 0% vs. 53% [P=0.0002]). In addition, compared with HPV51-negative LMS, HPV51-positive LMS were significantly enriched for genomic alterations in ATRX (55% vs. 24%, P=0.027) and TSC1 (18% vs. 0.6%, P=0.0047). All HPV51-positive LMS were in women; median age was 54 years at surgery (range: 23 to 74 y). All known primary sites were from the gynecologic tract or adjacent anogenital area, including 5 cases of vaginal primary site. Histology was heterogeneous, with evaluable cases showing predominant epithelioid (n=5) and spindle (n=5) morphology. In situ hybridization confirmed the presence of high-risk HPV E6/E7 mRNA in tumor cells in three of three evaluable cases harboring HPV51 genomic sequences. Overall, in our pan-LMS analysis, HPV reads were identified in a subset of TP53/RB1-wildtype LMS. For all HPV51-associated LMS, the striking absence of any detectable TP53 or RB1 mutations and predilection for the female lower reproductive tract supports our hypothesis that high-risk HPV can be an alternative tumorigenic mechanism in this distinct class of LMS., Competing Interests: Conflicts of Interest and Source of Funding: E.A.W., M.M., J.Y.T., M.C.H., D.A.M., B.B.S., B.M.A., J.S.R., S.H.R., D.I.L., and J.A.E. are employees or consultants of Foundation Medicine Inc., a wholly owned subsidiary of Roche Holdings Inc. and Roche Finance Ltd, and these employees have equity interest in an affiliate of these Roche entities. For the remaining authors none were declared., (Copyright © 2022 The Author(s). Published by Wolters Kluwer Health, Inc.)

Background: Liquid biopsy is a powerful tool that can enable treatment decisions for metastatic prostate cancer patients with difficult-to-biopsy tumors. However, the detection of genomic alterations via liquid biopsy is limited by the fraction (tumor fraction [TF]) of circulating tumor DNA (ctDNA) within the total cell-free DNA content. While prior work has preliminarily correlated TF with clinical features of prostate cancer, we sought to validate and provide additional resolution, such that a clinical practitioner might anticipate the probability of successful liquid biopsy profiling leveraging commonly assessed clinical and laboratory features., Methods: A total of 813 liquid biopsy specimens were assessable, with 545 associated with a PSA prostate specific antigen measurement, collected in standard-of-care settings across approximately 280 US academic or community-based cancer clinics from September 2018 to July 2021. Deidentified data were captured into a real-world clinico-genomic database (CGDB). Comprehensive genomic profiling (CGP) was performed on extracted cell-free DNA from liquid biopsy samples., Results: In multivariable models, higher PSA level, lower hemoglobin, lower albumin, higher alkaline phosphatase (all p < 0.001), and collection of liquid biopsy blood draw within 60 days of new treatment initiation (p = 0.002) were the most strongly associated features with higher TF. At PSA levels of <5 ng/ml, 43% of patients had a TF of <1% indicating an increased likelihood of unevaluable results. Conversely, at PSA levels of >5 ng/ml, 78% of patients had a TF of at least 1% and 46% had a TF of ≥10%, suggesting improved sensitivity for detection of targetable alterations., Conclusions: Universal genomic profiling of prostate cancers will require complementary use of liquid biopsy and tumor tissue profiling for suitable patients. The likelihood of adequate ctDNA shedding into plasma is one consideration when deciding whether to pursue CGP via liquid biopsy versus tumor profiling. Our real-world data suggest that PSA < 5 ng/ml is associated with lower ctDNA yield on liquid biopsy, potentially increasing the incidence of negative results or a need for confirmation with tissue testing., (© 2022 The Authors. The Prostate published by Wiley Periodicals LLC.)

Background: Among patients with breast carcinoma who have metastatic disease, 15%-30% will eventually develop brain metastases. We examined the genomic landscape of a large cohort of patients with breast carcinoma brain metastases (BCBMs) and compared it with a cohort of patients with primary breast carcinomas (BCs)., Material and Methods: We retrospectively analyzed 733 BCBMs tested with comprehensive genomic profiling (CGP) and compared them with 10,772 primary breast carcinomas (not-paired) specimens. For a subset of 16 triple-negative breast carcinoma (TNBC)-brain metastasis samples, programmed death-ligand 1 (PD-L1) immunohistochemistry (IHC) was performed concurrently., Results: A total of 733 consecutive BCBMs were analyzed. Compared with primary BCs, BCBMs were enriched for genomic alterations in TP53 (72.0%, 528/733), ERBB2 (25.6%, 188/733), RAD21 (14.1%, 103/733), NF1 (9.0%, 66/733), BRCA1 (7.8%, 57/733), and ESR1 (6.3%,46/733) (p < .05 for all comparisons). Immune checkpoint inhibitor biomarkers such as high tumor mutational burden (TMB-high; 16.2%, 119/733); high microsatellite instability (1.9%, 14/733); CD274 amplification (3.6%, 27/733); and apolipoprotein B mRNA editing enzyme, catalytic polypeptide-like mutational signature (5.9%, 43/733) were significantly higher in the BCBM cohort compared with the primary BC cohort (p < .05 for all comparisons). When using both CGP and PD-L1 IHC, 37.5% (6/16) of patients with TNBC brain metastasis were eligible for atezolizumab based on PD-L1 IHC, and 18.8% (3/16) were eligible for pembrolizumab based on TMB-high status., Conclusion: We found a high prevalence of clinically relevant genomic alterations in patients with BCBM, suggesting that tissue acquisition (surgery) and/or cerebrospinal fluid for CGP in addition to CGP of the primary tumor may be clinically warranted., Implications for Practice: This study found a high prevalence of clinically relevant genomic alterations in patients with breast carcinoma brain metastasis (BCBM), suggesting that tissue acquisition (surgery) and/or cerebrospinal fluid for comprehensive genomic profiling (CGP) in addition to CGP of the primary tumor may be clinically warranted. In addition, this study identified higher positive rates for FDA-approved immunotherapy biomarkers detected by CGP in patients with BCBM, opening a possibility of new on-label treatments. Last, this study noted limited correlation between tumor mutational burden and PD-L1 immunohistochemistry (IHC), which shows the importance of testing patients with triple-negative BCBM for immune checkpoint inhibitor eligibility with both PD-L1 IHC and CGP., (© 2021 AlphaMed Press.)

Detection of somatic genetic drivers is important for risk stratification and treatment selection in pediatric leukemias; however, newly recognized genetic markers may not be detected by routine karyotyping and fluorescence in situ hybridization (FISH). To identify the combination of assays that provides the highest detection rate for clinically significant molecular abnormalities, we tested 160 B- lymphoblastic leukemia (B-ALL) by karyotyping, FISH, chromosomal microarray analysis (CMA) and the custom next-generation sequencing (NGS) panel, OncoKids Ⓡ . In addition, we tested 40 myeloid malignancies with karyotyping, chromosomal microarray analysis (CMA), and OncoKids Ⓡ ; 36/40 myeloid malignancies were also tested with FISH. In B-ALL, individual testing methods had the following diagnostic yields for the key genetic drivers: karyotype 34%; basic FISH panel 45%; FISH panel with IGH and CRLF2 probes 65%; CMA 48%; OncoKids Ⓡ 39%. CMA and OncoKids Ⓡ testing allowed detection of key genetic drivers in 42% of the samples that remained unknown upon testing by conventional methods. In myeloid malignancies, OncoKids Ⓡ had the highest yield for detection of both primary and secondary DNA mutations and RNA fusions. Our data highlights the complementarity between CMA and NGS and conventional cytogenetics/FISH in pediatric leukemia diagnostics. Due to rapid turn-around-time, FISH may be useful as an initial screening method in B-ALL. Our data also suggests NGS testing with a comprehensive panel, despite a longer turnaround time, is a good alternative to karyotyping and FISH in pediatric AML due to its superior detection rate., Competing Interests: Declaration of Competing Interest Matthew C. Hiemenz is an employee of Foundation Medicine, Inc., a wholly owned subsidiary of Roche Holdings Inc. and Roche Finance Ltd., and has an equity interest in an affiliate of these Roche entities., (Copyright © 2021. Published by Elsevier Inc.)

Introduction: Several studies have shown clinical outcomes data that support the use of CD274 (PD-L1 ) copy-number (CN) gains and/or losses as a biomarker for immune checkpoint inhibitor (ICPI). Here, we present the landscape of CD274 CN changes across a large cohort of solid tumor cases and correlate these with PD-L1 protein expression by immunohistochemistry., Methods: We analyzed all cases that underwent comprehensive genomic profiling (CGP) testing at Foundation Medicine between August 2014 and June 2020. CD274 CN changes were correlated with PD-L1 expression in tumor types where there were Food and Drug Administration approved companion diagnostic (CDx) claims and the CDx assay was used to assess PD-L1 expression., Results: In all, 244 584 samples representing 290 solid tumor types were included in the study. Overall, 17.6% (42 983/244 584) had CD274 CN gains (>specimen ploidy), 44.6% (108 970/244 584) were CD274 CN neutral, and 37.9% (92 631/244 584) had CD274 CN loss. Using different CN cut offs to define CD274 positivity resulted in different prevalence estimates: ploidy +1, 17.4% (42 636/244 584); ploidy +2, 6.2% (15 183/244 584); ploidy +3, 2.2% (5375/244 584); ploidy +4, 1.1% (2712/244 584); and ploidy +8, 0.2% (434/244 584). The prevalence of CN changes and CN positivity varied based on tumor type. CD274 CN gains were significantly associated with PD-L1 positivity in NSCLC, urothelial carcinoma, breast carcinoma, cervical carcinoma, esophagus squamous cell carcinoma (SCC) and head and neck SCC (ORs 3.3, 3.0, 2.0, 4.5. 3.8, 8.4, 1.4, respectively; p<0.05) and with microsatellite instability status in only clinically relevant tumor types (gastric adenocarcinoma, colorectal adenocarcinoma, uterine endometrial adenocarcinoma, esophageal adenocarcinoma and gastroesophageal junction adenocarcinoma (OR: 5.2, 1.9, 3.2, 3.7 and 6.5, respectively; p<0.05)). Conversely, CD274 CN changes were not significantly correlated with tumor mutational burden in almost all the tumor types., Conclusion: CD274 CN changes and PD-L1 expression were highly correlated in multiple tumor types. These prevalence data on CD274 CN changes across a large cohort of different solid tumors can be used to design future clinical studies to assess whether CD274 CN changes could be a potential biomarker for ICPI., Competing Interests: Competing interests: All authors of the manuscript are employees of Foundation Medicine, which is a wholly owned subsidiary of Roche and receives stock from Roche., (© Author(s) (or their employer(s)) 2021. Re-use permitted under CC BY-NC. No commercial re-use. See rights and permissions. Published by BMJ.)

Background: Recent large-scale genomic studies have revealed a spectrum of genetic variants associated with specific subtypes of central nervous system (CNS) tumors. The aim of this study was to determine the clinical utility of comprehensive genomic profiling of pediatric, adolescent and young adult (AYA) CNS tumors in a prospective setting, including detection of DNA sequence variants, gene fusions, copy number alterations (CNAs), and loss of heterozygosity., Methods: OncoKids, a comprehensive DNA- and RNA-based next-generation sequencing (NGS) panel, in conjunction with chromosomal microarray analysis (CMA) was employed to detect diagnostic, prognostic, and therapeutic markers. NGS was performed on 222 specimens from 212 patients. Clinical CMA data were analyzed in parallel for 66% (146/222) of cases., Results: NGS demonstrated clinically significant alterations in 66% (147/222) of cases. Diagnostic markers were identified in 62% (138/222) of cases. Prognostic information and targetable genomic alterations were identified in 22% (49/222) and 18% (41/222) of cases, respectively. Diagnostic or prognostic CNAs were revealed by CMA in 69% (101/146) of cases. Importantly, clinically significant CNAs were detected in 57% (34/60) of cases with noncontributory NGS results. Germline cancer predisposition testing was indicated for 27% (57/212) of patients. Follow-up germline testing was performed for 20 patients which confirmed a germline pathogenic/likely pathogenic variant in 9 cases: TP53 (2), NF1 (2), SMARCB1 (1), NF2 (1), MSH6 (1), PMS2 (1), and a patient with 47,XXY Klinefelter syndrome., Conclusions: Our results demonstrate the significant clinical utility of integrating genomic profiling into routine clinical testing for pediatric and AYA patients with CNS tumors., (© The Author(s) 2021. Published by Oxford University Press, the Society for Neuro-Oncology and the European Association of Neuro-Oncology.)

Background: Disorders in the PIK3CA-related overgrowth spectrum because of somatic mosaicism are associated with segmental overgrowth of the body in conjunction with vascular, skeletal, and brain malformations such as hemimegalencephaly. A pathogenic variant may only be detectable in affected tissue and not in peripheral blood or saliva samples; therefore archival tissue may be the only relevant available specimen for testing. Although this is a common approach for cancer testing, it is not typically used for constitutional genetic disorders., Methods: PIK3CA mosaicism was assessed with a custom pediatric oncology next-generation sequencing panel (OncoKids) designed to capture somatic mutations in pediatric malignancies. The panel covers a wide range of targets including PIK3CA and AKT1 hotspots. We used OncoKids on archival formalin-fixed, paraffin-embedded or frozen samples from seven patients with facial hemihypertrophy and lipomas, hemimegalencephaly, or hemihypertrophy with a lymphovascular malformation. The age of the archival tissue examined by next-generation sequencing ranged from two to 13 years (median 5 years). Every patient had clinical manifestations within the PIK3CA-related overgrowth spectrum and had a sample of an affected tissue available for testing from a prior surgical intervention., Results: PIK3CA mosaicism was detected in all seven patients and the mutant allele fraction was lower in the lymphovascular malformation tissues (8% to 11%) than in brain (20% to 32%) and lipomatous (16% to 23%) tissues., Conclusions: Our study highlights the clinical utility of using a robust, oncology-focused next-generation sequencing assay to identify PIK3CA mosaicism in noncancer cases. It is feasible to use archival samples that are more than a decade old to obtain a molecular diagnosis, which can then be used to improve health care management., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Widespread indications for use of molecular diagnostics in various aspects of clinical medicine have driven proliferation of testing. The rapid adoption and continuous technological evolution of molecular diagnostics have often strained the development and maintenance of a functional underlying framework of coding, coverage, and reimbursement policies, thereby presenting challenges to various stakeholders, including molecular professionals, payers, and patients. A multidisciplinary working group convened by the Association for Molecular Pathology Economic Affairs Committee was tasked to describe the complex landscape of molecular pathology economics and highlight opportunities for member engagement. In this article, on the basis of review and synthesis of government regulations and procedures, published payer policy documents, peer-reviewed literature, and expert consensus, the Working Group navigates the ecosystem of molecular pathology economics in terms of stakeholders, coding systems and processes, coverage policy determination, and pricing mechanisms. The composition and interrelatedness of various working groups and committees are emphasized to highlight the functional underpinnings of the system. Molecular professionals must be conversant in the language and complex inner workings of molecular pathology economics to lead successful, viable laboratories and advocate effectively for policy development on their behalf. This overview is provided to be a resource to molecular professionals as they navigate the reimbursement landscape., (Copyright © 2020 Association for Molecular Pathology and American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.)

DICER1 syndrome is a hereditary cancer predisposition syndrome caused by deleterious germline DICER1 mutations. Characteristic "hotspot" somatic mutations of DICER1 have been identified in DICER1-associated tumors. With the exception of genitourinary embryonal rhabdomyosarcoma and anaplastic sarcoma of the kidney, sarcomas are rarely reported in DICER1 syndrome. Herein, we report the clinical, histopathologic, and molecular findings of a germline DICER1-associated ovarian sarcoma in a 5-year-old female, a somatic DICER1-associated metastatic peritoneal sarcoma in a 16-year-old female, and a somatic DICER1-associated primary intracranial sarcoma in a 4-year-old male. A comprehensive review of the literature, including 83 DICER1-associated sarcomas, illustrates an unequivocal histologic pattern mimicking pleuropulmonary blastoma, regardless of the site of origin. The features include undifferentiated small round blue cells, poorly differentiated spindle cells, and large bizarre pleomorphic cells (anaplasia), often with rhabdomyoblastic and/or chondroid differentiation, and rare bone/osteoid formation. This unique heterogeneous histologic pattern should raise suspicion for pathogenic DICER1 mutation(s) warranting a detailed review of the family history and DICER1 mutation analysis. In addition to expanding the phenotypic spectrum of DICER1-associated conditions, identification of pathogenic DICER1 variants facilitates optimized genetic counseling, caregiver education and judicious imaging-based surveillance.

Germline pathogenic variants in CBL are associated with an autosomal dominant RASopathy and an increased risk for malignancies, particularly juvenile myelomonocytic leukemia. Herein, we describe a patient with clinical features of a Noonan-spectrum disorder who developed embryonal rhabdomyosarcoma of the bladder at age two years. Tumor analysis using the OncoKids ® cancer panel revealed a CBL pathogenic variant: NM&#95;005188.3:c.1100A>C (p.Gln367Pro). Sanger sequencing of peripheral blood DNA confirmed a de novo heterozygous germline variant. This is the first report of embryonal rhabdomyosarcoma in association with a germline CBL pathogenic variant, further broadening the CBL cancer predisposition spectrum., (Published by Elsevier Inc.)

The OncoKids panel is an amplification-based next-generation sequencing assay designed to detect diagnostic, prognostic, and therapeutic markers across the spectrum of pediatric malignancies, including leukemias, sarcomas, brain tumors, and embryonal tumors. This panel uses low input amounts of DNA (20 ng) and RNA (20 ng) and is compatible with formalin-fixed, paraffin-embedded and frozen tissue, bone marrow, and peripheral blood. The DNA content of this panel covers the full coding regions of 44 cancer predisposition loci, tumor suppressor genes, and oncogenes; hotspots for mutations in 82 genes; and amplification events in 24 genes. The RNA content includes 1421 targeted gene fusions. We describe the validation of this panel by using a large cohort of 192 unique clinical samples that included a wide range of tumor types and alterations. Robust performance was observed for analytical sensitivity, reproducibility, and limit of detection studies. The results from this study support the use of OncoKids for routine clinical testing of a wide variety of pediatric malignancies., (Copyright © 2018 American Society for Investigative Pathology and the Association for Molecular Pathology. Published by Elsevier Inc. All rights reserved.)

A pediatric patient diagnosed initially with B-lymphoblastic leukemia (B-ALL) relapsed with lineage switch to acute myeloid leukemia (AML) after chimeric antigen receptor T-cell (CAR-T) therapy and hematopoietic stem cell transplant. A TCF3-ZNF384 fusion was identified at diagnosis, persisted through B-ALL relapse, and was also present in the AML relapse cell population. ZNF384-rearrangements define a molecular subtype of B-ALL characterized by a pro-B-cell immunophenotype; furthermore, ZNF384-rearrangements are prevalent in mixed-phenotype acute leukemias. Lineage switch following CAR-T therapy has been described in patients with KMT2A (mixed lineage leukemia) rearrangements, but not previously in any patient with ZNF384 fusion., (© 2018 Wiley Periodicals, Inc.)

As our understanding of the driver mutations necessary for initiation and progression of cancers improves, we gain critical information on how specific molecular profiles of a tumor may predict responsiveness to therapeutic agents or provide knowledge about prognosis. At our institution a tumor genotyping program was established as part of routine clinical care, screening both hematologic and solid tumors for a wide spectrum of mutations using two next-generation sequencing (NGS) panels: a custom, 33 gene hematological malignancies panel for use with peripheral blood and bone marrow, and a commercially produced solid tumor panel for use with formalin-fixed paraffin-embedded tissue that targets 47 genes commonly mutated in cancer. Our workflow includes a pathologist review of the biopsy to ensure there is adequate amount of tumor for the assay followed by customized DNA extraction is performed on the specimen. Quality control of the specimen includes steps for quantity, quality and integrity and only after the extracted DNA passes these metrics an amplicon library is generated and sequenced. The resulting data is analyzed through an in-house bioinformatics pipeline and the variants are reviewed and interpreted for pathogenicity. Here we provide a snapshot of the utility of each panel using two clinical cases to provide insight into how a well-designed NGS workflow can contribute to optimizing clinical outcomes.

Next-generation sequencing (NGS) is a powerful platform for identifying cancer mutations. Routine clinical adoption of NGS requires optimized quality control metrics to ensure accurate results. To assess the robustness of our clinical NGS pipeline, we analyzed the results of 304 solid tumor and hematologic malignancy specimens tested simultaneously by NGS and one or more targeted single-gene tests (EGFR, KRAS, BRAF, NPM1, FLT3, and JAK2). For samples that passed our validated tumor percentage and DNA quality and quantity thresholds, there was perfect concordance between NGS and targeted single-gene tests with the exception of two FLT3 internal tandem duplications that fell below the stringent pre-established reporting threshold but were readily detected by manual inspection. In addition, NGS identified clinically significant mutations not covered by single-gene tests. These findings confirm NGS as a reliable platform for routine clinical use when appropriate quality control metrics, such as tumor percentage and DNA quality cutoffs, are in place. Based on our findings, we suggest a simple workflow that should facilitate adoption of clinical oncologic NGS services at other institutions.

In the United States, recent judicial interpretation of interstate licensure laws has found pathologists guilty of malpractice and, more importantly, the criminal practice of medicine without a license. These judgments against pathologists highlight the need for a timely and comprehensive survey of licensure requirements and laws regulating the interstate practice of pathology. For all 50 states, each state medical practice act and state medical board website was reviewed. In addition, each medical board was directly contacted by electronic mail, telephone, or US registered mail for information regarding specific legislation or guidelines related to the interstate practice of pathology. On the basis of this information, states were grouped according to similarities in legislation and medical board regulations. This comprehensive survey has determined that states define the practice of pathology on the basis of the geographic location of the patient at the time of surgery or phlebotomy. The majority of states (n=32) and the District of Columbia allow for a physician with an out-of-state license to perform limited consultation to a physician with the specific state license. Several states (n=5) prohibit physicians from consultation without a license for the specific state. Overall, these results reveal the heterogeneity of licensure requirements between states. Pathologists who either practice in multiple states, send cases to out-of-state consultants, or serve as consultants themselves should familiarize themselves with the medical licensure laws of the states from which they receive or send cases.

Infiltrating astrocytomas and oligoastrocytomas of low to anaplastic grade (WHO grades II and III), in spite of being associated with a wide range of clinical outcomes, can be difficult to subclassify and grade by the current histopathologic criteria. Unlike oligodendrogliomas and anaplastic oligodendrogliomas that can be identified by the 1p/19q codeletion and the more malignant glioblastomas (WHO grade IV astrocytomas) that can be diagnosed solely based on objective features on routine hematoxylin and eosin sections, no such objective criteria exist for the subclassification of grade II-III astrocytomas and oligoastrocytomas (A+OA II-III). In this study, we evaluated the prognostic and predictive value of the stem cell marker nestin in adult A+OA II-III (n = 50) using immunohistochemistry and computer-assisted analysis on tissue microarrays. In addition, the correlation between nestin mRNA level and total survival was analyzed in the NCI Rembrandt database. The results showed that high nestin expression is a strong adverse prognostic factor for total survival (p = 0.0004). The strength of the correlation was comparable to but independent of the isocitrate dehydrogenase 1/2 (IDH 1/2) mutation status. Histopathological grading and subclassification did not correlate significantly with outcome, although the interpretation of this finding is limited by the fact that grade III tumors were treated more aggressively than grade II tumors. These results suggest that nestin level and IDH 1/2 mutation status are strong prognostic features in A+OA II-III and possibly more helpful for treatment planning than routine histopathological variables such as oligodendroglial component (astrocytoma vs. oligoastrocytoma) and WHO grade (grade II vs. III).

Rearrangements of JAK2 are rare and have been described in various hematological neoplasms. We report a novel JAK2 rearrangement resulting from a t(9;22)(p24;q11.2) in a 14-year-old male with a diagnosis of B lymphoblastic leukemia. He was treated with Children's Oncology Group's protocol (AALL0232) but failed to achieve remission by day 29. He underwent a second induction and entered remission. His clinical course suggested that this JAK2 rearrangement might portend an unfavorable prognosis. This case brings the total number of JAK2 rearranged lymphoblastic leukemia cases in the literature to seven. The molecular genetic and clinicopathologic features of these cases were reviewed., (Copyright © 2010 Elsevier Ltd. All rights reserved.)