The challenges and opportunities of offering and integrating training in clinical molecular genetics and clinical cytogenetics: A survey of LGG Fellowship Program Directors.

Purpose: The specialty of Laboratory Genetics and Genomics (LGG) was created in 2017 in an effort to reflect the increasing convergence in technologies and approaches between clinical molecular genetics and clinical cytogenetics. However, there has not yet been any formal evaluation of the merging of these disciplines and the challenges faced by Program Directors (PDs) tasked with ensuring the successful training of laboratory geneticists under the new model. Methods: An electronic multi-question Qualtrics survey was created and was sent to the PD for each of the Accreditation Council for Graduate Medical Education-accredited LGG fellowship programs at the time. The data were collected, and the responses were aggregated for each question. Results: All of the responding PDs had started training at least 1 LGG fellow. PDs noted challenges with funding, staff shortages, molecular/cytogenetics content integration, limited total training time, increased remote work, increased sendout testing, and a lack of prior cytogenetics knowledge among incoming fellows. Conclusion: This survey attempted to assess the challenges that LGG PDs have been facing in offering and integrating clinical molecular genetics and clinical cytogenetics fellowship training. Common challenges between programs were noted, and a set of 6 concluding comments are provided to facilitate future discussion.

Preemptive Pharmacogenetic-Guided Metoprolol Management for Postoperative Atrial Fibrillation in Cardiac Surgery: The Preemptive Pharmacogenetic-Guided Metoprolol Management for Atrial Fibrillation in Cardiac Surgery Pilot Trial.

OBJECTIVES: To test the hypothesis that implementation of a cytochrome P-450 2D6 (CYP2D6) genotype-guided perioperative metoprolol administration will reduce the risk of postoperative atrial fibrillation (AF), the authors conducted the Preemptive Pharmacogenetic-Guided Metoprolol Management for Atrial Fibrillation in Cardiac Surgery pilot study. DESIGN: Clinical pilot trial. SETTING: Single academic center. PARTICIPANTS: Seventy-three cardiac surgery patients. MEASUREMENTS AND MAIN RESULTS: Patients were classified as normal, intermediate, poor, or ultrarapid metabolizers after testing for their CYP2D6 genotype. A clinical decision support tool in the electronic health record advised providers on CYP2D6 genotype-guided metoprolol dosing. Using historical data, the Bayesian method was used to compare the incidence of postoperative AF in patients with altered metabolizer status to the reference incidence. A logistic regression analysis was performed to study the association between the metabolizer status and postoperative AF while controlling for the Multicenter Study of Perioperative Ischemia AF Risk Index. Of the 73 patients, 30% (n = 22) developed postoperative AF; 89% (n = 65) were normal metabolizers; 11% (n = 8) were poor/intermediate metabolizers; and there were no ultrarapid metabolizer patients identified. The estimated rate of postoperative AF in patients with altered metabolizer status was 30% (95% CI 8%-60%), compared with the historical reference incidence (27%). In the risk-adjusted analysis, there was insufficient evidence to conclude that modifying metoprolol dosing based on poor/intermediate metabolizer status was associated significantly with the odds of postoperative AF (odds ratio 0.82, 95% CI 0.15-4.55, p = 0.82). CONCLUSIONS: A CYP2D6 genotype-guided metoprolol management was not associated with a reduction of postoperative AF after cardiac surgery.

Assessments of Somatic Variant Classification Using the Association for Molecular Pathology/American Society of Clinical Oncology/College of American Pathologists Guidelines: A Report from the Association for Molecular Pathology.

To assess the clinical implementation of the 2017 Standards and Guidelines for the Interpretation and Reporting of Sequence Variants in Cancer: A Joint Consensus Recommendation of the Association for Molecular Pathology, American Society of Clinical Oncology, and College of American Pathologists, identify content that may result in classification inconsistencies, and evaluate implementation barriers, an Association for Molecular Pathology Working Group conducted variant interpretation challenges and a guideline implementation survey. A total of 134 participants participated in the variant interpretation challenges, consisting of 11 variants in four cancer cases. Results demonstrate 86% (range, 54% to 94%) of the respondents correctly classified clinically significant variants, variants of uncertain significance, and benign/likely benign variants; however, only 59% (range, 39% to 84%) of responses agreed with the working group's consensus intended responses regarding both tiers and categories of clinical significance. In the implementation survey, 71% (157/220) of respondents have implemented the 2017 guidelines for variant classification and reporting either with or without modifications. Collectively, this study demonstrates that, although they may not yet be optimized, the 2017 guideline recommendations are being adopted for standardized somatic variant classification. The working group identified significant areas for future guideline improvement, including the need for a more granular and comprehensive classification system and education resources to meet the growing needs of both laboratory professionals and medical oncologists.

Impact of Updating Pharmacogenetic Results: Lessons Learned from the PREDICT Program.

Pharmacogenomic (PGx) evidence for selective serotonin reuptake inhibitors (SSRIs) continues to evolve. For sites offering testing, maintaining up-to-date interpretations and implementing new clinical decision support (CDS) driven by existing results creates practical and technical challenges. Vanderbilt University Medical Center initiated panel testing in 2010, added CYP2D6 testing in 2017, and released CDS for SSRIs in 2020. We systematically reinterpreted historic CYP2C19 and CYP2D6 genotypes to update phenotypes to current nomenclature and to launch provider CDS and patient-oriented content for SSRIs. Chart review was conducted to identify and recontact providers caring for patients with current SSRI therapy and new actionable recommendations. A total of 15,619 patients' PGx results were reprocessed. Of the non-deceased patients reprocessed, 21% (n = 3278) resulted in CYP2C19*1/*17 reinterpretations. Among 289 patients with an actionable recommendation and SSRI medication prescription, 31.8% (n = 92) did not necessitate contact of a clinician, while 43.2% (n = 125) resulted in clinician contacted, and for 25% (n = 72) no appropriate clinician was able to be identified. Maintenance of up-to-date interpretations and recommendations for PGx results over the lifetime of a patient requires continuous effort. Reprocessing is a key strategy for maintenance and expansion of PGx content to be periodically considered and implemented.

A Tutorial for Pharmacogenomics Implementation Through End-to-End Clinical Decision Support Based on Ten Years of Experience from PREDICT.

Vanderbilt University Medical Center implemented pharmacogenomics (PGx) testing with the Pharmacogenomic Resource for Enhanced Decisions in Care and Treatment (PREDICT) initiative in 2010. This tutorial reviews the laboratory considerations, technical infrastructure, and programmatic support required to deliver panel-based PGx testing across a large health system with examples and experiences from the first decade of the PREDICT initiative. From the time of inception, automated clinical decision support (CDS) has been a critical capability for delivering PGx results to the point-of-care. Key features of the CDS include human-readable interpretations and clinical guidance that is anticipatory, actionable, and adaptable to changes in the scientific literature. Implementing CDS requires that structured results from the laboratory be encoded in standards-based messages that are securely ingested by electronic health records. Translating results to guidance also requires an informatics infrastructure with multiple components: (1) to manage the interpretation of raw genomic data to "star allele" results to expected phenotype, (2) to define the rules that associate a phenotype with recommended changes to clinical care, and (3) to manage and update the knowledge base. Knowledge base management is key to processing new results with the latest guidelines, and to ensure that historical genomic results can be reinterpreted with revised CDS. We recommend that these components be deployed with institutional authorization, programmatic support, and clinician education to govern the CDS content and policies around delivery.

CYP2D6 Genotype-guided Metoprolol Therapy in Cardiac Surgery Patients: Rationale and Design of the Pharmacogenetic-guided Metoprolol Management for Postoperative Atrial Fibrillation in Cardiac Surgery (PREEMPTIVE) Pilot Study.

OBJECTIVES: The Preemptive Pharmacogenetic-guided Metoprolol Management for Atrial Fibrillation in Cardiac Surgery (PREEMPTIVE) pilot trial aims to use existing institutional resources to develop a process for integrating CYP2D6 pharmacogenetic test results into the patient electronic health record, to develop an evidence-based clinical decision support tool to facilitate CYP2D6 genotype-guided metoprolol administration in the cardiac surgery setting, and to determine the impact of implementing this CYP2D6 genotype-guided integrated approach on the incidence of postoperative atrial fibrillation (AF), provider, and cost outcomes. DESIGN: One-arm Bayesian adaptive design clinical trial. SETTING: Single center, university hospital. PARTICIPANTS: The authors will screen (including CYP2D6 genotype) up to 600 (264 ± 144 expected under the adaptive design) cardiac surgery patients, and enroll up to 200 (88 ± 48 expected) poor, intermediate, and ultrarapid CYP2D6 metabolizers over a period of 2 years at a tertiary academic center. INTERVENTIONS: All consented and enrolled patients will receive the intervention of CYP2D6 genotype-guided metoprolol management based on CYP2D6 phenotype classified as a poor, intermediate, extensive (normal), or ultrarapid metabolizer. MEASUREMENTS AND MAIN RESULTS: The primary outcome will be the incidence of postoperative AF. Secondary outcomes relating to rates of CYP2D6 genotype-guided prescription changes, costs, lengths of stay, and implementation metrics also will be investigated. CONCLUSIONS: The PREEMPTIVE pilot study is the first perioperative pilot trial to provide essential information for the design of a future, large-scale trial comparing CYP2D6 genotype-guided metoprolol management with a nontailored strategy in terms of managing AF. In addition, secondary outcomes regarding implementation, clinical benefit, safety, and cost-effectiveness in patients undergoing cardiac surgery will be examined.

Clinicopathologic correlates of MYD88 L265P mutation and programmed cell death (PD-1) pathway in primary central nervous system lymphoma.

Primary central nervous system lymphoma (PCNSL) patients have a poorer prognosis than systemic lymphoma. Gain-of-function MYD88 c.794T > C (p. L265P) mutation and programed cell death-1 (PD-1) pathway alterations are potential targetable pathways. Our study objective was to determine the clinicopathologic correlates of MYD88 mutation and PD-1 alterations in PCNSL and the impact of Epstein-Barr virus (EBV) infection. We studied 53 cases including 13 EBV-associated (EBVpos) PCNSL, 49% harbored MYD88 mutation, none seen in EBVpos PCNSL. MYD88 protein expression did not correlate with MYD88 mutation. T-cell and macrophage infiltration was common. All PD-L1-positive tumors were EBVpos. Two PD-L1 positive tumors showed 9p24.1/PD-L1 locus alterations by Fluorescence In Situ Hybridization. T cells and macrophages expressed PD-1 and/or PD-L1 in 98% and 83% cases, respectively. MYD88 mutation or protein expression and PD-1 or PD-L1 expression did not predict outcome. We hypothesize that EBVpos PCNSL has a distinct activation mechanism, independent of genetic alterations.

The impact of variant classification on the clinical management of hereditary cancer syndromes.

PURPOSE: The reclassification of genetic variants poses a significant challenge for laboratories and clinicians. Variant review has resulted in the reclassification of variants of unknown significance as well as the reclassification of previously established pathogenic and likely pathogenic variants. These reclassifications have the potential to alter the clinical management of patients with hereditary cancer syndromes. METHODS: Results were reviewed for 1694 patients seen for hereditary cancer evaluation between August 2012 and May 2017 to determine the frequency and types of variant reclassification. Patients with reclassifications with high potential for impact were monitored for alterations in organ surveillance, prophylactic surgery, and cascade testing. RESULTS: One hundred forty-two variants were reclassified representing 124/1694 (7.3%) patients; 11.3% of reclassifications (16/142) had a high potential for clinical impact with 94% (15/16) altering clinical management of patients with 56% (9/16) changing multiple areas of management. CONCLUSION: While reclassifications are rare, the impact on clinical management is profound. In many cases, patients with downgraded pathogenic/likely pathogenic variants had years of unnecessary organ surveillance and underwent unneeded surgical intervention. In addition, cascade testing misidentified those at risk for developing cancers, thereby altering the management across generations. The frequency and types of alterations to clinical management highlight the need for timely variant reclassification.

Deciphering Elevated Microsatellite Alterations at Selected Tetra/Pentanucleotide Repeats, Microsatellite Instability, and Loss of Heterozygosity in Colorectal Cancers.

Elevated microsatellite alterations at selected tetranucleotide repeats (EMAST) are common in colorectal cancers (CRCs). The association between EMAST and classic mono/dinucleotide microsatellite instability (MSI) is unknown. We assessed the stability of 13 tetranucleotide and three pentanucleotide repeat markers in tumor and normal tissue from 22 MSI-high and 107 microsatellite-stable CRC samples. When present, instability was observed at tetra/pentanucleotide repeats and was defined as elevated microsatellite alterations at selected tetra/pentanucleotide repeats-high (EMASTP-H; ≥30% instability), -low (EMASTP-L; <30% instability), or -stable (EMASTP-S). EMASTP instability, including high and low, was observed in 50 of 123 CRCs (41%), including all MSI-high tumors and 28 of 101 microsatellite-stable tumors (28%). MSI-high CRCs were more likely to be EMASTP-H compared with microsatellite-stable tumors with EMASTP instability. Tetranucleotide markers VWA and D13S317 were the two most frequently altered loci. Loss of heterozygosity was more common in EMASTP-L/S than in EMASTP-H CRCs. Frequencies of loss of heterozygosity at three loci were different between EMASTP-L and EMASTP-S tumors. In addition, right-sided tumor site, large tumor size, high tumor grade, and the presence of Crohn-like reaction were significantly associated with EMASTP-H CRCs. However, there were no differences in clinicopathologic features between EMASTP-L and EMASTP-S tumors. In summary, more CRCs exhibited genomic instability as EMASTP than as MSI. EMASTP instability may prove to be an important prognostic/therapeutic indicator in CRCs.

Clinical prognostic value of the isocitrate dehydrogenase 1 single-nucleotide polymorphism rs11554137 in glioblastoma.

The presence of the single-nucleotide polymorphism (SNP) rs11554137:C>T in the IDH1 gene is associated with a significantly lower survival in acute myeloid leukemia patients. The impact of its presence in glioblastoma on patient survival is unclear. We retrospectively reviewed 171 adult (> 18 years of age) patients treated at a single, tertiary academic center for supratentorial glioblastoma (WHO grade IV) between 2013 and 2017. We conducted Kaplan-Meier overall and progression free survival analyses based on the IDH1 and IDH2 gene status of patients' glioblastoma (IDH wild type, mutant, and IDH1 rs11554137:C>T SNP). Multivariate Cox survival analyses were conducted accounting for age at diagnosis, preoperative Karnofsky performance status score, treatment (extent of resection, postoperative radiotherapy, and temozolomide), IDH gene variant, and MGMT promoter methylation status. Presence of rs11554137:C>T SNP in glioblastoma samples did not correlate with presence of IDH1 mutation. Patients with rs11554137:C>T SNP did not have histories of prior lower-grade gliomas. Patients with IDH mutant glioblastoma had a distinctly higher survival profile than both rs11554137:C>T SNP and IDH wild type glioblastomas. No survival difference was noted between patients with glioblastoma harboring the SNP and patients with IDH wild type glioblastoma. In this study, clinical prognostication in glioblastoma patients was largely dependent on the classification of IDH mutant and wild type glioblastoma, and not on the presence of IDH1 rs11554137:C>T SNP in the tumor.

Authors' Reply.

Authors' Reply to the Letter to the Editor by Montgomery et al (Identification of Germline Variants in Tumor Genomic Sequencing Analysis. J Mol Diagn 2017, 19:XXXX-XXXX).

Primary Carcinoid Tumor of the Renal Pelvis Arising From Intestinal Metaplasia: An Unusual Histogenetic Pathway?

OBJECTIVES: Primary carcinoid tumor of the renal pelvis is a rare neoplasm with few cases reported in the literature. Here we present the clinical and histopathologic findings of a primary carcinoid tumor arising in the left renal pelvis of a horseshoe kidney in a 61-year-old female patient. MATERIALS AND METHODS: Pathologic features were evaluated with standard hematoxylin and eosin sections and immunohistochemical studies. A literature review was performed to place our case in context to previous reports. RESULTS: The tumor was associated with intestinal metaplasia with high-grade dysplasia and neuroendocrine hyperplasia. Molecular testing for microsatellite instability and loss of heterozygosity were negative. CONCLUSIONS: This report portrays a unique presentation of carcinoid tumor arising from intestinal metaplasia of the pelvic urothelium, and supports its histogenesis from urothelial intestinal metaplasia and neuroendocrine hyperplasia.

Differences in Microsatellite Instability Profiles between Endometrioid and Colorectal Cancers: A Potential Cause for False-Negative Results?

Colorectal (CRCs) and endometrioid (EMCs) cancers in patients with Lynch syndrome exhibit microsatellite instability (MSI) detected by PCR or immunohistochemistry (IHC). While both assays are equally sensitive for CRCs, some suggest that PCR has a higher false-negative rate than IHC in EMCs. We assessed the MSI profiles of 91 EMC and 311 CRC specimens using five mononucleotide repeat markers: BAT25, BAT26, NR21, NR24, and MONO27. EMCs with high MSI (MSI-H) showed a mean left shift of 3 nucleotides (nt), which was significantly different from 6 nt in CRCs. A shift of 1 nt was observed in multiple markers in 76% of MSI-H EMCs, whereas only 12% of MSI-H CRCs displayed a 1-nt shift in one of five markers. IHC against four mismatch repair proteins was performed in 78 EMCs. Loss of staining in one or more proteins was detected in 18 of 19 tumors that were MSI-H by PCR. When EMC tumor cell burden was diluted to <30%, MSI-H was no longer observed in two of three EMCs with a mean nucleotide shift of 1 nt. These results indicate that EMC and CRC MSI profiles are different and that caution should be exercised when interpreting the results, as subtle, 1-nt changes may be missed. These findings provide a potential cause of previously reported discordant MSI and IHC results in EMCs.

Standards and Guidelines for the Interpretation and Reporting of Sequence Variants in Cancer: A Joint Consensus Recommendation of the Association for Molecular Pathology, American Society of Clinical Oncology, and College of American Pathologists.

Widespread clinical laboratory implementation of next-generation sequencing-based cancer testing has highlighted the importance and potential benefits of standardizing the interpretation and reporting of molecular results among laboratories. A multidisciplinary working group tasked to assess the current status of next-generation sequencing-based cancer testing and establish standardized consensus classification, annotation, interpretation, and reporting conventions for somatic sequence variants was convened by the Association for Molecular Pathology with liaison representation from the American College of Medical Genetics and Genomics, American Society of Clinical Oncology, and College of American Pathologists. On the basis of the results of professional surveys, literature review, and the Working Group's subject matter expert consensus, a four-tiered system to categorize somatic sequence variations based on their clinical significances is proposed: tier I, variants with strong clinical significance; tier II, variants with potential clinical significance; tier III, variants of unknown clinical significance; and tier IV, variants deemed benign or likely benign. Cancer genomics is a rapidly evolving field; therefore, the clinical significance of any variant in therapy, diagnosis, or prognosis should be reevaluated on an ongoing basis. Reporting of genomic variants should follow standard nomenclature, with testing method and limitations clearly described. Clinical recommendations should be concise and correlate with histological and clinical findings.

Mutation profiles of synchronous colorectal cancers from a patient with Lynch syndrome suggest distinct oncogenic pathways.

Patients with Lynch syndrome often present with multiple synchronous or metachronous colorectal cancers (CRCs). The presence of multiple CRCs with distinct genetic profiles and driver mutations could complicate treatment as each cancer may respond differently to therapy. Studies of sporadic CRCs suggested that synchronous tumors have distinct etiologies, but could not rule out differences in genetic background. The presence of multiple cancers in a patient with a predisposing mutation provides an opportunity to profile synchronous cancers in the same genetic background. Here, we describe the case of a patient with Lynch syndrome that presented with six synchronous CRCs. Microsatellite instability (MSI) and genomic profiling indicated that each lesion had a unique pattern of instability and a distinct profile of affected genes. These findings support the idea that in Lynch syndrome, synchronous CRCs can develop in parallel with distinct mutation profiles and that these differences may inform treatment decisions.

Characterization of breast cancers with PI3K mutations in an academic practice setting using SNaPshot profiling.

Mutations in the PIK3CA gene are common in breast cancer and represent a clinically useful therapeutic target. Several larger, population-based studies have shown a positive prognostic significance associated with these mutations. This study aims to further identify characteristics of patients harboring PIK3CA mutations while evaluating the clinical impact of genomic testing for these mutations. Tumors from 312 patients at Vanderbilt-Ingram Cancer Center were analyzed for PIK3CA mutations using a multiplex screening assay (SNaPshot). Mutation rates, receptor status, histopathologic characteristics, and time to recurrence were assessed. The number of patients participating in clinical trials, specifically trials relating to the PIK3CA mutation, was examined. Statistically significant differences between wild-type and mutated tumors were determined using the Wilcoxon, Pearson, and Fischer exact tests. The PIK3CA mutation was found in 25 % of tumors tested. Patients with PIK3CA mutations were significantly more likely to express hormone receptors, be of lower combined histological grade, and have a reduced time to recurrence. Patients found to have a PIK3CA mutation were significantly more likely to enter a PIK3CA-specific clinical trial. In addition to confirming previously established positive prognostic characteristics of tumors harboring PIK3CA mutations, this study demonstrates the feasibility and utility of mutation profiling in a clinical setting. PIK3CA mutation testing impacted treatment and resulted in more patients entering mutation-specific clinical trials.

Endomyometriosis ("Uterus-like mass") in an XY Male: Case Report With Molecular Confirmation and Literature Review.

Male endometriosis and endomyometriosis (also termed "uterus-like mass") are 2 unusual manifestations of endometriosis. We report a case of male endomyometriosis with immunohistochemical and molecular confirmation. A 52-year-old man presented with stabbing pelvic pain. Computed tomography scan showed a mass in the right inguinal area, at the site of prior hernia repair. The lesion was tubular in shape, with a thick muscular wall and a central blood-filled lumen. Microscopically, the tissue showed layers of concentric smooth muscle, with endometrial glands and stroma lining the lumen. Many theories have been proposed regarding the etiology of both endomyometriosis and male endometriosis, including remnant rests of Müllerian tissue and metaplasia. Cases of male endometriosis typically have been linked to estrogen therapy for prostate cancer. Our patient had a history of cirrhosis and took spironolactone, possibly leading to an altered hormonal state that interacted with a reactive/metaplastic process at a site of prior surgery.

Beyond histology: translating tumor genotypes into clinically effective targeted therapies.

Increased understanding of intertumoral heterogeneity at the genomic level has led to significant advancements in the treatment of solid tumors. Functional genomic alterations conferring sensitivity to targeted therapies can take many forms, and appropriate methods and tools are needed to detect these alterations. This review provides an update on genetic variability among solid tumors of similar histologic classification, using non-small cell lung cancer and melanoma as examples. We also discuss relevant technological platforms for discovery and diagnosis of clinically actionable variants and highlight the implications of specific genomic alterations for response to targeted therapy.