Tumor mutational burden (TMB) measurements aid in identifying patients who are likely to benefit from immunotherapy; however, there is empirical variability across panel assays and factors ...contributing to this variability have not been comprehensively investigated. Identifying sources of variability can help facilitate comparability across different panel assays, which may aid in broader adoption of panel assays and development of clinical applications.
Twenty-nine tumor samples and 10 human-derived cell lines were processed and distributed to 16 laboratories; each used their own bioinformatics pipelines to calculate TMB and compare to whole exome results. Additionally, theoretical positive percent agreement (PPA) and negative percent agreement (NPA) of TMB were estimated. The impact of filtering pathogenic and germline variants on TMB estimates was assessed. Calibration curves specific to each panel assay were developed to facilitate translation of panel TMB values to whole exome sequencing (WES) TMB values.
Panel sizes >667 Kb are necessary to maintain adequate PPA and NPA for calling TMB high versus TMB low across the range of cut-offs used in practice. Failure to filter out pathogenic variants when estimating panel TMB resulted in overestimating TMB relative to WES for all assays. Filtering out potential germline variants at >0% population minor allele frequency resulted in the strongest correlation to WES TMB. Application of a calibration approach derived from The Cancer Genome Atlas data, tailored to each panel assay, reduced the spread of panel TMB values around the WES TMB as reflected in lower root mean squared error (RMSE) for 26/29 (90%) of the clinical samples.
Estimation of TMB varies across different panels, with panel size, gene content, and bioinformatics pipelines contributing to empirical variability. Statistical calibration can achieve more consistent results across panels and allows for comparison of TMB values across various panel assays. To promote reproducibility and comparability across assays, a software tool was developed and made publicly available.
•Estimation of TMB varies across different panels, with panel size, gene content and bioinformatics pipelines contributing to empirical variability.•Panel sizes greater than 667Kb are necessary to maintain adequate PPA and NPA for calling TMB high versus TMB low across the range of cutoffs used in practice.•Statistical calibration can achieve more consistent results across panels and allows for comparison of TMB values across various panel assays.
Tumor-only sequencing, implemented for the identification of somatic variants, is oftentimes used for the detection of actionable germline variants. We sought to determine whether tumor-only ...sequencing assays are suitable for detection of actionable germline variants, given their importance for the delivery of targeted therapies and risk-reducing measures.
The detection of germline variants affecting moderate- and high-penetrance cancer susceptibility genes (CSGs) by tumor-only sequencing was compared to clinical germline testing in 21 333 cancer patients who underwent tumor and germline testing using the Food and Drug Administration (FDA)-authorized Memorial Sloan Kettering-Integrated Mutation Profiling of Actionable Targets (MSK-IMPACT) assay. Seven homologous recombination deficiency (HRD), two DNA damage response (DDR) and four mismatch repair (MMR) genes, as well as NF1, RB1 and TP53 were included in the analysis. FDA-authorized and New York State Department of Health-approved sequencing methods for germline, tumor/normal and tumor-only sequencing assays and analytical pipelines were employed.
In patients who underwent tumor and germline sequencing, as compared to clinical genetic testing, tumor-only sequencing failed to detect 10.5% of clinically actionable pathogenic germline variants in CSGs, including 18.8%, 12.8% and 7.3% of germline variants in MMR, DDR and HRD genes, respectively. The sensitivity for detection of pathogenic germline variants by tumor-only sequencing was 89.5%. Whilst the vast majority of pathogenic germline exonic single-nucleotide variants (SNVs) and small indels were detected by tumor-only sequencing, large percentages of germline copy number variants, intronic variants and repetitive element insertions were not detected.
Tumor-only sequencing is adequate for the detection of clinically actionable germline variants, particularly for SNVs and small indels; however, a small subset of alterations affecting HRD, DDR and MMR genes may not be detected optimally. Therefore, for high-risk patients with negative tumor-only sequencing results, clinical genetic testing could be considered given the impact of these variants on therapy and genetic counseling.
•In a cohort of >21 000 patients, tumor-only sequencing failed to detect 10.5% of pathogenic germline variants in 16 CSGs.•18.8%, 12.8% and 7.3% of pathogenic germline variants in MMR, DDR and HRD genes were not detected by tumor-only sequencing.•These undetected P/LP variants may affect the use of targeted therapies, preventative measures and familial surveillance.•For patients with negative tumor sequencing results, genetic testing is needed to guide therapy and genetic counseling.
Cases of Clostridiodes difficile infection (CDI) diagnosed after hospital discharge account for a substantial proportion of new infections. It is unclear whether post-discharge infections originate ...from hospital-based transmission.
This was a Retrospective cohort study at a tertiary-care cancer center (non-outbreak setting). For all laboratory-identified cases of CDI in 2015–2016, patients with post-discharge (PD) CDI within eight weeks of their hospital stay were included in the study. Isolates from PD-CDI cases and their CDI-positive unit-based contacts were first genotyped by multilocus sequence typing (MLST). Common strains were further examined by core genome sequencing (CGS) to evaluate transmission links.
Of 173 cases examined by MLST, 50% of PD cases matched previous unit contacts. Next, 34 isolates, including 16 PD cases and their 18-unit contacts were examined by CGS. None were ≤3 single-nucleotide variants apart. Seventy percent of PD cases had in-hospital antibiotic exposure before CDI onset in the community.
Our study results suggest that symptomatic CDI cases are not a substantial source of transmission to PD cases. Frequent antibiotic exposure in post-discharge CDI cases is an important target for surveillance and stewardship efforts.
The utility of cancer whole genome and transcriptome sequencing (cWGTS) in oncology is increasingly recognized. However, implementation of cWGTS is challenged by the need to deliver results within ...clinically relevant timeframes, concerns about assay sensitivity, reporting and prioritization of findings. In a prospective research study we develop a workflow that reports comprehensive cWGTS results in 9 days. Comparison of cWGTS to diagnostic panel assays demonstrates the potential of cWGTS to capture all clinically reported mutations with comparable sensitivity in a single workflow. Benchmarking identifies a minimum of 80× as optimal depth for clinical WGS sequencing. Integration of germline, somatic DNA and RNA-seq data enable data-driven variant prioritization and reporting, with oncogenic findings reported in 54% more patients than standard of care. These results establish key technical considerations for the implementation of cWGTS as an integrated test in clinical oncology.
Kinase fusions are rare and poorly characterized in breast cancer (BC). We aimed to characterize kinase fusions within a large cohort of advanced BC.
A total of 4854 patients with BC were analyzed by ...Memorial Sloan Kettering-Integrated Mutation Profiling of Actionable Cancer Targets (MSK-IMPACT) targeted DNAseq and MSK-Fusion targeted RNAseq during the study time period.
Twenty-seven of 4854 (0.6%) patients harbored fusions: 11 FGFR (five FGFR2, three FGFR3, three FGFR1), five BRAF, four NTRK1, two RET, two ROS1, one ALK, one ERBB2, and one MET. A history of endocrine therapy was present in 15 (56%) of fusion-positive BC; eight of the 15 cases had available pre-treatment samples, of which six were fusion-negative. None of the fusion-positive BC samples harbored ESR1 hotspot mutations. Two patients with acquired LMNA-NTRK1 fusions and metastatic disease received larotrectinib and demonstrated clinical benefit.
Kinase fusions in BC are extremely rare, and appear to be enriched in hormone-resistant, metastatic carcinomas and mutually exclusive with ESR1 mutations. The present study expands the spectrum of genetic alterations activating mitogen-activated protein kinase (MAPK) signaling that can substitute for ESR1 mutations in this setting. Molecular testing at progression after endocrine therapy should include fusion testing, particularly in the absence of ESR1 hotspot alterations, in an effort to identify additional therapeutic options which may provide substantial clinical benefit.
•Kinase fusions in breast cancer are extremely rare.•Kinase fusions in breast cancer appear enriched in ESR1 wild-type metastases after progression on endocrine therapy.•Fusion testing of metastatic breast cancers upon progression after endocrine therapy may provide new therapeutic options.
Inherited mutations in DNA-repair genes were found in nearly 12% of men with metastatic prostate cancer, as compared with 2.7% in an unselected general population.
Carcinoma of the prostate is a ...common cancer with a wide spectrum of clinical behavior that ranges from decades of indolence to rapid metastatic progression and lethality.
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Prostate cancer is also among the most heritable of human cancers, with 57% (95% confidence interval CI, 51 to 63) of the interindividual variation in risk attributed to genetic factors.
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Thus far, genomewide association studies have identified more than 100 common variants that account for approximately 33% of the excess familial prostate cancer risk.
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Mutations in other genes, including
BRCA1, BRCA2, MSH2,
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and
HOXB13,
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account for a small proportion of . . .
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