Reliable Pan-Cancer Microsatellite Instability Assessment by Using Targeted Next-Generation Sequencing Data

Abstract

Purpose Microsatellite instability (MSI)/mismatch repair (MMR) status is increasingly important in the management of patients with cancer to predict response to immune checkpoint inhibitors. We determined MSI status from large-panel clinical targeted next-generation sequencing (NGS) data across various solid cancer types. Methods The MSI statuses of 12,288 advanced solid cancers consecutively sequenced with Memorial Sloan Kettering-Integrated Mutation Profiling of Actionable Cancer Targets clinical NGS assay were inferred by using MSIsensor, a program that reports the percentage of unstable microsatellites as a score. Cutoff score determination and sensitivity/specificity were based on MSI polymerase chain reaction (PCR) and MMR immunohistochemistry. Results By using an MSIsensor score ≥ 10 to define MSI high (MSI-H), 83 (8%) of 996 colorectal cancers (CRCs) and 42 (16%) of 260 uterine endometrioid cancers (UECs) were MSI-H. Validation against MSI PCR and/or MMR immunohistochemistry performed for 138 (24 MSI-H, 114 microsatellite stable [MSS]) CRCs, and 40 (15 MSI-H, 25 MSS) UECs showed a concordance of 99.4%. MSIsensor also identified 68 MSI-H/MMR-deficient (MMR-D) non-CRC/UECs. Of 9,591 non-CRC/UEC tumors with MSS MSIsensor status, 456 (4.8%) had slightly elevated scores(≥3 and <10) of which 96.6% with available material were confirmed to be MSS by MSI PCR. MSI-H was also detected and confirmed in three non-CRC/UECs with low exonic mutation burden (< 20). MSIsensor correctly scored all 15 polymerase ε ultra-mutated cancers as negative for MSI. Conclusion MSI status can be reliably inferred by MSIsensor from large-panel targeted NGS data. Concurrent MSI testing by NGS is resource efficient, is potentially more sensitive for MMR-D than MSI PCR, and allows identification of MSI-H across various cancers not typically screened, as highlighted by the finding that 35% (68 of 193) of all MSI-H tumors were non-CRC/ UEC.

Accurate Pan-Cancer Molecular Diagnosis of Microsatellite Instability by Single-Molecule Molecular Inversion Probe Capture and High-Throughput Sequencing

“…Notably, this discordant endometrial tumor was also classified as MSS by MSI-PCR, and the consistency between these two assays raises the possibility that the tumor’s molecular signature may legitimately reflect a mutational process that is distinct from MSI (10,23). Regardless of this possibility, our findings reinforce the narrow validity of MSI-PCR only within colorectal cancers (15–17) and demonstrate that informed selection of larger numbers of informative markers by smMIP can recapitulate the pan-cancer performance of exome or genome-scale NGS diagnostic approaches for MSI (10,22,24,25). …”

“…Several groups (10,22,23–25) have alternatively used next-generation sequencing (NGS) to assay MSI by examining dozens to hundreds of repetitive loci that are incidentally sequenced along with regions of interest during targeted gene enrichment or exome sequencing. By virtue of their ability to broadly interrogate the genome, these approaches offer substantial advantages over conventional methods in terms of diagnostic accuracy and applicability across cancer types (10,22,24,25).…”

“…By virtue of their ability to broadly interrogate the genome, these approaches offer substantial advantages over conventional methods in terms of diagnostic accuracy and applicability across cancer types (10,22,24,25). Nevertheless, determination of MSI by NGS currently requires large-scale targeted gene enrichment or exome sequencing data, which is material-limiting for many clinical specimens and results in a high cost of testing.…”

MSIsensor-pro: fast, accurate and matched-normal-sample-free detection of microsatellite instability

“…Recently, several next-generation-sequencing (NGS)-based methods have been developed, which show much better time and cost efficiency and are highly consistent with the gold standards [3][4][5][11][12][13][14][15] . For instance, MSIsensor 11 , an FDA-approved MSI detection solution of MSK-IMPACT 16 , achieved 99.4% concordance and high sensitivity 17 . However, these NGS methods do have limitations, such as requiring matched normal samples as control (sometimes inaccessible), being computational expensive, and being affected by low sequencing depths and low tumor purities 9 .…”

“…The sequencing data of samples with low coverage and low tumor purities are common challenges for robust MSI detections in clinical applications 17 GHz and 32 GB Memory. MSIsensor-pro and MSIsensor required only 4 and 15 minutes, respectively, performances which were significantly faster than mSINGS (94 minutes) and mantis (119 minutes).…”