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Both microsatellite length and sequence context determine frameshift mutation rates in defective DNA mismatch repair.

ABSTRACT: It is generally accepted that longer microsatellites mutate more frequently in defective DNA mismatch repair (MMR) than shorter microsatellites. Indeed, we have previously observed that the A10 microsatellite of transforming growth factor beta type II receptor (TGFBR2) frameshifts -1 bp at a faster rate than the A8 microsatellite of activin type II receptor (ACVR2), although both genes become frameshift-mutated in >80% of MMR-defective colorectal cancers. To experimentally determine the effect of microsatellite length upon frameshift mutation in gene-specific sequence contexts, we altered the microsatellite length within TGFBR2 exon 3 and ACVR2 exon 10, generating A7, A10 and A13 constructs. These constructs were cloned 1 bp out of frame of EGFP, allowing a -1 bp frameshift to drive EGFP expression, and stably transfected into MMR-deficient cells. Subsequent non-fluorescent cells were sorted, cultured for 7-35 days and harvested for EGFP analysis and DNA sequencing. Longer microsatellites within TGFBR2 and ACVR2 showed significantly higher mutation rates than shorter ones, with TGFBR2 A13, A10 and A7 frameshifts measured at 22.38x10(-4), 2.17x10(-4) and 0.13x10(-4), respectively. Surprisingly, shorter ACVR2 constructs showed three times higher mutation rates at A7 and A10 lengths than identical length TGFBR2 constructs but comparably lower at the A13 length, suggesting influences from both microsatellite length as well as the sequence context. Furthermore, the TGFBR2 A13 construct mutated into 33% A11 sequences (-2 bp) in addition to expected A12 (-1 bp), indicating that this construct undergoes continual subsequent frameshift mutation. These data demonstrate experimentally that both the length of a mononucleotide microsatellite and its sequence context influence mutation rate in defective DNA MMR.

SUBMITTER: Chung H

PROVIDER: S-EPMC2912546 | biostudies-literature | 2010 Jul

REPOSITORIES: biostudies-literature

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Both microsatellite length and sequence context determine frameshift mutation rates in defective DNA mismatch repair.

Chung Heekyung H Lopez Claudia G CG Holmstrom Joy J Young Dennis J DJ Lai Jenny F JF Ream-Robinson Deena D Carethers John M JM

Human molecular genetics 20100423 13

It is generally accepted that longer microsatellites mutate more frequently in defective DNA mismatch repair (MMR) than shorter microsatellites. Indeed, we have previously observed that the A10 microsatellite of transforming growth factor beta type II receptor (TGFBR2) frameshifts -1 bp at a faster rate than the A8 microsatellite of activin type II receptor (ACVR2), although both genes become frameshift-mutated in >80% of MMR-defective colorectal cancers. To experimentally determine the effect o ...[more]

PMID: 20418486

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Project description:BackgroundMonoclonal antibodies targeting programmed death ligand 1 (PD-L1) signaling currently approved for defective mismatch repair (dMMR)/microsatellite instability high (MSI-H) tumors must be delivered by intravenous infusion. Envafolimab, a humanized single-domain anti-PD-L1 antibody fused to an Fc fragment, represents a potential advance because it can be conveniently administered subcutaneously.MethodsThis open-label, single-arm, phase 2 study evaluated the efficacy and safety of envafolimab in patients with previously treated advanced dMMR/MSI-H tumors from 25 clinical sites across China. Adults with histologically confirmed locally advanced or metastatic malignant dMMR/MSI-H solid tumors received weekly 150 mg subcutaneous envafolimab injections in a 28-day treatment cycle. The primary efficacy endpoint was the objective response rate (assessed by a blinded independent review committee). Secondary efficacy outcomes were disease control rate, duration of response, progression-free survival, and overall survival.ResultsOne hundred and three patients (65 with colorectal cancer, 18 with gastric cancer, and 20 with other solid tumors) were enrolled. Median follow-up was 11.5 months. The objective response rate was 42.7% (95% confidence interval [CI] 33.0-52.8), and the disease control rate was 66.0% (95% CI 56.0-75.1). Median duration of response was not reached; the duration of response rate at 12 months was 92.2% (95% CI 77.5-97.4). Median progression-free survival was 11.1 months (95% CI 5.5 to not evaluable). Overall survival at 12 months was 74.6% (95% CI 64.7-82.1). Sixteen patients (16%) had at least one grade 3 or 4 related treatment-emergent adverse event. No grade 5 treatment-emergent adverse events related to envafolimab were reported. Injection site reactions, all grade 1-2, were reported in nine patients (9%), but there were no infusion reactions. Eight patients (8%) had grade 3 or 4 immune-related adverse events.ConclusionsThis is the first pivotal phase 2 study to examine the efficacy and safety of a single-domain immune checkpoint antibody in the treatment of cancer. Envafolimab was effective and had acceptable safety in the treatment of previously treated advanced dMMR/MSI-H solid tumors. As the first single-domain PD-L1-targeting antibody administered by rapid subcutaneous injection, envafolimab has the potential to be a significant advance in the treatment of cancer. Trial registration ClinicalTrials.gov, NCT03667170. Registered 10 September 2018-Retrospectively registered, https://clinicaltrials.gov/ct2/show/NCT03667170 .

Project description:BackgroundGenomic biomarkers that predict response to anti-PD1 therapy in prostate cancer are needed. Frameshift mutations are predicted to generate more neoantigens than missense mutations; therefore, we hypothesized that the number or proportion of tumor frameshift mutations would correlate with response to anti-PD1 therapy in prostate cancer.MethodsTo enrich for response to anti-PD1 therapy, we assembled a multicenter cohort of 65 men with mismatch repair-deficient (dMMR) prostate cancer. Patient characteristics and outcomes were determined by retrospective chart review. Clinical somatic DNA sequencing was used to determine tumor mutational burden (TMB), frameshift mutation burden, and frameshift mutation proportion (FSP), which were correlated to outcomes on anti-PD1 treatment. We subsequently used data from a clinical trial of pembrolizumab in patients with nonprostatic dMMR cancers of various histologies as a biomarker validation cohort.ResultsNineteen of 65 patients with dMMR metastatic castration-resistant prostate cancer were treated with anti-PD1 therapy. The PSA50 response rate was 65%, and the median progression-free survival (PFS) was 24 (95% confidence interval 16-54) weeks. Tumor FSP, more than overall TMB, correlated most strongly with prolonged PFS and overall survival (OS) on anti-PD1 treatment and with density of CD8+ tumor-infiltrating lymphocytes. High FSP similarly identified patients with longer PFS as well as OS on anti-PD1 therapy in a validation cohort.ConclusionTumor FSP correlated with prolonged efficacy of anti-PD1 treatment among patients with dMMR cancers and may represent a new biomarker of immune checkpoint inhibitor sensitivity.Implications for practiceGiven the modest efficacy of immune checkpoint inhibition (ICI) in unselected patients with advanced prostate cancer, biomarkers of ICI sensitivity are needed. To facilitate biomarker discovery, a cohort of patients with DNA mismatch repair-deficient (dMMR) prostate cancer was assembled, as these patients are enriched for responses to ICI. A high response rate to anti-PD1 therapy in these patients was observed; however, these responses were not durable in most patients. Notably, tumor frameshift mutation proportion (FSP) was identified as a novel biomarker that was associated with prolonged response to anti-PD1 therapy in this cohort. This finding was validated in a separate cohort of patients with nonprostatic dMMR cancers of various primary histologies. This works suggests that FSP predicts response to anti-PD1 therapy in dMMR cancers, which should be validated prospectively in larger independent cohorts.

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Both microsatellite length and sequence context determine frameshift mutation rates in defective DNA mismatch repair.

Publications

Both microsatellite length and sequence context determine frameshift mutation rates in defective DNA mismatch repair.

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