MSI-H Tumors Research Articles

Relationship between the number of positive MSI markers and the efficacy of NIVO+IPI therapy in MSI-H gastric cancer: A subgroup analysis of NO LIMIT study (WJOG13320G/CA209-7W7).

386 Background: Although high instability of microsatellite lesion (MSI-H) is an established biomarker for response to immune checkpoint inhibitors, the relationship between the number of MSI markers and the efficacy of immune checkpoint inhibitors in MSI-H tumors has not been well elucidated. NO LIMIT (WJOG13320G/CA209-7W7) is a phase II trial of nivolumab plus low-dose ipilimumab (NIVO+IPI) for MSI-H advanced gastric or esophagogastric junction cancer (GC) in the first-line setting.1 Here, we investigated the relationship between the number of positive MSI markers and the efficacy of NIVO+IPI in NO LIMIT study. Methods: Eligible patients were unresectable advanced, recurrent, or metastatic GC with confirmed as MSI-H with the MSI-IVD Kit (FALCO), where two or more of the five MSI markers are defined as positive. Nivolumab (240 mg) biweekly and ipilimumab (1 mg/kg) every six weeks were given until disease progression or unacceptable toxicity. The primary endpoint was the objective response rate (ORR) assessed by a blinded independent central review. Secondary endpoints included progression-free survival (PFS), overall survival (OS), safety, and the association between number of positive MSI markers and clinical efficacy (ORR, PFS, OS). The analysis was performed 18 weeks after the first dose in the last patient, with a median follow-up of 9 months (range, 4.0-18.0). Results: The breakdown of MSI markers in the 29 patients enrolled in NO LIMIT was as follows: BAT-26 (26/26, 100%), NR-21 (25/29, 86.2%), BAT-25 (25/29, 86.2%), MONO-27 (29/29, 100%), NR-24 (27/29, 89.4%). Since most of the cases were positive for all five of them (the number of cases with 2, 3, 4, and 5 positive markers was 2, 2, 1, and 24, respectively), we dichotomized cases into group A (all five positive) and group B (the others). Response rates in groups A vs. B were 62.5% (15/24) vs. 60.0% (3/5), respectively. There was no difference in PFS or OS between the two groups. Conclusions: The number of MSI markers is not related to the efficacy of NIVO+IPI in NO LIMIT study. Further follow-up is needed to confirm the relationship between survival. 1. Muro K, et al. ESMO Congress 2023. Clinical trial information: jRCT2080225304 . [Table: see text]

Trends in utilization of MSI testing and reporting among patients with gastrointestinal cancers: A review of NCDB Database.

221 Background: MSI-H tumors have a different disease biology and clinical outcomes compared to MSS/MSI-low tumors. The frequency and pattern for MSI testing and reporting is unknown. The aim of this study was to characterize the trends of MSI testing and reporting among GI cancers. Methods: From the National Cancer Database, we identified all GI cancer patients from 2010 to 2016 to determine the number of patients where MSS/MSI low or MSI-H data were available. We describe the basic tumor and disease characteristics and treatment utilization for the two groups. Results: Between 2010-2016, MSI testing was done in 140,602 patients in colon, small intestine and rectal cancers among all GI cancers. MSI testing increased significantly during this study period (11,400 in 2010; 15,021 in 2011; 17,968 in 2012; 20,810 in 2013; 21,973 in 2014; 25,012 in 2015; 28,418 in 2016). We identified a total of 120,349 patients with MSI status reported, among these three tumor types, most commonly among colon followed by rectal and then small intestinal cancers. Overall 111,694 (92.8%) were MSS/MSI-low and 8,655 were MSI-H (7.2%). Among patients with reported MSI status, 52.1% were males and 47.9% were females. The mean age was 64.4 years. Most patients were whites (87.6%) and had insurance (96.8%), no existing comorbidities (70.4%), and grade 2 (70.8%) and AJCC Clinical Stage 4 (26.1%) GI cancer. A higher proportion of GI cancer patients with MSS/MSI-low utilized chemotherapy as first course compared to those with MSI high (48.2% vs 34.0%) whereas a lower proportion of GI cancer patients with MSS/MSI-low underwent a surgical procedure compared to those with MSI high (94.7% vs 97.1%). MSI status was reported most frequently in comprehensive community cancer program (40.3%) and academic center (34.8%), followed by integrated network cancer program (16.8%) and Community Cancer program (8.1%). Conclusions: The utilization of MSI testing has increased significantly between 2010 and 2016. Among all GI cancer patients, MSI status was most frequently reported in colon, rectum and small intestinal cancers. Comprehensive community cancer centers and academic centers reported MSI status more frequently than Integrated Network Cancer program and Community Cancer program.

Perceptions of cancer prevention options for Lynch syndrome (LS) in a multi-national convenience sample.

77 Background: LS is among the most common cancer (CA) syndromes affecting 1:280 individuals and portends high lifetime CA risk. Clinical trials are currently evaluating whether frameshift peptide vaccination (FSPVAX) could reduce LS CA risk. FSPVAX enhances immune response to diverse FSP neoantigens in MSI-H tumors and has shown promising results in an MSH2-mutant mouse model and metastatic MSI-H CRC. However, negative public attitudes towards vaccination and evidence suggesting low uptake of ASA chemoprevention (CP) among patients with LS could undermine success of this novel therapy. Here, we benchmark attitudes towards FSPVAX for CA prevention against colonoscopy (COLO) and aspirin or NSAID chemoprevention (CP). Methods: The PREVENTLynchII survey (IRB 20-8014) was distributed electronically to 1) participants in the Fox Chase Cancer Center (FCCC) Risk Registry and 2) members of two online LS support groups. Patients were invited to complete the one-time survey after providing informed consent. Demographic and personal CA history were collected. Attitudes were measured on 9-point scales indicating low vs high agreement with belief statements: e.g. “Colonoscopy is a convenient way to lower my risk of LS-related CA.” Results: Overall 296 participants at FCCC, nationally, and internationally completed the survey. Median age was 52.6 years [23-78], 84% female, and 95% White. Affected genes included MLH1 (20.3%), MSH2/EPCAM (35.5%), MSH6 (28.7%), and PMS2 (15.5%). Use of CP was uncommon (32.1% and 5.1% respectively). Stratified analyses of COLO perceptions by sex, age, and personal history of CA showed no differences. Interestingly, US pts viewed ASA as more convenient than non-US, and with lower risk of side effects (p=0.007), but were no more likely to use or recommend CP. Compared to COLO, FSPVAX was overall viewed as more convenient, but side effect concerns were higher, and perceptions of efficacy were lower (all p<0.001), while compared to CP, FSPVAX was less convenient and fostered greater concerns about side effects, but was viewed as having higher potential preventive efficacy (all p<0.001). FSPVAX perceptions showed high variability: pts >50 (p=0.003) and CA pts (p=0.01) saw FSP as more convenient (p=0.003) than younger and unaffected pts. CA pts reported higher reassurance than unaffected (p=0.04), while those >50 were more likely to participate in an FSPVAX study (p=0.04). US pts perceived more side effect risk of FSPVAX than non-US (p=0.003). Conclusions: Concerns about ASA CP and FSPVAX convenience, side effects, and efficacy are common among US and international patients with LS. [Table: see text]

A Panel-Based Mutational Signature of Mismatch Repair Deficiency is Associated With Durable Response to Pembrolizumab in Metastatic Castration-Resistant Prostate Cancer

Introduction/BackgroundImmune checkpoint inhibitors (ICIs) have limited efficacy in prostate cancer (PCa). Better biomarkers are needed to predict responses to ICIs. We sought to demonstrate that a panel-based mutational signature identifies mismatch repair (MMR) deficient (MMRd) PCa and is a biomarker of response to pembrolizumab. Patients and MethodsClinico-genomic data was obtained for 2664 patients with PCa sequenced at Dana-Farber Cancer Institute (DFCI) and Memorial Sloan Kettering (MSK). Clinical outcomes were collected for patients with metastatic castration-resistant PCa (mCRPC) treated with pembrolizumab at DFCI. SigMA was used to characterize tumors as MMRd or MMR proficient (MMRp). The concordance between MMRd with microsatellite instability (MSI-H) was assessed. Radiographic progression-free survival (rPFS) and overall survival (OS) were collected for patients treated with pembrolizumab. Event-time distributions were estimated using Kaplan-Meier methodology. ResultsAcross both cohorts, 100% (DFCI: 12/12; MSK: 43/43) of MSI-H tumors were MMRd. However, 14% (2/14) and 9.1% (6/66) of MMRd tumors in the DFCI and MSK cohorts respectively were microsatellite stable (MSS), and 26% (17/66) were MSI-indeterminate in the MSK cohort. Among patients treated with pembrolizumab, those with MMRd (n = 5) versus MMRp (n = 14) mCRPC experienced markedly improved rPFS (HR = 0.088, 95% CI: 0.011-0.70; P = .0064) and OS (HR = 0.11, 95% CI: 0.014-0.80; P = .010) from start of treatment. Four patients with MMRd experienced remissions of >= 2.5 years. ConclusionSigMA detects additional cases of MMRd as compared to MSI testing in PCa and identifies patients likely to experience durable response to pembrolizumab.

Molecular differences with therapeutic implications in early-onset compared to average-onset cholangiocarcinoma.

536 Background: Early-onset cholangiocarcinoma (eoCCA) is among early-onset cancers with the fastest rising rates, yet little is known about its biology. We sought to compare the molecular characteristics of eoCCA with average-onset CCA (aoCCA) with an age cut-off of 50 years, utilizing a real-world multi-omics dataset. Methods: The study comprised patients whose tumors underwent molecular analysis at Caris Life Sciences (Phoenix, AZ) using whole exome and whole transcriptome analyses. Patients were categorized by age as eoCCA defined as <50 years and aoCCA >50 years. Gene expression profiles were analyzed for transcriptional signatures predictive of immunotherapy response including the T-cell inflamed score (TIS) and interferon-gamma (IFG) score. P values (adjusted for multiple testing) were considered significant at False Discovery Rate (FDR) P<0.05 for molecular comparisons and FDR P<0.25 for Gene Set Enrichment Analysis (GSEA). Insurance claims data was used for survival comparison using Kaplan-Meier estimates. Results: The study included 5587 patients - 453 patients with eoCCA and 5134 with aoCCA (Table). Of targetable mutations ( FGFR2, IDH1, IDH2, ERBB2, BRCA, BRAF), FGFR2 fusion was significantly more prevalent in eoCCA (15.7% vs 5.9% in aoCCA, FDR P<0.001). Rates trended higher in eoCCAfor MSI-H tumors (4.1% vs 2.4%), and high tumor mutational burden (6.1% vs 5.1%) but not significantly different (FDR P=1). The IFG score (Fold Change FC: 1.1, FDR P=0.01) and TIS (FC:17.3, FDR P=0.03) were significantly higher in aoCCA vs eoCCA. On GSEA, angiogenesis was enriched in eoCCA (normalized enrichment score NES=1.51, FDR P=0.16) while IFG (NES= -1.58, FDR P =0.06) and inflammatory response (NES= -1.46, FDR P=0.18) were enriched in aoCCA. Median OS was longer in eoCCA (16.5 vs 13.3 months, Hazard Ratio (HR) 0.86, 95%CI 0.78-0.95, P=0.004). Among patients treated with immunotherapy, median OS was longer in patients with eoCCA (19.2 months, n=19) vs aoCCA (7.6 months, n=150) - HR 0.52, 95%CI 0.28-0.94, p=0.03. No survival difference was identified in patients on chemotherapy (HR 0.91, 95%CI 0.76-1.08, P=0.28). Conclusions: In the largest age-stratified analysis of molecular characteristics of CCA, we identified crucial differences, including higher prevalence of FGFR2 fusions and significant differences in immunotherapy-related markers, angiogenesis enrichment, and inflammatory response. Patients with eoCCA experienced better outcomes with immunotherapy even though immune-oncology-relevant markers favored aoCCA. Our findings, especially higher FGFR2 fusion prevalence in eoCCA, underscore the need for NGS testing and the potential for age-tailored therapeutic strategies. [Table: see text]

Microsatellite instability screening in colorectal carcinoma: immunohistochemical analysis of MMR proteins in correlation with clinicopathological features and Ki-67 protein expression

BackgroundDefects in mismatch repair (MMR) system or microsatellite instability (MSI) and detected in colorectal carcinoma (CRC), both in sporadic and more frequently in hereditary cases. Immunohistochemistry (IHC) is the most frequent method for MMR protein deficiency screening in CRCs. In this study, we aimed to evaluate immunohistochemical expression of MMR and Ki-67 in colorectal carcinoma with clinicopathological features.MethodsIn this study, we evaluated the immunohistochemical expression of MMR proteins including MSH6, MSH2, PMS2 and MLH1 in 50 resection materials with colorectal carcinoma. Their expression is correlated with clinicopathological features of patients together, with Ki-67 protein expression in attempt to screen the most significant predictor of microsatellite instability.ResultsOf the 50 cases of cancer colon, 28% were classified as MSI-H, 20% were MSI-L, and 52% were MSS. The most frequent pattern in MSI-H tumors was concurrent loss of MSH6 and PMS2 proteins. There was a significant correlation between MMR protein expression pattern with tumor size, grade, T-classification and stage (0.015, 0.0515, 0.0162 and 0.0391), respectively. MSI-H tumors were located more frequently in right colon, early TNM stage and poorly differentiated and infrequent distant metastases. There was a significant correlation between Ki-67 high expression and MSI status patterns in their common biological aspects distinct from MSI-negative tumors.ConclusionsMismatch repair defective colorectal carcinoma has characteristics clinicopathological features different from MSS tumors. The role of immunohistochemistry (IHC) for MSI evaluation is the easiest and effective way for evaluation of MMR deficiency in colorectal carcinoma.

Detecting Microsatellite Instability in Endometrial, Colon, and Stomach Cancers Using Targeted NGS.

To develop a method for testing the MSI based on targeted NGS. Based on the results of previous studies, 81 microsatellite loci with high variability in MSI-H tumors were selected, and a method for calculating the MSI score was developed. Using the MSI score, we defined the MSI status in endometral (162), colon (153), and stomach (190) cancers. Accuracy of the MSI scores was evaluated by comparison with MMR immunohistochemistry for 137 endometrium (63 dMMR and 74 pMMR), 76 colon (29 dMMR and 47 pMMR), and 81 stomach (8 dMMR and 73 pMMR) cancers. Classification of MSS and MSI-H tumors was performed with AUC (0.99), sensitivity (92%), and specificity (98%) for all tumors without division into types. The accuracy of MSI testing in endometrial cancer was lower than for stomach and colon cancer (0.98, 87%, and 100%, respectively). The use of 27 loci only, the most informative for endometrial cancer, increased the overall accuracy (1.00, 99%, and 99%). Comparison of MSI score values in 505 tumors showed that MSI score is significantly higher in colon (p < 10-5) and stomach (p = 0.008) cancer compared with endometrial cancer. The MSI score accurately determines MSI status for endometrial, colon, and stomach cancers and can be used to quantify the degree of MSI.

Clinicopathological and molecular analysis of microsatellite instability in prostate cancer: a multi-institutional study in China.

Microsatellite instability (MSI), or mismatch repair-deficiency (dMMR), is rare in prostate cancers (PCas). The histological and molecular features of PCas with MSI/dMMR are incompletely described. Thus, we sought to identify the characteristics of PCas with MSI/dMMR. We analyzed 1,141 primary treatment-naive PCas by MMR-related protein immunohistochemistry (MLH1, PMS2, MSH2, and MSH6). We identified eight cases exhibiting MSI/dMMR (0.7%, 8/1141). Of these, six tumors had both MSH2 and MSH6 protein loss, one had both MLH1 and PMS2 protein loss, and one had only MSH6 loss. Histologically, MSI/dMMR-PCas frequently demonstrated high histological grade (Grade Group 4 or 5), ductal/intraductal histology (6/8 cases), pleomorphic giant-cell features (4/8 cases), and conspicuous tumor lymphocytic infiltration (8/8 cases). Polymerase chain reaction-based analysis of seven MSI/dMMR tumors revealed two MSI-H tumors with loss of both MSH2 and MSH6 proteins. Subsequently, the seven cases underwent next-generation sequencing (NGS) analysis with a highly validated targeted panel; four were MSI. All cases had a high tumor mutation burden (median: 45.3 mutations/Mb). Overall, the MSI/dMMR-PCas showed a high frequency of DNA damage-repair pathway gene changes, including five with pathogenic somatic or germline MMR gene mutations. Activating mutations in the MAPK pathway, PI3K pathway, and WNT/β-catenin pathway were common. TMPRSS2::ERG rearrangement was identified in one case (1/7, 14.3%). Several pathological features are associated with MSI/dMMR in PCas. Identification of these features may help to select patients for genetic screening. As MSI/dMMR-PCas are enriched for actionable mutations, patients should be offered NGS to guide standard-of-care treatment.

1513MO A phase II study of nivolumab plus low dose ipilimumab as first -line therapy in patients with advanced gastric or esophago-gastric junction MSI-H tumor: First results of the NO LIMIT study (WJOG13320G/CA209-7W7)

1513MO A phase II study of nivolumab plus low dose ipilimumab as first -line therapy in patients with advanced gastric or esophago-gastric junction MSI-H tumor: First results of the NO LIMIT study (WJOG13320G/CA209-7W7)

Postoperative circulating tumor DNA-based molecular residual disease in patients with BRAF V600E and MSI-H colorectal cancer: Updated results from GALAXY study in the CIRCULATE-Japan.

32 Background: Postoperative circulating tumor DNA (ctDNA)-based molecular residual disease (MRD) is reported to be associated with a high risk of recurrence. Here, we present an updated analysis of MRD detection and correlations with BRAF and MSI status in radically resected, stage II-IV colorectal cancer (CRC) from the observational GALAXY study (UMIN000039205). Methods: Serial ctDNA was analyzed using personalized tumor-informed assay (Signatera bespoke multiplex-PCR NGS assay) at 1 (4-week MRD time point), 3, 6, 9, 12, 18, and 24 months after surgery or until recurrence (whichever comes first). CT scans were conducted every 6 months. BRAF and MSI status were determined by central assessment until September 2021 and per the standard of care after October 2021. The primary endpoint was disease-free survival (DFS), defined as the time between the date of surgery and date of diagnosis with relapse or death due to any cause. Results: Among 3,615 CRC patients (pts) who were enrolled between May 2020 and April 2022 in GALAXY study, 2,083 pts that met the inclusion criteria were analyzed. The median follow-up period was 16.3 months. Of 2,083 pts included in the analysis, 60 pts (2.9%) had BRAF V600E mutant and MSS tumors, 100 pts (4.8%) had BRAF wt and MSI-H tumors, and 115 patients (5.5%) had BRAF V600E mutant and MSI-H tumors. In the overall population, 286 (14%) were ctDNA-positive at the 4-week MRD time point and 1,797 (86%) were ctDNA-negative. Pts with ctDNA-positivity at the 4-week demonstrated inferior DFS and were 12 times more likely to recur, compared to ctDNA-negative pts (HR: 12, 95%CI: 9.1-15%; p&lt;0.001). Pts with BRAF wt and MSI-H tumors had significantly better DFS compared to pts with BRAF wt and MSS tumors (HR: 4.14, 95 %CI: 1.3-12.9, p=0.015). Pts with BRAF V600E mutant and MSI-H tumors had significantly better DFS compared to pts with BRAF wt and MSS tumors (HR: 4.80, 95 %CI: 1.5-15, p=0.007). On the other hand, ctDNA-positivity was associated with significantly shorter DFS in pts with BRAF V600E mutant and MSS tumors (HR: 2.33, 95%CI: 1.03-5.3, p=0.043) and BRAF V600E mutant and MSI-H tumors (HR:7.54, 95%CI:2.37-24, p&lt;0.001) compared to pts with BRAF wt and MSS tumors. Multivariate analysis in DFS showed that ctDNA positivity was significantly associated with poor prognosis (HR: 11.68, 95%CI: 8.61-15.85, p&lt;0.001), outperforming other clinicopathologic factors such as BRAF and MSI status. Conclusions: ctDNA status at the postoperative MRD time point is the most prognostic risk factor of DFS regardless of BRAF V600E or MSI-H status. Pts with positive postoperative ctDNA should be examined carefully due to a high risk of recurrence. ctDNA-guided adjuvant strategies will further be established by the ongoing randomized VEGA and ALTAIR studies in CIRCULATE-Japan. Clinical trial information: UMIN000039205 .

The results of treatment for resectable gastric cancer with microsatellite instability

Background. microsatellite instability (MSI) is a prognostic marker of survival in many malignant diseases and show resistance to chemotherapy at early stages of colorectal cancer and show no benefits from chemotherapy at early stages of colorectal cancer. However, the role of MSI in resectable gastric cancer (GC) remains unknown.Aim. To study the results of treatment of resectable gastric cancer with microsatellite instability.Materials and methods. The study included 286 patients with resectable gC who received treatment at the N. N. Blokhin national medical Research Center of Oncology. All patients underwent PCR testing for MSI-H in 5 markers (BAT25, BAT26, NR21, NR24, NR27). Tumor regression grades (TRG) were evaluated according to the mandard tumour regression score, including disease-free survival and overall survival.Results. MSI indicated in 27 cases (9.44 %) out of 286 resectable gastric cancer. In group patients who received only surgical treatment, 2-year disease-free survival in patients with MSI-H was 77.80 % versus 88.29 % in MSS patients (hazard ratio (HR) 1.82, 95 % confidence interval (CI) 0.37–8.82, p = 0.45), 2-year overall survival in patients with MSI-H was 88.90 % versus 95.36 % in MSS patients (HR 2.03, 95 % CI 0.20–19.8, p = 0.54). In patients who received perioperative chemotherapy, 28.57 % (4 / 14) had progression in MSI-H tumor versus 3.61 % (6 / 166) in MSS tumor (p &lt;0.001). In group patients who received treatment combined with chemotherapy, 2-year disease-free survival in patients with MSI-H was 59.60 % versus 67.36 % (HR 1.96, CI 95 % 0.88–4.35, p = 0.09), 2-year overall survival in patients with MSI-H was 67.30 % versus 85.86 % in MSS patients (HR 1.86, 95 % CI 0.64–5.41, p = 0.25)Conclusion. MSI-H is not a favorable prognosis factor in patients with resectable GC who are treated surgically combined with chemotherapy. The prevalence of progression in patients with MSI-H-status is higher than MSS-status with perioperative chemotherapy (FLOT / FOLFIRINOX).

Microsatellite instability states serve as predictive biomarkers for tumors chemotherapy sensitivity

There is an urgent need for markers to predict the efficacy of different chemotherapy drugs. Herein, we examined whether microsatellite instability (MSI) status can predict tumor multidrug sensitivity and explored the underlying mechanisms. We downloaded data from several public databases. Drug sensitivity was compared between the high microsatellite instability (MSI-H) and microsatellite-stable/low microsatellite instability (MSS/MSI-L) groups. In addition, we performed pathway enrichment analysis and cellular chemosensitivity assays to explore the mechanisms by which MSI status may affect drug sensitivity and assessed the differences between drug-treated and control cell lines. We found that multiple MSI-H tumors were more sensitive to a variety of chemotherapy drugs than MSS/MSI-L tumors, and especially for CRC, chemosensitivity is enhanced through the downregulation of DDR pathways such as NHEJ. Additional DNA damage caused by chemotherapeutic drugs results in further downregulation of DDR pathways and enhances drug sensitivity, forming a cycle of increasing drug sensitivity.

Molecular profile of SWI/SNF complex in Chinese patients with ovarian cancer.

e17538 Background: The SWI/SNF chromatin remodeling complexes are involved in cancer initiation and progression. ARID1A, ARID1B, and SMARCA4, the essential ATPase subunit of the SWI/SNF chromatin remodeling complex, play important roles in altering chromatin structure for various cellular functions, including transcription, DNA synthesis, and repair. In ovarian cancer (OC), mutations in several SWI/SNF complexes were found, but the molecular and clinical characteristics of SWI/SNF complexes in Chinese ovarian cancer patients have never been reported. Herein, we explored the profiles of SWI/SNF mutation in the Chinese population. Methods: Genetic mutations were reviewed in OC patients who underwent hybridization capture-based next-generation sequencing (NGS). SWI/SNF mutant tumors (MT) were compared to wild-type (WT), and other co-mutations were also evaluated. The pathogenicity of germline mutations was categorized based on American College of Medical Genetics and Genomics (ACMG) guidelines. Results: Among 461 OC patients, mutation in at least one evaluated SWI/SNF gene was identified in 23% of cases. The higher frequency of SWI/SNF alterations were ARID1A (41.03%), ARID1B (13.33%), SMARCA4 (11.28%). The co-mutated genes mainly were TP53, PIK3CA, PTEN. In addition, the MT group tended to have more BRCA1/2 somatic mutations (16.34% vs. 7.84%, χ2＝5.42, p &lt; 0.05) and less BRCA1/2 germline pathogenic mutations (8.7% vs. 17.2%, χ2＝4.40, p &lt; 0.05) compared to WT group. There was a significant statistical difference in TMB value that the MT group was higher than the WT group (11.1 vs. 2.7, p &lt; 0.001). In the MT group, the TMB of patients co-mutated with TP53, PIK3CA, or PTEN mutations were 10.2, 17.3, and 32.3, respectively. The proportion of SWI/SNF alterations in MSI-H tumors was higher than those without mutations (10.0% vs. 0.8%, χ2＝19.29, p &lt; 0.001). Conclusions: SWI/SNF mutant tumors tended to be associated with higher TMB (11.1 vs. 2.7), a higher proportion of MSI-H (10% vs. 0.8%), and more BRCA1/2 somatic mutations ( 16.34% vs. 7.84%) in OC patients. Patients with SWI/SNF alterations combined with PTEN mutation have extremely high TMB (TMB=32.3), indicating they may benefit more from immunotherapy.

Preoperative pembrolizumab for MSI high, EBV positive or PD-L1 positive locally advanced gastric cancer followed by surgery and adjuvant chemoradiation with pembrolizumab: Interim results of a phase 2 multi-center trial.

e16073 Background: Certain subtypes of gastric cancer are less responsive to perioperative chemotherapy. Preliminary data suggest that patients with microsatellite unstable tumors (MSI-H), EBV expressing tumors, and tumors with high PD-L1 expression may benefit from immunotherapy. This trial evaluates the benefit of PD-1 checkpoint immunotherapy in this subset of patients with operable gastric cancer. Methods: This is an interim analysis of this Phase II multicenter clinical trial (NCT03257163). Patients with clinically staged T2-T4, N0-N3, M0 gastric adenocarcinoma with MSI-H, EBV positive, or PDL1 expression with CPS&gt;/= 1% were included. Patients receive two cycles of neoadjuvant pembrolizumab followed by gastrectomy. Postoperatively, patients receive one cycle of adjuvant capecitabine + pembrolizumab, then chemoradiation (45 Gy in 25 fractions) with capecitabine + pembrolizumab, then two cycles capecitabine + pembrolizumab, followed by 1-year total of pembrolizumab. Primary endpoint is DFS. Results: 33 patients have been enrolled (planned = 40), median age 67.5 years (range, 44 – 88 years), median follow up duration 18.1 months (range, 2.3-42.5 months). Patients were 30.3% Hispanic, 27% Asian, 24% Black, 24% White. Of the 33 patients, 12 were MSI-H, 3 EBV (+), and 27 had PDL1 expression with CPS &gt;/= 1%. Tumors were of advanced clinical stage with 84.8% ≥ cT3 and 57.6% cN(+). All patients received two cycles of neoadjuvant pembrolizumab. Six patients did not undergo surgical resection: peritoneal disease (n=2), soft tissue metastases (n=1), locally unresectable (n=2), and frailty (n=1). Of the 27 patients who completed surgery, treatment effect was seen in 46%, down-staging in 50%, and complete pathologic response seen in 3 patients, 2 of whom has MSI-H tumors and one whose tumor was PDL1+. Two patients did not receive adjuvant therapy due to postoperative complications. A total of 20 patients have received optimal therapy as defined as postoperative RT and &gt;/=2 cycles of adjuvant pembrolizumab. Four patients had adjuvant pembrolizumab monotherapy (after chemoradiation) stopped due to rash (n=2), pneumonia (n=1), and colitis (n=1). The estimated DFS at 24 months is 60% for the entire cohort (n=33). For the optimal therapy cohort (n=20), the DFS at 24 months is 80%. Conclusions: In this biomarker-driven cohort of operable gastric adenocarcinoma patients, results are promising for patients who receive two cycles of preoperative pembrolizumab followed by postoperative concurrent pembrolizumab with chemoradiation therapy, and chemotherapy. This study presents a novel and effective paradigm for the management of localized gastric cancer and warrants additional evaluation. Clinical trial information: NCT03257163 .

Exploring the prognostic and predictive impact of genomic loss of heterozygosity (LOH) in patients with metastatic colorectal cancer (mCRC).

3576 Background: Genomic LOH consists in the loss of chromosomal regions and is associated with the homologous recombination repair (HRR) system deficiency (dHRR). In ovarian cancer, LOH-high predicts benefit from platinum-based chemotherapy and PARP inhibitors. In mCRC, the role of LOH has been poorly investigated. Methods: An NGS-based assay (FoundationOne CDx; Foundation Medicine, Inc. Cambrige, MA) was used to determine the percentage of genomic LOH and the presence of pathogenetic mutations in HRR-related genes in archival chemo-naïve tumor tissues of mCRC patients included in a real-world registry and in the AtezoTRIBE (NCT03721653) and AVETRIC (NCT04513951) studies. Both these trials assessed the combination of an anti-PDL1 (atezolizumab or avelumab) with the triplet FOLFOXIRI plus bevacizumab or cetuximab. The prespecified cut-off of ≥14.08 for LOH-high described for mCRC (Sokol et al., JCO Precis Oncol 2020) was adopted. Tumors with at least one biallelic alteration in any of the 27 genes involved in the HRR pathway (Riaz et al., Nat Commun 2017) included in the FoundationOne CDx panel (BARD1, BRCA1, BRCA2, BRIP1, MRE11A, NBN, PALB2, RAD51, RAD51B, RAD51C, RAD51D, RAD52, RAD54L, RBBP8, XRCC2, ABL1, ATM, ATR, BAP1, CDK12, DNMT3A, ERCC4, FANCA, FANCC, FANCG, FANCL, PARP1) were defined dHRR. Results: Overall, 196 samples were analysed. None of 7 MSI-H tumors were classified as LOH-high. Fourteen (7%) and 6 (3%) of 189 MSS tumors were classified as LOH-high and dHRR, respectively. In particular, in LOH-high subgroup, 3 (21%) tumors were defined dHRR, while 3 (50%) tumors were classified LOH-high among dHRR tumors. LOH-high tumors were more frequently BRAF mutated (p = 0.019) and dHRR (p = 0.006) compared to LOH-low. Among patients receiving triplet chemotherapy +/- biologic agent alone (N = 55) or with an anti-PDL1 (N = 58), an interaction effect was shown between the effect of the addition of the checkpoint inhibitor and LOH with higher benefit in the LOH-high subgroup (N = 10) in terms of both PFS (pinteraction= 0.002) and OS (pinteraction&lt; 0.001). In the cohort of patients not receiving the anti-PDL1, longer PFS was observed in patients with LOH-low (N = 125) respect to LOH-high (N = 6) tumors (12.1 vs 5.1 months, HR: 0.11, 95%CI: 0.04-0.26, p &lt; 0.001). No differences in PFS or OS were reported between patients treated with first-line oxaliplatin- (N = 40) vs irinotecan-based doublets (N = 25) or with the triplet FOLFOXIRI (N = 55) vs doublets (N = 65) according to LOH status. No prognostic or predictive impact of HRR deficiency was shown. Conclusions: In MSS mCRC, LOH-high was associated with biallelic alterations in the HRR system, worse prognosis and higher benefit from the addition of anti-PDL1 agents to chemotherapy. Considering the low number of LOH-high tumors in our study, these results deserve confirmation in larger cohorts.

Clinical and molecular characterization of microsatellite stable (MSS) tumors in patients with Lynch syndrome (LS).

10517 Background: LS, caused by germline pathogenic variants in DNA mismatch repair genes ( MLH1, MSH2, MSH6, PMS2, EPCAM), is characterized by near universal microsatellite instability (MSI) /mismatch repair deficiency (dMMR) of associated tumors. Accurate MSI/dMMR assessment is critical to determine immune checkpoint blockade (ICB) eligibility but discordant MSI and immunohistochemical analysis of MMR status (IHC) is described. We sought to characterize MSS tumors in LS pts. Methods: Cancer-affected LS pts consented to tumor and matched normal DNA sequencing via MSK-IMPACT (NCT01775072), from 01/2014 to 10/2022 were identified with MSI status inferred via MSIsensor. Microsatellite stable (MSS) or indeterminate (MSI-I) tumors were further assessed using MiMSI, a multiple instance learning tool demonstrating high sensitivity for MSI even with low sample tumor purity. MSIsensor and MiMSI status were correlated with MMR IHC. Results: Of 405 LS cancer-affected pts, 120 had MSS, 59 MSI-I and 226 MSI-H tumors using MSIsensor. Tumor MSI status varied by LS gene with MSH6 (MSS: 33 % (40/120), MSI-I: 42 % (25/59), MSI-H: 14%, (33/226)) and PMS2 (MSS: 42% (50/120), MSI-I: 12% (7/59), MSI-H: 12%, (28/226)) more frequently associated with MSS/MSI-I than MSI-H tumors (p = 0.005) Utilizing orthogonal analysis 43% (63/147) of MSS/MSI-I tumors were re-classified as either MSI or dMMR. MiMSI was available on 106/147 MSS/MSI-I tumors resulting in the re-classification of 36 (28/38 MSI-I, 8/68 MSS tumors) as MSI. IHC was concordant with reclassified MSI status in 23/36 tumors. 5/36 reclassified cases (MSI-I on MSIsensor) had pMMR on IHC, in 2 of these cases MSI was driven by POLE/POLD1 somatic mutations. IHC was unavailable for 8/36 tumors reclassified by MiMSI. 18/36 (50%) tumors reclassified by MiMSI had low tumor content in profiled samples a known limitation of MSIsensor. Tumors reclassified as MSI on MiMSI included those considered part of the LS cancer spectrum (8 CRC, 3 gastroesophageal, 8 endometrial, 3 ovarian, 3 urothelial, 2 brain) in addition to cancers not clearly LS associated. 6 tumors classified as MSS/MSI-I on MSIsensor were MSI-I on MiMSI, 2/6 were dMMR on IHC. Amongst the 41 MSS/MSI-I tumors on MSIsensor for which only orthogonal IHC was available, 25 (11 MSS, 14 MSI-I) demonstrated protein loss consistent with the implicated LS gene. Low tumor content was noted in 44% (11/25) of tumors reclassified by IHC. Of 34 tumors found to be both MSS and MMR-proficient on orthogonal analysis, the majority (88%) were in PMS2 or MSH6 carriers, representing 9 tumor types of which 4 are considered part of the LS cancer spectrum. Conclusions: MSS tumors occur in LS pts, especially in lower-penetrance MSH6 and PMS2 carriers. Given increasing ICB utilization for MSI/dMMR tumors, orthogonal testing methods are critical to comprehensively identify all LS pts who may derive benefit from ICB particularly when sample tumor purity is low.

Microsatellite instability in sarcomas: Which tests are ideal for diagnosis, and a systematic review.

e23519 Background: There is a lack of clinically relevant biomarkers to inform immunotherapy for patients with advanced sarcoma. High levels of microsatellite instability (MSI-H) is a histology-agnostic biomarker of response to pembrolizumab. However, the frequency of MSI-H in sarcoma is low - ranging from &lt; 0.5% to 4%. MSI status can be assessed through Next Generation Sequencing (NGS), immunohistochemistry (IHC), or PCR followed by fragment analysis with capillary electrophoresis (PCR). To date, both the College of American Pathologists and the European Society of Medical Oncology have not determined the best method to evaluate MSI in sarcomas. We performed a systematic review to evaluate the methods to diagnose MSI-H in sarcomas and to evaluate the impact of this biomarker in treatment with pembrolizumab. Methods: We performed a literature search using the PubMed database to identify all English full-text available articles between January 2020 and December 2022, with the terms “sarcoma and microsatellite instability” and “sarcoma and pembrolizumab”. One independent reviewer screened the studies, two independent reviewers analyzed the selected studies, and disparities were resolved with a fourth reviewer. Results: Concerning the diagnosis of MSI-H in sarcomas, 187 abstracts were found, and 31 articles were identified for full analysis. MSI status was reported for 7958 patients - 7762 with a single test and 196 with more than one test. NGS was used for 6649, IHC for 1222, and PCR for 285. MSI-H tumors were 25 (0.4%), 38 (3.1%), and 21 (7.33%) for each technique, respectively. Two investigators performed the three available tests to evaluate concordance, with one concordant case and one discordant case. 194 cases were evaluated for both IHC and PCR, with 4 discordant results. No article that described NGS results performed a second test. To analyze the results of pembrolizumab therapy in MSI-H sarcomas, we analyzed 199 abstracts of which we selected 13 prospective and 31 retrospective papers for full analysis. The K cohort of Keynote-158 is the only prospective trial concerning therapies guided by MSI-H in sarcoma (n = 14 patients, ORR = 33%). Regarding retrospective data, we identified 2 case reports of the use of pembrolizumab in patients with advanced sarcoma – both achieved a response to therapy. Conclusions: There is limited data to establish the best diagnostic test for MSI-H in patients with sarcoma. The concordance between NGS, IHC, and PCR seems to be high, but discrepancies have been described and need to be further evaluated. There is only one prospective trial that enrolled patients with sarcomas utilizing MSI-H as a biomarker. Albeit only a small number of patients were enrolled (n = 14), efficacy is consistent with that seen in other histologies, and scarce retrospective data corroborate the potential role of MSI-H as a predictive biomarker of pembrolizumab efficacy in sarcoma.

Multi-omic analysis in normal colon organoids highlights MSH4 as a novel marker of defective mismatch repair in Lynch syndrome and microsatellite instability.

Lynch syndrome (LS) is a hereditary condition that increases the risk of colorectal (CRC) and extracolonic cancers that exhibit microsatellite instability (MSI-H). MSI-H is driven by defective mismatch repair (dMMR), and approximately 15% of nonhereditary CRCs also exhibit MSI-H. Here, we aimed to better define mechanisms underlying tumor initiation in LS and MSI-H cancers through multi-omic analyses of LS normal colon organoids and MSI-H tumors. Right (n = 35) and left (n = 23) colon organoids generated from normal colon biopsies at routine colonoscopy of LS and healthy individuals were subjected to Illumina EPIC array. Differentially methylated region (DMR) analysis was performed by DMRcate. RNA-sequencing (n = 16) and bisulfite-sequencing (n = 15) were performed on a subset of right colon organoids. CRISPR-cas9-mediated editing of MMR genes in colon organoids of healthy individuals was followed by quantitative PCR of MSH4. The relationship between MSH4 expression and tumor mutational burden was further explored in three independent tumor data sets. We identified a hypermethylated region of MSH4 in both the right and left colon organoids of LS versus healthy controls, which we validated using bisulfite-sequencing. DMR analysis in three gastrointestinal and one endometrial data set revealed that this region was also hypermethylated in MSI-H versus microsatellite stable (MSS) tumors. MSH4 expression was increased in colon organoids of LS versus healthy subjects and in publicly available MSI-H versus MSS tumors across four RNA-seq and four microarray data sets. CRISPR-cas9 editing of MLH1 and MSH2, but not MSH6, in normal colon organoids significantly increased MSH4 expression. MSH4 expression was significantly associated with tumor mutational burden in three publicly available data sets. Our findings implicate DNA methylation and gene expression differences of MSH4 as a marker of dMMR and as a potential novel biomarker of LS. Our study of LS colon organoids supports the hypothesis that dMMR exists in the colons of LS subjects prior to CRC.

Abstract 6652: Overcoming immune resistance in DNA mismatch repair deficient tumors

Abstract Patients with Lynch Syndrome, an inherited mismatch repair deficiency, have an increased risk for developing microsatellite unstable (MSI-H) cancers. Our team recently identified shared immunogenic frameshift peptides in patient tumors that can be targets of T cell surveillance and preventative MSI-H cancer vaccines. We hypothesize that in Lynch Syndrome some premalignant lesions are capable of evading immune surveillance from cytotoxic T lymphocytes due to immune checkpoint expression and immunosuppression from tumor, myeloid and stromal cell populations. Immune checkpoint blockade (ICB) showed promising results in the treatment of MSI-H tumors and are being evaluated in the adjuvant setting of patients with Lynch syndrome post surgical resection. However, a significant percentage of advanced MSI-H tumors resist ICB suggesting evolving mechanisms of immune resistance. We will use a MSI-H in vivo mouse model and additionally develop 3D spheroids cocultured with immune cells to characterize the underlying immune resistance mechanisms. The MSI-H mouse model will first be applied to identify shared frameshifts in MSI-H tumors by WES to develop vaccination approaches preventing tumor growth. Then, we will quantify MSI-H tumor growth and immune infiltration in the MSI-H mouse model, with or without ICB. We will analyze the presence of immunosuppressive pathways and myeloid subsets in high growth resisting tumors by immunohistochemistry, scRNAseq, spatial transcriptomics and flow cytometry. In addition, we will perform short-term in vitro spheroid-splenic cell co-cultures to better characterize the early immune resistance mechanisms. We will also perform these spheroid-immune cell co-cultures using the immune cells taken from mice after vaccination or ICB to identify the early myeloid immune resistance mechanisms. Overall, the identification of shared immune frameshifts in MSI-H tumors will allow developing peptide vaccination approaches to prevent tumor growth. Further, the identification of myeloid resistance pathways will guide therapeutic strategies against tumor resisting to ICB and/or vaccination. Citation Format: Guillaume Mestrallet. Overcoming immune resistance in DNA mismatch repair deficient tumors [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 6652.

Urachal carcinoma: The journey so far and the road ahead.

Urachal carcinoma, a rare cancer arising from urachus, accounts for about 1% of bladder cancer. The diagnosis at stage I shows about 63% 5-year survival whereas only 8% of the patients at stage IV shows a 5-year survival. Above 90% of urachal carcinomas are adenocarcinomas and most of the urachal carcinoma cases are invasive, showing a high resemblance to adenocarcinoma of various origins, making it hard for a conclusive diagnosis. Even though inconclusive, immunohistochemistry can play a significant role in identifying urachal carcinoma. Most cases show the biomarkers CK20 and CDX2, whereas CK7 and β-catenin are expressed at a lesser frequency. Due to the few cases available, there is a lack of evidence regarding specific markers differentiating urachal carcinoma from colorectal or primary bladder adenocarcinomas. In addition to immunohistochemistry, genomic characterization is emerging to play a role in the classification and treatment of the disease. Urachal carcinoma has been reported to have a molecular level similarity with colorectal malignancies regarding certain gene expressions. The TP53 mutations inactivating the tumor suppressor can probably be explored as a possible target in treating urachal carcinoma. Additionally, certain targets identified in gastric and breast cancer along with anti-HER2 treatment strategies can be explored. Immuno-oncology utilizes immune checkpoint inhibitors for the treatment of MSI-H tumors whereas a combination of tyrosine kinase inhibitors along with immune checkpoint inhibitors are being studied to treat MSI stable tumors. The article is an in-depth overview of urachal carcinoma addressing the current landscape with an emphasis on the future scenario.