Microsatellite Stable Tumors Research Articles

Consensus molecular subtyping of metastatic colorectal cancer expands biomarker-directed therapeutic benefit for patients with CMS1 and CMS2 tumors.

We developed a whole transcriptome sequencing (WTS)-based Consensus Molecular Subtypes (CMS) classifier using FFPE tissue and investigated its prognostic and predictive utility in a large clinico-genomic database of CRC patients (n = 24,939). The classifier was trained against the original CMS datasets using an SVM model and validated in an independent blinded TCGA dataset (88.0% accuracy). Kaplan-Meier estimates of overall survival (OS) and time-on-treatment (TOT) were calculated for each CMS (p < 0.05 considered significant). CMS2 tumors were enriched on left-side of colon and conferred the longest median OS. In RAS-wildtype mCRC, left-sided tumors and CMS2 classification were associated with longer TOT with anti-EGFR antibodies (cetuximab and panitumumab). When restricting to only CMS2, there was no significant difference in TOT between right- versus left-sided tumors. CMS1 tumors were associated with a longer median TOT with pembrolizumab relative to other CMS groups, even when analyzing only microsatellite stable (MSS) tumors. A WTS-based CMS classifier allowed investigation of a large multi-institutional clinico-genomic mCRC cohort, suggesting anti-EGFR therapy benefit for right-sided RAS-WT CMS2 tumors and immune checkpoint inhibitor benefit for MSS CMS1. Routine CMS classification of CRC provides important treatment associations that should be further investigated.

TROP2 in colorectal carcinoma: associations with histopathology, molecular phenotype, and patient prognosis.

Antibody-drug conjugates (ADCs) directed to trophoblast cell surface antigen 2 (TROP2) have gained approval as a therapeutic option for advanced triple-negative breast cancer, and TROP2 expression has been linked to unfavourable outcomes in various malignancies. In colorectal carcinoma (CRC), there is still a lack of comprehensive studies on its expression frequency and its prognostic implications in relation to the main clinicopathological parameters. We examined the expression of TROP2 in a large cohort of 1,052 CRC cases and correlated our findings with histopathological and molecular parameters, tumour stage, and patient outcomes. TROP2 was heterogeneously expressed in 214/1,052 CRCs (20.3%), with only a fraction of strongly positive tumours. TROP2 expression significantly correlated with an invasive histological phenotype (e.g. increased tumour budding/aggressive histopathological subtypes), advanced tumour stage, microsatellite stable tumours, and p53 alterations. While TROP2 expression was prognostic in univariable analyses of the overall cohort (e.g. for disease-free survival, p < 0.001), it exhibited distinct variations among important clinicopathological subgroups (e.g. right- versus left-sided CRC, microsatellite stable versus unstable CRC, Union for International Cancer Control [UICC] stages) and lost its significance in multivariable analyses that included stage and CRC histopathology. In summary, TROP2 is quite frequently expressed in CRC and associated with an aggressive histopathological phenotype and microsatellite stable tumours. Future clinical trials investigating anti-TROP2 ADCs should acknowledge the observed intratumoural heterogeneity, given that only a subset of TROP2-expressing CRC show strong positivity. The prognostic implications of TROP2 are complex and show substantial variations across crucial clinicopathological subgroups, thus indicating that TROP2 is a suboptimal parameter to predict patient prognosis.

Comprehensive Analysis of Microsatellite Instability in Canine Cancers: Implications for Comparative Oncology and Personalized Veterinary Medicine.

Microsatellite instability (MSI) is a crucial feature in cancer biology, yet its prevalence and significance in canine cancers remain largely unexplored. This study conducted a comprehensive analysis of MSI across 10 distinct canine cancer histotypes using whole-exome sequencing data from 692 tumor-normal sample pairs. MSI was detected in 64% of tumors, with prevalence varying significantly among cancer types. B-cell lymphomas exhibited the highest MSI burden, contrasting with human studies. A novel "MSI-burden" score was developed, correlating significantly with tumor mutational burden. MSI-high (MSI-H) tumors showed elevated somatic mutation counts compared to MSI-low and microsatellite stable tumors. The study identified 3632 recurrent MSI-affected genomic regions across cancer types. Notably, seven of the ten cancer types exhibited MSI-H tumors, with prevalence ranging from 1.5% in melanomas to 37% in B-cell lymphomas. These findings highlight the potential importance of MSI in canine cancer biology and suggest opportunities for targeted therapies, particularly immunotherapies. The high prevalence of MSI in canine cancers, especially in B-cell lymphomas, warrants further investigation into its mechanistic role and potential as a biomarker for prognosis and treatment response.

Mesothelin expression correlates with elevated inhibitory immune activity in patients with colorectal cancer

The expression of the protein Mesothelin (MSLN) is highly variable in several malignancies, including colorectal cancer (CRC), and high levels are associated with aggressive clinicopathological features and worse patient survival. Colorectal cancer is both a common and deadly cancer; being the third most common in incidence and second most common cause of cancer-related death. While systemic therapy remains the primary therapeutic option for most patients with stage IV (metastatic; m) CRC, their disease eventually becomes treatment refractory, and 85% succumb within 5 years. Microsatellite-stable (MSS) CRC tumors, which constitute more than 90% of patients with mCRC, are generally refractory to immunotherapeutic interventions. In our current work, we characterize MSLN levels in CRC, specifically correlating expression with clinical outcomes in relevant CRC subtypes, and explore how MSLN expression impacts the status of immune activation and suppression in the peritumoral microenvironment. Higher MSLN expression is prevalent in CMS1 and CMS4 CRC subtypes and correlates with higher gene mutation rates across the patient cohorts. Further, MSLN-high patients exhibit increased M1/M2 macrophage infiltration, PD-L1 staining, immune-inhibitory gene expression, enrichment in inflammatory, TGF-β, IL6/JAK/STAT3, IL2/STAT5 signaling pathways, and mutation in KRAS and FBXW7. Together, these results suggest that MSLN protein is a potential target for antigen-specific therapy and supports investigation into its tumorigenic effects to identify possible therapeutic interventions for patients with high MSLN expressing MSS CRC.

Chromosomal copy number based stratification of gastric cancer has added prognostic value to Lauren’s histological classification

BackgroundThe Cancer Genome Atlas (TCGA) recognizes four molecular subgroups of gastric cancer: Epstein-Barr virus (EBV) positive, microsatellite instable (MSI), genomically stable (GS), and chromosomal instable (CIN). Since a GS/CIN classifier is lacking, alternative markers such as Lauren’s histopathology or CDH1/p53 immunohistochemistry are commonly applied. Here we compared survival of gastric cancer subgroups determined by four methods.Methods309 EBV negative and microsatellite stable tumors were included from the Dutch D1/D2 trial and assigned to subgroups by: (i) TCGA’s specific chromosomal copy number aberrations, (ii) genome instability index (GII), (iii) Lauren’s classification, and (iv) CDH1/p53 immunohistochemistry. Subgroups were associated with cancer-related survival (CRS).ResultsFive-year CRS was 42.0% for diffuse and 49.5% for patients with intestinal type tumors, and 57.8% for GS and 41.6% for patients with CIN tumors. Classification by GII or CDH1/p53 IHC did not correlate with CRS. The combination of TCGA and Lauren classifications resulted in four distinct subgroups. Five-year CRS for GS-intestinal (n = 24), GS-diffuse (n = 57), CIN-intestinal (n = 142) and CIN-diffuse (n = 86) was 61.4%, 56.5%, 47.6%, and 31.5%, respectively.ConclusionsTCGA’s GS and CIN subgroups have additional prognostic value to Lauren’s classification in resectable gastric cancer. GS-intestinal, GS-diffuse, CIN-intestinal and CIN-diffuse are suggested stratification variables for future studies.

Immune checkpoint inhibitors for patients with mismatch-repair deficient or microsatellite instability-high advanced cancers: a meta-analysis of phase I-III clinical trials.

Mismatch-repair deficient (dMMR) and microsatellite instability-high (MSI-H) cancers are associated with an increased number of somatic mutations, which can render tumors more susceptible to immune checkpoint blockade. However, a comprehensive evaluation of the efficacy profile of immune checkpoint inhibitors in this patient population across multiple cancer types is lacking. This study aims to address this knowledge gap by synthesizing data from Phase I-III clinical trials. A systematic search was conducted in PubMed, Embase, the Cochrane Central Register of Controlled Trials and Google Scholar from inception until June, 2024. Eligible studies included randomized controlled trials (RCTs), non-randomized comparative studies, and single-arm trials investigating immune checkpoint inhibitors in patients with dMMR/MSI-H advanced cancers. The primary outcome was objective response rate (ORR), and the secondary outcomes included disease control rate (DCR), 1-year, 2-year, and 3-year overall survival (OS) and progression-free survival (PFS) rates. Subgroup analyses were conducted for the primary outcome stratified by major study characteristics. Of the 10802 identified studies, 19 trials in 25 studies totaling 2052 participants met the inclusion criteria and were included in the meta-analysis. The pooled ORR was 41.7% (95% CI, 35.7%-47.7%). The pooled DCR was 68.9% (95% CI, 62.2%-75.7%). The pooled 12-month, 24-month and 36-month OS rates were 29.1% (95% CI, 19.9%-38.3%), 35.8% (95% CI, 23.6%-48.0%), and 35.8% (95% CI, 23.6%-48.0%), respectively. The pooled 12-month, 24-month and 36-month PFS rates were 46.4% (95% CI, 39.1%-53.8%), 67.0% (95% CI, 55.2%-78.8%), and 63.1% (95% CI, 37.3%-88.9%), respectively. The study establishes the therapeutic potential of immune checkpoint inhibitors in dMMR/MSI-H advanced cancers, highlighting the importance of MSI status in this context. Further head-to-head comparisons are needed to conclusively determine MSI's predictive power relative to proficient mismatch-repair/ microsatellite stable (pMMR/MSS) tumors.

The Landscape and Prognosis of Microsatellite Stable (MSS) Esophageal, Gastro-Esophageal Junction and Gastric Adenocarcinomas with High Tumor Mutation Burden (TMB)

Background A minority of patients with MSS tumors present a high tumor mutation burden (TMB) without underlying MMR defects. Methods Publicly available genomic series were assessed for identification of patients with MSS gastric gastroesophageal junction, and esophageal adenocarcinomas and a high TMB, defined as more than 10 mutations per Mb. These were compared with MSS cancers and a low TMB for genetic alterations and for survival outcomes. Results Patients with MSS cancers with high TMB in the MSK series were older but did not differ in other clinicopathologic parameters compared with MSS patients with low TMB. Mutations in tumor suppressors TP53 and APC and oncogenes KRAS and ERBB4 as well as amplifications of ERBB2 were more prevalent in the high TMB group of MSS cancers. Mutations in DDR associated genes, in epigenetic modifiers and in genes associated with immune response were more prevalent in the hIgh TMB group patients. However, high TMB was not associated with an improved survival in MSS gastric/gastroesophageal junction/esophageal adenocarcinomas (Log Rank p = 0.5). Conclusion MSS Gastric/gastroesophageal junction/esophageal adenocarcinomas with TMB above 10 mutations per Mb possess a genomic landscape with increased alteration frequencies in common gastroesophageal cancer genes and pathways.

Efficacy and safety of combining short-course neoadjuvant chemoradiotherapy with envafolimab in locally advanced rectal cancer patients with microsatellite stability: A phase II PRECAM experimental study.

Conventional neoadjuvant chemoradiotherapy (nCRT) yields a pathologic complete response (pCR) rate of 15%-30% for locally advanced rectal cancer (LARC). This study ventures to shift this paradigm by incorporating short-course nCRT with immunotherapy, specifically Envafolimab, to achieve improved treatment efficacy and possibly redefine the standard of care for LARC. The PRECAM study is a prospective, single-arm, phase 2 clinical trial for LARC in patients with microsatellite stable (MSS) tumors. Participants received short-course radiotherapy (25Gy/5f), followed by two cycles of CAPEOX chemotherapy and six weekly doses of Envafolimab, a PD-L1 antibody, before total mesorectal excision surgery. The primary endpoint was the pCR rate. From April to December 2022, 34 patients were enrolled, of whom 32 completed the study, each diagnosed with an MSS rectal adenocarcinoma. All patients underwent preoperative CRT combined with Envafolimab. Remarkably, a pCR rate of 62.5% (20/32) was attained, and a significant pathologic response rate of 75% (24/32) was achieved. Additionally, 21 of 32 participants achieved a neoadjuvant rectal (NAR) score below 8, suggesting an effective treatment response. Common adverse events included tenesmus (78.1%), diarrhea (62.5%), and leukocyte decrease (40.6%). Two Grade 3 adverse events were noted, one related to liver function abnormality and the other to a decrease in platelet count. Surgical procedures were performed in all cases, with minor complications, including ileus, infections, and anastomotic leakage. As of this report, there have been no reported cases of recurrence or death during the follow-up period, ranging from 12 to 20 months. In LARC patients exhibiting MSS tumors, combining short-course nCRT with Envafolimab demonstrated favorable efficacy, leading to a significant pCR rate. Minor adverse effects and surgical complications were observed. These preliminary but promising results underscore the potential of this approach and call for further exploration and validation through a randomized controlled trial.

Genomic, immunologic, and prognostic associations of TROP2 (TACSTD2) expression in solid tumors.

TROP2 (TACSTD2) expression is associated with decreased overall survival (OS) in some solid tumors, and the TROP2-targeting antibody-drug conjugate (ADC) sacituzumab govitecan has been approved in breast and urothelial carcinomas. We aimed to explore the multi-omic landscape associated with TACSTD2 gene expression in various solid tumors to identify patients most likely to benefit from this approach. Breast (N = 11246), colorectal (N = 15425), hepatocellular (N = 433), pancreatic (N = 5488), and urothelial (N = 4125) tumors were stratified into quartiles by TACSTD2 gene expression, analyzed by next-generation DNA sequencing, whole transcriptome sequencing, and immunohistochemistry at Caris Life Sciences (Phoenix, AZ). Survival data were obtained from insurance claims, and Kaplan-Meier estimates were calculated for molecularly defined cohorts. Several pathogenic mutations were associated with TACSTD2-high tumors, including TP53 in breast, colorectal (CRC), pancreatic, and hepatocellular cancers; KRAS in pancreatic and CRC cancers; ARID1A and FGFR3 in urothelial cancer; and CTNNB1 in hepatocellular cancer. TACSTD2-low breast tumors were enriched for copy number amplifications in CCND1 and FGF/R family member genes. TACSTD2 high was generally associated with more immune cell infiltration and greater T-cell inflammation scores. Patients with TACSTD2-high breast, CRC, and pancreatic cancers demonstrated a significantly shorter OS than TACSTD2-low tumors. This was restricted to CRC with microsatellite stable tumors and patients with pancreatic cancer with KRAS-mutant tumors. Patients with breast cancer with TACSTD2-high tumors also experienced significantly worse OS following immune checkpoint inhibitors. TACSTD2 expression is associated with key driver alterations and a more active immune microenvironment, suggesting possible combinatorial strategies with TROP2-targeting ADCs plus immunotherapy in various solid tumors.

Evolutionary history of adenomas to colorectal cancer in FAP families.

Familial adenomatous polyposis (FAP) is a genetic syndrome characterized by multiple polyps at various evolutionary stages, which, if left untreated, inevitably progress to colorectal cancer (CRC). In this study, we present a comprehensive analysis of the evolutionary history of FAP-CRC from precancerous adenoma to carcinoma. Tissues were collected from gastrointestinal endoscopy or surgical resection. Exome sequencing was performed on multiple regions of adenocarcinoma (n = 8), villous adenoma (n = 10), tubular adenoma (n = 9) and blood samples were obtained from 9 patients belonging to 7 Chinese FAP families. Phylogenetic trees were reconstructed, and evolutionary analysis was conducted to reveal the temporal sequence of events leading to CRC. Inherited germline mutation sites in APC gene were identified in FAP01 (p.S1281*, COSM19212), FAP03 (p.S384Tfs*19), FAP04 (p.E1538*, COSM6041693), FAP05 (p.Q1062*, COSM3696862), and FAP07-FAP09 (p.V677Sfs*3). Notably, p.V677Sfs*3 mutation was recognized as a novel germline mutation in APC, supported by evidence of genotype-phenotype correlation in pedigree analysis. Adenomas exhibited lower mutational rates than FAP-CRC and displayed recurrent alterations in well-known chromosomal instability (CIN) genes (APC, RAS, SMAD4 and TP53) and DNA damage repair genes (SUZ12, KMT2C, BCLAF1, RUNX1, and ARID1B), suggesting the presence of genomic instability. Furthermore, a progressive increase in the HRD score (a measure of "genomic scars") was observed from tubular adenomas to villous adenomas and ultimately to carcinomas. TP53 emerged as the primary driver gene for adenoma-carcinoma transition, with driver mutations consistently appearing simultaneously rather than sequentially acquired from adenomas to carcinomas. Clonal evolution demonstrated that liver metastases can originate from the same cancer-primed cell present in a primary cancerous lesion. We identified a novel pathogenic variant in APC, namely, p.V677Sfs*3. The process of carcinogenesis in FAP-CRC supports the classical cancerization model, where an initial APC mutation leads to the activation of the WNT signaling pathway and CIN. Subsequently, additional mutations occur in other putative CIN genes (e.g., DNA repair, chromatin remodeling), ultimately leading to the development of microsatellite stable (MSS) tumors. Our study provides a comprehensive understanding of the genomic landscapes that underlie the transition from adenoma to carcinoma.

Characterization of Wnt Signaling Pathway Aberrations in Metastatic Prostate Cancer.

Wnt (wingless-type) signaling pathway (WSP) alterations have been identified in patients with prostate cancer and are implicated in disease progression and hormonal resistance. In this study, we utilized a multi-institutional dataset to characterize molecular alterations in the canonical and noncanonical WSPs in prostate cancer. Patients with prostate cancer who underwent tissue-based genomic sequencing were investigated. Tumors with somatic activating mutations in CTNNB1 or RSPO2 or inactivating mutations in either APC or RNF43 were characterized as having aberrant canonical Wnt signaling (WSP-activated). Overall survival analyses were restricted to microsatellite-stable (MSS) tumors lacking RNF43 G659fs* mutations. We also investigated noncanonical WSP by evaluation of ROR1, ROR2, and WNT5 in WSP-activated versus WSP wild-type (WSP-WT) tumors. Of 4,138 prostate cancer samples, 3,684 were MSS. Among MSS tumors, 42.4% were from metastatic sites, of which 19.1% were WSP activated, and 57.6% were from the prostate, of which 10.1% were WSP activated. WSP-activated tumors were more prevalent in metastatic sites than in primary prostate cancer. WSP-activated prostate cancer exhibited more SPOP mutations and higher expression of canonical WSP activators than WSP-WT tumors. ROR1 gene expression was elevated in WSP-activated tumors from both primary and metastatic sites. M2 macrophages predominated the tumor microenvironment in WSP-activated tumors. There was no significant difference in overall survival between patients with WSP-activated and WSP-WT prostate cancer. WSP-activated prostate cancer demonstrated a more immunosuppressed tumor microenvironment and a pronounced upregulation of ROR1 gene expression, underscoring its potential involvement in the crosstalk between canonical and noncanonical WSPs. Implications: Our findings may provide a rationale for developing novel therapeutic strategies targeting Wnt-activated prostate cancer.

Transcriptome-based response predictor to identify potential responders among patients with negative standard markers for response to immune checkpoint blockers.

2666 Background: Immune checkpoint blockers (ICB) are revolutionizing cancer treatment, and are being approved for an increasingly wide range of cancer types. The most common biomarkers currently in use to select patients for ICB are PD-L1 expression on tumor cells, as measured by IHC, and tumor mutational burden (TMB) and microsatellite instability (MSI), both measured by NGS. However, some patients that are negative for these markers still respond to ICB. This calls for complementary biomarkers to better identify responders to ICB, especially in patients that are negative to current biomarkers. Here, we focus on predicting response to anti-PD1 in patients with negative PD-L1, TMB or MSI. Methods: We employ ENLIGHT, our transcriptome-based precision oncology platform, which identifies and utilizes clinically relevant genetic interactions to predict a patient’s response to a wide range of targeted drugs, including ICB. ENLIGHT generates an individual ICB response score calculated from a 10-gene expression signature - the ENLIGHT Matching Score (EMS). Patients with EMS above a predetermined threshold are considered matched by ENLIGHT to anti-PD-1 treatments. We have previously shown, based on more than 1000 cases analyzed retrospectively, that this signature can identify responders to anti-PD-1 with high accuracy. Here, we use ENLIGHT to perform a retrospective analysis of 125 cases from three different datasets, who had low PD-L1 presentation (&lt;1%), low TMB (&lt; 10) or microsatellite stable tumors (MSS), and were treated with anti-PD-1, to specifically assess ENLIGHT’s performance in this biomarker-negative sub-population. Results: Patients who responded to anti-PD-1 treatments had significantly higher EMS in all three datasets. Correspondingly, ENLIGHT is highly predictive of response to anti-PD-1 in patients with negative ICB markers in the three datasets (ROC AUCs of 0.80, 0.84, 0.77, respectively, for low PDL1, low TMB and MSS). It is important to note that ENLIGHT was not trained on any of these datasets, and is applied as-is using previously published parameters. Overall, we find that patients who are ENLIGHT-Matched to anti-PD1 in this biomarker-negative cohort are almost 3 times more likely to respond than patients who are not (31% vs. 11%, p=5e-13). Conclusions: ENLIGHT is a powerful tool for predicting response to anti-PD-1 treatment in patients with negative standard biomarkers for ICB, a currently unmet need with considerable clinical importance.

Predictors of nodal positivity in clinically under-staged patients with colon cancer: A National Cancer Database study and proposal of a predictive scoring system

BackgroundColon cancer pathological and clinical staging may be disoncordant. This study assessed patients with colon cancer in whom the nodal status was clinically understaged. MethodsPatients with stage I-III clinical node-negative colon cancer from the National Cancer Database were included. Regression analyses were conducted to elucidate risk factors for clinical nodal understaging and a scoring system was developed to identify high-risk patients. ResultsThe study included 94,945 patients with 78.4 ​% of patients correctly staged and 21.6 ​% clinically understaged. The predictors of nodal positivity in clinically understaged patients were age <65 (OR 1.43), left-sided tumors (OR 1.41), elevated CEA (OR 2.03), moderately (OR 1.81) or poorly/undifferentiated tumors (OR 3.76), T1 tumors (OR 1.29), signet-ring cell histology (OR 2.26), and microsatellite-stable tumors (OR 1.4). ConclusionPatients with colon cancer and the above factors are more likely to have their nodal status clinically understaged. A scoring system has been developed to identify high-risk patients.

Deep learning for dual detection of microsatellite instability and POLE mutations in colorectal cancer histopathology

In the spectrum of colorectal tumors, microsatellite-stable (MSS) tumors with DNA polymerase ε (POLE) mutations exhibit a hypermutated profile, holding the potential to respond to immunotherapy similarly to their microsatellite-instable (MSI) counterparts. Yet, due to their rarity and the associated testing costs, systematic screening for these mutations is not commonly pursued. Notably, the histopathological phenotype resulting from POLE mutations is theorized to resemble that of MSI. This resemblance not only could facilitate their detection by a transformer-based Deep Learning (DL) system trained on MSI pathology slides, but also indicates the possibility for MSS patients with POLE mutations to access enhanced treatment options, which might otherwise be overlooked. To harness this potential, we trained a Deep Learning classifier on a large dataset with the ground truth for microsatellite status and subsequently validated its capabilities for MSI and POLE detection across three external cohorts. Our model accurately identified MSI status in both the internal and external resection cohorts using pathology images alone. Notably, with a classification threshold of 0.5, over 75% of POLE driver mutant patients in the external resection cohorts were flagged as “positive” by a DL system trained on MSI status. In a clinical setting, deploying this DL model as a preliminary screening tool could facilitate the efficient identification of clinically relevant MSI and POLE mutations in colorectal tumors, in one go.

MITO END-3: efficacy of avelumab immunotherapy according to molecular profiling in first-line endometrial cancer therapy

Immunotherapy combined with chemotherapy significantly improves progression-free survival (PFS) compared to first-line chemotherapy alone in advanced endometrial cancer (EC), with a much larger effect size in microsatellite instability-high (MSI-H) cases. New biomarkers might help to select patients who may have benefit among those with a microsatellite-stable (MSS) tumor. In a pre-planned translational analysis of the MITO END-3 trial, we assessed the significance of genomic abnormalities in patients randomized to standard carboplatin/paclitaxel without or with avelumab. Out of 125 randomized patients, 109 had samples eligible for next-generation sequencing analysis, and 102 had MSI tested. According to The Cancer Genome Atlas (TCGA), there were 29 cases with MSI-H, 26 with MSS TP53 wild type (wt), 47 with MSS TP53 mutated (mut), and 1 case with POLE mutation. Four mutated genes were present in >30% of cases: TP53, PIK3CA, ARID1A, and PTEN. Eleven patients (10%) had a BRCA1/2 mutation (five in MSI-H and six in MSS). High tumor mutational burden (≥10 muts/Mb) was observed in all MSI-H patients, in 4 out of 47 MSS/TP53 mut, and no case in the MSS/TP53 wt category. The effect of avelumab on PFS significantly varied according to TCGA categories, being favorable in MSI-H and worst in MSS/TP53 mut (P interaction= 0.003); a similar non-significant trend was seen in survival analysis. ARID1A and PTEN also showed a statistically significant interaction with treatment effect, which was better in the presence of the mutation (ARID1A P interaction= 0.01; PTEN P interaction= 0.002). The MITO END-3 trial results suggest that TP53 mutation is associated with a poor effect of avelumab, while mutations of PTEN and ARID1A are related to a positive effect of the drug in patients with advanced EC.

Intrahepatic cholangiocarcinoma with FGFR alterations: A series of Chinese cases with an emphasis on their clinicopathologic and genetic features

Intrahepatic Cholangiocarcinoma (iCCA) with FGFR alterations is relatively rare, and its identification is important in the era of targeted therapy. We collected a large series of FGFR-altered cases in the Chinese population and characterized their clinicopathological and genetic features. Among the 18 FGFR-altered cases out of 260 iCCAs, 10 were males and 8 were females, ranging in age from 35 to 74 years (mean, 57.3 years; median, 58 years). Pathologically, they include 9 cases of large duct (LD, 50 %) and small duct (SD, 50 %) types each. All of them (100 %, 18/18) showed microsatellite stable (MSS) and low tumor mutation burden (TMB). Genetically, FGFR alterations involved FGFR1 (20 %), FGFR2 (70 %), and FGFR3 (10 %), with FGFR2 rearrangement accounting for the most (11/18). The most frequently altered genes/biological processes were development/proliferation-related pathways (44 %), chromatin organization (20 %), and tumor suppressors (32 %). Our study further revealed the clinicopathological and genetic features of FGFR-altered iCCA and demonstrated that its occurrence may show regional or ethnic variability and is less common in the Chinese population. A significant number of LD-type iCCA cases also have FGFR alterations rather than the SD type.

Abstract CT217: First-line durvalumab and tremelimumab with chemotherapy in RAS-mutated metastatic colorectal cancer: a phase 1b-2 trial

Abstract Metastatic colorectal cancer (mCRC) remains a difficult-to-treat disease. While patients with microsatellite instable metastatic colorectal cancer (mCRC) benefit from immune checkpoint blockade, chemotherapy with targeted therapies remains the only therapeutic option for microsatellite stable (MSS) tumors. The single arm, phase IB/II MEDITREME trial evaluated the safety and efficacy of durvalumab plus tremelimumab combined with mFOLFOX6 chemotherapy in first line, in 57 patients with RAS-mutant unresectable mCRC. Safety was the primary objective of phase IB; no safety issue was observed. The phase II primary objective of efficacy in terms of 3-month PFS in MSS patients was met, with 3-month PFS of 90.7% [95% CI: 79.2-96%]. For secondary objectives, response rate was 64.5%, median PFS was 8.2 months [95% CI: 5.9-8.6] and overall survival was not reached in MSS patients. We observed higher tumor mutational burden and lower genomic instability in responders. Integrated transcriptomic analysis underlined that high immune signature, and low epithelio-mesenchymal transition were associated with better outcome. Immunomonitoring showed induction of neoantigen and NY-ESO1 and TERT blood tumor specific T cell response, associated with better PFS. The combination of durvalumab-tremelimumab with mFOLFOX6 was tolerable with promising clinical activity in MSS mCRC. Clinicaltrials.gov identifier: NCT03202758. Citation Format: Marion Thibaudin, Jean-David Fumet, Emeric Limagne, Léa Hampe, Susy Daumoine, Valentin Derangère, Caroline Laheurte, Olivier Adotevi, Caroline Truntzer, François Ghiringhelli. First-line durvalumab and tremelimumab with chemotherapy in RAS-mutated metastatic colorectal cancer: a phase 1b-2 trial [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 2 (Late-Breaking, Clinical Trial, and Invited Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(7_Suppl):Abstract nr CT217.

Abstract NG07: Association between somatic microsatellite instability, hypermutation status, and specific T cell subsets in colorectal cancer tumors

Abstract NG07: Association between somatic microsatellite instability, hypermutation status, and specific T cell subsets in colorectal cancer tumors

Abstract 1375: Abundance of KLRB1+ (CD161) T cells in anti-PD1 non responders coupled with enhanced tumor cytotoxicity of anti-CD161 (IMT-009) with anti-PD1 makes it a rational target for combination with anti-PD-(L)1 immunotherapy

Abstract Background: Intra-tumoral CD4+ and CD8+ T cells expressing inhibitory receptor CD161 have an effector memory phenotype with high cytotoxic potential. CD161 inhibits T and NK cell function in the presence of its ligand, CLEC2D. This suppression can be recovered by treatment with IMT-009, a first in class anti-CD161 antibody. Single cell transcriptomic analysis of anti-PD1 refractory microsatellite stable colorectal tumors identified presence of CD161+ CD4+ cytotoxic T cells and CD161+ PD1+ exhausted CD8 T cells suggesting that anti-CD161 treatment may provide benefit in these tumors both as a monotherapy and in combination with anti-PD1. It was also observed that CD161+ CD4/CD8T cells are more abundant in anti-PD1/L1 non responders compared with responders in hepatocellular carcinoma, cutaneous squamous cell carcinoma, clear cell renal carcinoma and breast cancer, providing a rationale for targeting CD161. Using engineered ex-vivo tumor models and dissociated human tumors, we have demonstrated that IMT-009 reverses CLEC2D mediated inhibition and restores primary T cell function. Additionally, co-treatment of IMT-009 with anti-PD1 further enhanced T cell mediated cytotoxicity, providing rationale for studies combining IMT-009 and anti PD1 in the clinic. Methods: Using the Snakemake pipeline developed in house, we processed publicly available scRNAseq data. Cells were clustered by Louvian method followed by cell type annotation by Seurat, and gene expression and gene signature levels per cluster were plotted. For ex-vivo assays, antigen specific CD8 T cells expanded from HLA-A2:01 donors were co-cultured with HLA-A2:01 expressing MDA-MB 231 triple negative breast cancer cell line or Raji Burkitt Lymphoma model with HLA-A2:01 allelic overexpression. Target tumor cells were engineered to express the ligands CLEC2D or PDL1. These cell lines were made to overexpress luciferase for monitoring target cell death using a luminescence readout. Additionally, dissociated tumor cells (DTCs) were cultured in presence of the treatment molecules. Intracellular staining of cytotoxic T cells along with detection of IFNγ, IL-2, TNFα and Granzyme B levels in culture supernatants by MSD/ELISA were used to assess activation profile of tumor infiltrated T cells. Results: Combining anti-PD1 and IMT-009 significantly enhanced cytotoxic benefit compared to individual monotherapy in both Raji and MDA-MB-231 target cell line models. IFNγ levels in supernatants of DTCs treated with this combination were elevated as compared to individual monotherapy. Together with scRNAseq data of increased abundance of KLRB1 (CD161) expressing T cells in anti PD1 non-responder patients, our data provide rationale for testing combination of IMT-009 with anti-PD1 in patients that may not respond to anti PD1 alone. Citation Format: Adwitiya Kar, Tim Nieuwenhuis, Meixue Duan, Siamak Tabrizi, Matthew Huggins, Elizabeth Scanlon, Cynthia Hession, Matthew Bernstein, Michael Ross, Ming Tang, Shruti Malu. Abundance of KLRB1+ (CD161) T cells in anti-PD1 non responders coupled with enhanced tumor cytotoxicity of anti-CD161 (IMT-009) with anti-PD1 makes it a rational target for combination with anti-PD-(L)1 immunotherapy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 1375.

Abstract 7004: DNA Methylation analysis of African American colorectal cancers reveal race-specific alterations

Abstract African Americans (AAs) have among the highest incidence and mortality rates of colorectal cancer (CRC) in the US. They present with more right-sided, microsatellite stable (MSS) tumors and are diagnosed at earlier ages than non-Hispanic Whites (NHW). While DNA methylation changes and their significance have been previously described in CRC, much less is known about the unique DNA methylation changes that occur in AA CRCs compared to NHW CRCs. In the current study, we analyzed DNA methylation changes in AA patients and compared those changes to data from NHWs. AA patients diagnosed with a MSS colorectal tumor at the Yale New Health System were included in the study. A pathologist reviewed archival slides, and selected areas from CRCs (n=160) and adjacent normal tissue (n=42) were cored (median age=59; 82 females, 78 males). DNA was bisulfite converted and analysis performed using EPIC arrays. The 450K methylation array from NHW CRCs was also analyzed for 217 CRCs and 20 normal tissues obtained from TCGA (median age=68; 103 females, 114 males). Differentially methylated regions (DMRs) with |Δβ|&amp;gt;0.1, FDR corrected p-value &amp;lt;0.05 between tumor and normal tissue were identified using SeSAMe. We identified 4881 DMRs (57% hyper- and 44% hypomethylated) between AA CRC and normal tissue. Of these, 46% were in CpG Islands, 15% in CpG Shores, 4% in CpG shelves, and 36% in open sea. Most DMRs (71%) were in promoter regions, with 18% in gene bodies, 5% in enhancer regions, and 6% in intergenic regions. Next, we performed additional DMR analysis using overlapping probes from both arrays to compare NHW with AA data. We identified 2239 DMRs between AA CRC and normal tissue compared to 4138 in NHW. Of the 2239 AA DMRs, 1419 (66%) were common with NHW DMRs, 790 (34%) were unique to AAs. There were many overlapping pathways and genes; however, the DMRs impacting these pathways differed between AA and NHW CRCs. Hypermethylation of LCK, a tyrosine kinase involved in T-cell receptor signaling, was found in 60% of AA CRCs compared to 5.9% of NHW CRCs. LCK hypermethylation leads to CD4/8 T-cell inactivation and linked to poor outcomes in lung cancer. We observed AA-specific hypermethylation of SSTR1 in 57.2% of AA CRCs compared to 27.6% of NHW CRCs. Somatostasin (SST) suppresses tumor growth by regulating certain factors involved in cell proliferation. Hypermethylation of SSTR1 has been associated with various malignancies including gastric cancer. Lastly, promoter hypermethylation of 3 protocadherin genes was found in 65.1% of AAs compared to 33.1% NHWs. Protocadherins regulate WNT signaling and hypermethylation is associated with poor outcomes in multiple cancers. Taken together, these results show AA-specific CRC methylation changes compared with NHW CRCs. Further work is warranted to investigate the potential functional and clinical implications of these methylation alterations, some of which may represent therapeutic targets and biomarkers. Citation Format: Seeta Rajpara, David N. Buckley, Rosa Xicola, Reger Mikaeel, Mary K. Yagle, Baris Kerimoglu, Curtis Thorne, Dante A. Bellomo, Megha Padi, Xavier Llor, Nathan Ellis, Bodour Salhia. DNA Methylation analysis of African American colorectal cancers reveal race-specific alterations [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 7004.