MSI-high Tumors Research Articles

Abstract A052: RNF43 p.G659fs mutation in MSI-high colorectal cancer leads to natural killer cell dysfunction

Abstract RNF43_p.G659fsV*41 mutation (RNF43659mut) is found in approximately 8% of colorectal cancers (CRCs) and is enriched in microsatellite-instability high (MSI-high) tumors. Previously, we reported that RNF43659mut promotes tumor growth independent of WNT signaling and activates PI3K/AKT signaling in CRC, conferring vulnerability to PI3K/MTOR inhibition. In addition, RNF43659mut was associated with decreased interferon signaling suggesting a link with anti-tumor immunity. Thus, in the current work we investigated the role of RNF43659mut in CRC immunomodulation. The immune-cell landscape was evaluated in RNF43659mut compared with RNF43-wild type (wt) MSI-high CRCs in two cohorts; single-cell (sc)-RNA-sequencing data from treatment-naïve primary tumors from 34 patients (cohort-1) and multiplex immunofluorescence / whole exome sequencing data from an independent cohort of 91 patients (cohort-2). Healthy donor blood-derived natural killer (NK) cell responses and cytotoxicity were investigated in co-cultures with RNF43659mut isogenic tumor lines and with patient-derived organoids harboring RNF43659mut or overexpressing RNF43wt. A third patient cohort with MSI-high CRCs treated with immune checkpoint blockade was analyzed to understand the impact of RNF43659mut on immunotherapy efficacy. Interrogation of sc-RNASeq data in cohort-1 revealed a significant enrichment of NK cells in the RNF43659mut patient samples compared to the RNF43wt ones. This finding was also confirmed by multiplex immunofluorescence in cohort-2. Single-cell transcriptomic analyses in cohort-1 showed increased inhibitory markers on NK cells despite their enrichment in RNF43659mut tumors. RNAseq analysis of sorted NK cells co-cultured with RNF43659mut isogenic tumor lines further supported their dysfunctional state. Flow cytometric analyses of NK cell maturation, activation, and inhibitory markers in NK cells co-cultured with RNF43659mut isogenic tumor lines and organoids highlighted decreased levels of NCAM1 (CD56)dim and NCAM1 (CD56)bright subsets, FCGR3A (CD16), and KLRK1 (NKG2D), along with increased levels of KLRC1 (NKG2A), KIR3DL1 (NKB1) and TIGIT inhibitory markers. There was reduced NK cell mediated tumor cell killing in RNF43659mut isogenic tumor lines and organoids. We postulated that aberrant signaling of the PI3K/AKT pathway with RNF43659mut could lead to NK cell dysfunction. Indeed, siRNA-mediated blocking of PI3K/AKT pathway restored the NK-cell dysfunctional phenotype and tumor cell killing in co-cultures with RNF43659mut isogenic tumor lines and organoids. Conversely, overexpression of WT RNF43 in RNF43659mut organoids downregulated PI3K/AKT signaling and increased NK cell cytotoxicity. Lastly, MSI-high CRC tumors harboring RNF43659mut in patients treated with immunotherapy showed less benefit from immune checkpoint blockade. PI3K/AKT activation due to RNF43659mut in MSI-high CRC leads to NK cell dysfunction and reduced NK cell cytotoxicity. This could be responsible for immunotherapy failure, warranting further investigation. Citation Format: Pushpamali De Silva, Samantha Fitzgerald, Jules Cazaubiel, Matan Hofree, Tomotaka Ugai, Juha P Väyrynen, Dane Ford-Roshon, Lishan Fang, Ana Garrido-Castro, Nir Hacohen, Kimmie Ng, Shuji Ogino, Jonathan A Nowak, Marios Giannakis. RNF43 p.G659fs mutation in MSI-high colorectal cancer leads to natural killer cell dysfunction [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Tumor Immunology and Immunotherapy; 2024 Oct 18-21; Boston, MA. Philadelphia (PA): AACR; Cancer Immunol Res 2024;12(10 Suppl):Abstract nr A052.

Role of CD47 gene expression in colorectal cancer: a comprehensive molecular profiling study

BackgroundIn patients with colorectal cancer (CRC), the therapeutic effects of conventional immune checkpoint inhibitors targeting the adaptive immune system are largely limited to those with microsatellite instability-high tumors. Meanwhile, new...

Immune microenvironmental heterogeneity according to tumor DNA methylation phenotypes in microsatellite instability-high colorectal cancers

The detailed association between tumor DNA methylation, including CpG island methylation, and tumor immunity is poorly understood. CpG island methylator phenotype (CIMP) is observed typically in sporadic colorectal cancers (CRCs) with microsatellite instability-high (MSI-H). Here, we investigated the differential features of the tumor immune microenvironment according to CIMP status in MSI-H CRCs. CIMP-high (CIMP-H) or CIMP-low/negative (CIMP-L/0) status was determined using MethyLight assay in 133 MSI-H CRCs. All MSI-H CRCs were subjected to digital pathology-based quantification of CD3 + /CD8 + /CD4 + /FoxP3 + /CD68 + /CD204 + /CD177 + tumor-infiltrating immune cells using whole-slide immunohistochemistry. Programmed death-ligand 1 (PD-L1) immunohistochemistry was evaluated using the tumor proportion score (TPS) and combined positive score (CPS). Representative cases were analyzed using whole-exome and RNA-sequencing. In 133 MSI-H CRCs, significantly higher densities of CD8 + tumor-infiltrating lymphocytes (TILs) were observed in CIMP-H tumors compared with CIMP-L/0 tumors. PD-L1 TPS and CPS in CIMP-H tumors were higher than in CIMP-L/0 tumors. Next-generation sequencing revealed that, compared with CIMP-L/0 tumors, CIMP-H tumors had higher fractions of CD8 + T cells/cytotoxic lymphocytes, higher cytolytic activity scores, and activated immune-mediated cell killing pathways. In contrast to CIMP-L/0 tumors, most CIMP-H tumors were identified as consensus molecular subtype 1, an immunogenic transcriptomic subtype of CRC. However, there were no differences in tumor mutational burden (TMB) between CIMP-H and CIMP-L/0 tumors in MSI-H CRCs. In conclusion, CIMP-H is associated with abundant cytotoxic CD8 + TILs and PD-L1 overexpression independent of TMB in MSI-H CRCs, suggesting that CIMP-H tumors represent a typical immune-hot subtype and are optimal candidates for immunotherapy in MSI-H tumors.

Identifying Actionable Alterations in KRAS Wild-Type Pancreatic Cancer.

The 5-year relative survival rate for pancreatic cancer is currently the lowest among all cancer types with a dismal 13%. A Kirsten rat sarcoma virus (KRAS) gene mutation is present in approximately 90% of patients with pancreatic cancer; however, KRAS-specific drugs are not yet widely used in clinical practice for pancreatic cancer, specifically the KRASG12D variant. Advances in genomic testing revealed an opportunity to detect genetic alterations in a subset of patients with no KRAS mutation termed KRAS wild-type. Patients with KRAS wild-type tumors have a propensity to express driver alterations, hence paving the way for utilizing a targeted therapy approach either via clinical trials or standard-of-care drugs. These alterations include fusions, amplifications, translocations, rearrangements and microsatellite instability-high tumors and can be as high as 11% in some studies. Here, we discuss some of the most notable alterations in KRAS wild-type and highlight promising clinical trials.

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.

WRN Helicase: Is There More to MSI-H than Immunotherapy?

In this issue, Picco and colleagues provide further evidence that WRN inhibitors are synthetically lethal in microsatellite instability-high (MSI-H) cancers and function by blocking the helicase domain of select WRN residues. They demonstrate that WRN inhibitors may be even more effective in a subset of MSI-high tumors with (TA)n repeat expansions, which represents a possible strategy in clinical development. See related article by Picco et al., p. 1457 (1).

RPL22 is a tumor suppressor in MSI-high cancers and a splicing regulator of MDM4

Microsatellite instability-high (MSI-H) tumors are malignant tumors that, despite harboring a high mutational burden, often have intact TP53. One of the most frequent mutations in MSI-H tumors is a frameshift mutation in RPL22, a ribosomal protein. Here, we identified RPL22 as a modulator of MDM4 splicing through an alternative splicing switch in exon 6. RPL22 loss increases MDM4 exon 6 inclusion and cell proliferation and augments resistance to the MDM inhibitor Nutlin-3a. RPL22 represses the expression of its paralog, RPL22L1, by mediating the splicing of a cryptic exon corresponding to a truncated transcript. Therefore, damaging mutations in RPL22 drive oncogenic MDM4 induction and reveal a common splicing circuit in MSI-H tumors that may inform therapeutic targeting of the MDM4-p53 axis and oncogenic RPL22L1 induction.

Molecular features and prognostic factors of locally advanced microsatellite instability-high gastric cancer.

Microsatellite instability-high (MSI-H) tumors are distinct molecular subtypes in gastric cancer. However, a few studies have comprehensively reported the molecular features of MSI-H tumors and their prognostic factors in locally advanced gastric cancer. This study aimed to clarify the molecular features and prognostic factors of locally advanced MSI-H gastric cancer. This study included 499 patients with locally advanced gastric cancer who underwent radical gastrectomy. We evaluated the MSI status and compared with previously published whole-exome sequencing, panel sequencing, and gene expression profiling data. Clinicopathological characteristics and molecular profiles were compared between patients with MSI-H and microsatellite stable (MSS) gastric cancer. A subgroup analysis of survival was performed in patients with MSI-H gastric cancer. MSI-H tumors were detected in 79 of 499 patients (15.8%). MSI-H tumors were associated with an increased tumor mutational burden, MLH1 downregulation, CD274 (PD-L1) upregulation, and enrichment of cell cycle pathways. Among patients with MSI-H gastric cancer, the disease-specific survival (DSS) tended to be better in the surgery plus tegafur, gimeracil, and oteracil potassium (S-1) adjuvant chemotherapy group than in the surgery alone group, especially for stage III patients. Furthermore, DSS was better in the T cell-inflamed gene expression signature-high group, and it tended to be worse in the non-solid type poorly differentiated adenocarcinoma group. The molecular features and prognostic factors of locally advanced MSI-H gastric cancer were clarified. S-1 adjuvant chemotherapy appears to be beneficial, and the T cell-inflamed gene expression signature and histopathological type are prognostic factors in MSI-H tumors.

Progress and prospects of biomarker-based targeted therapy and immune checkpoint inhibitors in advanced gastric cancer.

Gastric cancer and gastroesophageal junction cancer represent the leading cause of tumor-related death worldwide. Although advances in immunotherapy and molecular targeted therapy have expanded treatment options, they have not significantly altered the prognosis for patients with unresectable or metastatic gastric cancer. A minority of patients, particularly those with PD-L1-positive, HER-2-positive, or MSI-high tumors, may benefit more from immune checkpoint inhibitors and/or HER-2-directed therapies in advanced stages. However, for those lacking specific targets and unique molecular features, conventional chemotherapy remains the only recommended effective and durable regimen. In this review, we summarize the roles of various signaling pathways and further investigate the available targets. Then, the current results of phase II/III clinical trials in advanced gastric cancer, along with the superiorities and limitations of the existing biomarkers, are specifically discussed. Finally, we will offer our insights in precision treatment pattern when encountering the substantial challenges.

NCI MATCH (EAY131) arm Z1D: Retrospective evaluation of attributions of adverse events experienced on nivolumab in patients with MSI-high tumors.

e14711 Background: Machine learning algorithms for prediction of Adverse Events (AEs) due to immune checkpoint inhibitors (ICIs) rely on clinically annotated data to serve as gold standards. Clinical trials are the best sources of such data, but audits to assess validity are rare, compromising future machine learning efforts. We reviewed AEs reported on the National Cancer Institute Molecular Analysis for Therapy Choice (NCI-MATCH) EAY-131 Arm Z1D. Patients matched to Arm Z1D due to MSI-high status were treated with nivolumab. Routine and serious AEs were reported to NCI. Attribution data from AEs on 29 patients with tumor genomics suitable for machine learning approaches were obtained and reviewed retrospectively. Methods: Documented AEs were sorted by grade via the Common Terminology for Adverse Events (CTCAE) system and categorized as related or unrelated to ICIs. Primary source data reported to NCI were reviewed. If any ambiguity was found, trial sites were queried to provide additional information and documentation. Decisions were made by an experienced NCI medical officer with the benefit of additional data and site responses. All serious AEs and a subset of ambiguous routine AEs were assessed to determine if attribution conformed to expected attribution per the nivolumab package insert. Uncertain attribution required additional queries, including email or phone correspondence. Grade 1 or 2 events were only evaluated to determine if any attribution to ICIs was possible. If a higher grade event was attributed to ICIs, there was no further query for lower grade events on the same patient. Results: A total of 639 Grade 1, 152 Grade 2, 75 Grade 3, 10 Grade 4, and 5 Grade 5 events were reported among the 29 patients. One Grade 5 event, 5 Grade 4 events, 13 Grade 3 Events, 9 Grade 2 events, and 6 Grade 1 events were considered less likely to be attributed to ICIs compared with the initial sponsor or site investigator assessment of possibly, probably, or definitely related. Two Grade 4 events, 1 Grade 2 event, and 8 Grade 1 events were considered to be possibly, probably, or definitely due to ICIs compared with the initial sponsor or site assessment of less likely or unrelated. Site responsiveness to retrospective queries from the sponsor varied. Additional lab values, primary source documentation, and communication with site investigators were necessary to make modifications. A total of 11 AEs required additional queries to site investigators. Conclusions: Retrospective reviews of real-time site and sponsor assessments of attribution exhibit significant modification of initial attribution to ICIs in this review of 29 patients on Arm Z1D of the NCI MATCH trial. Machine learning approaches for prediction of AEs on patients treated with ICIs may require careful assessments of primary source data to develop adequate training sets, given the paucity of gold standard data available.

Phase 1/2 randomized, open-label, multicenter, Simon two-stage study of pelareorep combined with modified FOLFIRINOX +/- atezolizumab in patients with metastatic pancreatic ductal adenocarcinoma.

TPS4203 Background: Immune therapies benefit only the small subset of metastatic pancreatic ductal adenocarcinoma (mPDAC) patients who have microsatellite instability-high (MSI-H) tumors (approximately 1%). Most PDAC tumors are microsatellite stable (MSS) and have an immunologically “cold” phenotype with fewer genetic mutations, reduced immune cell infiltration, and downregulated immune checkpoint proteins. These attributes make most mPDAC tumors resistant to conventional immunotherapy including checkpoint inhibitors. Pelareorep is a naturally occurring, non-genetically modified reovirus that selectively infects cancer cells following intravenous administration. Upon infection, pelareorep’s double-stranded RNA genome stimulates a pro-inflammatory response that primes tumors for immunologic killing. This includes increased T cell infiltration into tumors, expansion of tumor-infiltrating lymphocyte clones, increased PD-L1 expression, and stimulation of tumor-directed innate and adaptive immune responses. Pelareorep combined with chemotherapy, checkpoint inhibitors, or both has shown promising efficacy in several malignancies, including mPDAC. This study is designed to evaluate pelareorep plus modified FOLFIRINOX with or without the PD-L1 inhibitor atezolizumab as first-line therapy in patients with mPDAC. Methods: This trial represents a new cohort within the ongoing GOBLET signal finding platform study in gastrointestinal cancers. This is a phase 1/2 randomized, open-label study that uses a screened selection design within a Simon two-stage approach. Eligible patients must be 18 years or older and have newly diagnosed, radiologically evaluable mPDAC with no prior chemotherapy and an ECOG performance status of 0 or 1. Patients are randomized 1:1 to receive either pelareorep + mFOLFIRINOX orpelareorep + mFOLFIRINOX + atezolizumab. The first stage enrolls 15 evaluable patients into each arm. If pre-specified efficacy criteria are met in Stage 1, one or both arms may be expanded to Stage 2 and enroll 17 additional evaluable patients per arm. The first 3-6 patients enrolled into both arms comprise a safety run-in that must be successfully concluded prior to enrolling additional patients. The treatment period consists of 4-week treatment cycles. The primary objectives are 1) safety and tolerability of pelareorep + mFOLFIRINOX with or without atezolizumab and 2) efficacy based on objective response rate per RECIST v1.1. Secondary objectives include duration of response, progression-free survival, and overall survival. In addition, tumor and blood samples are being collected to evaluate the immunological effects of treatment and to explore potential biomarkers of response. Clinical trial information: Eudra-CT: 2020-003996-16.

Immune checkpoint inhibitor-associated gastrointestinal adverse events in patients with colorectal cancer.

e14708 Background: Immune checkpoint inhibitors (ICI) are successfully used to treat microsatellite instability-high (MSI-H) tumors. While immune checkpoint inhibitor related colitis is a frequent and devastating immune related adverse event (irAE) of these agents, the incidence and characteristics of this inflammatory toxicity in patients with MSI-H colorectal cancers has not been examined. We aimed to describe the characteristics and clinical profile of patients diagnosed with luminal gastrointestinal irAE in patients treated with ICI for colorectal cancer in a tertiary cancer care center. Methods: This is a retrospective analysis that included adult cancer patients diagnosed with colorectal cancer that received ICI between 6/1/2014 and 12/31/2022. We report data on those that developed colitis as an irAE up to 3 months after the last dose of ICI confirmed by laboratory and/or imaging report. We included patients’ demographic characteristics, oncologic profile and outcomes as well as clinical course, endoscopic features as well as treatment and outcomes in terms of luminal gastrointestinal irAEs. Results: Out of 474 patients with colorectal cancer on ICI in our study period, only 18 developed a gastrointestinal irAE with an incidence of 3.8%. Patients were primarily Caucasian (88.8%) males (61.1%) with a median age of 69.5 years. The majority of these patients received combination therapy with anti-PD1/L1 and CTLA-4 (50%). 66.6 % received ICI for MSI-H colorectal cancer. 11.1% of our sample were noted to have a second cancer-melanoma. The major-ity of patients had grade 1-2 colitis (61.2%) and grade 1-2 diarrhea (88.8%). Only 5 patients underwent endoscopic evaluation of whom, 2 had ulcerative inflammation necessitating use of selective immunosuppressive therapy with biologics. 61.1% had to withhold cancer treatment due toxicity. With regards to other gastrointestinal irAEs among this population, 41.4% and 5.8% were noted to have liver and pancreas toxicity respectively, with a median CTCAE grade of severity 2. The most common treatment for these toxicities were steroids. Conclusions: Luminal gastrointestinal irAEs seem to occur less frequently and at a lower severity among patients with MSI-H colorectal cancer after checkpoint inhibitors exposure compared to the overall incidence of the same among other cancers reported in literature. Larger prospective studies are necessary to determine the role of tumor biology and the gut microbiome in the disease profile and severity of immune related adverse events of the GI organ system. [Table: see text]

Contrasting comprehensive genomic profiles of adenocarcinomas of the appendix in younger versus older patients.

3626 Background: Adenocarcinoma of the appendix (AAC) is a rare form of intestinal malignancy that can pursue an aggressive clinical course. Little is known about the clinical and biologic features of this disease when it presents in young patients under 40. Methods: 1,840 cases of clinically advanced AAC were selected and analyzed by hybrid capture based comprehensive genomic profiles (CGP) to evaluate all classes of genomic alterations (GA). MSI high status and tumor mutation burden (TMB) were determined from sequencing results and PD-L1 was measured by immunohistochemistry (Dako 22C3). Results: 135 (7.3%) of the AAC were identified in patients of 39 years of age or younger (“younger”). The mean age of the younger patients was 32.9 years and for the 40 and over (“older”) patients it was 60.4 years. The slight female gender preponderance was higher in the older patients than the younger patients (Table). The GA per sample were similar. The younger patients had a significantly higher frequency of KRAS GA (71.1% vs 60.4%; p=.017) and TP53 GA (56.3% vs 41.3%; p=.0008). The KRAS G12C mutation frequency was similarly low in both cohorts. Targetable GA in BRAF and ERBB2 were infrequent in both groups as were GA in PIK3CA, PTEN and FBXW7. GA in the APC gene was significantly more frequent in the older patients (10.8% vs 3.7%; p=.007). Biomarkers of immune-oncology (IO) drug efficacy were infrequently identified in both groups including MSI-High status, TMB levels and PD-L1 expression. Conclusions: At a 7.3% incidence, AAC is a relatively infrequent form of gastrointestinal malignancy in young patients. The data analysis shows significant differences in the genomics found in younger and older patients suffering from this disease. These results can have a crucial impact on current and future clinical trials used to design novel treatments. Further exploration into this subject can change the overall approach and even treatment guidelines for this type of cancer. [Table: see text]

Retrospective analysis of change in frequency of STK11 mutation in lung adenocarcinomas over a 10-year period.

8032 Background: Previous studies have linked the presence of inactivating mutations in the tumor suppressor gene STK11 with reduced efficacy of immune checkpoint inhibitor (ICI) treatments in lung adenocarcinoma (LA). We queried whether there has been a change in the STK11 mutation frequency over the last 10-year period that could potentially reflect the emergence of additional acquired resistance mutations. Methods: Over a 10-year period from 2013 through 2022, 109,763 clinically advanced LA underwent hybrid capture based comprehensive genomic profiling (CGP) to assess all classes of genomic alterations (GA) at a single reference laboratory (Foundation Medicine). MSI-high status and tumor mutational burden (TMB) were determined from the sequencing data. PD-L1 status was measured by immunohistochemistry (IHC) using the Dako 22C3 kit with TPS scoring. Statistical comparisons utilized the 2-tailed Chi Square method with the Yates’ correction. Results: The frequency of STK11 inactivating GA in all cases of LA significantly increased during the study period from 2013 to 2022 (16.1% vs 20.1%; p=0.0005). This increase appeared to be predominantly in KRAS wild type LA (10.1% vs 15.5%; p<0.0001). The STK11 GA frequency slightly declined in the KRAS mutated LA over the 10-year period (29.9% vs o 27.0%, P=0.25). During the same time period, the KEAP1 GA frequency increased from 12.6% to 14.9% (p= 0.076) and the frequency of combined STK11 and KEAP1 GA increased from 4.8% to 8.0% (p=.0006). The frequencies of MSI-high status, TMB and PD-L1 expression did not significantly change over the observation period. Conclusions: The continuously increasing approved indications for ICI-based treatments for both early stage and clinically advanced LA appears to be accompanied by a significant increase in the frequency of inactivating STK11 GA predominantly in the KRAS mutation negative LA population. This inactivation is associated with an attenuated response rate and progression-free survival in this population. Given the development of novel therapies focused on potential restoration of STK11 function by HDAC/CoREST inhibition, further study of the STK11 GA distribution in LA appears warranted. [Table: see text]

A machine learning algorithm based on multi-omics biomarkers for the detection of tumor microsatellite instability.

1554 Background: Microsatellite instability (MSI) occurs via defects in mismatch repair (MMR) and is characterized by high tumor mutational burden (TMB) and a robust inflammatory response. Accurate MSI-high classification of immune-evasive tumors is vital due to the effectiveness of immune checkpoint inhibitor (ICI) therapy, such as pembrolizumab, which is FDA approved for all MSI-high tumors irrespective of anatomic origin. Detection methods of MSI-status vary widely, including immunohistochemistry (IHC), polymerase chain reaction (PCR), and next-generation sequencing (NGS). We developed a machine learning (ML) model to predict MSI status in patients with solid tumors using comprehensive genomic and immune profiling (CGIP), independent of direct sequencing data from microsatellite sites. Methods: We analyzed samples from 1,838 patients with colorectal cancer (CRC) by CGIP, which included DNA panel testing (523 genes) for pathogenic single nucleotide variants (pSNV) and determination of TMB (mut/Mb), and RNA sequencing (397 genes) for gene expression (GEX). The Boruta algorithm was used to select key genomic and GEX changes associated with MSI status. A distributed gradient boosting algorithm created a predictive model that was trained on 70% of the cohort. The model was tested on the remaining CRC cohort (Test), and in the PanCancer Atlas CRCs (TCGA), endometrial adenocarcinomas (EMCA), and CRC with indeterminate MSI testing by CGIP. Performance of the trained model was assessed using sensitivity, specificity, and positive (PPV) and negative (NPV) predictive value. Results: Feature selection identified 79 genes with pSNVs, 63 GEX changes, and TMB as informative for MSI prediction. The model showed high predictive accuracy in differentiating MSI-H tumors. Of the 39 cases that failed MSI component of CGIP (Indeterminate), 17 had MMR IHC results available, of which 1 demonstrated loss of MLH1/PMS2 and the other 16 showed intact (normal) expression. Testing of this cohort identified 3 cases as MSI-high, at least one of which was confirmed by IHC. No cases with intact MMR protein IHC were identified as MSI-high by the algorithm. Conclusions: Our ML-driven approach accurately assessed MSI status of CRC and EMCA using CGIP data. This approach also identified potential cases with MSI-high status where direct sequencing of microsatellites failed. This study highlights a method to identify patients with potential MSI-high status for orthogonal screening with the MSI component of a test fails. [Table: see text]

Clinical significance of combined tumour-infiltrating lymphocytes and microsatellite instability status in colorectal cancer: a systematic review and network meta-analysis

Microsatellite instability (MSI) status and tumour-infiltrating lymphocytes (TIL) are established prognostic factors in colorectal cancer. Previous studies evaluating the combination of TIL and MSI status identified distinct colorectal cancer subtypes with unique prognostic associations. However, these studies were often limited by sample size, particularly for MSI-high (MSI-H) tumours, and there is no comprehensive summary of the available evidence. We aimed to review the literature to compare the survival outcomes associated with the subtypes derived from the integrated MSI-TIL classification in patients with colorectal cancer. In this systematic review and network meta-analysis, we searched PubMed, Embase, Scopus, and the Cochrane Library without language restrictions, for articles published between Jan 1, 1990, and March 13, 2024. Patient cohorts comparing different combinations of TIL (high or low) and MSI status (MSI or microsatellite stable [MSS]) in patients with surgically resected colorectal cancer were included. Studies were excluded if they focused on neoadjuvant therapy or on other immune markers such as B cells or macrophages. Methodological quality assessment was done with the Newcastle-Ottawa scale; data appraisal and extraction was done independently by two reviewers. Summary estimates were extracted from published reports. The primary outcomes were overall survival, disease-free survival, and cancer-specific survival. A frequentist network meta-analysis was done to compare hazard ratios (HRs) and 95% CI for each outcome. The MSI-TIL subgroups were prognostically ranked based on P-score, bias, magnitude, and precision of associations with each outcome. The protocol is registered with PROSPERO (CRD42023461108). Of 302 studies initially identified, 21 studies (comprising 14 028 patients) were included in the systematic review and 19 (13 029 patients) in the meta-analysis. Nine studies were identified with a low risk of bias and the remaining ten had a moderate risk of bias. The MSI-TIL-high (MSI-TIL-H) subtype exhibited longer overall survival (HR 0·45, 95% CI 0·34-0·61; I2=77·7%), disease-free survival (0·43, 0·32-0·58; I2=61·6%), and cancer-specific survival (0·53, 0·43-0·66; I2=0%), followed by the MSS-TIL-H subtype for overall survival (HR 0·53, 0·41-0·69; I2=77·7%), disease-free survival (0·52, 0·41-0·64; I2=61·6%), and cancer-specific survival (0·55, 0·47-0·64; I2=0%) than did patients with MSS-TIL-low tumours (MSS-TIL-L). Patients with the MSI-TIL-L subtype had similar overall survival (0·88, 0·66-1·18; I2=77·7%) and disease-free survival (0·93, 0·69-1·26; I2=61·6%), but a modestly longer cancer-specific survival (0·72, 0·57-0·90; I2=0%) than did the MSS-TIL-L subtype. Results from the direct and indirect evidence were strongly congruous. The findings from this network meta-analysis suggest that better survival was only observed among patients with TIL-H colorectal cancer, regardless of MSI or MSS status. The integrated MSI-TIL classification should be further explored as a predictive tool for clinical decision-making in early-stage colorectal cancer. German Research Council (HO 5117/2-2).

Identifying modifiable risk factors to prevent aggressive colorectal cancer.

It remains unclear if pre-diagnostic factors influence the developmental pathways of colorectal cancer (CRC) that could enhance tumor aggressiveness. This study used prospective data from 205,489 cancer-free US health professionals to investigate the associations of 31 known or putative risk factors with the risk of aggressive CRC. Tumor aggressiveness was characterized by three endpoints: aggressive CRC (cancer that causes death within 5 years of diagnosis), fatal CRC, and tumor stage at diagnosis. The data augmentation method was used to assess the difference in the associations between risk factors and endpoints. We documented 3201 CRC cases, of which 899 were aggressive. The protective associations of undergoing lower endoscopy (hazard ratios [HR] 0.43, 95% confidence interval (CI) 0.37, 0.49 for aggressive versus HR 0.61, 95% CI 0.56, 0.67 for non-aggressive) and regular use of aspirin (HR 0.70, 95% CI 0.61, 0.81 versus HR 0.84, 95% CI 0.77, 0.92) were stronger for aggressive than non-aggressive CRC (pHeterogeneity <0.05). Lower intake of whole grains or cereal fiber and greater dietary inflammatory potential were associated with a higher risk of aggressive but not non-aggressive CRC. The remaining risk factors showed comparable associations with aggressive CRC and non-aggressive CRC. Aggressive cases were more likely to have KRAS-mutated tumors but less likely to have distal or MSI-high tumors (p < .007). Similar results were observed for fatal CRC and advanced tumor stages at diagnosis. These findings provide initial evidence for the role of pre-diagnostic risk factors in the pathogenesis of aggressive CRC and suggest research priorities for preventive interventions.

Advancing Personalized Medicine in the Treatment of Locally Advanced Rectal Cancer.

Rectal cancer presents a significant burden globally, often requiring multimodal therapy for locally advanced cases. Long-course chemoradiotherapy (LCRT) and short-course radiotherapy (SCRT) followed by surgery have been conventional neoadjuvant approaches. Recent trials favor LCRT due to improved local control. However, distant tumor recurrence remains a concern, prompting the exploration of total neoadjuvant therapy (TNT) as a comprehensive treatment strategy. Immune checkpoint inhibitors (ICIs) show promise, particularly in mismatch repair-deficient (dMMR) or microsatellite instability-high (MSI-H) tumors, potentially revolutionizing neoadjuvant regimens. Nonoperative management (NOM) represents a viable alternative post-neoadjuvant therapy for selected patients achieving complete clinical response (cCR). Additionally, monitoring minimal residual disease (MRD) using circulating tumor DNA (ctDNA) emerges as a non-invasive method for the assessment of treatment response. This review synthesizes current evidence on TNT, ICIs, NOM, and ctDNA, elucidating their implications for rectal cancer management and highlighting avenues for future research and clinical application.

Somatic mutations that affect early genetic progression and immune microenvironment in gastric carcinoma

Gastric carcinoma (GC) is a high heterogeneity and malignant tumor with a poor prognosis. The current implementation of immunotherapy in GC is limited due to the insufficient exploration of immune-related mutations and speculated early mutation events. Therefore, we performed whole-exome sequencing on 40 patients with GC to explore their genetic characteristics, shedding light on the order of genetic events, somatic mutations impacting the immune microenvironment, and potential biomarkers for immunotherapy. Regarding genetic events, TP53 disruptions were identified as frequent and early events in GC progression, often occurring alongside other gene mutations. The mutations occurring in GANS, SMAD4, and POLE were early independent events. Patients harboring CSMD3, FAT4, FLG, KMT2C, LRP1B, MUC5B, MUC16, PLEC, RNF43, SYNE1, TP53, TTN, XIRP2, and ZFHX4 mutations tended to have decreased B cells, T cells, macrophage, neutrophil, and dendritic cells infiltration, except for the ARID1A gene mutations. We also found patients with microsatellite instability-high tumors had higher homologous recombination deficiency (HRD) scores. HRD showed a positive correlation with tumor mutational burden, which might serve as indirect evidence supporting the potential of HRD as a biomarker for GC. These findings highlighted GC's high heterogeneity and complexity and provided valuable insights into the somatic mutations that affect early genetic progression and immune microenvironment.

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