CDH1 Mutations Research Articles

The Gastric Cancer Registry: A multi-omic cellular and molecular resource for cancer biomarker and therapeutic discovery.

491 Background: The Gastric Cancer Registry (GCR) is a comprehensive clinical and genomic data resource focused on gastric cancer (GC) patients and individuals with a family history of GC or a germline CDH1 mutation. As part of the GCR’s ongoing enrollment, patients contribute cancer history information, archival tumor samples and other biospecimens. The cancer samples undergo extensive molecular and cellular analysis that includes genomic sequencing and spatial analysis. We have generated a robust dataset, publicly accessible online through the GCR Genome Explorer (GCR-GE). This genomic, molecular and cellular resource provides a rich data set that accelerates multidisciplinary research aimed at improving detection and treatment strategies for GC. GC disproportionately affects Latin American populations - we have included new cohorts from Latin America. The GCR enables researchers, clinicians and patients to address specific questions about the molecular and cellular basis of GC. Methods: GC samples underwent whole genome, whole exome, and bulk RNA sequencing were conducted. The genomic features identified include copy number variation, gene mutations, gene expression, microbiome composition, estimation of tumor-infiltrating immune cells, tumor neoantigens, MSI/MSS status, and HLA subtypes. In addition to genomic data, clinical data was compiled from survey responses and pathology reports and integrated into the GCR-GE. We have incorporated single cell and spatial analysis that includes diverse cell types, single cell gene expression and cellular architecture in native tumor tissues. This data was released to the GCR Genome Explorer website. Results: A total of 817 subjects have enrolled in the GCR including 598 diagnosed with GC and 219 at high-risk for GC through family history of GC and/or a CDH1 mutation. Of the 598 with GC, some met multiple criteria including at least 40 with a family history of GC, 10 with a CDH1 mutation, and 18 with both a family history of GC and a CDH1 mutation. The GCR-GE includes data from 257 gastric tumors in addition to external datasets from TCGA. In the latest data release, clinical and genomic data from 78 gastric tumors from patients in Latin America were integrated into the GCR-GE. Conclusions: The GCR is a comprehensive database of crucial clinical and genomic data necessary for driving forward GC research. Through global expansion, the GCR has generated a more representative dataset by increasing racial diversity and including underrepresented populations that are at higher risk for GC.

Molecular profiling of a gastroesophageal adenocarcinoma (GEA) multicohort using next-generation sequencing (NGS).

494 Background: The mutational landscape of GEA is rapidly evolving with the identification of promising molecular targets. However, histological subtypes are insufficient to capture the molecular heterogeneity in these patients (pts). We aim to assess the histopathological and molecular features in a GEA cohort using NGS techniques, and to evaluate the mutational frequency and its prognostic correlation. Methods: We conducted a retrospective study on GEA from 2019 to 2024. Immunohistochemistry (IHC) was used to evaluate HER2 expression, microsatellite instability (MSI), Epstein-Barr virus (EBV) and PD-L1 status. In-house and commercial NGS platforms were used to detect molecular alterations from tumor samples. Cox regression model was used to analyze overall survival (OS) based on the clinicopathological and molecular subgroups. Results: Using NGS techniques, we identified the following alterations with potential clinical relevance in the 115-pts included: pathogenic mutations in PIK3CA (5.2%), BRCA1/2 (3.5%) and POLD1 (0.9%), along with copy number alterations (CNA) in ERBB2 (10.4%), EGFR (8.7%), MET (5.2%) and FGFR2 (4.3%) genes, among others. Additionally, a high tumor mutational burden (≥ 13 Mut/Mb) was observed in 6 pts (5.2%). Clinical characteristics are detailed in the table. We detected a higher rate of alterations in CDH1 (10.42% vs 4.48%; p=0.22) and PIK3CA (8.33% vs 2.99%; p=0.20) genes in early (<50y) vs late-onset (≥50y) GEA. Additionally, we identified a significantly higher mutation frequency in diffuse vs intestinal tumors; ARID1A (17.6% vs 2.2%; p=0.01) and CDH1 (11.8% vs 0%; p=0.02) genes. Also, pts with peritoneal involvement showed a greater prevalence of mutations in CDH1 (12% vs 0%; p=0.04) and RHOA (10% vs 0%; p=0.06) compared to those with liver disease. We observed a significantly worse OS in pts with diffuse vs intestinal tumors (15.7m vs 19.1m; p=0.04) and a trend toward shorter survival in pts with CDKN2A mutations (11.97m vs 17.83m; p=0.42). Although not significantly, CNA in ERBB2 were associated with a better prognosis (22.2m vs 16.2m; p=0.09). Conclusions: Our results support the utility of NGS platforms in detecting novel molecular targets with prognostic and clinical impact, beyond ERBB2 . Additionally, we observed poorer prognosis in diffuse subtype tumors, which exhibited a higher frequency of mutations in ARID1A and CDH1 genes. Clinicopathological characteristics of the 115-pts with GEA. N=115 (%) AgeMedian years [range] 54 [18-83] Sex Male 69 (60.0) Tumor site Gastric 72 (62.6) Gastroesophageal junction 37 (32.2) Esophagus 6 (5.2) Histological subtype Diffuse or pooly cohesive 51 (44.3) Intestinal or tubular 46 (40.0) Others 18 (15.7) Stage at diagnosis Advanced 74 (66.9) Localized 41 (33.1) Metastatic site Peritoneum 53 (46.1) Liver 34 (29.6) Molecular profile (IHC) HER2 (+) 20 (17.4) MSI 5 (4.3) EBV (+) 2 (1.8) PDL1 (CPS) ≥1 35 (30.4)

Phase 2 trial of defactinib in combination with avutometinib in patients with advanced diffuse-type gastric cancer.

TPS505 Background: Diffuse-type gastric cancer (DGC) accounts for approximately one third of gastric cancer diagnoses. DGC is characterized by poor differentiation, discohesive growth, frequent peritoneal spread, chemoresistance, and inferior survival compared to intestinal-type gastric cancer. Moreover, DGC is commonly genomically stable and often lacks clinically actionable targets such as ERBB2 amplification. Alterations enriched in DGC including loss-of-function CDH1 mutations and gain-of-function RHOA mutations can facilitate increased focal adhesion kinase (FAK) signaling that drives tumor proliferation, invasiveness, and metastasis. Recent preclinical studies demonstrated that the FAK inhibitor defactinib can inhibitor DGC growth in vitro and in vivo . FAK inhibitor monotherapy leads to compensatory mitogen-activated protein kinase (MAPK) pathway upregulation, which can be overcome with combined FAK and MEK inhibition. In preclinical DGC xenograft models, defactinib plus the RAF/MEK clamp avutometinib led to deep tumor regressions, suggesting synergistic activity of the combination. The goal of this study is to investigate the efficacy and tolerability of avutometinib plus defactinib in patients with advanced DGC refractory to standard therapy. Methods: This investigator-initiated, multicenter phase 2 trial will enroll 27 patients with metastatic/unresectable gastric or gastroesophageal junction carcinoma classified as diffuse, poorly cohesive, signet ring cell, or mixed type with progression on at least one line of therapy including platinum/fluorouracil chemotherapy. Patients with known pathogenic CDH1 and/or RHOA alterations regardless of histology are also eligible. Patients must have ECOG PS 0/1 and can have either measurable or evaluable non-measurable disease per RECIST 1.1 criteria. Avutometinib is given 3.2 mg orally twice weekly and defactinib is given 200 mg orally twice a day, both for 3 weeks on/1 week off for each 28-day cycle. The primary endpoint is 6-month PFS rate, with secondary endpoints including ORR, OS, DOR, and tolerability. Exploratory studies will interrogate FAK/MAPK pathway activation, ctDNA dynamics, and the impact of the combination of avutometinib/defactinib on the tumor microenvironment from pre- and on-treatment biopsies. The trial was opened for enrollment in August 2024. Clinical trial information: NCT06487221 .

Molecular characteristics of early-onset versus average-onset gastroesophageal adenocarcinoma.

496 Background: Incidence of early-onset gastroesophageal adenocarcinoma (eoGEA, ages <50 years) has been increasing in the United States since the 1990s, the etiology of which is still unclear, and limited data exists on molecular drivers. This study evaluates somatic and germline profiles in eoGEA compared to average-onset GEA (aoGEA, ages≥ 50 years). Methods: This is a retrospective, cross-sectional study utilizing data from de-identified records of GEA (esophageal, gastroesophageal junction and gastric adenocarcinomas) patients who underwent somatic NGS testing via the Tempus xT assay (595-648 gene DNA panel) from 12/2017 to 07/2024. This assay assesses somatic tumor mutations (including SNVs, Indels, CNVs, and select SVs), MSI, TMB, and incidentally detected germline SNVs and small indels in matched normal tissue. Clinical characteristics and immunotherapy markers were compared between eoGEA and aoGEA by Wilcoxon Rank Sum or Chi-squared test. Somatic and germline mutations were compared between two age groups with false discovery rate adjustments. Results: We compared the demographic, clinical, and molecular features of a total of 5863 patients with eoGEA (n=785, median age 43) and aoGEA (n=5078, median age 68). Over 80% of patients were diagnosed with stage IV disease. The eoGEA group had a higher proportion of females, non-white, and Hispanic or Latino patients compared to the aoGEA group (p<0.001). Prevalence of MSI-H/ dMMR, and TMB-H was lower in eoGEA vs aoGEA (p<0.001), while PD-L1 expression was similar. Prevalence of somatic CDH1 , CDKN2A , and TP53 SNVs, and FGFR2 and KRAS CNAs were higher in eoGEA vs aoGEA (q<0.001). In a subset of 3286 patients with tumor/normal match testing (eoGEA=464 and aoGEA=2822), eoGEA also had a higher prevalence of germline CDH1 mutations (q<0.001). Conclusions: eoGEA has a unique somatic and germline mutation profile compared to aoGEA; further study evaluating the molecular landscape including epigenetic changes could shed light on underlying mechanisms responsible for the rising incidence of eoGEA. eoGEA (n=785) aoGEA (n=5078) p-value/q-value* Baseline characteristics Median age at diagnosis (IQR) 43 (38, 47) 68 (61,74) <0.001 Gender: male 509 (65%) 3798 (75%) <0.001 Race (white) 280 (69%) 2347 (80%) <0.001 Ethnicity (not hispanic or latino) 206 (60%) 1768 (86% <0.001 Stage IV 495 (85%) 2883 (81%) 0.073 Immunologic markers TMB ≥ 10 25 (3.2%) 519(10%) <0.001 MSI-H 13 (1.7%) 261 (5.1%) <0.001 dMMR 7 (2.2%) 120 (5.9%) 0.007 Somatic mutations profiles * CDH1 126 (16%) 334 (6.6%) <0.001 TP53 515 (66%) 3764 (74%) <0.001 CDKN2A 101 (13%) 1011 (20%) <0.001 KRAS 92 (12%) 931 (18%) <0.001 NOTCH1 12 (1.5%) 228 (4.5%) 0.002 Germline mutation profiles * eoGEA (N=464) aoGEA (N= 2,822) Overall prevalence 46 (9.9%) 207 (7.3%) CDH1 10 (2.2%) 8 (0.3%) 0.001 TP53 3 (0.6%) 0 (0%) 0.039 BRCA2 2 (0.4%) 38 (1.3%) 0.6 BRIP1 4 (0.9%) 10 (0.4%) 0.6

Genomic Analysis Reveals Racial and Age-Related Differences in the Somatic Landscape of Breast Cancer and the Association with Socioeconomic Factors.

Cancer genomics consortia have identified somatic drivers of breast cancer subtypes. However, these studies have predominantly included older, non-Black women, and the related socioeconomic status (SES) data is limited. Increased representation and depth of social data are crucial for understanding how health inequity is intertwined with somatic landscapes. Here, we conducted targeted sequencing on primary tumors from the Carolina Breast Cancer Study (N = 357; 52% Black, 47% <50) and compared the results to The Cancer Genome Atlas (N = 948; 18% Black, 27% <50). Race (Black vs. non-Black), age, and SES were evaluated in association with mutations, copy number alterations, and aneuploidy using generalized linear models. Pathway dysfunction was also assessed by aggregating mutation and copy number alterations. Adjusting for age, Black participants (N =350) were significantly more likely to have TP53 and FAT1 mutations and less likely to have PIK3CA, CDH1, DDR2, and GATA3 mutations than non-Black participants. Younger participants had more GATA3 alterations and fewer KMT2C, PTEN, MAP3K1 and CDH1 alterations. Black participants had significant enrichment for MYC (8q) and PIK3CA (3q26) amplifications and higher total aneuploidy, but age was not associated with copy number variation. SES was associated with different patterns of alteration in Black versus non-Black women. Overall, Black participants showed modest differences in TP53, PIK3CA, and other alterations that further varied by SES. Race is a social construct, and varying distributions of etiologic factors across social strata may predispose Black, young, and low SES women to cancer subtypes characterized by these alterations.

Outcomes of minimally invasive and open prophylactic gastrectomy for hereditary diffuse gastric cancer

BackgroundMutations in the CDH1 gene predispose individuals to hereditary diffuse gastric cancer. As these tumors can evade endoscopic screening, prophylactic total gastrectomy is often recommended. Since skill with minimally invasive surgery (MIS) has progressed, we compared CDH1 mutation carriers who underwent open vs MIS total gastrectomy. MethodsA retrospective review of 48 CDH1 carriers who underwent total gastrectomy from May 2004 to April 2023 was performed. Eight patients were excluded because they were symptomatic prior to surgery and had advanced signet ring cell adenocarcinoma. ResultsTwenty-eight open and 12 MIS total gastrectomy patients were included; one MIS case was converted to open. The groups were comparable regarding age, comorbidities, and pre-operative carcinoma identified (42 % vs 36 %). Blood loss was lower with MIS gastrectomy (200 vs 23 mL) while operative time was longer (163 vs 286 minutes). The number of lymph nodes harvested (18 vs 23) and the percentage with carcinoma (86 % vs 92 %) were not different between open and MIS approaches. Length of stay was shorter after MIS gastrectomy (7 vs 5 days). In the MIS group, there were no major post-operative complications (2.5 % open) or readmissions within 90 days (11 % open). Subsequent surgery or dilation was infrequent (18 % vs 8 %). Less weight loss was seen after MIS gastrectomy, reaching significance at 9 months post-operatively (-25 % vs −13 %). ConclusionsMIS total gastrectomy is the preferred operation for CDH1 carriers, resulting in shorter hospitalization without compromising pathology or safety. SynopsisMinimally invasive total gastrectomy performed for patients with CDH1 mutations has minimal short- or long-term complications and was associated with shorter length of stay and less weight loss than open total gastrectomy, without compromising lymph node yield or margin status.

Development of a streamlined NGS-based TCGA classification scheme for gastric cancer and its implications for personalized therapy.

The Cancer Genome Atlas (TCGA) has identified four distinct molecular subtypes of gastric cancer (GC) with prognostic significance: Epstein-Barr virus (EBV)-positive, microsatellite instability (MSI)-high, genomically stable (GS), and chromosomal instability (CIN). Unfortunately, the complex analysis required for TCGA classification limits its practical use in clinical settings. Our study sought to devise a next-generation sequencing (NGS)-based method to classify GC more efficiently, serving as a promising biomarker for prognosis and immunotherapy efficacy. This study was a retrospective observation study, and we employed 2 independent GC cohorts. The 3DMed cohort (n=765), comprising data on 733 cancer-related genes along with 4 EBV-encoded genes, was utilized to develop the new NGS classification. Additionally, the secondary Korean cohort (n=55), which includes both genomic data and information on immune checkpoint inhibitor (ICI) treatment, was employed to establish a correlation between NGS subtypes and ICI responsiveness. In the 3DMed cohort, we identified 5.2% EBV, 4.6% MSI, 30.6% GS, and 59.6% CIN subtypes. The MSI subtype exhibited the highest number of mutation events, along with the highest tumor mutational burden (TMB) and strong programmed cell death ligand 1 (PD-L1) expression. CIN tumors showed extensive copy number variations (CNVs) and genomic heterogeneity. The EBV subtype presented recurrent ARID1A and PIK3CA mutations and fewer TP53 mutations. GS tumors exhibited specific mutations in CDH1 and ARID1A. In the Korean cohort, ICIs were most effective in MSI and EBV cases, showing disease control rates of 100%, compared to 62.9% in GS and 12.5% in CIN subtypes. The NGS method successfully maps the mutational landscape of GC, providing a practical TCGA classification surrogate to optimize patient-specific treatment strategies.

Large-scale analysis of CDH1 mutations defines a distinctive molecular subset with treatment implications in gastric cancer.

Although histological and molecular classifications have been extensively studied for gastric cancer (GC), targeted therapies for GC remain limited. CDH1 mutations (MT) are characteristic of genomically stable GC and are associated with poor prognosis, but lack effective or targeted therapies. Here, we showed the overall mutation frequency of CDH1 was 9.7% (155 of 1596). CDH1-MT GC showed significantly lower rates of PD-L1 positivity (CPS score ≥1) than CDH1-wildtype (WT) GC (56.7% vs. 73.3%, p < 0.05). Compared to CDH1-WT GC, mutations of ARID1A, WRN, POT1, CDK12, and FANCC were significantly higher, while TP53 and APC were significantly lower in CDH1-MT GC (p < 0.05); The rates of KRAS and HER2 amplifications were significantly lower, while CRKL and IGF1R amplifications were significantly higher in CDH1-MT GC, compared to CDH1-WT GC (p < 0.05). Frequently altered genes in CDH1-MT GC were especially enriched in DNA damage repair and cell cycle checkpoint pathways. Inhibition of E-cadherin sensitized GC cell lines to PARP and Wee1 inhibitors by disrupting DNA damage repair pathway and cell cycle checkpoint. This is the largest study to investigate the distinct genomic landscape of CDH1-MT GC. Our data indicated GC patients with CDH1 mutations could potentially benefit from agents targeting PARP and Wee1.

A Systematic Review of Surgical and Pathological Outcomes in Patients With a CDH1 Mutation Undergoing Total Gastrectomy.

CDH1 (E-cadherin) genetic mutations are associated with a 30%-70% increased lifetime risk of hereditary diffuse gastric cancer (HDGC). Although prophylactic total gastrectomy (PTG) reduces long-term risk of gastric cancer, the associated morbidity and mortality remain unclear. This systematic review aims to characterise postoperative surgical outcomes in patients undergoing total gastrectomy. A systematic literature search was performed for studies reporting endoscopic surveillance, surgical and pathological outcomes for patients with CDH1 mutation undergoing a total gastrectomy. Thirty-nine studies included 1849 patients, of which 96% had a CDH1 (n = 1777) or CTNNA1 (n = 3) mutation. Endoscopy outcomes were reported for 1640 patients. Cancer foci were identified in 32% (n = 523/1640) and 71% of these patients went on to have a total gastrectomy (n = 369/523). The remaining 78% of patients did not have cancer foci detected on endoscopy (n = 1117/1640). Of these patients, 62% underwent a total gastrectomy (n = 688/1117) and 81% were found to have cancer on surgical histology (n = 556/688). Pathological staging was reported for 790 patients undergoing surgery, of which 68% had pT1 disease (n = 537). Postoperative complications were reported for 430 patients across 23 studies, with the most common complications being anastomotic strictures (25%), anastomotic leaks (13%), wound infections (12%) and pulmonary complications (11%). Only one postoperative death was reported within 30 days. Rates of early cancers are high in CDH1 patients undergoing PTG, highlighting the need for improvement in reliable endoscopic surveillance. Although postoperative mortality in this surgical cohort remains low, high rates of postoperative complications warrant careful patient counselling.

Spatially Resolved Niche and Tumor Microenvironmental Alterations in Gastric Cancer Peritoneal Metastases

Background and aimsPeritoneal metastasis (PM) in gastric cancer (GC) is associated with poor prognosis and significant morbidity. We sought to understand the genomic, transcriptomic, and tumor microenvironment (TME) features that contribute to peritoneal organotropism in GC. MethodsWe conducted a comprehensive multi-omic analysis of 548 samples from 326 patients, including primary tumors, matched normal tissues, peritoneal metastases, and adjacent-normal peritoneal tissues. We employed whole exome sequencing, whole transcriptome sequencing, and digital spatial profiling to investigate molecular alterations, gene expression patterns, and TME characteristics associated with PM. ResultsOur analysis identified specific genomic alterations in primary tumors, including mutations in ELF3, CDH1, and PIGR, and TME signatures, such as stromal infiltration and M2 macrophage enrichment, associated with increased risk of PM. We observed distinct transcriptional programs and immune compositions in GCPM compared to liver metastases, highlighting the importance of the TME in transcoelomic metastasis. We found differential expression of therapeutic targets between primary tumors and PM, with lower CLDN18.2 and FGFR2b expression in PM. We unravel the roles of the TME in niche reprogramming within the peritoneum, and provide evidence of pre-metastatic niche conditioning even in early GC without clinical PM. These findings were further validated using a humanized mouse model, which demonstrated niche remodeling in the peritoneum during transcoelomic metastasis. ConclusionOur study provides a comprehensive molecular characterization of GCPM and unveils key biological principles underlying transcoelomic metastasis. The identified predictive markers, therapeutic targets, and TME alterations offer potential avenues for targeted interventions and improved patient outcomes.

E-Cadherin Mutational Landscape and Outcomes in Breast Invasive Lobular Carcinoma

Invasive lobular carcinomas (ILC) are characterized by the loss of E-cadherin expression and CDH1 gene inactivation. Diagnostic reproducibility for this tumor type is currently suboptimal and could be improved by a better understanding of its histomolecular and clinical heterogeneity. We have analyzed the relationship between the presence, type, or position of CDH1 mutations, E-cadherin expression, and clinicopathological features (including outcome) in a retrospective series of 251 primary ILC with a long follow-up (median: 9.5 years). The mutational status of E-cadherin gene (CDH1) was determined by RNA sequencing from frozen tumor samples. E-cadherin immunohistochemistry (IHC) was performed with antibodies directed against the intracellular domain (clone 4A2C7) and the extracellular domain (clone NCH38). IHC expression of p120 and β-catenin was also assessed in E-cadherin diffusely positive cases. Three major patterns of E-cadherin membrane expression were identified by IHC, with good agreement between the 2 clones (overall concordance: 83.8%, Kappa 0.67): null/focal expression (≤10%) (72.8% of cases for 4A2C7 and 83.8% for NCH38), heterogeneous expression (11%-89%) (19.2% of cases for 4A2C7 and 6.9% for NCH38), and diffuse expression (≥90%) (8% of cases for 4A2C7 and 9.3% for NCH38). E-cadherin membranous expression, when present, was abnormal (incomplete labeling and/or reduced intensity). ILC with diffuse E-cadherin expression showed abnormal β-catenin or p120-catenin staining in 21% of cases. Interestingly, these cases with diffusely expressed E-cadherin had a CDH1 mutation rate as high as the E-cadherin null/focal cases (∼70%) but were enriched in nontruncating mutations. Regarding CDH1 mutation location, intracytoplasmic domain mutations correlated with a divergent E-cadherin IHC phenotype between the 2 antibodies (4A2C7 ≤ 10%/NCH38 ≥ 10%). Clinico-pathological correlation analyses found that stromal amount (inversely correlated with tumor cellularity) and tumor-infiltrating lymphocytes were less abundant in ILC with E-cadherin null/focal cases. In addition, CDH1 truncating mutations were associated with radiohistologic size discordance and were identified in multivariate survival analysis as an independent poor prognosis factor in terms of metastasis risk and breast cancer-related mortality. Overall, our study highlights the importance of the precise mutational status of CDH1 in the clinical, radiological, histologic, and phenotypic expression of lobular carcinomas. These findings should be taken into account in future attempts to improve diagnostic criteria or methods for ILC, as well as for clinicobiological studies dedicated to this tumor type.

Double Heterozygosity for Germline Mutations in Chinese Breast Cancer Patients.

Double pathogenic mutations occurring in an individual are considered a rare event. The introduction of a multiple-gene panel at Hong Kong Hereditary Breast Cancer Family Registry has allowed the identification of pathogenic variants in multiple genes, providing more information on clinical management and surveillance to the proband and their family members. Breast cancer patients who are double heterozygous (DH) for different hereditary breast and ovarian cancer syndrome (HBCO)-related genes were identified from a cohort of 3649 Chinese patients. Nine patients (0.25%) were observed to have germline DH mutations in ATM, BRCA1, BRCA2, BRIP1, CDH1, CHEK2, MSH6, PALB2, and TP53. Three probands were diagnosed with unilateral breast cancer, two patients were diagnosed with bilateral breast cancer, and four patients had multiple primary cancers. The median age for breast cancer diagnosis was an early age of 36 years. Chinese DH carriers did not show worse phenotypes or have a significantly downhill clinical presentation. However, seven out of nine (77.8%) of our DH carriers harbored a BRCA1 mutation, and four of them (44.4%) developed bilateral breast cancer, suggesting Chinese DH individuals may have a higher chance of having bilateral breast cancer than other populations (p = 0.0237).

Silencing immune-infiltrating biomarker CCDC80 inhibits malignant characterization and tumor formation in gastric cancer

ObjectiveTumor immune infiltration leads to poor prognosis of gastric cancer patients and seriously affects the life quality of gastric cancer patients. This study was based on bioinformatics to screen prognostic biomarkers in patients with high degree of immune invasion of gastric cancer. Meanwhile, the action of biomarker CCDC80 was explored in gastric cancer by cell and tumorigenesis experiments, to provide reference for the cure of gastric cancer patients.MethodsData sets and clinical massage on gastric cancer were collected from TCGA database and GEO database. ConsensusClusterPlus was used to cluster gastric cancer patients based on the 28 immune cells infiltration in ssGSEA. R “Limma” package was applied to analyze differential mRNAs between Cluster 1 and Cluster 2. Differential expression genes were screened by single factor analysis. Stemness markers (SERPINF1, DCN, CCDC80, FBLN5, SPARCL1, CCL14, DPYSL3) were identified for differential expression genes. Prognostic value of CCDC80 was evaluated in gastric cancer. Differences in genomic mutation and tumor microenvironment immune infiltration were assessed between high or low CCDC80. Finally, gastric cancer cells (HGC-27 and MKN-45) were selected to evaluate the action of silencing CCDC80 on malignant characterization, macrophage polarization, and tumor formation.ResultsBioinformatics analysis showed that CCDC80, as a stemness marker, was significantly overexpressed in gastric cancer. CCDC80 was also related to the degree of gastric cancer immune invasion. CCDC80 was up-expressed in cells of gastric cancer. Silencing CCDC80 inhibited malignant characterization and subcutaneous tumor formation of gastric cancer cells. High expression of CCDC80 was positive correspondence with immune invasion. Silencing CCDC80 inhibited M2 polarization and promoted M1 polarization in tumor tissues. In addition, gastric cancer patients were likely to have mutations in CDH1, ACTRT1, GANAB, and CDH10 genes in the High-CCDC80 group.ConclusionSilencing CCDC80, a prognostic biomarker in patients with immune invasion of gastric cancer, could effectively inhibit the malignant characterization, M2 polarization, and tumor formation of gastric cancer.

Molecular and immunological characterization of androgen receptor expression in different breast cancer subtypes.

1114 Background: While estrogen receptor (ER) is well-studied in breast cancer (BC), the role of androgen receptor (AR) in prognosis and therapy response is less understood. Here, we aimed to characterize the clinicopathologic and molecular features of AR gene expression in BC subtypes. Methods: 10,728 BC samples were tested by NGS (592, NextSeq; WES, NovaSeq) and WTS (NovaSeq; Caris Life Sciences, Phoenix, AZ). MSI was tested by IHC and NGS. TMB-high was designated as &gt;10 somatic mutations/MB. Tumors with AR-high(H) and AR-low(L) RNA expression were classified by top and bottom quartile, respectively. Real world overall survival (OS) and treatment-associated survival was obtained from insurance claims and calculated from treatment start to last contact using Kaplan-Meier estimates. Statistical significance was determined by chi-square and Mann-Whitney U test with p-values adjusted for multiple comparisons ( q&lt;.05). Results: Lobular carcinoma had higher AR expression (2.37-fold) and AR positivity by IHC (93.6% vs 63.6%) compared to ductal carcinoma ( q&lt;.05). Luminal A (LumA) had higher AR expression and IHC positivity compared to HER2-enriched, basal-like, and normal-like tumors ( q&lt;.05). AR-H tumors were significantly ( p&lt;.05) enriched for PIK3CA mutations in LumA (60.3% vs 29.8%), LumB (48.1% vs 23.2%), HER2-enriched (51.2% vs 39.3%), and basal-like (23.8% vs 8.3%); CDH1 mutations in LumA (26% vs 10.3%) and normal-like (55.5% vs 4.8%); MAP3K1 in LumA (17.1% vs 5.3%); ESR1 in LumB (17.9% vs 10.7%); and NF1 in HER2-enriched (19.3% vs 7.1%). AR-H had higher immune infiltration for LumA (B cells, M2 Mφ, neutrophils, NK cells, DC), LumB (B cells, M2 Mφ, neutrophils, NK cells, CD4+ T cells, DC), basal-like (NK cells, CD4+ T cells), and normal-like (NK cells, DC; q&lt;.05) than AR-L. Conversely, AR-H HER2-enriched tumors had lower immune infiltration (B cells, M1 and M2 Mφ, NK cells, CD8+ T cells, DC) and fewer TMB-high (15.1% vs 23.2%), dMMR/MSI-high (0.3% vs 2%), and PD-L1+ tumors (15% vs 40.8%; p&lt;.05) compared to AR-L (all q&lt;.05). Post-doxorubicin survival was longer for patients with AR-H tumors; however, opposing trends were observed for patients with LumB (75.5 vs 56.3 m; p=.081) and HER2-enriched tumors (47.9 vs 65.2 m; p.071). AR-H was associated with longer post-dox survival in patients with LumB PIK3CA-mt tumors (81.7 vs 39.3 m; p=.036), but shorter post-dox survival in HER2-enriched PIK3CA-wt (50.4 vs 70.4 m) and PIK3CA-mt cohorts (40.8 vs 58.0 m; p=.18). Conclusions: Findings from our updated analysis based on PAM50 designation suggest a strong association between AR and PIK3CA mutations across subtypes and suggest that distinct AR-associated alterations in the immune microenvironment require further study. Exploration of specific mutations and immune-oncology markers associated with AR-H may aid in molecularly selected clinical trial design for advanced BC patients.

Serial Cell-Free DNA Sequencing in ROS1 Fusion-Positive Lung Cancers During Treatment With Entrectinib.

Patients with metastatic ROS1 fusion-positive non-small cell lung cancer (NSCLC) are effectively treated with entrectinib, a multikinase inhibitor. Whether serial targeted gene panel sequencing of cell-free DNA (cfDNA) can identify response and progression along with mechanisms of acquired resistance to entrectinib is underexplored. In patients with ROS1 fusion-positive NSCLC, coclinical trial plasma samples were collected before treatment, after two cycles, and after progression on entrectinib (global phase II clinical trial, ClinicalTrials.gov identifier: NCT02568267). Samples underwent cfDNA analysis using MSK-ACCESS. Variant allele frequencies of detectable alterations were correlated with objective response per RECIST v1.1 criteria. Twelve patients were included, with best response as partial response (n = 9, 75%), stable disease (n = 2, 17%), and progressive disease (PD; n = 1, 8%). A ROS1 fusion was variably detected in cfDNA; however, patients without a ROS1 fusion in cfDNA had no other somatic alterations detected, indicative of possible low cfDNA shedding. Clearance of the enrolling ROS1 fusion or concurrent non-ROS1 alterations (TP53, CDH1, NF1, or ARID1A mutations) was observed in response to entrectinib therapy. Radiologic PD was accompanied by redemonstration of a ROS1 fusion or non-ROS1 alterations. On-target resistance was rare; only one patient acquired ROS1 G2032R at the time of progression. Several patients acquired new off-target likely oncogenic alterations, including a truncating alteration in NF1. Serial cfDNA monitoring may complement radiographic assessments as determinants of response and resistance to entrectinib in ROS1 fusion-positive lung cancers in addition to detecting putative resistance mechanisms on progression.

Mutation characteristics of cancer susceptibility genes in Chinese ovarian cancer patients.

The association between mutations in susceptibility genes and the occurrence of ovarian cancer has been extensively studied. Previous research has primarily concentrated on genes involved in the homologous recombination repair pathway, particularly BRCA1 and BRCA2. However, a wider range of genes related to the DNA damage response pathways has not been fully explored. To investigate the mutation characteristics of cancer susceptibility genes in the Chinese ovarian cancer population and the associations between gene mutations and clinical data, this study initially gathered a total of 1171 Chinese ovarian cancer samples and compiled a dataset of germline mutations in 171 genes. In this study, it was determined that MC1R and PRKDC were high-frequency ovarian cancer susceptibility genes in the Chinese population, exhibiting notable distinctions from those in European and American populations; moreover high-frequency mutation genes, such as MC1R: c.359T>C and PRKDC: c.10681T>A, typically had high-frequency mutation sites. Furthermore, we identified c.8187G>T as a characteristic mutation of BRCA2 in the Chinese population, and the CHEK2 mutation was significantly associated with the early onset of ovarian cancer, while the CDH1 and FAM175A mutations were more prevalent in Northeast China. Additionally, Fanconi anemia pathway-related genes were significantly associated with ovarian carcinogenesis. In summary, this research provided fundamental data support for the optimization of ovarian cancer gene screening policies and the determination of treatment, and contributed to the precise intervention and management of patients.

Proteogenomic characterization of difficult-to-treat breast cancer with tumor cells enriched through laser microdissection

BackgroundBreast cancer (BC) is the most commonly diagnosed cancer and the leading cause of cancer death among women globally. Despite advances, there is considerable variation in clinical outcomes for patients with non-luminal A tumors, classified as difficult-to-treat breast cancers (DTBC). This study aims to delineate the proteogenomic landscape of DTBC tumors compared to luminal A (LumA) tumors.MethodsWe retrospectively collected a total of 117 untreated primary breast tumor specimens, focusing on DTBC subtypes. Breast tumors were processed by laser microdissection (LMD) to enrich tumor cells. DNA, RNA, and protein were simultaneously extracted from each tumor preparation, followed by whole genome sequencing, paired-end RNA sequencing, global proteomics and phosphoproteomics. Differential feature analysis, pathway analysis and survival analysis were performed to better understand DTBC and investigate biomarkers.ResultsWe observed distinct variations in gene mutations, structural variations, and chromosomal alterations between DTBC and LumA breast tumors. DTBC tumors predominantly had more mutations in TP53, PLXNB3, Zinc finger genes, and fewer mutations in SDC2, CDH1, PIK3CA, SVIL, and PTEN. Notably, Cytoband 1q21, which contains numerous cell proliferation-related genes, was significantly amplified in the DTBC tumors. LMD successfully minimized stromal components and increased RNA–protein concordance, as evidenced by stromal score comparisons and proteomic analysis. Distinct DTBC and LumA-enriched clusters were observed by proteomic and phosphoproteomic clustering analysis, some with survival differences. Phosphoproteomics identified two distinct phosphoproteomic profiles for high relapse-risk and low relapse-risk basal-like tumors, involving several genes known to be associated with breast cancer oncogenesis and progression, including KIAA1522, DCK, FOXO3, MYO9B, ARID1A, EPRS, ZC3HAV1, and RBM14. Lastly, an integrated pathway analysis of multi-omics data highlighted a robust enrichment of proliferation pathways in DTBC tumors.ConclusionsThis study provides an integrated proteogenomic characterization of DTBC vs LumA with tumor cells enriched through laser microdissection. We identified many common features of DTBC tumors and the phosphopeptides that could serve as potential biomarkers for high/low relapse-risk basal-like BC and possibly guide treatment selections.

CDH1 methylation analysis in invasive lobular breast carcinomas with and without gene mutation

The proposed role of CDH1 (E-cadherin gene) methylation as a mechanism of gene inactivation in invasive lobular carcinoma (ILC) remains inconclusive. For many years, CDH1 promoter hypermethylation has been regarded as a mechanism for gene inactivation in ILC. However, this assumption has primarily relied on non-quantitative assays, which have reported CDH1 methylation frequencies ranging from 26 to 93% at CpG sites within the island region. Few studies employing quantitative methods and covering CpG island shores, regions of relatively low CpG density situated proximal to conventional promoter CpGs, have been conducted, revealing lower percentages of methylation ranging from 0 to 51%. Therefore, using the quantitative pyrosequencing method, we examined CDH1 methylation in the island region and shores in E-cadherin deficient ILC cases (15 with CDH1 mutation and 22 non-mutated), 19 cases of invasive breast carcinomas non-special type (IBC-NSTs), and five cases of usual ductal hyperplasia (UDH). Our analysis revealed CDH1 methylation frequencies ranging from 3 to 64%, with no significant increase in methylation levels in any group of ILCs (median = 12%) compared to IBC-NST (median = 15%). In addition, considering the poorly studied association between the number of tumor-infiltrating lymphocytes (TILs) and CDH1 methylation in breast cancer, we undertook a thorough analysis within our dataset. Our findings revealed a positive correlation between CDH1 methylation and the presence of TILs (r = 0.5; p-value < 0.05), shedding light on an aspect of breast cancer biology warranting further investigation. These findings challenge CDH1 methylation as a CDH1 inactivation mechanism in ILC and highlight TILs as a potential confounding factor in gene methylation.

Abstract PO5-25-12: CDH1 mutations predict resistance to neoadjuvant taxane therapy

Abstract Background: Despite recent advances in personalized medicine, conventional chemotherapy remains a backbone in breast cancer therapy. Thus, identifying markers predicting sensitivity or resistance to individual chemotherapeutics is of great importance. Methods: In the EpiTax neoadjuvant trial, enrolling patients between 1997-2003, patients with primary breast cancers (T2 &amp;gt;4cm, T3/T4 and/or N2/N3) were randomized to epirubicin 90mg/m2/3W or paclitaxel 200mg/m²/3W monotherapy, with cross-over in case of inferior response. Pre-treatment snap-frozen tumor biopsies from 223 patients were analyzed by targeted NGS of a 360 gene panel. Endpoint for comparison was clinical response to the first regimen, since pCR was rare due to the large tumor sizes at inclusion. For validation purposes we performed targeted sequencing of tumor samples from a total of 478 patients included in the Gepar Trio (n=132), Quattro (n=171) and Quinto (n=175) trials, in which patients with &amp;gt;2cm tumors received neoadjuvant anthracycline/taxane combination regimens. Here, the primary endpoint was clinical response to combined treatment, but since these tumors were smaller than in the EpiTax-trial, pCR was included as a secondary endpoint. In addition, experimental validations were performed -by CDH1 knock-down and CRISPR/Cas9 knock-out in cell line models. Results: In samples from the EpiTax-trial, CDH1 mutations predicted an inferior response (trend across response groups; cPD, cSD, cPR and cCR) in the paclitaxel arm (p=0.01) as well as the epirubicin arm (p=0.04). The predictive value was observed within the subgroup of ER-positive cases (both for paclitaxel (p=0.005) and epirubicin (p=0.003)) but not among ER-negative tumors. The majority of CDH1 mutations (24/34=71%) were observed in lobular cancers. While lobular histology predicted resistance to paclitaxel (but not epirubicin), CDH1 mutations predicted resistance also within the subgroup of lobular cases (p=0.002), demonstrating CDH1 mutations to be an independent predictor and not only a co-variate to lobular histology. As assessing functionally linked genes, mutations in GATA3, a transcriptional regulator of CDH1, were predominantly observed in ductal cancers, and were not predictive of resistance to any compound. Yet, combining GATA3 and CDH1 mutations into a composite biomarker predicted resistance to both paclitaxel (p=0.007) and epirubicin (p=0.01), especially in ER-positive cases (p=0.002 and p=0.0004, respectively). While EMT-signatures had predictive value, this effect was largely dominated by CDH1, while other EMT-related genes had limited impact on response. In the independent validation cohort from the Gepar trials, selected with enrichment for lobular cancers (34%), CDH1 mutations were not significantly associated with resistance to therapy (p=0.19) although predicted lack of pCR (p=0.01). Combining GATA3 and CDH1 mutations predicted lack of clinical response (p=0.05) and lack of pCR (p=0.0007) respectively in this cohort. In the in vitro analyses, resistance to paclitaxel was observed in three different breast cancer cell lines upon siRNA mediated knock-down of CDH1, as well as in a CRISPR/Cas9 mediated CDH1 knock-out model, as measured by growth rate, induction of apoptosis, G2 arrest, mitochondrial respiration and tubulin stability. For anthracyclines, similar effects were observed for mitochondrial respiration. Conclusions: In conclusion, mutations in CDH1 predicted resistance to paclitaxel and epirubicin. Our data suggest that CDH1 mutations should be explored further as a predictive biomarker for potential application. Citation Format: Stian Knappskog, Reham Helwa, Sivaramakrishna Rachakonda, Liv B. Gansmo, Carsten Denkert, Lucy R. Yates, Christine Solbach, Michael Untch, Bruno V. Sinn, Anne-Sophie Litmeyer, Beyhan Ataseven, Jens Huober, David C. Wedge, Thomas Karn, Oleksii Nikolaienko, Frederik Marmé, Peter A. Fasching, Hans Petter Eikesdal, Elmar Stickeler, Christian Schem, Paul Jank, Marion van Mackelenbergh, Volkmar Müller, Baerbel Felder, Johannes Holtschmidt, Peter J. Campbell, Sibylle Loibl, Per Lonning. CDH1 mutations predict resistance to neoadjuvant taxane therapy [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PO5-25-12.

Abstract GS03-04: Novel Mechanisms of CDH1 Inactivation in Breast Invasive Lobular Carcinoma Unveiled by the Integration of Artificial Intelligence and Genomics

Abstract Background: Invasive lobular carcinoma (ILC) of the breast is the second most common histologic subtype of breast cancer (BC), following invasive ductal carcinoma of no special type (IDC-NST). The hallmark histologic feature of ILC is cellular discohesiveness, the result of bi-allelic inactivation of CDH1, and represents an important genotypic-phenotypic correlation in BC. Although most ILCs harbor CDH1 loss-of function mutations associated to loss-of-heterozygosity (LOH) of the wild type-allele, a subset of ILCs lack these alterations despite displaying a typical lobular phenotype. Here, we sought to identify alternative molecular mechanisms converging on CDH1 inactivation by employing an integrative artificial intelligence (AI) and genomics approach. Materials and Methods: A genomics-driven AI-based algorithm using hematoxylin and eosin (H&amp;E) whole slide images (WSIs) as input, previously developed to detect bi-allelic CDH1 mutations (inactivating mutation associated to LOH) in BC was employed. WSIs of 1,057 BCs including ILCs (n=187) and non-lobular BCs (n=870) previously subjected to FDA-cleared tumor/normal targeted sequencing were subjected to analysis with the AI-based algorithm. Cases predicted to harbor CDH1 bi-allelic mutations by the AI-model but lacking CDH1 bi-allelic mutations by targeted sequencing were assessed through targeted sequencing data re-analysis, CDH1 gene promoter methylation evaluation and/or whole genome sequencing analysis. Results: AI-based analysis WSIs corresponding to 1,057 BCs resulted in the identification of 34 cases found to lack CDH1 bi-allelic mutations by targeted sequencing but predicted to harbor these genetic alterations by the AI-based model. CDH1 gene promoter methylation assessment revealed CDH1 promoter methylation in 18 cases. Targeted sequencing data reanalysis revealed other genetic mechanisms of CDH1 inactivation including CDH1 homozygous deletions (n=3), intragenic deletion with LOH (n=1), and likely pathogenic non-coding CDH1 alterations associated with LOH (n=2). WGS analysis of an ILC revealed a novel deleterious CDH1 fusion stemming from translocation t(13;16), resulting in loss of the 5’UTR, transcription start site and exons 1 and 2 of CDH1, associated with complete loss of E-cadherin protein expression. Taken together, we identified alternative/novel mechanisms of bi-allelic CDH1inactivation in 74% (25/34) cases analyzed. Conclusions: By applying an AI-based algorithm trained to detect a genetic alteration (i.e., CDH1 bi-allelic mutations), we were able to identify alternative epigenetic and genetic molecular mechanisms of CDH1 inactivation in ILCs, including novel non-coding CDH1 genetic alterations and a new inactivating CDH1 fusion gene. These findings indicate that molecular mechanisms affecting a single gene or process converging on the same phenotype can be unveiled by the integration of AI and genomics, highlighting the robustness of this approach for the discovery of novel biology. Citation Format: Fresia Pareja, Higinio Dopeso, Yikan Wang, Andrea Gazzo, David Brown, Pier Selenica, Jan Bernhard, Fatemeh Derakhshan, Edaise M. da Silva, Lorraine Colon-Cartagena, Thais Basili, Antonio Marra, Jillian Sue, Qiqi Ye, Arnaud Da Cruz Paula, Selma Yeni, Xin Pei, Hunter Green, Kaitlyn Gill, Yingjie Zhu, Matthew Lee, Ran Godrich, Adam Casson, Britta Weigelt, Nadeem Riaz, Hanna Y Wen, Edi Brogi, Matthew Hanna, Diana Mandelker, Jeremy Kunz, Brandon Rothrock, Sarat Chandarlapaty, Christopher Kanan, Gerard Oakley III, David Klimstra, Thomas Fuchs, Jorge Reis-Filho. Novel Mechanisms of CDH1 Inactivation in Breast Invasive Lobular Carcinoma Unveiled by the Integration of Artificial Intelligence and Genomics [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr GS03-04.