Doublecortin-like kinase 1 (DCLK1) is a proposed driver of gastric cancer (GC) that phosphorylates serine and threonine residues. Here, we showed that the kinase activity of DCLK1 orchestrated cancer cell–intrinsic and–extrinsic processes that led to pro-invasive and pro-metastatic reprogramming of GC cells. Inhibition of the kinase activity of DCLK1 reduced the growth of subcutaneous xenograft tumors formed from MKN1 human gastric carcinoma cells in mice and decreased the abundance of the stromal markers α-Sma, vimentin, and collagen. Similar effects were seen in mice with xenograft tumors formed from MKN1 cells expressing a kinase-inactive DCLK1 mutant (MKN1 D511N ). MKN1 D511N cells also had reduced in vitro migratory potential and stemness compared with control cells. Mice orthotopically grafted with MKN1 cells overexpressing DCLK1 (MKN1 DCLK1 ) showed increased invasiveness and had a greater incidence of lung metastases compared with those grafted with control MKN1 cells. Mechanistically, we showed that the chemokine CXCL12 acted downstream of DCLK1 in cultured MKN1 cells and in mice subcutaneously implanted with gastric tumors formed by MKN1 DCLK1 cells. Moreover, inhibition of the kinase activity of DCLK1 or the expression of DCLK1 D511N reversed the pro-tumorigenic and pro-metastatic phenotype. Together, this study establishes DCLK1 as a broadly acting and potentially targetable promoter of GC.

Although aberrant activation of the KRAS and PI3K pathway alongside TP53 mutations account for frequent aberrations in human gastric cancers, neither the sequence nor the individual contributions of these mutations have been clarified. Here, we establish an allelic series of mice to afford conditional expression in the glandular epithelium of KrasG12D;Pik3caH1047R or Trp53R172H and/or ablation of Pten or Trp53. We find that KrasG12D;Pik3caH1047R is sufficient to induce adenomas and that lesions progress to carcinoma when also harboring Pten deletions. An additional challenge with either Trp53 loss- or gain-of-function alleles further accelerated tumor progression and triggered metastatic disease. While tumor-intrinsic STAT3 signaling in response to gp130 family cytokines remained as a gatekeeper for all stages of tumor development, metastatic progression required a mutant Trp53-induced interleukin (IL)-11 to IL-6 dependency switch. Consistent with the poorer survival of patients with high IL-6 expression, we identify IL-6/STAT3 signaling as a therapeutic vulnerability for TP53-mutant gastric cancer.

XLS file - 22KB, Ontological enrichment in epithelial and stromal compartments in PMCC_gastric datset.

GIM is a persistent, premalignant lesion whereby gastric mucosa is replaced by metaplastic mucosa resembling intestinal tissue, arising in the setting of chronic inflammation, particularly in the context of Helicobacter pylori. While the overall rates of progression to gastric adenocarcinoma are low, estimated at from 0.25 to 2.5%, there are features that confer a much higher risk and warrant follow-up. In this review, we collate and summarise the current knowledge regarding the pathogenesis of GIM, and the clinical, endoscopic and histologic risk factors for cancer. We examine the current state-of-practice with regard to the diagnosis and management of GIM, which varies widely in the published guidelines and in practice. We consider the emerging evidence in population studies, artificial intelligence and molecular markers, which will guide future models of care. The ultimate goal is to increase the detection of early gastric dysplasia/neoplasia that can be cured while avoiding unnecessary surveillance in very low-risk individuals.

Abstract Background Intestinal metaplasia (IM) is considered a key pivot point in the Correa model of gastric cancer (GC). It is histologically subtyped into the complete and incomplete subtypes, the latter being associated with a greater risk of progression. However, the clinical utility of IM subtyping remains unclear, partially due to the absence of reliable defining biomarkers. Methods Based on gene expression data and existing literature, we selected CD10 and Das1 as candidate biomarkers to distinguish complete and incomplete IM glands in tissues from patients without GC (IM-GC) and patients with GC (IM + GC). Immunohistochemical staining of individually subtyped IM glands was scored after blinding by two researchers using tissue belonging to both IM-GC and IM + GC patients. Whole tissue Das1 staining was further assessed using digital image quantification (cellSens Dimension, Olympus). Results Across both cohorts CD10 stained the IM brush border and was shown to have a high sensitivity (87.5% and 94.9% in IM-GC and IM + GC patients respectively) and specificity (100.0% and 96.7% respectively) with an overall AUROC of 0.944 for complete IM glands. By contrast Das1 stained mainly goblet cells and the apical membrane of epithelial cells, mostly of incomplete IM glands with a low sensitivity (28.6% and 29.3% in IM-GC and IM + GC patients respectively) but high specificity (98.3% and 85.1% respectively) and an overall AUROC of 0.603 for incomplete IM glands. A combined logistic regression model showed a significant increase in AUROC for detecting complete IM glands (0.955 vs 0.970). Whole tissue digital quantification of Das1 staining showed a significant association with incomplete IM compared to complete IM, both in IM-GC and in IM + GC patients (p = 0.016 and p = 0.009 respectively, Mann–Whitney test and unpaired t test used). Additionally, complete IM in IM + GC patients exhibited significantly more Das1 staining than in IM-GC patients (p = 0.019, Mann–Whitney test). Conclusions These findings suggest that CD10 is an outstanding biomarker for complete IM and Das1 may be useful as a secondary biomarker for IM glands at greater risk of progression irrespective of IM subtype. Overall, the clinical use of these biomarkers could lead to improved patient stratification and targeted surveillance.

Gastric cancer (GC) is the 3rd leading cause of cancer mortality worldwide, therefore providing novel diagnostic and treatment options is crucial for at risk groups. The serine/threonine kinase doublecortin-like kinase 1 (DCLK1) is a proposed driver of GC with frequent amplification and somatic missense mutations yet the molecular mechanism how DCLK1 mediates tumorigenesis is poorly understood. We report how DCLK1 expression orchestrates complementary cancer cell intrinsic and extrinsic processes leading to a comprehensive pro-invasive and pro-metastatic reprogramming of cancer cells and tumor stroma in a DCLK1 kinase-dependent manner. Mechanistically, we identify the chemokine CXCL12 as a key promoter of the pro-tumorigenic properties downstream of DCLK1. Importantly, inhibition of the DCLK1 kinase domain reverses the pro-tumorigenic and pro-metastatic phenotype. Together, this study establishes DCLK1 as a promising, targetable master regulator of GC. Teaser DCLK1 is a druggable cancer driver of GC

Metastasis is considered one of the hallmarks of cancer and enhanced tumor invasion and metastasis is significantly associated with cancer mortality. Metastasis occurs via a series of integrated processes involving tumor cells and the tumor microenvironment. The innate immune components of the microenvironment have been shown to engage with tumor cells and not only regulate their proliferation and survival, but also modulate the surrounding environment to enable cancer progression. In the era of immune therapies, it is critical to understand how different innate immune cell populations are involved in this process. This review summarizes recent literature describing the roles of innate immune cells during the tumor metastatic cascade.

Although the incidence of gastric cancer (GC) is declining, it remains a significant global problem. GC is thought to arise via a series of histologically distinct premalignant stages and triggered by chronic Helicobacter pylori infection. Key to this pathway is intestinal metaplasia (IM), an intermediate stage immediately preceding dysplasia and a known risk factor in the progression to GC. This chapter focuses primarily on IM, its histological subtypes, clinical implications, and role in GC progression, and discusses the global surveillance and management guidelines for individuals with IM. The latter part of the chapter addresses the diagnosis and treatment of early GC which is a developing field in Western countries where GC is often diagnosed at an advanced stage.

Abstract Gastric cancer (GC) is a heterogeneous disease in which diverse genetic, genomic, and epigenetic alterations can accumulate in different molecular and histologic subtypes. Tumor microenvironment (TME) also contributes to the heterogeneity of GC. To investigate what molecular features of tumor cells drive GC heterogeneity, we developed an integrative causal model, called Integrative Sequential Causality Test (ISCT), to identify key regulators of GC by integrating DNA methylation, copy number variation, and transcriptomic data. Applying ISCT to three GC cohorts that contain methylation, CNV, and gene expression data, 11 common methylation-driven key regulators were identified: ADHFE1, CDO1, CRYAB, FSTL1, GTP, PKP3, PTPRCAP, RAB25, RHOH, SFN, and SORD. Based on these 11 genes, gastric tumors resolved into three groups that were associated with known molecular subtypes, Lauren classification, tumor stage, and patient survival, suggesting significance of the methylation-driven key regulators in molecular and histologic heterogeneity of GC. We also investigated the relationship between TME and the methylation-driven key regulators and showed that both immune/stromal proportions in TME and tumor cell genomics variations contributed to expression variations of the methylation-driven key regulators. Especially, FSTL1, significantly associated with patient survival and tumor progression as well as stromal proportion in TME, was expressed at high level in both stromal and cancer cells, indicating its potential role in mediating tumor-stroma interactions. In summary, this study suggests that genetic, genomic, and epigenetic alterations as well as their interactions with TME contribute to heterogeneity of GC. Citation Format: Seungyeul Yoo, Quan Chen, Li Wang, Wenhui Wang, Ankur Chakravarthy, Rita Busuttil, Alex Boussioutas, Tim R. Fenton, Jiangwen Zhang, Xiaodan Fan, Seut-Yi Leung, Jun Zhu. Molecular heterogeneity of gastric cancer explained by methylation-driven key regulators [abstract]. In: Proceedings of the AACR Special Conference on Tumor Immunology and Immunotherapy; 2019 Nov 17-20; Boston, MA. Philadelphia (PA): AACR; Cancer Immunol Res 2020;8(3 Suppl):Abstract nr B103.

Abstract Objectives To facilitate disease prognosis and improve precise immunotherapy of gastric cancer (GC) patients, a comprehensive study integrating immune cellular and molecular analyses on tumor tissues and peripheral blood was performed. Methods The association of GC patients’ outcomes and the immune context of their tumors was explored using multiplex immunohistochemistry (mIHC) and transcriptome profiling. Potential immune dysfunction mechanism/s in the tumors on the systemic level was further examined using mass cytometry (CyTOF) in complementary peripheral blood from selected patients. GC cohorts with mIHC and gene expression profiling data were also used as validation cohorts. Results Increased CD4+FOXP3+ T‐cell density in the GC tumor correlated with prolonged survival. Interestingly, CD4+FOXP3+ T cells had a close interaction with CD8+ T cells rather than tumor cells. High densities of CD4+FOXP3+ T cells and CD8+ T cells (High‐High) independently predicted prolonged patient survival. Furthermore, the interferon‐gamma (IFN‐γ) gene signature and PDL1 expression were up‐regulated in this group. Importantly, a subgroup of genomically stable (GS) tumors and tumors with chromosomal instability (CIN) within this High‐High group also had excellent survival. The High‐High GS/CIN tumors were coupled with increased frequencies of Tbet+CD4+ T cells and central memory CD4+ T cells in the peripheral blood. Conclusion These novel findings identify the combination of CD8+ T cells and FOXP3+CD4+ T cells as a significant prognostic marker for GC patients, which also could potentially be targeted and applied in the combination therapy with immune checkpoint blockades in precision medicine.

Background: The association between the survival or efficacy of chemotherapy and the Lauren subtype of gastric cancer (GC) remains unclear. We aimed to clarify whether patients with different Lauren subtypes have different survival after treatment with systemic chemotherapy: intestinal gastric cancer (IGC) patients survived better than patients with mixed type gastric cancer (MGC) or diffuse gastric cancer (DGC) after treatment with systemic chemotherapy. Patients & methods: Relevant studies for the meta-analysis were identified through searching Pubmed, Embase, Cochrane and Ovid up to March 2020. We also included our own prospectively collected cohort of patients that were followed over a 10-year period. Sub-group and sensitivity analyses were also performed. Results: In our prospective cohort, the overall survival (OS) of IGC patients receiving systemic chemotherapy (chemoIGC) [median OS 5.01 years, interquartile range (IQR) 2.63–6.71] was significantly higher than that of DGC patients receiving the same chemotherapy (chemoDGC) (median OS 1.33 years, IQR 0.78–3.33, p = 0.0001). After adjusting for age, gender and cancer stage, there was a significant difference in OS in patients treated with chemotherapy based on the Lauren classification of GC {hazard ratio (HR) for OS of the IGC versus DGC 0.33, [95% confidence interval (CI), 0.17–0.65; p < 0.001]}. In the IGC patients, the adjusted HR associated with chemotherapy was 0.26 (95% CI, 0.12–0.56; p = 0.001), whereas the association was 0.64 (95% CI, 0.30–1.33; p = 0.23) in the DGC patient group. In our meta-analysis, 33 studies comprising 10,246 patients treated with systemic chemotherapy (chemoIGC n = 4888, chemoDGC n = 5358) met all the selection criteria. While we accounted for much of the heterogeneity in these studies, we found that chemoIGC patients showed significantly improved OS [HR, 0.76 (95% CI, 0.71–0.82); p < 0.00001] when compared with similarly treated chemoDGC patients. Conclusion: Our results support the consideration of Lauren subtype when prescribing systemic chemotherapy for GC, particularly for MGC or DGC, which may not benefit from chemotherapy. Lauren classification should be considered to stratify chemotherapy regimens to GC patients in future clinical trials, with particular relevance to MGC or DGC, which is more difficult to treat with current regimens.

Supplemental material, Supplementary_Information_for_TAM for A cohort study and meta-analysis of the evidence for consideration of Lauren subtype when prescribing adjuvant or palliative chemotherapy for gastric cancer by Kunning Wang, Enxiao Li, Rita A. Busuttil, Joseph C. Kong, Sharon Pattison, Joseph J. Y. Sung, Jun Yu, Emad M. El-Omar, Julie A. Simpson and Alex Boussioutas in Therapeutic Advances in Medical Oncology

OBJECTIVE:Gastric cancer patients generally have a poor outcome, particularly those with advanced-stage disease which is defined by the increased invasion of cancer locally and is associated with higher metastatic potential. This study aimed to identify genes that were functional in the most fundamental hallmark of cancer, namely invasion. We then wanted to assess their value as biomarkers of gastric cancer progression and recurrence.DESIGN:Data from a cohort of patients profiled on cDNA expression arrays was interrogated using K-means analysis. This genomic approach classified the data based on patterns of gene expression allowing the identification of the genes most correlated with the invasion of GC. We evaluated the functional role of a key protein from this analysis in invasion and as a biomarker of recurrence after curative resection.RESULTS:Expression of secreted frizzled-related protein 4 (SFRP4) was identified as directly proportional to gastric cancer invasion. This finding was validated in multiple, independent datasets and its functional role in invasion was also confirmed using invasion assays. A change in serum levels of SFRP4 after curative resection, when coupled with AJCC stage, can accurately predict the risk of disease recurrence after curative therapy in an assay we termed PredictR.CONCLUSIONS:This simple ELISA-based assay can help predict recurrence of disease after curative gastric cancer surgery irrespective of adjuvant therapy. The results require further evaluation in a prospective trial but would help in the rational prescription of cancer therapies and surveillance to prevent under or over treatment of patients after curative resection.

Gastric cancer (GC) is a heterogeneous disease of diverse genetic, genomic, and epigenetic alterations. Tumor microenvironment (TME) also contributes to the heterogeneity of GC. To investigate GC heterogeneity, we developed an Integrative Sequential Causality Test (ISCT) to identify key regulators of GC by integrating DNA methylation, copy number variation, and transcriptomic data. Applying ISCT to three GC cohorts containing methylation, CNV and transcriptomic data, 11 common methylation-driven key regulators ( ADHFE1, CDO1, CRYAB, FSTL1, GPT, PKP3, PTPRCAP, RAB25, RHOH, SFN, and SORD ) were identified. Based on these 11 genes, gastric tumors were clustered into 3 clusters which were associated with known molecular subtypes, Lauren classification, tumor stage, and patient survival, suggesting significance of the methylation-driven key regulators in molecular and histological heterogeneity of GC. We further showed that chemotherapy benefit was different in the 3 GC clusters and varied depending on the tumor stage. Both immune/stromal proportions in TME and tumor cell genomic variations contributed to expression variations of the 11 methylation-driven key regulators and to the GC heterogeneity.

Tumor-associated macrophages (TAMs), one of the most abundant immune components in gastric cancer (GC), are difficult to characterize due to their heterogeneity. Multiple approaches have been used to elucidate the issue, however, due to the tissue-destructive nature of most of these methods, the spatial distribution of TAMs in situ remains unclear. Here we probe the relationship between tumor context and TAM heterogeneity by multiplex immunohistochemistry of 56 human GC cases. Using distinct expression marker profiles on TAMs, we report seven predominant populations distributed between tumor and non-tumor tissue. TAM population-associated gene signatures reflect their heterogeneity and polarization in situ. Increased density of CD163+ (CD206−) TAMs with concurrent high CD68 expression is associated with upregulated immune-signaling and improved patient survival by univariate, but not multivariate analysis. CD68-only and CD206+ TAMs are correlated with high PDL1 expression.

Gastric cancer is a leading cause of cancer death worldwide, with advanced stage being correlated to the level of tumour invasion and metastasis. Current research is heavily focused on the identification and development of efficacious therapeutics targeting these fundamental hallmarks of cancer, however there are currently no animal models that mimic the invasive phenotypes observed in humans. To address this we have developed an orthotopic mouse model whereby gastric cancer cell lines are tagged with luciferase and injected into the subserosal layer of the stomach. This allows for the monitoring of primary tumour growth and metastasis in real-time as well as quantitation of the degree of tumour invasion through the stomach wall by immunohistochemistry. We have three models based on the degree of invasion and metastasis that are cell line specific: The AGS cells develop into invasive tumours by 4-weeks with no evidence of metastases, MKN45 cells are moderately metastatic with minimal invasion till week 2 and MKN28 cells are highly metastatic and fully invasive by week 1. These models have utility as a tool for testing the efficacy of anti-tumour, anti-invasive and anti-metastatic therapies in the setting of gastric cancer, which currently has poor treatment options.