Inflammation-associated Cancer Research Articles

Cytokine-centered strategies to boost cancer immunotherapy.

Cytokines have gained attention in oncology in recent years, especially in the context of immunotherapy. Although immune checkpoint inhibitors (ICIs) have revolutionized the treatment of cancer, there are still some challenges to be faced, such as the lack of predictive biomarkers as well as the emergence of resistance and their severe side effects. In this Viewpoint, we discuss the potential of cytokines, the soluble mediators of cancer-associated inflammation, in immunotherapy. Indeed, both the activation and inhibition of cytokines have been suggested as potential strategies to overcome immunotherapy resistance. In addition, serum levels of certain cytokines can predict response to immunotherapy, and cytokine inhibition could also contribute to prevent side effects induced by ICIs. Thus, although further research is still required, data support that cytokine-based therapies could be an attractive therapeutic strategy for cancer patients treated with immunotherapy in the near future.

Unlocking the potential: FKK6 as a microbial mimicry-based therapy for chronic inflammation-associated colorectal cancer in a murine model

Unlocking the potential: FKK6 as a microbial mimicry-based therapy for chronic inflammation-associated colorectal cancer in a murine model

P0144 Unveiling the role of the MFSD2A protein in contributing to the resolution of colitis-associated cancer inflammation

Abstract Background The intrinsic connection between inflammation and tumor promotion is well-established in colorectal cancer (CRC), where chronic inflammation, like ulcerative colitis (UC), plays a significant role in intestinal carcinogenesis (1). UC patients are at higher risk of colitis-associated cancer, emphasizing the inflammation-cancer connection. Previous studies from our laboratory revealed that UC patients with active inflammation have impaired production of inflammation-resolving DHA-derived metabolites due to a defect in the endothelial Major Facilitator Superfamily Domain containing 2A (MFSD2A) (2). Given the MFSD2A’s role in reducing colonic inflammation, we hypothesize it also counteracts cancer-associated inflammation. Methods Human Intestinal Microvascular Endothelial Cells (HIMEC) isolated from CRC and healthy samples were transduced with lentiviruses carrying either the GFP-tagged MFSD2A encoding sequence (MFSD2A-OE), or the relative GFP control, or the MFSD2A-targeting shRNAs, or the scramble as control, then analyzed by transcriptomics and lipidomics. A colon-adenocarcinoma cell line (Caco-2) was cocultured with either MFSD2A-OE or GFP CRC HIMEC to evaluate their proliferation rate. An orthotopic model of CRC was performed by intrarectally injecting CD1 nude mice with a cell mixture of Caco-2 and CRC HIMEC overexpressing either MFSD2A or GFP as control. Collected tumors were analyzed by flow cytometry and lipidomics (3). Results CRC HIMEC displayed a pro-resolving signature at the lipidomic level, consistently with higher MFSD2A expression as compared to the healthy. However, gene expression analysis shows a pro-inflammatory profile in CRC HIMEC, suggesting that MFSD2A levels alone are insufficient to induce a fully pro-resolving phenotype. We performed loss of function experiments to assess whether the pro-resolving profile in CRC HIMEC depends on MFSD2A. MFSD2A silencing reduces pro-resolving functions, confirming its role in balancing pro-inflammatory and pro-resolving signals in CRC regarding lipid production. Caco-2 co-cultured with MFSD2A-OE CRC HIMEC displayed a decrease in their proliferation rate after 48 and 72 hours of co-culture, indicating that endothelial MFSD2A overexpression reduces tumor cell proliferation. By comparison with animals injected with Caco-2 and GFP HIMEC, mice injected with a Caco-2/MFSD2A-OE CRC HIMEC mixture developed tumors with reduced weight and size, and increased pro-resolving lipid release. Additionally, flow cytometry showed increased M2 macrophage polarization, indicating a shift toward a pro-resolving immune response. Conclusion These data suggested that MFSD2A contrasts cancer-associated inflammation, ensuring the correct balance between pro-inflammatory and pro-resolving milieu.

OP03 Mapping the observable IBDverse: Using massive-scale single-cell RNA sequencing of gut and blood samples to nominate genes, cell types and pathways driving IBD susceptibility

Abstract Background Genome-wide association studies (GWAS) have identified 370 loci associated with inflammatory bowel disease (IBD) susceptibility1. The majority of these are driven by non-coding variants, thought to dysregulate the expression of a nearby gene. The dysregulated disease effector gene, and the operative cell-type(s) is often unknown. Previous efforts to nominate effector genes have relied on identifying expression quantitative trait loci (eQTL) and expression-linked genes (eGenes). However, gene expression is often derived from healthy individuals, a single, non-primary site, and measured using bulk RNA sequencing. This likely misses cell-, site-, or disease-specific genetic effects, and has nominated disease effector genes for only a minority of IBD loci. Methods To better characterise the universe of IBD effector genes, we created IBDverse - the largest collection of single-cell RNAseq data from the terminal ileum (TI, nCD=119, nHealthy=243), rectum (nHealthy=275) and blood (nCD=95) - a total of >2.4 million high quality transcriptomes from 732 samples across 396 individuals. After grouping cells into different cell types, we mapped genetic variants associated with gene expression changes. Disease effector genes were nominated by identifying expression-linked genetic effects likely driven by the same causal variant as an IBD GWAS signal, as determined by colocalisation analysis2. Results Across all tissues, we identify 102 transcriptionally distinct cell types (Fig 1), and genetic effects associated with the expression of >17,000 genes. For 507 genes, the genetic association for their expression was strongly associated with genetic effects for IBD susceptibility, suggesting them to be drivers of disease risk. Remarkably, this doubles the previous best efforts to nominate disease effector genes3, doing so at >52% of IBD susceptibility loci. As well as their nomination, this study also helps contextualise disease effector expression variation. Disease effector genes were detectable in only a single tissue or cell annotation in >40% of cases, highlighting the need for high resolution data obtained across multiple tissues. We also find the first evidence for upregulation of RASGRP1, a gene previously correlated with exacerbated colitis in mice4, to drive Crohn’s disease susceptibility. As this is identified in a subset of tissue-resident memory T cells, and only from the TI, this also suggests this cell type and tissue as governing this effect. Conclusion This study serves as a step change in the understanding of cellular heterogeneity of these tissues, provides a compendium of genetic regulatory effects across constituent cell types, and quantifies the relevance of these to IBD susceptibility. References Liu et al., (2023). Nat. Genetics. ‘Genetic architecture of the inflammatory bowel diseases across East Asian and European ancestries’ Giambartolemi et al., (2014) PLOS Genetics. ‘Bayesian test for colocalisation between pairs of genetic association studies using summary statistics’ Panousis et al., (2024) Nat. Comms (in press). Wang et al., (2022) Nat. Comms. ‘RasGRP1 promotes the acute inflammatory response and restricts inflammation-associated cancer cell growth’

Alpha-1 Antitrypsin as a Regulatory Protease Inhibitor Modulating Inflammation and Shaping the Tumor Microenvironment in Cancer.

Alpha-1 antitrypsin (AAT) is a key serine protease inhibitor for regulating proteases such as neutrophil elastase. AAT restrains the pulmonary matrix from enzymatic degradation, and a deficiency in AAT leads to inflammatory tissue damage in the lungs, resulting in chronic obstructive pulmonary disease. Due to the crucial biological function of AAT, the emerging research interest in this protein has shifted to its role in cancer-associated inflammation and the dynamics of the tumor microenvironment. However, the lack of comprehensive reviews in this field hinders our understanding of AAT as an essential immune modulator with great potential in cancer immunotherapy. Therefore, in this review, we have elucidated the pivotal roles of AAT in inflammation and the tumor microenvironment, including the structure and molecular properties of AAT, its molecular functions in the regulation of the inflammatory response and tumor microenvironment, and its clinical implications in cancer including its diagnosis, prognosis, and therapeutic intervention. This review seeks to bridge the gap in the understanding of AAT between inflammatory diseases and cancer, and to foster deeper investigations into its translational potential in cancer immunotherapy in the future.

Prostaglandin E2 and Akt promote stemness in Apc mutant Dclk1+ cells to give rise to colitis-associated cancer.

Loss of the tumor suppressor gene Apc in Lgr5+ intestinal stem cells results in aberrant Wnt signaling and colonic tumorigenesis. In the setting of injury, however, we and others have also shown that non-stem cells can also give rise to colonic tumors. The mechanism by which inflammation leads to cellular plasticity and cancer, however, remains largely unknown. RNA expression analysis of Wnt, COX and Akt signaling was assessed in patients with quiescent or active UC and patients with UC-associated neoplasia using available datasets. The role of COX signaling in colonic tumorigenesis was examined using epithelial and Dclk1+ cell specific conditional COX-1 knockout mice and pharmacologic treatment with different NSAIDs. In this study, we show that prostaglandins and phospho-Akt are key inflammatory mediators that promote stemness in Apc mutant Dclk1+ cells that give rise to colorectal cancer. Moreover, prostaglandin E2 (PGE2) and Akt are increased in colitis in both mice and humans, leading to inflammation-associated dysplasia upon activation of Wnt signaling. Importantly, inhibition of epithelial derived COX-1 by Aspirin or conditional knockout in Dclk1+ cells reduced PGE2 levels and prevented the development of inflammation-associated colorectal cancer. Our data shows that epithelial and Dclk1+ cell derived COX-1 plays an important role in inflammation-associated tumorigenesis. Importantly, low dose Aspirin was effective in chemo-prevention through inhibition of COX-1 that reduced colitis-associated cancer.

RORc expressing immune cells negatively regulate tertiary lymphoid structure formation and support their pro-tumorigenic functions.

RORc-expressing immune cells play important roles in inflammation, autoimmune disease and cancer. They are required for lymphoid organogenesis and have been implicated in tertiary lymphoid structure (TLS) formation. TLSs are formed in many cancer types and have been correlated with better prognosis and response to immunotherapy. In liver cancer, some TLSs are pro-tumorigenic as they harbor tumor progenitor cells and support their growth. The processes involved in TLS development and acquisition of pro- or anti-tumorigenic roles are largely unknown. This study aims to explore the role of RORc-expressing cells in TLS development in the context of inflammation-associated liver cancer. IKKβ(EE)Hep mice, exhibiting chronic liver inflammation, TLS formation and liver cancer, were crossed with RORc knockout mice to explore RORc's effect on TLS and tumor formation. TLS phenotypes were analyzed using transcriptional, proteomic, and immunohistochemical techniques. CD4, CD8, and B cell depletions were used to assess their contribution to liver TLS and tumor formation. RORc-expressing cells are detected within TLSs of both human patients and mice developing intrahepatic cholangiocarcinoma. In mice, these cells negatively regulate TLS formation, as excess TLSs form in their absence. CD4 cells are essential for liver TLS formation, while B cells are required for TLS formation specifically in the absence of RORc-expressing cells. Importantly, in chronically inflamed livers lacking RORc-expressing cells, TLSs become anti-tumorigenic, reducing tumor load. Anti-tumorigenic TLSs revealed enrichment of exhausted CD8 cells with effector functions, germinal center B cells and plasma cells. B cells are key in limiting tumor development, possibly via tumor-directed antibodies. RORc-expressing cells negatively regulate B cell responses and facilitate the pro-tumorigenic functions of hepatic TLSs.

A proinflammatory stem cell niche drives myelofibrosis through a targetable galectin-1 axis.

Myeloproliferative neoplasms are stem cell-driven cancers associated with a large burden of morbidity and mortality. Most patients present with early-stage disease, but a substantial proportion progress to myelofibrosis or secondary leukemia, advanced cancers with a poor prognosis and high symptom burden. Currently, it remains difficult to predict progression, and therapies that reliably prevent or reverse fibrosis are lacking. A major bottleneck to the discovery of disease-modifying therapies has been an incomplete understanding of the interplay between perturbed cellular and molecular states. Several cell types have individually been implicated, but a comprehensive analysis of myelofibrotic bone marrow is lacking. We therefore mapped the cross-talk between bone marrow cell types in myelofibrotic bone marrow. We found that inflammation and fibrosis are orchestrated by a "quartet" of immune and stromal cell lineages, with basophils and mast cells creating a TNF signaling hub, communicating with megakaryocytes, mesenchymal stromal cells, and proinflammatory fibroblasts. We identified the β-galactoside-binding protein galectin-1 as a biomarker of progression to myelofibrosis and poor survival in multiple patient cohorts and as a promising therapeutic target, with reduced myeloproliferation and fibrosis in vitro and in vivo and improved survival after galectin-1 inhibition. In human bone marrow organoids, TNF increased galectin-1 expression, suggesting a feedback loop wherein the proinflammatory myeloproliferative neoplasm clone creates a self-reinforcing niche, fueling progression to advanced disease. This study provides a resource for studying hematopoietic cell-niche interactions, with relevance for cancer-associated inflammation and disorders of tissue fibrosis.

ERK5 mediates pro-tumorigenic phenotype in non-small lung cancer cells induced by PGE2

Lung cancer is the leading cause of cancer-related deaths worldwide, with non-small cell lung cancer (NSCLC) constituting approximately 84 % of all lung cancer cases. The role of inflammation in the initiation and progression of NSCLC tumors has been the focus of extensive research. Among the various inflammatory mediators, prostaglandin E2 (PGE2) plays a pivotal role in promoting the aggressiveness of epithelial tumors through multiple mechanisms, including the stimulation of growth, evasion of apoptosis, invasion, and induction of angiogenesis. The Extracellular signal-Regulated Kinase 5 (ERK5), the last discovered member among conventional mitogen-activated protein kinases (MAPK), is implicated in cancer-associated inflammation. In this study, we explored whether ERK5 is involved in the process of tumorigenesis induced by PGE2. Using A549 and PC9 NSCLC cell lines, we found that PGE2 triggers the activation of ERK5 via the EP1 receptor. Moreover, both genetic and pharmacological inhibition of ERK5 reduced PGE2-induced proliferation, migration, invasion and stemness of A549 and PC9 cells, indicating that ERK5 plays a critical role in PGE2-induced tumorigenesis. In summary, our study underscores the pivotal role of the PGE2/EP1/ERK5 axis in driving the malignancy of NSCLC cells in vitro. Targeting this axis holds promise as a potential avenue for developing novel therapeutic strategies aimed at controlling the advancement of NSCLC.

The m6A methyltransferase METTL3 modifies Kcnk6 promoting on inflammation associated carcinogenesis is essential for colon homeostasis and defense system through histone lactylation dependent YTHDF2 binding

Inflammation induces tumor formation and plays a crucial role in tumor progression and prognosis. KCNK6, by regulating K(+) efflux to reduce NLRP3 Inflammasome-induced lung injury, relaxes the aorta. This study aims to elucidate the effects and biological mechanism of KCNK6 in inflammation-associated carcinogenesis, which may be essential for colon homeostasis and the defense system. To induce colitis, mice were given 3.0% Dextran Sodium Sulfate (DSS) in their drinking water for 7 days. The Azoxymethane (AOM) +DSS method was used to induce colon cancer in the mice model. Bone marrow-derived macrophages (BMDM) from Kcnk6-/- mice, AW264.7 cells, and human colon cancer HCT116 and Caco2 cells were used as in vitro models. The loss of Kcnk6 prevented spontaneous colitis and restored mucosal integrity and homeostatic molecules. Additionally, the loss of Kcnk6 reduced the severity of AOM/DSS-induced carcinogenesis. Kcnk6 promoted cell viability and proliferation in HCT-116 or Caco-2 cells. The loss of Kcnk6 inhibited the levels of inflammatory factors in BMDM cells. Kcnk6 accelerated potassium channel activity, inducing NLRP3 inflammasome activation. METTL3-mediated m6A modification increased Kcnk6 stability in a YTHDF2-dependent manner. Histone lactylation activated the transcription of YTHDF2/Kcnk6. Our study revealed the important role of Kcnk6 in inflammation-associated carcinogenesis progression. The m6A methyltransferase METTL3 and histone lactylation increased Kcnk6 stability in a YTHDF2-dependent manner, providing a potential strategy for inflammation-associated carcinogenesis or colorectal cancer therapy.

Mood, Anxiety, and Cognitive Alterations in Cancer Patients.

To analyze the cytokine profile in cerebrospinal fluid (CSF), as well as mood, anxiety, and cognition profiles in patients with CC. One hundred and nine individuals were evaluated, 37 controls, 18 CWC, and 54 CC patients. Assessments included BDI, HADS, Digit Span, FAS-verbal, Animals/WMS-R, Matrix Reasoning and Vocabulary (WASI), and QLQ-C30. The CC group exhibited 62.96% depression and probable anxiety/depression, with 75.92% showing attention deficits. The CC and CWC groups demonstrated significant cognitive impairment on the WASI-Vocabulary test (CWC: 13.4 ± 2.2; CC: 15.9 ± 1.1) compared to the control group (Ct: 22.8 ± 1.6; p = 0.0002). In the QLQ-C30 scores, the CC group reported a greater perceived loss of quality of life and health deterioration (score of 17.5 ± 2.6) and lower scores on the Functional Scale (49.8 ± 4.5). The CC group had 18.52% illiteracy, 18.52% incomplete higher education, and 22.22% complete elementary education. The CC group also had lower weight (Ct: 67.8 ± 1.4; CWC: 61.7 ± 3.1; CC: 59.6 ± 1.7; p = 0.0023) and BMI (CC: 21.5 [18.3; 24.8]; Ct: 24.9 [23; 25.8]; p = 0.0021) compared to controls. Cytokines detected in the CSF were MCP-1, VEGF, IL-8, IP-10, and MIP-1β. Higher concentrations of MCP-1 were found in cancer patients (CSC: 571.2 ± 105.8; CC: 399.5 ± 65.9; Ct: 1477 ± 0.1; p < 0.0001), along with lower levels of MIP-1β (CC: 4345 [3060; 7353]) and VEGF (CC: 48.3 ± 2.0; CWC: 49.8 ± 3.8; Ct: 64.8 ± 3.2; p < 0.0001). The level of mental impairment (mood, anxiety, and cognitive deficits) correlated with cancer-associated and cachexia-associated inflammation, weight loss, low BMI, elevated C-reactive protein (CRP), leukocytosis, lymphopenia, anemia, hypoalbuminemia, and low scores on the QLQ-C30 questionnaire (Global Health Status, Functional Scale, Symptom Scale). The CC group exhibited a higher prevalence of depression/anxiety, a stronger correlation between depression and inflammation, and greater cognitive impairment in attention, reasoning, and language, alongside lower average educational attainment. The low concentration of certain cytokines in the CSF combined with elevated systemic CRP in cancer and cachexia, associated with mental disorders, presents a paradox that requires further investigation. Higher concentrations of the cytokine MCP-1 in cancer patient groups indicated a positive correlation with the preservation of language abilities in these patients.

Elevated expression of Golgi Transport 1B promotes the progression of cervical cancer by activating NF-κB signaling pathway via interaction with TBK1.

As a preventable disease, cervical cancer (cervical squamous cell carcinoma and endocervical adenocarcinoma - CESC) remains a tumor with high morbidity and mortality worldwide, underscoring the pressing need for effective treatment strategies. This research identified Golgi transport 1B (GOLT1B) as a critical gene involved in the development of cervical cancer. Gene Expression Omnibus (GEO) datasets were investigated to determine the upregulation of GOLT1B in cervical cancer tissue compared to normal tissue. Besides, GOLT1B was found to predict poor prognosis in cervical cancer by utilizing Gene Expression Profiling Interactive Analysis (GEPIA). The functional assay indicated that GOLT1B promoted CESC viability and migration in vitro and in vivo. RNA sequencing results suggested that GOLT1B likely influenced NF-κB pathway. The subsequent western blot and dual luciferase reporter assay revealed the interaction between GOLT1B and TBK1, modulating the NF-κB pathway. More importantly, GOLT1B was also found to regulate immune cells infiltration, suggesting its potential role in tumor microenvironment. In conclusion, GOLT1B promotes CESC progression via interaction with TBK1 and augmentation of NF-κB signaling-mediated cancer-associated inflammation, which provides us a new approach to CESC target therapy.

Functional variants of the pentraxin 3 gene are associated with the metastasis and progression of prostate cancer.

Age, ethnic background and genetic components have been identified as the established risks for prostate cancer (PCa). Pentraxin 3 (PTX3), originally identified as a pattern-recognition molecule for defence against infectious agents, has multiple functions in tissue repair and in the regulation of cancer-associated inflammation. In this study, we sought to investigate the impact of PTX3 gene variants on the development of PCa. Genotypes of four common single-nucleotide polymorphisms (SNPs) of PTX3 gene, including rs1840680, rs2305619, rs3816527 and rs2120243, were profiled among 705 PCa patients and 705 ethnicity-matched controls. In this study, we found that patients who carry at least one minor allele (C) of rs3816527 (AC and CC) tended to develop advanced forms of diseases (clinical large T stage, OR, 1.593, p = 0.032; pathologically-confirmed nodal spread, OR, 1.987, p = 0.011; metastatic tumour, OR, 3.896, p = 0.032) as compared with those homologous for the major allele (AA). Further stratification analysis showed that such association of rs3816527 with lymphatic and distal metastasis of PCa was accentuated in the younger age group (≤65 at diagnosis) but not seen in the older age group (>65 at diagnosis), suggesting an age-specific effect of PTX3 variants. Prediction of PTX3 protein structure implied that polymorphism may alter the quaternary organization and oligomerization of PTX3 protein. Moreover, our gene silencing experiments and survey of public datasets revealed that elevation of PTX3 levels in PCa was required for cell migration and associated with tumour metastasis. Our results highlight an association of PTX3 rs3816527 with the progression of PCa.

Unlocking the Potential: FKK6 as a Microbial Mimicry-Based Therapy for Chronic Inflammation-Associated Colorectal Cancer in a Murine Model.

Chronic intestinal inflammation significantly contributes to the development of colorectal cancer (CRC) and remains a pertinent clinical challenge, necessitating novel therapeutic approaches. Indole-based microbial metabolite mimics FKK6, which is a ligand and agonist of the pregnane X receptor (PXR), was recently demonstrated to have PXR-dependent anti-inflammatory and protective effects in a mouse model of dextran sodium sulfate (DSS)-induced acute colitis. Here, we examined the therapeutic potential of FKK6 in a mouse model (C57BL/6 FVB humanized PXR mice) of colitis-associated colon cancer (CAC) induced by azoxymethane (AOM) and dextran sodium sulfate (DSS). FKK6 (2 mg/kg) displayed substantial anti-tumor activity, as revealed by reduced size and number of colon tumors, improved colon histopathology, and decreased expression of tumor markers (c-MYC, β-catenin, Ki-67, cyclin D) in the colon. In addition, we carried out the chronic toxicity (30 days) assessment of FKK6 (1 mg/kg and 2 mg/kg) in C57BL/6 mice. Histological examination of tissues, biochemical blood analyses, and immunohistochemical staining for Ki-67 and γ-H2AX showed no difference between FKK6-treated and control mice. Comparative metabolomic analyses in mice exposed for 5 days to DSS and administered with FKK6 (0.4 mg/kg) revealed no significant effects on several classes of metabolites in the mouse fecal metabolome. Ames and micronucleus tests showed no genotoxic and mutagenic potential of FKK6 in vitro . In conclusion, anticancer effects of FKK6 in AOM/DSS-induced CAC, together with FKK6 safety data from in vitro tests and in vivo chronic toxicity study, and comparative metabolomic study, are supportive of the potential therapeutic use of FKK6 in the treatment of CAC.

Cyclophilin J limits linear ubiquitin signaling and controls colorectal cancer progression

Exorbitant sustained inflammation is closely linked to inflammation-associated disorders, including cancer. The initiation of gastrointestinal cancers such as colorectal cancer (CRC) is frequently accelerated by uncontrollable chronic inflammation which is triggered by excessive activation of nuclear factor kappa-B (NF-κB) signaling. Linear ubiquitin chains play an important role in activating canonical NF-κB pathway. The only known E3 complex, linear ubiquitin assembly complex (LUBAC) is responsible for the synthesis of linear ubiquitin chains, thus leading to the activation of NF-κB axis and promoting the development of inflammation and inflammation-associated cancers. We report here cyclophilin J (CYPJ) is a negative regulator of the LUBAC. The N-terminus of CYPJ binds to the second Npl4-like zinc finger (NZF2) domain of HOIP and the ubiquitin-like (UBL) domain of SHARPIN to disrupt the interaction between HOIP and SHARPIN and thus restrains linear ubiquitin chain synthesis and NF-κB activation. Cypj-deficient mice are highly susceptible to dextran sulfate sodium (DSS)-induced colitis and DSS plus azoxymethane (AOM)-induced colon cancer. Moreover, CYPJ expression is induced by hypoxia. Patients with high expression of both CYPJ and hypoxia-inducible factor-1α (HIF-1α) have longer overall survival and progression-free survival. These results implicate CYPJ as an unexpected robust attenuator of inflammation-driven tumorigenesis that exerts its effects by controlling linear ubiquitin chain synthesis in NF-κB signal pathway.

Single-Cell Profiling Reveals the Impact of Genetic Alterations on the Differentiation of Inflammation-Induced Murine Colon Tumors.

Genetic mutations and chronic inflammation of the colon contribute to the development of colorectal cancer (CRC). Using a murine model of inflammation-induced colon tumorigenesis, we determined how genetic mutations alter colon tumor cell differentiation. Inflammation induced by enterotoxigenic Bacteroides fragilis (ETBF) colonization of multiple intestinal neoplasia (MinApcΔ716/+) mice triggers loss of heterozygosity of Apc causing colon tumor formation. Here, we report that the addition of BRAFV600E mutation (BRAFF-V600ELgr5tm1(Cre/ERT2)CleMinApcΔ716/+, BLM) or knocking out Msh2 (Msh2LoxP/LoxPVil1-creMinApcΔ716/+, MSH2KO) in the Min model altered colon tumor differentiation. Using single-cell RNA sequencing, we uncovered the differences between BLM, Min, and MSH2KO tumors at a single-cell resolution. BLM tumors showed an increase in differentiated tumor epithelial cell lineages and a reduction in the tumor stem cell population. Interestingly, the tumor stem cell population of BLM tumors had revival colon stem cell characteristics with low WNT signaling and an increase in RevCSC marker gene expression. In contrast, MSH2KO tumors were characterized by an increased tumor stem cell population that had higher WNT signaling activity compared to Min tumors. Furthermore, overall BLM tumors had higher expression of transcription factors that drive differentiation, such as Cdx2, than Min tumors. Using RNA velocity, we identified additional potential regulators of BLM tumor differentiation such as NDRG1. The role of CDX2 and NDRG1 as putative regulators for BLM tumor cell differentiation was verified using organoids derived from BLM tumors. Our results demonstrate the critical connections between genetic mutations and cell differentiation in inflammation-induced colon tumorigenesis. Understanding such roles will deepen our understanding of inflammation-associated colon cancer.

Inflammation and cancer: friend or foe?

Chronic inflammation plays a crucial role in the onset and progression of pathologies like neurodegenerative and cardiovascular diseases, diabetes, and cancer, since tumor development and chronic inflammation are linked, sharing common signaling pathways. At least 20% of breast and colorectal cancers are associated with chronic inflammation triggered by infections, irritants, or autoimmune diseases. Obesity, chronic inflammation, and cancer interconnection underscore the importance of population-based interventions in maintaining healthy body weight, to disrupt this axis. Given that the dietary inflammatory index is correlated with an increased risk of cancer, adopting an anti-inflammatory diet supplemented with nutraceuticals may be useful for cancer prevention. Natural products and their derivatives offer promising antitumor activity with favorable adverse effect profiles; however, the development of natural bioactive drugs is challenging due to their variability and complexity, requiring rigorous research processes. It has been shown that combining anti-inflammatory products, such as non-steroidal anti-inflammatory drugs (NSAIDs), corticosteroids, and statins, with plant-derived products demonstrate clinical utility as accessible adjuvants to traditional therapeutic approaches, with known safety profiles. Pharmacological approaches targeting multiple proteins involved in inflammation and cancer pathogenesis emerge as a particularly promising option. Given the systemic and multifactorial nature of inflammation, comprehensive strategies are essential for long term success in cancer therapy. To gain insights into carcinogenic phenomena and discover diagnostic or clinically relevant biomarkers, is pivotal to understand genetic variability, environmental exposure, dietary habits, and TME composition, to establish therapeutic approaches based on molecular and genetic analysis. Furthermore, the use of endocannabinoid, cannabinoid, and prostamide-type compounds as potential therapeutic targets or biomarkers requires further investigation. This review aims to elucidate the role of specific etiological agents and mediators contributing to persistent inflammatory reactions in tumor development. It explores potential therapeutic strategies for cancer treatment, emphasizing the urgent need for cost-effective approaches to address cancer-associated inflammation.

D239Y germline variant of NTHL1 glycosylase increases susceptibility to colorectal cancer in mice

Introduction: Persistent genotoxic stress leads to spontaneous DNA damage, which occurs on the order of 104−105 events per cell per day. The base excision repair is the first responder in the repair mechanisms after DNA damage, with DNA glycosylase being the first step in the repair process by binding to the DNA, cleaving the N-glycosidic bond and removing the oxidized pyrimidine DNA lesions. NTHL1 is a pivotal bifunctional DNA glycosylase enzyme involved in base excision repair mechanism. NTHL1 tumor syndrome is characterized by an increased lifetime risk for colorectal cancer (CRC), breast cancer, and colorectal polyposis. A rare rs3087468 polymorphism (D239Y) exhibits no glycosylase activity and may act as a dominant-negative mutation. Hypothesis: NTHL1-D239Y variant conforms susceptibility to inflammation-associated colorectal cancer. Methods: 6-8 weeks/old male and female C57BL/6 wild-type (WT, NTHL1WT/WT), heterozygous (NTHL1WT/D239Y), or homozygous (NTHL1D239Y/D239Y) knock-in mice were injected with 7.5mg/kg azoxymethane (AOM) followed by three cycles of alternating 2.5% dextran sulfate sodium (DSS, 5 days) and normal drinking water (14 days). We monitored body weight, collected fecal samples for microbial 16S rRNA amplicon sequencing, and analyzed colon length and number/size of polyps at euthanasia. The histology score was assessed by an unbiased pathologist to evaluate inflammation and tumor development. Blood samples were used to evaluate micronuclei formation. Results: NTHL1WT/D239Y and NTHL1D239Y/D239Y mice showed more pronounced weight loss starting from the 2nd DSS cycle. The number and size of polyps in the distal colon of NTHL1WT/D239Y and NTHL1D239Y/D239Y male mice were increased compared to WT littermates. NTHL1D239Y/D239Y females, but not males, showed increased polyp number compared to NTHL1WT/D239Y mice. No statistical difference was observed in the polyps’ size. Tumors extending into the proximal colon were observed uniquely in NTHL1D239Y/D239Y females. On the other hand, only NTHL1WT/D239Y males developed high adenomas in the distal colon compared to WT males. Interestingly, only NTHL1D239Y/D239Y males showed statistical difference in micronucleated normochromatic erythrocytes compared to WT mice. NTHL1 genotype did not significantly affect fecal microbial α- or β-diversity or colitis score and colon length in either gender. Conclusion: Our data suggest that NTHL1-D239Y variant confers a complex, zygosity- and sex-dependent susceptibility risk for inflammation-associated colorectal cancer that may be intrinsic and independent of changes in the gut microbiota. In females, it promotes more proximal tumor location, while in males, it promotes the size and tumor progression. 5P30CA023074 (JBS), Phoenix Women's Board of the Steele Children's Research Center (FKG). This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Abstract 2180: Modulation of immune infiltration patterns in inflammation-associated colorectal cancer through rice bran-based dietary strategies

Abstract The correlation between gut microbiota dysbiosis and inflammation-associated colorectal cancer (CRC) has generated considerable interest in dietary interventions targeting immune modulation. Our prior study demonstrated the colon cancer protective effects of rice bran (RB) and ex vivo Bifidobacterium longum fermented rice bran (FRB) diet intervention against azoxymethane/dextran sodium sulfate (AOM/DSS)-induced CRC in gut microbiota-intact specific pathogen-free mice (SPF). RB showed higher levels of protection compared to FRB against CRC, as evidenced by diminished neoplastic lesion size and restored colonic epithelial integrity. The major objective of the present study was to leverage cutting-edge multispectral imaging technique for simultaneous detection of multiple immune cells markers and for a comprehensive analysis of distinct immune landscapes. We also performed spatial correlations within the tumor microenvironment of AOM/DSS-SPF mice undergoing RB and FRB dietary interventions. The primary focus was on T cell subtypes including cytotoxic T cells, helper T cells, regulatory T (Treg) cells, natural killer (NK) cells, NKT cells, B cells and macrophages (MΦ) subtypes (M1 MΦ, and M2 MΦ) to uncover the intricacies of the immune landscape and the interplay for CRC prevention through dietary intervention. The results indicated that RB intervention for 15 weeks led to reductions in overall T cell subtypes in the colonic tissue, particularly Treg cells, which are associated with protecting tumors against immune surveillance in CRC. On the other hand, FRB intervention also resulted in reductions in overall T cell subtypes but exhibited higher infiltration of NK cells and proinflammatory M1 macrophages in the colonic tissue after 15 weeks. While an increased presence of NK cells might be beneficial in countering CRC, the infiltration of M1 macrophages could potentially lessen the impact of FRB compared to RB intervention in CRC protection. The outcomes suggest that the better protective efficacy of non-fermented RB diets is linked to the modulation of Treg cell infiltration, whereas the efficacy of the FRB diet is to some extent limited by the presence of M1 macrophages. Overall, this study provides valuable insights into the complex immune landscape influenced by RB and FRB interventions in a murine model of inflammation-induced CRC, and a novel mechanism for the immunomodulatory effects of dietary RB interactions with gut microbiota. In the future, observing immune modulation throughout diet intervention would enhance our understanding of the role played by these immune cells. Citation Format: Robin Kumar, Lakshmi Sai Pratyusha Bugata, Md. Imtiazul Kabir, Rajesh Agarwal, Elizabeth P. Ryan, Komal Raina. Modulation of immune infiltration patterns in inflammation-associated colorectal cancer through rice bran-based dietary strategies [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 2180.

Abstract 1440: Retinoblastoma binding protein 9 regulates intestinal inflammation and inflammation-associated carcinogenesis

Abstract The intestinal epithelium constitutes the mucosal barrier, and its dysfunction promotes microbial invasion from the gut lumen, which further induces the intestinal inflammation. Chronic intestinal inflammation, as observed in poorly controlled inflammatory bowel diseases (IBD), leads to the development of inflammation-associated cancer. Thus, a deeper understanding of the regulatory system that controls intestinal homeostasis is essential. Retinoblastoma-binding protein 9 (RBBP9) is expressed in a range of human cancer cells. RBBP9 was initially reported to have the ability to regulate the cell cycle pathway. More recent studies have identified RBBP9 as a serine hydrolase (SH) that promotes cancer progression by subverting the anti-proliferative function of TGFβ in a pancreatic cancer model. However, little is known about the roles RBBP9 plays in the development of intestinal cancer. In this study, we investigated the role of RBBP9 in inflammation and inflammation-associated cancer in the intestine. Analyses of surgically resected human samples and publicly available datasets demonstrated that RBBP9 expression was reduced in patient samples of ulcerative colitis (UC) and colitis-associated cancer (CAC). Genetically knockout mice for Rbbp9 (Rbbp9−/−) were susceptible to experimental colitis induced by dextran sulfate sodium (DSS). Furthermore, Rbbp9−/− mice exhibited enhanced azoxymethane/DSS-induced tumorigenesis, a model for human CAC. Mechanistically, RNA sequencing of these tumors showed increased enrichment of inflammation-associated signatures, including interferon signaling pathways. Ex vivo organoid experiments revealed the suppressive function of RBBP9 in interferon signaling-mediated apoptosis of the intestinal epithelium. Simultaneous inhibition of RBBP9 and STAT1, a downstream transcription factor of interferon signaling, reverted the apoptotic phenotype of Rbbp9−/− organoids. Collectively, our data provide insight into the understanding of the crucial role of RBBP9 in protecting against intestinal inflammation and inflammation-related tumorigenesis via regulating the interferon signaling-mediated apoptotic pathway. Citation Format: Yuki Nakanishi, Kensuke Hamada, Mayuki Omatsu, Kosuke Iwane, Hiroki Kitamoto, Maokoto Okabe, Yu Muta, Shuji Yamamoto, Akihisa Fukuda, Hiroaki Kasashima, Hiroshi Seno. Retinoblastoma binding protein 9 regulates intestinal inflammation and inflammation-associated carcinogenesis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 1440.