Telomere dysfunction alters intestinal stem cell dynamics to promote cancer.

Telomere dynamics are linked to aging hallmarks, and age-associated telomere loss fuels the development of epithelial cancers. In Apc-mutant mice, the onset of DNA damage associated with telomere dysfunction has been shown to accelerate adenoma initiation via unknown mechanisms. Here, we observed that Apc-mutant mice engineered to experience telomere dysfunction show accelerated adenoma formation resulting from augmented cell competition and clonal expansion. Mechanistically, telomere dysfunction induces the repression of EZH2, resulting in the derepression of Wnt antagonists, which causes the differentiation of adjacent stem cells and a relative growth advantage to Apc-deficient telomere dysfunctional cells. Correspondingly, in this mouse model, GSK3ß inhibition countered the actions of Wnt antagonists on intestinal stem cells, resulting in impaired adenoma formation of telomere dysfunctional Apc-mutant cells. Thus, telomere dysfunction contributes to cancer initiation through altered stem cell dynamics, identifying an interception strategy for human APC-mutant cancers with shortened telomeres.

Histone demethylase KDM5D upregulation drives sex differences in colon cancer.

Sex exerts a profound impact on cancer incidence, spectrum and outcomes, yet the molecular and genetic bases of such sex differences are ill-defined and presumptively ascribed to X-chromosome genes and sex hormones1. Such sex differences are particularly prominent in colorectal cancer (CRC) in which men experience higher metastases and mortality. A murine CRC model, engineered with an inducible transgene encoding oncogenic mutant KRASG12D and conditional null alleles of Apc and Trp53 tumour suppressors (designated iKAP)2, revealed higher metastases and worse outcomes specifically in males with oncogenic mutant KRAS (KRAS*) CRC. Integrated cross-species molecular and transcriptomic analyses identified Y-chromosome gene histone demethylase KDM5D as a transcriptionally upregulated gene driven by KRAS*-mediated activation of the STAT4 transcription factor. KDM5D-dependent chromatin mark and transcriptome changes showed repression of regulators of the epithelial cell tight junction and major histocompatibility complex class I complex components. Deletion of Kdm5d in iKAP cancer cells increased tight junction integrity, decreased cell invasiveness and enhanced cancer cell killing by CD8+ T cells. Conversely, iAP mice engineered with a Kdm5d transgene to provide constitutive Kdm5d expression specifically in iAP cancer cells showed an increased propensity for more invasive tumours in vivo. Thus, KRAS*-STAT4-mediated upregulation of Y chromosome KDM5D contributes substantially to the sex differences in KRAS* CRC by means of its disruption of cancer cell adhesion properties and tumour immunity, providing an actionable therapeutic strategy for metastasis risk reduction for men afflicted with KRAS* CRC.

Synthetic Essentiality of Tryptophan 2,3-Dioxygenase 2 in APC-Mutated Colorectal Cancer.

Inactivation of adenomatous polyposis coli (APC) is common across many cancer types and serves as a critical initiating event in most sporadic colorectal cancers. APC deficiency activates WNT signaling, which remains an elusive target for cancer therapy, prompting us to apply the synthetic essentiality framework to identify druggable vulnerabilities for APC-deficient cancers. Tryptophan 2,3-dioxygenase 2 (TDO2) was identified as a synthetic essential effector of APC-deficient colorectal cancer. Mechanistically, APC deficiency results in the TCF4/ß-catenin-mediated upregulation of TDO2 gene transcription. TDO2 in turn activates the Kyn-AhR pathway, which increases glycolysis to drive anabolic cancer cell growth and CXCL5 secretion to recruit macrophages into the tumor microenvironment. Therapeutically, APC-deficient colorectal cancer models were susceptible to TDO2 depletion or pharmacologic inhibition, which impaired cancer cell proliferation and enhanced antitumor immune profiles. Thus, APC deficiency activates a TCF4-TDO2-AhR-CXCL5 circuit that affects multiple cancer hallmarks via autonomous and nonautonomous mechanisms and illuminates a genotype-specific vulnerability in colorectal cancer. SIGNIFICANCE: This study identifies critical effectors in the maintenance of APC-deficient colorectal cancer and demonstrates the relationship between APC/WNT pathway and kynurenine pathway signaling. It further determines the tumor-associated macrophage biology in APC-deficient colorectal cancer, informing genotype-specific therapeutic targets and the use of TDO2 inhibitors. This article is highlighted in the In This Issue feature, p. 1599.

A bipartite element with allele-specific functions safeguards DNA methylation imprints at the Dlk1-Dio3 locus.

Loss of imprinting (LOI) results in severe developmental defects, but the mechanisms preventing LOI remain incompletely understood. Here, we dissect the functional components of the imprinting control region of the essential Dlk1-Dio3 locus (called IG-DMR) in pluripotent stem cells. We demonstrate that the IG-DMR consists of two antagonistic elements: a paternally methylated CpG island that prevents recruitment of TET dioxygenases and a maternally unmethylated non-canonical enhancer that ensures expression of the Gtl2 lncRNA by counteracting de novo DNA methyltransferases. Genetic or epigenetic editing of these elements leads to distinct LOI phenotypes with characteristic alternations of allele-specific gene expression, DNA methylation, and 3D chromatin topology. Although repression of the Gtl2 promoter results in dysregulated imprinting, the stability of LOI phenotypes depends on the IG-DMR, suggesting a functional hierarchy. These findings establish the IG-DMR as a bipartite control element that maintains imprinting by allele-specific restriction of the DNA (de)methylation machinery.

Tumor cell-derived SPON2 promotes M2-polarized tumor-associated macrophage infiltration and cancer progression by activating PYK2 in CRC.

BACKGROUND: Tumor-associated macrophages (TAMs) are key regulators of the complex interplay between cancer and the immune microenvironment. Tumor cell-derived spondin 2 (SPON2) is an extracellular matrix glycoprotein that has complicated roles in recruitment of macrophages and neutrophils during inflammation. Overexpression of SPON2 has been shown to promote tumor cell migration in colorectal cancer (CRC). However, the mechanism by which SPON2 regulates the accumulation of TAMs in the tumor microenvironment (TME) of CRC is unknown. METHODS: Immunohistochemistry was used to examine SPON2 expression in clinical CRC tissues. In vitro migration assays, transendothelial migration assays (iTEM), and cell adhesion assays were used to investigate the effects of SPON2 on monocyte/macrophage migration. Subcutaneous tumor formation and orthotopic implantation assays were performed in C57 BL/6 mice to confirm the effects of SPON2 on TAM infiltration in tumors. RESULTS: SPON2 expression is positively correlated with M2-TAM infiltration in clinical CRC tumors and poor prognosis of CRC patients. In addition, SPON2 promotes cytoskeletal remodeling and transendothelial migration of monocytes by activating integrin ß1/PYK2 axis. SPON2 may indirectly induce M2-polarization through upregulating cytokines including IL10, CCL2 and CSF1 expression in tumor cells. Blocking M2 polarization and Macrophage depletion inhibited the SPON2-induced tumors growth and invasion. Furthermore, blocking the SPON2/integrin ß1/PYK2 axis impairs the transendothelial migration of monocytes and cancer-promoting functions of TAMs in vivo. CONCLUSIONS: Our findings demonstrate that SPON2-driven M2-TAM infiltration plays an important role during CRC tumor growth and metastasis. SPON2 may be a valuable biomarker guiding the use of macrophage-targeting strategies and a potential therapeutic target in advanced CRC.

AR-negative prostate cancer is vulnerable to loss of JMJD1C demethylase.

Prostate cancer is a leading cause of cancer-related mortality in men. The widespread use of androgen receptor (AR) inhibitors has generated an increased incidence of AR-negative prostate cancer, triggering the need for effective therapies for such patients. Here, analysis of public genome-wide CRISPR screens in human prostate cancer cell lines identified histone demethylase JMJD1C (KDM3C) as an AR-negative context-specific vulnerability. Secondary validation studies in multiple cell lines and organoids, including isogenic models, confirmed that small hairpin RNA (shRNA)-mediated depletion of JMJD1C potently inhibited growth specifically in AR-negative prostate cancer cells. To explore the cooperative interactions of AR and JMJD1C, we performed comparative transcriptomics of 1) isogenic AR-positive versus AR-negative prostate cancer cells, 2) AR-positive versus AR-negative prostate cancer tumors, and 3) isogenic JMJD1C-expressing versus JMJD1C-depleted AR-negative prostate cancer cells. Loss of AR or JMJD1C generates a modest tumor necrosis factor alpha (TNFα) signature, whereas combined loss of AR and JMJD1C strongly up-regulates the TNFα signature in human prostate cancer, suggesting TNFα signaling as a point of convergence for the combined actions of AR and JMJD1C. Correspondingly, AR-negative prostate cancer cells showed exquisite sensitivity to TNFα treatment and, conversely, TNFα pathway inhibition via inhibition of its downstream effector MAP4K4 partially reversed the growth defect of JMJD1C-depleted AR-negative prostate cancer cells. Given the deleterious systemic side effects of TNFα therapy in humans and the viability of JMJD1C-knockout mice, the identification of JMJD1C inhibition as a specific vulnerability in AR-negative prostate cancer may provide an alternative drug target for prostate cancer patients progressing on AR inhibitor therapy.

Genetic and biological hallmarks of colorectal cancer.

Colorectal cancer has served as a genetic and biological paradigm for the evolution of solid tumors, and these insights have illuminated early detection, risk stratification, prevention, and treatment principles. Employing the hallmarks of cancer framework, we provide a conceptual framework to understand how genetic alterations in colorectal cancer drive cancer cell biology properties and shape the heterotypic interactions across cells in the tumor microenvironment. This review details research advances pertaining to the genetics and biology of colorectal cancer, emerging concepts gleaned from immune and single-cell profiling, and critical advances and remaining knowledge gaps influencing the development of effective therapies for this cancer that remains a major public health burden.

H3K27ac bookmarking promotes rapid post-mitotic activation of the pluripotent stem cell program without impacting 3D chromatin reorganization.

During self-renewal, cell-type-defining features are drastically perturbed in mitosis and must be faithfully reestablished upon G1 entry, a process that remains largely elusive. Here, we characterized at a genome-wide scale the dynamic transcriptional and architectural resetting of mouse pluripotent stem cells (PSCs) upon mitotic exit. We captured distinct waves of transcriptional reactivation with rapid induction of stem cell genes and transient activation of lineage-specific genes. Topological reorganization at different hierarchical levels also occurred in an asynchronous manner and showed partial coordination with transcriptional resetting. Globally, rapid transcriptional and architectural resetting associated with mitotic retention of H3K27 acetylation, supporting a bookmarking function. Indeed, mitotic depletion of H3K27ac impaired the early reactivation of bookmarked, stem-cell-associated genes. However, 3D chromatin reorganization remained largely unaffected, suggesting that these processes are driven by distinct forces upon mitotic exit. This study uncovers principles and mediators of PSC molecular resetting during self-renewal.

Telomerase Reverse Transcriptase Preserves Neuron Survival and Cognition in Alzheimer's Disease Models.

Amyloid-induced neurodegeneration plays a central role in Alzheimer's disease (AD) pathogenesis. Here, we show that telomerase reverse transcriptase (TERT) haploinsufficiency decreases BDNF and increases amyloid-ß (Aß) precursor in murine brain. Moreover, prior to disease onset, the TERT locus sustains accumulation of repressive epigenetic marks in murine and human AD neurons, implicating TERT repression in amyloid-induced neurodegeneration. To test the impact of sustained TERT expression on AD pathobiology, AD mouse models were engineered to maintain physiological levels of TERT in adult neurons, resulting in reduced Aß accumulation, improved spine morphology, and preserved cognitive function. Mechanistically, integrated profiling revealed that TERT interacts with ß-catenin and RNA polymerase II at gene promoters and upregulates gene networks governing synaptic signaling and learning processes. These TERT-directed transcriptional activities do not require its catalytic activity nor telomerase RNA. These findings provide genetic proof-of-concept for somatic TERT gene activation therapy in attenuating AD progression including cognitive decline.

ROP18-Mediated Transcriptional Reprogramming of HEK293T Cell Reveals New Roles of ROP18 in the Interplay Between Toxoplasma gondii and the Host Cell.

Toxoplasma gondii secretes a number of virulence-related effector proteins, such as the rhoptry protein 18 (ROP18). To further broaden our understanding of the molecular functions of ROP18, we examined the transcriptional response of human embryonic kidney cells (HEK293T) to ROP18 of type I T. gondii RH strain. Using RNA-sequencing, we compared the transcriptome of ROP18-expressing HEK293T cells to control HEK293T cells. Our analysis revealed that ROP18 altered the expression of 750 genes (467 upregulated genes and 283 downregulated genes) in HEK293T cells. Gene ontology (GO) and pathway enrichment analyses showed that differentially expressed genes (DEGs) were significantly enriched in extracellular matrix- and immune-related GO terms and pathways. KEGG pathway enrichment analysis revealed that DEGs were involved in several disease-related pathways, such as nervous system diseases and eye disease. ROP18 significantly increased the alternative splicing pattern "retained intron" and altered the expression of 144 transcription factors (TFs). These results provide new insight into how ROP18 may influence biological processes in the host cells via altering the expression of genes, TFs, and pathways. More in vitro and in vivo studies are required to substantiate these findings.

Telomere dysfunction activates YAP1 to drive tissue inflammation.

Germline telomere maintenance defects are associated with an increased incidence of inflammatory diseases in humans, yet whether and how telomere dysfunction causes inflammation are not known. Here, we show that telomere dysfunction drives pATM/c-ABL-mediated activation of the YAP1 transcription factor, up-regulating the major pro-inflammatory factor, pro-IL-18. The colonic microbiome stimulates cytosolic receptors activating caspase-1 which cleaves pro-IL-18 into mature IL-18, leading to recruitment of interferon (IFN)-γ-secreting T cells and intestinal inflammation. Correspondingly, patients with germline telomere maintenance defects exhibit DNA damage (γH2AX) signaling together with elevated YAP1 and IL-18 expression. In mice with telomere dysfunction, telomerase reactivation in the intestinal epithelium or pharmacological inhibition of ATM, YAP1, or caspase-1 as well as antibiotic treatment, dramatically reduces IL-18 and intestinal inflammation. Thus, telomere dysfunction-induced activation of the ATM-YAP1-pro-IL-18 pathway in epithelium is a key instigator of tissue inflammation.

Chromatin Regulator CHD1 Remodels the Immunosuppressive Tumor Microenvironment in PTEN-Deficient Prostate Cancer.

Genetic inactivation of PTEN is common in prostate cancer and correlates with poorer prognosis. We previously identified CHD1 as an essential gene in PTEN-deficient cancer cells. Here, we sought definitive in vivo genetic evidence for, and mechanistic understanding of, the essential role of CHD1 in PTEN-deficient prostate cancer. In Pten and Pten/Smad4 genetically engineered mouse models, prostate-specific deletion of Chd1 resulted in markedly delayed tumor progression and prolonged survival. Chd1 deletion was associated with profound tumor microenvironment (TME) remodeling characterized by reduced myeloid-derived suppressor cells (MDSC) and increased CD8+ T cells. Further analysis identified IL6 as a key transcriptional target of CHD1, which plays a major role in recruitment of immunosuppressive MDSCs. Given the prominent role of MDSCs in suppressing responsiveness to immune checkpoint inhibitors (ICI), our genetic and tumor biological findings support combined testing of anti-IL6 and ICI therapies, specifically in PTEN-deficient prostate cancer. SIGNIFICANCE: We demonstrate a critical role of CHD1 in MDSC recruitment and discover CHD1/IL6 as a major regulator of the immunosuppressive TME of PTEN-deficient prostate cancer. Pharmacologic inhibition of IL6 in combination with immune checkpoint blockade elicits robust antitumor responses in prostate cancer.This article is highlighted in the In This Issue feature, p. 1241.

Tumor Microenvironment Remodeling Enables Bypass of Oncogenic KRAS Dependency in Pancreatic Cancer.

Oncogenic KRAS (KRAS*) is a key tumor maintenance gene in pancreatic ductal adenocarcinoma (PDAC), motivating pharmacologic targeting of KRAS* and its effectors. Here, we explored mechanisms involving the tumor microenvironment (TME) as a potential basis for resistance to targeting KRAS*. Using the inducible Kras G12D;Trp53 -/- PDAC mouse model, gain-of-function screens of epigenetic regulators identified HDAC5 as the top hit enabling KRAS* independent tumor growth. HDAC5-driven escaper tumors showed a prominent neutrophil-to-macrophage switch relative to KRAS*-driven tumors. Mechanistically, HDAC5 represses Socs3, a negative regulator of chemokine CCL2, resulting in increased CCL2, which recruits CCR2+ macrophages. Correspondingly, enforced Ccl2 promotes macrophage recruitment into the TME and enables tumor recurrence following KRAS* extinction. These tumor-associated macrophages in turn provide cancer cells with trophic support including TGFß to enable KRAS* bypass in a SMAD4-dependent manner. Our work uncovers a KRAS* resistance mechanism involving immune cell remodeling of the PDAC TME. SIGNIFICANCE: Although KRAS* is required for PDAC tumor maintenance, tumors can recur following KRAS* extinction. The capacity of PDAC cancer cells to alter the TME myeloid cell composition to support KRAS*-independent tumor growth illuminates novel therapeutic targets that may enhance the effectiveness of therapies targeting KRAS* and its pathway components.See related commentary by Carr and Fernandez-Zapico, p. 910.This article is highlighted in the In This Issue feature, p. 890.

iTRAQ-based Quantitative Proteomics Analysis Identifies Host Pathways Modulated during Toxoplasma gondii Infection in Swine.

Toxoplasma gondii is a leading cause of foodborne illness and consumption of undercooked pig meat is a major risk factor for acquiring toxoplasmosis, which causes a substantial burden on society. Here, we used isobaric tags for relative and absolute quantification (iTRAQ) labelling coupled with liquid chromatography-tandem mass spectrometry (LC-MS/MS) to identify cellular proteins and pathways altered during T. gondii infection in pigs. We also used parallel reaction monitoring-based LC-MS/MS to verify the levels of protein expression of infected spleens and mesenteric lymph nodes (MLNs). At 6 days post-infection (dpi), 156, 391, 170, 292, and 200 differentially expressed proteins (DEPs) were detected in the brain, liver, lung, MLNs and spleen, respectively. At 18 dpi, 339, 351, 483, 388, and 303 DEPs were detected in the brain, liver, lung, MLNs and spleen, respectively. Although proteins involved in immune responses were upregulated in all infected tissues, protein expression signature in infected livers was dominated by downregulation of the metabolic processes. By weighted gene co-expression network analysis, we could further show that all proteins were clustered into 25 co-expression modules and that the pink module significantly correlated with the infection status. We also identified 163 potential anti-T. gondii proteins (PATPs) and provided evidence that two PATPs (HSP70.2 and PDIA3) can reduce T. gondii burden in porcine macrophages in vitro. This comprehensive proteomics analysis reveals new facets in the pathogenesis of T. gondii infection and identifies key proteins that may contribute to the pig's defense against this infection.

Transcriptional changes in Toxoplasma gondii in response to treatment with monensin.

BACKGROUND: Infection with the apicomplexan protozoan parasite T. gondii can cause severe and potentially fatal cerebral and ocular disease, especially in immunocompromised individuals. The anticoccidial ionophore drug monensin has been shown to have anti-Toxoplasma gondii properties. However, the comprehensive molecular mechanisms that underlie the effect of monensin on T. gondii are still largely unknown. We hypothesized that analysis of T. gondii transcriptional changes induced by monensin treatment can reveal new aspects of the mechanism of action of monensin against T. gondii. METHODS: Porcine kidney (PK)-15 cells were infected with tachyzoites of T. gondii RH strain. Three hours post-infection, PK-15 cells were treated with 0.1 µM monensin, while control cells were treated with medium only. PK-15 cells containing intracellular tachyzoites were harvested at 6 and 24 h post-treatment, and the transcriptomic profiles of T. gondii-infected PK-15 cells were examined using high-throughput RNA sequencing (RNA-seq). Quantitative real-time PCR was used to verify the expression of 15 differentially expressed genes (DEGs) identified by RNA-seq analysis. RESULTS: A total of 4868 downregulated genes and three upregulated genes were identified in monensin-treated T. gondii, indicating that most of T. gondii genes were suppressed by monensin. Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis of T. gondii DEGs showed that T. gondii metabolic and cellular pathways were significantly downregulated. Spliceosome, ribosome, and protein processing in endoplasmic reticulum were the top three most significantly enriched pathways out of the 30 highly enriched pathways detected in T. gondii. This result suggests that monensin, via down-regulation of protein biosynthesis in T. gondii, can limit the parasite growth and proliferation. CONCLUSIONS: Our findings provide a comprehensive insight into T. gondii genes and pathways with altered expression following monensin treatment. These data can be further explored to achieve better understanding of the specific mechanism of action of monensin against T. gondii.

The Multitasking Fasciola gigantica Cathepsin B Interferes With Various Functions of Goat Peripheral Blood Mononuclear Cells in vitro.

Cathepsin B, a lysosomal cysteine protease, is thought to be involved in the pathogenesis of Fasciola gigantica infection, but its exact role remains unclear. In the present study, a recombinant F. gigantica cathepsin B (rFgCatB) protein was expressed in the methylotrophic yeast Pichia pastoris. Western blot analysis confirmed the reactivity of the purified rFgCatB protein to serum from F. gigantica-infected goats. The effects of serial concentrations (10, 20, 40, 80, and 160 µg/ml) of rFgCatB on various functions of goat peripheral blood mononuclear cells (PBMCs) were examined. We demonstrated that rFgCatB protein can specifically bind to the surface of PBMCs. In addition, rFgCatB increased the expression of cytokines (IL-2, IL-4, IL-10, IL-17, TGF-ß, and IFN-γ), and increased nitric oxide production and cell apoptosis, but reduced cell viability. These data show that rFgCatB can influence cellular and immunological functions of goat PBMCs. Further characterization of the posttranslational modification and assessment of rFgCatB in immunogenicity studies is warranted.

Global Transcriptome Profiling of Multiple Porcine Organs Reveals Toxoplasma gondii-Induced Transcriptional Landscapes.

We characterized the porcine tissue transcriptional landscapes that follow Toxoplasma gondii infection. RNAs were isolated from liver, spleen, cerebral cortex, lung, and mesenteric lymph nodes (MLNs) of T. gondii-infected and uninfected (control) pigs at days 6 and 18 postinfection, and were analyzed using next-generation sequencing (RNA-seq). T. gondii altered the expression of 178, 476, 199, 201, and 362 transcripts at 6 dpi and 217, 223, 347, 119, and 161 at 18 dpi in the infected brain, liver, lung, MLNs and spleen, respectively. The differentially expressed transcripts (DETs) were grouped into five expression patterns and 10 sub-clusters. Gene Ontology enrichment and pathway analysis revealed that immune-related genes dominated the overall transcriptomic signature and that metabolic processes, such as steroid biosynthesis, and metabolism of lipid and carboxylic acid, were downregulated in infected tissues. Co-expression network analysis identified transcriptional modules associated with host immune response to infection. These findings not only show how T. gondii infection alters porcine transcriptome in a tissue-specific manner, but also offer a gateway for testing new hypotheses regarding human response to T. gondii infection.

Symbiotic Macrophage-Glioma Cell Interactions Reveal Synthetic Lethality in PTEN-Null Glioma.

Heterotypic interactions across diverse cell types can enable tumor progression and hold the potential to expand therapeutic interventions. Here, combined profiling and functional studies of glioma cells in glioblastoma multiforme (GBM) models establish that PTEN deficiency activates YAP1, which directly upregulates lysyl oxidase (LOX) expression. Mechanistically, secreted LOX functions as a potent macrophage chemoattractant via activation of the ß1 integrin-PYK2 pathway in macrophages. These infiltrating macrophages secrete SPP1, which sustains glioma cell survival and stimulates angiogenesis. In PTEN-null GBM models, LOX inhibition markedly suppresses macrophage infiltration and tumor progression. Correspondingly, YAP1-LOX and ß1 integrin-SPP1 signaling correlates positively with higher macrophage density and lower overall survival in GBM patients. This symbiotic glioma-macrophage interplay provides therapeutic targets specifically for PTEN-deficient GBM.

KRAS-IRF2 Axis Drives Immune Suppression and Immune Therapy Resistance in Colorectal Cancer.

The biological functions and mechanisms of oncogenic KRASG12D (KRAS∗) in resistance to immune checkpoint blockade (ICB) therapy are not fully understood. We demonstrate that KRAS∗ represses the expression of interferon regulatory factor 2 (IRF2), which in turn directly represses CXCL3 expression. KRAS∗-mediated repression of IRF2 results in high expression of CXCL3, which binds to CXCR2 on myeloid-derived suppressor cells and promotes their migration to the tumor microenvironment. Anti-PD-1 resistance of KRAS∗-expressing tumors can be overcome by enforced IRF2 expression or by inhibition of CXCR2. Colorectal cancer (CRC) showing higher IRF2 expression exhibited increased responsiveness to anti-PD-1 therapy. The KRAS∗-IRF2-CXCL3-CXCR2 axis provides a framework for patient selection and combination therapies to enhance the effectiveness of ICB therapy in CRC.