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Background Long non-coding RNAs (lncRNAs) play important roles in the biology of colorectal cancer (CRC). There are several lncRNAs associated with invasion and metastasis have been characterized in CRC. However, studies focusing on the precise molecular mechanisms by which lncRNAs function in lymph node (LN) metastasis in CRC are still limited. Methods In this study, by analyzing TCGA dataset, we identified that AC244100.2 (termed CCL14-AS), a novel lncRNA enriched in the cytoplasm, was negatively correlated with LN metastasis and unfavorable prognosis of CRC. In situ hybridization was used to examine CCL14-AS expression in clinical CRC tissues. Various functional experiments including migration assay and wound-healing assay were used to investigate the effects of CCL14-AS on CRC cells migration. The nude mice popliteal lymph node metastasis model assay further confirmed the effects of CCL14-AS in vivo. Results CCL14-AS expression was significantly downregulated in CRC tissues compared to adjacent normal tissues. In addition, low CCL14-AS expression was correlated with advanced T classification, LN metastasis, distant metastasis, and shorter disease-free survival of CRC patients. Functionally, CCL14-AS overexpression inhibited the invasiveness of CRC cells in vitro and LN metastasis in nude mice. On the contrary, knockdown of CCL14-AS promoted the invasiveness and LN metastasis abilities of CRC cells. Mechanistically, CCL14-AS downregulated the expression of MEP1A via interacting with MEP1A mRNA and reduced its stability. Overexpression of MEP1A rescued the invasiveness and LN metastasis abilities in CCL14-AS-overexpressing CRC cells. Moreover, the expression levels of CCL14-AS was negatively correlated with that of MEP1A in CRC tissues. Conclusions We identified a novel lncRNA, CCL14-AS, as a potential tumor suppressor in CRC. Our findings supported a model in which the CCL14-AS/MEP1A axis serves as critical regulator in CRC progression, suggesting a novel biomarker and therapeutic target in advanced 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 β1/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 β1/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.

Breast cancer (BC) is a serious threat to women’s health and metastasis is the major cause of BC-associated mortality. Various techniques are currently used to preoperatively describe the metastatic status of tumors, based on which a comprehensive treatment protocol was determined. However, accurately staging a tumor before surgery remains a challenge, which may lead to the miss of optimal treatment options. More severely, the failure to detect and remove occult micrometastases often causes tumor recurrences. There is an urgent need to develop a more precise and non-invasive strategy for the detection of the tumor metastasis in lymph nodes and distant organs. Based on the facts that tumor metastasis is closely related to the primary tumor microenvironment (TME) evolutions and that metabolomics profiling of the circulatory system can precisely reflect subtle changes within TME, we suppose whether metabolomic technology can be used to achieve non-invasive and real-time monitoring of BC metastatic status. In this study, the metastasis status of BC mouse models with different tumor-bearing times was firstly depicted to mimic clinical anatomic TNM staging system. Metabolomic profiling together with metastasis-related changes in TME among tumor-bearing mice with different metastatic status was conducted. A range of differential metabolites reflecting tumor metastatic states were screened and in vivo experiments proved that two main metastasis-driving factors in TME, TGF-β and hypoxia, were closely related to the regular changes of these metabolites. The differential metabolites level changes were also preliminarily confirmed in a limited number of clinical BC samples. Metabolite lysoPC (16:0) was found to be useful for clinical N stage diagnosis and the possible cause of its changes was analyzed by bioinformatics techniques.

BackgroundForkhead box S1 (FOXS1) is a member of the forkhead box (FOX) transcriptional factor superfamily. The biological roles and underlying regulatory mechanism of FOXS1 in CRC remain unclear.MethodsBioinformatics analysis, Western blotting, real-time PCR, and immunohistochemistry (IHC) were used to detect the expression FOXS1 in CRC. MTT assay, transwell assay, human umbilical vein endothelial cell tube formation assay, and chicken chorioallantoic membrane assay were performed to investigate the effects of FOXS1 on proliferation, invasion, and angiogenesis. Additionally, tumor formation assay and orthotopic implantation assay were used to investigate the effects of FOXS1 on tumor growth and metastasis in vivo. Furthermore, gene set enrichment analysis (GSEA) was used to analyze the correlation between FOXS1 and EMT or angiogenesis. The correlation between FOXS1 and CXCL8 expression was analyzed in clinical CRC samples using IHC.ResultsThe results showed that FOXS1 expression was upregulated in CRC tissues compared with adjacent normal intestine tissues. A high FOXS1 expression is positively correlated with poor survival. FOXS1 promoted the malignant behavior of CRC cancer cells in vitro, including proliferation, invasion, and angiogenesis. In addition, FOXS1 promoted tumor growth and metastasis in nude mice. Mechanistically, FOXS1 upregulated the expression of C–X–C motif chemokine ligand 8 (CXCL8) at the transcriptional level. Knockdown of CXCL8 blocked FOXS1 induced the enhancement of the EMT and angiogenesis. GSEAs in public CRC datasets revealed strong correlations between FOXS1 expression and EMT marker and angiogenesis markers. IHC showed that FOXS1 expression was positively correlated with CXCL8 expression and CD31 expression in clinical CRC samples.ConclusionThe results suggest that FOXS1 promotes angiogenesis and metastasis by upregulating CXCL8 in CRC. Interference with the FOXS1/CXCL8 axis may serve as a potential therapeutic target for the treatment of metastatic CRC.

Tumor metastasis is one of the major causes of high mortality in patients with hepatocellular carcinoma (HCC). Sustained activation of STAT3 signaling plays a critical role in HCC metastasis. RNA binding protein (RBP)–mediated posttranscriptional regulation is involved in the precise control of signal transduction, including STAT3 signaling. In this study, we investigated whether RBPs are important regulators of HCC metastasis. The RBP MEX3C was found to be significantly upregulated in highly metastatic HCC and correlated with poor prognosis in HCC. Mechanistically, MEX3C increased JAK2/STAT3 pathway activity by downregulating SOCS3, a major negative regulator of JAK2/STAT3 signaling. MEX3C interacted with the 3′UTR of SOCS3 and recruited CNOT7 to ubiquitinate and accelerate decay of SOCS3 mRNA. Treatment with MEX3C-specific antisense oligonucleotide significantly inhibited JAK2/STAT3 pathway activation, suppressing HCC migration in vitro and metastasis in vivo. These findings highlight a novel mRNA decay-mediated mechanism for the disruption of SOCS3-driven negative regulation of JAK2/STAT3 signaling, suggesting MEX3C may be a potential prognostic biomarker and promising therapeutic target in HCC. Significance: This study reveals that RNA-binding protein MEX3C induces SOCS3 mRNA decay to promote JAK2/STAT3 activation and tumor metastasis in hepatocellular carcinoma, identifying MEX3C targeting as a potential approach for treating metastatic disease.

Triple-negative breast cancer (TNBC) is the most aggressive subtype of breast cancer with a high risk of metastasis and recurrence. Although chemotherapy has greatly improved the clinical outcome of TNBC patients, acquired drug resistance remains a huge challenge for TNBC treatment. Breast cancer stem cells (BCSCs) play a critical role in breast cancer development, metastasis, recurrence, and chemotherapy resistance. Thus, it is of great importance to decipher the underlying molecular mechanism of BCSCs regulation for TNBC drug resistance. In this study, we demonstrate that the F-box protein FBXL2 is a critical negative regulator of BCSCs stemness and that downregulation of FBXL2 plays a causal role in TNBC drug resistance. We show that expression levels of FBXL2 significantly influence CD44

Gender exerts a profound impact on cancer incidence, spectrum and outcomes, yet the molecular genetic bases of such sex differences are ill-defined and presumptively ascribed to X-chromosome genes and sex hormones. Such sex differences are particularly prominent in colorectal cancer (CRC) where 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 tumor suppressors (designated iKAP), 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 up-regulated gene driven by KRAS*-mediated activation of the STAT4 transcription factor. KDM5D-dependent chromatin mark and transcriptome changes showed repression of regulators of epithelial cell tight junction and MHC 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 exhibited an increased propensity for more invasive tumors in vivo. Thus, KRAS*-STAT4-mediated upregulation of Y chromosome KDM5D contributes significantly to the sex differences in KRAS* CRC via its disruption of cancer cell adhesion properties and tumor immunity, thus providing an actionable therapeutic strategy for metastasis risk reduction for men afflicted with KRAS* CRC. Citation Format: Jiexi Li, Zhengdao Lan, Wenting Liao, Jielin Liu, Shan Jiang, Hong Seok Shim, Max Slotnik, Kyle A. LaBella, Wen-Hao Hsu, Xiaoying Shang, Denise Spring, Y Alan Wang, Ronald A. DePinho. Histone demethylase KDM5D drives sex-specific differences in colorectal cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 3515.

Gender exerts a profound impact on cancer incidence, spectrum and outcomes, yet the molecular genetic bases of such sex differences are ill-defined and presumptively ascribed to X-chromosome genes and sex hormones. Such sex differences are particularly prominent in colorectal cancer (CRC) where men experience higher metastases and mortality. A murine CRC model, engineered with an inducible oncogenic Kras transgene (KRAS*) and conditional null alleles of Apc and Trp53 tumor suppressors (designated iKAP), revealed higher metastases and worse outcomes specifically in males with KRAS* CRC. Integrated cross-species molecular and transcriptomic analyses identified Y-chromosome gene histone demethylase KDM5D as a transcriptionally up-regulated gene driven by KRAS*-mediated activation of the STAT4 transcription factor. KDM5D-dependent chromatin mark and transcriptome changes showed repression of regulators of epithelial cell tight junction and MHC 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 exhibited an increased propensity for more invasive tumors in vivo. Thus, KRAS*-STAT4-mediated upregulation of Y chromosome KDM5D contributes significantly to the sex differences in KRAS* CRC via its disruption of cancer cell adhesion properties and tumor immunity, thus providing an actionable therapeutic strategy for metastasis risk reduction for men afflicted with KRAS* CRC. Citation Format: Jiexi Li, Zhengdao Lan, Wenting Liao, James W. Horner, Jielin Liu, Shan Jiang, Hong S. Shim, Max Slotnik, Kyle A. LaBella, Wen-Hao Hsu, Denise J. Spring, Y. Alan Wang, Ronald A. DePinho. Histone demethylase KDM5D drives sex-specific differences in colorectal cancer [abstract]. In: Proceedings of the AACR Special Conference on Colorectal Cancer; 2022 Oct 1-4; Portland, OR. Philadelphia (PA): AACR; Cancer Res 2022;82(23 Suppl_1):Abstract nr PR011.

Transducin-like enhancer of Split3 (TLE3) serves as a transcriptional corepressor during cell differentiation and shows multiple roles in different kinds of cancers. Recently, TLE3 together with many other genes involved in Wnt/β-catenin pathway were detected hyper-methylated in colorectal cancer (CRC). However, the potential role and the underlying mechanism of TLE3 in CRC progression remain scarce. Gene expression profiles were analyzed in The Cancer Genome Atlas (TCGA) microarray dataset of 41 normal colorectal intestine tissues and 465 CRC tissues. Western blot and Real-time Quantitative PCR (RT-qPCR) were respectively performed to detect protein and mRNA expression in 8 pairs of CRC tissue and matched adjacent normal mucosa. Immunohistochemistry (IHC) was conducted to evaluate TLE3 protein expression in 105 paraffin-embedded, archived human CRC tissues from patients, whose survival data were analyzed with Kaplan-Meier method. In vitro experiments including MTT assay, colony formation assay, and soft agar formation assay were used to investigate the effects of TLE3 on CRC cell growth and proliferation. Additionally, subcutaneous tumorigenesis assay was performed in nude mice to confirm the effects of TLE3 in vivo. Furthermore, gene set enrichment analysis (GSEA) was run to explore potential mechanism of TLE3 in CRC, and then we measured the distribution of CRC cell cycle phases and apoptosis by flow cytometry, as well as the impacts of TLE3 on MAPK and AKT signaling pathways by Western blot and RT-qPCR. TLE3 was significantly down-regulated in 465 CRC tissues compared with 41 normal tissues. Both protein and mRNA expressions of TLE3 were down-regulated in CRC compared with matched adjacent normal mucosa. Lower expression of TLE3 was significantly associated with poorer survival of patients with CRC. Besides, knock down of TLE3 promoted CRC cell growth and proliferation, while overexpression of TLE3 showed suppressive effects. Furthermore, overexpression of TLE3 caused G1-S phase transition arrest, inhibition of MAPK and AKT pathways, and up-regulation of p21Cip1/WAF1 and p27Kip1. This study indicated that TLE3 repressed CRC proliferation partly through inhibition of MAPK and AKT signaling pathways, suggesting the possibility of TLE3 as a biomarker for CRC prognosis.

The ubiquitin-specific protease (USP) family is the largest group of cysteine proteases. Cancer genomic analysis identified frequent amplification of USP21 (22%) in human pancreatic ductal adenocarcinoma (PDAC). USP21 overexpression correlates with human PDAC progression, and enforced expression of USP21 accelerates murine PDAC tumor growth and drives PanIN to PDAC progression in immortalized human pancreatic ductal cells. Conversely, depletion of USP21 impairs PDAC tumor growth. Mechanistically, USP21 deubiquitinates and stabilizes the TCF/LEF transcription factor TCF7, which promotes cancer cell stemness. Our work identifies and validates USP21 as a PDAC oncogene, providing a potential druggable target for this intractable disease.

Background Approximately 1–2% of non-small cell lung cancer (NSCLC) patients harbor RET (rearranged during transfection) fusions. The oncogenic RET fusions could lead to constitutive kinase activation and oncogenesis. Methods 1746 Chinese NSCLC patients were analyzed in this study. Tumor tissues were collected, and were formalin fixed, paraffin-embedded (FFPE) and archived. Peripheral blood (PB) samples were also collected from each patient as control. In addition, we selected 17 of them for cfDNA NGS testing and 14 tumor samples for immunohistochemistry testing using PD-L1 rabbit monoclonal antibody, clones 28-8 (Abcam, Cambridge, UK). Results Of the 1746 NSCLC cases, RET rearrangements were identified in 25 cases (1.43%) with locally advanced or metastatic NSCLC, of which 20 (80%) were female. We found that 14 out of 25 patients had an KIF5B-RET fusion, with KIF5B exon15-RET exon12, KIF5B exon23-RET exon12, and KIF5B exon24-RET exon11 detected in 14, 3, and 1 patients, respectively. We also identified one novel RET fusion partner PLCE1 and 4 intergenic-breakpoint fusions. Conclusion In this study, using the hybrid capture based next generation sequencing (NGS) techniques, we revealed the genomic profiling for the patients with RET fusion-positive NSCLC. To the best of our knowledge, this is the first study that exhibited the detailed breakpoints of Chinese NSCLC patients with RET rearrangement, and we found a novel new partner PLCE1. The results provided genomic information for patients with RET fusion which is significant for personalized clinical management in the era of precision medicine.

e22519 Background: Human papillomavirus (HPV) is a family of small double-stranded DNA viruses. It is associated with a series of cancer types, such as cervical, oropharyngeal, anal, vulvar, vaginal, and penile cancers. In the world, nearly 70% of HPV-related cancer cases are HPV types 16 and 18. In this study, we investigated HPV infection prevalence among a cohort of Chinese solid tumor patients. Methods: The study enrolled thousands of Chinese patients with different types of cancers. We finally collected 4,606 tumor tissue DNA (ttDNA), 7,296 plasma cell-free DNA (cfDNA), and 7,509 peripheral blood mononuclear cell genomic DNA (gDNA) samples, respectively. Using a targeted capture technique that covering the whole reference genome of HPV16 and HPV18 and the next generation sequencing (NGS) technology, we calculated the percentage of HPV16 and HPV 18 reads in all sequencing reads for each sample. Finally, we explored the prevalence of HPV infection in each cancer type. Results: Based on different types of DNA samples, the HPV infection rate was 38.75%, 28.62%, and 10.43% in ttDNA, cfDNA, and gDNA samples, respectively. Besides, the magnitudes of the mean percentage of HPV reads in all the sequencing reads in ttDNA, cfDNA and gDNA samples were 10-5, 10-6 and 10-8, respectively, while in the HPV infection positive the percentages of ttDNA, cfDNA, and gDNA samples increased to 10-4, 10-5, and 10-7, respectively. The prevalence of HPV infection rate in different cancer types was shown in table. Conclusions: ttDNA and cfDNA are better material for HPV infective detection compared with gDNA, and NGS is capable of HPV detection.[Table: see text]

Patients with colorectal cancer (CRC) often develop malignant regrowth of metastatic dormant tumor cells in liver years after primary treatment. FBX8 is involved in suppressing tumor metastasis. Short-term chemotherapy experiments and liver metastasis mice model of orthotopic injection into the cecum were performed to construct the dormant models. GST-pull-down assay, Co-IP and immunofluorescence were used to confirm the bindings among FBX8 and its substrates. FBX8 upregulated the expression of epithelial and stemness markers, while downregulated the expression of mesenchymal and proliferative markers associated with tumor cell dormancy. FBX8 promoted the maintenance of metastatic dormancy of CRC cells. Mechanistically, FBX8 directly bound to HIF-1α, CDK4 and C-myc through its Sec7 domain and led to the ubiquitin degradation of these proteins, thereby inhibiting cell cycle progression, proliferation, angiogenesis, and metastasis. Clinically, FBX8 expression was negatively correlated with the HIF-1α, CDK4, and c-Myc in CRC tissues. Our study reveals a novel mechanism of FBX8 in regulating tumor metastatic dormancy in liver and provides new strategies for the treatment of CRC metastasis.

Background cAMP responsive element binding protein 5 (CREB5) is a transcriptional activator in eukaryotic cells that can regulate gene expression. Previously, we found that CREB5 was involved in the occurrence and development of colorectal cancer (CRC) using bioinformatics analysis. However, the biological roles and underlying regulatory mechanism of CREB5 in CRC remain unclear. Methods Real-time PCR, western blotting, and immunohistochemistry were used to examine CREB5 expression. In vitro experiments including migration assay, wound-healing assay, chicken chorioallantoic membrane assay, and human umbilical vein endothelial cells tube formation assay were used to investigate the effects of CREB5 on CRC cell migration and tumor angiogenesis ability. Additionally, an orthotopic implantation assay was performed in nude mice to confirm the effects of CREB5 in vivo. Furthermore, gene set enrichment analysis was performed to explore the potential mechanism of CREB5 in CRC. Results We found that CREB5 expression was highly upregulated in CRC. CREB5 overexpression was positively correlated with advanced WHO stages and TNM stages and shorter survival in CRC patients. Moreover, CREB5 overexpression promoted while CREB5 silencing reduced the invasiveness and metastatic capacity of CRC cells both in vitro and in vivo. Furthermore, CREB5 directly interacted with the MET promoter and activated the hepatocyte growth factor-MET signalling pathway. Importantly, inhibition of MET reduced the invasion and metastasis of CREB5-overexpressing CRC cells, suggesting that CREB5 promotes metastasis mainly through activation of MET signalling. Conclusion Our study demonstrates a crucial role for CREB5 in CRC metastasis by directly upregulating MET expression. CREB5 may be both a potential prognostic marker and a therapeutic target to effectively overcome metastasis in CRC.

Anaplastic lymphoma kinase (ALK) fusion events account for ~3–7% genetic alterations in patients with non-small cell lung cancer (NSCLC). In this study, we identified the ALK fusion patterns and a novel ALK fusion partner in 44 ALK positive NSCLC patients using a customized HapOncoCDx panel, and identified ALK fusion partners. The most common partner is EML4, forming the variant 1 (v1, E13:A20, 18/44), variant 2 (v2, E20:A20, 5/44), and variant 3 (v3, E6:A20, 13/44). Moreover, we detected a new ALK fusion partner HMBOX1. At the mutation level, TP53 is the most frequently mutated gene (24%), followed by ALK (12%) and STED2 (12%). The median tumor mutation burden (TMB) of these samples is 2.29 mutations/Mb, ranging from 0.76 mut/Mb to 16.79 muts/Mb. We further elaborately portrayed the TP53 mutation sites on the peptide sequence of the encoded protein by lollipop. The mutational signature and copy number alterations (CNAs) of the samples were also analyzed. The CNA events were found in 13 (13/44) patients, and the most commonly amplified genes were MDM2 (n = 4/13) and TERT (n = 4/13). Together, these results may guide personalized clinical management of patients with ALK fusion in the era of precision medicine.

Cancer-derived exosomes are considered a major driver of cancer-induced pre-metastatic niche formation at foreign sites, but the mechanisms remain unclear. Here, we show that miR-25-3p, a metastasis-promoting miRNA of colorectal cancer (CRC), can be transferred from CRC cells to endothelial cells via exosomes. Exosomal miR-25-3p regulates the expression of VEGFR2, ZO-1, occludin and Claudin5 in endothelial cells by targeting KLF2 and KLF4, consequently promotes vascular permeability and angiogenesis. In addition, exosomal miR-25-3p from CRC cells dramatically induces vascular leakiness and enhances CRC metastasis in liver and lung of mice. Moreover, the expression level of miR-25-3p from circulating exosomes is significantly higher in CRC patients with metastasis than those without metastasis. Our work suggests that exosomal miR-25-3p is involved in pre-metastatic niche formation and may be used as a blood-based biomarker for CRC metastasis., The mechanisms underlying pre-metastatic niche formation by cancer derived exosomes is unclear. Here they show that colorectal cancer (CRC) derived exosomal miR-25-3p promotes vascular leakiness and angiogenesis, CRC metastasis, and is upregulated in CRC pateints with metastasis, and suggest miR-25-3p as a biomarker for CRC metastasis.

Jade family PHD finger 3 (JADE3) plays a role in inducing histone acetylation during transcription, and is involved in the progression of several human cancers; however, its role in colon cancer remains unclear. Herein, we found that JADE3 was markedly upregulated in colon cancer tissues and significantly correlated with cancer progression, and predicted shorter patient survival. Further, JADE3 was expressed much higher in colon cancer cell lines that are enriched with a stem-like signature. Overexpression of JADE3 increased, while silencing JADE3 reduced cancer stem cell-like traits in colon cancer cells in vitro and in vivo. Importantly, silencing of JADE3 strongly impaired the tumor initiating capacity of colon cancer cells in vivo. Furthermore, JADE3 interacted with the promoters of colon stem cell marker LGR5 and activated its transcription, by increasing the occupancy of p300 acetyltransferase and histone acetylation on the promoters. Finally, we found that JADE3 expression was substantially induced by Wnt/β-catenin signaling. These findings suggest an oncogenic role of JADE3 by regulating cancer stem cell-like traits in the colon cancer, and therefore JADE3 might be a potential therapeutic target for the treatment of colon cancer.

Recently, invadopodia have been increasingly recognized as important drivers of local invasion and metastasis. Cortactin, as an actin-binding protein, is closely associated with invadopodia through interacting with proteins. Formin-like 2 (FMNL2), a member of diaphanous-related formins which act as nucleation factors, plays an important role in tumor progression. But whether cortactin can interact with FMNL2 to promote invadopodia formation and the role of FMNL2 in invadopodia formation are still unknown. Here we found that cortactin directly bound to FMNL2 and elevated the activities of actin polymerization and recycling endosome motility. FMNL2 was necessary for invadopodia formation and function in CRC cells. The interaction of cortactin and FMNL2 could further promote the invadopodia formation and matrix degradation. The stimulation of EGF/cdc42 enhanced the combination of cortactin and FMNL2 to intensify the numbers of invadopodia and the degrees of matrix degradation. In vivo, induction of invadopodia formation via cortactin is essential for the ability of FMNL2 to promote CRC metastasis. Furthermore, up-regulations of FMNL2 and cortactin were highly linked in CRC tissues. Collectively, our work demonstrates a brand-new mechanism of cortacin and FMNL2 at invadopodia in CRC.

In this issue of Cancer Cell, Aftab et al. identify a pro-inflammatory cytokine, IL-33, that is released as a chemoattractant for type 2 immune cells in response to the intratumoral mycobiome. Depletion of fungi or deletion of IL-33 in cancer cells significantly decreases pancreatic ductal adenocarcinoma (PDAC) tumor progression and increases survival.

Radioresistance hampers success in the treatment of patients with advanced colorectal cancer (CRC). Improving our understanding of the underlying mechanisms of radioresistance could increase patients’ response to irradiation (IR). MicroRNAs are a class of small RNAs involved in tumor therapy response to radiation. Here we found that miR-214 was markedly decreased in CRC cell lines and blood of CRC patients after IR exposure. Meanwhile, autophagy was enhanced in irradiated CRC cells. Mechanically, ATG12 was predicted and identified as a direct target of miR-214 by dual luciferase assay, qPCR, and Western blot. In vitro and in vivo experiments showed that miR-214 promoted radiosensitivity by inhibiting IR-induced autophagy. Restoration of ATG12 attenuated miR-214-mediated inhibition of cell growth and survival in response to IR. Importantly, miR-214 was highly expressed in radiosensitive CRC specimens and negatively correlated with plasma level of CEA. Moreover, ATG12 and LC3 expressions were increased in radioresistant CRC specimens. Our study elucidates that miR-214 promotes radiosensitivity by inhibition of ATG12-mediated autophagy in CRC. Importantly, miR-214 is a determinant of CRC irradiation response and may serve as a potential therapeutic target in CRC treatment.

Purpose: To investigate the role and the underlying mechanism of scaffold attachment factor B (SAFB) in the progression of colorectal cancer (CRC). Experimental Design: SAFB expression was analyzed in the Cancer Outlier Profile Analysis of Oncomine and in 175 paraffin-embedded archived CRC tissues. Gene Ontology analyses were performed to explore the mechanism of SAFB in CRC progression. Western blot, RT-PCR, luciferase assay, and chromatin immunoprecipitation (ChIP) were used to detect the regulation of transforming growth factor-β–activated kinase 1 (TAK1) and NF-κB signaling by SAFB. The role of SAFB in invasion, metastasis, and angiogenesis was investigated using in vitro and in vivo assays. The relationship between SAFB and TAK1 was analyzed in CRC tissues. Results: SAFB was downregulated in CRC tissues, and low expression of SAFB was significantly associated with an aggressive phenotype and poorer survival of CRC patients. The downregulation of SAFB activated NF-κB signaling by targeting the TAK1 promoter. Ectopic expression of SAFB inhibited the development of aggressive features and metastasis of CRC cells both in vitro and in vivo. The overexpression of TAK1 could rescue the aggressive features in SAFB-overexpressed cells. Furthermore, the expression of SAFB in CRC tissues was negatively correlated with the expression of TAK1- and NF-κB–related genes. Conclusions: Our results show that SAFB regulated the activity of NF-κB signaling in CRC by targeting TAK1. This novel mechanism provides a comprehensive understanding of both SAFB and the NF-κB signaling pathway in the progression of CRC and indicates that the SAFB–TAK1–NF-κB axis is a potential target for early therapeutic intervention in CRC progression. Clin Cancer Res; 23(22); 7108–18. ©2017 AACR.

Background: Diaphanous-related formins (DRFs), actin necleator, have been known to participate in the progression of cancer cells. We previously reported that FMNL2 (Formin-like2), a member of DRFs, was a positive regulator in colorectal cancer (CRC) metastasis, yet proteins and pathways required for the function of this pro-invasive DRFs remain to be identified. Methods: The relationship between FMNL2 and COMMD10 was examined using Co-IP, GST pull-down, immunofluorescence and in vitro ubiquitination assay. The in vitro and in vivo function of COMMD10 in CRC was evaluated using CCK-8 proliferation assay, plate colony formation, cell cycle, apoptosis and animal models. The inhibition of NF-κB signalling by COMMD10 was detected using dual-luciferase reporter assay and western blotting. Co-IP, GST pull-down and nuclear protein extraction assay were performed to evaluate the effect on p65 by COMMD10. Real-time PCR and western blotting were performed to detect expressions of FMNL2, COMMD10 and p65 in paired tissues. Results: FMNL2 targets COMMD10 for ubiquitin-mediated proteasome degradation in CRC cells. COMMD10 targets p65 NF-κB (nuclear factor-κB) subunit and reduces its nuclear translocation, thereby leading to the inactivation of NF-κB pathway and suppression of CRC invasion and metastasis. Inhibition of NF-κB signalling by COMMD10 is necessary for FMNL2-mediated CRC cell behaviours. Downregulation of COMMD10 predicts poor prognosis of CRC patients. The expressions of FMNL2, COMMD10 and p65 are highly linked in CRC tissues. Conclusions: These data demonstrate that the FMNL2/COMMD10/p65 axis acts as a critical regulator in the maintenance of metastatic phenotypes and is strongly associated with negative clinical outcomes.

Hypoxic preconditioning (HPC) is well‑known to exert a protective effect against hypoxic injury; however, the underlying molecular mechanism remains unclear. The present study utilized a serum metabolomics approach to detect the alterations associated with HPC. In the present study, an animal model of HPC was established by exposing adult BALB/c mice to acute repetitive hypoxia four times. The serum samples were collected by orbital blood sampling. Metabolite profiling was performed using ultra‑performance liquid chromatography‑quadrupole time‑of‑flight mass spectrometry (UPLC‑QTOFMS), in conjunction with univariate and multivariate statistical analyses. The results of the present study confirmed that the HPC mouse model was established and refined, suggesting significant differences between the control and HPC groups at the molecular levels. HPC caused significant metabolic alterations, as represented by the significant upregulation of valine, methionine, tyrosine, isoleucine, phenylalanine, lysophosphatidylcholine (LysoPC; 16:1), LysoPC (22:6), linoelaidylcarnitine, palmitoylcarnitine, octadecenoylcarnitine, taurine, arachidonic acid, linoleic acid, oleic acid and palmitic acid, and the downregulation of acetylcarnitine, malate, citrate and succinate. Using MetaboAnalyst 3.0, a number of key metabolic pathways were observed to be acutely perturbed, including valine, leucine and isoleucine biosynthesis, in addition to taurine, hypotaurine, phenylalanine, linoleic acid and arachidonic acid metabolism. The results of the present study provided novel insights into the mechanisms involved in the acclimatization of organisms to hypoxia, and demonstrated the protective mechanism of HPC.

F-box proteins are critical components of the SKP1-CUL1-F-box (SCF) E3 ubiquitin ligases and involved in the ubiquitin-dependent proteolytic pathway. Dysregulation of F-box protein-mediated proteolysis often leads to human malignancies. F-box only protein 8 (FBX8), a novel component of F-box proteins, is down-regulated in several tumors and closely correlates with tumor progression. However, little is known about its function, regulatory mechanisms and substrates in the progression of colorectal carcinoma (CRC). Combining microRNA (miRNA) assay, functional characterization, mechanistic studies with clinical validation, we identify FBX8 as a CRC metastasis suppressor downstream of miR-223, a metastasis promoting miRNA that is transcriptionally regulated by Myocyte enhancer factor (MEF2A). mTOR is a substrate of FBX8 for ubiquitin-mediated degradation and is required for FBX8 induced cell proliferation and invasion in CRC cells. FBX8 is down-regulated in human CRC tissues and correlates with MEF2A, miR-223 and mTOR expression levels. Notably, low FBX8 expression status in CRC tissues was a significant prognostic factor for poor overall survival of patients. These findings illustrate FBX8 as a metastasis suppressor that functions through mTOR signaling pathway and has significant prognostic power.

Background Ubinuclein-2 (UBN2) is a nuclear protein that interacts with many transcription factors. The molecular role and mechanism of UBN2 in the development and progression of cancers, including colorectal cancer (CRC), is not well understood. The current study explored the role of UBN2 in the development and progression CRC. Methods Oncomine network and The Cancer Genome Atlas (TCGA) database were downloaded and Gene Set Enrichment Analysis (GSEA) was performed to compare the UBN2′s expression between normal and tumor tissues, as well as the potential correlation of UBN2 expression with signaling pathways. Immunohistochemistry (IHC), qRT-PCR and Western blotting were performed to determine the expression of UBN2 in CRC tissues or cell lines. In vitro proliferation and invasion assays, and orthotopic mouse metastatic model were used to analyze the effect of UBN2 on the development and progression of CRC. Results The analysis of UBN2 expression using Oncomine network showed that UBN2 was upregulated in CRC tissues compared to matched adjacent normal intestinal epithelial tissues. IHC, qRT-PCR and Western blotting confirmed that UBN2 expression is higher in CRC tissues compared with matched adjacent normal intestinal epithelial tissues. In addition, analyses of TCGA data revealed that high UBN2 expression was associated with advanced stages of lymph node metastasis, distant metastasis, and short survival time in CRC patients. IHC showed that high UBN2 expression is correlated with advanced stages of CRC. Moreover, UBN2 is highly expressed in the liver metastatic lesions. Furthermore, knockdown of UBN2 inhibited the growth, invasiveness and metastasis of CRC cells via regulation of the Ras/MAPK signaling pathway. Conclusion The current study demonstrates that UBN2 promotes tumor progression in CRC. UBN2 may be used as a promising biomarker for predicting the prognosis of CRC patients. Electronic supplementary material The online version of this article (10.1186/s12935-019-0848-4) contains supplementary material, which is available to authorized users.

Background Cancer associated fibroblasts (CAFs) are key stroma cells that play dominant roles in tumor progression. However, the CAFs-derived molecular determinants that regulate colorectal cancer (CRC) metastasis and chemoresistance have not been fully characterized. Methods CAFs and NFs were obtained from fresh CRC and adjacent normal tissues. Exosomes were isolated from conditioned medium and serum of CRC patients using ultracentrifugation method and ExoQuick Exosome Precipitation Solution kit, and characterized by transmission electronic microscopy, nanosight and western blot. MicroRNA microarray was employed to identify differentially expressed miRNAs in exosomes secreted by CAFs or NFs. The internalization of exosomes, transfer of miR-92a-3p was observed by immunofluorescence. Boyden chamber migration and invasion, cell counting kit-8, flow cytometry, plate colony formation, sphere formation assays, tail vein injection and primary colon cancer liver metastasis assays were employed to explore the effect of NFs, CAFs and exosomes secreted by them on epithelial-mesenchymal transition, stemness, metastasis and chemotherapy resistance of CRC. Luciferase report assay, real-time qPCR, western blot, immunofluorescence, and immunohistochemistry staining were employed to explore the regulation of CRC metastasis and chemotherapy resistance by miR-92a-3p, FBXW7 and MOAP1. Results CAFs promote the stemness, epithelial-mesenchymal transition (EMT), metastasis and chemotherapy resistance of CRC cells. Importantly, CAFs exert their roles by directly transferring exosomes to CRC cells, leading to a significant increase of miR-92a-3p level in CRC cells. Mechanically, increased expression of miR-92a-3p activates Wnt/β-catenin pathway and inhibits mitochondrial apoptosis by directly inhibiting FBXW7 and MOAP1, contributing to cell stemness, EMT, metastasis and 5-FU/L-OHP resistance in CRC. Clinically, miR-92a-3p expression is significantly increased in CRC tissues and negatively correlated with the levels of FBXW7 and MOAP1 in CRC specimens, and high expression of exosomal miR-92a-3p in serum was highly linked with metastasis and chemotherapy resistance in CRC patients. Conclusions CAFs secreted exosomes promote metastasis and chemotherapy resistance of CRC. Inhibiting exosomal miR-92a-3p provides an alternative modality for the prediction and treatment of metastasis and chemotherapy resistance in CRC. Electronic supplementary material The online version of this article (10.1186/s12943-019-1019-x) contains supplementary material, which is available to authorized users.

e13068 Background: Breast cancer (BC) is the most commonly diagnosed cancer in 2020, with an estimated 2.3 million new cases and the leading cause of cancer death in women. The cytoskeleton is a dynamic system of protein filaments that is regulated by dozens of proteins which are, in turn, regulated by phosphoinositides. Cytoskeletal regulation and vesicle trafficking (CRVT) signaling pathway is associated with many changes in the structure and movements of cells. Few studies addressing the association between genetic mutation of the CRVT pathway and the disease prognosis have been reported up to now. Here we investigate the genetic status of CRVT pathway as a biomarker for predicting overall survival (OS) and relapse-free survival (RFS) for breast cancer patients. Methods: We used cBioPortal platform to analyze a cohort of 10646 breast cancer samples. Mutations in 7 genes ( APC, BAP1, KEAP1, PAK1, PTEN, SRC, STAT3 ) are the most common indication in the CRVT signaling pathway. We define overall survival (OS) as the time between the procedure date when the tumor specimen was collected and the date of death or last follow-up visit. And we define relapse-free survival (RFS) as the time from surgical resection to relapse of disease. We estimated and compared the survival curves of these two groups using Kaplan-Meier method and log-rank tests. The statistical significance level was set to 5% ( p < 0.05). Results: Genomic alterations of the CRVT-indicator genes were found in 2582 (n = 2582/10646, 24.25%) samples with sorts of alterations identified including variants of non-synonymous mutations, splicing mutations, short in-frame insertion and deletion, short frame shift insertion and deletion, and copy number gain and loss. PTEN was most commonly altered (n = 2329), followed by PAK1 (n = 863), APC (n = 718), STAT3 (n = 341), KEAP1 (n = 317), BAP1(n = 292), and SRC (n = 278). Based on the result, patients were divided into the CRVT and non-CRVT group. The median OS of the CRVT group was 141.57 moth, which was significantly shorter than the OS of the non-CRVT group (log-rank test p= 0.000662). The median RFS of the CRVT group was 167.76 moth, which was also significantly shorter than the RFS of the non-CRVT group (log-rank test p= 0.0169). Conclusions: To the best of our knowledge, this study is the first investigation to evaluate the prognostic value of CRVT pathway mutation in patients with breast cancer. Our investigation shows that CRVT pathway mutation could significantly predict poor prognosis in breast cancer patients. The result of our analysis should be better confirmed with additional relevant research in the future using updated analyses.

Background: MiR-214 is aberrantly regulated in several tumours, but its underlying mechanisms in colorectal cancer (CRC) metastasis remain largely unknown. This study aimed to demonstrate the function and potential mechanism of miR-214 in regulating invasion and metastasis of CRC. Methods: The transcription factor and targets of miR-214 were predicted by bioinformatics and validated using ChIP and dual-luciferase reporter assay. DNA methylation status was explored using bisulphite sequencing PCR. The in vitro and in vivo function of miR-214 in CRC was evaluated using MTT, plate colony formation, Matrigel invasion and animal models. Real-time PCR or western blotting was performed to detect FOXD3, miR-214 and MED19 expressions in CRC cells and clinical specimens. Results: MiR-214 was downregulated in CRC and was significantly correlated with lymphatic metastasis. Downregulation of miR-214 might due to promoter hypermethylation in CRC. FOXD3 was validated as a transcription factor of miR-214 by ChIP assay. Dual-luciferase assay identified MED19 as a target of miR-214 in CRC. In vitro and in vivo experiments showed that miR-214 mediated the inhibiting effect of FOXD3 on proliferation, invasion and metastasis by targeting MED19. Spearman's correlation analysis showed a positive correlation between FOXD3 and miR-214, and negative correlations between FOXD3 and MED19, miR-214 and MED19 in CRC cells and clinical specimens. Conclusions: FOXD3/miR-214/MED19 axis is important for the regulation of growth, invasion and metastasis of CRC. Targeting the miR-214-mediated axis might be helpful for the treatment of CRC.

// Ya-Ping Ye 1, 2, 3, * , Ping Wu 1, 2, 3, * , Chun-cai Gu 1, 2, 3, * , Dan-ling Deng 1, 2, 3 , Hong-Li Jiao 1, 2, 3, Ting-Ting Li 1, 2, 3 , Shu-Yang Wang 1, 2, 3 , Yong-Xia Wang 1, 2, 3 , Zhi-Yuan Xiao 1, 2, 3 , Wen-ting Wei 1, 2, 3 , Yan-Ru Chen 1, 2, 3 , Jun-Feng Qiu 1, 2, 3 , Run-Wei Yang 1, 2, 3 , Jie Lin 1, 2, 3 , Li Liang 1, 2, 3 , Wen-Ting Liao 1, 2, 3 , Yan-Qing Ding 1, 2, 3 1 Department of Pathology, Nanfang Hospital, Southern Medical University, Guangzhou, Guangdong, China 2 Department of Pathology, School of Basic Medical Sciences, Southern Medical University, Guangzhou, Guangdong, China 3 Guangdong Province Key Laboratory of Molecular Tumor Pathology, Guangzhou, Guangdong, China * These authors have contributed equally to this work Correspondence to: Wen-Ting Liao, email: liaowt2002@gmail.com Yan-Qing Ding, email: dyq@fimmu.com Keywords: miR-450b-5p, Wnt/β-Catenin pathway, colorectal cancer, progression, KRAS Received: May 13, 2016 Accepted: July 14, 2016 Published: August 2, 2016 ABSTRACT The development and progression of CRC are regarded as a complicated network and progressive event including genetic and/or epigenetic alterations. Recent researches revealed that MicroRNAs are biomarkers and regulators of CRC progression. Analyses of published microarray datasets revealed that miR-450b-5p was highly up-regulated in CRC tissues. In addition, high expression of miR-450b-5p was significantly associated with KRAS mutation. However, the role of miR-450b-5p in the progression of CRC remains unknown. Here, we sought to validate the expression of miR-450b-5p in CRC tissues and investigate the role and underlying mechanism of miR-450b-5p in the progression of CRC. The results revealed that miR-450b-5p was up-regulated in CRC tissues, high expression level of miR-450b-5p was positively associated with poor differentiation, advanced TNM classification and poor prognosis. Moreover, miR-450b-5p was especially high in KRAS-mutated cell lines and could be up-regulated by KRAS/AP-1 signaling. Functional validation revealed that overexpression of miR-450b-5p promoted cell proliferation and tumor growth while inhibited apoptosis of CRC cells. Furthermore, we demonstrated that miR-450b-5p directly bound the 3’-UTRs of SFRP2 and SIAH1, and activated Wnt/β-Catenin signaling. In conclusion, miR-450b-5p induced by oncogenic KRAS is required for colorectal cancer progression. Collectively, our work helped to understand the precise role of miR-450b-5p in the progression of CRC, and might promote the development of new therapeutic strategies against CRC.

// Huijun Xie 1, 2, 3, * , Xiaoli Ren 1, 2, 3, * , Sainan Xin 1, 2, 3, * , Xiaoliang Lan 4 , Guifeng Lu 1, 2, 3 , Yuan Lin 1, 2, 3 , Shaoshan Yang 1, 2, 3 , Zhicheng Zeng 1, 2, 3 , Wenting Liao 1, 2, 3 , Yan-Qing Ding 1, 2, 3 , Li Liang 1, 2, 3 1 Department of Pathology, Nanfang Hospital, Guangzhou, Guangdong, China 2 Department of Pathology, School of Basic Medical Sciences, Southern Medical University, Guangzhou, Guangdong, China 3 Guangdong Province Key Laboratory of Molecular Tumor Pathology, Guangzhou, Guangdong, China 4 Department of General Surgery, Nanfang Hospital, Guangzhou, Guangdong, China * These authors contributed equally to this work Correspondence to: Li Liang, email: redsnow007@hotmail.com Yan-Qing Ding, email: dyq@fimmu.com Keywords: hsa_circ_001569, miR-145, colorectal cancer, FMNL2, BAG4 Received: December 08, 2015 Accepted: March 07, 2016 Published: April 05, 2016 ABSTRACT Circular RNAs (circRNAs), a large class of RNAs, have recently shown huge capabilities as gene regulators in mammals. Some of them bind with microRNAs (miRNAs) and act as natural miRNA sponges to inhibit related miRNAs’ activities. Here we showed that hsa_circ_001569 acted as a positive regulator in cell proliferation and invasion of colorectal cancer (CRC). Moreover, hsa_circ_001569 was identified as a sponge of miR-145 and up-regulated miR-145 functional targets E2F5, BAG4 and FMNL2. In CRC tissues, circ_001569 negatively correlated with miR-145, and miR-145 correlated negatively with E2F5, BAG4 and FMNL2 expressions. Our study reveals a novel regulatory mechanism of circ_001569 in cell proliferation and invasion in CRC, provides a comprehensive landscape of circ_001569 that will facilitate further biomarker discoveries in the progression of CRC.

Colorectal cancer (CRC) is one of the most common malignancies and is the second leading cause of cancer death in humans. Tumour suppressor candidate 3 (TUSC3) plays an important role in embryogenesis and metabolism. Deletion of TUSC3 often causes non-syndromic mental retardation. Even though TUSC3 deregulation is frequently observed in epithelial cancers, the function of TUSC3 in CRC has remained unknown. In this study, we observed greater expression of TUSC3 at the mRNA and protein level in clinical colorectal tumour samples compared with paired normal tissues. Gain- and loss-of-function analyses were performed to evaluate the functional significance of TUSC3 in CRC initiation and progression. Immunoblotting, immunofluorescence, and co-immunoprecipitation analyses were used to identify potential pathways with which TUSC3 might be involved. Overexpression of TUSC3 in CRC cells induced epithelial-mesenchymal transition (EMT) in CRC cells, accompanied by down-regulation of the epithelial marker, E-cadherin, and up-regulation of the mesenchymal marker, vimentin. Increased proliferation, migration, and invasion, as well as accelerated xenograft tumour growth, were observed in TUSC3-overexpressing CRC cells, while opposite effects were achieved in TUSC3-silenced cells. In conclusion, our study demonstrated the oncogenic role of TUSC3 in CRC and showed that TUSC3 may be responsible for alternations in the proliferation ability, aggressiveness, and invasive/metastatic potential of CRC through regulating the MAPK, PI3K/Akt, and Wnt/β-catenin signalling pathways.

This presentation will highlight our use of mouse models to dissect molecular and cellular interactions in the tumor microenvironment, including (i) the illumination of the role of myeloid cells in driving immune suppression in prostate cancer, and the impact of combined MDSC-targeted and immune checkpoint blockade therapies in the treatment of metastatic castration-resistant prostate cancer; (ii) the generation and analysis of oncogenic Kras in an inducible colorectal cancer mouse model, revealing the role of IRF2 in the recruitment of myeloid cells into the tumor microenvironment and informing patient selection for more effective ICB therapy in CRC; (iii) the concept of “synthetic essentiality” (a means by which to target specific tumor-suppressor gene deficiencies in cancer), exemplified by the identification of the role of CHD1 in modulation of the tumor microenvironment in Pten-deficient prostate cancer; (iv) the discovery of a symbiotic interaction between glioma cell (LOX) and macrophage (SPP1) in PTEN-null glioma, as well as macrophage reprogramming in immune suppression in glioblastoma; and (v) the role of KRAS in cancer metabolism, in particular how the tumor microenvironment may support KRAS-directed cancer cell metabolism and provide a nonautonomous mechanism enabling escape from Kras-dependent tumor growth. These illustrative examples provide translational opportunities to improve cancer patient treatment and survival. Citation Format: Adam Boutin, Peiwen Chen, Prasenjit Dey, Pingping Hou, Wen-Ting Liao, Xin Lu, Haoqiang Ying, Di Zhao, Ronald A. DePinho. Modeling and understanding tumor biologic mechanisms [abstract]. In: Proceedings of the AACR Special Conference on the Evolving Landscape of Cancer Modeling; 2020 Mar 2-5; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2020;80(11 Suppl):Abstract nr IA01.

e13546 Background: Liver cancer is the sixth most common cancer in the world and the fourth most common cause of cancer mortality. Genomic alterations including homologous recombination deficiency (HRD) mutations often lead to aberrant regulations of signaling pathways, which play an important role in tumorigenesis. The prognostic value of HRD mutation and its correlation with overall survival in liver cancer is unclear. Methods: We used cBioPortal platform to analyze a cohort of 978 liver cancer samples. Mutations in 13 genes (BRCA1, BRCA2, ATM, BARD1, BRIP1, CHEK1, CHEK2, ABRAXAS1, MRE11, NBN, PALB2, RAD51C, RAD51D) were screened due to the fact that they are the most common indication of deficiency in the homologous recombination (HR) DNA repair pathway. We define overall survival (OS) as the time between the sample collection (surgical) date and the date of death or the last follow-up visit. We conducted Kaplan-Meier survival analysis, and the statistical differences were compared by log-rank tests. Results: 228 of the cases harbored at least one mutation of the HRD-indicator genes. Based on the result, patients were divided into the HRD and non-HRD group. The median OS of the HRD group was 45.47, which is significantly shorter than the OS of the non-HRD group (log-rank test p = 1.6196e-4, Table). Conclusions: HRD mutations are significantly associated with a shorter OS in liver cancer patients. HRD mutation was a negative prognostic factor for overall survival in liver cancer, but the underlying mechanisms remain to be explored. [Table: see text]

e13544 Background: Hepatitis B virus (HBV) infection in hepatocytes may induce chronic liver damage, which further leads to hepatocellular carcinoma. HBV infection was also reported in other cell types including peripheral blood mononuclear cells (PBMCs). Moreover, epidemiological evidence suggested that HBV infection was associated with various types of cancers. In this study, we investigated whether HBV DNA integration was observed in extrahepatic tumor cells. Methods: The study enrolled thousands of Chinese patients with different types of cancers. Peripheral blood and tumor tissue samples were collected, plasma cfDNA, PBMC gDNA and tumor tissue DNA were extracted. By targeted capture and next-generation sequencing, we sequenced the cancer-related genes and identified HBV DNA integration events in these samples. Moreover, the subtype of HBV captured in the host genome was verified, and the ratio of reads mapped to the HBV genome was calculated. Results: Samples with more than 0.1% of the sequencing reads mapped to the HBV genome were identified as HBV positive cases. Thirty-two liver cancer patients were subtype-b HBV positive, among which 8 tumor samples exhibited HBV genome integration. Three colorectal cancer patients were infected by subtype-c HBV, only one tumor sample carried HBV DNA integration. Two lung cancer patients were shown to be HBV positive; one was infected by subtype-b HBV and the other carried subtype-c HBV. Subtype-b and-c HBV genome was observed in the tumor sample of two cholangiocarcinoma patients, whereas subtype-b HBV DNA was identified in the PBMC of one gastrointestinal stromal cancer patient. Conclusions: HBV genome integration was identified in the tumor cells of lung, colorectal, biliary ducts, and gastrointestinal stromal cancer. The mechanisms by which the HBV genome integrates into these tissues requires further investigation.

The precise biologic functions and mechanisms of oncogenic KrasG12D (Kras*) in therapeutic resistance, including immune therapy, are not understood. Here, utilizing a colorectal cancer (CRC) mouse model engineered with an inducible Kras* transgene and conditional null APC and Trp53 alleles (iKAP) that we recently established (Boutin et al., Gene & Development 2017), we observed that KRAS*-expressing invasive tumors possess a prominent immune-suppressive profile characterized by a preponderance of myeloid-derived suppressor cells (MDSCs). Integrated transcriptomic and genomic analyses identified the Interferon Regulatory Factor 2 (IRF2) gene as a key downstream target of KRAS* mediating tumor progression and immune suppression. Mechanistically, integrated transcriptomics, cytokine profiling and ChIP-seq analysis identified CXCL3 as a key direct target of IRF2, and pharmacologic inhibition of CXCL3 or its receptor CXCR2 resulted in suppression of MDSC migration and infiltration in KRAS*-expressing CRC tumors. Translational studies established that de novo anti-PD1 resistance of KRAS*-expressing tumors can be overcome by enforced IRF2 expression or by treatment with small-molecule CXCR2 antagonist SX-682. Correspondingly, CRC patient tumors with higher IRF2 expression show increased response to anti-PD1 therapy. This KRAS*-mediated immune suppressive mechanism provides a framework for patient selection and combination therapies to enhance the effectiveness of immune checkpoint blockade therapy in advanced CRC. Citation Format: Wenting Liao, Michael Overman, Adam Boutin, Scott Kopetz, Ron DePinho, Alan Wang. KRAS-IRF2 axis drives immune suppression and immune therapy resistance in colorectal cancer [abstract]. In: Proceedings of the AACR Special Conference on Targeting RAS-Driven Cancers; 2018 Dec 9-12; San Diego, CA. Philadelphia (PA): AACR; Mol Cancer Res 2020;18(5_Suppl):Abstract nr B52.

Oncogenic Kras (Kras*) plays a key role in tumor maintenance and therapeutic resistance in several cancer types, although the precise biologic functions and mechanisms are not completely understood. We had previously shown in pancreatic ductal adenocarcinoma that some tumors underwent spontaneous relapse and were devoid of Kras* expression and downstream canonical MAPK signaling, instead acquiring amplification and overexpression of the transcriptional coactivator Yap1. Functional studies established the role of Yap1 and the transcriptional factor Tead2 in driving Kras*-independent tumor maintenance. In a second previous study, we developed a faithful colorectal cancer (CRC) mouse model and showed that Kras* drove invasion and maintenance in CRC. Here, utilizing this CRC mouse model, we demonstrate that KRAS* promotes a pronounced immune-suppressive profile distinguished by a predominance of myeloid-derived suppressor cells (MDSCs). We performed integrated transcriptomic and genomic analyses that identified IRF2 (Interferon Regulatory Factor 2) as a key KRAS* effector mediating immune suppression. IRF2 suppresses MDSC migration and infiltration in KRAS*-expressing CRC tumors by targeting the CXCL3/CXCR2 axis. We also observed that de novo resistance of KRAS*-expressing tumors to anti-PD1 therapy could be overcome by enforced IRF2 expression or by treatment with SX-682, a small-molecule CXCR2 antagonist. In addition, CRC patient tumors with higher IRF2 expression showed increased response to anti-PD1 therapy. This KRAS*-mediated immune-suppressive mechanism provides a framework for CRC patient selection and combination therapies to enhance the effectiveness of immune checkpoint blockade therapy in advanced disease. Citation Format: Wen-Ting Liao, Michael J. Overman, Adam Boutin, Prasenjit Dey, Di Zhao, Guocan Wang, Jiexi Li, Zhengdao Lan, Jun Li, Xiaoying Shang, Ming Tang, Shan Jiang, Xingdi Ma, Peiwen Chen, Riham Katkhuda, Krittiya Korphaisarn, Deepavali Chakravarti, Qing Chang, Jianhua Zhang, Dipen M. Maru, Dean Y. Maeda, John A. Zebala, Scott Kopetz, Y. Alan Wang, Ronald A. DePinho. Context-dependent role of KRAS in GI malignancies [abstract]. In: Proceedings of the AACR Special Conference on Targeting RAS-Driven Cancers; 2018 Dec 9-12; San Diego, CA. Philadelphia (PA): AACR; Mol Cancer Res 2020;18(5_Suppl):Abstract nr IA27.

As an inhibitor of heat shock proteins (HSPs), KNK437 has been reported to play an anti-tumor role in several cancers. But its therapeutic effect and mechanisms in colorectal cancer (CRC) remain unclear. Here, KNK437 sharply inhibited the level of DnaJ heat shock protein family (Hsp40) member A1 (DNAJA1), followed by DNAJB1, but had little effect on the levels of HSP27, HSP105, HSP90, and HSP70 in CRC cells. DNAJA1 promoted CRC cell proliferation in vitro and tumor growth and metastasis in vivo. Mechanistically, DNAJA1 was activated by E2F transcription factor 1 (E2F1) and then promoted cell cycle by stabilizing cell division cycle protein 45 (CDC45), which could be reversed by KNK437. DNAJA1 was significantly upregulated in CRC tissues and positively correlated with serosa invasion, lymph node metastasis. High level of DNAJA1 predicted poor prognosis for CRC patients. Its expression was highly linked with E2F1 and CDC45 in CRC tissues. More importantly, KNK437 significantly suppressed the growth of DNAJA1 expressing tumor in vivo. The combined treatment of KNK437 with 5-FU/L-OHP chemotherapy reduced liver metastasis of CRC. These data reveal a novel mechanism of KNK437 in anti-tumor therapy of CRC and provides a newly therapeutic strategy with potential translation to the CRC patients.

Background Colorectal cancer (CRC) is one of the most common digestive malignant tumors, and DMTN is a transcriptionally differentially expressed gene that was identified using CRC mRNA sequencing data from The Cancer Genome Atlas (TCGA). Our preliminary work suggested that the expression of DMTN was downregulated in CRC, and the Rac1 signaling pathway was significantly enriched in CRC tissues with low DMTN expression. However, the specific functions and underlying molecular mechanisms of DMTN in the progression of CRC and the upstream factors regulating the downregulation of the gene remain unclear. Methods DMTN expression was analyzed in CRC tissues, and the relationship between DMTN expression and the clinicopathological parameters was analyzed. In vitro and in vivo experimental models were used to detect the effects of DMTN dysregulation on invasion and metastasis of CRC cells. GSEA assay was performed to explore the mechanism of DMTN in invasion and metastasis of CRC. Westernblot, Co-IP and GST-Pull-Down assay were used to detect the interaction between DMTN and ARHGEF2, as well as the activation of the RAC1 signaling. Bisulfite genomic sequence (BSP) assay was used to test the degree of methylation of DMTN gene promoter in CRC tissues. Results We found that the expression of DMTN was significantly decreased in CRC tissues, and the downregulation of DMTN was associated with advanced progression and poor survival and was regarded as an independent predictive factor of CRC patient prognosis. The overexpression of DMTN inhibited, while the knockdown of DMTN promoted, invasion and metastasis in CRC cells. Moreover, hypermethylation and the deletion of DMTN relieved binding to the ARHGEF2 protein, activated the Rac1 signaling pathway, regulated actin cytoskeletal rearrangements, and promoted the invasion and metastasis of CRC cells. Conclusion Our study demonstrated that the downregulation of DMTN promoted the metastasis of colorectal cancer cells by regulating the actin cytoskeleton through RAC1 signaling activation, potentially providing a new therapeutic target to enable cancer precision medicine for CRC patients. Electronic supplementary material The online version of this article (10.1186/s13046-018-0958-1) contains supplementary material, which is available to authorized users.

Forkhead box F1 (FOXF1) has been recently implicated in the progression and metastasis of lung cancer and breast cancer. However, the biological functions and underlying mechanisms by which FOXF1 regulates the progression of colorectal cancer (CRC) are largely unknown. As shown in our previous study, FOXF1 is upregulated in 182 CRC tissues, and elevated FOXF1 expression is significantly associated with microvessel density and advanced TNM (T = primary tumour; N = regional lymph nodes; M = distant metastasis) stages. In this study, 43 CRC tissues collected from patients who underwent treatment with first-line standard chemotherapeutic regimens in combination with bevacizumab were used to explore the correlation between FOXF1 expression and resistance to bevacizumab. In addition, FOXF1 regulated angiogenesis by inducing the transcription of vascular endothelial growth factor A1 (VEGFA) in vitro and in vivo. Furthermore, upregulation of FOXF1 enhanced bevacizumab resistance in CRC, and inhibition of VEGFA attenuated angiogenesis and bevacizumab resistance in FOXF1-overexpressing CRC cells. These results suggest that FOXF1 plays critical roles in CRC angiogenesis and bevacizumab resistance by inducing VEGFA transcription and that FOXF1 represents a potentially new therapeutic strategy and biomarker for anti-angiogenic therapy against CRC.

Forkhead Box F1 (FOXF1) has been recently implicated in cancer progression and metastasis of lung cancer and breast cancer. However, the biological functions and underlying mechanisms of FOXF1 in the regulation of the progression of colorectal cancer (CRC) are largely unknown. We showed that FOXF1 was up-regulated in 93 paraffin-embedded archived human CRC tissue, and both high expression and nuclear location of FOXF1 were significantly associated with the aggressive characteristics and poorer survival of CRC patients. The GSEA analysis showed that the higher level of FOXF1 was positively associated with an enrichment of EMT gene signatures, and exogenous overexpression of FOXF1 induced EMT by transcriptionally activating SNAI1. Exogenous overexpression FOXF1 functionally promoted invasion and metastasis features of CRC cells, and inhibition of SNAI1 attenuates the invasive phenotype and metastatic potential of FOXF1-overexpressing CRC cells. Furthermore, the results of the tissue chip showed that the expression of FOXF1 was positively correlated with SNAI1 in CRC tissues chip. These results suggested that FOXF1 plays a critical role in CRC metastasis by inducing EMT via transcriptional activation of SNAI1, highlighting a potential new therapeutic strategy for CRC.

Metastatic progression is the main contributor to the poor prognosis of colorectal cancer (CRC). Thus, identifying the determinants of CRC metastasis will be of great significance. Based on our previous bioinformatics analysis, Syntaxin2 (STX2) may be upregulated and correlated with the poor prognosis of CRC patients. In this study, we found that STX2 expression was associated with CRC invasion and metastasis and poor patient survival. Gain- and loss-of-function analyses demonstrated that STX2 functioned as a key oncogene by promoting CRC invasion and metastasis. Mechanistically, STX2 selectively interacted with tumor necrosis factor receptor-associated factor 6 (TRAF6) and activated the nuclear transcription factor-κB (NF-κB) signaling pathway. Furthermore, chromatin immunoprecipitation (ChIP) analysis revealed that NF-κB directly bound to the STX2 promoter and drove STX2 transcription. Therefore, STX2 activated the NF-κB pathway, and in turn, NF-κB increased STX2 expression, forming a positive signaling loop that eventually promoted CRC metastasis. Collectively, our results reveal STX2 as a crucial modulator of the aggressive CRC phenotype and highlight STX2 as a potential prognostic biomarker and therapeutic target for combating CRC metastasis.

The chemotherapy drug 5-fluorouracil (5-FU) is fundamental for the treatment of colorectal cancer (CRC); however, drug resistance to 5-FU may occasionally occur. Abnormal expression of Forkhead box C2 gene (Foxc2) has been identified in several human cancers, but the role of Foxc2 in the progression of CRC remains unclear. The present study established a stable Foxc2-short hairpin (sh)RNA cell line, which was confirmed by western blot analysis and quantitative polymerase chain reaction. The Foxc2-shRNA cells were treated with 5-FU and the cell viability was determined by an MTT assay. Western blot analysis was performed to investigate the signaling pathway involved in 5-FU treatment. The present study identified that 5-FU increased the percentage of apoptotic CRC cells among the Foxc2/RNA interference-transfected cells compared with cells transfected with an empty vector. Therefore, the downregulation of Foxc2, induced by 5-FU, may enhance apoptosis by the downregulation of apoptotic factors, including B cell lymphoma-2 and pro-caspase-3, in Foxc2-shRNA CRC cells. Furthermore, the mitogen-activated protein kinase (MAPK) and phosphatidylinositide 3-kinases/protein kinase B (PI3K/AKT) pathways were essential for the sensitization effect of Foxc2 to 5-FU treatment. Overall, these findings reveal the mechanisms behind Foxc2 depletion and 5-FU treatment of CRC and suggest that Foxc2 enhances resistance to apoptosis, induced by 5-FU, through the activation of MAPK and P13K/AKT pathways, and may serve as a valuable clinical prognostic marker for CRC.

DAB2IP has been identified as a tumor suppressor in several cancers but its oncogenic role and transcriptionally regulatory mechanisms in the progression of colorectal carcinoma (CRC) remain unknown. In this study, DAB2IP was down-regulated in CRC tissues and a valuable prognostic marker for survival of CRC patients, especially in the late stage. Moreover, DAB2IP was sufficient to suppress proliferation, epithelial-mesenchymal transition (EMT), invasion and metastasis in CRC. Mechanically, the linear complex of EZH2/HDAC1/Snail contributed to DAB2IP silencing in CRC cells. The study further proved that the positive feedback loop between Snail and DAB2IP existed in CRC cells and DAB2IP was required for Snail-induced aggressive cell behaviors. Finally, DAB2IP correlated negatively with Snail and EZH2 expressions in CRC tissues. Our findings reveal the suppressive role and a novel regulatory mechanism of DAB2IP expression in the progression of CRC. DAB2IP may be a potential, novel therapeutic and prognostic target for clinical CRC patients.

The Leucine zipper tumor suppressor gene 1 (LZTS1/FEZ1) gene was originally identified as a potential tumor suppressor. However, the expression pattern and the role of LZTS1 in the progression of colorectal cancer (CRC) have not been well characterized. Herein, we reported that LZTS1 was markedly reduced in CRC tissues compared with matched adjacent normal intestine epithelial tissues. In analysis of 160 CRC specimens, we revealed that decreased expression of LZTS1 was correlated to aggressive characteristics and poor survival of patients with CRC. Moreover, we found that expression of LZTS1 in CRC cells significantly inhibited cell proliferation in vitro and prohibited tumor growth in vitro. On the contrary, silence of LZTS1 promoted cell proliferation and tumor growth in CRC cells. Furthermore, we demonstrated that LZTS1 inhibited cell proliferation and tumor growth in CRC in part via suppression of AMT–mTOR, subsequently down-regulating p27Kip and up-regulating cyclin D1. These findings suggest that LZTS1 plays a potential tumor suppressor role in CRC progression and represents a valuable clinical prognostic marker of this disease.

// Yuling Li 1,2,* , Zhenbing Lv 1,3,4,* , Guoyang He 1,2 , Jianmei Wang 1,2 , Xiaojing Zhang 1,5 , Guifeng Lu 1,2 , Xiaoli Ren 1,2 , Feifei Wang 1,2 , Xiaohui Zhu 1,2 , Yi Ding 6 , Wenting Liao 1,2 , Yanqing Ding 1,2 and Li Liang 1,2 1 Department of Pathology, Nanfang Hospital, Southern Medical University, Guangzhou, Guangdong Province, People’s Republic of China 2 Guangdong Province Key Laboratory of Molecular Tumor Pathology, Guangzhou, Guangdong Province, People’s Republic of China 3 Department of General Surgery Two, Nanchong Central Hospital, Nanchong city, Sichuan Province, People’s Republic of China 4 The Second Clinical School of Northern Sichuan Medical College, Nanchong city, Sichuan Province, People’s Republic of China 5 Department of Pathology, Shenzhen University, Shenzhen, People’s Republic of China 6 Department of Radiotherapy, Nanfang Hospital, Southern Medical University, Guangzhou, Guangdong Province, People’s Republic of China * These authors contributed equally to this work Correspondence to: Li Liang, email: // Keywords : miR-371-5p, SOX17, SOX2, metastasis, colorectal cancer Received : January 15, 2015 Accepted : February 10, 2015 Published : March 15, 2015 Abstract Cancer stem cells (CSCs) and EMT-type cells, which share molecular characteristics with CSCs, have been believed to play critical roles in tumor metastasis. Although much progress has been garnered in elucidating the molecular pathways that trigger EMT, stemness and metastasis, a number of key mechanistic gaps remain elusive. In the study, miR-371-5p was obviously down-regulated in primary CRC tissues compared with matched adjacent normal mucosa and correlated significantly with differentiation, tumor size, lymphatic and liver metastases. MiR-371-5p could attenuate proliferation, invasion in vitro and metastasis in vivo in CRC cells. It also suppressed EMT by regulating Wnt /β-catenin signaling and strongly decreased the CRC stemness phenotypes. Moreover, demethylation of SOX17 induced miR-371-5p expression and consequently suppressed its direct target SOX2 in CRC cells. MiR-371-5p was necessary for SOX17 mediated cancer-related traits and SOX2 was a functional target of miR-371-5p. A positive relationship between SOX17 and miR-371-5p expression and a negative one between miR-371-5p and SOX2 expression were observed in CRC cell lines and tissues. In conclusion, we identified miR-371-5p as an important “oncosuppressor” in CRC progression and elucidated a novel mechanism of the SOX17/miR-371-5p/SOX2 axis in the regulation of EMT, stemness and metastasis, which may be a potential therapeutic target.

Background miRNAs are regarded as molecular biomarkers and therapeutic targets for colorectal cancer (CRC), a series of miRNAs have been proven to involve into CRC carcinogenesis, invasion and metastasis. Aberrant miR-422a expression and its roles have been reported in some cancers. However, the function and underlying mechanism of miR-422a in the progression of CRC remain largely unknown. Methods Real-time PCR were used to quantify miR-422a expression in CRC tissues. Both vivo and vitro functional assays showed miR-422a inhibits CRC cell proliferation. Target prediction program (miRBase) and luciferase reporter assays were conducted to confirm the target genes AKT1 and MAPK1 of miR-422a. Specimens from 50 patients with CRC were analyzed for the correlation between the expression of miR-422a and the expression of the target genes AKT1 and MAPK1 by real-time PCR. Results MiR-422a was down‑regulated in CRC tissues and cell lines. Ectopic expression of miR-422a inhibited cell proliferation and tumor growth ability; inhibition of endogenous miR-422a, by contrast, promoted cell proliferation and tumor growth ability of CRC cells. MiR-422a directly targets 3′-UTR of the AKT1 and MAPK1, down-regulation of miR-422a led to the activation of Raf/MEK/ERK and PI3K/AKT signaling pathways to promote cell proliferation in CRC. In addition, miR-422a expression was negatively correlated with the expressions of AKT1 and MAPK1 in CRC tissues. Conclusion miR-422a inhibits cell proliferation in colorectal cancer by targeting AKT1 and MAPK1. Electronic supplementary material The online version of this article (doi:10.1186/s12935-017-0461-3) contains supplementary material, which is available to authorized users.

Metastasis is the major cause of death in colorectal cancer (CRC). Although multiple genes have been identified to be responsible for the development of CRC, the molecular changes that enable CRC cells to undergo early local invasion and to form distant metastatic colonies still remain largely unknown. Herein, we investigated the role of Forkhead box protein C2 (FOXC2) and explored the underlying mechanisms in invasion and metastasis of CRC. We show that both high FOXC2 expression and nuclear localization of FOXC2 are significantly correlated with advanced TNM (T=primary tumor; N=regional lymph nodes; M=distant metastasis) stages. FOXC2 enhanced the invasive abilities of CRC cells in vitro and promoted local invasion and distant metastasis in an orthotopic mouse metastatic model of CRC. Microarray analysis revealed that overexpression of FOXC2 increased the proto-oncogene MET tyrosine kinase expression and activated the hepatocyte growth factor (HGF)-MET signaling pathway. Furthermore, luciferase reporter assays and chromatin immunoprecipitation assays revealed that FOXC2 directly associated with MET promoter to increase the transcriptional activity of MET. Inhibition of MET attenuates the invasive phenotype and metastatic potential of FOXC2-overexpressing CRC cells, indicating that MET is a major mediator of FOXC2-promoted metastasis. In addition, FOXC2 expression was positively correlated with MET expression in CRC tissue samples. Our findings suggest that FOXC2 has a crucial role in CRC metastasis by regulating HGF-MET signaling via inducing MET expression, highlighting FOXC2 as a potential therapeutic target for preventing or reducing metastasis in CRC.

A hallmark of pancreatic ductal adenocarcinoma (PDAC) is an exuberant stroma comprised of diverse cell types that enable or suppress tumor progression. This Kras*-driven heterotypic signaling circuit in the early and advance tumor microenvironment enables cooperative protumorigenic interactions in early steps of the neoplastic transformation process as well as in promoting growth of established tumors, providing novel therapeutic targets in the Kras* pathway for this intractable disease. Here, we explored the role of Kras* in protumorigenic signaling interactions between cancer cells and host cells in tumor microenvironment. Early-stage malignant lesions and advanced tumors show close proximity of Kras* expressing cancer cells and abundant invading TH2 (CD4+ Gata-3+) cells, which can produce IL-4 and IL-13. Single-cell analysis of low- and high-grade IPMN and PDAC patient samples revealed CD4+ T-cell infiltration and higher percentage of the CD4+ T cells are Gata3+ TH2 subtypes compared to T-bet+ TH1 cells. Similarly, imaging mass spectrometry analysis showed presence of CD4+ T cells in close proximity to cancer cells. Mechanistic studies identified Jak1-Stat6 as the primary signaling pathway downstream of IL-4/IL-13 activated IL-4 receptor complexes in cancer cells. Integrated transcriptomic and chromatin immunoprecipitation sequencing (ChIP-seq) profiling identified cMyc, specifically its upstream enhancer element, as a direct target of activated Stat6. Metabolomic analysis demonstrated that IL-4/13STAT6cMyc upregulation drives increased glycolysis along with upregulated expression of key glycolysis enzymes, forging a link between Kras*-driven glycolysis via paracrine activation of cMyc. Citation Format: Prasenjit Dey, Jun Li, Jianhua Zhang, Huamin Wang, Wen-Ting Liao, Vincent P Bernard, Giannicola Genovese, Anirban Maitra, Anders Strom, Ronald A. DePinho. CD4 T cells mediated protumorigenic pathway in pancreatic cancer [abstract]. In: Proceedings of the AACR Special Conference on Pancreatic Cancer: Advances in Science and Clinical Care; 2019 Sept 6-9; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2019;79(24 Suppl):Abstract nr A09.