Downregulation of lncRNA MIR17HG reduced tumorigenicity and Treg-mediated immune escape of non-small-cell lung cancer cells through targeting the miR-17-5p/RUNX3 axis.

Long noncoding RNA MIR17HG was involved with the progression of non-small-cell lung cancer (NSCLC), but specific mechanisms of MIR17HG-mediated immune escape of NSCLC cells were still unknown. The present study investigated the function of MIR17HG on regulatory T cell (Treg)-mediated immune escape and the underlying mechanisms in NSCLC. Expression of MIR17HG and miR-17-5p in NSCLC tissue samples were detected using quantitative real-time PCR (qRT-PCR). A549 and H1299 cells were transfected with sh-MIR17HG, miR-17-5p inhibitor, or sh-MIR17HG + miR-17-5p inhibitor, followed by cocultured with Tregs. Cell proliferation was measured using 5-ethynyl-20-deoxyuridine (Edu) staining assay and cell counting kit-8 (CCK-8) assay. Flow cytometry was used for determining positive numbers of FOXP3+CD4+/CD25+/CD8+ Tregs. Through subcutaneous injection with transfected A549 cells, a xenograft nude mouse model was established. Weights and volumes of xenograft tumors were evaluated. Additionally, the expressions of immune-related factors including transforming growth factor beta (TGF-ß), vascular endothelial growth factor A (VEGF-A), interleukin-10 (IL-10), IL-4, and interferon-gamma (IFN-γ) in cultured cells, were evaluated by enzyme-linked immunosorbent assay and western blot analysis. Then, miR-17-5p was decreased and MIR17HG was enhanced in both NSCLC tissues and cell lines. MIR17HG knockdown significantly suppressed cell proliferation, tumorigenicity, and immune capacity of Tregs in A549 and H1299 cells, whereas sh-MIR17HG significantly reduced expression levels of VEGF-A, TGF-ß, IL-4, and IL-10 but promoted the IFN-γ level in vitro and in vivo. Moreover, downregulation of miR-17-5p significantly reversed the effects of sh-MIR17HG. Additionally, we identified that runt- related transcription factor 3 (RUNX3) was a target of miR-17-5p, and sh-MIR17HG and miR-17-5p mimics downregulated RUNX3 expression. In conclusion, downregulation of MIR17HG suppresses tumorigenicity and Treg-mediated immune escape in NSCLC through downregulating the miR-17-5p/RUNX3 axis, indicating that this axis contains potential biomarkers for NSCLC.

Runx3 Regulates CD8+ T Cell Local Expansion and CD43 Glycosylation in Mice by H1N1 Influenza A Virus Infection.

We have recently reported that transcription factor Runx3 is required for pulmonary generation of CD8+ cytotoxic T lymphocytes (CTLs) that play a crucial role in the clearance of influenza A virus (IAV). To understand the underlying mechanisms, we determined the effects of Runx3 knockout (KO) on CD8+ T cell local expansion and phenotypes using an inducible general Runx3 KO mouse model. We found that in contrast to the lungs, Runx3 general KO promoted enlargement of lung-draining mediastinal lymph node (mLN) and enhanced CD8+ and CD4+ T cell expansion during H1N1 IAV infection. We further found that Runx3 deficiency greatly inhibited core 2 O-glycosylation of selectin ligand CD43 on activated CD8+ T cells but minimally affected the cell surface expression of CD43, activation markers (CD44 and CD69) and cell adhesion molecules (CD11a and CD54). Runx3 KO had a minor effect on lung effector CD8+ T cell death by IAV infection. Our findings indicate that Runx3 differently regulates CD8+ T cell expansion in mLNs and lungs by H1N1 IAV infection. Runx3 is required for CD43 core 2 O-glycosylation on activated CD8+ T cells, and the involved Runx3 signal pathway may mediate CD8+ T cell phenotype for pulmonary generation of CTLs.

Runx3 Restoration Regresses K-Ras-Activated Mouse Lung Cancers and Inhibits Recurrence.

Oncogenic K-RAS mutations occur in approximately 25% of human lung cancers and are most frequently found in codon 12 (G12C, G12V, and G12D). Mutated K-RAS inhibitors have shown beneficial results in many patients; however, the inhibitors specifically target K-RASG12C and acquired resistance is a common occurrence. Therefore, new treatments targeting all kinds of oncogenic K-RAS mutations with a durable response are needed. RUNX3 acts as a pioneer factor of the restriction (R)-point, which is critical for the life and death of cells. RUNX3 is inactivated in most K-RAS-activated mouse and human lung cancers. Deletion of mouse lung Runx3 induces adenomas (ADs) and facilitates the development of K-Ras-activated adenocarcinomas (ADCs). In this study, conditional restoration of Runx3 in an established K-Ras-activated mouse lung cancer model regressed both ADs and ADCs and suppressed cancer recurrence, markedly increasing mouse survival. Runx3 restoration suppressed K-Ras-activated lung cancer mainly through Arf-p53 pathway-mediated apoptosis and partly through p53-independent inhibition of proliferation. This study provides in vivo evidence supporting RUNX3 as a therapeutic tool for the treatment of K-RAS-activated lung cancers with a durable response.

RUNX3 inactivates oncogenic MYC through disruption of MYC/MAX complex and subsequent recruitment of GSK3ß-FBXW7 cascade.

MYC is one of the most commonly dysregulated proto-oncogenes in cancer. MYC promotes cancer initiation and maintenance by regulating multiple biological processes, such as proliferation and stem cell function. Here, we show that developmental regulator RUNX3 targets MYC protein for rapid degradation through the glycogen synthase kinase-3 beta-F-box/WD repeat-containing protein 7 (GSK3ß-FBXW7) proteolytic pathway. The evolutionarily conserved Runt domain of RUNX3 interacts directly with the basic helix-loop-helix leucine zipper of MYC, resulting in the disruption of MYC/MAX and MYC/MIZ-1 interactions, enhanced GSK3ß-mediated phosphorylation of MYC protein at threonine-58 and its subsequent degradation via the ubiquitin-proteasomal pathway. We therefore uncover a previously unknown mode of MYC destabilization by RUNX3 and provide an explanation as to why RUNX3 inhibits early-stage cancer development in gastrointestinal and lung mouse cancer models.

C/ebpα represses the oncogenic Runx3-Myc axis in p53-deficient osteosarcoma development.

Osteosarcoma (OS) is characterized by TP53 mutations in humans. In mice, loss of p53 triggers OS development, and osteoprogenitor-specific p53-deleted mice are widely used to study the process of osteosarcomagenesis. However, the molecular mechanisms underlying the initiation or progression of OS following or parallel to p53 inactivation remain largely unknown. Here, we examined the role of transcription factors involved in adipogenesis (adipo-TFs) in p53-deficient OS and identified a novel tumor suppressive molecular mechanism mediated by C/ebpα. C/ebpα specifically interacts with Runx3, a p53 deficiency-dependent oncogene, and, in the same manner as p53, decreases the activity of the oncogenic axis of OS, Runx3-Myc, by inhibiting Runx3 DNA binding. The identification of a novel molecular role for C/ebpα in p53-deficient osteosarcomagenesis underscores the importance of the Runx-Myc oncogenic axis as a therapeutic target for OS.

Clinical significance of determining the hypermethylation of the RUNX3 gene promoter and its cohypermethylation with the BRCA1 gene for patients with breast cancer.

PURPOSE: The aim of this study was to assess the clinical significance of RUNX3 gene hypermethylation in the pathogenetic mechanisms of breast cancer in women, taking into account its cohypermethylation with the BRCA1 gene. METHODS: This study included 74 women with newly diagnosed breast cancer (samples from female primary breast carcinomas and paired peripheral blood samples) and 62 women without oncological pathology-control group (peripheral blood samples). Epigenetic testing for hypermethylation status studying was performed in all samples on freshly collected material with the addition of a preservative before the storage and DNA isolation. RESULTS: Hypermethylation of the RUNX3 gene promoter region was detected in 71.6% samples of breast cancer tissue and in 35.13% samples of blood. The RUNX3 gene promoter region hypermethylation was significantly higher among breast cancer patients compared to the control group. The frequency of cohypermethylation in RUNX3 and BRCA1 genes was significantly increased in breast cancer tissues compared to the blood of patients. CONCLUSION: A significantly increased frequency of the hypermethylation of the RUNX3 gene promoter region and its cohypermethylation with the BRCA1 gene promoter region was found in tumor tissue and blood samples from patients with breast cancer, in contrast to the control group. The identified differences indicate the importance of further investigations of suppressor genes cohypermethylation in patients with breast cancer. Further large-scale studies are needed to find out whether the detected hypermethylation and cohypermethylation of the RUNX3 gene promoter region will have an impact on the treatment strategy in patients.

The RUNX Family of Proteins, DNA Repair, and Cancer.

The RUNX family of transcription factors, including RUNX1, RUNX2, and RUNX3, are key regulators of development and can function as either tumor suppressors or oncogenes in cancer. Emerging evidence suggests that the dysregulation of RUNX genes can promote genomic instability in both leukemia and solid cancers by impairing DNA repair mechanisms. RUNX proteins control the cellular response to DNA damage by regulating the p53, Fanconi anemia, and oxidative stress repair pathways through transcriptional or non-transcriptional mechanisms. This review highlights the importance of RUNX-dependent DNA repair regulation in human cancers.

p53 Deficiency-Dependent Oncogenicity of Runx3.

The RUNX transcription factors are frequently dysregulated in human cancers, suggesting their potential as attractive targets for drug treatment. However, all three transcription factors have been described as both tumor suppressors and oncogenes, indicating the need to determine their molecular mechanisms of action. Although RUNX3 has long been considered a tumor suppressor in human cancers, several recent studies have shown that RUNX3 is upregulated during the development or progression of various malignant tumors, suggesting it may act as a "conditional" oncogene. Resolving this paradox and understanding how a single gene can exhibit both oncogenic and tumor-suppressive properties is essential for successful drug targeting of RUNX. This review describes the evidence for the activities of RUNX3 in human cancer and proposes an explanation for the duality of RUNX3 involving the status of p53. In this model, p53 deficiency causes RUNX3 to become oncogenic, leading to aberrant upregulation of MYC.

Prox2 and Runx3 vagal sensory neurons regulate esophageal motility.

Vagal sensory neurons monitor mechanical and chemical stimuli in the gastrointestinal tract. Major efforts are underway to assign physiological functions to the many distinct subtypes of vagal sensory neurons. Here, we use genetically guided anatomical tracing, optogenetics, and electrophysiology to identify and characterize vagal sensory neuron subtypes expressing Prox2 and Runx3 in mice. We show that three of these neuronal subtypes innervate the esophagus and stomach in regionalized patterns, where they form intraganglionic laminar endings. Electrophysiological analysis revealed that they are low-threshold mechanoreceptors but possess different adaptation properties. Lastly, genetic ablation of Prox2 and Runx3 neurons demonstrated their essential roles for esophageal peristalsis in freely behaving mice. Our work defines the identity and function of the vagal neurons that provide mechanosensory feedback from the esophagus to the brain and could lead to better understanding and treatment of esophageal motility disorders.

MicroRNA-17 Family Targets RUNX3 to Increase Proliferation and Migration of Hepatocellular Carcinoma.

Hepatocellular carcinoma (HCC) is one common cancer in the world. Previous studies have shown that miR-17 family members are elevated in most tumors and promote tumor progression. However, there is no comprehensive analysis of the expression and functional mechanism of the microRNA-17 (miR-17) family in HCC. The aim of this study is to comprehensively analyze the function of the miR-17 family in HCC and the molecular mechanism of its role. Bioinfoimatics analysis of the miR-17 family expression profile and its relationship to clinical significance using The Cancer Genome Atlas (TCGA) database, and this result was confirmed using quantitative real-time polymerase chain reaction. miR-17 family members were tested for functional effects through transfection of miRNA precursors and inhibitors, and monitoring cell viability and migration by cell count and wound healing assays. In addition, we using dual-luciferase assay and Western blot demonstrated the targeting relationship between the miRNA-17 family and RUNX3. These members of miR-17 family were highly expressed in HCC tissues, and the overexpression of the miR-17 family promoted the proliferation and migration of SMMC-7721 cells, whereas treatment with anti-miR17 inhibitors caused the opposite effects. Notably, we also found that inhibitors anti-each member of miR-17 can suppress the expression of the entire family member. In addition, they can bind to the 3' untranslated region of RUNX3 to regulate its expression at the translational level. Our results proved that miR-17 family has oncogenic characteristics, overexpression every member of the family contributed to HCC cell proliferation and migration by reducing the translation of RUNX3.

Role of RUNX3 in Restriction Point Regulation.

A cell cycle is a series of events that takes place in a cell as it grows and divides. At the G1 phase of cell cycle, cells monitor their cumulative exposure to specific signals and make the critical decision to pass through the restriction (R)-point. The R-point decision-making machinery is fundamental to normal differentiation, apoptosis, and G1-S transition. Deregulation of this machinery is markedly associated with tumorigenesis. Therefore, identification of the molecular mechanisms that govern the R-point decision is one of the fundamental issues in tumor biology. RUNX3 is one of the genes frequently inactivated in tumors by epigenetic alterations. In particular, RUNX3 is downregulated in most K-RAS-activated human and mouse lung adenocarcinomas (ADCs). Targeted inactivation of Runx3 in the mouse lung induces adenomas (ADs), and markedly shortens the latency of ADC formation induced by oncogenic K-Ras. RUNX3 participates in the transient formation of R-point-associated activator (RPA-RX3-AC) complexes, which measure the duration of RAS signals and thereby protect cells against oncogenic RAS. This review focuses on the molecular mechanism by which the R-point participates in oncogenic surveillance.

RUNX3 Meets the Ubiquitin-Proteasome System in Cancer.

RUNX3 is a transcription factor with regulatory roles in cell proliferation and development. While largely characterized as a tumor suppressor, RUNX3 can also be oncogenic in certain cancers. Many factors account for the tumor suppressor function of RUNX3, which is reflected by its ability to suppress cancer cell proliferation after expression-restoration, and its inactivation in cancer cells. Ubiquitination and proteasomal degradation represent a major mechanism for the inactivation of RUNX3 and the suppression of cancer cell proliferation. On the one hand, RUNX3 has been shown to facilitate the ubiquitination and proteasomal degradation of oncogenic proteins. On the other hand, RUNX3 can be inactivated through the ubiquitin-proteasome system. This review encapsulates two facets of RUNX3 in cancer: how RUNX3 suppresses cell proliferation by facilitating the ubiquitination and proteasomal degradation of oncogenic proteins, and how RUNX3 is degraded itself through interacting RNA-, protein-, and pathogen-mediated ubiquitination and proteasomal degradation.

RUNX3 improves CAR-T cell phenotype and reduces cytokine release while maintaining CAR-T function.

CAR-T therapy has shown successful in the treatment of certain types of hematological malignancy, while the efficacy of CAR-T cell in treating solid tumors has been limited due to the exhaustion of CAR-T caused by the tumor microenvironment in solid tumors. Therefore, improving the exhaustion of CAR-T cell is one of the inspiring strategies for CAR-T treatment of solid tumors. As an important regulator in T cell immunity, the transcription factor RUNX3 not only negatively regulates the terminal differentiation T-bet gene, reducing the ultimate differentiation of T cells, but also increases the residency of T cells in non-lymphoid tissues and tumors. By overexpressing RUNX3 in CAR-T cells, we found that increasing the expression of RUNX3 maintained the low differentiation of CAR-T cells, further improving the exhaustion of CAR-T cells during antigen stimulation. In vitro, we found that RUNX3 could reduce the release of cytokines while maintaining CAR-T cells function. In re-challenge experiments, CAR-T cells overexpressing RUNX3 (Runx3-OE CAR-T) were safer than conventional CAR-T cells, while RUNX3 could also maintain the anti-tumor efficacy of CAR-T cells in vivo. Collectively, we found that Runx3-OE CAR-T cells can improve CAR-T phenotype and reduce cytokines release while maintaining CAR-T cells function, which may improve the safety of CAR-T therapy in clinical trials.

Genetically encoded Runx3 and CD4+ intestinal epithelial lymphocyte deficiencies link SKG mouse and human predisposition to spondyloarthropathy.

Disturbances in immune regulation, intestinal dysbiosis and inflammation characterize ankylosing spondylitis (AS), which is associated with RUNX3 loss-of-function variants. ZAP70W163C mutant (SKG) mice have reduced ZAP70 signaling, spondyloarthritis and ileitis. In small intestine, Foxp3+ regulatory T cells (Treg) and CD4+CD8αα+TCRαß+ intraepithelial lymphocytes (CD4-IEL) control inflammation. TGF-ß and retinoic acid (RA)-producing dendritic cells and MHC-class II+ intestinal epithelial cells (IEC) are required for Treg and CD4-IEL differentiation from CD4+ conventional or Treg precursors, with upregulation of Runx3 and suppression of ThPOK. We show in SKG mouse ileum, that ZAP70W163C or ZAP70 inhibition prevented CD4-IEL but not Treg differentiation, dysregulating Runx3 and ThPOK. TGF-ß/RA-mediated CD4-IEL development, T-cell IFN-γ production, MHC class-II+ IEC, tissue-resident memory T-cell and Runx3-regulated genes were reduced. In AS intestine, CD4-IEL were decreased, while in AS blood CD4+CD8+ T cells were reduced and Treg increased. Thus, genetically-encoded TCR signaling dysfunction links intestinal T-cell immunodeficiency in mouse and human spondyloarthropathy.

Inducible general knockout of Runx3 profoundly reduces pulmonary cytotoxic CD8+ T cells with minimal effect on outcomes in mice following influenza infection.

Respiratory viruses pose a continuing and substantive threat to human health globally. Host innate and adaptive immune responses are the critical antiviral defense mechanisms to control viral replication and spread. The present study is designed to determine the role of transcription factor Runx3 in the host immune response to influenza A virus (IAV) infection. As Runx3 is required for embryonic development, we generated an inducible Runx3 global knockout (KO) mouse model and found that Runx3 KO in adult C57BL/6 mice minimally affected thymic function under normal conditions and survival was at least 250 days post Runx3 deletion. We applied the mouse model to IAV infection and found that Runx3 KO resulted in a huge reduction (>85%) in numbers of total and antigen-specific pulmonary CD8+ cytotoxic T cells during IAV infection, while it had a minor effect on pulmonary generation of CD4+ T cells. To our surprise, this general KO of Runx3 did not significantly alter viral clearance and animal survival following IAV infection. Interestingly, we found that Runx3 KO significantly increased the numbers of pulmonary innate immune cells such as macrophages and neutrophils and the production of pro-inflammatory cytokines during IAV infection. We further found that Runx3 was strongly detected in CCR2+ immune cells in IAV-infected mouse lungs and was induced in activated macrophages and dendritic cells (DCs). As pulmonary CD8+ cytotoxic T cells play a central role in the clearance of IAV, our findings suggest that Runx3 KO may enhance host innate immunity to compensate for the loss of pulmonary CD8+ cytotoxic T cells during IAV infection.

Methylation of RUNX3 Promoter 2 in the Whole Blood of Children with Ulcerative Colitis.

Ulcerative colitis (UC) results from a complex interplay between the environment, gut microbiota, host genetics, and immunity. Runt-related transcription factor 3 (RUNX3) regulates Th1/Th2 balance and, thus, the synthesis of cytokines and inflammation. We aimed to analyze the dependence of RUNX3 promoter 2 (P2) methylation level on: age, sex, body mass index (BMI), C-reactive protein (CRP), serum albumin, disease duration, Pediatric Ulcerative Colitis Activity Index (PUCAI), the Paris classification, and exposure to medications. This multicenter, cross-sectional study recruited hospitalized children with UC. Methylation of RUNX3 P2 was measured with methylation-sensitive restriction enzymes in the whole blood DNA. Sixty-four children were enrolled, with a mean age of 14.5 ± 2.8 years. Half of them were female (51.6%), and the average BMI Z-score was -0.44 ± 1.14. The mean methylation of RUNX3 P2 was 54.1 ± 13.3%. The methylation level of RUNX3 P2 did not correlate with age, sex, nutritional status, CRP, albumin, PUCAI, or the extent of colitis (Paris E1-E4). RUNX3 P2 methylation did not differ between patients recruited within two and a half months of diagnosis and children who had UC for at least a year. Current or past exposure to biologics, immunosuppressants, or steroids was not associated with RUNX3 P2 methylation. Methylation of RUNX3 promoter 2 in whole blood DNA does not seem to be associated with the characteristics of UC in children.

RUNX3-Regulated GALNT6 Promotes the Migration and Invasion of Hepatocellular Carcinoma Cells by Mediating O-Glycosylation of MUC1.

Background: Hepatocellular carcinoma (HCC) is a leading cause of cancer-related death worldwide. Dysregulation of messenger RNAs (mRNA) has been recognized to be associated with HCC carcinogenesis and development. Polypeptide GalNAc Transferase 6 (GALNT6), an O-type glycosyltransferase, has been confirmed as tumor promoter in different cancers. However, the function of GALNT6 in HCC remains to be studied. Methods: RT-qPCR and western blot experiments were, respectively, performed for evaluating RNA expressions and protein levels. Supported by bioinformatics analysis, mechanism assays were conducted for validating the potential relation between different genes. Functional assays were implemented to analyze HCC cell migration and invasion after different transfections. Results: GALNT6 was aberrantly upregulated in HCC cells. Knockdown of GALNT6 could repress HCC cell migration and invasion. RUNX3 was verified to bind to GALNT6 promoter and activate GALNT6 transcription. GALNT6 depletion led to inhibited O-glycosylation and aggravated degradation of MUC1. MUC1 overexpression could rescue the impeded HCC cell migration and invasion induced by GALNT6 knockdown. Conclusion: To sum up, GALNT6 transcriptionally activated by RUNX3 mediated the O-glycosylation of MUC1, thus exerting promoting influence on HCC cell migration and invasion.

RUNX3-regulated circRNA METTL3 inhibits colorectal cancer proliferation and metastasis via miR-107/PER3 axis.

Colorectal cancer (CRC) is one of the most prevalent and lethal malignancies. Exploring the underlying molecular mechanisms is very helpful for the development of new therapy. Here, we investigated the function of circMETTL3/miR-107/PER3 in CRC. Human CRC tissues from diagnosed CRC patients and six CRC cell lines, one normal human colon cell line were used. qRT-PCR and western blotting were performed to determine expression levels of RUNX3, circMETTL3, miR-107, PER3, and proliferation-, and migration-related proteins. CCK-8, colony formation assay, transwell assay, and scratch wound assay were utilized to assess CRC cell proliferation and invasion. ChIP, EMSA, biotin-pull down, RIP assay, and dual luciferase reporter assay were performed to validate interactions of RUNX3/METTL3 promoter, circMETTL3/miR-107, and miR-107/PER3. FISH was used to characterize circMETTL3. MSP was employed to measure methylation level. Nude mouse xenograft model was used to determine the effects on tumor growth and metastasis in vivo. RUNX3, circMETTL3, and PER3 were diminished while miR-107 was elevated in CRC tissues and cells. Low levels of RUNX3 and circMETTL3 correlated with poor prognosis of CRC. Overexpression of RUNX3, circMETTL3, or PER3 suppressed while miR-107 mimics promoted, CRC cell proliferation and invasion, as well as tumor growth and metastasis in vivo. Mechanistically, RUNX3 bound to METTL3 promoter and activated circMETTL3 transcription. circMETTL3 directly bound with miR-107 which targeted PER3. circMETTL3/miR-107 regulated CRC cell proliferation and invasion via PER3. CircMETTL3, transcriptionally activated by RUNX3, restrains CRC development and metastasis via acting as a miR-107 sponge to regulate PER3 signaling.

[RUNX3 regulates trastuzumab resistance of gastric cancer cells: a metabolomic analysis based on UPLC-Q Exactive Focus Orbitrap mass spectrometry].

OBJECTIVE: To explore the role of Runt-related transcription factor 3 (RUNX3) in metabolic regulation of trastuzumab-resistant gastric cancer cells and investigate the mechanism of RUNX3 knockdown-mediated reversal of trastuzumab resistance. METHODS: We performed a metabolomic analysis of trastuzumab-resistant gastric cancer cells (NCI N87R) and RUNX3 knockdown cells (NCI N87R/RUNX3) using ultra performance liquid chromatography (UPLC) coupled with Q Exactive Focus Orbitrap mass spectrometry (MS). Multivariate combined with univariate analyses and MS/MS ion spectrums were used to screen the differential variables. MetaboAnalyst 5.0 database was employed for pathway enrichment analysis. Differential metabolites-genes regulatory relationships were constructed based on OmicsNet database. The changes in GSH/GSSG and NADPH/NADP ratios in NCI N87R/RUNX3 cells were measured using detection kits. RESULTS: The metabolic profile of NCI N87R cells was significantly altered after RUNX3 knockdown, with 81 differential metabolites identified to contribute significantly to the classification, among which 43 metabolites were increased and 38 were decreased (P < 0.01). In NCI N87R cells, RUNX3 knockdown resulted in noticeable alterations in 8 pathways involving glutamine metabolism, glycolysis, glycerophospholipid, nicotinate-nicotinamide and glutathione metabolism, causing also significant reduction of intracellular GSH/GSSG and NADPH/NADP ratios (P < 0.01). The differential metabolites-genes network revealed a regulatory relationship between the metabolic molecules and genes. CONCLUSION: RUNX3 reverses trastuzumab resistance in gastric cancer cells by regulating energy metabolism and oxidation-reduction homeostasis and may serve as a potential therapeutic target for trastuzumab-resistant gastric cancer.

Increased expression of RUNX3 inhibits normal human myeloid development.

RUNX3 is a transcription factor dysregulated in acute myeloid leukemia (AML). However, its role in normal myeloid development and leukemia is poorly understood. Here we investigate RUNX3 expression in both settings and the impact of its dysregulation on myelopoiesis. We found that RUNX3 mRNA expression was stable during hematopoiesis but decreased with granulocytic differentiation. In AML, RUNX3 mRNA was overexpressed in many disease subtypes, but downregulated in AML with core binding factor abnormalities, such as RUNX1::ETO. Overexpression of RUNX3 in human hematopoietic stem and progenitor cells (HSPC) inhibited myeloid differentiation, particularly of the granulocytic lineage. Proliferation and myeloid colony formation were also inhibited. Conversely, RUNX3 knockdown did not impact the myeloid growth and development of human HSPC. Overexpression of RUNX3 in the context of RUNX1::ETO did not rescue the RUNX1::ETO-mediated block in differentiation. RNA-sequencing showed that RUNX3 overexpression downregulates key developmental genes, such as KIT and RUNX1, while upregulating lymphoid genes, such as KLRB1 and TBX21. Overall, these data show that increased RUNX3 expression observed in AML could contribute to the developmental arrest characteristic of this disease, possibly by driving a competing transcriptional program favoring a lymphoid fate.