Interferon-induced transmembrane protein 2 promotes epithelial-mesenchymal transition by activating transforming growth factor-ß1/small mother against decapentaplegic 2 signaling in gastric cancer.

BACKGROUND: Gastric cancer (GC) is one of the most prevalent malignancy around the world. Primary tumor cells are enabled to invade and migrate into adjacent normal tissues to form secondary tumors. Epithelial-mesenchymal transitions (EMT) plays a pivotal role in facilitating tumor progression. Abundant evidence suggested that the transforming growth factor-ß1 (TGF-ß1) triggered the process of EMT. Nonetheless, the precise molecular mechanisms underlying EMT requires further elucidation, and there still lacks effective specific therapeutic target. In our recent research, we demonstrated that the interferon (IFN)-induced transmembrane protein 2 (IFITM2) promoted the growth and metastasis of GC. However, it remains unclear whether IFITM2 involves in TGF-ß1 mediated EMT in GC. METHODS AND RESULTS: In the present research, we investigated the functional role of IFITM2 in EMT process and TGF-ß1 signaling pathway in two GC cell lines. We noticed that silencing IFITM2 can effectively inhibit TGF-ß1 signaling mediated EMT by regulating down stream small mother against decapentaplegic (SMAD) 2/3 and transcription factors.This finding was further determined in both tumor tissues from GC patients and normal tissues adjacent to cancer. Our data demonstrated the key role of IFITM2 in TGF-ß1 signaling and EMT in GC. CONCLUSION: The findings enriched our understanding of the underlying mechanism in EMT during the progression of GC. In addition, IFITM2 would be a potential target for treating GC and other malignant tumors.

The lncRNA MACC1-AS1 promotes gastric cancer cell metabolic plasticity via AMPK/Lin28 mediated mRNA stability of MACC1.

BACKGROUND: Metabolic plasticity has been increasingly thought to be a determinant of tumor growth and metastasis. MACC1, a transcriptional regulator of MET, was recognized as an oncogene in gastric cancer (GC); however, its transcriptional or post-translational regulation was not clear. We previously reported the metabolic role of MACC1 in glycolysis to promote GC progression. MACC1-AS1 is the antisense lncRNA of MACC1, yet its function was previously unknown. METHODS: We profiled and analyzed the expression of MACC1-AS1 utilizing the TCGA database as well as in situ hybridization using 123 pairs of GC tissues and matched adjacent normal gastric mucosa tissues (ANTs). The biological role of MACC1-AS1 in cell growth and metastasis was determined by performing in vitro and in vivo functional experiments. Glycolysis and antioxidant capabilities were assayed to examine its metabolic function. Further, the specific regulatory effect of MACC1-AS1 on MACC1 was explored transcriptionally and post-transcriptionally. RESULTS: MACC1-AS1 was shown to be expressed significantly higher in GC tissues than in ANTs, which predicted poor prognosis in GC patients. MACC1-AS1 promoted GC cell proliferation and inhibited cell apoptosis under metabolic stress. Mechanistically, MACC1-AS1 stabilized MACC1 mRNA and post-transcriptionally augmented MACC1 expression. Further, MACC1-AS1 was shown to mediate metabolic plasticity through MACC1 upregulation and subsequent enhanced glycolysis and anti-oxidative capabilities, and this was suggested to be coordinated by the AMPK/Lin28 pathway. CONCLUSIONS: Elevated expression of MACC1-AS1 in gastric cancer tissues is linked to poor prognosis and promotes malignant phenotype upon cancer cells. MACC1-AS1 is elevated under metabolic stress and facilitates metabolic plasticity by promoting MACC1 expression through mRNA stabilization. Our study implicates lncRNA MACC1-AS1 as a valuable biomarker for GC diagnosis and prognosis.

ROS signaling under metabolic stress: cross-talk between AMPK and AKT pathway.

Cancer cells are frequently confronted with metabolic stress in tumor microenvironments due to their rapid growth and limited nutrient supply. Metabolic stress induces cell death through ROS-induced apoptosis. However, cancer cells can adapt to it by altering the metabolic pathways. AMPK and AKT are two primary effectors in response to metabolic stress: AMPK acts as an energy-sensing factor which rewires metabolism and maintains redox balance. AKT broadly promotes energy production in the nutrient abundance milieu, but the role of AKT under metabolic stress is in dispute. Recent studies show that AMPK and AKT display antagonistic roles under metabolic stress. Metabolic stress-induced ROS signaling lies in the hub between metabolic reprogramming and redox homeostasis. Here, we highlight the cross-talk between AMPK and AKT and their regulation on ROS production and elimination, which summarizes the mechanism of cancer cell adaptability under ROS stress and suggests potential options for cancer therapeutics.

MACC1 mediates chemotherapy sensitivity of 5-FU and cisplatin via regulating MCT1 expression in gastric cancer.

Chemotherapeutic insensitivity is a main obstacle for effective treatment of gastric cancer (GC), the underlying mechanism remains to be investigated. Metastasis-associated in colon cancer-1 (MACC1), a transcription factor highly expressed in GC, is found to be related to chemotherapy sensitivity. Monocarboxylate transporter 1 (MCT1), a plasma membrane protein co-transporting lactate and H+, mediates drug sensitivity by regulating lactate metabolism. Targeting MCT1 has recently been regarded as a promising way to treat cancers and MCT1 inhibitor has entered the clinical trial for GC treatment. However, the correlation of these two genes and their combined effects on chemotherapy sensitivity has not been clarified. In this study, we found that MACC1 and MCT1 were both highly expressed in GC and exhibited a positive correlation in clinical samples. Further, we demonstrated that MACC1 could mediate sensitivity of 5-FU and cisplatin in GC cells, and MACC1 mediated MCT1 regulation was closely related to this sensitivity. A MCT1 inhibitor AZD3965 recovered the sensitivity of 5-FU and cisplatin in GC cells which overexpressed MACC1. These results suggested that MACC1 could influence the chemotherapy sensitivity by regulating MCT1 expression, providing new ideas and strategy for GC treatment.

High baseline tumor burden-associated macrophages promote an immunosuppressive microenvironment and reduce the efficacy of immune checkpoint inhibitors through the IGFBP2-STAT3-PD-L1 pathway.

BACKGROUND: Several clinical studies have uncovered a negative correlation between baseline tumor burden and the efficacy of immune checkpoint inhibitor (ICI) treatment. This study aimed to uncover the specific mechanisms underlying the difference in sensitivity to ICI treatment between tumors with high (HTB) and low (LTB) tumor burden. METHODS: For in vivo studies, several mouse models of subcutaneous tumors were established, and transcriptome sequencing, immunohistochemistry, and flow cytometry assays were used to detect the immune status in these subcutaneous tumors. For in vitro experiments, co-culture models, cytokine antibody arrays, western blotting, flow cytometry, and enzyme-linked immunosorbent assays were used to explore the underlying molecular mechanisms RESULTS: We found that MC38 or B16 subcutaneous tumors from the HTB group did not show any response to anti-programmed cell death protein-1 (PD-1) therapy. Through flow cytometry assays, we found that the infiltration with CD8+ T cells was significantly decreased whereas M2-like macrophages were enriched in subcutaneous tumors of HTB groups compared with those of LTB group. These changes were not affected by the initial number of injected tumor cells or tumor age, nor could they be reversed by surgical tumor reduction. Intraperitoneal colony-stimulating factor 1 receptor (CSF-1R) inhibitor PLX3397 injection at different time points of tumor growth only had an effect when administered in the early tumor stage to maintain the "heat" of the tumor microenvironment during the process of tumor growth, thereby achieving a response to ICI treatment when the tumor grew to a large size. Mechanistically, we found that insulin-like growth factor binding protein 2 (IGFBP2) expression levels were significantly elevated in HTB tumor tissues. IGFBP2 promoted the programmed death-ligand 1 (PD-L1) expression in M2-like macrophages by activating signal transducer and activator of transcription 3 (STAT3), and PD-L1+ M2-like macrophages exerted an immunosuppressive effect by inhibiting the proliferation and activation of CD8+ T cells in a PD-L1-dependent fashion. CONCLUSIONS: This study suggested that the low efficacy of ICI treatment in HTB tumors is mainly attributed to the intratumoral accumulation of PD-L1+ M2-like macrophages via the IGFBP2-STAT3-PD-L1 signaling pathway and their substantial inhibitory effects on T cell proliferation and activation.

Matrix Stiffness Triggers Lipid Metabolic Cross-talk between Tumor and Stromal Cells to Mediate Bevacizumab Resistance in Colorectal Cancer Liver Metastases.

Bevacizumab is an anti-VEGF monoclonal antibody that plays an important role in the combination treatment of advanced colorectal cancer. However, resistance remains a major hurdle limiting bevacizumab efficacy, highlighting the importance of identifying a mechanism of antiangiogenic therapy resistance. Here, we investigated biophysical properties of the extracellular matrix (ECM) related to metabolic processes and acquired resistance to bevacizumab. Evaluation of paired pre- and posttreatment samples of liver metastases from 20 colorectal cancer patients treated with combination bevacizumab therapy, including 10 responders and 10 nonresponders, indicated that ECM deposition in liver metastases and a highly activated fatty acid oxidation (FAO) pathway were elevated in nonresponders after antiangiogenic therapy compared with responders. In mouse models of liver metastatic colorectal cancer (mCRC), anti-VEGF increased ECM deposition and FAO in colorectal cancer cells, and treatment with the FAO inhibitor etomoxir enhanced the efficacy of antiangiogenic therapy. Hepatic stellate cells (HSC) were essential for matrix stiffness-mediated FAO in colon cancer cells. Matrix stiffness activated lipolysis in HSCs via the focal adhesion kinase (FAK)/yes-associated protein (YAP) pathway, and free fatty acids secreted by HSCs were absorbed as metabolic substrates and activated FAO in colon cancer cells. Suppressing HSC lipolysis using FAK and YAP inhibition enhanced the efficacy of anti-VEGF therapy. Together, these results indicate that bevacizumab-induced ECM remodeling triggers lipid metabolic cross-talk between colon cancer cells and HSCs. This metabolic mechanism of bevacizumab resistance mediated by the physical tumor microenvironment represents a potential therapeutic target for reversing drug resistance. SIGNIFICANCE: Extracellular matrix stiffening drives bevacizumab resistance by stimulating hepatic stellate cells to provide fuel for mCRC cells in the liver, indicating a potential metabolism-based therapeutic strategy for overcoming resistance.

Evaluation of stromal cell infiltration in the tumor microenvironment enable prediction of treatment sensitivity and prognosis in colon cancer.

Current clinical factors for screening candidates that might benefit from adjuvant chemotherapy in colon cancer are inadequate. Tumor microenvironment, especially the stromal components, has the potential to determine treatment response. However, clinical translation of the tumor-associated stromal characterization into a practical biomarker for helping treatment decision has not been established. Using machine learning, we established a novel 31-gene signature, called stromal cell infiltration intensity score (SIIS), to distinguish patients characterized by the enrichment of abundant stromal cells in five colon cancer datasets from GEO (N = 990). Patients with high-SIIS were at higher risk for recurrence and mortality, and could not benefit from adjuvant chemotherapy due to their intrinsic drug resistance; however, the opposite was reported for patients with low-SIIS. The role of SIIS in detection of patients with high stromal cell infiltration and reduced drug efficiency was consistently validated in the TCGA-COAD cohort (N = 382), Sun Yat-sen University Cancer Center cohort (N = 30), and could also be observed in TCGA pan-cancer settings (N = 4898) and four independent immunotherapy cohorts (N = 467). Based on multi-omics data analysis and the CRISPR library screen, we reported that lack of gene mutation, hypomethylation in ADCY4 promoter region, activation of WNT-PCP pathway and SIAH2-GPX3 axis were potential mechanisms responsible for the chemoresistance of patients within high-SIIS group. Our findings demonstrated that SIIS provide an important reference for those making treatment decisions for such special patients.

CRIP1 cooperates with BRCA2 to drive the nuclear enrichment of RAD51 and to facilitate homologous repair upon DNA damage induced by chemotherapy.

Homologous recombination (HR) repair is an important determinant of chemosensitivity. However, the mechanisms underlying HR regulation remain largely unknown. Cysteine-rich intestinal protein 1 (CRIP1) is a member of the LIM/double-zinc finger protein family and is overexpressed and associated with prognosis in several tumor types. However, to date, the functional role of CRIP1 in cancer biology is poorly understood. Here we found that CRIP1 downregulation causes HR repair deficiency with concomitant increase in cell sensitivity to cisplatin, epirubicin, and the poly ADP-ribose polymerase (PARP) inhibitor olaparib in gastric cancer cells. Mechanistically, upon DNA damage, CRIP1 is deubiquitinated and upregulated by activated AKT signaling. CRIP1, in turn, promotes nuclear enrichment of RAD51, which is a prerequisite step for HR commencement, by stabilizing BRCA2 to counteract FBXO5-targeted RAD51 degradation and by binding to the core domain of RAD51 (RAD51184-257) in coordination with BRCA2, to facilitate nuclear export signal masking interactions between BRCA2 and RAD51. Moreover, through mass spectrometry screening, we found that KPNA4 is at least one of the carriers controlling the nucleo-cytoplasmic distribution of the CRIP1-BRCA2-RAD51 complex in response to chemotherapy. Consistent with these findings, RAD51 inhibitors block the CRIP1-mediated HR process, thereby restoring chemotherapy sensitivity of gastric cancer cells with high CRIP1 expression. Analysis of patient specimens revealed an abnormally high level of CRIP1 expression in GC tissues compared to that in the adjacent normal mucosa and a significant negative association between CRIP1 expression and survival time in patient cohorts with different types of solid tumors undergoing genotoxic treatments. In conclusion, our study suggests an essential function of CRIP1 in promoting HR repair and facilitating gastric cancer cell adaptation to genotoxic therapy.

Cysteine Dioxygenase 1 Mediates Erastin-Induced Ferroptosis in Human Gastric Cancer Cells.

BACKGROUND: Ferroptosis is a recently discovered form of iron-dependent nonapoptotic cell death. It is characterized by loss of the activity of the lipid repair enzyme, glutathione peroxidase 4 (GPX4), and accumulation of lethal reactive lipid oxygen species. However, we still know relatively little about ferroptosis and its molecular mechanism in gastric cancer (GC) cells. Here, we demonstrate that erastin, a classic inducer of ferroptosis, induces this form of cell death in GC cells and that cysteine dioxygenase 1 (CDO1) plays an important role in this process. METHODS: We performed quantitative real-time polymerase chain reaction, Western blotting, cell viability assay, reactive oxygen species (ROS) assay, glutathione assay, lipid peroxidation assay, RNAi and gene transfection, immunofluorescent staining, dual-luciferase reporter assay, transmission electron microscopy, and chromatin immunoprecipitation assay to study the regulation of ferroptosis in GC cells. Mouse xenograft assay was used to figure out the mechanism in vivo. RESULTS: Silencing CDO1 inhibited erastin-induced ferroptosis in GC cells both in vitro and in vivo. Suppression of CDO1 restored cellular GSH levels, prevented ROS generation, and reduced malondialdehyde, one of the end products of lipid peroxidation. In addition, silencing COO1 maintained mitochondrial morphologic stability in erastin-treated cells. Mechanistically, c-Myb transcriptionally regulated CDO1, and inhibition of CDO1 expression upregulated GPX4 expression. CONCLUSIONS: Our findings give a better understanding of ferroptosis and its molecular mechanism in GC cells, gaining insight into ferroptosis-mediated cancer treatment.