Inhibition of Glutamine Utilization Synergizes with Immune Checkpoint Inhibitor to Promote Antitumor Immunity.

Despite its outstanding clinical success, immune checkpoint blockade remains ineffective in many patients. Accordingly, combination therapy capable of achieving greater antitumor immunity is urgently required. Here, we report that limiting glutamine metabolism in cancer cells bolsters the effectiveness of anti-programmed death ligand-1 (PD-L1) antibody. Inhibition of glutamine utilization increased PD-L1 levels in cancer cells, thereby inactivating co-cultured T cells. Under glutamine-limited conditions, reduced cellular GSH levels caused an upregulation of PD-L1 expression by impairing SERCA activity, which activates the calcium/NF-κB signaling cascade. Consequently, in tumors grown in immunocompetent mice, inhibition of glutamine metabolism decreased the antitumor activity of T cells. In combination with anti-PD-L1, however, glutamine depletion strongly promoted the antitumor efficacy of T cells in vitro and in vivo due to simultaneous increases in Fas/CD95 levels. Our results demonstrate the relevance of cancer glutamine metabolism to antitumor immunity and suggest that co-targeting of glutamine metabolism and PD-L1 represents a promising therapeutic approach.

Unveiling Prognostic RNA Biomarkers through a Multi-Cohort Study in Colorectal Cancer.

Because cancer is a leading cause of death and is thought to be caused by genetic errors or genomic instability in many circumstances, there have been studies exploring cancer's genetic basis using microarray and RNA-seq methods, linking gene expression data to patient survival. This research introduces a methodological framework, combining heterogeneous gene expression data, random forest selection, and pathway analysis, alongside clinical information and Cox regression analysis, to discover prognostic biomarkers. Heterogeneous gene expression data for colorectal cancer were collected from TCGA-COAD (RNA-seq), and GSE17536 and GSE39582 (microarray), and were integrated with Entrez Gene IDs. Using Cox regression analysis and random forest, genes with consistent hazard ratios and significantly affecting patient survivability were chosen. Predictive accuracy was evaluated using ROC curves. Pathway analysis identified potential RNA biomarkers. The authors identified 28 RNA biomarkers. Pathway analysis revealed enrichment in cancer-related pathways, notably EGFR downstream signaling and IGF1R signaling. Three RNA biomarkers (ZEB1-AS1, PI4K2A, and ITGB8-AS1) and two clinical biomarkers (stage and age) were chosen for a prognostic model, improving predictive performance compared to using clinical biomarkers alone. Despite biomarker identification challenges, this study underscores integration of heterogenous gene expression data for discovery.

Oncogenic KRAS signaling activates mTORC1 through COUP-TFII-mediated lactate production.

Oncogenic signals contribute to enhanced glycolysis and mTORC1 activity, leading to rapid cell proliferation in cancer. Regulation of glycolysis and mTORC1 by PI3K/Akt signaling is well established, but how KRAS-induced MEK signaling regulates these pathways remains poorly understood. Here, we report a role for MEK-driven lactate production in mTORC1 activation in KRAS-activated cells. KRAS/MEK-induced upregulation of the chicken ovalbumin upstream promoter transcriptional factor II (COUP-TFII) increases the expression of lactate dehydrogenase A (LDHA), resulting in lactate production and mTORC1 activation. Further, lactate inhibits the interaction of TSC2 and Rheb, leading to the cellular activation of mTORC1 irrespective of growth factor stimulation. These findings suggest that COUP-TFII is a novel oncogenic mediator, connecting KRAS signaling and glycolysis, and leading to mTORC1 activation and cellular growth.

Glutamine-derived aspartate is required for eIF5A hypusination-mediated translation of HIF-1α to induce the polarization of tumor-associated macrophages.

Tumor-associated macrophages (TAMs) are vital contributors to the growth, metastasis, and therapeutic resistance of various cancers, including hepatocellular carcinoma (HCC). However, the exact phenotype of TAMs and the mechanisms underlying their modulation for therapeutic purposes have not been determined. Here, we present compelling evidence that glutamine-derived aspartate in TAMs stimulates spermidine production through the polyamine synthesis pathway, thereby increasing the translation efficiency of HIF-1α via eIF5A hypusination. Consequently, augmented translation of HIF-1α drives TAMs to undergo an increase glycolysis and acquire a metabolic phenotype distinct from that of M2 macrophages. Finally, eIF5A levels in tumor stromal lesions were greater than those in nontumor stromal lesions. Additionally, a higher degree of tumor stromal eIF5A hypusination was significantly associated with a more advanced tumor stage. Taken together, these data highlight the potential of inhibiting hypusinated eIF5A by targeting glutamine metabolism in TAMs, thereby opening a promising avenue for the development of novel therapeutic approaches for HCC.

Targeting YAP Activity and Glutamine Metabolism Cooperatively Suppresses Tumor Progression by Preventing Extracellular Matrix Accumulation.

Cancer cells use multiple mechanisms to evade the effects of glutamine metabolism inhibitors. The pathways that govern responses to alterations in glutamine availability within the tumor may represent therapeutic targets for combinatorial strategies with these inhibitors. Here, we showed that targeting glutamine utilization stimulated Yes-associated protein (YAP) signaling in cancer cells by reducing cyclic adenosine monophosphate (cAMP)/protein kinase A (PKA)-dependent phosphorylation of large tumor suppressor (LATS). Elevated YAP activation induced extracellular matrix (ECM) deposition by increasing secretion of connective tissue growth factor (CTGF) that promoted production of fibronectin and collagen by surrounding fibroblasts. Consequently, inhibiting YAP synergized with inhibition of glutamine utilization to effectively suppress tumor growth in vivo, along with a concurrent decrease in ECM deposition. Blocking ECM remodeling also augmented the tumor suppressive effects of the glutamine utilization inhibitor. Collectively, these data reveal mechanisms by which targeting glutamine utilization increases ECM accumulation and identify potential strategies to reduce ECM levels and increase the efficacy of glutamine metabolism inhibitors.

VGLL3 expression is associated with macrophage infiltration and predicts poor prognosis in epithelial ovarian cancer.

Background/objective: High-grade serous ovarian carcinoma (HGSOC) is the most common histologic type of epithelial ovarian cancer (EOC). Due to its poor survival outcomes, it is essential to identify novel biomarkers and therapeutic targets. The hippo pathway is crucial in various cancers, including gynaecological cancers. Herein, we examined the expression of the key genes of the hippo pathway and their relationship with clinicopathological significance, immune cells infiltration and the prognosis of HGSOC. Methods: The Cancer Genome Atlas (TCGA) and Gene Expression Omnibus (GEO) data were curated to analyse the mRNA expression as well as the clinicopathological association and correlation with immune cell infiltration in HGSOC. The protein levels of significant genes in the HGSOC tissue were analysed using Tissue Microarray (TMA)-based immunohistochemistry. Finally, DEGs pathway analysis was performed to find the signalling pathways associated with VGLL3. Results: VGLL3 mRNA expression was significantly correlated with both advanced tumor stage and poor overall survival (OS) (p=0.046 and p=0.003, respectively). The result of IHC analysis also supported the association of VGLL3 protein with poor OS. Further, VGLL3 expression was significantly associated with tumor infiltrating macrophages. VGLL3 expression and macrophages infiltration were both found to be independent prognostic factors (p=0.003 and p=0.024, respectively) for HGSOC. VGLL3 was associated with four known and three novel cancer-related signalling pathways, thus implying that VGLL3 is involved in the deregulation of many genes and pathways. Conclusion: Our study revealed that VGLL3 may play a distinct role in clinical outcomes and immune cell infiltration in patients with HGSOC and that it could potentially be a prognostic marker of EOC.

Macropinocytosis is an alternative pathway of cysteine acquisition and mitigates sorafenib-induced ferroptosis in hepatocellular carcinoma.

BACKGROUND: Macropinocytosis, an important nutrient-scavenging pathway in certain cancer cells, allows cells to compensate for intracellular amino acid deficiency under nutrient-poor conditions. Ferroptosis caused by cysteine depletion plays a pivotal role in sorafenib responses during hepatocellular carcinoma (HCC) therapy. However, it is not known whether macropinocytosis functions as an alternative pathway to acquire cysteine in sorafenib-treated HCC, and whether it subsequently mitigates sorafenib-induced ferroptosis. This study aimed to investigate whether sorafenib drives macropinocytosis induction, and how macropinocytosis confers ferroptosis resistance on HCC cells. METHODS: Macropinocytosis, both in HCC cells and HCC tissues, was evaluated by measuring TMR-dextran uptake or lysosomal degradation of DQ-BSA, and ferroptosis was evaluated via C11-BODIPY fluorescence and 4-HNE staining. Sorafenib-induced ferroptosis and macropinocytosis were validated in tumor tissues taken from HCC patients who underwent ultrasound-guided needle biopsy. RESULTS: Sorafenib increased macropinocytosis in human HCC specimens and xenografted HCC tissues. Sorafenib-induced mitochondrial dysfunction was responsible for activation of PI3K-RAC1-PAK1 signaling, and amplified macropinocytosis in HCC. Importantly, macropinocytosis prevented sorafenib-induced ferroptosis by replenishing intracellular cysteine that was depleted by sorafenib treatment; this rendered HCC cells resistant to sorafenib. Finally, inhibition of macropinocytosis by amiloride markedly enhanced the anti-tumor effect of sorafenib, and sensitized resistant tumors to sorafenib. CONCLUSION: In summary, sorafenib induced macropinocytosis, which conferred drug resistance by mitigating sorafenib-induced ferroptosis. Thus, targeting macropinocytosis is a promising therapeutic strategy to facilitate ferroptosis-based therapy for HCC.

DN200434, an orally available inverse agonist of estrogen-related receptor γ, induces ferroptosis in sorafenib-resistant hepatocellular carcinoma.

Sorafenib, originally identified as an inhibitor of multiple oncogenic kinases, induces ferroptosis in hepatocellular carcinoma (HCC) cells. Several pathways that mitigate sorafenib-induced ferroptosis confer drug resistance; thus strategies that enhance ferroptosis increase sorafenib efficacy. Orphan nuclear receptor estrogen-related receptor γ (ERRγ) is upregulated in human HCC tissues and plays a role in cancer cell proliferation. The aim of this study was to determine whether inhibition of ERRγ with DN200434, an orally available inverse agonist, can overcome resistance to sorafenib through induction of ferroptosis. Sorafenib-resistant HCC cells were less sensitive to sorafenibinduced ferroptosis and showed significantly higher ERRγ levels than sorafenib-sensitive HCC cells. DN200434 induced lipid peroxidation and ferroptosis in sorafenib-resistant HCC cells. Mechanistically, DN200434 increased mitochondrial ROS generation by reducing glutathione/glutathione disulfide levels, which subsequently reduced mTOR activity and GPX4 levels. DN200434 induced amplification of the antitumor effects of sorafenib was confirmed in a tumor xenograft model. The present results indicate that DN200434 may be a novel therapeutic strategy to re-sensitize HCC cells to sorafenib. [BMB Reports 2022; 55(11): 547-552].