ABCC2 induces metabolic vulnerability and cellular ferroptosis via enhanced glutathione efflux in gastric cancer.

BACKGROUND: Although it is traditionally believed that ATP binding cassette subfamily C member 2 (ABCC2) is a multidrug resistance-associated protein correlated with a worse prognosis, our previous and several other studies demonstrated the contrary to be true in gastric cancer (GC). We aim to explore the underlying mechanism of this discovery. METHODS: Our study utilized whole-exome sequencing (WES), RNA sequencing, and droplet digital PCR (ddPCR) analysis of 80 gastric cancer samples, along with comprehensive immunohistochemical (IHC) analysis of 1044 human GC tissue samples.By utilizing CRISPRCas9 to genetically modify cell lines with the ABCC2-24C > T (rs717620) point mutation and conducting dual-luciferase reporter assays, we identified that transcription factors SOX9 and ETS1 serve as negative regulators of ABCC2 expression. Seahorse assay and mass spectrometry were used to discover altered metabolic patterns. Gain and loss-of-function experiments in GC cell lines and preclinical models were carried out to validate ABCC2 biological function. RESULTS: ABCC2 high expression correlated with better prognosis, and rs717620 can influence ABCC2 expression by disrupting the binding of ETS1 and SOX9. Gain and loss-of-function experiments in GC cell lines demonstrated amino acid deprivation reduces proliferation, migration, and drug resistance in ABCC2-high GC cells. ABCC2 leads to reduced intracellular amino acid pools and disruption of cellular energy metabolism. This phenomenon depended on ABCC2-mediated GSH extrusion, resulting in alterations in redox status, thereby increasing the cell's susceptibility to ferroptosis. Furthermore, patient-derived organoids and patient-derived tumor-like cell clusters were used to observe impact of ABCC2 on therapeutic effect. In the xenograft model with high ABCC2 expression, we observed that constricting amino acid intake in conjunction with GPX4 inactivation resulted in notable tumor regression. CONCLUSIONS: Our findings demonstrate a significant role of ABCC2 in amino acid metabolism and ferroptosis by mediating GSH efflux in GC. This discovery underlines the potential of combining multiple ferroptosis targets as a promising therapeutic strategy for GC with high ABCC2 expression. HIGHLIGHTS: ABCC2 plays a crucial role in inducing metabolic vulnerability and ferroptosis in gastric cancer through enhanced glutathione efflux. The ABCC2 24C > T polymorphism is a key factor influencing its expression. These results highlight the potential of ABCC2 as a predictive biomarker and therapeutic target in gastric cancer.

Using Normalized Carcinoembryonic Antigen and Carbohydrate Antigen 19 to Predict and Monitor the Efficacy of Neoadjuvant Chemotherapy in Locally Advanced Gastric Cancer.

Carcinoembryonic antigen (CEA) and carbohydrate antigen 19-9 (CA19-9) are established prognostic biomarkers for patients with gastric cancer. However, their potential as predictive markers for neoadjuvant chemotherapy (NACT) efficacy has not been fully elucidated. METHODS: We conducted a retrospective analysis to determine values of CEA and CA19-9 prior to NACT (pre-NACT) and after NACT (post-NACT) in 399 patients with locally advanced gastric cancer (LAGC) who received intended NACT and surgery. RESULTS: Among the 399 patients who underwent NACT plus surgery, 132 patients (33.1%) had elevated pre-NACT CEA/CA19-9 values. Furthermore, either pre-NACT or post-NACT CEA /CA19-9 levels were significantly associated with prognosis (p = 0.0023) compared to patients with non-elevated levels. Moreover, among the patients, a significant proportion (73/132, 55.3%) achieved normalized CEA/CA19-9 following NACT, which is a strong marker of a favorable treatment response and survival benefits. In addition, the patients with normalized CEA/CA19-9 also had a prolonged survival compared to those who underwent surgery first (p = 0.0140), which may be attributed to the clearance of micro-metastatic foci. Additionally, the magnitude of CEA/CA19-9 changes did not exhibit a statistically significant prognostic value. CONCLUSIONS: Normalization of CEA/CA19-9 is a strong biomarker for the effectiveness of treatment, and can thus be exploited to prolong the long-term survival of patients with LAGC.

Identification of Immunogenic Cell-Death-Related Subtypes and Development of a Prognostic Signature in Gastric Cancer.

BACKGROUND: Immunogenic cell death (ICD) is considered a promising type of regulated cell death and exerts effects by activating the adaptive immune response, reshaping the tumor environment (TME) and improving therapeutic efficacy. However, the potential roles and prognostic value of ICD-associated genes in gastric cancer (GC) remain unclear. METHODS: The RNA expression data and clinical information of 1090 GC patients from six cohorts were collected. Consensus clustering was used to identify three distinct molecular subtypes. Then, a robust prognostic ICD_score for predicting prognosis was built via WGCNA and LASSO Cox regression according to the TCGA cohort, and the predictive capability of the ICD_score in GC patients was validated in the other cohorts. ICD-related immune features were analyzed using a CIBERSORT method and verified by immunofluorescence. RESULTS: We found that ICD-related gene variations were correlated with clinical outcomes, tumor immune microenvironment (TIME) characteristics and treatment response. We then constructed an ICD signature that classifies cases as low- and high-ICD_score groups. The high-ICD_score group indicates unfavorable OS, a more advanced TNM stage, and presents an immune-suppressed phenotype, which has more infiltrations of pro-tumor immune cells, such as macrophages, which was verified by immunofluorescence. In addition, a nomogram containing the ICD_score showed a high predictive accuracy with AUCs of 0.715, 0.731 and 0.8 on Years 1, 3, and 5. CONCLUSION: We performed the first and synthesis ICD analysis in GC and built a clinical application tool based on the ICD signature, which paved a new path for assessing prognosis and guiding individual treatment.

Aspartate ß-Hydroxylase Serves as a Prognostic Biomarker for Neoadjuvant Chemotherapy in Gastric Cancer.

Neoadjuvant chemotherapy (NACT) has been established as being an effective treatment for advanced gastric cancer (GC), while the predictive biomarker of NACT efficacy remains under investigation. Aspartate ß-hydroxylase (ASPH) represents an attractive target which is a highly conserved transmembrane enzyme overexpressed in human GC, and participates in the malignant transformation by promoting tumor cell motility. Here, we evaluated the expression of ASPH by immunohistochemistry in 350 GC tissues (including samples for NACT) and found that ASPH expression was higher in patients undergoing NACT compared with patients without NACT pre-operation. The OS and PFS time of ASPH-intensely positive patients was significantly shorter than that of the negative patients in the NACT group, while the difference was not significant in patients without NACT. We showed that ASPH knockout enhanced the inhibitory effects of chemotherapeutic drugs on the cell proliferation, migration, and invasion in vitro and suppressed tumor progression in vivo. Co-immunoprecipitation revealed that ASPH might interact with LAPTM4B to perform chemotherapeutic drug resistance. Our results suggested that ASPH might serve as a candidate biomarker to predict prognosis and a novel therapeutic target for gastric cancer patients treated with neoadjuvant chemotherapy.

Clinical Significance and Immune Infiltration Analyses of the Cuproptosis-Related Human Copper Proteome in Gastric Cancer.

BACKGROUND: The human copper Cu proteome, also termed Cu-binding proteins (CBP), is responsible for transporting "free" Cu to the cell that is related to cuproptosis. However, their role in gastric cancer (GC) has not been reported. METHODS: RNA expression data of 946 GC patients were collected. A series of machine learning and bioinformatic approaches were combined to build a CBP signature to predict survival and immune microenvironment and guide the priority treatment. Immunohistochemistry and multicolor immunofluorescence (mIF) in 1076 resection slides were used to verify immune features. RESULTS: A CBP signature was constructed using the machine learning method from TCGA that classifies cases as CBP_low and CBP_high groups. Multivariable Cox analysis confirmed that the CBP signature was an independent prognostic factor in the training and validation cohorts. Additionally, GC patients with low CBPscores showed an increase in anti-tumor immune microenvironment, which was further verified by mIF in pathological resections following immunotherapy. Importantly, patients with low CBPscores had higher levels of TMB/MSI and responded well to immunotherapy. CONCLUSIONS: We conducted the first and comprehensive CBP analysis of GC patients and established a clinically feasible CBP signature for predicting survival and response to treatment, which will be helpful for guiding personalized medicine.

A Novel Classification and Scoring Method Based on Immune-Related Transcription Factor Regulation Patterns in Gastric Cancer.

Background: Transcription factors (TFs) play a crucial role in tumorigenesis and anti-tumor immunity. However, the potential role of large-scale transcription factor regulation patterns in the progression in gastric cancer (GC) is unknown. Methods: We comprehensively assessed the relevance of immune-related TF (IRTF) regulation patterns in anti-tumor immunity and immunotherapy in 1,136 gastric cancer (GC) patients, and evaluated the IRTF score based on IRTF regulation patterns using random forests. Results: Two distinct IRTF regulation patterns were identified, which demonstrating the distinct characteristics in clinical phenotypes, tumor immune microenvironment (TIME), immunogenicity and prognosis in GC. Subsequently, the IRTF score was established to quantify the IRTF regulation pattern for each GC patient. Analysis of large conventional therapy cohorts showed low IRTF score was associated with a better prognosis. In addition, analysis of multiple immunotherapy cohorts showed low IRTF score was also linked to enhanced response to immunotherapy. Conclusion: TF regulation patterns were found to play an important role in the complex immune regulatory relationships in GC. Evaluation of the IRTF regulation patterns in patients will enhance our understanding of immune specificities, and thus, provide effective strategies for personalized therapy.

Novel prognostic marker LINC00205 promotes tumorigenesis and metastasis by competitively suppressing miRNA-26a in gastric cancer.

Rapid proliferation and metastasis of gastric cancer (GC) resulted in a poor prognosis in the clinic. Previous studies elucidated that long non-coding RNA (LncRNA) LINC00205 was upregulated in various tumors and participated in tumor progression. The aim of our study was to investigate the regulating role of LINC00205 in tumorigenesis and metastasis of GC. Both public datasets and our data showed that the LINC00205 was highly expressed in GC tissues and several cell lines. Notably, GC patients with high level of LINC00205 had a poor prognosis in our cohort. Mechanistically, knockdown of LINC00205 by shRNAs suppressed GC cells proliferation, migration, invasion remarkably, and induced cell cycle arrest. Based on bioinformatics prediction, we found that LINC00205 might act as a competitive endogenous RNA (ceRNA) through targeting miR-26a. The level of miR-26a had negatively correlated with LINC00205 expression and was decreased among GC cell lines, tissues, and serum samples. Our results for the first time confirmed that miR-26a was a direct target of LINC00205 and might have the potential to become a plasma marker for clinical tumor diagnosis. Indeed, LINC00205 knockdown resulted in the dramatic promotion of miR-26a expression as well as inhibition of miR-26a potential downstream targets, such as HMGA2, EZH2, and USP15. These targets were essential for cell survival and epithelial-mesenchymal transition. Importantly, LINC00205 was able to remodel the miR-26a-mediated downstream silence, which identified a new mechanism of malignant transformation of GC cells. In conclusion, this study revealed the regulating role of the LINC00205/miR-26a axis in GC progression and provided a new potential therapeutic strategy for GC treatment.