Acetylation of aldehyde dehydrogenase ALDH1L2 regulates cellular redox balance and the chemosensitivity of colorectal cancer to 5-fluorouracil.

Folate-mediated one-carbon metabolism (FOCM) is crucial in sustaining rapid proliferation and survival of cancer cells. The folate cycle depends on a series of key cellular enzymes, including aldehyde dehydrogenase 1 family member L2 (ALDH1L2) that is usually overexpressed in cancer cells, but the regulatory mechanism of ALDH1L2 remains undefined. In this study, we observed the significant overexpression of ALDH1L2 in colorectal cancer (CRC) tissues, which is associated with poor prognosis. Mechanistically, we identified that the acetylation of ALDH1L2 at the K70 site is an important regulatory mechanism inhibiting the enzymatic activity of ALDH1L2 and disturbing cellular redox balance. Moreover, we revealed that sirtuins 3 (SIRT3) directly binds and deacetylates ALDH1L2 to increase its activity. Interestingly, the chemotherapeutic agent 5-fluorouracil (5-Fu) inhibits the expression of SIRT3 and increases the acetylation levels of ALDH1L2 in colorectal cancer cells. 5-Fu-induced ALDH1L2 acetylation sufficiently inhibits its enzymatic activity and the production of NADPH and GSH, thereby leading to oxidative stress-induced apoptosis and suppressing tumor growth in mice. Furthermore, the K70Q mutant of ALDH1L2 sensitizes cancer cells to 5-Fu both in vitro and in vivo through perturbing cellular redox and serine metabolism. Our findings reveal an unknown 5-Fu-SIRT3-ALDH1L2 axis regulating redox homeostasis, and suggest that targeting ALDH1L2 is a promising therapeutic strategy to sensitize tumor cells to chemotherapeutic agents.

Long noncoding RNA DLGAP1-AS2 promotes tumorigenesis and metastasis by regulating the Trim21/ELOA/LHPP axis in colorectal cancer.

BACKGROUND: Long noncoding RNAs (lncRNAs) have driven research focused on their effects as oncogenes or tumor suppressors involved in carcinogenesis. However, the functions and mechanisms of most lncRNAs in colorectal cancer (CRC) remain unclear. METHODS: The expression of DLGAP1-AS2 was assessed by quantitative RT-PCR in multiple CRC cohorts. The impacts of DLGAP1-AS2 on CRC growth and metastasis were evaluated by a series of in vitro and in vivo assays. Furthermore, the underlying mechanism of DLGAP1-AS2 in CRC was revealed by RNA pull down, RNA immunoprecipitation, RNA sequencing, luciferase assays, chromatin immunoprecipitation, and rescue experiments. RESULTS: We discovered that DLGAP1-AS2 promoted CRC tumorigenesis and metastasis by physically interacting with Elongin A (ELOA) and inhibiting its protein stability by promoting tripartite motif containing 21 (Trim21)-mediated ubiquitination modification and degradation of ELOA. In particular, we revealed that DLGAP1-AS2 decreases phospholysine phosphohistidine inorganic pyrophosphate phosphatase (LHPP) expression by inhibiting ELOA-mediated transcriptional activating of LHPP and thus blocking LHPP-dependent suppression of the AKT signaling pathway. In addition, we also demonstrated that DLGAP1-AS2 was bound and stabilized by cleavage and polyadenylation specificity factor (CPSF2) and cleavage stimulation factor (CSTF3). CONCLUSIONS: The discovery of DLGAP1-AS2, a promising prognostic biomarker, reveals a new dimension into the molecular pathogenesis of CRC and provides a prospective treatment target for this disease.

A novel high-risk subpopulation identified by CTSL and ZBTB7B in gastric cancer.

BACKGROUND: Gastric cancer (GC) is characterised by a heterogeneous tumour microenvironment (TME) that is closely associated with the response to treatment, especially immunotherapies. However, most previous GC molecular subtyping systems need complex gene signatures and examination methods, restricting their clinical applications. Thus, we developed a new TME-based molecular subtype using only two genes. METHODS: Nine independent GC cohorts at the tissue- or single-cell level with more than 2000 patients were used in this study, including data we examined by single-cell sequencing, quantitative RT-PCR and immunochemistry/immunofluorescence staining. Nine different methods, five existing molecular subtypes and a series of signatures were used to evaluate the TME and molecular characteristics of GC. RESULTS: We established a CTSL/ZBTB7B subtyping system and uncovered the novel CTSLHighZBTB7BLow high-risk subgroup, but characterised by relative higher immune cell infiltration and lower tumour purity. This subgroup demonstrate higher levels of immune checkpoints and more enrichment of cancer-related pathways compared with other cases. CONCLUSIONS: We identified a high-risk subpopulation with unique TME features based on expressions of CTSL and ZBTB7B, suggesting a counterbalancing phenotype between immunostimulatory and immunosuppressive mechanisms. This subtyping system could be used to select treatment and management strategies for GC.

SLCO4A1-AS1 promotes colorectal tumourigenesis by regulating Cdk2/c-Myc signalling.

BACKGROUND: SLCO4A1-AS1 was found to be upregulated in several cancer types, including colorectal cancer (CRC). However, the detailed roles of SLCO4A1-AS1 in CRC remain to be elucidated. Therefore, we investigated the functions, mechanism, and clinical significance of SLCO4A1-AS1 in colorectal tumourigenesis. METHODS: We measured the expression of SLCO4A1-AS1 in CRC tissues using qRT-PCR and determined its correlation with patient prognosis. Promoter methylation analyses were used to assess the methylation status of SLCO4A1-AS1. Gain- and loss-of-function assays were used to evaluate the effects of SLCO4A1-AS1 on CRC growth in vitro and in vivo. RNA pull-down, RNA immunoprecipitation, RNA-seq, luciferase reporter and immunohistochemistry assays were performed to identify the molecular mechanism of SLCO4A1-AS1 in CRC. RESULTS: SLCO4A1-AS1 was frequently upregulated in CRC tissues based on multiple CRC cohorts and was associated with poor prognoses. Aberrant overexpression of SLCO4A1-AS1 in CRC is partly attributed to the DNA hypomethylation of its promoter. Ectopic SLCO4A1-AS1 expression promoted CRC cell growth, whereas SLCO4A1-AS1 knockdown repressed CRC proliferation both in vitro and in vivo. Mechanistic investigations revealed that SLCO4A1-AS1 functions as a molecular scaffold to strengthen the interaction between Hsp90 and Cdk2, promoting the protein stability of Cdk2. The SLCO4A1-AS1-induced increase in Cdk2 levels activates the c-Myc signalling pathway by promoting the phosphorylation of c-Myc at Ser62, resulting in increased tumour growth. CONCLUSIONS: Our data demonstrate that SLCO4A1-AS1 acts as an oncogene in CRC by regulating the Hsp90/Cdk2/c-Myc axis, supporting SLCO4A1-AS1 as a potential therapeutic target and prognostic factor for CRC.

MicroRNA-204-5p: A pivotal tumor suppressor.

MicroRNAs (miRNAs) are a class of non-coding single-stranded RNA molecules with a length of approximately 18-25 nt nucleotides that regulate gene expression post-transcriptionally. MiR-204-5p originates from the sixth intron of the transient receptor potential cation channel subfamily M member 3 (TRPM3) gene. MiR-204-5p is frequently downregulated in various cancer types and is related to the clinicopathological characteristics and prognosis of cancer patients. So far, many studies have determined that miR-204-5p functions as a tumor suppressor for its extensive and powerful capacity to inhibit tumor proliferation, metastasis, autophagy, and chemoresistance in multiple cancer types. MiR-204-5p appears to be a promising prognostic biomarker and a therapeutic target for human cancers. This review summarized the latest advances on the role of miR-204-5p in human cancers.

SNHG15 promotes chemoresistance and glycolysis in colorectal cancer.

Long noncoding RNAs (lncRNAs) play an important role in tumor progression. Small nucleolar RNA host gene 15 (SNHG15) is a lncRNA that has been confirmed to play an oncogenic role in multiple cancer types. However, its role in glycolysis and chemoresistance in colorectal cancer (CRC) is unclear. The expression of SNHG15 in CRC was analyzed using data from The Cancer Genome Atlas (TCGA) and Gene Expression Omnibus (GEO) databases by bioinformatics methods. Cell Counting Kit-8 (CCK-8) and colony formation assays were used to evaluate cell viability. Cell sensitivity to 5-fluorouracil (5-FU) was detected by CCK-8. Glucose absorption and lactate production were used to evaluate the impact of SNHG15 on glycolysis. RNA-seq, real-time fluorescence quantitative reverse transcription PCR (RT-qPCR) and Western blotting (WB) were used to reveal the potential molecular mechanism of SNHG15 in CRC. SNHG15 was upregulated in CRC tissues compared with paired noncancerous tissues. Ectopic SNHG15 expression increased proliferation, 5-FU chemoresistance, and glycolysis in CRC cells. In contrast, SNHG15 knockdown inhibited CRC proliferation, 5-FU chemoresistance and glycolysis. Multiple pathways, including apoptosis and glycolysis, were potentially regulated by SNHG15 based on RNA-seq and pathway enrichment analyses. RT-qPCR and WB experiments confirmed that SNHG15 promoted the expression of TYMS, BCL2, GLUT1 and PKM2 in CRC cells. In conclusion, SNHG15 promotes 5-FU chemoresistance and glycolysis in CRC by potentially regulating the expression of TYMS, BCL2, GLUT1 and PKM2 and appears to be a new target for cancer therapy.

EXOSC8 promotes colorectal cancer tumorigenesis via regulating ribosome biogenesis-related processes.

Extensive protein synthesis is necessary for uncontrolled cancer cell proliferation, requiring hyperactive ribosome biogenesis. Our previous Pan-cancer study has identified EXOSC8 as a potential copy number variation (CNV)-driven rRNA metabolism-related oncogene in colorectal cancer (CRC). Herein, we further investigated proliferation-prompting functions and mechanisms of EXOSC8 in CRC by performing in silico analyses and wet-lab experiments. We uncovered that increased EXOSC8 expression and CNV levels are strongly associated with ribosome biogenesis-related factor levels in CRC, including ribosome proteins (RPs), eukaryotic translation initiation factors and RNA polymerase I/III. EXOSC8 silence decreases nucleolar protein and proliferation marker levels, as well as rRNA/DNA and global protein syntheses. Clinically, EXOSC8 is upregulated across human cancers, particularly CNV-driven upregulation in CRC was markedly associated with poor clinical outcomes. Mechanistically, EXOSC8 knockdown increased p53 levels in CRC, and the oncogenic proliferation phenotypes of EXOSC8 depended on p53 in vitro and in vivo. We discovered that EXOSC8 knockdown in CRC cells triggers ribosomal stress, nucleolar RPL5/11 being released into the nucleoplasm and "hijacking" Mdm2 to block its E3 ubiquitin ligase function, thus releasing and activating p53. Furthermore, our therapeutic experiments provided initial evidence that EXOSC8 might serve as a potential therapeutic target in CRC. Our findings revealed, for the first time, that the RNA exosome gene (EXOSC8) promotes CRC tumorigenesis by regulating cancer-related ribosome biogenesis in CRC. This study further extends our previous Pan-cancer study of the rRNA metabolism-related genes. The inhibition of EXOSC8 is a novel therapeutic strategy for the RPs-Mdm2-p53 ribosome biogenesis surveillance pathway in CRC.

Comprehensive Characterization of Integrin Subunit Genes in Human Cancers.

Although integrin subunit genes (ITGs) have been reported to be associated with some human cancer types, a systematic assessment of ITGs across human cancers is lacking. Hence, we performed comprehensive analyses to investigate mRNA expression, copy number variation (CNV), DNA methylation, mutation, and clinical landscapes of ITGs in more than 8000 cancer patients from The Cancer Genome Atlas (TCGA) dataset. Landscapes of ITGs were established across 20 human cancer types. We observed that ITGs are extensively dysregulated with heterogeneity in different system cancer types, part of which are driven by CNV, DNA hypomethylation or mutation. Furthermore, dysregulated prognosis-related ITGs were systematically identified in each cancer type, including ITGA11 in stomach adenocarcinoma (STAD). The models based on dysregulated ITGs with clinical relevance and TNM staging indexes are good indicators in STAD and head and neck squamous cell carcinoma. Finally, ITGA11 is overexpressed and associated with poor survival in STAD cases from the TCGA and additionally Gene Expression Omnibus cohorts. Functionally, ITGA11 knockdown inhibits malignant phenotypes in STAD cell lines AGS and MKN45, demonstrating the oncogenic role of ITGA11 in STAD. Together, this study highlights the important roles of ITGs in tumorigenesis as potential prognostic biomarkers, and provide an effective resource that identifies cancer-related genes of ITGs in human cancers.

Hsa_circ_0062682 Promotes Serine Metabolism and Tumor Growth in Colorectal Cancer by Regulating the miR-940/PHGDH Axis.

Colorectal cancer (CRC) is one of the most common malignancies globally. Increasing evidence indicates that circular RNAs (circRNAs) play a pivotal role in various cancers. The present study focused on exploring the role of a functionally unknown circRNA, hsa_circ_0062682 (circ_0062682), in CRC. By online analyses and experimental validations, we showed that circ_0062682 expression was aberrantly increased in CRC tissues compared with paired normal tissues. Increased expression of circ_0062682 in CRC notably correlated with a poor prognosis and advanced tumor stage. Functional experiments showed that circ_0062682 knockdown reduced CRC growth both in vitro and in vivo. Mechanistically, we revealed that circ_0062682 could sponge miR-940 and identified D-3-phosphoglycerate dehydrogenase (PHGDH), a key oxidoreductase involved in serine biosynthesis, as a novel target of miR-940. Silencing miR-940 expression could mimic the inhibitory effect of circ_0062682 knockdown on CRC proliferation. The expression of PHGDH was downregulated in circ_0062682-depleted or miR-940 overexpressing CRC cells at both the mRNA and protein levels. Circ_0062682 knockdown suppressed CRC growth by decreasing PHGDH expression and serine production via miR-940. Taken together, these data demonstrate, for the first time, that circ_0062682 promotes serine metabolism and tumor growth in CRC by regulating the miR-940/PHGDH axis, suggesting circ_0062682 as a potential novel therapeutic target for CRC.

SNHG17 promotes colorectal tumorigenesis and metastasis via regulating Trim23-PES1 axis and miR-339-5p-FOSL2-SNHG17 positive feedback loop.

BACKGROUND: Small nucleolar RNA host gene (SNHG) long noncoding RNAs (lncRNAs) are frequently dysregulated in human cancers and involved in tumorigenesis and progression. SNHG17 has been reported as a candidate oncogene in several cancer types, however, its regulatory role in colorectal cancer (CRC) is unclear. METHODS: SNHG17 expression in multiple CRC cohorts was assessed by RT-qPCR or bioinformatic analyses. Cell viability was evaluated using Cell Counting Kit-8 (CCK-8) and colony formation assays. Cell mobility and invasiveness were assessed by Transwell assays. Tumor xenograft and metastasis models were applied to confirm the effects of SNHG17 on CRC tumorigenesis and metastasis in vivo. Immunohistochemistry staining was used to measure protein expression in cancer tissues. RNA pull-down, RNA immunoprecipitation, chromatin immunoprecipitation, and dual luciferase assays were used to investigate the molecular mechanism of SNHG17 in CRC. RESULTS: Using multiple cohorts, we confirmed that SNHG17 is aberrantly upregulated in CRC and correlated with poor survival. In vitro and in vivo functional assays indicated that SNHG17 facilitates CRC proliferation and metastasis. SNHG17 impedes PES1 degradation by inhibiting Trim23-mediated ubiquitination of PES1. SNHG17 upregulates FOSL2 by sponging miR-339-5p, and FOSL2 transcription activates SNHG17 expression, uncovering a SNHG17-miR-339-5p-FOSL2-SNHG17 positive feedback loop. CONCLUSIONS: We identified SNHG17 as an oncogenic lncRNA in CRC and identified abnormal upregulation of SNHG17 as a prognostic risk factor for CRC. Our mechanistic investigations demonstrated, for the first time, that SNHG17 promotes tumor growth and metastasis through two different regulatory mechanisms, SNHG17-Trim23-PES1 axis and SNHG17-miR-339-5p-FOSL2-SNHG17 positive feedback loop, which may be exploited for CRC therapy.

Long noncoding RNA MCM3AP-AS1 enhances cell proliferation and metastasis in colorectal cancer by regulating miR-193a-5p/SENP1.

BACKGROUND: Accumulating evidences have shown that long noncoding RNAs (lncRNAs) play key roles in many diseases, including cancer. Several studies reported that MCM3AP antisense RNA 1 (MCM3AP-AS1) was associated with the tumorigenesis and progression. However, the specific function and mechanism of MCM3AP-AS1 in colorectal cancer (CRC) have not been fully understood. METHODS: The expression of MCM3AP-AS1 was detected by quantitative reverse transcription PCR (RT-qPCR) in CRC tissues and matched noncancerous tissues (NCTs). CCK-8 assay, colony formation assay, transwell assay, xenograft and lung metastasis mouse models were used to examine the tumor-promoting function of MCM3AP-AS1 in vitro and in vivo. The binding relationship between MCM3AP-AS1, miR-193a-5p and sentrin-specific peptidase 1 (SENP1) were screened and identified by databases, RT-qPCR, dual luciferase reporter assay and western blot. RESULTS: In the present study, we got that the expression of MCM3AP-AS1 was higher in CRC tissues than in paired NCTs, and increased MCM3AP-AS1 expression was associated with adverse outcomes in CRC patients. Functional experiments in vitro revealed that silencing of MCM3AP-AS1 could inhibit the proliferation, colony formation, migratory, and invasive abilities of CRC cells. The mouse models of xenograft and lung metastasis further confirmed that in vivo silencing MCM3AP-AS1 could significantly inhibit the growth and metastasis of CRC. Further mechanism studies indicated that MCM3AP-AS1 could sponge miR-193a-5p and inhibit the activity of it. What is more, SENP1 was proved to be a novel target of miR-193a-5p and could be upregulated by MCM3AP-AS1. At last, we observed that SENP1 overexpression in CRC tissues was closely related to unfavorable prognosis. CONCLUSION: Taken together, we identified in CRC the MCM3AP-AS1/miR-193a-5p/SENP1 regulatory axis, which affords a therapeutic possibility for CRC.