Distinctive tumorigenic significance and innovative oncology targets of SUMOylation.

Protein SUMOylation, a post-translational modification, intricately regulates diverse biological processes including gene expression, cell cycle progression, signaling pathway transduction, DNA damage response, and RNA metabolism. This modification contributes to the acquisition of tumorigenicity and the maintenance of cancer hallmarks. In malignancies, protein SUMOylation is triggered by various cellular stresses, promoting tumor initiation and progression. This augmentation is orchestrated through its specific regulatory mechanisms and characteristic biological functions. This review focuses on elucidating the fundamental regulatory mechanisms and pathological functions of the SUMO pathway in tumor pathogenesis and malignant evolution, with particular emphasis on the tumorigenic potential of SUMOylation. Furthermore, we underscore the potential therapeutic benefits of targeting the SUMO pathway, paving the way for innovative anti-tumor strategies by perturbing this dynamic and reversible modifying process.

Regulatory mechanisms and therapeutic implications of insulin-like growth factor 2 mRNA-binding proteins, the emerging crucial m6A regulators of tumors.

Insulin-like growth factor 2 mRNA-binding proteins (IGF2BPs) serve essential biological functions as post-transcriptional performers, participating in the acquisition or maintenance of tumor hallmarks due to their distinct protein structures. Emerging evidence indicates that IGF2BPs belong to the class III type of RNA N6-methyladenosine (m6A) modification readers, controlling RNA stability, storage, localization, metabolism, and translation in multiple vital bioprocesses, particularly tumorigenesis and tumor progression. Here, we discuss the underlying regulatory mechanisms and pathological functions of IGF2BPs which act as m6A readers in the context of tumor pathogenesis and multidrug resistance. Furthermore, we highlight the potential of IGF2BPs as drug targets in clinical tumor treatment. Hence, precise and novel tumor therapeutic approaches could be uncovered by targeting epigenetic heterogeneity.

The interplay between non-coding RNAs and alternative splicing: from regulatory mechanism to therapeutic implications in cancer.

Alternative splicing (AS) is a common and conserved process in eukaryotic gene regulation. It occurs in approximately 95% of multi-exon genes, greatly enriching the complexity and diversity of mRNAs and proteins. Recent studies have found that in addition to coding RNAs, non-coding RNAs (ncRNAs) are also inextricably linked with AS. Multiple different types of ncRNAs are generated by AS of precursor long non-coding (pre-lncRNAs) or precursor messenger RNAs (pre-mRNAs). Furthermore, ncRNAs, as a novel class of regulators, can participate in AS regulation by interacting with the cis-acting elements or trans-acting factors. Several studies have implicated abnormal expression of ncRNAs and ncRNA-related AS events in the initiation, progression, and therapy resistance in various types of cancers. Therefore, owing to their roles in mediating drug resistance, ncRNAs, AS-related factors and AS-related novel antigens may serve as promising therapeutic targets in cancer treatment. In this review, we summarize the interaction between ncRNAs and AS processes, emphasizing their great influences on cancer, especially on chemoresistance, and highlighting their potential values in clinical treatment.

The m6A reader IGF2BP2 regulates glutamine metabolism and represents a therapeutic target in acute myeloid leukemia.

N6-Methyladenosine (m6A) modification and its modulators play critical roles and show promise as therapeutic targets in human cancers, including acute myeloid leukemia (AML). IGF2BP2 was recently reported as an m6A binding protein that enhances mRNA stability and translation. However, its function in AML remains largely elusive. Here we report the oncogenic role and the therapeutic targeting of IGF2BP2 in AML. High expression of IGF2BP2 is observed in AML and associates with unfavorable prognosis. IGF2BP2 promotes AML development and self-renewal of leukemia stem/initiation cells by regulating expression of critical targets (e.g., MYC, GPT2, and SLC1A5) in the glutamine metabolism pathways in an m6A-dependent manner. Inhibiting IGF2BP2 with our recently identified small-molecule compound (CWI1-2) shows promising anti-leukemia effects in vitro and in vivo. Collectively, our results reveal a role of IGF2BP2 and m6A modification in amino acid metabolism and highlight the potential of targeting IGF2BP2 as a promising therapeutic strategy in AML.

Circular RNA MTCL1 promotes advanced laryngeal squamous cell carcinoma progression by inhibiting C1QBP ubiquitin degradation and mediating beta-catenin activation.

BACKGROUND: Circular RNAs (circRNAs) are involved in regulatory processes of ubiquitination and deubiquitination in various tumors at post-transcriptional epigenetic modification level. However, the underlying mechanism and its biological functions of circRNAs in the advanced laryngeal squamous cell carcinoma (LSCC) remain obscure. METHODS: RNA sequencing and quantitative real-time PCR (qRT-PCR) assays were applied to screen for circRNAs differentially expressed in LSCC tissues and cell lines. The candidate RNA-binding proteins and target signalling pathway were detected by RNA pull-down and mass spectrometry, in situ hybridization (ISH), immunohistochemistry (IHC), qRT-PCR assays, and bioinformatics analysis. The functional roles of these molecules were investigated using in vitro and in vivo experiments including EdU, transwell, wound healing, western blot assays, and the xenograft mice models. The molecular mechanisms were identified using RNA pull-down assays, RNA immunoprecipitation (RIP), Co-IP, ISH, Ubiquitination assay, bioinformatics analysis, and the rescue experiments. RESULTS: Here, we unveil that microtubule cross-linking factor 1 circRNA (circMTCL1, circ0000825) exerts its critical oncogenic functions by promoting complement C1q-binding protein (C1QBP)-dependent ubiquitin degradation and subsequently activating Wnt/ß-catenin signalling in laryngeal carcinoma initiation and development. Specifically, circMTCL1 was remarkably up-regulated in the paired tissues of patients with LSCC (n = 67), which predicted a worse clinical outcome. Functionally, circMTCL1 exerted oncogenic biological charactersistics by promoting cell proliferative capability and invasive and migrative abilities. Ectopic circMTCL1 augumented cell proliferation, migration, and invasion of LSCC cells, and this effect could be reversed by C1QBP knocking down in vitro and in vivo. Mechanistically, circMTCL1 directly recruited C1QBP protein by harboring the specific recognized sequence (+ 159 - + 210), thereby accelerating the translation of C1QBP expression by inhibiting its ubiquitin-proteasome-mediated degradation. Importantly, the direct interaction of C1QBP with ß-catenin protein was enhanced via suppressing the ß-catenin phosphorylation and accelerating its accumulation in cytoplasm and nucleus. CONCLUSION: Our findings manifested a novel circMTCL1-C1QBP-ß-catenin signaling axis involving in LSCC tumorigenesis and progression, which shed new light on circRNAs-ubiquitous acidic glycoprotein mediated ubiquitin degradation and provided strategies and targets in the therapeutic intervention of LSCC.

[Corrigendum] MicroRNA-148a inhibits breast cancer migration and invasion by directly targeting WNT-1.

Subsequently to the publication of the above article, an interested reader drew to the authors' attention that the data panel for the MDA­MB­231/migration/NC experiment in Fig. 2B on p. 1428 was strikingly similar to the data shown for the MDA­MB­231/invasion/Blank experiment in Fig. 2C, such that these data appeared to have been derived from the same original source. The authors have referred back to their original data, and realize that the data panel was selected incorrectly for Fig. 2B. The corrected version of Fig. 2, showing the correct data for the MDA­MB­231/migration/NC experiment in Fig. 2B, is shown on the next page. The authors regret the error that was made during the preparation of this figure, and can confirm that the error in the assembly of this figure did not adversely affect the conclusions reported in the study. The authors are grateful to the Editor of Oncology Reports for allowing them the opportunity to publish a Corrigendum, and all the authors agree to this Corrigendum. Furthermore, they apologize to the readership for any inconvenience caused. [the original article was published in Oncology Reports 35: 1425­1432, 2016; DOI: 10.3892/or.2015.4502].

A novel HIF-2α targeted inhibitor suppresses hypoxia-induced breast cancer stemness via SOD2-mtROS-PDI/GPR78-UPRER axis.

Hypoxic tumor microenvironment (TME) plays critical roles in induction of cancer stem cell-like phenotype in breast cancer and contribute to chemoresistance. However, the mechanism underlying stemness reprogramming of breast cancer cells (BCs) by hypoxic TME remains largely unknown. In the present study, we illustrated that HIF-2α, but not HIF-1α, induces stemness in BCs under hypoxia through SOD2-mtROS-PDI/GRP78-UPRER pathway, linking mitochondrial metabolic state to endoplasmic reticulum (ER) response via mitochondrial reactive oxygen species (mtROS) level. HIF-2α activates endoplasmic reticulum unfolded protein response (UPRER) in drug-sensitive MCF7 and T47D cells to induce drug-resistant stem-like phenotype. Genetic depletion or pharmacological inhibition (YQ-0629) of HIF-2α abolished hypoxia-induced stem-like phenotype in vitro and in vivo. Mechanistically, HIF-2α activates transcription of superoxide dismutase 2 (SOD2) under hypoxia and thereby decreases mtROS level. With less mtROS transported to endoplasmic reticulum, the expression and activity of protein disulfide isomerase (PDI) is suppressed, allowing glucose-regulated protein 78 (GRP78) to dissociate from receptor proteins of UPRER and bind misfolded protein to activate UPRER, which eventually confer chemoresistance and stem-like properties to BCs. Moreover, the increase in mtROS and PDI levels caused by HIF-2α knockdown and the subsequent UPRER inhibition could be substantially rescued by mitoTEMPOL (a mtROS scavenger), 16F16 (a PDI inhibitor), or GRP78 overexpression. Overall, we reported the critical roles of HIF-2α-SOD2-mtROS-PDI/GRP78-UPRER axis in mediating hypoxia-induced stemness in BCs, highlighting the interaction between organelles and providing evidence for further development of targeted HIF-2α inhibitor as a promising therapeutic strategy for chemoresistant breast cancer.

LINC01021 maintains tumorigenicity by enhancing N6-methyladenosine reader IMP2 dependent stabilization of MSX1 and JARID2: implication in colorectal cancer.

Insulin-like growth factor-2 mRNA-binding protein 2 (IGF2BP2, also known as IMP2), a novel class III N6-methyladenosine (m6A) reader, has recently gained attention due to its critical functions in recognizing and stabilizing m6A modified oncogenic transcripts. However, whether and how long non-coding RNAs (lncRNAs) facilitate IMP2's role as m6A "reader" remains elusive, particularly in colorectal cancer (CRC). Here, we demonstrated that oncogenic LINC021 specifically bound with the m6A "reader" IMP2 protein and enhanced the mRNA stability of MSX1 and JARID2 in an m6A regulatory manner during CRC tumorigenesis and pathogenesis. Specifically, a remarkable upregulation of LINC021 was confirmed in CRC cell lines and clinical tissues (n = 130). High level of LINC021acted as an independent prognostic predictor for CRC clinical outcomes. Functional assays demonstrated that LINC021 exerted its functions as an oncogene to aggravate CRC malignant phenotypes including enhanced cell proliferation, colony formation, migration capabilities, and reduced cell apoptosis. Mechanistically, LINC021 directly recognized IMP2 protein, the latter enhanced the mRNA stability of transcripts such as MSX1 and JARID2 by recognizing their m6A-modified element RGGAC. Thus, these findings uncovered an essential LINC021/IMP2/MSX1 and JARID2 signaling axis in CRC tumorigenesis, which provided profound insights into our understanding of m6A modification regulated by lncRNA in CRC initiation and progression and shed light on the targeting of this axis for CRC treatment.

The Relationship Between the Network of Non-coding RNAs-Molecular Targets and N6-Methyladenosine Modification in Colorectal Cancer.

Recent accumulating researches implicate that non-coding RNAs (ncRNAs) including microRNA (miRNA), circular RNA (circRNA), and long non-coding RNA (lncRNAs) play crucial roles in colorectal cancer (CRC) initiation and development. Notably, N6-methyladenosine (m6A) methylation, the critical posttranscriptional modulators, exerts various functions in ncRNA metabolism such as stability and degradation. However, the interaction regulation network among ncRNAs and the interplay with m6A-related regulators has not been well documented, particularly in CRC. Here, we summarize the interaction networks and sub-networks of ncRNAs in CRC based on a data-driven approach from the publications (IF > 6) in the last quinquennium (2016-2021). Further, we extend the regulatory pattern between the core m6A regulators and m6A-related ncRNAs in the context of CRC metastasis and progression. Thus, our review will highlight the clinical potential of ncRNAs and m6A modifiers as promising biomarkers and therapeutic targets for improving the diagnostic precision and treatment of CRC.

N6-methyladenosine reader IMP2 stabilizes the ZFAS1/OLA1 axis and activates the Warburg effect: implication in colorectal cancer.

BACKGROUND: Accumulating evidence shows that N6-methyladenine (m6A) modulators contribute to the etiology and progression of colorectal cancer (CRC). However, the exact mechanisms of m6A reader involved in glycolytic metabolism remain vague. This article aimed to crosstalk the m6A reader with glycolytic metabolism and reveal a new mechanism for the progression of CRC. METHODS: The relationship between candidate lncRNA and m6A reader was analyzed by bioinformatics, ISH and IHC assays. In vivo and in vitro studies (including MTT, CFA, trans-well, apoptosis, western blot, qRT-PCR and xenograft mouse models) were utilized to explore the biological functions of these indicators. Lactate detection, ATP activity detection and ECAR assays were used to verify the biological function of the downstream target. The bioinformatics, RNA stability, RIP experiments and RNA pull-down assays were used to explore the potential molecular mechanisms. RESULTS: We identified that the crosstalk of the m6A reader IMP2 with long-noncoding RNA (lncRNA) ZFAS1 in an m6A modulation-dependent manner, subsequently augmented the recruitment of Obg-like ATPase 1 (OLA1) and adenosine triphosphate (ATP) hydrolysis and glycolysis during CRC proliferation and progression. Specifically, IMP2 and ZFAS1 are significantly overexpressed with elevated m6A levels in CRC cells and paired CRC cohorts (n = 144). These indicators could be independent biomarkers for CRC prognostic prediction. Notably, IMP2 regulated ZFAS1 expression and enhanced CRC cell proliferation, colony formation, and apoptosis inhibition; thus, it was oncogenic. Mechanistically, ZFAS1 is modified at adenosine +843 within the RGGAC/RRACH element in an m6A-dependent manner. Thus, direct interaction between the KH3-4 domain of IMP2 and ZFAS1 where IMP2 serves as a reader for m6A-modified ZFAS1 and promotes the RNA stability of ZFAS1 is critical for CRC development. More importantly, stabilized ZFAS1 recognizes the OBG-type functional domain of OLA1, which facilitated the exposure of ATP-binding sites (NVGKST, 32-37), enhanced its protein activity, and ultimately accelerated ATP hydrolysis and the Warburg effect. CONCLUSIONS: Our findings reveal a new cancer-promoting mechanism, that is, the critical modulation network underlying m6A readers stabilizes lncRNAs, and they jointly promote mitochondrial energy metabolism in the pathogenesis of CRC.

Identification of Target PTEN-Based miR-425 and miR-576 as Potential Diagnostic and Immunotherapeutic Biomarkers of Colorectal Cancer With Liver Metastasis.

A major complication of colorectal cancer (CRC), one of the most common and fatal types of cancers, is secondary liver metastasis. For patients with this fate, there are very few biomarkers available in clinical application, and the disease remains incurable. Recently, increasing studies demonstrated that tumorigenesis and development are closely related to immune escape, indicating that the roles of immune-related indicators might have been neglected in the past in colorectal cancer liver metastases (CRLM). Here, we unveil that elevated miR-425 and miR-576 promote CRLM through inhibiting PTEN-mediated cellular immune function. Specifically, miR-425 and miR-576 were identified for their significant upregulation in CRLM compared with the primary CRC tissues based on GSE81581 (n = 8) and GSE44121 (n = 18) datasets. Besides, we determined that the two microRNAs (miRNAs) coparticipated in restraining P53 and transforming growth factor beta (TGF-ß) signaling pathways associated with tumor metastasis, and both shortened the overall survival of the patients with metastatic susceptibility. Notably, in situ hybridization on relatively large samples of paired CRC tissues (n = 157) not only substantiated that the expression of miR-425 and miR-576 was dramatically upregulated in CRLM but also revealed that they were closely related to tumor deterioration, especially liver metastases. Moreover, we further confirmed that the combination of miR-425 and miR-576 was an effective predictive model for liver metastases and poor clinical outcomes. Mechanically, downregulated PTEN (GSE81558, n = 6) was verified to be a shared target of miR-425 and miR-576 acting as metastasis-related oncogenes, on account of the presence of binding sites (+2928-+2934 and +4371-+4378, respectively) and the collaborative suppression of P53/TGF-ß signaling in CRLM, which was further confirmed in CRC cells (HCT116 and SW480) based on systematic molecular biology experiments. Importantly, the target PTEN was strongly associated with microsatellite instability, tumor microenvironment, and immune cell infiltration. Thus, we speculate that miR-425 and miR-576 are novel biomarkers for CRLM prevention and immunotherapy and upstream inhibitors of the PTEN-P53/TGF-ß function axis.

Analysis of DNA methylation-driven genes for predicting the prognosis of patients with colorectal cancer.

Aberrant promoter methylation and ensuing abnormal gene expression are important epigenetic mechanisms that contribute to colorectal oncogenesis. Yet, the prognostic significance of such methylation-driven genes in colorectal cancer (CRC) remains obscure. Herein, a total of 181 genes were identified as the methylation-driven molecular features of CRC by integrated analysis of the expression profiles and the matched DNA methylation data from The Cancer Genome Atlas (TCGA) database. Among them, a five-gene signature (POU4F1, NOVA1, MAGEA1, SLCO4C1, and IZUMO2) was developed as a risk assessment model for predicting the clinical outcomes in CRC. The Kaplan-Meier analysis and Harrell's C index demonstrated that the risk assessment model significantly distinguished the patients in high or low-risk groups (p-value < 0.0001 log-rank test, HR: 2.034, 95% CI: 1.419-2.916, C index: 0.655). The sensitivity and specificity were validated by the receiver operating characteristic (ROC) analysis. Furthermore, different pharmaceutical treatment responses were observed between the high-risk and low-risk groups. Indeed, the methylation-driven gene signature could act as an independent prognostic evaluation biomarker for assessing the OS of CRC patients and guiding the pharmaceutical treatment. Compared with known biomarkers, the methylation-driven gene signature could reveal cross-omics molecular features for improving clinical stratification and prognosis.

Colon cancer-specific diagnostic and prognostic biomarkers based on genome-wide abnormal DNA methylation.

Abnormal DNA methylation is a major early contributor to colon cancer (COAD) development. We conducted a cohort-based systematic investigation of genome-wide DNA methylation using 299 COAD and 38 normal tissue samples from TCGA. Through conditional screening and machine learning with a training cohort, we identified one hypomethylated and nine hypermethylated differentially methylated CpG sites as potential diagnostic biomarkers, and used them to construct a COAD-specific diagnostic model. Unlike previous models, our model precisely distinguished COAD from nine other cancer types (e.g., breast cancer and liver cancer; error rate ≤ 0.05) and from normal tissues in the training cohort (AUC = 1). The diagnostic model was verified using a validation cohort from The Cancer Genome Atlas (AUC = 1) and five independent cohorts from the Gene Expression Omnibus (AUC ≥ 0.951). Using Cox regression analyses, we established a prognostic model based on six CpG sites in the training cohort, and verified the model in the validation cohort. The prognostic model sensitively predicted patients' survival (p ≤ 0.00011, AUC ≥ 0.792) independently of important clinicopathological characteristics of COAD (e.g., gender and age). Thus, our DNA methylation analysis provided precise biomarkers and models for the early diagnosis and prognostic evaluation of COAD.

Long non-coding RNA ZFAS1 promotes colorectal cancer tumorigenesis and development through DDX21-POLR1B regulatory axis.

Increasing evidence supports long non-coding RNA-ZFAS1 as master protein regulators involved in a variety of human cancers. However, the molecular mechanism is not fully understood in colorectal cancer (CRC) and remains to be elucidated. Here, we uncovered a previously unreported mechanism linking RNA helicase DDX21 regulated by lncRNA ZFAS1 in control of POLR1B expression in CRC initiation and progression. Specifically, ZFAS1 exerted its oncogenic functions and was significantly up-regulated accompanied by elevated DDX21, POLR1B expression in CRC cells and tissues, which further closely associated with poor clinical outcomes. Notably, ZFAS1 knockdown dramatically suppressed CRC cell proliferation, invasion, migration, and increased cell apoptosis, which were contrary to the effect caused by ZFAS1 up-regulation. We further revealed that the inhibitory effect caused by ZFAS1 knockdown could be reversed by DDX21 overexpression in vitro and in vivo. Mechanistically, our research found that ZFAS1 could directly recruit DDX21 protein by harboring the specific motif (AAGA or CAGA). Finally, POLR1B was identified as the downstream target of DDX21 regulated by ZFAS1, which was also up-regulated in CRC cells and tissues and closely related to poor prognosis. The unrecognized ZFAS1/DDX21/POLR1B signaling regulation axis may provide new biomarkers and targets for CRC treatment and prognostic evaluation.

LNC473 Regulating APAF1 IRES-Dependent Translation via Competitive Sponging miR574 and miR15b: Implications in Colorectal Cancer.

A growing number of studies have focused on the involvement of non-coding RNAs (ncRNAs) in the internal ribosome entry site (IRES)-mediated translation in tumorigenesis; however, the underlying mechanisms in colorectal cancer (CRC) remain elusive. In this study, we show that LINC00473 (LNC473) exerted its functions as a tumor suppressor in promoting apoptotic protease-activating factor 1 (APAF1) IRES activity through competitively sponging miR574-5p and miR15b-5p in CRC initiation and pathogenesis. Specifically, LNC473 and its downstream target APAF1 were significantly downregulated accompanied by upregulated miR574-5p and miR15b-5p in CRC cells and tissues, which had a significant prognostic impact on clinical outcomes in our CRC cohort (n = 157). Furthermore, ectopic LNC473 significantly sponged endogenous miR574-5p or miR15b-5p and thereby inhibited cell proliferation and colony formation capacity, and it accelerated cell apoptosis through activating the APAF1-CASP9-CASP3 pathway. Notably, LNC473 overexpression resulted in dramatic promotion of APAF1 IRES activity and translation, whereas rescue experiments confirmed the recovery by the existence of LNC473 and miR574/15b-5p. Mechanistically, LNC473 overexpression promoted IRES binding domain exposure and removed the constraints controlling from miR574-5p and miR15b-5p, and subsequently enhanced IRES-mediated APAF1 expression in vitro and in vivo. Therefore, our results uncover a novel LNC473-miR574/miR15b-APAF1 signaling axis, which provides new targets and crosstalk regulation mechanism for CRC prevention and treatment.

LNC942 promoting METTL14-mediated m6A methylation in breast cancer cell proliferation and progression.

Increasing evidence supports that long noncoding RNAs (lncRNAs) act as master regulators involved in tumorigenesis and development at the N6-methyladenine (m6A) epigenetic modification level. However, the underlying regulatory mechanism in breast cancer (BRCA) remains elusive. Here, we unveil that LINC00942 (LNC942) exerts its functions as an oncogene in promoting METTL14-mediated m6A methylation and regulating the expression and stability of its target genes CXCR4 and CYP1B1 in BRCA initiation and progression. Specifically, LNC942 and METTL14 were significantly upregulated accompanied with the upregulation of m6A levels in BRCA cells and our included BRCA cohorts (n = 150). Functionally, LNC942 elicits potent oncogenic effects on promoting cell proliferation and colony formation and inhibiting cell apoptosis, subsequently elevating METTL14-mediated m6A methylation levels and its associated mRNA stability and protein expression of CXCR4 and CYP1B1 in BRCA cells. Mechanistically, LNC942 directly recruits METTL14 protein by harboring the specific recognize sequence (+176-+265), thereby stabilized the expression of downstream targets of LNC942 including CXCR4 and CYP1B1 through posttranscriptional m6A methylation modification in vitro and in vivo. Therefore, our results uncover a novel LNC942-METTL14-CXCR4/CYP1B1 signaling axis, which provides new targets and crosstalk m6A epigenetic modification mechanism for BRCA prevention and treatment.

Long noncoding RNA ZFAS1 promoting small nucleolar RNA-mediated 2'-O-methylation via NOP58 recruitment in colorectal cancer.

BACKGROUND: Increasing evidence supports the role of small nucleolar RNAs (snoRNAs) and long non-coding RNAs (lncRNAs) as master gene regulators at the epigenetic modification level. However, the underlying mechanism of these functional ncRNAs in colorectal cancer (CRC) has not been well investigated. METHODS: The dysregulated expression profiling of lncRNAs-snoRNAs-mRNAs and their correlations and co-expression enrichment were assessed by GeneChip microarray analysis. The candidate lncRNAs, snoRNAs, and target genes were detected by in situ hybridization (ISH), RT-PCR, qPCR and immunofluorescence (IF) assays. The biological functions of these factors were investigated using in vitro and in vivo studies that included CCK8, trans-well, cell apoptosis, IF assay, western blot method, and the xenograft mice models. rRNA 2'-O-methylation (Me) activities were determined by the RTL-P assay and a novel double-stranded primer based on the single-stranded toehold (DPBST) assay. The underlying molecular mechanisms were explored by bioinformatics and RNA stability, RNA fluorescence ISH, RNA pull-down and translation inhibition assays. RESULTS: To demonstrate the involvement of lncRNA and snoRNAs in 2'-O-Me modification during tumorigenesis, we uncovered a previously unreported mechanism linking the snoRNPs NOP58 regulated by ZFAS1 in control of SNORD12C, SNORD78 mediated rRNA 2'-O-Me activities in CRC initiation and development. Specifically, ZFAS1 exerts its oncogenic functions and significantly up-regulated accompanied by elevated NOP58, SNORD12C/78 expression in CRC cells and tissues. ZFAS1 knockdown suppressed CRC cell proliferation, migration, and increased cell apoptosis, and this inhibitory effect could be reversed by NOP58 overexpression in vitro and in vivo. Mechanistically, the NOP58 protein could be recognized by the specific motif (AAGA or CAGA) of ZFAS1. This event accelerates the assembly of SNORD12C/78 to allow for further guiding of 2'-O-Me at the corresponding Gm3878 and Gm4593 sites. Importantly, silencing SNORD12C or 78 reduced the rRNAs 2'-O-Me activities, which could be rescued by overexpression ZFAS1, and this subsequently inhibits the RNA stability and translation activity of their downstream targets (e.g., EIF4A3 and LAMC2). CONCLUSION: The novel ZFAS1-NOP58-SNORD12C/78-EIF4A3/LAMC2 signaling axis that functions in CRC tumorigenesis provides a better understanding regarding the role of lncRNA-snoRNP-mediated rRNAs 2'-O-Me activities for the prevention and treatment of CRC.

Prognostic evaluation of colorectal cancer using three new comprehensive indexes related to infection, anemia and coagulation derived from peripheral blood.

Background: Many indicators of peripheral blood in routine blood test (BRT) results of colorectal cancer (CRC) patients are related to prognosis. Currently, indexes such as NLR (Neutrophil-to- Lymphocyte Ratio), PLR (Platelet-to-Lymphocyte Ratio) and LMR (Lymphocyte-to-Monocyte ratio) evaluate the survival risk of patients by assessing the inflammatory - immune status of CRCs. These indexes are more comprehensive and accurate than independent estimates. We hope to design more effective indexes through fully considering the correlation and significance between BRT indicators and prognosis, so as to play a guiding role in clinical malignant estimation of CRCs. Methods: 701 CRCs in training set and 256 CRCs in test set were included in the study samples, and their clinical data, tumor pathology results and peripheral blood routine results were collected. The prognosis, progression, and survival status of all patients were determined after follow-up. Above data were used for statistical analysis and designing new indexes. Results: It was found that high NE, MONO, RDW-CV/SD and PLT in peripheral blood indicated poor prognosis of DFS and OS. Conversely, CRCs with postoperative tumor progression or death had lower LY, EO, RBC, HGB, HCT, MCV, MCH, MCHC, PDW, and P-LCR. IRR, ARR and CRR related to infection, anemia and coagulation were designed respectively using the largest AUC indicators (P<0.05) selected by ROC curve. The formula: IRR= (NE*MONO)/(LY*EO); ARR= (HGB*MCHC)/RDW-CV; CRR=PLT/PDW. Results of Kaplan­Meier survival analysis and multivariate COX proportional hazard analysis adjusted for age, gender, TNM stage, infiltration, adhesion showed IRR, ARR, CRR were all able to be used as the evaluation standard of survival of CRC. The result was also authenticated in the test set. Conclusion: We designed three different prognostic indexes of colorectal cancer, IRR, ARR and CRR, which could be used as risk indicators of CRC prognosis, tumor progression and survival.

Identification of functional circRNA/miRNA/mRNA regulatory network for exploring prospective therapy strategy of colorectal cancer.

Circular RNA (circRNA) has been reported to have great scientific significance and clinical value in multiple cancers including colorectal cancer (CRC). However, the biological function of most circRNAs in CRC is still in its infancy. Herein, we discovered the differential expressed circRNAs (DECs) between CRC tissues and matched adjacent using deep RNA sequencing and further confirmed the DECs expression by combining with another Gene Expression Omnibus dataset. Furthermore, we validated the expression of the top four upregulated circRNAs (hsa_circ_0030632, hsa_circ_0004887, hsa_circ_0001550, and hsa_circ_0001681) in both of paired CRC tissues and CRC cell lines. Then, a circRNA/microRNA/messenger RNA regulatory network was established and the Gene Ontology analysis and Kyoto Encyclopedia of Genes and Genomes pathway enrichment analysis showed these four circRNAs participated in various biological processed including apoptotic process and multiple metabolic processes. Moreover, based on the regulatory network, three bioactive compounds (pergolide, pivampicillin, and methylergometrine) for the treatment of CRC were also found. In conclusion, this study improved our understanding of circRNAs and may also facilitate the finding of promising targets and biomarkers in CRC.