Wemics: A Single-Base Resolution Methylation Quantification Method for Enhanced Prediction of Epigenetic Regulation.

DNA methylation, an epigenetic mechanism that alters gene expression without changing DNA sequence, is essential for organism development and key biological processes like genomic imprinting and X-chromosome inactivation. Despite tremendous efforts in DNA methylation research, accurate quantification of cytosine methylation remains a challenge. Here, a single-base methylation quantification approach is introduced by weighting methylation of consecutive CpG sites (Wemics) in genomic regions. Wemics quantification of DNA methylation better predicts its regulatory impact on gene transcription and identifies differentially methylated regions (DMRs) with more biological relevance. Most Wemics-quantified DMRs in lung cancer are epigenetically conserved and recurrently occurred in other primary cancers from The Cancer Genome Atlas (TCGA), and their aberrant alterations can serve as promising pan-cancer diagnostic markers. It is further revealed that these detected DMRs are enriched in transcription factor (TF) binding motifs, and methylation of these TF binding motifs and TF expression synergistically regulate target gene expression. Using Wemics on epigenomic-transcriptomic data from the large lung cancer cohort, a dozen novel genes with oncogenic potential are discovered that are upregulated by hypomethylation but overlooked by other quantification methods. These findings increase the understanding of the epigenetic mechanism by which DNA methylation regulates gene expression.

Linc00673-V3 positively regulates autophagy by promoting Smad3-mediated LC3B transcription in NSCLC.

Since its first discovery, long noncoding RNA Linc00673 has been linked to carcinogenesis and metastasis of various human cancers. Linc00673 had five transcriptional isoforms and their biological functions remained to be explored. Here we have reported that Linc00673-V3, one of the isoforms of Linc00673, promoted non-small cell lung cancer chemoresistance, and increased Linc00673-V3 expression level was associated with enhanced autophagy. Mechanistically, we discerned the existence of a stem-loop configuration engendered by the 1-100-nt and 2200-2275-nt fragments within Linc00673-V3. This structure inherently interacted with Smad3, thereby impeding its ubiquitination and subsequent degradation orchestrated by E3 ligase STUB1. The accumulation of Smad3 contributed to autophagy via up-regulation of LC3B transcription and ultimately conferred chemoresistance in NSCLC. Our results revealed a novel transcriptional regulation network between Linc00673-V3, Smad3, and LC3B, which provided an important insight into the interplay between autophagy regulation and non-canonical function of Smad3. Furthermore, the results from in vivo experiments suggested Linc00673-V3 targeted antisense oligonucleotide as a promising therapeutic strategy to overcome chemotherapy resistance in NSCLC.

Pan-cancer tRNA-derived fragment CAT1 coordinates RBPMS to stabilize NOTCH2 mRNA to promote tumorigenesis.

Transfer RNA-derived fragments (tRFs) are a class of small non-coding regulatory RNAs that are involved in the pathophysiology of many diseases. However, the role of tRFs in cancer progression remains largely elusive. Here, we demonstrate that a pan-cancer 3'-tRF, CAT1 (cancer associated tRF 1), is ubiquitously upregulated in tumors and associated with poor prognosis of a variety of cancers, including lung cancer. The upregulated CAT1 in cancer cells binds to RNA-binding protein with multiple splicing (RBPMS) and displaces NOTCH2 association from RBPMS, thereby inhibiting the subsequent CCR4-NOT deadenylation-complex-mediated NOTCH2 mRNA decay. The CAT1-enhanced NOTCH2 expression promotes lung cancer cell proliferation and metastasis in vitro and in vivo. In addition, plasma CAT1 levels are substantially increased in patients with lung cancer compared to non-cancer control subjects. Our findings reveal an intrinsic connection between cancer-specific upregulation of CAT1 and cancer progression, show the regulation of NOTCH signaling in cancer by a 3'-tRF, and highlight its great clinical potential.

Specific intracellular retention of circSKA3 promotes colorectal cancer metastasis by attenuating ubiquitination and degradation of SLUG.

Our previous study demonstrated that tumor-suppressor circular RNAs (circRNAs) can be specifically secreted outside of colorectal cancer (CRC) cells within exosomes to maintain tumor cell fitness. However, whether tumor-driving circRNAs can be specifically retained in cells to facilitate tumor progression remains unknown. In this study, circRNA-seq showed that circSKA3 was significantly upregulated in CRC tissues but downregulated in serum samples from CRC patients. In addition, circSKA3 promoted CRC progression in vitro and in vivo and was retained in CRC cells via a specific cellmotif element. Interestingly, the cellmotif element was also the site of interaction of circSKA3 with SLUG, which inhibited SLUG ubiquitination degradation and promoted CRC epithelial-mesenchymal transition (EMT). Moreover, FUS was identified as a key circularization regulator of circSKA3 that bound to the key element. Finally, we designed and synthesized specific antisense oligonucleotides (ASOs) targeting circularization and cellmotif elements, which repressed circSKA3 expression, abolished the SLUG-circSKA3 interaction, and further inhibited CRC EMT and metastasis in vitro and in vivo.

Uncertainty-aware refinement framework for ovarian tumor segmentation in CECT volume.

BACKGROUND: Ovarian cancer is a highly lethal gynecological disease. Accurate and automated segmentation of ovarian tumors in contrast-enhanced computed tomography (CECT) images is crucial in the radiotherapy treatment of ovarian cancer, enabling radiologists to evaluate cancer progression and develop timely therapeutic plans. However, automatic ovarian tumor segmentation is challenging due to factors such as inhomogeneous background, ambiguous tumor boundaries, and imbalanced foreground-background, all of which contribute to high predictive uncertainty for a segmentation model. PURPOSE: To tackle these challenges, we propose an uncertainty-aware refinement framework that aims to estimate and refine regions with high predictive uncertainty for accurate ovarian tumor segmentation in CECT images. METHODS: To this end, we first employ an approximate Bayesian network to detect coarse regions of interest (ROIs) of both ovarian tumors and uncertain regions. These ROIs allow a subsequent segmentation network to narrow down the search area for tumors and prioritize uncertain regions, resulting in precise segmentation of ovarian tumors. Meanwhile, the framework integrates two guidance modules that learn two implicit functions capable of mapping query features sampled according to their uncertainty to organ or boundary manifolds, guiding the segmentation network to facilitate information encoding of uncertain regions. RESULTS: Firstly, 367 CECT images are collected from the same hospital for experiments. Dice score, Jaccard, Recall, Positive predictive value (PPV), 95% Hausdorff distance (HD95) and Average symmetric surface distance (ASSD) for the testing group of 77 cases are 86.31%, 73.93%, 83.95%, 86.03%, 15.17  mm and 2.57  mm, all of which are significantly better than that of the other state-of-the-art models. And results of visual comparison shows that the compared methods have more mis-segmentation than our method. Furthermore, our method achieves a Dice score that is at least 20% higher than the Dice scores of other compared methods when tumor volumes are less than 20 cm3 , indicating better recognition ability to small regions by our method. And then, 38 CECT images are collected from another hospital to form an external testing group. Our approach consistently outperform the compared methods significantly, with the external testing group exhibiting substantial improvements across key evaluation metrics: Dice score (83.74%), Jaccard (69.55%), Recall (82.12%), PPV (81.61%), HD95 (12.31 mm), and ASSD (2.32 mm), robustly establishing its superior performance. CONCLUSIONS: Experimental results demonstrate that the framework significantly outperforms the compared state-of-the-art methods, with decreased under- or over-segmentation and better small tumor identification. It has the potential for clinical application.

Identification of novel protein biomarkers and drug targets for colorectal cancer by integrating human plasma proteome with genome.

BACKGROUND: The proteome is a major source of therapeutic targets. We conducted a proteome-wide Mendelian randomization (MR) study to identify candidate protein markers and therapeutic targets for colorectal cancer (CRC). METHODS: Protein quantitative trait loci (pQTLs) were derived from seven published genome-wide association studies (GWASs) on plasma proteome, and summary-level data were extracted for 4853 circulating protein markers. Genetic associations with CRC were obtained from a large-scale GWAS meta-analysis (16,871 cases and 26,328 controls), the FinnGen cohort (4957 cases and 304,197 controls), and the UK Biobank (9276 cases and 477,069 controls). Colocalization and summary-data-based MR (SMR) analyses were performed sequentially to verify the causal role of candidate proteins. Single cell-type expression analysis, protein-protein interaction (PPI), and druggability evaluation were further conducted to detect the specific cell type with enrichment expression and prioritize potential therapeutic targets. RESULTS: Collectively, genetically predicted levels of 13 proteins were associated with CRC risk. Elevated levels of two proteins (GREM1, CHRDL2) and decreased levels of 11 proteins were associated with an increased risk of CRC, among which four (GREM1, CLSTN3, CSF2RA, CD86) were prioritized with the most convincing evidence. These protein-coding genes are mainly expressed in tissue stem cells, epithelial cells, and monocytes in colon tumor tissue. Two interactive pairs of proteins (GREM1 and CHRDL2; MMP2 and TIMP2) were identified to be involved in osteoclast differentiation and tumorigenesis pathways; four proteins (POLR2F, CSF2RA, CD86, MMP2) have been targeted for drug development on autoimmune diseases and other cancers, with the potentials of being repurposed as therapeutic targets for CRC. CONCLUSIONS: This study identified several protein biomarkers to be associated with CRC risk and provided new insights into the etiology and promising targets for the development of screening biomarkers and therapeutic drugs for CRC.

AMER1 deficiency promotes the distant metastasis of colorectal cancer by inhibiting SLC7A11- and FTL-mediated ferroptosis.

The crosstalk between ferroptosis and cancer metastasis remains unclear. Here, we identify AMER1 as a key regulator of ferroptosis. AMER1 loss causes resistance to ferroptosis in colorectal cancer (CRC) cells. Interestingly, AMER1-deficient CRC cells preferentially form distant metastases, while AMER1-naive CRC cells mainly invade lymph nodes. Moreover, the ferroptosis inhibitor liproxstatin-1 effectively promotes hematogenous transfer of AMER1-naive cells. Mechanistically, AMER1 binds to SLC7A11 and ferritin light chain (FTL) and recruits ß-TrCP1/2, which degrade SLC7A11 and FTL by ubiquitination. Therefore, AMER1 deficiency increases cellular cystine levels but decreases the pool of labile free iron, thereby enhancing resistance to ferroptosis in CRC cells. Thus, AMER1 deficiency increases the survival of CRC cells in the blood under conditions of high oxidative stress and then promotes hematogenous metastasis of CRC. In conclusion, AMER1 mediates the crosstalk between ferroptosis and cancer metastasis, which provides a window of opportunity for treating metastatic colorectal cancer patients with AMER1 mutations.

Loss of GLTSCR1 causes congenital heart defects by regulating NPPA transcription.

Precise and specific spatiotemporal domains of gene expression regulation are critical for embryonic development. Recent studies have identified GLTSCR1 as a gene transcriptional elongation regulator in cancer research. However, the function of GLTSCR1, especially in embryonic development, remains poorly understood. Here, we found that GLTSCR1 was essential for cardiac development because Gltscr1 knockout (Gltscr1-/-) led to embryonic lethality in mice with severe congenital heart defects (CHDs). Ventricular septal defect and double outflow right ventricular were also observed in neural crest cells with conditional deletion of Gltscr1, which were associated with neonatal lethality in mice. Mechanistically, GLTSCR1 deletion promoted NPPA expression by coordinating the CHD risk G allele of rs56153133 in the NPPA enhancer and releasing the transcription factor ZNF740-binding site on the NPPA promoter. These findings demonstrated that GLTSCR1 acts as a candidate CHD-related gene.

WMDS.net: a network control framework for identifying key players in transcriptome programs.

MOTIVATION: Mammalian cells can be transcriptionally reprogramed to other cellular phenotypes. Controllability of such complex transitions in transcriptional networks underlying cellular phenotypes is an inherent biological characteristic. This network controllability can be interpreted by operating a few key regulators to guide the transcriptional program from one state to another. Finding the key regulators in the transcriptional program can provide key insights into the network state transition underlying cellular phenotypes. RESULTS: To address this challenge, here, we proposed to identify the key regulators in the transcriptional co-expression network as a minimum dominating set (MDS) of driver nodes that can fully control the network state transition. Based on the theory of structural controllability, we developed a weighted MDS network model (WMDS.net) to find the driver nodes of differential gene co-expression networks. The weight of WMDS.net integrates the degree of nodes in the network and the significance of gene co-expression difference between two physiological states into the measurement of node controllability of the transcriptional network. To confirm its validity, we applied WMDS.net to the discovery of cancer driver genes in RNA-seq datasets from The Cancer Genome Atlas. WMDS.net is powerful among various cancer datasets and outperformed the other top-tier tools with a better balance between precision and recall. AVAILABILITY AND IMPLEMENTATION: https://github.com/chaofen123/WMDS.net. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

LncRNA SFTA1P promotes cervical cancer progression by interaction with PTBP1 to facilitate TPM4 mRNA degradation.

Long non-coding RNAs (lncRNAs) play key roles in cancer development and progression. However, the biological function and clinical significance of most lncRNAs in cervical cancer remain elusive. In this study, we explore the function and mechanism of lncRNA surfactant associated 1 (SFTA1P) in cervical cancer. We firstly identified SFTA1P by analyzing the RNA sequencing data of cervical cancer from our previous study and from The Cancer Genome Atlas (TCGA). We then verified SFTA1P expression by qRT-PCR. The cell proliferation and migration capacity of SFTA1P was assessed by using CCK-8, colony formation, transwell and wound healing assays. RNA pull-down, RNA immunoprecipitation (RIP), RNA stability and western blot assays were used to reveal potential mechanisms. Athymic nude mice were used to evaluate tumorigenicity and metastasis in vivo. SFTA1P is upregulated in cervical tumor tissues and its high expression is associated with poor prognosis. Biologically, knockdown of SFTA1P inhibited the proliferation, migration, and invasion of cervical cancer cells in vitro, as well as tumorigenesis and metastasis in vivo. Mechanistically, SFTA1P was shown to interact with polypyrimidine tract binding protein 1 (PTBP1) to regulate the stability of tropomyosin 4 (TPM4) mRNA, thereby resulting in malignant cell phenotypes. TPM4 knockdown could attenuate the suppression of cell progression induced by either SFTA1P or PTBP1 knockdown. Our findings demonstrate that SFTA1P can promote tumor progression by mediating the degradation of TPM4 mRNA through its interaction with PTBP1 protein. This provides a potential therapeutic strategy to target the SFTA1P-PTBP1-TPM4 axis in cervical cancer.

LCAT1 is an oncogenic LncRNA by stabilizing the IGF2BP2-CDC6 axis.

Long non-coding RNAs (lncRNAs) is known to play vital roles in modulating tumorigenesis. We previously reported that LCAT1, a novel lncRNA, promotes the growth and metastasis of lung cancer cells both in vitro and in vivo. However, the underlying mechanism(s) of LCAT1 as an oncogenic regulator remains elusive. Here, we showed that LCAT1 physically interacts with and stabilizes IGF2BP2, an m6A reader protein, by preventing its degradation via autolysosomes. IGF2BP2 is overexpressed in lung cancer tissues, which is associated with poor survival of non-small cell lung cancer patients, suggesting its oncogenic role. Biologically, IGF2BP2 depletion inhibits growth and survival as well as the migration of lung cancer cells. Mechanistically, the LCAT1/IGF2BP2 complex increased the levels of CDC6, a key cell cycle regulator, by stabilizing its mRNA in an m6A-dependent manner. Like IGF2BP2, CDC6 is also overexpressed in lung cancer tissues with poor patient survival, and CDC6 knockdown has oncogenic inhibitory activity. Taken together, the LCAT1-IGF2BP2-CDC6 axis appears to play a vital role in promoting the growth and migration of lung cancer cells, and is a potential therapeutic target for lung cancer. Importantly, our finding also highlights a previously unknown critical role of LCAT1 in m6A-dependent gene regulation by preventing autolytic degradation of IGF2BP2.

SR Splicing Factors Promote Cancer via Multiple Regulatory Mechanisms.

Substantial emerging evidence supports that dysregulated RNA metabolism is associated with tumor initiation and development. Serine/Arginine-Rich proteins (SR) are a number of ultraconserved and structurally related proteins that contain a characteristic RS domain rich in arginine and serine residues. SR proteins perform a critical role in spliceosome assembling and conformational transformation, contributing to precise alternative RNA splicing. Moreover, SR proteins have been reported to participate in multiple other RNA-processing-related mechanisms than RNA splicing, such as genome stability, RNA export, and translation. The dysregulation of SR proteins has been reported to contribute to tumorigenesis through multiple mechanisms. Here we reviewed the different biological roles of SR proteins and strategies for functional rectification of SR proteins that may serve as potential therapeutic approaches for cancer.

ENO2 Promotes Colorectal Cancer Metastasis by Interacting with the LncRNA CYTOR and Activating YAP1-Induced EMT.

The glycolytic enzyme enolase 2 (ENO2) is dysregulated in many types of cancer. However, the roles and detailed molecular mechanism of ENO2 in colorectal cancer (CRC) metastasis remain unclear. Here, we performed a comprehensive analysis of ENO2 expression in 184 local CRC samples and samples from the TCGA and GEO databases and found that ENO2 upregulation in CRC samples was negatively associated with prognosis. By knocking down and overexpressing ENO2, we found that ENO2 promoted CRC cell migration and invasion, which is dependent on its interaction with the long noncoding RNA (lncRNA) CYTOR, but did not depend on glycolysis regulation. Furthermore, CYTOR mediated ENO2 binding to large tumor suppressor 1 (LATS1) and competitively inhibited the phosphorylation of Yes-associated protein 1 (YAP1), which ultimately triggered epithelial-mesenchymal transition (EMT). Collectively, these findings highlight the molecular mechanism of the ENO2-CYTOR interaction, and ENO2 could be considered a potential therapeutic target for CRC.

Long Noncoding RNA MIR4435-2HG Suppresses Colorectal Cancer Initiation and Progression By Reprogramming Neutrophils.

MIR4435-2HG, also known as LINC00978, has previously been described as an oncogenic long noncoding RNA (lncRNA). However, we show here that Mir4435-2hg depletion promoted colorectal tumorigenesis and progression in in vivo models of colitis-associated colorectal cancer, spontaneous intestinal adenomatous polyposis, and subcutaneous tumors. Alteration of MIR4435-2HG in colorectal cancer cells did not change the potential for cell proliferation, migration, or invasion in vitro. RNAscope assays showed that most MIR4435-2HG was located in the tumor stroma, which caused high expression of MIR4435-2HG in colorectal cancer tumor tissue. Transcriptome analysis of colorectal cancer tissues from wild-type and Mir4435-2hg-deficient mice revealed Mir4435-2hg as a tumor suppressor gene that regulated the immune microenvironment. Loss of Mir4435-2hg led to a decline in neutrophils and elevation of polymorphonuclear myeloid-derived suppressor cells (PMN-MDSC). In tissue-specific Mir4435-2hg knockout mice, we confirmed that Mir4435-2hg depletion in neutrophils, but not in intestinal epithelial cells, promoted colorectal cancer progression. Mechanistically, Mir4435-2hg depletion enhanced the immunosuppressive ability of PMN-MDSCs by disturbing their fatty acid metabolism. These findings suggest that MIR4435-2HG is a tumor-suppressing lncRNA whose deficiency could increase tumor-infiltrating PMN-MDSCs and enhance the immunosuppressive potential of PMN-MDSCs to promote colorectal cancer development. This provides a theoretical basis for further illustrating the pathogenesis of colorectal cancer and a potential antitumor immunotherapy target.

TP53 mutation is enriched in colorectal cancer liver metastasis in the context of polyclonal seeding.

Cancer metastasis accounts for the majority of cancer motility burden. For colorectal cancer (CRC), the liver is the most common site of distant metastasis. It is still little known that cancer genomic mutations, which are a cell-intrinsic and heritable property, are enriched in CRC liver metastasis. Here, we try to answer the question in the context of polyclonal seeding. In this study, we sequenced 18 pairs of colorectal cancer primary tumors and their matched liver metastasis samples. Together with public available sequencing data, we compared the mutations in 113 primary and metastasis pairs. The TP53 mutation variant allele frequency (VAF) was significantly increased in metastasis compared to the paired primary tumor, although most of the frequently observed mutations in liver metastasis foci were concordant with their matched CRC primary tumors. The results support late metastasis and polyclonal seeding. Consequently, we quantitatively compared the intratumor heterogeneity (ITH) between primary and metastasis tumors, and with the help of in silico metastasis simulation, we inferred that more than 10 cells take part in the CRC liver metastasis.

circFBXO7/miR-96-5p/MTSS1 axis is an important regulator in the Wnt signaling pathway in ovarian cancer.

BACKGROUND: CircRNAs are a novel class of evolutionarily conserved noncoding RNA molecules that form covalently closed continuous loop structures without 5' caps and 3' poly(A) tails. Accumulating evidence suggests that circRNAs play important regulatory roles in cancer and are promising biomarkers for cancer diagnosis and prognosis, as well as targets for cancer therapy. In this study, we identify and explore the role of a novel circRNA, circFBXO7, in ovarian cancer. METHODS: rRNA-depleted RNA-sequencing was performed to identify differentially expressed circRNAs between ovarian cancerous and normal tissues. qRT-PCR and single-molecule RNA in-situ hybridization was used to quantify circFBXO7 expression in tumor tissues. The association of circFBXO7 expression with patient prognosis was evaluated by Kaplan-Meier survival analysis. The biological function of circFBXO7 was also investigated using loss-of-function and gain-of-function assays in vivo and in vitro. Luciferase reporter and TOP/FOP-Flash reporter assays were then conducted together with RNA immunoprecipitation and western blot to assess the circFBXO7/miR-96-5p/MTSS1/Wnt/ß-catenin axis. RESULTS: circFBXO7 was downregulated in ovarian cancer which was associated with poor prognosis. Biologically, circFBXO7 overexpression significantly suppressed ovarian cancer cell proliferation, migration, and invasion in vitro, and inhibited tumor growth and metastasis in vivo, whereas its knockdown exerted an opposite role. Mechanistically, circFBXO7 functioned as a competing endogenous RNA for miR-96-5p to regulate the expression of MTSS1. Consequently, downregulation of MTSS1 led to excessive accumulation of ß-catenin and increased phosphorylation of GSK3ß, leading to the translocation of ß-catenin to the nucleus, thereby activating the Wnt/ß-catenin signaling pathway and ultimately promoting ovarian cancer progression. CONCLUSIONS: Our findings indicate that circFBXO7 acts as a bone fide tumor suppressor in ovarian cancer and that the circFBXO7/miR-96-5p/MTSS1 axis is an important regulator in the Wnt/ß-catenin signaling pathway which may provide a promising target for ovarian cancer therapy.

LncRNA CASC15, MiR-23b Cluster and SMAD3 form a Novel Positive Feedback Loop to promote Epithelial-Mesenchymal Transition and Metastasis in Ovarian Cancer.

Cancer Susceptibility Candidate 15 (CASC15), which is a newly identified long noncoding RNA crucial for epigenetic regulation in human tumors, was found to be associated with poor prognosis of the patients with ovarian cancer by utilizing The Cancer Genome Atlas and Gene Expression Omnibus database. Therefore, the purpose of this paper was to explore the functional role and latent molecular mechanism of CASC15 in the progression of ovarian cancer. In vitro and in vivo experiments validated CASC15 as an oncogenic lncRNA in ovarian cancer, which could enhance metastasis through TGF-ß-induced epithelial-mesenchymal transition progress. MiR-23b-3p and miR-24-3p, which are members of the miR-23b cluster, were identified to directly target CASC15 through luciferase assays. Further mechanistic investigations indicated that CASC15-mediated miR-23b-3p/miR-24-3p sequestration cooperatively upregulated SMAD3 expression, which, in turn, would permit increased CASC15 mRNA level as a transcription activation factor. This study first described a miR-23b-3p/miR-24-3p-mediated positive feedback loop between CASC15 and SMAD3, which may reflect the underlying molecular mechanism of CASC15's oncogenic function in ovarian cancer.

Tumor-suppressive circRHOBTB3 is excreted out of cells via exosome to sustain colorectal cancer cell fitness.

BACKGROUND & AIMS: To clarify the biological roles, circularization process and secretion pathway of circRHOBTB3 in colorectal cancer (CRC) progression. METHODS: We performed a comprehensive analysis of circRNA levels in serum exosomes from multiple types of cancer patients in public databases and verified the higher level of circRHOBTB3 in CRC sera versus healthy donors by RT-qPCR. Then, the function of circRHOBTB3 in CRC was investigated in vitro and in vivo. RNA-seq and RNA pull-down assays together with mass spectrometry identified the downstream signals and the binding proteins of circRHOBTB3. Finally, Antisense oligonucleotides (ASOs) were designed to target circularization and secretion elements of circRHOBTB3 for CRC therapy. RESULTS: circRHOBTB3 levels were increased in the sera but was downregulated in tissue samples in CRC, and the downregulation was associated with poor prognosis. Furthermore, circRHOBTB3 acts a tumor-suppressive circRNA by repressing metabolic pathways, intracellular ROS production in CRC. Several key elements were discovered to regulate circRHOBTB3 circularization and exosomal secretion. Moreover, SNF8 was identified that sorts circRHOBTB3 into exosomes. Interestingly, we found that CRC cells could actively secrete more circRHOBTB3 than normal cells. According to the sequence of regulatory elements for circularization and exosomal secretion, we designed and synthesized ASOs, which increased circRHOBTB3 expression and blocked circRHOBTB3 exosomal secretion. More importantly, ASOs could inhibit CRC growth and metastasis in vitro and in vivo. CONCLUSIONS: circRHOBTB3 plays a tumor-suppressive role in CRC and has to be excreted out of cells to sustain cancer cell fitness. ASOs targeting regulatory elements for circularization and exosomal secretion will become a novel antitumor strategy.

GLTSCR1 coordinates alternative splicing and transcription elongation of ZO1 to regulate colorectal cancer progression.

Alternative splicing (AS) and transcription elongation are vital biological processes, and their dysregulation causes multiple diseases, including tumors. However, the coregulatory mechanism of AS and transcription elongation in tumors remains unclear. This study demonstrates a novel AS pattern of tight junction protein 1 (ZO1) regulated by the RNA polymerase II elongation rate in colorectal cancer (CRC). Glioma tumor suppressor candidate region gene 1 (GLTSCR1) decreases the transcription elongation rate of ZO1 to provide a time window for binding of the splicing factor HuR to the specific motif in intron 22 of ZO1 and spliceosome recognition of the weak 3' and 5' splice sites in exon 23 to promote exon 23 inclusion. Since exon 23 inclusion in ZO1 suppresses migration and invasion of CRC cells, our findings suggest a novel potential therapeutic target for CRC.

The interaction between S100A2 and KPNA2 mediates NFYA nuclear import and is a novel therapeutic target for colorectal cancer metastasis.

Nucleocytoplasmic transport of proteins is disrupted and dysregulated in cancer cells. Nuclear pore complexes and cargo proteins are two main transportation regulators. However, the mechanism regulating nucleocytoplasmic transport in cancer remains elusive. Here, we identified a S100A2/KPNA2 cotransport complex that transports the tumor-associated transcription factor NFYA in colorectal cancer (CRC). Through the S100A2/KNPA2 complex, depending on its interaction with S100A2, NFYA is transported to the nucleus and inhibits the transcriptional activity of E-cadherin, which in turn promotes CRC metastasis. Targeting the S100A2/KPNA2 binding sites with the specific inhibitor delanzomib is a potential therapeutic approach for CRC.