A patient-derived organoid-based study identified an ASO targeting SNORD14E for endometrial cancer through reducing aberrant FOXM1 Expression and ß-catenin nuclear accumulation.

BACKGROUND: Most of the endometrial cancer (EC) patients are diagnosis in early stage with a good prognosis while the patients with locally advanced recurrent or metastatic result in a poor prognosis. Adjuvant therapy could benefit the prognosis of patients with high-risk factors. Unfortunately, the molecular classification of great prognostic value has not yet reached an agreement and need to be further refined. The present study aims to identify new targets that have prognostic value in EC based on the method of EC patient-derived organ-like organs (PDOs), and further investigate their efficacy and mechanism. METHODS: The Cancer Genome Atlas (TCGA) database was used to determine SNORD14E expression. The effects of SNORD14E were investigated using CCK8, Transwell, wound-healing assays, and a xenograft model experiment; apoptosis was measured by flow cytometry. Antisense oligonucleotide (ASO) targeting SNORD14E was designed and patient-derived organoids (PDO) models in EC patients was established. A xenograft mouse and PDO model were employed to evaluate the effects of ASO targeting SNORD14E. RNA-seq, Nm-seq, and RNA immunoprecipitation (RIP) experiments were employed to confirm the alternative splicing (AS) and modification induced by SNORD14E. A minigene reporter gene assay was conducted to confirm AS and splicing factors on a variable exon. Actinomycin-d (Act-D) and Reverse Transcription at Low deoxy-ribonucleoside triphosphate concentrations followed by PCR (RTL-P) were utilized to confirm the effects of 2'-O methylation modification on FOXM1. RESULTS: We found that SNORD14E was overexpressed in EC tissues and patients with high expressed SNORD14E were distributed in the TCGA biomolecular classification subgroups without difference. Further, SNORD14E could reduce disease-free survival (DFS) and recurrence free survival (RFS) of EC patients. SNORD14E promoted proliferation, migration, and invasion and inhibited the apoptosis of EC cells in vitro. ASOs targeting SNORD14E inhibited cell proliferation, migration, invasion while promoted cell apoptosis. ASOs targeting SNORD14E inhibited tumor growth in the xenograft mouse model. TCGA-UCEC database showed that the proportion of patients with high expression of SNORD14E in middle-high risk and high-risk patients recommended by EMSO-ESGO-ESTRO guidelines for adjuvant therapy is more than 50%. Next, we enrolled 8 cases of high-risk and high-risk EC patients according to EMSO-ESGO-ESTRO guidelines and successfully constructed EC-PDOs. ASOs targeting SNORD14E inhibited the EC-PDO growth. Mechanistically, SNORD14E could recognize the mRNA of FOXM1 and recruit SRSF1 to promote the shearing of the variable exon VIIa of FOXM1, resulting in the overexpression of the FOXM1 malignant subtypes FOXM1b and FOXM1c. In addition, SNORD14E modified FOXM1 mRNA with 2`-O-methylation, which prolonged the half-life of FOXM1 mRNA. The nucleus accumulation of ß-catenin caused by aberrant FOXM1 expression led to EC progression. CONCLUSIONS: ASO targeting SNORD14E can be an effective treatment for EC.

SNORA73B promotes endometrial cancer progression through targeting MIB1 and regulating host gene RCC1 alternative splicing.

Endometrial cancer (EC) is a common gynaecological malignant tumour with unclear pathogenesis. Small nucleolar RNA (snoRNA) is involved in many biological processes, including those of cancers. Using the Cancer Genome Atlas (TCGA) database, the expression pattern of a snoRNA, SNORA73B, was analysed. The biological functions of SNORA73B were assessed by in vitro proliferation, apoptosis, migration, and invasion assays and in vivo by the xenograft model. RNA sequencing (RNA-seq) and RNA immunoprecipitation assays were performed to determine the relationship between SNORA73B and its target genes. High-performance liquid chromatography (HPLC) was performed to detect the pseudouridine content of the mindbomb E3 ubiquitin protein ligase 1 gene (MIB1). The stability of MIB1 mRNA was evaluated using a transcription inhibitor, actinomycin D. By performing co-immunoprecipitation assays, the change in the ubiquitin levels of the Jagged canonical Notch ligand 1 (Jag 1), caused by SNORA73B and MIB1, was identified. RNA-seq and qRT-PCR were performed to detect the alternative splicing of the regulator of the chromosome condensation 1 gene (RCC1). The TCGA database analysis showed that SNORA73B was highly expressed in EC. SNORA73B promoted cell proliferation, migration, and invasion and inhibited apoptosis. SNORA73B modified the pseudouridine content in MIB1 and increased the stability of MIB1 mRNA and protein; thus, it affected Jag 1 ubiquitination and further activated the Notch pathway. SNORA73B also affected the alternative splicing of RCC1, increasing the number of transcripts, RCC1-T2 and RCC1-T3, which promoted cell proliferation, migration, and invasion. SNORA73B can be a potential target for EC.

Small Nucleolar RNA and C/D Box 15B Regulate the TRIM25/P53 Complex to Promote the Development of Endometrial Cancer.

Background: Endometrial cancer is associated with a high mortality rate, which warrants the identification of novel diagnostic markers and therapeutic targets. The aim of this study is to evaluate the role of SNORD15B in the development of endometrial cancer and explore the potential underlying mechanisms. Methods: Bioinformatics was used to analyze the expression level and prognostic relevance of SNORD15B in endometrial cancer. The Ishikawa and HEC-1B cells were respectively transfected with SNORD15B expression plasmid and an antisense oligonucleotide, or the corresponding empty vector and a nonspecific sequence. The malignant phenotype of the suitably transfected cells was assessed by standard in vitro functional assays and the establishment of in vivo xenografts. The expression levels of the specific markers were analyzed with RT-qPCR and western blotting. The subcellular localization of P53 was determined by analyzing the nuclear and cytoplasmic fractions. RIP, Co-IP, and immunohistochemistry were performed as per standard protocols. Results: SNORD15B was overexpressed in the endometrial cancer tissues and correlated to a poor prognosis. Ectopic expression of SNORD15B in Ishikawa cells inhibited apoptosis, increased the proliferation, invasion, and migration in vitro, and enhanced their tumorigenicity in vivo. SNORD15B overexpression also upregulated TRIM25 and accelerated P53 accumulation in the cytoplasm of the endometrial cancer cells. Conclusion: SNORD15B functions as an oncogene in endometrial cancer by targeting the TRIM25/P53 complex and blocking the nuclear translocation of P53.

Circ-NOLC1 promotes epithelial ovarian cancer tumorigenesis and progression by binding ESRP1 and modulating CDK1 and RhoA expression.

Circular RNAs (circRNAs) play important roles in cancer tumorigenesis and progression, representing prognostic biomarkers and therapeutic targets. In this case, we demonstrated the role of circ-NOLC1 in epithelial ovarian cancer (EOC). Our results have shown that Circ-NOLC1 expression was higher in EOC tissues than in normal tissues, and was positively associated with FIGO stage, differentiation. Among ovarian cancer cell lines, circ-NOLC1 expression was the highest in A2780, and lowest in CAOV3. Overexpression of circ-NOLC1 in CAOV3 cells increased cell proliferation, migration, and invasion ability, whereas silencing of circ-NOLC1 in A2780 cells had the opposite effect: however, neither circ-NOLC1 downregulation nor overexpression influenced NOLC1 mRNA expression. In nude mice with subcutaneous tumors, circ-NOLC1 downregulation decreased tumor growth. Bioinformatic analysis and RNA-binding protein immunoprecipitation showed that circ-NOLC1 could bind to ESRP1. In addition, the overexpression of circ-NOLC1 significantly increased ESRP1, RhoA, and CDK1 protein and mRNA expression level; circ-NOLC1 downregulation had the opposite effects. The tumor-promoting effect of circ-NOLC1 was inhibited by knockdown of ESRP1, CDK1, or RhoA expression in circ-NOLC1-overexpressing cells, which might act by modulating RhoA and CDK1 expression. In conclusion, our study demonstrated that Circ-NOLC1 might promote EOC tumorigenesis and development by binding ESRP1 and modulating CDK1 and RhoA expression.