Circulating circTOLLIP serves as a diagnostic biomarker for liquid biopsy in non-small cell lung cancer.

BACKGROUND: Circular RNAs (CircRNAs) have been found to possess vital functions in tumorigenesis of various cancer types, including non-small cell lung cancer (NSCLC). The aim of this study was to identify and explore the diagnostic values of the newly found Toll interacting protein (TOLLIP)-derived circRNA (circTOLLIP) for liquid biopsy in NSCLC. METHODS: RNase R and actinomycin D assays were conducted to confirm the existence and stability of circTOLLIP. RT-qPCR was performed to identify the expression levels of circTOLLIP in NSCLC tumor tissues, whole blood, and cell lines. The diagnostic values were evaluated by receiver operating characteristic (ROC) curve analysis. RESULTS: CircTOLLIP was screened as a candidate biomarker and was found to be significantly down-regulated in both NSCLC tissues and cell lines. Interestingly, circulating circTOLLIP was also lower-expressed in the whole blood of patients with NSCLC compared to that of patients with benign lung disease and healthy controls. Importantly, the circulating circTOLLIP represented better diagnostic values in comparison to the traditional tumor markers (NSE, CYFR21-1, and CA72-4), and showed higher stability even though the whole blood was exposed to various tough conditions. CONCLUSIONS: Our findings indicate that circTOLLIP can be used as a non-invasive biomarker to distinguish early-stage NSCLC from benign lung diseases and from healthy controls, suggesting the potential application of circTOLLIP for liquid biopsy in NSCLC.

LncRNA DPP10-AS1 promotes malignant processes through epigenetically activating its cognate gene DPP10 and predicts poor prognosis in lung cancer patients.

OBJECTIVE: The purpose of this study was to explore the function and gene expression regulation of the newly identified lncRNA DPP10-AS1 in lung cancer, and its potential value as a prognostic biomarker. METHODS: qRT-PCR and Western blot were conducted to detect the expression of DDP10-AS1 and DPP10 in lung cancer cell lines and tissues. The effects of DDP10-AS1 on DPP10 expression, cell growth, invasion, apoptosis, and in vivo tumor growth were investigated in lung cancer cells by Western blot, rescue experiments, colony formation, flow cytometry, and xenograft animal experiments. RESULTS: The novel antisense lncRNA DPP10-AS1 was found to be highly expressed in cancer tissues (P < 0.0001), and its upregulation predicted poor prognosis in patients with lung cancer (P = 0.0025). Notably, DPP10-AS1 promoted lung cancer cell growth, colony formation, and cell cycle progression, and repressed apoptosis in lung cancer cells by upregulating DPP10 expression. Additionally, DPP10-AS1 facilitated lung tumor growth via upregulation of DPP10 protein in a xenograft mouse model. Importantly, DPP10-AS1 positively regulated DPP10 gene expression, and both were coordinately upregulated in lung cancer tissues. Mechanically, DPP10-AS1 was found to associate with DPP10 mRNA but did not enhance DPP10 mRNA stability. Hypomethylation of DPP10-AS1 and DPP10 contributed to their coordinate upregulation in lung cancer. CONCLUSIONS: These findings indicated that the upregulation of the antisense lncRNA DPP10-AS1 promotes lung cancer malignant processes and facilitates tumorigenesis by epigenetically regulating its cognate sense gene DPP10. DPP10-AS1 may serve as a candidate prognostic biomarker and a potential therapeutic target in lung cancer.

lncRNA JPX/miR-33a-5p/Twist1 axis regulates tumorigenesis and metastasis of lung cancer by activating Wnt/ß-catenin signaling.

BACKGROUND: MicroRNAs (miRNAs) and Twist1-induced epithelial-mesenchymal transition (EMT) in cancer cell dissemination are well established, but the involvement of long noncoding RNAs (lncRNAs) in Twist1-mediated signaling remains largely unknown. METHODS: RT-qPCR and western blotting were conducted to detect the expression levels of lncRNA JPX and Twist1 in lung cancer cell lines and tissues. The impact of JPX on Twist1 expression, cell growth, invasion, apoptosis, and in vivo tumor growth were investigated in lung cancer cells by western blotting, rescue experiments, colony formation assay, flow cytometry, and xenograft animal experiment. RESULTS: We observed that lncRNA JPX was upregulated in lung cancer metastatic tissues and was closely correlated with tumor size and an advanced stage. Functionally, JPX promoted lung cancer cell proliferation in vitro and facilitated lung tumor growth in vivo. Additionally, JPX upregulated Twist1 by competitively sponging miR-33a-5p and subsequently induced EMT and lung cancer cell invasion. Interestingly, JPX and Twist1 were coordinately upregulated in lung cancer tissues and cells. Mechanically, the JPX/miR-33a-5p/Twist1 axis participated in EMT progression by activating Wnt/ß-catenin signaling. CONCLUSIONS: These findings suggest that lncRNA JPX, a mediator of Twist1 signaling, could predispose lung cancer cells to metastasis and may serve as a potential target for targeted therapy.

CRL3-SPOP ubiquitin ligase complex suppresses the growth of diffuse large B-cell lymphoma by negatively regulating the MyD88/NF-κB signaling.

Recurrent oncogenic mutations of MyD88 have been identified in a variety of lymphoid malignancies. Gain-of-function mutations of MyD88 constitutively activate downstream NF-κB signaling pathways, resulting in increased cellular proliferation and survival. However, whether MyD88 activity can be aberrantly regulated in MyD88-wild-type lymphoid malignancies remains poorly understood. SPOP is an adaptor protein of CUL3-based E3 ubiquitin ligase complex and frequently mutated genes in prostate and endometrial cancers. In this study, we reveal that SPOP binds to and induces the nondegradative ubiquitination of MyD88 by recognizing an atypical SPOP-binding motif in MyD88. This modification blocks Myddosome assembly and downstream NF-κB activation. SPOP is mutated in a subset of lymphoid malignancies, including diffuse large B-cell lymphoma (DLBCL). Lymphoid malignancies-associated SPOP mutants exhibited impaired binding to MyD88 and suppression of NF-κB activation. The DLBCL-associated, SPOP-binding defective mutants of MyD88 escaped from SPOP-mediated ubiquitination, and their effect on NF-κB activation is stronger than that of wild-type MyD88. Moreover, SPOP suppresses DLBCL cell growth in vitro and tumor xenograft in vivo by inhibiting the MyD88/NF-κB signaling. Therefore, SPOP acts as a tumor suppressor in DLBCL. Mutations in the SPOP-MyD88 binding interface may disrupt the SPOP-MyD88 regulatory axis and promote aberrant MyD88/NF-κB activation and cell growth in DLCBL.

Liquid biopsy: Circulating exosomal long noncoding RNAs in cancer.

Despite many advances in diagnostics and multimodal treatment (surgery, radiotherapy, chemotherapy), cancer still remains one of the most important public health challenges worldwide because of the associated morbidity and mortality. Liquid biopsy has been developed to detect cancer at an early stage based on minimally invasive and serial body fluid tests with the advantage of following tumor evolution in real time. Circulating tumor cells (CTCs), circulating tumor DNA (ctDNA), circulating cell-free noncoding RNAs (cfRNAs) and circulating exosomes represent the major components of liquid biopsy analysis. Liquid biopsy already has been implemented in cancer management, and most studies thus far are mainly focused on CTCs and ctDNA. In fact, the circulating long noncoding RNAs (lncRNAs) in exosomes have been discovered and confirmed to be closely related to tumorigenesis, metastasis and therapy. Thus this review is mainly focused on the clinical potential of circulating exosomal lncRNAs as a source of liquid biopsy biomarkers in cancer diagnosis, prognosis, and response to treatment, offering novel insights into the precision medicine of oncology.

Circular RNAs Serve as Novel Biomarkers and Therapeutic Targets in Cancers.

Circular RNAs (circRNAs) are a class of non-coding RNAs (ncRNAs) that structurally form closed loops without 5'-end cap and 3'-end poly(A) tail unlike linear RNAs. CircRNAs are widely present in eukaryotic cells with the capabilities of structural stability, high abundance and cell- /tissue-specific expression. A growing body of researches suggest that the dysregulated circRNAs are intimately relevant to the occurrence and development of cancer. In this review, we mainly discuss the differentially expressed circRNAs in cancer tissues, plasma and exosomes, which makes it possible for clinicians to use certain circRNAs as novel biomarkers for cancer diagnosis and prognosis. In particular, we primarily focus on circRNAs as potential therapeutic targets, which will provide promising applications in cancer gene therapy.

A two-miRNA signature (miR-33a-5p and miR-128-3p) in whole blood as potential biomarker for early diagnosis of lung cancer.

MicroRNAs (MiRNAs) have been found to be dysregulated in lung cancer tissues compared to their matched paracancerous tissues. However, the roles of miRNAs in peripheral blood as potential biomarkers for early diagnosis of lung cancer remain poorly understood. Here we found that miR-33a-5p and miR-128-3p were down-regulated in lung cancer tissues and cell lines. The expression levels of miR-33a-5p and miR-128-3p in lung cancer tissues were significantly correlated to TNM stages. MiR-128-3p in lung cancer tissues was also remarkably related to smoking and tumor size. The relative expression levels of miR-33a-5p and miR-128-3p were positively correlated in lung cancer tissues. Notably, miR-33a-5p and miR-128-3p in whole blood of lung cancer patients or early-stage lung cancer patients (TNM stage I-II) were lowly expressed as compared with that in healthy controls. The receiver operating characteristic curve (ROC) analyses revealed higher area under the ROC curve (AUC) values and higher sensitivity/specificity of miR-33a-5p and miR-128-3p alone and in combination were superior to that of traditional tumor markers (CYFR21-1, NSE and CA72-4). Importantly, both miR-33a-5p and miR-128-3p in whole blood were highly stable even under different harsh conditions. The results demonstrate that tumor suppressor miR-33a-5p/miR-128-3p in whole blood can serve as novel biomarkers for the early detection of lung cancer.

Insights into the Noncoding RNA-encoded Peptides.

BACKGROUND: Considerable evidence has indicated that most noncoding RNA (ncRNA) transcripts act directly as functional RNAs rather than as encoded peptides. However, a recent study of ribosome occupancy reported that many large intergenic ncRNAs (lincRNAs) are bound by ribosomes, raising the possibility that they are translated into proteins. These lincRNAs contain either Internal Ribosomal Entry Sites (IRES) or short Open Reading Frames (sORFs), and other translation elements, which can be translated into peptides with physiological functions. CONCLUSION: This review discusses three major types of ncRNA-encoded peptides (ncPEPs). First, microRNA(miRNA)-encoded peptides (miPEPs) are translated from their primary transcripts of miRNA genes and can promote the development of plant roots. Second, a long ncRNA(lncRNA) containing sORF encodes a 90 residues long regulatory peptide. Upon the amino acid response, the lncRNA-encoded peptide reduces the activity of mammalian Target Rapamycin complex 1 (mTORC1) and promotes muscle regeneration. Third, a circular RNA (circRNA) of the Na+/Ca2+ exchanger gene 1 (NCX1) exon 2 transcript encodes a truncated NCX1 protein exhibiting the Na+/Ca2+ exchange activity. It is also of worth noting that the majority of ncRNAs is not translated. This review summarizes the current understanding of the translatability of ncRNAs and the functions of ncPEPs in cellular processes, which may provide novel insights into the roles of ncPEPs.

Circulating lncRNA XLOC_009167 serves as a diagnostic biomarker to predict lung cancer.

BACKGROUND: Long noncoding RNAs (lncRNAs) have been proven to be involved in carcinogenesis and to be released into peripheral blood. Our objective was to develop a circulating lncRNA as a novel biomarker to predict lung cancer. METHODS: We analyzed the lncRNA expression profile in lung cancer patients by lncRNA array and identified lncRNA XLOC_009167 as a circulating biomarker using qRT-PCR in whole blood of lung cancer patients. The diagnostic value of was analyzed by area under curve (AUC) and the receiver operating characteristic (ROC) test. RESULTS: LncRNA XLOC_009167 was screened as a candidate biomarker for lung cancer and was up-regulated in both lung cancer tissues and cell lines. Notably, lncRNA XLOC_009167 in whole blood of lung cancer patients was highly expressed as compared with that in healthy controls or in patients with pneumonia. The values of AUC of lung cancer vs. healthy controls, and that of lung cancer vs. pneumonia were 0.7398 (95%CI = 0.6493 to 0.8303) and 0.7005 (95%CI = 0.5771 to 0.8240), respectively. Intriguingly, the ROC showed lncRNA XLOC_009167 was a better diagnostic potential compared to the traditional biomarkers (CYFR21-1, NSE and CA72-4), and the circulating lncRNA XLOC_009167 was found to be stable in whole blood under different conditions. CONCLUSIONS: LncRNA XLOC_009167 could serve as a novel diagnostic biomarker to distinguish lung cancer from benign lung disease and healthy controls.

CRISPR/Cas9-mediated noncoding RNA editing in human cancers.

Cancer is characterized by multiple genetic and epigenetic alterations, including a higher prevalence of mutations of oncogenes and/or tumor suppressors. Mounting evidences have shown that noncoding RNAs (ncRNAs) are involved in the epigenetic regulation of cancer genes and their associated pathways. The clustered regularly interspaced short palindromic repeats (CRISPR)-associated nuclease 9 (CRISPR/Cas9) system, a revolutionary genome-editing technology, has shed light on ncRNA-based cancer therapy. Here, we briefly introduce the classifications and mechanisms of CRISPR/Cas9 system. Importantly, we mainly focused on the applications of CRISPR/Cas9 system as a molecular tool for ncRNA (microRNA, long noncoding RNA and circular RNA, etc.) editing in human cancers, and the novel techniques that are based on CRISPR/Cas9 system. Additionally, the off-target effects and the corresponding solutions as well as the challenges toward CRISPR/Cas9 were also evaluated and discussed. Long- and short-ncRNAs have been employed as targets in precision oncology, and CRISPR/Cas9-mediated ncRNA editing may provide an excellent way to cure cancer.

[Circulating long noncoding RNAs as biomarkers in tumor diagnosis].

Long noncoding RNAs (lncRNAs) are involved in vital life processes of gene expression, epigenetic regulation and X-chromosome inactivation. lncRNAs are also closely associated with tumor initiation and progression. Moreover, lncRNAs may enter human circulation system in the form of microvesicle or exosome, or in combination with RNA binding protein. Interestingly, the circulating lncRNAs are widely existed in body fluids, such as blood and urine. We review the origin of circulating lncRNAs, and the detection methods as potential biomarkers. We focus on the early diagnosis value of circulating lncRNAs as tumor biomarkers in lung, breast, gastric, liver, colorectal and prostate cancers. Compared with the traditional biomarkers, the circulating lncRNAs show the unique advantages and clinical values as novel biomarkers.

Pecanex functions as a competitive endogenous RNA of S-phase kinase associated protein 2 in lung cancer.

Investigating the RNA-RNA interactions involving in the initiation and progression of non-small cell lung cancer (NSCLC) may provide promising diagnostic and targeted therapeutic strategies. Here, we showed that pecanex (PCNX) positively regulates the mRNA and protein expressions of S-phase kinase associated protein 2 (Skp2) in miRNA- and 3' UTR-dependent manners. And miR-26, miR-182, miR-340 and miR-506 were verified as the common miRNAs shared by Skp2 and PCNX. Intriguingly, we initially uncovered that PCNX-3' UTR promotes cell growth, proliferation and cell cycle progression, and suppresses apoptosis of lung cancer cells, which is consistent with the oncogenic activity of Skp2-3' UTR. Consequently, PCNX was identified as a competitive endogenous RNA (ceRNA) of Skp2. Moreover, knockdown of PCNX inhibits EGF-induced Akt phosphorylation, which can be reversed by the silencing of Dicer. Finally, we further discovered that Skp2 and PCNX are coordinately upregulated in lung cancer tissues compared with the adjacent non-tumor tissues. Our study establishes for the first time the oncogenic property of PCNX-3' UTR and Skp2-3' UTR, and the PCNX-miRNA-Skp2 regulatory pattern, which may offer a molecular basis for the diagnosis and targeted therapy in NSCLC.