Detection of a long non-coding RNA (CCAT1) in living cells and human adenocarcinoma of colon tissues using FIT–PNA molecular beacons

Abstract

Although the function and mechanism of action of long non-coding RNAs (lncRNA) is still not completely known, studies have shown their potential role in the control of gene expression and regulation, in cellular proliferation and invasiveness at the transcriptional level via multiple mechanisms. Recently, colon cancer associated transcript 1 (CCAT1) lncRNA was found to be expressed in colorectal cancer (CRC) tumors but not in normal tissue. This study aimed to study the ability of a CCAT1-specific peptide nucleic acid (PNA) based molecular beacons (TO-PNA-MB) to serve as a diagnostic probe for in vitro, ex vivo, and in situ (human colon biopsies) detection of CRC. The data showed enhanced fluorescence upon in vitro hybridization to RNA extracted from CCAT1 expressing cells (HT-29, SW-480) compared to control cells (SK-Mel-2). Uptake of TO-PNA-MBs into cells was achieved by covalently attaching cell penetrating peptides (CPPs) to the TO-PNA-MB probes. In situ hybridization of selected TO-PNA-MB in human CRC specimens was shown to detect CCAT1 expression in all (4/4) subjects with pre-cancerous adenomas, and in all (8/8) patients with invasive adenocarcinoma (penetrating the bowel wall) tumors. The results showed that CCAT1 TO-PNA-MB is a powerful diagnostic tool for the specific identification of CRC, suggesting that with the aid of an appropriate pharmaceutical vehicle, real time in vivo imaging is feasible. TO-PNA-MB may enable identifying occult metastatic disease during surgery, or differentiating in real time in vivo imaging, between benign and malignant lesions.

Overexpression of long non-coding RNA-CTD903 inhibits colorectal cancer invasion and migration by repressing Wnt/β-catenin signaling and predicts favorable prognosis

“…LncRNA-H19 can increase cell invasion and metastasis in bladder cancer through associating with EZH2 and inhibiting E-cadherin (19), while LncRNA-EBIC is found to promote tumor invasion by similar mechanism in cervical cancer (18). Aberrant expressed lncRNAs also unravelled their importance in CRC (20)(21)(22)(23)(24)(25). However, most of lncRNA studies focused on correlations of clinicopathological characteristics and prognosis, the mechanisms are rarely clarified, especially for mCRC.…”

LncRNAs: Emerging Biomarkers in Gastric Cancer

“…Recently, a large quantity of lncRNAs have been identified and studies on the biological functions of lncRNAs on various kind of human cancers have been exponentially growing [96,97]. Although many key questions about lncRNAs remain unsolved, exploration on lncRNAs field gives us a better understanding on cancer diagnosis and treatment.…”

Long noncoding RNAs: Novel players in colorectal cancer

“…Accordingly, a plasmid composed of the H19 gene regulatory element that drives the expression of diphtheria toxin (DT-A) gene has been developed and it is undergoing clinical testing as a treatment for CRC and other cancers [103,104]. Moreover, Kam and colleagues [105] showed that a well designed peptide nucleic acid (PNA)-molecular beacon (PNA-MB) complementary to lncRNA-CCAT1 can be used as a tool for imaging cell lines and detecting malignancies in human CRC biopsies in situ, indicating the potential diagnostic application of lncRNAs in CRC. Taken together, although the research of lncRNAs is presently at its infancy and only a small number of lncRNAs have been characterized, dysregulated lncRNAs in CRC could still become an important factor for both the diagnosis and prognosis of CRC.…”