Identification of a methylomics-associated nomogram for predicting overall survival of stage I-II lung adenocarcinoma.

The aim of this paper was to identify DNA methylation based biomarkers for predicting overall survival (OS) of stage I-II lung adenocarcinoma (LUAD) patients. Methylation profile data of patients with stage I-II LUAD from The Cancer Genome Atlas (TCGA) database was used to determine methylation sites-based hallmark for stage I-II LUAD patients' OS. The patients were separated into training and validation datasets by using median risk score as cutoff. Univariate Cox, least absolute shrinkage and selection operator (LASSO) and multivariate Cox analyses were employed to develop a DNA methylation signature for OS of patients with stage I-II LUAD. As a result, an 11-DNA methylation signature was determined to be critically associated with the OS of patients with stage I-II LUAD. Analysis of receiver operating characteristics (ROC) suggested a high prognostic effectiveness of the 11-DNA methylation signature in patients with stage I-II LUAD (AUC at 1, 3, 5 years in training set were (0.849, 0.879, 0.831, respectively), validation set (0.742, 0.807, 0.904, respectively), entire TCGA dataset (0.747, 0.818, 0.870, respectively). Kaplan-Meier survival analyses exhibited that survival was significantly longer in the low-risk cohort compared to the high-risk cohort in the training dataset (P = 7e - 07), in the validation dataset (P = 1e - 08), and in the all-cohort dataset (P = 6e - 14). In addition, a nomogram was developed based on molecular factor (methylation risk score) as well as clinical factors (age and cancer status) (AUC at 1, 3, 5 years entire TCGA dataset were 0.770, 0.849, 0.979, respectively). The result verified that our methylomics-associated nomogram had a strong robustness for predicting stage I-II LUAD patients' OS. Furthermore, the nomogram combined clinical and molecular factors to determine an individualized probability of recurrence for patients with stage I-II LUAD, which stood for a major advance in the field of personalized medicine for pulmonary oncology. Collectively, we successfully identified a DNA methylation biomarker and a DNA methylation-based nomogram to predict the OS of patients with stage I-II LUAD.

Free-radical ring closure to conformationally locked α-L-carba-LNAs and synthesis of their oligos: nuclease stability, target RNA specificity, and elicitation of RNase H.

A new class of conformationally constrained nucleosides, α-L-ribo-carbocyclic LNA thymidine (α-L-carba-LNA-T, LNA is an abbreviation of locked nucleic acid) analogues and a novel "double-locked" α-L-ribo-configured tetracyclic thymidine (6,7'-methylene-bridged-α-L-carba-LNA-T) in which both the sugar puckering and glycosidic torsion are simultaneously constrained, have been synthesized through a key step involving 5-exo free-radical intramolecular cyclization. These α-L-carba-LNA analogues have been subsequently transformed to corresponding phosphoramidites and incorporated into isosequential antisense oligonucleotides (AONs), which have then been examined for the thermal denaturation of their duplexes, nuclease stability, and RNase H recruitment capabilities. Introduction of a single 6',7'-substituted α-L-carba-LNA-T modification in the AON strand of AON/RNA heteroduplex led to T(m) reduction by 2-3 °C as compared to the native heteroduplex, whereas the parent 2'-oxa-α-L-LNA-T modification at the identical position in the AON strand has been found to lead to an increase in the T(m) by 3-5 °C. This suggests that the 6' and 7' substitutions lead to much reduced thermal stability for the modified heteroduplex, especially the hydrophobic 7'-methyl on α-L-carba-LNA, which is located in the major groove of the duplex. All of the AONs incorporating 6',7'-substituted α-L-carba-LNA-T have, however, showed considerably improved nuclease stability toward 3'-exonuclease (SVPDE) and in human blood serum compared to the 2'-oxa-α-L-LNA-T incorporated AONs. The hybrid duplexes that are formed by 6',7'-substituted α-L-carba-LNA-T-modified AONs with complementary RNA have been found to recruit RNase H with higher efficiency than those of the ß-D-LNA-T or ß-D-carba-LNA-T-modified counterparts. These greatly improved nuclease resistances and efficient RNase H recruitment capabilities elevate the α-L-carba-LNA-modified nucleotides into a new class of locked nucleic acids for potential RNA targeting therapeutics.

miR-135 promotes proliferation and stemness of oesophageal squamous cell carcinoma by targeting RERG.

MicroRNA (miRNA) plays an important role in tumourigenesis and cancer development by regulating oncogenes or tumour suppressor that are implicated in cell cycle, cell mobility and even cell senescence. Due to the resistance to enzymes that could degrade nucleotides, miRNAs have been considered proper for diagnosis and prognosis evaluation of cancer. The present study was designed to investigate miRNA associated with ESCC and identified effective miRNAs, which could serve as biomarker or targets. We first performed miRNA profiling to identify a subset of dysregulated miRNAs in ESCC. miR-135, miR-451 and miR-186 were the most differentially expressed miRNAs. Subsequent RT-PCR validated that miR-135 was upregulated in ESCC cell lines TE2 and TE9, implying the promise as a prognostic and diagnostic marker. The Cox regression analysis suggests the correlation of miR-135 expression and tumour stages. Survival analysis demonstrated metastatic samples largely have higher miR-135 expression. Downregulation of miR-135 suppressed proliferation and invasion of TE2 and TE9 cell lines. Subsequent target prediction combined with functional enrichment analysis identified "Small GTPase superfamily" that are possibly targeted by miR-135, which offers candidates for further investigation. Finally, RERG was identified as a target of miR-135. Downregulation of RERG could inhibit the cell proliferation and sphere formation ability of TE2 and TE9. Taken together, miR-135 was proved to promote tumour development of ESCC, which promises the prospect of using miR-135 as a biomarker indicator in diagnosis and prognosis.