Keele Research Repository

Introduction
Of the total human DNA only 1.2% o codes for proteins, with the rest only being transcribed into regulatory RNA, known as non-coding RNA. Long non-coding RNAs (lncRNAs) are non-coding RNAs that have no or limited protein-coding potential, are >200 nt in length and include- among other categories- long/large intergenic/intervening RNAs (lincRNAs), intronic lncRNAs, natural antisense transcripts (NATs) and pseudogenes. lncRNAs are key regulators of several cellular processes and have been associated with a variety of diseases, including cancer. Metformin (N, N-dimethyl biguanide) is an oral biguanide already in clinical use against diabetes and has also been suggested to decrease the incidence of various cancers, including neuroblastoma, the most common extracranial paediatric cancer arising at the sympathetic nervous system.

Material and methods
The present study aimed at assessing the differential expression of protein-coding and, primarily, non-protein-coding RNAs between control untreated SH-SY5Y neuroblastoma cells and cells resistant to 3 mM Metformin via a paired-end RNA sequencing approach.

Results and discussions
In the first data set (untreated cells compared to cells resistant to 3 mM Metformin), out of 13 741 genes measured, 7855 genes were found to be differentially expressed with 108 of them falling within the lncRNA category. Among these 33 comprise lincRNAs, 24 are NATs and 18 pseudogenes. In the second data set (untreated cells compared to cells treated with 20 mM Metformin) 13 481 genes were tested, of which 5652 showed perturbed expression. Among them, 86 belong to the lncRNA category, and in particular, 33 are lincRNAs, 19 are NATs and 13 are pseudogenes. Interestingly, 34 of the assessed lncRNAs display differential expression in both data sets. The results were obtained using a threshold of 0.05 for statistical significance (p-value) and a log fold change of expression with an absolute value of at least 0.6.Neuroblastoma cells’ response to Metformin, as well as the acquisition of resistance to the drug, trigger the differential expression of a great diversity of lncRNAs.

Conclusion
Given that Metformin is an appealing and promising therapeutic approach against neuroblastoma, these lncRNAs could, in turn, be used as molecular biomarkers towards better prediction, prognosis and diagnosis of the disease. Moreover, such an approach would be of interest as part of a combination therapy.