Published online Dec 27, 2020. doi: 10.4254/wjh.v12.i12.1211
Peer-review started: June 30, 2020
First decision: September 14, 2020
Revised: September 30, 2020
Accepted: October 29, 2020
Article in press: October 29, 2020
Published online: December 27, 2020
Hepatocellular carcinoma (HCC) develops in an inflammatory milieu containing tumor infiltrating lymphocytes, thus boosting tumor immunogenicity and provides an aspect for developing immunotherapies against HCC. However, immunotherapies have a modest response in HCC, accordingly combinatorial therapies with epigenetic immunomodulation may be a promising modality. Growing scientific evidence has suggested a modulatory role for miRNAs and long non-coding ribonucleic acids (lncRNAs) on programmed cell-death protein 1 (PD-1)/programmed death ligand 1 (PD-L1) immune checkpoint in HCC.
HCC is considered a therapy-resistant disease, and is frequently diagnosed at an advanced stage. Thus, the development of a novel therapeutic modality is essential. It is noteworthy that immune checkpoint blockade therapy in HCC is gaining attention. Additionally, given the wide range of non-coding RNAs (ncRNAs) that orchestrate PD-1/PD-L1 immune checkpoint, we investigated how selected ncRNAs regulate PD-1/PD-L1 immune checkpoint. Hence, the therapeutic potential of combining epigenetic immunomodulation through ncRNAs with immune checkpoint blockade was studied.
This study aimed at exploring potential upstream regulatory ncRNAs of immune checkpoint PD-1/PD-L1. Hence, the potential of combining immune checkpoint blockade with epigenetic immunomodulation was investigated.
Based on bioinformatics software and the literature, ncRNAs including miR-155-5p and miR-194-5p as well as lncRNAs X-inactive specific transcript (XIST) and MALAT-1 were selected. 23 HCC tissue biopsies and 10 healthy donor tissue biopsies were used to screen the expression of PD-L1 as well as lncRNAs XIST and MALAT-1. To study the interaction between miR-155-5p, miR-194-5p, lncRNAs XIST and MALAT-1, as well as PD-L1 mRNA, a series of transfections and co-transfections of the Huh-7 cell line was carried out. Quantified real-time polymerase chain reaction was then utilized to study the abundance of selected ncRNAs as well as PD-L1 transcripts in Huh-7 cells in the transfections experiments.
Based on bioinformatics software and the literature, we hypothesized that a potential upstream regulatory pathway to immune checkpoint PD-L1 is present in HCC, composed of both miRNAs, tumor suppressor miR-194-5p and oncomiR-155-5p, as well as both lncRNAs XIST and MALAT-1. Following the screening of 23 HCC biopsies, PD-L1 and XIST were found to be significantly upregulated compared to healthy controls; however, MALAT-1 was barely detected. Induced expression of miR-194-5p and miR-155-5p in the Huh-7 cell line showed the same pattern of upregulation of both PD-L1 transcript and XIST. However, ectopic expression of the respective miRNAs had a paradoxical impact on MALAT-1 abundance, i.e. miR-194-5p induced the upregulation of MALAT-1 while miR-155-5p downregulated the abundance of MALAT-1. Knockdown of XIST had no impact on PD-L1 expression; however, following knockdown of the negative regulator of X-inactive specific transcript (TSIX), PD-L1 expression was elevated, and MALAT-1 activity was abolished. Upon co-transfection of miR-194-5p with siMALAT-1, PD-L1 expression was elevated. On the other hand, co-transfection of miR-194-5p with siXIST did not have an impact on PD-L1 expression. Following co-transfection of miR-194 with siTSIX, PD-L1 expression was upregulated. Interestingly, the same PD-L1 expression pattern was revealed following the oncomiR-155-5p co-transfection series.
In conclusion, this study reported the controversial role of miR-194-5p in HCC and despite its paradoxical function of a tumour suppressor and having oncogenic activity in HCC, both had the same impact on upregulation of XIST. LncXIST is thought to be an intermediate player whose upregulation increased PD-L1 transcript abundance.
Although further investigations are needed, this study proposes a novel competing endogenous RNA circuit made up of both miR-155-5p and miR-194-5p as well as lncXIST and PD-L1 mRNA. This circuit could be regarded as a potential therapeutic target in HCC.