Abstract
Long non-coding RNAs (lncRNAs) play key roles in human diseases, including cancer. Functional studies of the lncRNA HOTAIR (HOX transcript antisense RNA) provide compelling evidence for therapeutic targeting of HOTAIR in cancer, but targeting lncRNAs in vivo has proven to be difficult. In the current study, we describe a peptide nucleic acids (PNA)-based approach to block the ability of HOTAIR to interact with EZH2 and subsequently inhibit HOTAIR-EZH2 activity and resensitize resistant ovarian tumors to platinum. Treatment of HOTAIR-overexpressing ovarian and breast cancer cell lines with PNAs decreased invasion and increased chemotherapy sensitivity. Furthermore, the mechanism of action correlated with reduced nuclear factor-kappaB (NF-κB) activation and decreased expression of NF-κB target genes matrix metalloprotease 9 and interleukin 6. To deliver the anti-lncRNA to the acidic (pH approximately 6) tumor microenvironment, PNAs were conjugated to pH-low insertion peptide (pHLIP). Treatment of mice harboring platinum-resistant ovarian tumor xenografts with pHLIP-PNA constructs suppressed HOTAIR activity, reduced tumor formation and improved survival. This first report on pHLIP-PNA lncRNA targeting solid tumors in vivo suggests a novel cancer therapeutic approach.
Highlights
Non-protein coding RNAs make up the majority of the transcripts in the genome and both small ncRNAs (200 nucleotides) are considered to be functionally important
PNA4 was used as the control peptide nucleic acids (PNA), as it had no effect on HOTAIR-EZH2 interaction
As a positive correlation between tumor growth and the pro-inflammatory cytokine IL- 6 has been described[30] (Fig. 3C), and we recently demonstrated that HOTAIR upregulated both IL-6 and matrix metalloproteinase 9 (MMP-9) in ovarian cancer cells[9], it was of interest to examine the effect of PNA3 on IL-6 in vivo
Summary
Inhibiting HOTAIR and EZH2 alters platinum sensitivity and cell behaviors. Inhibiting either. No change in cell survival was observed between pHLIP-PNA3 and pHLIP-control under normal pH; in pH 6, pHLIP-PNA3 decreased (P < 0.05) survival (Fig. 4C), indicating HOTAIR targeting and altered CDDP-sensitivity under acidic conditions. As a positive correlation between tumor growth and the pro-inflammatory cytokine IL- 6 has been described[30] (Fig. 3C), and we recently demonstrated that HOTAIR upregulated both IL-6 and MMP-9 in ovarian cancer cells[9], it was of interest to examine the effect of PNA3 on IL-6 in vivo. The results demonstrate that pHLIP-PNA3-mediated HOTAIR inhibition reduces ovarian tumor levels of IL-6, MMP-9, and ALDH1A1, increases CDDP sensitivity and improves overall survival. No apparent histological changes (based on H&E staining) were seen (Supplementary Fig. S4H), but H&E slides of ovarian tumors from CDDP-treated mice exhibited fewer cells (Supplementary Fig. S4H), indicating cell death
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