Down-modulation of cancer targets using locked nucleic acid (LNA)-based antisense oligonucleotides without transfection

Abstract

Usually, small interfering RNAs and most antisense molecules need mechanical or chemical delivery methods to down-modulate the targeted mRNA. However, these delivery approaches complicate the interpretations of biological consequences. We show that locked nucleic acid (LNA)-based antisense oligonucleotides (LNA–ONs) readily down-modulate genes of interest in multiple cell lines without any delivery means. The down-modulation of genes was quick, robust, long-lasting and specific followed by potent down-modulation of protein. The efficiency of the effect varied among the 30 tumor cell lines investigated. The most robust effects were found in those cells where nuclear localization of the LNA–ON was clearly observed. Importantly, without using any delivery agent, we demonstrated that HER3 mRNA and protein could be efficiently down-modulated in cells and a tumor xenograft model. These data provide a simple and efficient approach to identify potential drug targets and animal models. Further elucidation of the mechanism of cellular uptake and trafficking of LNA–ONs may enhance not only the therapeutic values of this platform but also antisense molecules in general.

Downregulation of HER3 by a Novel Antisense Oligonucleotide, EZN-3920, Improves the Antitumor Activity of EGFR and HER2 Tyrosine Kinase Inhibitors in Animal Models

“…The data were consistent with the function of HER3 in cancer cells. Recently, we also showed that EZN-3920 was effective in downmodulating mRNA and HER3 protein in the absence of lipofection (30). The effect was specific as EZN-3920 did not downmodulate EGFR or HER2 (see below).…”

“…Beyond that, target inhibition in tumors derived from 15PC3 cells after i.v. administration of EZN-3920 was detected (30). In addition, the compound downmodulated HER3 and inhibited tumor growth in a mouse model of mammary carcinoma driven by the polyomavirus middle T oncogene (21).…”

“…S4). Moreover, as heterodimerization of HER2 with HER3 activates PI3K/AKT pathway, we next tested the combination of EZN-3920 with a PI3KCA (PI3K constitutively active)-specific antisense molecule EZN-4150 (30). The combination of these 2 LNAs showed greater antitumor activity than either agent administered alone (Fig.…”

6BIO Enhances Oligonucleotide Activity in Cells: A Potential Combinatorial Anti-androgen Receptor Therapy in Prostate Cancer Cells

“…The value of the CI ranged between 0.37 and 0.30 , and HER3 (data not shown). 16,37,38 The effects of the combination of 6BIO and the ASO on the targeting on b-catenin were of particular interest in view of the potential capability of 6BIO to activate the Wnt/b-catenin pathway 39 through the suppression of glycogen synthase kinase (GSK)-3b, a b-catenin inhibitor. 39 Nevertheless, the treatment of LNCaP cells with 6BIO and a b-catenin ASO (b-Cat-ASO+ 6BIO) still decreased the b-catenin mRNA expression by approximately 90% compared with a 65% reduction with ASO alone (Figure 4B, compare b-Cat-ASO+6BIO with b-Cat-ASO; n = 3; p < 0.001).…”

Reduced Expression of the Androgen Receptor by Third Generation of Antisense Shows Antitumor Activity in Models of Prostate Cancer

“…In particular, LNA-ASOs have shown very high binding affinity to mRNA, excellent potency for target mRNA downmodulation, improved resistance to nuclease digestion, and excellent stability in plasma and tissues in preclinical studies (10). These features allow LNAASOs simplistically prepared in saline to be highly effective in vitro and in vivo (11,12). The LNA technology has been used to design antisense molecules to hypoxiainducible factor-1a (HIF-1a; ref.…”