Abstract 602: Silencing of the androgen receptor (AR) with a novel mRNA antisense oligonucleotide causes antitumor effects in xenograft models of prostate cancer

Background: The AR is a ligand-activated transcriptional factor that plays an important role in prostate cancer. While androgen-deprivation therapies typically inhibit tumor growth, tumor recurrence due to the castration resistant form of the disease frequently occurs and leads to mortality. Nevertheless, castration-resistant tumors are still dependent on a functional (AR). Therefore, inhibition of AR expression, rather than systemic androgen deprivation, may provide a novel strategy for treatment of advanced prostate cancer. We have developed a novel locked nucleic acid (LNA)-based antisense oligonucleotide (ON), EZN-4176, which down-modulated AR and inhibited tumor growth. Methods: The target mRNA down-modulation, growth inhibition, luciferase activity, protein inhibition, and reduction of prostate specific antigen (PSA; an AR downstream target and biomarker of prostate cancer) were evaluated by qRT-PCR, MTS, SteadyGlo, western blot analysis, and ELISA assay, respectively, in LNCaP cancer cell lines. A mismatched LNA antisense oligo (EZN-4176MM) and an AR negative cell line (PC-3 prostate cancer) served as controls. Additionally, the effect of EZN-4176 on DHT-induced AR transcriptional activity was evaluated in LNCaP cells containing an AR-responsive luciferase reporter. In vivo, therapeutic efficacy of EZN-4176 was evaluated in the AR-positive CWR22 (androgen dependent). Results: In vitro, in the absence of any transfection reagent, EZN-4176 but not the mismatched control (EZN-4176MM) inhibited the growth of DHT-induced LNCaP cells. The growth reduction correlated with AR down-modulation and its down-stream target gene expression and AR-responsive luciferase activity. In vivo, EZN-4176, prepared in the absence of a delivery vehicle, demonstrated dose-dependent inhibition of mRNA of AR, PSA, and TMPRSS2 as well as protein level of AR in androgen-sensitive CWR22 tumor xenografts, resulting in significant tumor growth inhibition (TGI) (∼55%) (similar to high-dose Casodex ® [bicalutamide] treatment). Cy5.5-labeled EZN-4176 was shown to localize in subcutaneous CWR22 or LNCaP tumor xenografts using xenogen imaging system. In castration-resistant tumor cells, EZN-4176 potently and specifically inhibited the growth of C4-2b cells as well as down-regulated AR-luciferase reporter systems. More importantly, AR and PSA mRNA were inhibited in both castration-resistant C4-2b and 22RV1 tumor xenografts. Conclusions: Our data suggest that AR mRNA down modulation by EZN-4176 inhibits prostate tumor growth and provides a novel strategy to treat advanced prostate cancer. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 602.