Abstract 3677: Nanoparticle mediated delivery of siRNA targeting RRM2 inhibits tumor growth of head and neck and lung cancers (2010)

Abstract Inhibition of specific genes by small interfering RNAs (siRNA) offers a promising therapeutic approach in oncology. However, siRNA therapy is hindered by poor stability under physiological conditions and limited intracellular uptake. Ribonucleotide reductase subunit M2 (RRM2) is an essential protein for DNA synthesis and is responsible for the reduction of ribonucleotides to deoxyribonucleotides, providing a balanced supply of precursors for DNA synthesis and repair. In this study we investigated the efficacy and mechanism of action of a targeted nanoparticle formulation of siRNA against RRM2. The clinical version of this nanoparticle is denoted as CALAA-01 and is currently used in a Phase I clinical trial. The nanoparticle consists of a cyclodextrin-containing polymer, a polyethylene glycol steric stabilization agent, and human transferrin as a targeting ligand for binding to transferrin receptors (TfR) that are typically up regulated in cancer cells. Initially, we screened several head and neck squamous cell carcinoma (HNSCC) and lung cancer cell lines and found various degrees of expression of TfR and RRM2. Suppression of RRM2 by transfection of 5 nM siRNA for 72 hours in vitro strongly inhibited the cell growth in HNSCC (Tu212∼75% and M4e∼60%) and in lung cancer cell lines (A549 and H460 ∼80%). Using xenograft (Tu212) as an in-vivo model, we found nanoparticles (10mg/Kg) injected via the tail vein in four dosing schedules (Day 1, 3, 8 and 10) significantly reduced RRM2 expression and its activity in xenograft tumors. RRM2 activity was reduced by an average 1.88-fold in the RRM2 siRNA treated group compare to the formulated control siRNA group (p=0.04). We monitored tumor growth in treated and control groups (8 mice in each group) for 28 days and found that tumor growth reduced by an average 2.9- fold in the treated group compared to formulated control siRNA (p=0.004). The nanoparticle delivered RRM2 siRNA significantly suppressed cell proliferation and induced apoptosis as evidenced by xenograft tumor tissue staining with Ki67 and terminal deoxynucleotidyl transferase dUTP nick and labeling, respectively. Mechanistic studies (in-vitro) have revealed siRNA-mediated suppression of RRM2 increases apoptosis in a p53-independent manner. Interestingly, suppression of RRM2 by 5nM siRNA for 72 hours increased activation of caspase 9 and caspase 3 at least 2-fold with concomitant increase in the levels of cleaved poly-ADP-ribose polymerase (PARP), thus suggesting a vital role for p73 in the observed apoptosis through an intrinsic pathway. Thus this study demonstrates that targeted nanoparticles can deliver RRM2 siRNA to head and neck tumors in mice and highlights RRM2 as an excellent target to induce potent apoptosis and tumor growth inhibition. (Supported by NIH/NCI grant U54CA119338-04) 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 3677.