期刊
DNA AND CELL BIOLOGY
卷 30, 期 9, 页码 715-726出版社
MARY ANN LIEBERT, INC
DOI: 10.1089/dna.2011.1240
关键词
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Bifunctional small hairpin RNAs (bi-shRNAs) are functional miRNA/siRNA composites that are optimized for posttranscriptional gene silencing through concurrent mRNA cleavage-dependent and -independent mechanisms (Rao et al., 2010). We have generated a novel bi-shRNA using the miR30 scaffold that is highly effective for knockdown of human stathmin (STMN1) mRNA. STMN1 overexpression well documented in human solid cancers correlates with their poor prognosis. Transfection with the bi-shSTMN1-encoding expression plasmid (pbi-shSTMN1) markedly reduced CCL-247 human colorectal cancer and SK-Mel-28 melanoma cell growth in vitro (Rao et al., 2010). We now examine in vivo the antitumor efficacy of this RNA interference-based approach with human tumor xenografted athymic mice. A single intratumoral (IT) injection of pbi-shSTMN1 (8 mu g) reduced CCL-247 tumor xenograft growth by 44% at 7 days when delivered as a 1,2-dioleoyl-3-trimethylammoniopropane: cholesterol liposomal complex. Extended growth reductions (57% at day 15; p < 0.05) were achieved with three daily treatments of the same construct. STMN1 protein reduction was confirmed by immunoblot analysis. IT treatments with pbi-shSTMN1 similarly inhibited the growth of tumorgrafts derived from low-passage primary melanoma (>= 70% reduction for 2 weeks) and abrogated osteosarcoma tumorgraft growth, with the mature bi-shRNA effector molecule detectable for up to 16 days after last injection. Antitumor efficacy was evident for up to 25 days posttreatment in the melanoma tumorgraft model. The maximum tolerated dose by IT injection of >92 mu g (Human equivalent dose [HED] of >0.3mg/kg) in CCL-247 tumor xenograft-bearing athymic mice was similar to 10-fold higher than the extrapolated IC50 of 9 mu g (HED of 0.03 mg/kg). Healthy, immunocompetent rats were used as biorelevant models for systemic safety assessments. The observed maximum tolerated dose of <100 mu g for intravenously injected pbi-shSTMN1 (mouse equivalent of <26.5 mu g; HED of <0.09 mg/kg) confirmed systemic safety of the therapeutic dose, hence supporting early-phase assessments of clinical safety and preliminary efficacy.
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