Abstract 948:In vivoevaluation of IGF2 RNA interference in a mouse xenograft model of paclitaxel-resistant human ovarian cancer.

Introduction. We previously showed that paclitaxel-resistance of human ovarian carcinoma cell lines was reverted by knockdown of IGF2 with RNA interference. The aim of this study was to determine if our paclitaxel-resistant cell line HEY-T30 was resistant to paclitaxel treatment in vivo and if this resistance could be reverted by IGF2 knockdown through short hairpin RNA. Methods. Paclitaxel-resistant HEY-T30 cells, derived in our laboratory from HEY ovarian carcinoma cells, were transfected with a plasmid containing shRNA to target IGF2 (shIGF2-p) or a nontargeting shRNA (shScrambled). Stably transfected clones were evaluated by reverse transcription real-time PCR for IGF2 mRNA expression and by sulforhodamine cytotoxicity assay for paclitaxel sensitivity. For in vivo experiments, HEY, HEY-T30, shIGF2-p or shScrambled cells were suspended in OptiMem and one million cells/animal injected subcutaneously into 4-week-old female nude mice. Tumors were allowed to grow to a mean volume of 125 mm3, prior to treatment initiation with paclitaxel (20 mg/kg administered intraperitoneally every 3 days x 5 doses) or vehicle (5% dextrose in water; D5W). Animals were weighed and tumors were measured with digital calipers three times per week. Effect of treatment was calculated as effect=1-(T/C) at the indicated time point, where T is the mean tumor volume of the paclitaxel treatment group and C is the mean tumor volume of the vehicle control group. Significance was calculated with a 2-way repeated measures ANOVA with a Bonferroni multiple comparisons post-test. Results. IGF2 mRNA levels of shIGF2-p cells were comparable to HEY, whereas shScrambled was similar to HEY-T30. The IC50s (concentration of 50% inhibition of proliferation) for paclitaxel in vitro were HEY: 2.3 nM; HEY-T30: 164.0 nM; shIGF2-p: 13.89 nM; shScrambled: 140.0 nM. HEY xenografts responded well to paclitaxel treatment, showing a significant growth retardation compared to the D5W group beginning at 9 days after treatment initiation (p=0.0005), with an effect of treatment of 83.2% at day 19. In contrast, HEY-T30 xenografts showed no significant difference in tumor volume between the paclitaxel-treated and the D5W group at any time point, confirming its resistance to paclitaxel (the effect of treatment was 5.5%). shScrambled xenografts were similarly resistant to paclitaxel as HEY-T30 xenografts. shIGF2-p xenografts responded well to paclitaxel treatment, showing a difference in tumor volume from day 15 onward (day 15, p=0.0457; day 19, p=0.0006) with an effect of treatment of 64.1% after 22 days. Discussion. HEY-T30 xenografts are resistant to paclitaxel treatment, whereas HEY xenografts respond well. IGF2 knockdown by shRNA restores paclitaxel sensitivity to the HEY-T30 xenografts. These in vivo results confirm our prior observations in cell lines and suggest a novel target for the treatment of paclitaxel-resistant disease. Citation Format: Jurriaan Brouwer-Visser, Jiyeon Lee, Maria J. Cossio, Gloria S. Huang. In vivo evaluation of IGF2 RNA interference in a mouse xenograft model of paclitaxel-resistant human ovarian cancer. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 948. doi:10.1158/1538-7445.AM2013-948