Supplementary Fig. S2. A. HuR mRNA levels measured by RT-PCR in A2780 transiently-transfected with siHuR and siCtrl. B. Constitutively expressed shHuR maintains inhibition of HuR expression in ovarian tumor cells. Western blots of total cell lysates of parental and shHuR-constitutively expressing A2780, OVCAR5, and OVCA420 cells at various passage numbers. C. HuR mRNA levels measured by RT-PCR in late passages of A2780 and OVCAR5 cells expressing shHuRc or shCtrl. * = p<0.0005
Funding
Marsha Rivkin Center for Ovarian Cancer Research
Sharpe-Strumia Research Foundation of the Bryn Mawr Hospital
Sarah Parvin Foundation
ARTICLE ABSTRACT
Growing evidence shows that cancer cells use mRNA-binding proteins and miRNAs to posttranscriptionally regulate signaling pathways to adapt to harsh tumor microenvironments. In ovarian cancer, cytoplasmic accumulation of mRNA-binding protein HuR (ELAVL1) is associated with poor prognosis. In this study, we observed high HuR expression in ovarian cancer cells compared with ovarian primary cells, providing a rationale for targeting HuR. RNAi-mediated silencing of HuR in ovarian cancer cells significantly decreased cell proliferation and anchorage-independent growth, and impaired migration and invasion. In addition, HuR-depleted human ovarian xenografts were smaller than control tumors. A biodistribution study showed effective tumor-targeting by a novel Cy3-labeled folic acid (FA)-derivatized DNA dendrimer nanocarrier (3DNA). We combined siRNAs against HuR with FA-3DNA and found that systemic administration of the resultant FA-3DNA-siHuR conjugates to ovarian tumor–bearing mice suppressed tumor growth and ascites development, significantly prolonging lifespan. NanoString gene expression analysis identified multiple HuR-regulated genes that function in many essential cellular and molecular pathways, an attractive feature of candidate therapeutic targets. Taken together, these results are the first to demonstrate the versatility of the 3DNA nanocarrier for in vivo–targeted delivery of a cancer therapeutic and support further preclinical investigation of this system adapted to siHuR-targeted therapy for ovarian cancer. Cancer Res; 76(6); 1549–59. ©2016 AACR.
