RRC ID 67254
Author Subramanian N, Kanwar JR, Kanwar RK, Sreemanthula J, Biswas J, Khetan V, Krishnakumar S.
Title EpCAM Aptamer-siRNA Chimera Targets and Regress Epithelial Cancer.
Journal PLoS One
Abstract Epithelial cell adhesion molecule (EpCAM), a cancer stem cell (CSC) marker is over expressed in epithelial cancers and in retinoblastoma (RB). We fabricated an EpCAM targeting aptamer-siRNA chimera and investigated its anti-tumor property and EpCAM intracellular domain (EpICD) mediated signaling in epithelial cancer. The anti-tumor efficacy of EpCAM aptamer-siEpCAM chimera (EpApt-siEp) was evaluated by qPCR, northern and Western blotting in WERI-Rb1- RB cell line, primary RB tumor cells and in MCF7- breast cancer cell line. Anti-tumor activity of EpApt-siEp was studied in vivo using epithelial cancer (MCF7) mice xenograft model. The mechanism and pathways involved in the anti-tumor activity was further studied using protein arrays and qPCR. EpApt-siEp chimera was processed in vitro by dicer enzyme. Treatment of the WERI-Rb1 and MCF7 cells with EpApt-siEp revealed statistically significant down regulation of EpCAM expression (P<0.005) and concomitant reduction in cellular proliferation. In primary RB cells cultured from RB tumors, EpApt-siEp silenced EpCAM, significantly inhibited (P<0.01) cell proliferation and induced cytotoxicity. Knockdown of EpICD expressed in RB primary tumors led to repression of pluripotency markers, SOX2, OCT4, NANOG, and CD133. In vivo studies showed complete tumor growth regression without any toxicity in animals (P<0.001) and tumor tissues showed significant downregulation (P<0.05) of EpCAM, MRP1, ABCG2, stathmin, survivin and upregulation of ATM (P<0.05) leading to apoptosis by intrinsic pathway with minor alteration in cytokines. Our results revealed that EpApt-siEp potentially eradicated EpCAM positive cancer cells through CSC marker suppression and apoptosis, while sparing normal EpCAM negative adjacent cells.
Volume 10(7)
Pages e0132407
Published 2015-1-1
DOI 10.1371/journal.pone.0132407
PII PONE-D-14-43570
PMID 26176230
PMC PMC4503753
MeSH Animals Antigens, Neoplasm / chemistry Antigens, Neoplasm / metabolism* Apoptosis Aptamers, Nucleotide / metabolism* Base Sequence Biomarkers, Tumor / metabolism Breast Neoplasms / metabolism Breast Neoplasms / pathology* Cell Adhesion Molecules / chemistry Cell Adhesion Molecules / metabolism* Cell Line, Tumor Cell Proliferation Endocytosis Epithelial Cell Adhesion Molecule Female Gene Knockdown Techniques Humans Immunohistochemistry Immunomodulation Kinetics MCF-7 Cells Mice Models, Biological Molecular Sequence Data Neoplastic Stem Cells / metabolism Protein Array Analysis Protein Structure, Tertiary RNA, Small Interfering / metabolism* Retinoblastoma Ribonuclease III / metabolism Xenograft Model Antitumor Assays
IF 2.74
Human and Animal Cells WERI-Rb-1(RCB2146) MCF7(RCB1904)