RRC ID 60147
Author Athyala PK, Chitipothu S, Kanwar JR, Krishnakumar S, Narayanan J.
Title Synthesis of saporin-antibody conjugates for targeting EpCAM positive tumour cells.
Journal IET Nanobiotechnol
Abstract Epithelial cell adhesion molecule (EpCAM) is a transmembrane glycoprotein involved in cell proliferation and differentiation. Ribosomal inactivating proteins derived from plants specifically target ribosomes and irreversibly inhibit protein synthesis. EpCAM antibody and saporin were conjugated using 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride/N-hydroxysuccinimide chemistry. The mass of the conjugates were characterised using matrix-assisted laser desorption ionisation (MALDI). The saporin-EpCAM (SAP-EpAB) conjugates were tested in-vitro against MCF-7 (breast cancer cells), WERI-Rb1 (retinoblastoma) cells. The flow cytometry and fluorescence microscopy were performed to show the binding efficiency of SAP-EpAB conjugate. Whole transcriptome changes of sap-conjugate treated cells were studied using affymetrix microarrays. MALDI-TOF analysis and polyacrylamide gel electrophoresis confirmed the conjugation of SAP with EpCAM antibody. Flow cytometry and fluorescent microscopy analysis revealed the binding of SAP-EpAB conjugates to the MCF-7, WERI-Rb1 cells. Apoptosis assay by annexin-V has shown an increased apoptotic and necrotic population in conjugate treated cells. MTT assay confirmed the tumour cell death and had shown the IC50 value of 0.8 µg for conjugate in MCF-7 (breast cancer cells), and 1 µg for WERI-Rb1 (retinoblastoma) cells. The microarray analysis revealed downregulation of the tumourigenic genes and upregulation of pro-apoptotic genes leading to apoptosis of tumour cells.
Volume 13(1)
Pages 90-99
Published 2019-2-1
DOI 10.1049/iet-nbt.2018.5222
PMID 30964044
MeSH Antibodies / chemistry Antibodies / immunology Antibodies / metabolism* Antibodies / pharmacology Antineoplastic Agents / chemical synthesis Antineoplastic Agents / chemistry Antineoplastic Agents / metabolism* Antineoplastic Agents / pharmacology Apoptosis / drug effects Cell Line, Tumor Drug Delivery Systems / methods* Epithelial Cell Adhesion Molecule / immunology Epithelial Cell Adhesion Molecule / metabolism* Humans MCF-7 Cells Saporins / chemistry* Saporins / metabolism
IF 1.859
Times Cited 0
Resource
Human and Animal Cells WERI-Rb-1(RCB2146)