Minicell-Based Targeted Delivery of shRNA to Cancer Cells: An Experimental Protocol
Bacterial minicell has emerged as a novel targeted delivery system for RNAi-based therapeutics. In this chapter, we have described the detailed protocol for the preparation of minicell-based targeted delivery system for shRNA. Initially, minicell-producing parent bacterial cells were transformed with plasmid vector containing shRNA. Subsequently, shRNA-packaged minicells were purified from parent bacterial cells. Purified minicells were characterized by fluorescence microscopy and transmission electron microscopy. In the next step, targeting ligand was conjugated on the minicell surface for the active targeting of cancer cell surface receptors. Eventually, target-specific delivery of minicells was explored in vitro in selected cancer cell line and in vivo in mice bearing tumor xenograft.
KeywordsBacterial minicells shRNA delivery Folate receptor targeted delivery RNA interference Cancer
The authors are thankful to B. V. Patel Pharmaceutical Education and Research Development (PERD) Centre, Ahmedabad, for providing all the facility for the successful completion of the work. Authors are also grateful to Dr. Lawrence Rothfield (University of Connecticut, USA) for providing E. coli PB114 as a generous gift. Additionally, authors would like to thank CSIR, India, for providing financial assistance to Mehul Jivrajani as Senior Research Fellow (# 113353/2K12/1) to carry out this work. Authors are also thankful to NIRMA University, Ahmedabad, Gujarat, India.
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