Construction of folate-conjugated pRNA of bacteriophage phi29 DNA packaging motor for delivery of chimeric siRNA to nasopharyngeal carcinoma cells.
Nasopharyngeal carcinoma is a poorly differentiated upper respiratory tract cancer that highly expresses human folate receptors (hFR). Binding of folate to hFR triggers endocytosis. The folate was conjugated into adenosine 5'-monophosphate (AMP) by 1,6-hexanediamine linkages. After reverse HPLC to reach 93% purity, the folate-AMP, which can only be used for transcription initiation but not for chain extension, was incorporated into the 5'-end of bacteriophage phi29 motor pRNA. A 16:1 ratio of folate-AMP to ATP in transcription resulted in more than 60% of the pRNA containing folate. A pRNA with a 5'-overhang is needed to enhance the accessibility of the 5' folate for specific receptor binding. Utilizing the engineered left/right interlocking loops, polyvalent dimeric pRNA nanoparticles were constructed using RNA nanotechnology to carry folate, a detection marker, and siRNA targeting at an antiapoptosis factor. The chimeric pRNAs were processed into ds-siRNA by Dicer. Incubation of nasopharyngeal epidermal carcinoma (KB) cells with the dimer resulted in its entry into cancer cells, and the subsequent silencing of the target gene. Such a protein-free RNA nanoparticle with undetectable antigenicity has a potential for repeated long-term administration for nasopharyngeal carcinoma as the effectiveness and specificity were confirmed by ex vivo delivery in the animal trial.
Department of Pathobiology and Weldon School of Biomedical Engineering, Purdue University, West Lafayette, IN 47907, USA.
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D000249:Adenosine Monophosphate / Q000235:genetics / Q000378:metabolism; D000818:Animals; D017103:Bacillus Phages / Q000235:genetics*; D002352:Carrier Proteins / Q000378:metabolism; D002478:Cells, Cultured; D002678:Chimera; D042003:DNA Packaging; D058973:Folate Receptors, GPI-Anchored; D005492:Folic Acid / Q000235:genetics*; D020868:Gene Silencing; D005818:Genetic Engineering; D006801:Humans; D008297:Male; D051379:Mice; D008819:Mice, Nude; D049329:Nanostructures; D009303:Nasopharyngeal Neoplasms / Q000628:therapy*; D009374:Neoplasms, Experimental / Q000378:metabolism / Q000628:therapy; D034741:RNA, Small Interfering / Q000008:administration & dosage*; D011956:Receptors, Cell Surface / Q000378:metabolism
D002352:Carrier Proteins; D058973:Folate Receptors, GPI-Anchored; D034741:RNA, Small Interfering; D011956:Receptors, Cell Surface; D000249:Adenosine Monophosphate; D005492:Folic Acid