Quantum dot-mediated delivery of siRNA to inhibit sphingomyelinase activities in brain-derived cells.
The use of RNAi to suppress protein synthesis offers a potential way of reducing the level of enzymes or the synthesis of mutant toxic proteins but there are few tools currently available for their delivery. To address this problem, bioconjugated quantum dots (QDs) containing a hydrophobic component (N-palmitate) and a sequence VKIKK designed to traverse across cell membranes and visualize drug delivery were developed and tested on cell lines of brain origin. We used the Zn outer shell of the QD to bind HIS6in JB577 (W•G•Dap(N-Palmitoyl)•VKIKK•P9•G2•H6) and by a gel-shift assay showed that siRNAs would bind to the positively charged KIKK sequence. By comparing many peptides and QD coatings, we showed that the QD-JB577-siRNA construct was taken up by cells of nervous system origin, distributed throughout the cytosol, and inhibited protein synthesis, implying that JB577 was also promoting endosome egress. By attaching siRNA for luciferase in a cell line over-expressing luciferase, we showed 70% inhibition of mRNA after 24-48 h. To show more specific effects, we synthesized siRNA for neutral (NSMase2), acid (lysosomal ASMase) sphingomyelinase, and sphingosine kinase 1 (SK1), we demonstrated a dose-dependent inhibition of activity. These data suggest that QDs are a useful siRNA delivery tool and QD-siRNA could be a potential theranostic for a variety of diseases.
Department of Pediatrics, University of Chicago, Chicago, Illinois, USA.
Ted
T
Getz
Department of Pediatrics, University of Chicago, Chicago, Illinois, USA.
Jingdong
J
Qin
US Naval Research Labs, Washington, District of Columbia, USA.
Igor L
IL
Medintz
US Naval Research Labs, Washington, District of Columbia, USA.
James B
JB
Delehanty
US Naval Research Labs, Washington, District of Columbia, USA.
Kimihiro
K
Susumu
Scripps Research, La Jolla, California, USA.
Philip E
PE
Dawson
Department of Pediatrics, University of Chicago, Chicago, Illinois, USA.|Department of Biochemistry and Molecular Biology, University of Chicago, Chicago, Illinois, USA.
Glyn
G
Dawson
D000818:Animals; D001921:Brain / Q000187:drug effects* / Q000201:enzymology*; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects / Q000502:physiology; D004305:Dose-Response Relationship, Drug; D004789:Enzyme Activation / Q000187:drug effects / Q000502:physiology; D004791:Enzyme Inhibitors / Q000008:administration & dosage; D018014:Gene Transfer Techniques; D006801:Humans; D051379:Mice; D045663:Quantum Dots / Q000008:administration & dosage*; D034741:RNA, Small Interfering / Q000008:administration & dosage* / Q000235:genetics; D013108:Sphingomyelin Phosphodiesterase / Q000037:antagonists & inhibitors* / Q000378:metabolism*
D004791:Enzyme Inhibitors; D034741:RNA, Small Interfering; D013108:Sphingomyelin Phosphodiesterase
RNA interference; acid sphingomyelinase; neutral sphingomyelinase2; quantum dots