Journal
JOURNAL OF NEUROCHEMISTRY
Volume 139, Issue 5, Pages 872-885Publisher
WILEY-BLACKWELL
DOI: 10.1111/jnc.13841
Keywords
acid sphingomyelinase; neutral sphingomyelinase2; quantum dots; RNA interference
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Funding
- United States Public Health Service (USPHS) [NS36866-40, GM098871]
- Naval Research Laboratory (NRL)
- Naval Research Laboratory Nanoscience Institute (NRL NSI)
- Office of Naval Research (ONR)
- Defense Advanced Research Project Agency (DARPA)
- Defense Threat Research Agency (DTRA)
- Joint Science and Technology Office (JSTO)
- Military interdepartmental Purchase Request (MIPR) [B112582M]
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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 HIS6 in JB577 (W center dot G center dot Dap(N-Palmitoyl)center dot VKIKK center dot P-9 center dot G(2)center dot H-6) 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 24-48h. 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.
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