Creation of a novel peptide with enhanced nuclear localization in prostate and pancreatic cancer cell linesReport as inadecuate




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BMC Biotechnology

, 10:79

First Online: 28 October 2010Received: 11 February 2010Accepted: 28 October 2010

Abstract

BackgroundFor improved uptake of oligonucleotide-based therapy, the oligonucleotides often are coupled to peptides that facilitate entry into cells. To this end, novel cell-penetrating peptides CPPs were designed for mediating intracellular uptake of oligonucleotide-based therapeutics. The novel peptides were based on taking advantage of the nuclear localization properties of transcription factors in combination with a peptide that would bind putatively to cell surfaces. It was observed that adding a glutamate peptide to the N-terminus of the nuclear localization signal NLS of the Oct6 transcription factor resulted in a novel CPP with better uptake and better nuclear colocalization than any other peptide tested.

ResultsUptake of the novel peptide Glu-Oct6 by cancer cell lines was rapid in less than 1 hr, more than 60% of DU-145 cells were positive for FITC, complete by 4 hr, 99% of cells were positive for FITC, concentration-dependent, temperature-dependent, and inhibited by sodium azide NaN3. Substitution of Phe, Tyr, or Asn moieties for the glutamate portion of the novel peptide resulted in abrogation of novel CPP uptake; however none of the substituted peptides inhibited uptake of the novel CPP when coincubated with cells. Live-cell imaging and analysis by imaging flow cytometry revealed that the novel CPP accumulated in nuclei. Finally, the novel CPP was coupled to a carboxyfluorescein-labeled synthetic oligonucleotide, to see if the peptide could ferry a therapeutic payload into cells.

ConclusionsThese studies document the creation of a novel CPP consisting of a glutamate peptide coupled to the N-terminus of the Oct6 NLS; the novel CPP exhibited nuclear colocalization as well as uptake by prostate and pancreatic cancer cell lines.

Electronic supplementary materialThe online version of this article doi:10.1186-1472-6750-10-79 contains supplementary material, which is available to authorized users.

H Dan Lewis, Ali Husain contributed equally to this work.

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Author: H Dan Lewis - Ali Husain - Robert J Donnelly - Dimitrios Barlos - Sheraz Riaz - Kalyani Ginjupalli - Adetola Shodeinde -

Source: https://link.springer.com/article/10.1186/1472-6750-10-79



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