Creation of a novel peptide with enhanced nuclear localization in prostate and pancreatic cancer cell lines

TY - JOUR

T1 - Creation of a novel peptide with enhanced nuclear localization in prostate and pancreatic cancer cell lines

AU - Lewis, H. Dan

AU - Husain, Ali

AU - Donnelly, Robert J.

AU - Barlos, Dimitrios

AU - Riaz, Sheraz

AU - Ginjupalli, Kalyani

AU - Shodeinde, Adetola

AU - Barton, Beverly E.

N1 - Funding Information: The authors acknowledge Dr. Sukhwinder Singh for acquiring data on the Amnis ImageStream and Mr. Richard A. DeMarco of Amnis Corporation for help analyzing data files. This work was supported by NIH grant CA 121782 (BEB) and a Research & Development Merit Award from the Department of Veterans Affairs (BEB).

PY - 2010/10/28

Y1 - 2010/10/28

N2 - Background: For 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.Results: Uptake 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.Conclusions: These 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.

AB - Background: For 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.Results: Uptake 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.Conclusions: These 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.

UR - https://www.scopus.com/pages/publications/77958573892

UR - https://www.scopus.com/pages/publications/77958573892#tab=citedBy

U2 - 10.1186/1472-6750-10-79

DO - 10.1186/1472-6750-10-79

M3 - Article

C2 - 21029412

SN - 1472-6750

VL - 10

JO - BMC biotechnology

JF - BMC biotechnology

M1 - 79

ER -