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Dimeric Cationic Amphiphilic Polyproline Helices for Mitochondrial Targeting

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ABSTRACT

Purpose

Efficient delivery of therapeutic biopolymers across cell membranes remains a daunting challenge. The development of cell-penetrating peptides (CPPs) has been useful; however, many CPPs are found trapped within endosomes, limiting their use as delivery agents. We optimize a class of CPPs, cationic amphiphilic polyproline helices (CAPHs), for direct transport into cells with mitochondrial localization through dimerization.

Methods

The CAPH P11LRR used for this study has been found to enter cells by two distinct pathways: an endocytotic pathway was favored at low concentrations; internalization by direct transport was observed at higher concentrations. CAPH was dimerized to probe if direct transport within cells may be enhanced through increased association of CAPH with the membrane and through the association of individual peptides within the membrane.

Results

The dimerization of the CAPH was found to significantly increase cellular uptake over its monomeric counterpart, with a concomitant lowering of the concentration threshold favoring direct transport. Evidence for direct transport within cells and mitochondrial localization was observed.

Conclusions

CAPH cellular uptake efficiency can be significantly enhanced through peptide dimerization while favoring cell entry via direct transport at low concentration with low cell toxicity.

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Abbreviations

CAPH:

cationic amphiphilic polyproline helix

CPP:

cell-penetrating peptide

FBS:

fetal bovine serum

FCCP:

carbonylcyanide p-trifluoromethoxyphenylhydrazone

FL:

fluorescein

Fmoc:

9-fluorenylmethyloxycarbonyl

HATU:

N,N,N',N'-Tetramethyl-O-(7-azabenzotriazol-1-yl)uranium hexafluorophosphate

MTT:

3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide

NHS-FL:

5-(and 6)-carboxyfluorescein, succinimidyl ester

PBS:

phosphate-buffered saline

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Authors and Affiliations

  1. Department of Chemistry, Purdue University, 560 Oval Drive, West Lafayette, Indiana, 47907, USA

    Iris M. Geisler & Jean Chmielewski

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  1. Iris M. Geisler
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  2. Jean Chmielewski
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Correspondence to Jean Chmielewski.

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Geisler, I.M., Chmielewski, J. Dimeric Cationic Amphiphilic Polyproline Helices for Mitochondrial Targeting. Pharm Res 28, 2797–2807 (2011). https://doi.org/10.1007/s11095-011-0493-7

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  • DOI: https://doi.org/10.1007/s11095-011-0493-7

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