Identification of a Novel Cell-Penetrating Peptide from Human Phosphatidate Phosphatase LpIN3

Lim, Su-Ho; Kim, Won-Ju; Kim, Yeonho; Choi, Je-Min

doi:10.1007/s10059-012-0284-y

Cited by 25 publications

(15 citation statements)

References 30 publications

(33 reference statements)

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“…To examine the intracellular protein delivery activity of these CPP candidates, we initially constructed plasmids expressing enhanced green fluorescent proteins (EGFPs), including NP1-EGFP, NP2-EGFP, and EGFP without a CPP, and then purified the resulting proteins ( Supplementary Fig. 2 ), as previously described 26 . These proteins were examined by incubation with cultured cells to determine their intracellular transduction efficiency.…”

Section: Resultsmentioning

confidence: 99%

dNP2 is a blood–brain barrier-permeable peptide enabling ctCTLA-4 protein delivery to ameliorate experimental autoimmune encephalomyelitis

Lim

Kim

et al. 2015

Nat Commun

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Central nervous system (CNS)-infiltrating effector T cells play critical roles in the development and progression of multiple sclerosis (MS). However, current drugs for MS are very limited due to the difficulty of delivering drugs into the CNS. Here we identify a cell-permeable peptide, dNP2, which efficiently delivers proteins into mouse and human T cells, as well as various tissues. Moreover, it enters the brain tissue and resident cells through blood vessels by penetrating the tightly organized blood–brain barrier. The dNP2-conjugated cytoplasmic domain of cytotoxic T-lymphocyte antigen 4 (dNP2-ctCTLA-4) negatively regulates activated T cells and shows inhibitory effects on experimental autoimmune encephalomyelitis in both preventive and therapeutic mouse models, resulting in the reduction of demyelination and CNS-infiltrating T helper 1 and T helper 17 cells. Thus, this study demonstrates that dNP2 is a blood–brain barrier-permeable peptide and dNP2-ctCTLA-4 could be an effective agent for treating CNS inflammatory diseases such as MS.

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Section: Resultsmentioning

confidence: 99%

dNP2 is a blood–brain barrier-permeable peptide enabling ctCTLA-4 protein delivery to ameliorate experimental autoimmune encephalomyelitis

Lim

Kim

et al. 2015

Nat Commun

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“…ZFNs typically contain a single N-terminal Simian vacuolating virus 40 (SV40) NLS sequence (PKKKRKV) that mediates nuclear import but does not measurably contribute to its intrinsic cell-penetrating activity. 27 Because in some contexts NLS sequences possess an innate ability to cross cell membranes 45 and mediate protein transfection, 46 we hypothesized that tandem NLS repeats could enhance ZFN protein cell-permeability. To test this, we fused one, two, three, or four additional repeats of the SV40 NLS to the N-terminus of ZFN proteins that already contained one NLS and were designed to target the human CCR5 gene ( Figure 1a ).…”

Section: Resultsmentioning

confidence: 99%

Improved Cell-Penetrating Zinc-Finger Nuclease Proteins for Precision Genome Engineering

Liu

Gaj

Wallen

et al. 2015

Molecular Therapy - Nucleic Acids

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Safe, efficient, and broadly applicable methods for delivering site-specific nucleases into cells are needed in order for targeted genome editing to reach its full potential for basic research and medicine. We previously reported that zinc-finger nuclease (ZFN) proteins have the innate capacity to cross cell membranes and induce genome modification via their direct application to human cells. Here, we show that incorporation of tandem nuclear localization signal (NLS) repeats into the ZFN protein backbone enhances cell permeability nearly 13-fold and that single administration of multi-NLS ZFN proteins leads to genome modification rates of up to 26% in CD4+ T cells and 17% in CD34+ hematopoietic stem/progenitor cells. In addition, we show that multi-NLS ZFN proteins attenuate off-target effects and that codelivery of ZFN protein pairs facilitates dual gene modification frequencies of 20–30% in CD4+ T cells. These results illustrate the applicability of ZFN protein delivery for precision genome engineering.

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“…A traditional CPP, the trans-activator of transcription (TAT) of HIV-1, has been extensively conjugated on the surface of drug-loaded nanoparticles to increase the transport efficiency across the BBB to the CNS [23]. However, drawbacks of TAT include non-human origin, immunogenicity, and low delivery efficiency [24]. Recent studies have explored a new CPP called RF from screening of the 16 amino acid peptide library [25,26,27,28].…”

Section: Introductionmentioning

confidence: 99%

Enhancing Anticancer Effect of Gefitinib across the Blood–Brain Barrier Model Using Liposomes Modified with One α-Helical Cell-Penetrating Peptide or Glutathione and Tween 80

Lin¹,

Hong²,

Wang³

et al. 2016

IJMS

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Epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (TKI), such as gefitinib, have been demonstrated to effectively treat the patients of extracranial non-small cell lung cancer (NSCLC). However, these patients often develop brain metastasis (BM) during their disease course. The major obstacle to treat BM is the limited penetration of anticancer drugs across the blood–brain barrier (BBB). In the present study, we utilized gefitinib-loaded liposomes with different modifications to improve gefitinib delivery across the in vitro BBB model of bEnd.3 cells. Gefitinib was encapsulated in small unilamellar liposomes modified with glutathione (GSH) and Tween 80 (SUV-G+T; one ligand plus one surfactant) or RF (SUV-RF; one α-helical cell-penetrating peptide). GSH, Tween 80, and RF were tested by the sulforhodamine B (SRB) assay to find their non-cytotoxic concentrations on bEnd.3 cells. The enhancement on gefitinib across the BBB was evaluated by cytotoxicity assay on human lung adenocarcinoma PC9 cells under the bEnd.3 cells grown on the transwell inserts. Our findings showed that gefitinib incorporated in SUV-G+T or SUV-RF across the bEnd.3 cells significantly reduced the viability of PC9 cells more than that of free gefitinib. Furthermore, SUV-RF showed no cytotoxicity on bEnd.3 cells and did not affect the transendothelial electrical resistance (TEER) and transendothelial permeability of sodium fluorescein across the BBB model. Moreover, flow cytometry and confocal laser scanning microscopy were employed to evaluate the endocytosis pathways of SUV-RF. The results indicated that the uptake into bEnd.3 cells was mainly through adsorptive-mediated mechanism via electrostatic interaction and partially through clathrin-mediated endocytosis. In conclusion, cell penetrating peptide-conjugated SUV-RF shed light on improving drug transport across the BBB via modulating the transcytosis pathway(s).

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Identification of a Novel Cell-Penetrating Peptide from Human Phosphatidate Phosphatase LpIN3

Cited by 25 publications

References 30 publications

dNP2 is a blood–brain barrier-permeable peptide enabling ctCTLA-4 protein delivery to ameliorate experimental autoimmune encephalomyelitis

dNP2 is a blood–brain barrier-permeable peptide enabling ctCTLA-4 protein delivery to ameliorate experimental autoimmune encephalomyelitis

Improved Cell-Penetrating Zinc-Finger Nuclease Proteins for Precision Genome Engineering

Enhancing Anticancer Effect of Gefitinib across the Blood–Brain Barrier Model Using Liposomes Modified with One α-Helical Cell-Penetrating Peptide or Glutathione and Tween 80

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