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Characterization of a novel cytotoxic cell-penetrating peptide derived from p14ARF protein
Stockholm University, Faculty of Science, Department of Neurochemistry.
Stockholm University, Faculty of Science, Department of Neurochemistry.
Stockholm University, Faculty of Science, Department of Neurochemistry.
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2008 (English)In: Molecular Therapy, ISSN 1525-0016, E-ISSN 1525-0024, Vol. 16, no 1, 115-123 p.Article in journal (Refereed) Published
Place, publisher, year, edition, pages
2008. Vol. 16, no 1, 115-123 p.
Identifiers
URN: urn:nbn:se:su:diva-24648ISI: 000251821000020OAI: oai:DiVA.org:su-24648DiVA: diva2:197994
Note
Part of urn:nbn:se:su:diva-7287Available from: 2008-01-24 Created: 2008-01-11 Last updated: 2017-12-13Bibliographically approved
In thesis
1. Cell-penetrating peptides in protein mimicry and oligonucleotide delivery: Applications and mechanisms
Open this publication in new window or tab >>Cell-penetrating peptides in protein mimicry and oligonucleotide delivery: Applications and mechanisms
2008 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

The plasma membrane functions as a barrier, restricting entry of hydrophilic pharmaceutical agents. Cell-penetrating peptides (CPPs) are capable of transporting bioactive cargos into the cell and have consequently been extensively investigated for their mechanism of entry and capability to deliver various cargos spanning from peptides to plasmids.

The main aim of this thesis was to investigate the mechanism and capability of some of these CPPs to deliver mainly oligonucleotides and peptides into the cell. Oligonucleotides in the form of ds DNA decoy for sequestering of transcription factors or PNAs for redirection of splicing. In addition, peptides derived from the interaction interface of a tumor suppressor protein were investigated for their potential to combine a biological effect with internalization.

Peptides with or without any cargo were predominantly dependent on some form of endocytic mechanism for internalization, substantiated by using a functional assay, where all tested CPPs were associated with endocytosis for delivery of splice correcting PNAs. A new CPP, M918 proved most efficient in promoting splice correction and internalized mainly via macropinocytosis. In addition, TP10 efficiently delivered dsDNA decoy oligonucleotides for sequestering of the transcription factor Myc with a concomitant biological response, i.e. reduced proliferation. Finally, for the first time, to our knowledge, a novel pro-apoptotic peptide with cell-penetrating properties was designed from the tumor suppressor p14ARF, which decreased proliferation and induced apoptosis in cancer cell-lines, potentially mimicking the full-length protein. Altogether, this thesis highlights the functionality of CPPs and the possibility to develop new CPPs with improved or new properties, having the potential to advance delivery of therapeutic compounds.

Place, publisher, year, edition, pages
Stockholm: Institutionen för neurokemi, 2008. 62 p.
Keyword
peptide, oligonucleotide, cell-penetrating peptide, PNA, splicing
National Category
Neurosciences
Research subject
Neurochemistry and Molecular Neurobiology
Identifiers
urn:nbn:se:su:diva-7287 (URN)978-91-7155-511-3 (ISBN)
Public defence
2008-02-15, Magnélisalen, Kemiska övningslaboratoriet, Svante Arrhenius väg 12 A, Stockholm, 10:00 (English)
Opponent
Supervisors
Available from: 2008-01-24 Created: 2008-01-11 Last updated: 2009-09-18Bibliographically approved
2. Cell-penetrating peptides: Uptake, stability and biological activity
Open this publication in new window or tab >>Cell-penetrating peptides: Uptake, stability and biological activity
2011 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Cell-penetrating peptides (CPPs) have emerged as a group of remarkable delivery vectors for various hydrophilic macromolecules, otherwise excluded from cells due to the protective plasma membrane. Unbiased conclusions regarding e.g. uptake mechanism, intracellular distribution and cargo delivery efficacy is complicated by the use of different methodological parameters by different laboratories. The first paper in this thesis introduced unifying protocols enabling comparison of results from different research groups. One of these methods, HPLC, was used in paper II to investigate CPP uptake and degradation in yeasts. Both parameters varied depending on peptide and yeast species; however pVEC emerged as a promising delivery vector in yeast since it internalized into both species tested without concomitant degradation. Protein mimicry was another investigated phenomenon and in paper III a 22-mer peptide from the p14Arf protein (Arf (1-22)) was found to be sufficient for retaining its function as a tumor suppressor. This peptide comprised a combination of apoptogenic property and CPP in one unity, thus providing opportunity to conjugate cytotoxic agents boosting the tumoricidal activity. Surprisingly, a partially inverted control peptide to Arf (1-22), called M918, was found to be an extraordinary CPP. In paper IV, it was shown to be superior to well-established CPPs in delivery of both peptide nucleic acids and proteins. Albeit the promising results these two peptides displayed, their utility in vivo, as with all peptides, is hampered by rapid degradation. With the aim of improving their stability, Arf (1-22) and M918 were synthesized with D-amino acids in the reverse order, a modification called retro-inverso (RI) isomerization. Their cell-penetrating ability was retained, but the treated cells displayed unexpected morphological alterations indicative of apoptosis. The presented results demonstrate the versatility of CPPs, functioning as vectors in both yeast and mammalian cells and as protein mimicking peptides with biological activity. Their potential as drug delivery agents is obvious; however, peptide degradation is an issue that requires further improvements before clinical success is in reach.

Place, publisher, year, edition, pages
Stockholm: Department of Neurochemistry, Stockholm University, 2011. 99 p.
National Category
Natural Sciences
Research subject
Neurochemistry and Neurotoxicology
Identifiers
urn:nbn:se:su:diva-55664 (URN)978-91-7447-269-1 (ISBN)
Public defence
2011-05-06, Magnélisalen, Kemiska övningslaboratoriet, Svante Arrhenius väg 16 B, Stockholm, 13:00 (English)
Opponent
Supervisors
Note
At the time of the doctoral defense, the following papers were unpublished and had a status as follows: Paper 5: In press.Available from: 2011-04-14 Created: 2011-03-24 Last updated: 2011-03-24Bibliographically approved

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