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Peptide-Based Delivery of Oligonucleotides Across Blood-Brain Barrier Model
Stockholm University, Faculty of Science, Department of Neurochemistry. Mahidol University, Thailand.ORCID iD: 0000-0003-2265-557X
Stockholm University, Faculty of Science, Department of Neurochemistry.ORCID iD: 0000-0002-4604-6413
Stockholm University, Faculty of Science, Department of Neurochemistry.ORCID iD: 0000-0001-8947-6643
Show others and affiliations
2014 (English)In: International Journal of Peptide Research and Therapeutics, ISSN 1573-3904, Vol. 20, no 2, 169-178 p.Article in journal (Refereed) Published
Abstract [en]

Delivery of pharmaceutical agents across a blood–brain barrier (BBB) is a challenge for brain cancer therapy. In this study, an in vitro BBB model was utilized to study the delivery of oligonucleotides across brain endothelial cells targeting to glioma cells in a Transwell™ setup. A series of novel peptides were synthesized by covalent conjugation of cell-penetrating peptides with targeting peptides for delivery of gene-based therapeutics. These peptides were screened for passage across the Transwell™ and we found the most efficient peptide PepFect32 from originating PepFect 14 coupled with the targeting peptide angiopep-2. PepFect32/pDNA nanocomplexes exhibited high transcytosis across the BBB in vitro model and the highest transfection efficiency to glioma cells. In conclusion, PepFect32 revealed the most efficient peptide-based vector for pDNA delivery across in vitro BBB model.

Place, publisher, year, edition, pages
2014. Vol. 20, no 2, 169-178 p.
Keyword [en]
Blood–brain barrier model, Cell-penetrating peptide, bEnd.3, Glioma cells, Plasmid transfection, Gene-based therapy
National Category
Chemical Sciences
Research subject
Neurochemistry with Molecular Neurobiology
Identifiers
URN: urn:nbn:se:su:diva-100010DOI: 10.1007/s10989-013-9378-4ISI: 000334420100007OAI: oai:DiVA.org:su-100010DiVA: diva2:690404
Available from: 2014-01-23 Created: 2014-01-23 Last updated: 2017-05-05Bibliographically approved
In thesis
1. Rational design and applications of cell-penetrating peptides
Open this publication in new window or tab >>Rational design and applications of cell-penetrating peptides
2014 (English)Licentiate thesis, comprehensive summary (Other academic)
Abstract [en]

Cell-penetrating peptides are peptides capable of translocating the cellular membrane and entering the cell, either alone or together with a cargo. Potential applications of cell-penetrating peptides include drug delivery and gene therapy. This thesis is focused on the development of novel cell-penetrating peptides and applications for passage across the blood-brain barrier. We have developed a series of novel cell-penetrating peptides based on the model amphipathic peptide and modifications developed for the PepFect peptides. Our general goal is to improve our understanding of the structural requirements for efficient cell penetration and to apply this knowledge in the development of improved cell-penetrating peptides. We have also developed an in vitro model of the blood brain barrier based on brain endothelial cells grown on a semi-permeable membrane. This model has been used together with a series of novel peptides modified with targeting sequences in order to study the passage of peptides across the barrier and into an underlying layer of glioma cells.

Place, publisher, year, edition, pages
Stockholm: Department of Neurochemistry, 2014. 80 p.
National Category
Chemical Sciences Neurosciences
Research subject
Neurochemistry with Molecular Neurobiology
Identifiers
urn:nbn:se:su:diva-99849 (URN)978-91-7447-848-8 (ISBN)
Presentation
2014-02-04, C458, Department of Neurochemistry, Stockholm University, Stockholm, 12:15 (English)
Opponent
Supervisors
Note

At the time of the seminar, the papers were not published and had a status as follows: Paper 1: Accepted; Paper 2: Epub ahead of print.

Available from: 2014-01-23 Created: 2014-01-20 Last updated: 2015-03-16Bibliographically approved
2. Peptide-based delivery to glioblastoma cells studied by the blood-brain barrier model
Open this publication in new window or tab >>Peptide-based delivery to glioblastoma cells studied by the blood-brain barrier model
2016 (English)Licentiate thesis, comprehensive summary (Other academic)
Abstract [en]

Crossing the blood-brain barrier (BBB) is a great challenge for delivery of highly charged macromolecules such as nucleic acids and proteins to the brain. Cell-penetrating peptides (CPPs) are promising vectors to deliver various cargoes ranging from small molecules to large molecules such as antibodies, proteins, and nucleic acids. The BBB limits the passage of all large molecules to central nervous system (CNS), thus, CPP is a potential vector to use for oligonucleotide delivery across the BBB. In paper I, various CPPs were covalently conjugated with two different glioma-targeting peptides, glioma-homing peptide (gHo) and angiopep-2 (ANG). PepFect 32 (PF32), a conjugation between truncated PepFect 14 and ANG, was the most efficient vector to deliver plasmid DNA (pDNA) across a setup in vitro model of the BBB and showed the highest transfection in glioma cells. LRP-1 receptors, which are over-expressed in brain endothelial cells and glioma cells, were speculated to mediate the transcytosis of PF32:pDNA complexes across the BBB model since the ANG could target to LRP-1. In paper II, scavenger receptors class A and B (SCARA3, SCARA5, and SR-BI) were found to be expressed in the brain endothelial cells. Inhibition of these scavenger receptors led to a reduction of the transfection of PF32:pDNA complexes in the brain endothelial cells. Therefore, in the BBB model scavenger receptors also played a vital role as well as LRP-1 in the transport of oligonucleotides in the complex with peptide-based vector PF32.

In conclusion, PF32 is a potential vector to deliver pDNA across the BBB model and target to the glioma cells. The complexes of PF32:pDNA transport across the brain endothelial cells via receptor-mediated endocytosis pathway recognized by scavenger receptors and LRP-1. To improve the specificity and enhance the transport into the brain, the brain-homing devices are considered as a promising strategy for CNS drug delivery.

Place, publisher, year, edition, pages
Stockholm: Department of Neurochemistry, 2016. 33 p.
Keyword
peptide-based delivery, blood-brain barrier, glioblastoma
National Category
Other Chemistry Topics
Research subject
Neurochemistry with Molecular Neurobiology
Identifiers
urn:nbn:se:su:diva-129528 (URN)978-91-7649-432-5 (ISBN)
Presentation
2016-05-13, Heilbronnsalen, C458, Svante Arrhenius väg 16B, Stockholm, 13:00 (English)
Opponent
Supervisors
Available from: 2016-04-25 Created: 2016-04-25 Last updated: 2016-04-25Bibliographically approved
3. Cell-penetrating peptide based nanocomplexes for oligonucleotide delivery
Open this publication in new window or tab >>Cell-penetrating peptide based nanocomplexes for oligonucleotide delivery
2016 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Oligonucleotide-based drugs hold great promise for the treatment of many types of diseases, ranging from genetic disorders to viral infections and cancer. The problem is that efficient delivery across the cell membrane is required for oligonucleotides to have their desired effect. Cell-penetrating peptides (CPPs) provide a solution to this problem. CPPs are capable of transporting cargoes such as drugs or nucleic acids for gene therapy into the cell, either by covalent conjugation to the cargo or by non-covalent complex formation. This thesis is focused on the development of a class of peptides called PepFects, peptides with fatty acid modifications capable of forming nanoparticle-sized complexes with oligonucleotides. These complexes are efficiently internalized by many different cell types and are generally non-toxic and non-immunogenic.

We have developed a number of novel PepFect peptides and a quantitative structure-activity model to predict the biological effect of our peptides. In addition, the involvement of scavenger receptors class A in the endocytic uptake of PepFect complexes as well as other CPPs and polymeric transfection agents was studied. Lastly, we have developed a series of PepFect peptides for delivery across the blood-brain barrier and a model system mimicking the blood-brain barrier in order to evaluate the passage of these peptides.

The general aim of this thesis is to improve the understanding of intracellular delivery of oligonucleotides with PepFect peptides from both a chemical and a biological viewpoint, and further improve the efficacy of this delivery system with the long-term goal of making it useful in clinical settings. 

Place, publisher, year, edition, pages
Stockholm: Department of Neurochemistry, Stockholm University, 2016. 69 p.
Keyword
Cell-penetrating peptides, oligonucleotides, gene therapy, drug delivery, scavenger receptors, blood-brain barrier
National Category
Chemical Sciences Biochemistry and Molecular Biology
Research subject
Neurochemistry with Molecular Neurobiology
Identifiers
urn:nbn:se:su:diva-133794 (URN)978-91-7649-479-0 (ISBN)
Public defence
2016-11-04, Magnélisalen, Kemiska övningslaboratoriet, Svante Arrhenius väg 16 B, Stockholm, 10:00 (English)
Opponent
Supervisors
Available from: 2016-10-12 Created: 2016-09-15 Last updated: 2016-09-26Bibliographically approved
4. Cell-penetrating peptides targeting glioblastomas for nucleic acid delivery in the blood-brain barrier model
Open this publication in new window or tab >>Cell-penetrating peptides targeting glioblastomas for nucleic acid delivery in the blood-brain barrier model
2017 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Glioblastoma multiforme is the most aggressive form of malignant brain tumor with poor prognosis. The efficacy of brain cancer treatment by chemotherapeutics is limited by the blood-brain barrier (BBB) which allows less than 2% of the small molecules and blocks almost all the macromolecules to transport into the brain. Delivery of the large molecules such as proteins and nucleic acids across the BBB is a great challenge for brain-targeted drug delivery. To overcome this obstacle, cell-penetrating peptides (CPPs) were used as vectors for delivery of nucleic acids across the BBB targeting glioblastomas. The CPPs have shown such promising carriers to deliver various cargoes ranging from small molecules to large molecules into the cells. This thesis is focused on the development of glioblastoma-targeting vectors based on modifications of the CPPs and the targeting peptides. The peptide-based vectors were developed to improve the transport of the nucleic acids across the BBB and specifically target glioblastomas.

In this thesis, a series of peptide-based vectors targeting glioblastomas were synthesized and modified with targeting peptides by either covalent conjugation or non-covalent complex formation. The delivery of plasmid DNA (pDNA) in the complex with the peptide-based vectors was studied in the in vitro model of the BBB. The role of receptors expressed on the BBB was investigated. Scavenger receptors class A and B were found to be expressed on the BBB, and they were involved in the delivery of the pDNA across the BBB model. Moreover, various targeting peptides were modified with hexaglutamate to form non-covalent complexes with the CPPs for small interfering RNA (siRNA) delivery to glioblastoma cells. The non-covalent complex of the CPP and the targeting peptide showed greater gene-silencing efficiency than the consecutively covalent conjugation of the CPP and the targeting peptide for siRNA delivery to glioblastoma cells. Lastly, a number of novel, amphipathic peptides were developed based on the model amphipathic peptide. The prediction of the biological effect of the designed peptides using quantitative structure-activity relationship model showed a correlation with the experimental data.

Finally, the CPP-based nucleic acid delivery vectors with homing peptide strategy have a potential for the BBB shuttle and the future use as a glioblastoma-targeted drug carrier in the in vivo studies and the clinical applications.

Place, publisher, year, edition, pages
Stockholm: Department of Neurochemistry, Stockholm University, 2017
Keyword
Blood-brain barrier model, nucleic acid delivery, glioblastomas, cell-penetrating peptides
National Category
Chemical Sciences
Research subject
Neurochemistry with Molecular Neurobiology
Identifiers
urn:nbn:se:su:diva-142109 (URN)978-91-7649-725-8 (ISBN)978-91-7649-726-5 (ISBN)
Public defence
2017-06-09, Magnélisalen, Kemiska övningslaboratoriet, Svante Arrhenius väg 16 B, Stockholm, 10:00 (English)
Opponent
Supervisors
Note

At the time of the doctoral defense, the following paper was unpublished and had a status as follows: Paper 4: Manuscript.

Available from: 2017-05-17 Created: 2017-04-27 Last updated: 2017-05-17Bibliographically approved

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