Publikationsserver der Universitätsbibliothek Marburg

Titel:Entwicklung eines protektiven dermalen Drug Delivery Systems für therapeutische DNAzyme gegen Hauterkrankungen wie die aktinische Keratose
Autor:Marquardt, Kay
Weitere Beteiligte: Renz, Harald (Prof. Dr.) / Runkel, Frank (Prof. Dr.)
Veröffentlicht:2016
URI:https://archiv.ub.uni-marburg.de/diss/z2017/0002
DOI: https://doi.org/10.17192/z2017.0002
URN: urn:nbn:de:hebis:04-z2017-00025
DDC: Medizin, Gesundheit
Titel(trans.):Development of a protective dermal drug delivery system for therapeutic DNAzymes targeting skin diseases such as actinic keratosis
Publikationsdatum:2017-01-12
Lizenz:https://creativecommons.org/licenses/by/4.0

Dokument

Schlagwörter:
Hautkrankheit, Haut, DNAzyme, submicron emulsion, Pharmazeutische Technologie, Polyplex, Wirkstoff, Therapeutisches System, Submikronemulsion, polyplexe, DNase, DNS, Keratosis actinica, DNAzym

Zusammenfassung:
Die therapeutischen DNAzyme stellen als neue Wirkstoffgruppe einen vielversprechend Ansatz bei der Therapie von Hauterkrankungen dar. Derzeit sind fünf DNAzyme in der klinischen Wirkstoffprüfung von denen zukünftig wiederum zwei gegen Hauterkrankungen eingesetzt werden sollen. Für eine erfolgreiche Therapie muss der Wirkstoff an den Zielort in der Haut penetrieren und dort intakt vorliegen. Allerdings erschweren allgemein die Eigenschaften der Wirkstoffgruppe den Wirkstofftransport und den Erhalt der Wirkstoffintegrität. Zudem besitzt die Haut eine Barrierefunktion, die es zu überwinden gilt, um eine hohe Bioverfügbarkeit in den pathogen veränderten Hautzellen zu erreichen. Die Hautbarriere lässt sich in eine passive und aktive Barriere unterteilen. Während die aktive Barriere der Integrität des Wirkstoffes schadet, verhindert die passive Barriere die Wirkstoffdiffusion. In Bezug auf die aktive Barriere konnte nachgewiesen werden, dass das ungeschützte DNAzym auf der humanen Hautoberfläche durch DNase II degradiert wird. Durch eine Komplexierung mit Polykationen zu Polyplexen konnte der Wirkstoff auf der Haut geschützt werden. In Abhängigkeit der Eigenschaften des Polykations waren unterschiedliche ξ-Verhältnisse nötig, um den Schutz zu gewährleisten. Ein ξ-Verhältnis im Bereich von 1 2 und darüber hinaus im positiven Zeta-Potenzial erhöht die Chance auf einen ausreichenden Schutz. Als Polykationen eigneten sich insbesondere bioabbaubare Chitosan-Polymere, welche im Vergleich zu Polyethylenimin laut Literatur eine geringere Toxizität aufweisen. In Bezug auf die passive Hautbarriere konnte diese mittels Submicronemulsion (SME) überwunden werden. Selbst die größeren Polyplex-Partikel konnten durch die SME in die Haut penetrieren. Dabei erhöhten die Hilfsstoffe der SME die Penetration und erreichen eine Akkumulation des Wirkstoffes in dem Stratum corneum der intakten Haut. Somit stand das Stratum corneum als Wirkstoffreservoir zur Verfügung. Die Polyplexe wiederum erreichen auf zellulärer Ebene eine erhöhte Aufnahme des Wirkstoffes in HaCaT-Zellen. Die Kombination aus Chitosan-Polyplexen und SME führte zu dem Ergebnis eines protektiven dermalen DDS, das sowohl die Penetration als auch die Zellaufnahme fördert und darüber hinaus den Wirkstoff schützt.

Summary:
DNAzymes are a group of synthetic nucleic acid based APIs that have the ability to inhibit protein translation by targeting the transcript of a specific gene. DNAzymes have already been tested in clinical trials with five different diseases. Two of the DNAzymes have been tested for dermal application against atopic dermatitis and skin cancer (squamous-cell carcinoma, basal cell carcinoma). The development of an appropriate dermal drug delivery system (DDS) is challenging, because DNAzymes are prone to enzymatic degradation and have problematic properties in terms of skin penetration. The task of this thesis was to develop a dermal DDS that addresses both issues. The dermal DDS should maintain the integrity of DNAzymes while enhancing the penetration and uptake of DNAzymes into human skin and the targeted keratinozytes. First, to ensure the integrity of this particularly group of APIs, the degradation of DNAzymes on human skin had to be identified and analysed. The analysis revealed that unprotected DNAzymes were rapidly degraded by a DNase II like activity. Therefore, protective systems were developed to protect against DNase II. Especially, complexation of DNAzymes with polycations, forming polyplexes, was able to sufficiently maintain the integrity of the API. Depending on the properties of the polycations a specific ratio (ξ) between the oppositely charged DNAzymes and polycations was necessary to fully protect. A ξ-ratio of about 1 2 along with a positive zeta potential increased in general the chances of an appropriate protection. The most effective protection was achieved with polycations of biodegradable chitosan. To enhance the penetration of DNAzymes through the skin barrier the submicron emulsion (SME) was identified as ideal candidate and the SME was developed as carrier system for DNAzymes. The SME increased the penetration of API into skin. The DNAzymes accumulated especially in the stratum corneum due to the SME’s excipients. Consequently, the stratum corneum was able to act as an API reservoir. The SME and the chitosan polyplexes were finally combined to a so called protective dermal DDS. The protective dermal DDS enhanced the penetration into the skin and protected the DNAzymes. On cellular level the uptake of DNAzymes into the targeted keratinocytes were enhanced due to the polyplexes.

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