Follicular targeted drug delivery via nanocarriers in the treatment of (auto)inflammatory skin diseases

Today, it is widely known that nanoparticles (NPs) maximize follicular and minimize interfollicular drug delivery, thereby reducing systemic drug levels and severe side effects. Non-life-threatening skin diseases, like Alopecia Areata (AA), a reversible hair loss disorder associated with major negative effect on quality of life, lack of sufficient treatment, among other things caused by negative risk-benefit profiles of potent drugs such as Janus Kinase inhibitors, e.g., tofacitinib (TFB). Therefore, NPs are predestined to enable a targeted drug delivery into hair follicles (HFs) for a safer treatment against skin diseases like AA. As proof of concept, the NP deposition inside human scalp HFs was studied by adjustment of a quantification method for human forearm HFs. Investigations on healthy human, AAaffected hairless and hairy body donor scalp HFs showed that NP uptake into HFs depends on follicular density but is independent of hair shaft presence and hair type. Accordingly, TFB-loaded NPs were developed. Previously-synthesized squalenyl derivative NPs exceeded other approaches, demonstrating high drug loading capacity, biocompatibility, colloidal stability, and enhanced follicular delivery in an ex vivo pig ear model, which was supported by a biological effect in an in vivo dermatitis mouse model. Further investigations on follicular targeted delivery via NPs for a safe and potent treatment of skin diseases, leading to translation into the clinics, are highly desired.Nanopartikel (NP) maximieren den follikulären und minimieren den interfollikulären Arzneistofftransport. Sie reduzieren dabei systemische Wirkstoffspiegel und schwere Nebenwirkungen. Für nicht lebensbedrohliche Hauterkrankungen wie Alopecia Areata (AA) gibt es keine zufriedenstellende Behandlung, da unter anderem für potente Arzneistoffe wie Janus Kinase Inhibitor Tofacitinib (TFB) negative Nutzen-Risiko-Verhältnisse existieren. Daher ist ein gezielter Arzneistofftransport durch NP in Haarfollikeln (HF) prädestiniert, um eine sichere und effiziente Behandlung von AA zu ermöglichen. Die Machbarkeitsstudie untersuchte die NP-Deposition in menschlichen Kopfhaut-HF unter Zuhilfenahme einer Quantifizierungsmethode für menschliche Unterarm-HF. Sowohl Untersuchungen an gesunden, an AA-befallenen haarlosen als auch an Körperspenden Kopfhaut-HF zeigten, dass die NP-Aufnahme in HF abhängig ist von der Follikeldichte, jedoch nicht vom Haarschaft-Vorhandensein oder dem Haartyp. TFB beladene Squalen-Derivat-NP überzeugten durch hohe Arzneistoffbeladung, Biokompatibilität, kolloidale Stabilität und verbesserten follikulären Arzneistofftransport – gezeigt im Ex-vivo-Schweineohrmodell und unterstützt durch eine biologische Antwort im In-vivo-Dermatitis-Mausmodell. Weitere Untersuchungen des follikulären gezielten Arzneistofftransports mit NP zur Erreichung einer sicheren und effizienten Behandlung von Hauterkrankungen, welche zu einer Translation in die Klinik führen, sind sehnlichst erwünscht.Dr. Rolf M. Schwiete Stiftung, Mannheim, German

Novel anti-infective delivery systems for the treatment of pulmonary bacterial infections

Drug delivery systems (DDS) have the capacity to overcome biological barriers limiting the bioavailability of inhaled anti-infectives. This is important to eradicate bacterial infections and to prevent the development of bacterial resistance. Despite substantial efforts in the field, the current state-of-the-art often fails to achieve those goals, and we still observe an increase in bacterial resistance. In the context of drug delivery of pulmonary anti-infectives, this work proposes three novel strategies (i) polyplexes based on natural polysaccharides (starch, chitosan and cyclodextrin), (ii) self-assembly of amphiphilic excipients (farnesylated chitosan, squalenyl hydrogen sulfate), (iii) innovative excipient-free nano-assemblies of aminoglycoside antibiotics and farnesyl quorum sensing inhibitors. The latter invention allows an exceptional 100% co-loading capacity of established antibiotics and innovative pathoblockers without the need of any additional excipients. The biofilm eradicating efficacy of such DDS increased 16-fold compared to free actives. Finally, the interaction between the DDS and some important pulmonary biological barriers, such as biofilm, mucus, and macrophages, was also investigated. The knowledge gained in this thesis advances the research in pulmonary anti-infectives delivery beyond the state-of-the-art.Drug Delivery Systeme (DDS) haben die Fähigkeit, biologische Barrieren zu überwinden, welche die Bioverfügbarkeit inhalierter Antiinfektiva begrenzen. Dies ist wichtig, um bakterielle Infektionen zu beseitigen und die Entwicklung bakterieller Resistenz zu verhindern. Trotz erheblicher Anstrengungen auf diesem Gebiet werden diese Ziele vom derzeitigen Stand der Technik oft nicht erreicht, und wir beobachten immer noch eine Zunahme der bakteriellen Resistenz. Im Rahmen der pulmonalen Wirkstofffreisetzung von Antiinfektiva werden in dieser Arbeit drei neue Strategien vorgeschlagen: (i) Polyplexe auf der Basis natürlicher Polysaccharide (Stärke, Chitosan und Cyclodextrin), (ii) Selbstorganisations- Partikel aus amphiphilen Hilfsstoffen (farnesyliertes Chitosan, Squalenylhydrogensulfat), und (iii) innovative Hilfsstoff-freie Nano-Assemblys aus Aminoglycosid-Antibiotika und Farnesyl-Quorum-Sensing-Inhibitoren (QSI). Die letztgenannte Erfindung ermöglicht eine bemerkenswerte 100 %-ige Beladung, da sowohl das Antibiotikum als auch der Pathoblocker QSI Wirkstoffe darstellen und keine Hilfsstoffe mehr benötigt werden. Ein solches DDS verbessert die Biofilm-Elimination 16-fach im Vergleich zur Behandlung mit freien Wirkstoffen. Abschließend wurde die Interaktion zwischen den DDSs und einigen wichtigen biologischen Lungenbarrieren, wie Biofilm, Schleim und Makrophagen, untersucht, was die Forschung im Bereich der Verabreichung von pulmonalen Antiinfektiva weiter über den Stand der Technik hinaus vorantreibt.MARIE SKŁODOWSKA-CURIE ACTIONS_Project: "Design and Development of advanced NAnomedicines to overcome Biological BArriers and to treat severe diseases" (NABBA

Tofacitinib Loaded Squalenyl Nanoparticles for Targeted Follicular Delivery in Inflammatory Skin Diseases

Tofacitinib (TFB), a Janus kinase inhibitor, has shown excellent success off-label in treating various dermatological diseases, especially alopecia areata (AA). However, TFB’s safe and targeted delivery into hair follicles (HFs) is highly desirable due to its systemic adverse effects. Nanoparticles (NPs) can enhance targeted follicular drug delivery and minimize interfollicular permeation and thereby reduce systemic drug exposure. In this study, we report a facile method to assemble the stable and uniform 240 nm TFB loaded squalenyl derivative (SqD) nanoparticles (TFB SqD NPs) in aqueous solution, which allowed an excellent loading capacity (LC) of 20%. The SqD NPs showed an enhanced TFB delivery into HFs compared to the aqueous formulations of plain drug in an ex vivo pig ear model. Furthermore, the therapeutic efficacy of the TFB SqD NPs was studied in a mouse model of allergic dermatitis by ear swelling reduction and compared to TFB dissolved in a non-aqueous mixture of acetone and DMSO (7:1 v/v). Whereas such formulation would not be acceptable for use in the clinic, the TFB SqD NPs dispersed in water illustrated a better reduction in inflammatory effects than plain TFB’s aqueous formulation, implying both encouraging good in vivo efficacy and safety. These findings support the potential of TFB SqD NPs for developing a long-term topical therapy of AA

Scalable processes to manufacture nanoparticulate dosage forms for oral vaccination

Polymeric nanoparticles are promising drug delivery systems and antigen-carriers for vaccination. They may enhance the severity or the type of the antigen-specific immune response and may facilitate needle-free vaccination via the oral or respiratory route. However, the translation of nanoparticulate systems from bench to bedside remains a major challenge. Among the reasons are the limited knowledge and control of critical process parameters during early research, and the poor scalability to and reproducibility in clinical research and the commercial stage. This thesis presents a novel, easily scalable and potentially continuous method for manufacturing antigen-loaded polymeric nanoparticles. The method allows for effective tuning of the nanoparticle size with economically interesting yield, relevant antigen-loading capacity and retained antigen integrity across a batch size range of four orders of magnitude, but with limited loading efficiency. Nanoparticle properties were comparable between scales, but the process parameters were not found to be independent of or proportional to scale. Two continuous methods were developed to simultaneously prepare and dry such nanoparticles for improved process efficiency, and to manufacture enteric-matrix multiparticulates for oral dosing. Further optimization is required to achieve full scalability, improve the cost-effectiveness of the processes, and to demonstrate the functionality for an oral vaccine.Polymerbasierte Nanopartikel sind vielversprechende Trägersysteme für therapeutische Anwendungen und Impfungen. Solche Systeme können Umfang und Art der antigen-spezifischen Immunreaktion maßgeblich beeinflussen sowie die orale oder inhalative Gabe ermöglichen. Allerdings stellt die Translation von Forschungsergebnissen in die Klinik eine große Herausforderung dar, die von der eingeschränkten Kenntnis und Kontrolle kritischer Prozessparameter sowie der ungenügenden Skalierbarkeit zu klinischer Entwicklung und kommerzieller Herstellung erschwert wird. Hier wurde eine neue, skalierbare und potentiell kontinuierliche Herstellungsmethode für antigen-beladene polymerbasierte Nanopartikel entwickelt. Sie ermöglicht das Anpassen der Partikelgröße bei wirtschaftlich interessanter Ausbeute, relevanter Antigenbeladung und erhaltener Antigenintegrität über vier Größenordnungen von Losgrößen, allerdings bei limitierter Antigenausbeute. Ähnliche Partikeleigenschaften wurden über verschiedene Losgrößen erzielt, jedoch waren die Prozessparameter nicht unabhängig von oder proportional zum Prozessvolumen. Eine kontinuierliche Methode zur effizienten weil simultanen Generierung und Trocknung von Nanopartikeln wurde entwickelt, sowie eine weitere zur Herstellung magensaftresistenter multipartikulärer Pulver zur oralen Anwendung. Weitere Prozessoptimierung ist notwendig für vollständige Skalierbarkeit und verbesserte Wirtschaftlichkeit, sowie der Beweis der Anwendbarkeit für die orale Impfung

Synthesis and Biopharmaceutical Characterization of Amphiphilic Squalenyl Derivative Based Versatile Drug Delivery Platform.

Hepatitis C virus (HCV) has no animal reservoir, infecting only humans. To investigate species barrier determinants limiting infection of rodents, murine liver complementary DNA library screening was performed, identifying transmembrane proteins Cd302 and Cr1l as potent restrictors of HCV propagation. Combined ectopic expression in human hepatoma cells impeded HCV uptake and cooperatively mediated transcriptional dysregulation of a noncanonical program of immunity genes. Murine hepatocyte expression of both factors was constitutive and not interferon inducible, while differences in liver expression and the ability to restrict HCV were observed between the murine orthologs and their human counterparts. Genetic ablation of endogenous Cd302 expression in human HCV entry factor transgenic mice increased hepatocyte permissiveness for an adapted HCV strain and dysregulated expression of metabolic process and host defense genes. These findings highlight human-mouse differences in liver-intrinsic antiviral immunity and facilitate the development of next-generation murine models for preclinical testing of HCV vaccine candidates

Synthesis and Biopharmaceutical Characterization of Amphiphilic Squalenyl Derivative Based Versatile Drug Delivery Platform

Limited drug loading capacity (LC), mostly below 5% w/w, is a significant drawback of nanoparticulate drug delivery systems (DDS). Squalenoylation technology, which employs bioconjugation of squalenyl moiety and drug, allows self-assemble of nanoparticles (NPs) in aqueous media with significantly high LC (>30% w/w). The synthesis and particle preparation of squalenoylated prodrugs are, however, not facile for molecules with multiple reactive groups. Taking a different approach, we describe the synthesis of amphiphilic squalenyl derivatives (SqDs) as well as the physicochemical and biopharmaceutical characterizations of their self-assembled NPs as DDSs. The SqDs included in this study are (i) cationic squalenyl diethanolamine (ii) PEGylated SqD (PEG 750 Da), (iii) PEGylated SqD (PEG 3,000 Da), and (iv) anionic squalenyl hydrogen sulfate. All four SqDs self-assemble into NPs in a size range from 100 to 200 nm in an aqueous solution. Furthermore, all NP derivatives demonstrate appropriate biocompatibility and adequate colloidal stability in physiological relevant pH environments. The mucoprotein binding of PEGylated NPs is reduced compared to the charged NPs. Most importantly, this technology allows excellent LC (at maximum of 45% w/w) of a wide range of multifunctional compounds, varying in physicochemical properties and molecular weight. Interestingly, the drug release profile can be tuned by different loading methods. In summary, the SqD-based NPs appear as versatile drug delivery platforms