This work explored the possibilities to design CMS nanocarriers for two different types of problems, the encapsulation of small hydrophobic drugs and subsequent transport into skin or the retention and targeted release of a cationic analgesic. The aim of the first part was to design a set of nanocarriers for the encapsulation and penetration enhancement of these drugs and thereby fulfilling the following criteria: Ease of synthesis, non-toxic products and building blocks, good loading capacity, degradability, and degradation-based release and high penetration enhancement. To simplify the synthesis, the dendritic building block, which poses the focal point in the nanocarrier architecture, was exchanged. Instead of hPG-NH2, unfunctionalized hPG was used. This change in the synthesis process had two main impacts. First, this strategy spared three synthetic steps thereby making the procedure less time consuming. Secondly, the potentially toxic building block hPG-NH2 was avoided and thus could not harm the organism post-application as a degradation product. This change was the next step in decreasing the number of amine groups in the core and thus the toxicity ranging from PEI over hPG-NH2 to hPG.[83,85] The inner shell’s building block was varied in length to study the impact on encapsulation, melting temperature and penetration enhancement. Aliphatic diacids with a number of carbon atoms between 12 and 19 as well as a branched version of the one with 18 carbon atoms were used as inner shell building blocks. While, in previous studies, the length of the inner shell had already been altered with bigger steps (six C-atoms),[83] here the focus shifted more onto details and other structural features. The different systems showed an increase of the melting temperature (Tm), of the inner shell with increasing chain length. And while an amide bond in the structure significantly increased the Tm, the introduction of side arms led to its decrease. While the rigid molecule dexamethasone (DXM) showed no apparent correlation between chain length and loading capacity, the situation was different for the bulkier tacrolimus. Here, an optimum was found for a C15 inner shell. Interestingly, while the additional branching did not lead to results different from the linear ester-based architecture, the loading capacity plummeted when an amide-based nanocarrier was used. All examined nanocarriers were degraded by a lipase to a degree of more than 70% in two weeks, which was similar to other ester-based nanotransporters.[124,125] The degradation of DXM-loaded CMS nanotransporters led to the loss of their function as a solubilizer, more than 80% of the loading was released. Thus, the degradation can serve as a possible release mechanism. Various assays were carried out to determine toxicity. Of the investigated ester-based CMS architectures, CMS-E15 showed the least cytotoxicity. The others caused a decrease in cell viability, at least to some extent. Of the building blocks, the aliphatic diacids were only toxic after prolonged exposition, but of the PEG-conjugated diacids, cytotoxicity increased from C12 to C18, which was attributed to interaction with the cell membrane. The most toxic building block was C18-PEG350, also with regard to genotoxicity. The CMS-E18 nanocarrier, the product of this building block, also exhibited some ROS generation and genotoxicity. The overall least toxic, ester-based nanocarrier was CMS-E15, which was selected for in vivo testing and was nontoxic when applied to the skin of Sprague Dawley rats. After establishing the biocompatibility of the different candidates, the CMS nanocarriers were tested for skin penetration and penetration enhancement of several guest molecules on various skin models. Nile red-loaded and DXM-loaded CMS-nanocarriers were used to study the penetration enhancement on excised human skin. All nanocarriers proved to be superior over a formulation with base cream (NR) or LAW cream (DXM), which was in accordance with previous results.[85] The nanotransporters showed no dependence on inner shell chain length or type of bond for attachment. CMS-E15 was then loaded with dexamethasone and applied on an inflammatory skin model which had an upregulated interleukin 8 (IL-8) and IL-1β expression as part of their inflammatory condition. The application of the DXM-loaded nanocarriers reduced the expression of both ILs more effectively than a LAW cream with equal DXM content. The CMS nanocarrier was not only tested for application on skin, but for oral mucosa as well. Here, biocompatibility was of even higher importance, because the constant flow of saliva eventually flushed the CMS nanocarriers into the GI tract if not taken up by the mucosa. Along the penetration experiment, toxicity of the CMS nanotransporters was determined for gingival epithelial cells and found to be only cytotoxic at very high concentrations. The penetration enhancement of PCA-labeled dexamethasone encapsulated in CMS-E15 and the penetration of ICC-labeled, amide based CMS nanocarriers (CMS-ICC) was measured on buccal and masticatory mucosa. While penetration of unloaded CMS-ICC nanocarriers was limited to the stratum corneum on skin,[122] a small fraction also penetrated more deeply into viable tissues, in masticatory slightly more than in buccal mucosa. Although the conditions were not identical, this result might have indicated a higher penetrability for oral mucosa compared to skin, which might be related to a difference in the structure of both tissues. For the penetration enhancement of spin-labeled DXM (DXM-PCA) by CMS-E15, similar results were found as in skin. Even after a washing step, which simulated the flow of saliva in the oral cavity, penetration enhancement of the labeled drug was found. The CMS nanotransporters enhanced the penetration of DXM-PCA more than a cream formulation, and buccal mucosa generally seemed more penetrable than masticatory mucosa for DXM-PCA. Additionally, X-band electron paramagnetic resonance spectroscopy revealed that the guest molecule left its vehicle. This observation was in accordance with previous observations,[122] which indicated that nanocarrier and guest molecule penetrated up to different depths. Taken together, these three studies describe a step-wise selection process at which out of six different initial candidates, one was selected by criteria like ease of synthesis, drug loading capacity, and biocompatibility. The nanocarrier with the best overall performance and thus most promising for future applications was CMS-E15. The aim of the second part of this work was to design a nanocarrier for the encapsulation and controlled release of the drug U 50,488H (U50), which is a strong analgesic for the post-operational pain treatment but elicits side effects when it crosses the blood brain barrier. A formulation into a nanocarrier can help to increase circulation time and facilitate targeted release. After surgery, the tissue is inflamed and thereby acidified and fenestrated. To use these conditions for the targeted release of U50, a nanocarrier was synthesized that offered the possibility to encapsulate U50 efficiently and release it in a pH-dependent manner. As basic architectures, CMS-E15 and CMS-A18, which were the most promising candidates in previous studies, were used and extended by an anchor moiety. It was needed, because there was no possibility to bind the drug covalently to the nanocarrier. For this task, N-Z-L-protected aspartic acid was utilized, because it offered the possibility for ionic interactions, π-π stacking, and hydrogen bonds to U50. Further, it was possible to employ the carboxylic acid groups for a pH-dependent release. While many approaches in literature used a hydrophobic segment that was ionized upon a pH change and thus,[126–128] a lipophilic guest is expelled, here, the opposite is the case. Ionic interactions are interrupted and thus the ionic drug can diffuse out of the nanocarrier. After synthesis, the pKapp of the carboxylic acid groups was determined to be in the physiological range (4.4 for pCMS-A and 5.4 for pCMS-E) and the loading capacity (LC) and encapsulation efficiency (EE) for U50 free base was measured. While the EE at 16 wt% feed already exceeded 10% for all nanocarriers (> 60% EE), the feed was increased to 48 wt%. Under these conditions, the LC of all nanoarchitectures could be increased, especially for the pCMS, which reached up to 28% (> 50% EE). Subsequently, the release kinetics were studied under various conditions to determine the influence of the additional functionalization and factors like pH, the encapsulation of the free base or the methylsulfonate salt, or the presence of physiological salt concentration. While the latter two had no influence on the release kinetics, the functionalization with the binding patch was beneficial for the retention of U50. In the case of pCMS A, there was no difference in this effect at pH 5.5 and 7.4, but for pCMS E, a pH dependence was found. For this reason, pCMS-E was selected for an in vivo study in rats. Here, formulations of U50 with and without pCMS-E were compared regarding prolonged analgesia and reduced side effects. While the latter one was not the case, the formulation with additional pCMS-E resulted in the enhancement and the extension of the analgesic effect, which might indicate a longer blood circulation time caused by the nanocarrier. In a previous prodrug approach, morphine covalently attached to hPG was tested under similar conditions and the absence of a systemic side effect reported.[61] Taken together, these two studies indicated that to optimize post-operational pain treatment, nanocarrier architectures ideally have a sufficiently high molecular weight for a prolonged blood circulation time and bind the analgesic tightly enough to prevent premature release. If possible, this should be realized in a DDS approach, to make the drug interchangeable. When first reported, the core-multishell nanocarrier was termed universal, because it was suitable for both hydrophilic and hydrophobic guest molecules in both aqueous and organic media. Subsequent studies confirmed that and reported the encapsulation of even inorganic nanoparticles and explored the fundamental effects, such as the aggregation of nanoparticles and guest molecules, which were also described theoretically. In a more application-focused approach, many nanocarriers were designed for a certain guest, e.g., copper ions, or target, for instance, the endosome. Ideally both can be achieved, namely, the identification of a promising architecture and further insight into fundamental effects. In this work, this was achieved. In the first part, it could be shown that there is an optimum chain length for the encapsulation tacrolimus and that the addition of one CH2 group to the aliphatic chain can alter the nanocarrier’s properties dramatically. There also was a dependence of cytotoxicity on the length of the hydrophobic part of the amphiphilic double shell, which altogether identified CMS-E15 as the most promising candidate. Addressing oral mucosa with CMS showed that this is an interesting field for DDS like CMS nanocarriers, because even more efficient penetration seems to be possible. In the more application-focused second part, the pH-dependent retention effect that was measured for the best candidate pCMS-E. Even though the encapsulation of hydrophilic guest molecules remains a greater challenge than the formulation of hydrophobic ones, this showed that it was possible even for rather small nanocarrier systems (diameter ~10 nm) to bind a water-soluble drug. Additionally, it could be proven that the chemistry used to attach the anchor moiety influenced the pKapp and thus is crucial for the application. Different anchor molecules could be employed to further fine-tune the pH of release and the retention effect proven in this study might be improved by increasing the nanocarrier’s sizes. One advantage of the DDS-approach is that pCMS are a potential nanocarrier suitable for any aromatic, hydrophobic drug molecule that bears a positive charge, e.g. by an amino group. The past research conducted on CMS nanocarriers has shown that, starting from a universal nanocarrier, of which mainly fundamental effects have been studied, many subclasses and varieties have been realized to meet specific requirements. To be suitable for future applications, a universal nanocarrier will not be able to solve all the problems. It will be necessary to adapt the nanocarrier’s architecture to the requirements of drug and physiological condition. And to achieve that, a library of CMS nanocarriers is needed to study fundamental effects and to select a potential candidate for an application. It is important to design new architectures, because the human body’s complexity demands specialized DDS for specific applications. This work has not only extended the library, but also identified certain novel nanocarriers as promising candidates.
