Novel thiazolopyridine derivatives of diflapolin as dual sEH/FLAP inhibitors with improved solubility.

Inflammatory responses are orchestrated by a plethora of lipid mediators, and perturbations of their biosynthesis or degradation hinder resolution and lead to uncontrolled inflammation, which contributes to diverse pathologies. Small molecules that induce a switch from pro-inflammatory to anti-inflammatory lipid mediators are considered valuable for the treatment of chronic inflammatory diseases. Commonly used non-steroidal anti-inflammatory drugs (NSAIDs) are afflicted with side effects caused by the inhibition of beneficial prostanoid formation and redirection of arachidonic acid (AA) into alternative pathways. Multi-target inhibitors like diflapolin, the first dual inhibitor of soluble epoxide hydrolase (sEH) and 5-lipoxygenase-activating protein (FLAP), promise improved efficacy and safety but are confronted by poor solubility and bioavailability. Four series of derivatives bearing isomeric thiazolopyridines as bioisosteric replacement of the benzothiazole core and two series additionally containing mono- or diaza-isosteres of the phenylene spacer were designed and synthesized to improve solubility. The combination of thiazolo[5,4-b]pyridine, a pyridinylen spacer and a 3,5-Cl2-substituted terminal phenyl ring (46a) enhances solubility and FLAP antagonism, while preserving sEH inhibition. Moreover, the thiazolo[4,5-c]pyridine derivative 41b, although being a less potent sEH/FLAP inhibitor, additionally decreases thromboxane production in activated human peripheral blood mononuclear cells. We conclude that the introduction of nitrogen, depending on the position, not only enhances solubility and FLAP antagonism (46a), but also represents a valid strategy to expand the scope of application towards inhibition of thromboxane biosynthesis.

Synthesis and structure-activity relationships for some novel diflapolin derivatives with benzimidazole subunit.

A series of derivatives of the potent dual soluble epoxide hydrolase (sEH)/5-lipoxygenase-activating protein (FLAP) inhibitor diflapolin was designed, synthesised, and characterised. These novel compounds, which contain a benzimidazole subunit were evaluated for their inhibitory activity against sEH and FLAP. Molecular modelling tools were applied to analyse structure-activity relationships (SAR) on both targets and to predict solubility and gastrointestinal (GI) absorption. The most promising dual inhibitors of these series are 5a, 6b, and 6c.

Self-Emulsifying Drug Delivery Systems: Hydrophobic Drug Polymer Complexes Provide a Sustained Release in Vitro.

The aim of this study was to develop hydrophobic ionic drug polymer complexes in order to provide sustained drug release from self-emulsifying drug delivery systems (SEDDS). Captopril (CTL) was used as an anionic model drug to form ionic complexes with the cationic polymers Eudragit RS, RL, and E. Complexes of polymer to CTL charge ratio 1:1, 2:1, and 4:1 were incorporated in two SEDDS, namely FA which was 40% Kolliphor RH 40, 20% Kolliphor EL, and 40% castor oil and FB, which was 40% Kolliphor RH 40, 30% glycerol, 15% Kolliphor EL, and 15% castor oil. Blank and complex loaded SEDDS were characterized regarding their droplet size, polydispersity index (PDI), and zeta potential. Resazurin assay was performed on Caco-2 cells to evaluate the biocompatibility of SEDDS. Release of CTL from SEDDS was determined in release medium containing 0.2 mg/mL of 5,5'-dithiobis(2-nitrobenzoic acid) (DNTB) allowing quantification of free drug released into solution via a thiol/disulfide exchange reaction between CTL and DNTB forming a yellow dye. The droplet size of SEDDS FA and SEDDS FB were in the range of 100 ± 20 nm and 40 ± 10 nm, respectively, with a PDI < 0.5. The zeta potential of SEDDS FA and SEDDS FB increased after the incorporation of complexes. Cell viability remained above 80% after incubation with SEDDS FA and SEDDS FB in a concentration of 1% and 3% for 4 h. Without any polymer, CTL was entirely released from both SEDDS within seconds. In contrast, the higher the cationic lipophilic polymer to CTL ratio in SEDDS, the more sustained was the release of CTL. Among the polymers which were evaluated, Eudragit RL provided the most sustained release. SEDDS FA containing Eudragit RL and CTL in a ratio of 1:1 released 64.78 ± 8.28% of CTL, whereas SEDDS FB containing the same complex showed a release of 91.85 ± 1.17% within 1 h. Due to the formation of lipophilic ionic polymer complexes a sustained drug release from oily droplets formed by SEDDS can be achieved. Taking into account that drugs are otherwise instantly released from SEDDS, results of this study might open the door for numerous additional applications of SEDDS for which a sustained drug release is essential.

Thiolated PVP-Amphotericin B Complexes: An Innovative Approach toward Highly Mucoadhesive Gels for Mucosal Leishmaniasis Treatment.

The aim of this study was to synthesize polymeric excipients that can form mucoadhesive hydrogels containing amphotericin B (AmB) for the treatment of mucosal leishmaniasis. 2-(2-Acryloylaminoethyldisulfanyl)-nicotinic acid (ACENA) was copolymerized with N-vinyl pyrrolidone to obtain thiolated polyvinylpyrrolidone (PVP) that was then complexed with AmB to improve its solubility. The resulting structure of thiolated PVP was evaluated by 1H nuclear magnetic resonance to confirm S-protected thiol groups, and the average molecular mass was determined by size exclusion chromatography. Moreover, variants of thiolated PVP-AmB were studied for the thiol content, amount of complexed AmB, cytotoxicity, mucoadhesive properties, and antileishmaniasis activity. The highest achieved degree of thiolation was 772 ± 24.64 µmol/g, and the amount of complexed AmB was 27.05 ± 0.31 µmol per g of polymer. Thiolated PVP and thiolated PVP-AmB variants (0.5% m/v) showed no cytotoxicity, whereas the equivalent concentration of free AmB reduced Caco-2 cell viability to 70% within 24 h. Thiol-functionalized PVP and PVP-AmB complexes displayed 7.66- and 7.20-fold higher adhesion to the mucosal surface in comparison to unmodified PVP and PVP-AmB, respectively. In addition, variants of thiolated PVP-AmB complexes displayed 100% antileishmaniasis activity in comparison to the 80% killing efficiency of Fungizone, which has been applied in the equivalent AmB concentration of 0.2 µg/mL. Thiol-functionalized PVP proved to be a promising novel excipient for the delivery of AmB providing enhanced solubility and improved mucoadhesive properties which are beneficial for the treatment of mucosal leishmaniasis.

Synthesis and Characterization of Thiolated PVP-Iodine Complexes: Key to Highly Mucoadhesive Antimicrobial Gels.

The aim of this study was to synthesize iodine containing polymeric excipients for mucosal treatment of microbial infection exhibiting a prolonged mucosal residence time by forming an adhesive gel on the mucosal surface. In order to achieve this aim, 2-(2 acryloylamino-ethyldisulfanyl)-nicotinic acid (ACENA) was copolymerized with N-vinylpyrrolidone (NVP) to obtain thiolated polyvinylpyrrolidone (PVP) for complexation with iodine. The average molecular mass of different thiolated PVP variants was determined by size exclusion chromatography. The structure of thiolated PVP was confirmed by 1H NMR. Thiolated PVP variants were characterized for thiol content, cytotoxicity, iodine loading capacity, rheological behavior, and adhesion time on mucosa. The highest achieved degree of thiolation was 610 ± 43 µmol/g, and the maximum recorded iodine loading was 949 ± 31 µmol/g of polymer. Thiolated PVP variants (0.5% m/v) showed no toxicity after incubation on Caco-2 cells for the period of 3 and 24 h, respectively. Thiolated PVP and thiolated PVP-iodine complexes exhibited a 5.4- and 4.4-fold increased dynamic viscosity in porcine mucus in comparison to PVP and PVP-iodine complex, respectively. Compared to PVP and PVP-iodine complex thiol-functionalized PVP and PVP-iodine complexes demonstrated significantly prolonged attachment to mucosal surface over a period of 3 h. Thiol functionalized PVP proved to be a promising novel excipient for complexation with iodine and to exhibit strongly improved mucoadhesive properties.

Phenylbenzenesulfonates and -sulfonamides as 17ß-hydroxysteroid dehydrogenase type 2 inhibitors: Synthesis and SAR-analysis.

17ß-Hydroxysteroid dehydrogenase type 2 (17ß-HSD2) converts the potent estrogen estradiol into the weakly active keto form estrone. Because of its expression in bone, inhibition of 17ß-HSD2 provides an attractive strategy for the treatment of osteoporosis, a condition that is often caused by a decrease of the active sex steroids. Currently, there are no drugs on the market targeting 17ß-HSD2, but in multiple studies, synthesis and biological evaluation of promising 17ß-HSD2 inhibitors have been reported. Our previous work led to the identification of phenylbenzenesulfonamides and -sulfonates as new 17ß-HSD2 inhibitors by ligand-based pharmacophore modeling and virtual screening. In this study, new molecules representing this scaffold were synthesized and tested in vitro for their 17ß-HSD2 activity to derive more profound structure-activity relationship rules.

Can thiolation render a low molecular weight polymer of just 20-kDa mucoadhesive?

The objective was to investigate whether even low-molecular weight polymers (LMWPs) can be rendered mucoadhesive due to thiolation. Interceded by the double catalytic system carbodiimide/N-hydroxysuccinimide, cysteamine was covalently attached to a copolymer, poly(4-styrenesulfonic acid-co-maleic acid) (PSSA-MA) exhibiting a molecular weight of just 20 kDa. Depending on the amount of added N-hydroxysuccinimide and cysteamine, the resulting PSSA-MA-cysteamine (PC) conjugates exhibited increasing degree of thiolation, highest being "PC 2300" exhibiting 2300.16 ± 149.86 µmol thiol groups per gram of polymer (mean ± SD; n = 3). This newly developed thiolated polymer was evaluated regarding mucoadhesive, rheological and drug release properties as well from the toxicological point of view. Swelling behavior in 100 mM phosphate buffer pH 6.8 was improved up to 180-fold. Furthermore, due to thiolation, the mucoadhesive properties of the polymer were 240-fold improved. Rheological measurements of polymer/mucus mixtures confirmed results obtained by mucoadhesion studies. In comparison to unmodified polymer, PC 2300 showed 2.3-, 2.3- and 2.4-fold increase in dynamic viscosity, elastic modulus and viscous modulus, respectively. Sustained release of the model drug codeine HCl out of the thiomer was provided for 2.5 h (p < 0.05), whereas the drug was immediately released from the unmodified polymer. Moreover, the thiomer was found non-toxic over Caco-2 cells for a period of 6- and 24-h exposure. Findings of the present study provide evidence that due to thiolation LMWPs can be rendered highly mucoadhesive as well as cohesive and that a controlled drug release out of such polymers can be provided.

Experimentally validated HERG pharmacophore models as cardiotoxicity prediction tools.

The goal of this study was to design, experimentally validate, and apply a virtual screening workflow to identify novel hERG channel blockers. The hERG channel is an important antitarget in drug development since cardiotoxic risks remain as a major cause of attrition. A ligand-based pharmacophore model collection was developed and theoretically validated. The seven most complementary and suitable models were used for virtual screening of in-house and commercially available compound libraries. From the hit lists, 50 compounds were selected for experimental validation through bioactivity assessment using patch clamp techniques. Twenty compounds inhibited hERG channels expressed in HEK 293 cells with IC50 values ranging from 0.13 to 2.77 µM, attesting to the suitability of the models as cardiotoxicity prediction tools in a preclinical stage.

S-Protected Thiolated Chitosan versus Thiolated Chitosan as Cell Adhesive Biomaterials for Tissue Engineering.

Chitosan (Ch) and different Ch derivatives have been applied in tissue engineering (TE) because of their biocompatibility, favored mechanical properties, and cost-effectiveness. Most of them, however, lack cell adhesive properties that are crucial for TE. In this study, we aimed to design an S-protected thiolated Ch derivative exhibiting high cell adhesive properties serving as a scaffold for TE. 3-((2-Acetamido-3-methoxy-3-oxopropyl)dithio) propanoic acid was covalently attached to Ch via a carbodiimide-mediated reaction. Low-, medium-, and high-modified Chs (Ch-SS-1, Ch-SS-2, and Ch-SS-3) with 54, 107 and 140 µmol of ligand per gram of polymer, respectively, were tested. In parallel, three thiolated Chs, namely Ch-SH-1, Ch-SH-2, and Ch-SH-3, were prepared by conjugating N-acetyl cysteine to Ch at the same degree of modification to compare the effectiveness of disulfide versus thiol modification on cell adhesion. Ch-SS-1 showed better cell adhesion capability than Ch-SS-2 and Ch-SS-3. This can be explained by the more lipophilic surfaces of Ch-SS as a higher modification was made. Although Ch-SH-1, Ch-SH-2, and Ch-SH-3 were shown to be good substrates for cell adhesion, growth, and proliferation, Ch-SS polymers were superior to Ch-SH polymers in the formation of 3D cell cultures. Cryogels structured by Ch-SS-1 (SSg) resulted in homogeneous scaffolds with tunable pore size and mechanical properties by changing the mass ratio between Ch-SS-1 and heparin used as a cross-linker. SSg scaffolds possessing interconnected microporous structures showed good cell migration, adhesion, and proliferation. Therefore, Ch-SS can be used to construct tunable cryogel scaffolds that are suitable for 3D cell culture and TE.

Imine bond formation as a tool for incorporation of amikacin in self-emulsifying drug delivery systems (SEDDS).

AIM: The aim was to develop a self-emulsifying drug delivery system (SEDDS) for amikacin via imine bond formation with hydrophobic aldehydes. METHODS: Trans-2, cis-6-nonadienal, trans-cinnamaldehyde, citral and benzaldehyde were conjugated to amikacin at pH 8.5. Based on results of precipitation efficiency, Fourier-transform infrared spectroscopy (FTIR) and NMR analysis, amikacin-trans-cinnamaldehyde conjugates were further characterized regarding log Poctanol/water via HPLC. The release of amikacin from the amikacin-trans-cinnamaldehyde conjugates was examined through in vitro incubation with bovine serum albumin (BSA). SEDDS containing the amikacin-trans-cinnamaldehyde conjugates were tested regarding mean droplet size (MDS), polydispersity index (PDI), log DSEDDS/release medium and cell viability. RESULTS: Trans-cinnamaldehyde formed the most hydrophobic conjugates with amikacin whereas benzaldehyde did not form hydrophobic conjugates at all. Imine bond formation was confirmed by FTIR and NMR analysis. The highest increase in log P was achieved for the amikacin-trans-cinnamaldehyde conjugate in a molar ratio of 1:5, shifting from -8.58 up to 1.59. Incubation of this conjugate with BSA led to the formation of BSA-trans-cinnamaldehyde releasing in turn amikacin. SEDDS based on Capmul MCM, Cremophor EL and propylene glycol containing the conjugate demonstrated a MDS of 61.4 nm and PDI of 0.265. Log DSEDDS/release medium was calculated to be 3.38. Cell viability studies showed very good tolerability of conjugate loaded SEDDS in concentrations of 0.1% - 0.5%. CONCLUSION: Imine bond formation of amikacin with trans-cinnamaldehyde and the incorporation of the resulting conjugate into SEDDS represents a promising strategy for oral delivery of amikacin.

Antifungal Siderophore Conjugates for Theranostic Applications in Invasive Pulmonary Aspergillosis Using Low-Molecular TAFC Scaffolds.

Invasive pulmonary aspergillosis (IPA) is a life-threatening form of fungal infection, primarily in immunocompromised patients and associated with significant mortality. Diagnostic procedures are often invasive and/or time consuming and existing antifungals can be constrained by dose-limiting toxicity and drug interaction. In this study, we modified triacetylfusarinine C (TAFC), the main siderophore produced by the opportunistic pathogen Aspergillus fumigatus (A. fumigatus), with antifungal molecules to perform antifungal susceptibility tests and molecular imaging. A variation of small organic molecules (eflornithine, fludioxonil, thiomersal, fluoroorotic acid (FOA), cyanine 5 (Cy5) with antifungal activity were coupled to diacetylfusarinine C (DAFC), resulting in a "Trojan horse" to deliver antifungal compounds specifically into A. fumigatus hyphae by the major facilitator transporter MirB. Radioactive labeling with gallium-68 allowed us to perform in vitro characterization (distribution coefficient, stability, uptake assay) as well as biodistribution experiments and PET/CT imaging in an IPA rat infection model. Compounds chelated with stable gallium were used for antifungal susceptibility tests. [Ga]DAFC-fludioxonil, -FOA, and -Cy5 revealed a MirB-dependent active uptake with fungal growth inhibition at 16 µg/mL after 24 h. Visualization of an A. fumigatus infection in lungs of a rat was possible with gallium-68-labeled compounds using PET/CT. Heterogeneous biodistribution patterns revealed the immense influence of the antifungal moiety conjugated to DAFC. Overall, novel antifungal siderophore conjugates with promising fungal growth inhibition and the possibility to perform PET imaging combine both therapeutic and diagnostic potential in a theranostic compound for IPA caused by A. fumigatus.

Lysine-Based Biodegradable Surfactants: Increasing the Lipophilicity of Insulin by Hydrophobic Ion Paring.

AIM: The aim of this study was to evaluate biodegradable cationic surfactants based on lysine. METHODS: Lysine was esterified with cholesterol, oleyl alcohol and 1-decanol resulting in cholesteryl lysinate (CL), oleyl lysinate (OL) and decyl lysinate (DL). Esters were investigated regarding their log Dn-octanol/water, critical micelle concentration (CMC) and biodegradability. Hemolytic potential of CL, OL, DL and the already established hexadecyl lysinate (HL) was determined and complexes with insulin (INS) were formed by hydrophobic ion pairing (HIP). Lipophilic characteristics of ion-pairs were examined by analyzing their log Pn-butanol/water. RESULTS: Successful synthesis of CL, OL and DL was confirmed by IR, NMR and MS. Log D analysis revealed amphiphilic properties for the esters and a CMC of 0.01 mM, 2.0 mM and 6.0 mM was found for CL, OL and DL, respectively. Biodegradability was proven, as over 99% of OL and DL were degraded by isolated enzymes within 30 min and after 3 h 97% of CL was cleaved by membrane bound enzymes. OL as well as DL displayed no hemolytic effect and for CL cytotoxicity was significantly reduced in comparison to HL. INS/CL complex exhibited highest lipophilicity. CONCLUSION: Cholesterol-amino acid based surfactants seem to be promising agents for HIP.

Polyphosphate coatings: A promising strategy to overcome the polycation dilemma.

AIM: It was the aim of this study to develop a zeta potential changing drug delivery system by decorating lipid-based nanocarriers with a polycationic cell penetrating peptide (CPP) and subsequently masking these cationic substructures with polyphosphates. METHODS: In order to anchor the CPP poly-l-lysine (PLL) on the surface of the oily droplets of an o/w nanoemulsion, stearic acid was covalently attached to the peptide. The resulting CPP-decorated oily droplets were coated with phytic acid and tripolyphosphate. The elimination of these polyphosphates due to cleavage by alkaline phosphatase was monitored by the release of monophosphate from the surface of the nanocarriers, by the change in zeta potential and by cellular uptake studies on Caco-2 cells. RESULTS: Polyphosphate coated PLL-decorated nanocarriers exhibited a pronounced conversion of zeta potential from -14.1 mV to +4.2 mV in case of tripolyphosphate coated nanocarriers and from -9.9 mV to -2.6 mV in case of phytic acid coated nanocarriers. The cellular uptake on Caco-2 cells of the polyphosphate coated nanocarriers was 4-fold improved compared to the control nanocarriers. Furthermore, confocal images showed that the majority of nanodroplets distributed in cytoplasm not being internalized into lysosomes. CONCLUSION: Polyphosphate coating of CPP-decorated nanocarriers seems to be a promising and simple strategy to overcome the polycation dilemma.

S-protected thiolated hyaluronic acid: In-situ crosslinking hydrogels for 3D cell culture scaffold.

The purpose of this study was to synthesize S-protected thiolated hyaluronic acid (HA) and to evaluate its potential for 3D cell culture scaffold. S-protected thiolated HA was synthesized by the covalent attachment of N-acetyl-S-((3-((2,5-dioxopyrrolidin-1-yl)oxy)-3-oxopropyl)thio)cysteine hydrazide ligand to the HA. Hydrogels were characterized for texture, swelling behavior and rheological properties. Furthermore, the potential of S-protected thiolated HA hydrogels as a scaffold for tissue engineering was evaluated by cell proliferation studies with Caco-2 and NIH 3T3 cells. It showed enhanced cohesion upon addition of N-acetyl cysteine (NAC). Dynamic viscosity of S-protected thiolated HA hydrogel was increased up to 19.5-fold by addition of NAC and 10.1-fold after mixing with mucus. Furthermore, Caco-2 and NIH 3T3 cells encapsulated into hydrogels proliferated in-vitro. As this novel S-protected thiolated HA is stable towards oxidation and forms highly cohesive gels when getting into contact with endogenous thiols due to disulfide-crosslinking, it is a promising tool for 3D cell culture scaffold.

Tetradeca-thiolated cyclodextrins: Highly mucoadhesive and in-situ gelling oligomers with prolonged mucosal adhesion.

The purpose of this study was to synthesize a highly mucoadhesive tetradeca-thiolated ß-cyclodextrin (ß-CD) by replacement of all primary OH groups at C-6 position and all secondary OH groups at C-2 position of ß-CD backbone viaSH groups and to evaluate its rheological and mucoadhesive properties in-vitro. Primary and secondary OH groups of ß-CD were substituted by SH groups using a microwave-assisted method. The structure of tetradeca-thiolated ß-CD was confirmed by FTIR and 1H NMR spectroscopy. The modified ß-CD was evaluated for SH content, thiol stability towards oxidation and cytotoxicity. Moreover, the viscoelastic behavior of the modified oligomer was investigated via rheological studies with porcine intestinal mucus and fibrous structural protein keratin, whereas mucoadhesive properties were evaluated using different porcine mucosae. Tetradeca-thiolated ß-CD oligomer displayed 8144 ± 317 µmol thiol groups per gram. These thiol groups displaying a pKa value of 8.2 were stable at pH 4 but prone to oxidation at higher pH values. The newly synthesized thiolated CD did not show any cytotoxicity to Caco-2 cells at a concentration of 0.5% (m/v) within 24 h. Due to the addition of 0.5 and 2% (m/v) tetradeca-thiolated ß-CD to mucus and keratin, the dynamic viscosity was increased up to 7.6- and 5.9- fold, respectively, within 4 h at 37 °C. Moreover, in-vitro mucoadhesion studies of tetradeca-thiolated CD showed 78.6-, 60.3-, 62.3- and 49.3- fold improved mucoadhesion on intestinal, buccal, bladder and vaginal mucosa as compared to unmodified ß-CD, respectively. According to these results, tetradeca-thiolated ß-CD might be a promising auxiliary agent to provide a prolonged residence time of drug delivery systems on different mucosal surfaces.

Per-6-Thiolated Cyclodextrins: A Novel Type of Permeation Enhancing Excipients for BCS Class IV Drugs.

The purpose of the study was to develop a per-6-thiolated α-cyclodextrin (α-CD) by substituting all primary hydroxyl groups of α-CD with thiol groups and to assess its solubility-improving and permeation-enhancing properties for a BCS Class IV drug in vitro as well as in vivo. The primary hydroxyl groups of α-CD were replaced by iodine, followed by substitution with -SH groups. The structure of per-6-thiolated α-CD was approved by FT-IR and 1H NMR spectroscopy. The per-6-thiolated was characterized for thiol content, -SH stability, cytotoxicity, and solubility-improving properties by using the model BCS Class IV drug furosemide (FUR). The mucoadhesive properties of the thiolated oligomer were investigated via viscoelastic measurements with porcine mucus, whereas permeation-enhancing features were evaluated on the Caco-2 cell monolayer and rat gut mucosa. Furthermore, oral bioavailability studies were performed in rats. The per-6-thiolated α-CD oligomer displayed 4244 ± 402 µmol/g thiol groups. These -SH groups were stable at pH ≤ 4, exhibiting a pKa value of 8.1, but subject to oxidation at higher pH. Per-6-thiolated α-CD was not cytotoxic to Caco-2 cells in 0.5% (m/v) concentration within 24 h. It improved the solubility of FUR in the same manner as unmodified α-CD. The addition of per-6-thiolated α-CD (0.5% m/v) increased the mucus viscosity up to 5.8-fold at 37 °C within 4 h. Because of the incorporation in per-6-thiolated α-CD, the apparent permeability coefficient (Papp) of FUR was 6.87-fold improved on the Caco-2 cell monolayer and 6.55-fold on the intestinal mucosa. Moreover, in vivo studies showed a 4.9-fold improved oral bioavailability of FUR due to the incorporation in per-6-thiolated α-CD. These results indicate that per-6-thiolated α-CD would be a promising auxiliary agent for the mucosal delivery of, in particular, BCS Class IV drugs.

Characterization of an amino acid based biodegradable surfactant facilitating the incorporation of DNA into lipophilic delivery systems.

HYPOTHESIS: Lysine based cationic surfactants are well-tolerated tools for hydrophobic ion pairing (HIP) with DNA and its incorporation into lipophilic delivery systems. EXPERIMENTS: Di-Boc-lysine was esterified with 1-hexadecanol and the Boc-residues were cleaved off resulting in hexadecyl lysinate (HL). Subsequently, its Log POctanol/water and the critical micelle concentration (CMC) were determined. Degradability was evaluated utilizing trypsin and pancreas lipase as well as Caco-2 cells. Afterwards, the viability of Caco-2 cells upon incubation with HL was investigated. Finally, HL was ion-paired with plasmid DNA (pDNA, 6159 bp) and the obtained complex was incorporated into self-emulsifying drug delivery systems (SEDDS) for transfection studies on HEK-293 cells. FINDINGS: HL was synthesized with a yield of 53% and subsequent characterization revealed a Log PWater/Octanol of 0.05 and a CMC of 2.7 mM. Enzymatic degradation studies showed rapid degradation of HL by isolated enzymes and Caco-2 cells and cell viability experiments revealed no toxic effect of HL even in a concentration of 250 µg·ml-1 within 24 h. HIP with pDNA was the most efficient in a molar ratio of 6159:1 (HL:pDNA) equalling a charge ratio of 1:1. Formed complexes could be incorporated into SEDDS facilitating successful transfection of HEK-293 cells.

Less Reactive Thiol Ligands: Key towards Highly Mucoadhesive Drug Delivery Systems.

As less reactive s-protected thiomers can likely interpenetrate the mucus gel layer to a higher extent before getting immobilized via disulfide bond formation with mucins, it was the aim of this study to develop a novel type of s-protected thiomer based on the less reactive substructure cysteine-N-acetyl cysteine (Cys-NAC) in order to obtain improved mucoadhesive properties. For this purpose, two types of s-protected thiomers, polyacrylic acid-cysteine-mercaptonicotinic acid (PAA-Cys-MNA) and polyacrylic acid-cysteine-N-acetyl cysteine (PAA-Cys-NAC), were synthesized and characterized by Fourier-transform infrared spectroscopy (FT-IR) and the quantification of attached disulfide ligands. The viscosity of both products was measured in the presence of NAC and mucus. Both thiomers were also evaluated regarding swelling behavior, tensile studies and retention time on the porcine intestinal mucosa. The FT-IR spectra confirmed the successful attachment of Cys-MNA and Cys-NAC ligands to PAA. The number of attached sulfhydryl groups was in the range of 660-683 µmol/g. The viscosity of both s-protected thiomers increased due to the addition of increasing amounts of NAC. The viscosity of the mucus increased in the presence of 1% PAA-Cys-MNA and PAA-Cys-NAC 5.6- and 10.9-fold, respectively, in comparison to only 1% PAA. Both s-protected thiomers showed higher water uptake than unmodified PAA. The maximum detachment force (MDF) and the total work of adhesion (TWA) increased in the case of PAA-Cys-MNA up to 1.4- and 1.6-fold and up to 2.4- and 2.8-fold in the case of PAA-Cys-NAC. The retention of PAA, PAA-Cys-MNA, and PAA-Cys-NAC on porcine intestinal mucosa was 25%, 49%, and 76% within 3 h, respectively. The results of this study provide evidence that less reactive s-protected thiomers exhibit higher mucoadhesive properties than highly reactive s-protected thiomers.

Grafting of wool fibers through disulfide bonds: An advanced application of S-protected thiolated starch.

The purpose of this study is to develop a potential pathway for grafting polymers onto wool fibers based on thiol-disulfide exchange reactions. S-protected thiolated starch (PTS) was synthesized by coupling 3-(2-pyridyldithio) propanoic acid to starch through esterification, resulting in 417.3 ± 15.1 µmol ligand binding to 1 g of starch. PTS was labelled with fluorescein isothiocyanate (FITC) prior to grafting. Wool fibers were preactivated by raising the amount of thiol groups utilizing mild reducing agents. The highest degree of preactivation on the surface of wool fibers was achieved by a 0.2% (w/v) sodium borohydride and 1.5% (w/v) sodium bisulfite mixture pH 5.0 resulting in 182.6 ± 8.7 µmol thiol groups per gram of fibers. Different incubation times and ratios between FITC-labelled PTS and wool fibers were investigated. A graft yield of 58.5% was achieved at a ratio of 1:1.5 (w/w) between wool fibers and FITC-labelled PTS within 18 h of incubation. Successful coating of PTS on wool fibers was confirmed by confocal imaging, scanning electron microscopy and FT-IR. Mechanical properties of grafted wool fibers were tested regarding elongation and tensile strength. These results provide evidence for the potential of S-protected thiolated starch as a superior coating material for wool fibers.

Dual Inhibitory Action of a Novel AKR1C3 Inhibitor on Both Full-Length AR and the Variant AR-V7 in Enzalutamide Resistant Metastatic Castration Resistant Prostate Cancer.

The expanded use of second-generation antiandrogens revolutionized the treatment landscape of progressed prostate cancer. However, resistances to these novel drugs are already the next obstacle to be solved. Various previous studies depicted an involvement of the enzyme AKR1C3 in the process of castration resistance as well as in the resistance to 2nd generation antiandrogens like enzalutamide. In our study, we examined the potential of natural AKR1C3 inhibitors in various prostate cancer cell lines and a three-dimensional co-culture spheroid model consisting of cancer cells and cancer-associated fibroblasts (CAFs) mimicking enzalutamide resistant prostate cancer. One of our compounds, named MF-15, expressed strong antineoplastic effects especially in cell culture models with significant enzalutamide resistance. Furthermore, MF-15 exhibited a strong effect on androgen receptor (AR) signaling, including significant inhibition of AR activity, downregulation of androgen-regulated genes, lower prostate specific antigen (PSA) production, and decreased AR and AKR1C3 expression, indicating a bi-functional effect. Even more important, we demonstrated a persisting inhibition of AR activity in the presence of AR-V7 and further showed that MF-15 non-competitively binds within the DNA binding domain of the AR. The data suggest MF-15 as useful drug to overcome enzalutamide resistance.