Cristal Therapeutics (Netherlands)

Core-crosslinked polymeric micelles (CCPMs) are an attractiveclassof nanocarriers for drug delivery. Two crosslinking approaches toform CCPMs exist: either via a low-molecular-weight crosslinking agentto connect homogeneous polymer chains with reactive handles or viacross-reactive handles on polymers to link them to each other (complementarypolymers). Previously, CCPMs based on methoxy poly(ethylene glycol)-b-poly[N-(2-hydroxypropyl) methacrylamide-lactate](mPEG-b-PHPMAmLacn) modifiedwith thioesters were crosslinked via native chemical ligation (NCL,a reaction between a cysteine residue and thioester resulting in anamide bond) using a bifunctional cysteine containing crosslinker.These CCPMs are degradable under physiological conditions due to hydrolysisof the ester groups present in the crosslinks. The rapid onset ofdegradation observed previously, as measured by the light scatteringintensity, questions the effectiveness of crosslinking via a bifunctionalagent. Particularly due to the possibility of intrachain crosslinksthat can occur using such a small crosslinker, we investigated thedegradation mechanism of CCPMs generated via both approaches usingvarious analytical techniques. CCPMs based on complementary polymersdegraded slower at pH 7.4 and 37 °C than CCPMs with a crosslinker(the half-life of the light scattering intensity was approximately170 h versus 80 h, respectively). Through comparative analysis ofthe degradation profiles of the two different CCPMs, we conclude thatpartially ineffective intrachain crosslinks are likely formed usingthe small crosslinker, which contributed to more rapid CCPM degradation.Overall, this study shows that the type of crosslinking approach cansignificantly affect degradation kinetics, and this should be takeninto consideration when developing new degradable CCPM platforms.

Polymeric micelles (PMs) are promising platforms for enhanced tissue targeting of entrapped therapeutic agents. Strategies to circumvent premature release of entrapped drugs include cross-linking of the micellar core as well as covalent attachment of the drug cargo. The chemistry employed to obtain cross-linked micelles needs to be mild to also allow entrapment of fragile molecules, such as certain peptides, proteins, oligonucleotides, and fluorescent dyes. Native chemical ligation (NCL) is a mild bio-orthogonal reaction between a N-terminal cysteine residue and a thioester that proceeds under physiological conditions. Here, we designed a trifunctional cross-linker containing two cysteine residues for the micelle core-cross-linking reaction and an azide residue for ring-strained alkyne conjugation of fluorescent dyes. We applied this approach to thermosensitive methoxypolyethylene glycol-b-N-(2-hydroxypropyl)methacrylamide-lactate (mPEG-b-HPMAmLacn) based block copolymers of a core-cross-linked polymeric micelle (CCPM) system by attaching thioester residues (using ethyl thioglycolate-succinic anhydride, ETSA) for NCL cross-linking with the trifunctional cross-linker under physiological conditions. By use of mild copper-free click chemistry, we coupled fluorescent dyes, Sulfo.Cy5 and BODIPY, to the core via the azide residue present on the cross-linker by triazole ring formation. In addition, we employed a recently developed cycloheptyne strain promoted click reagent (TMTHSI, CliCr) in comparison to the frequently employed cyclooctyne derivative (DBCO), both achieving successful dye entrapment. The size of the resulting CCPMs could be tuned between 50 and 100 nm by varying the molecular weight of the thermosensitive block and ETSA content. In vitro cell experiments showed successful internalization of the dye entrapped CCPMs, which did not affect cell viability up to a polymer concentration of 2 mg/mL in PC3 cells. These fluorescent dye entrapped CCPMs can be applied in diagnostic imaging and the chemistry developed in this study serves as a steppingstone toward covalently entrapped fragile drug compounds with tunable release in CCPMs.

Polymeric micelles are increasingly explored for tumor-targeted drug delivery. CriPec® technology enables the generation of core‐crosslinked polymeric micelles (CCPMs) based on thermosensitive (mPEG-b-pHPMAmLacn) block copolymers, with high drug loading capacity, tailorable size, and controlled drug release kinetics. In this study, we decorated clinical-stage CCPM with the αvβ3 integrin-targeted cyclic arginine-glycine-aspartic acid (cRGD) peptide, which is one of the most well-known active targeting ligands evaluated preclinically and clinically. Using a panel of cell lines with different expression levels of the αvβ3 integrin receptor and exploring both static and dynamic incubation conditions, we studied the benefit of decorating CCPM with different densities of cRGD. We show that incubation time and temperature, as well as the expression levels of αvβ3 integrin by target cells, positively influence cRGD-CCPM uptake, as demonstated by immunofluorescence staining and fluorescence microscopy. We demonstrate that even very low decoration densities (i.e., 1 mol % cRGD) result in increased engagement and uptake by target cells as compared to peptide-free control CCPM, and that high cRGD decoration densities do not result in a proportional increase in internalization. In this context, it should be kept in mind that a more extensive presence of targeting ligands on the surface of nanomedicines may affect their pharmacokinetic and biodistribution profile. Thus, we suggest a relatively low cRGD decoration density as most suitable for in vivo application.Graphical

The restrictive nature of the blood-brain barrier (BBB) prevents efficient treatment of many brain diseases. Focused ultrasound in combination with microbubbles has shown to safely and transiently increase BBB permeability. Here, the potential of Acoustic Cluster Therapy (ACT®), a microbubble platform specifically engineered for theranostic purposes, to increase the permeability of the BBB and improve accumulation of IRDye® 800CW-PEG and core-crosslinked polymeric micelles (CCPM) in the murine brain, was studied. Contrast enhanced magnetic resonance imaging (MRI) showed increased BBB permeability in all animals after ACT®. Near infrared fluorescence (NIRF) images of excised brains 1 h post ACT® revealed an increased accumulation of the IRDye® 800CW-PEG (5.2-fold) and CCPM (3.7-fold) in ACT®-treated brains compared to control brains, which was retained up to 24 h post ACT®. Confocal laser scanning microscopy (CLSM) showed improved extravasation and penetration of CCPM into the brain parenchyma after ACT®. Histological examination of brain sections showed no treatment related tissue damage. This study demonstrated that ACT® increases the permeability of the BBB and enhances accumulation of macromolecules and clinically relevant nanoparticles to the brain, taking a principal step in enabling improved treatment of various brain diseases.

CPC634 is a novel nanoparticle entrapping docetaxel, developed to enhance the intratumoral chemotherapy exposure. This randomized cross-over study compared the intratumoral and plasma pharmacokinetics of CPC634 with conventional docetaxel. Adult patients with solid tumors were randomized to receive CPC634 (75 mg/m2) in cycle 1, and conventional docetaxel (75 mg/m2) in cycle 2 or vice versa. The study was powered to identify a 25% increase of intratumoral total docetaxel exposure after CPC634 infusion compared with conventional docetaxel. Four patients were allocated per tumor sampling time point, that is, 24, 48, 72, and 96 hours, 7 and 14 days after infusion during both cycles. Total docetaxel and released docetaxel from the nanoparticle were determined in tumor tissue derived from a metastatic lesion and in plasma. Pharmacokinetic data were analyzed using linear mixed modeling. In total, 24 evaluable patients were included. In the tumor, CPC634 exhibited a 461% higher total docetaxel (P < 0.001) and a comparable released docetaxel concentration (P = 0.43). Plasma AUCinf was 27% higher (P = 0.001) and C max was 91% lower (P < 0.001) for CPC634 released docetaxel. The median observed neutrophil count nadir after conventional docetaxel treatment was lower (0.50 × 109/L) compared with CPC634 (4.30 × 109/L; P < 0.001). Here, we demonstrated that CPC634 enhanced the intratumoral total docetaxel exposure compared with conventional docetaxel. The lower incidence of neutropenia during CPC634 treatment is presumably related to lower plasma C max of released docetaxel. The unique pharmacokinetic profile of CPC634 nanoparticles has the potential to improve docetaxel treatment. A phase II efficacy trial of CPC634 is currently ongoing.

Polymerisierbare und hydrolytisch spaltbare Derivate von Dexamethason (DEX, roter Punkt) wurden kovalent in kernverknüpften Polymer-Micellen bestehend aus einem temperaturempfindlichen Blockcopolymer (gelber und grauer Baustein) eingeschlossen. Die Freisetzungsrate konnte über den Oxidationsgrad des Thioethers im Linker des Wirkstoffderivats gesteuert werden. Mit den DEX-Micellen wurde entzündliche rheumatische Arthritis in zwei Tiermodellen effizient behandelt.