Injectable Biodegradable Silica Depot for Controlled Subcutaneous Delivery of Antisense Oligonucleotides with beyond Monthly Administration.

Single-stranded antisense oligonucleotides (ASOs) are typically administered subcutaneously once per week or monthly. Less frequent dosing would have strong potential to improve patient convenience and increase adherence and thereby for some diseases result in more optimal therapeutic outcomes. Several technologies are available to provide sustained drug release via subcutaneous (SC) administration. ASOs have a high aqueous solubility and require relatively high doses, which limits the options available substantially. In the present work, we show that an innovative biodegradable, nonporous silica-based matrix provides zero-order release in vivo (rats) for at least 4 weeks for compositions with ASO loads of up to about 100 mg/mL (0.5 mL injection) without any sign of initial burst. This implies that administration beyond once monthly can be feasible. For higher drug loads, substantial burst release was observed during the first week. The concentrations of unconjugated ASO levels in the liver were found to be comparable to corresponding bolus doses. Additionally, infusion using a minipump shows a higher liver exposure than SC bolus administration at the same dose level and, in addition, clear mRNA knockdown and circulating protein reduction comparable to SC bolus dosing, hence suggesting productive liver uptake for a slow-release administration.

Nano- and microcrystals of griseofulvin subcutaneously administered to rats resulted in improved bioavailability and sustained release.

Griseofulvin is a commonly used antifungal agent which is administered per oral (p.o.). The oral administration route, however, shows rather low bioavailability. The aim of this study was to improve the bioavailability and to evaluate and interpret the pharmacokinetic profiles after subcutaneous (s.c.) administration of crystalline griseofulvin nano- and microsuspensions. Both formulations were injected at 5 and 500 µmol/kg to rats. For the lower concentration, the profiles were similar after s.c. injection but extended as compared to p.o. administration. For the higher concentration, injection of microsuspension resulted in a maintained plateau whereas the nanosuspension resulted in an obvious peak exposure followed by extended elimination. Both suspensions showed improved exposure with dose. The differences in peak exposures between nano- and microparticles, at the high dose, were mainly ascribed to differences in dissolution rate, experimentally determined in vitro, using spectroscopic methods. The extended appearance in the circulation may depend on the physicochemical properties of the compound and the physiological conditions at the injection site. The bioavailability was improved for both formulations compared with an orally administered nanosuspension, suggesting the s.c. route to be a preferred administration option for compounds with low oral bioavailability regarding both overall exposure and extended efficacy.

Subcutaneous administration of nano- and microsuspensions of poorly soluble compounds to rats.

The aim of the present study was to evaluate and interpret the pharmacokinetic profiles of two compounds after subcutaneous (s.c.) administration. The compounds have similar physicochemical properties, but are a base (BA99) and an acid (AC88), respectively. The compounds were administered as nano- (5 and 500 µmol/kg) and microsuspensions (5 µmol/kg) s.c. to Sprague-Dawley rats. At the low dose, the exposure was higher for both compounds administered as nanocrystals compared to microparticles. The high dose of the compounds resulted in even higher exposure, but not in a dose-linear manner. The differences in exposure between nano- and microparticles were mainly ascribed to higher dissolution rate and improved solubility for smaller particles. In addition to differences in exposure, there were also differences in the elimination pattern. After s.c. injection of 5 µmol/kg of BA99 as nano- and microsuspensions, the elimination profile was similar as observed earlier after oral administration. However, after injection of the higher dose of BA99 and all formulations of AC88, an extended elimination profile was observed, forming a maintained plateau under the investigated time-period. Essentially, constant plasma levels were caused by a balanced equilibrium between total body clearance of the drug and supply rate of drug from the formulations.

Evaluation of exposure properties after injection of nanosuspensions and microsuspenions into the intraperitoneal space in rats.

In the present paper, BA99 and AC88 were used as model compounds for intraperitoneal (i.p.) administration to Sprague-Dawley rats. A major problem for the compounds, like many others newly developed pharmaceutical drugs, is the poor solubility in water. To solve solubility related problems, development of nanosuspensions is an attractive alternative. Both compounds are suitable for nanosuspensions, using the milling approach. After 2 weeks in freezer, the nanoparticles aggregated to form particles in the 400-2000 nm interval. However, following a 20 s ultrasonication step, the original particle sizes (about 200 nm) were obtained. Adding 5% mannitol before the samples were frozen abolished aggregation. It is also possible to freeze-dry the nanosuspension in the presence of 5% mannitol and re-disperse the formulation in water. Nanosuspensions of both compounds were injected i.p. to rats at 5 and 500 µmoL/kg. At the low dose, also a microsuspension was administered. I.p. administration resulted in overall improved C(max) for both AC88 and BA99 compared to s.c. and oral administration. I.p. is the preferred route of administration of tolerable drugs when a fast onset of action is desired and when a significant first passage metabolism occurs. The net charge of the active molecule appeared to affect the absorption kinetics. In the present work, the neutral molecule was favored over the negatively charged one.

Multiomics analysis of naturally efficacious lipid nanoparticle coronas reveals high-density lipoprotein is necessary for their function.

In terms of lipid nanoparticle (LNP) engineering, the relationship between particle composition, delivery efficacy, and the composition of the biocoronas that form around LNPs, is poorly understood. To explore this we analyze naturally efficacious biocorona compositions using an unbiased screening workflow. First, LNPs are complexed with plasma samples, from individual lean or obese male rats, and then functionally evaluated in vitro. Then, a fast, automated, and miniaturized method retrieves the LNPs with intact biocoronas, and multiomics analysis of the LNP-corona complexes reveals the particle corona content arising from each individual plasma sample. We find that the most efficacious LNP-corona complexes were enriched with high-density lipoprotein (HDL) and, compared to the commonly used corona-biomarker Apolipoprotein E, corona HDL content was a superior predictor of in-vivo activity. Using technically challenging and clinically relevant lipid nanoparticles, these methods reveal a previously unreported role for HDL as a source of ApoE and, form a framework for improving LNP therapeutic efficacy by controlling corona composition.

Co-administration of a nanosuspension of a poorly soluble basic compound and a solution of a proton pump inhibitor--the importance of gastrointestinal pH and solubility for the in vivo exposure.

In Sigfridsson et al. (2011, Drug Dev Ind Pharm, 37:243-251), there was no difference in plasma concentration of BA99 when administering the drug as nanosuspension or microsuspension and analyzing the blood samples by liquid chromatography-mass spectrometry. This was related to the dissolved amount of drug in the gastric tract, which was high enough to support fast absorption when the drug reached the small intestine. One single physicochemical property (pK(a), about 3 for BA99) abolished the benefit of small particles. These results were further confirmed in the present study, where a proton pump inhibitor, AZD0865, was used to elevate the gastric pH and then drastically decreased the gastric solubility. In this way, BA99 could be considered as a model compound for a neutral substance. By increasing the gastric pH to 5-6 and 8-9, respectively, in rats, the plasma concentrations of BA99, after administering nanosuspensions, were unchanged compared with untreated (i.e. no AZD0865) animals. For microsuspensions of the test compound, on the other hand, the exposure of BA99 was 2- to 3-fold lower than for nanosuspensions at both pHs. Moreover, the blood concentrations of BA99 administered as microsuspension were also 2- to 3-fold lower compared with untreated (no AZD0865) individuals receiving both nanoparticles and microparticles of BA99. Obviously, for neutral compounds, with similar physicochemical properties as the present compound, size reduction will be crucial for increased plasma exposure. For basic compounds, with similar physicochemical properties as the present compound, the crucial step for absorption is the dissolution and solubility in the gastric tract.

Particle size reduction and pharmacokinetic evaluation of a poorly soluble acid and a poorly soluble base during early development.

AIM: The aim of the present study was to find out if nanosuspensions were a better choice compared with microsuspensions, for the present substances with water solubility in the order of 2-3 µM (pH 6.8, small intestinal pH) and no permeability limitations. The ambition was also to understand what the higher solubility in the stomach for BA99 means in terms of absorption properties of the substance. METHOD: The pharmacokinetic parameters of a poorly soluble acid (AC88) and a poorly soluble base (BA99) administered orally as nanosuspensions have been compared with those from microsuspensions using rat as in vivo species. RESULTS: A significant difference was observed between the two suspensions for AC88 already at the lowest dose, 5 µmol/kg (the particle size of the nanosuspensions and the microsuspensions was about 200 nm and 14 µm, respectively). These results were further confirmed at a high dose (500 µmol/kg). However, for BA99, there were no significant differences between the two formulations at any dose investigated (the particle size of the nanosuspensions and the microsuspensions was about 280 nm and 12 µm, respectively). CONCLUSIONS: The study demonstrated a clear correlation between particle size and in vivo exposures for an acidic compound, the nanosuspensions providing the highest exposure. For a basic compound, on the other hand, with the present properties and doses, a microsuspension was sufficient. In the latter case, the higher solubility at gastric pH, because of the basic pK(a), limits the need for particle reduction.

A candidate drug administered subcutaneously to rodents as drug particles showing hepatic recirculation which influenced the sustained release process.

The aim of the present study was to evaluate and interpret the pharmacokinetic profiles after subcutaneous (s.c.) administration of crystalline AZ'72 nano- and microsuspensions to rodents. Both formulations were injected at 1.5 and 150 mg/kg to rats. For the lower dose, the profiles were similar after s.c. injection but extended as compared to oral administration. The overall exposure was higher for nanoparticles compared with microparticles during the investigated period. For the higher dose, injection of both suspensions resulted in maintained plateaus caused by the drug depots but, unexpectedly, at similar exposure levels. After addition of a further stabilizer, pluronic F127, nanosuspensions showed improved exposure with dose and higher exposure compared to larger particles in mice. Obviously, a stabilizer mixture that suits one delivery route is not necessarily optimal for another one. The differences in peak concentration (Cmax) between nano- and microparticles were mainly ascribed to differences in dissolution rate. Plasma profiles in mice showed curves with secondary absorption peaks after intravenous and oral administration, suggesting hepatic recirculation following both administration routes. This process, together with the depot formulation, complicates the analysis of absorption from s.c. administration, i.e. multiple processes were driving the plasma profile of AZ'72.

Particle size reduction for improvement of oral absorption of the poorly soluble drug UG558 in rats during early development.

BACKGROUND: The exposure of UG558 was not good enough using traditional microsuspensions. AIM: The aim of this study was to find out whether nanosuspensions were a better choice compared with a microsuspension, for an acidic substance with a water solubility in the order of 2 microM (pH 6.8, small intestinal pH) and no permeability limitations. METHODS: UG558 was ground by a planetary ball mill. The particle size was measured by laser diffraction and the stability of the particle sizes was followed. The pharmacokinetic parameters of UG558 administered as nanosuspension have been compared with those from microsuspension using rat as in vivo specie. Both formulations were administered orally. The nanosuspension was also administered intravenously. RESULTS: The particle size of the nanosuspensions was about 190 nm and about 12 microm for the microsuspensions. At the administered doses, solutions were no alternative (e.g. due to limited solubility). Already at the lowest dose, 5 micromol/kg (5 ml/kg), a significant difference was observed between the two suspensions. These results were further confirmed at a high dose (500 micromol/kg, 5 mL/kg). Thus, the study demonstrated a clear correlation between particle size and in vivo exposures, where the nanosuspensions provided the highest exposure. Furthermore, no adverse events were observed for the substance nor the nanosuspension formulations (i.e., the particles) in spite of the higher exposures obtained with the nanoparticles. To make it possible to calculate the bioavailability, 5 micromol/kg doses of the nanosuspensions (5 ml/kg) were also administered intravenously. No adverse events were observed. CONCLUSIONS: The nanoparticles have a larger surface, resulting in faster in vivo dissolution rate, faster absorption, and increased bioavailability, compared to microparticles. The lower overall bioavailability observed at the high dose, compared with the low dose, was due to a combination of low dissolution rate, low solubility, and a narrow intestinal absorption window for UG558.

Mass Spectrometry Imaging proves differential absorption profiles of well-characterised permeability markers along the crypt-villus axis.

Knowledge about the region-specific absorption profiles from the gastrointestinal tract of orally administered drugs is a critical factor guiding dosage form selection in drug development. We have used a novel approach to study three well-characterized permeability and absorption marker drugs in the intestine. Propranolol and metoprolol (highly permeable compounds) and atenolol (low-moderate permeability compound) were orally co-administered to rats. The site of drug absorption was revealed by high spatial resolution matrix-assisted laser desorption ionization mass spectrometry imaging (MALDI-MSI) and complemented by quantitative measurement of drug concentration in tissue homogenates. MALDI-MSI identified endogenous molecular markers that illustrated the villi structures and confirmed the different absorption sites assigned to histological landmarks for the three drugs. Propranolol and metoprolol showed a rapid absorption and shorter transit distance in contrast to atenolol, which was absorbed more slowly from more distal sites. This study provides novel insights into site specific absorption for each of the compounds along the crypt-villus axis, as well as confirming a proximal-distal absorption gradient along the intestine. The combined analytical approach allowed the quantification and spatial resolution of drug distribution in the intestine and provided experimental evidence for the suggested absorption behaviour of low and highly permeable compounds.