On the Production of Chitosan-Coated Polycaprolactone Nanoparticles in a Confined Impinging Jet Reactor

This work is focused on the synthesis of polycaprolactone nanoparticles, coated with chitosan, in a confined impinging jet reactor using the solvent displacement method. The role of the various reacting species was investigated, evidencing that a biocompatible polymer, for example, polycaprolactone, is required to support chitosan to obtain a monomodal particle size distribution, with low particle diameters. A surfactant is required to reduce the nanoparticle size (down to a mean diameter of about 260 nm) and obtain a positive zeta potential (about +31 mV), perfectly suitable for pharmaceutical applications. Different surfactants were tested, and Poloxamer 388 appeared to be preferable to polyvinyl alcohol. The effect of the concentration of Poloxamer 388 (in the range 0.5-5 mg mL^?1) and of chitosan (in the range 1.5-5 mg mL^?1) on both the mean particle size and zeta potential was also investigated, evidencing that chitosan concentration has the strongest effect on both parameters. Finally, the effect of solvent evaporation, quenching and feed flow rate was investigated, showing that the evaporation stage does not affect particle characteristics, quenching is required to avoid particle aggregation, and a minimum liquid flow rate of 80 mL min^?1 is required in the considered reactor to minimize the particle size.     

Prediction of Human Brain Penetration of P-glycoprotein and Breast Cancer Resistance Protein Substrates Using In Vitro Transporter Studies and Animal Models

Four P-glycoprotein (P-gp) and breast cancer resistance protein (BCRP) substrates with human cerebrospinal fluid (CSF) concentrations and preclinical neuropharmacokinetics were used to assess in vitro–in vivo extrapolation of brain penetration in preclinical species and the ability to predict human brain penetration. Unbound brain (C_b,u), unbound plasma (C_p,u), and CSF compound concentrations (C_CSF) were measured in rats and nonhuman primates (NHPs), and the unbound partition coefficients (C_b,u/C_p,u and C_CSF/C_p,u) were used to assess brain penetration. The results indicated that for P-gp and BCRP dual substrates, brain penetration was severally impaired in all species. In comparison, for P-gp substrates that are weak or non-BCRP substrates, improved brain penetration was observed in NHPs and humans than in rats. Overall, NHP appears to be more predictive of human brain penetration for P-gp substrates with weak or no interaction with BCRP than rat. Although C_CSF does not quantitatively correspond to C_b,u for efflux transporter substrates, it is mostly within 3-fold higher of C_b,u in rat and NHP, suggesting that C_CSF can be used as a surrogate for C_b,u. Taken together, a holistic approach including both in vitro transporter and in vivo neuropharmacokinetics data enables a better estimation of human brain penetration of P-gp/BCRP substrates.     

Long-Acting Profile of 4 Drugs in 1 Anti-HIV Nanosuspension in Nonhuman Primates for 5 Weeks After a Single Subcutaneous Injection

Daily oral antiretroviral therapy regimens produce limited drug exposure in tissues where residual HIV persists and suffer from poor patient adherence and disparate drug kinetics, which all negatively impact outcomes. To address this, we developed a tissue- and cell-targeted long-acting 4-in-1 nanosuspension composed of lopinavir (LPV), ritonavir, tenofovir (TFV), and lamivudine (3TC). In 4 macaques dosed subcutaneously, drug levels over 5 weeks in plasma, lymph node mononuclear cells (LNMCs), and peripheral blood mononuclear cells (PBMCs) were analyzed by liquid chromatography–tandem mass spectrometry. Plasma and PBMC levels of the active drugs (LPV, TFV, and 3TC) were sustained for 5 weeks; PBMC exposures to LPV, ritonavir, and 3TC were 12-, 16-, 42-fold higher than those in plasma. Apparent T_1/2z of LPV, TFV, and 3TC were 219.1, 63.1, and 136.3 h in plasma; 1045.7, 105.9, and 127.7 h in PBMCs. At day 8, LPV, TFV, and 3TC levels in LNMCs were 4.1-, 5.0-, and 1.9-fold higher than in those in PBMCs and much higher than in plasma. Therefore, 1 dose of a 4-drug nanosuspension exhibited persistent drug levels in LNMCs, PBMCs, and plasma for 5 weeks. With interspecies scaling and dose adjustment, this 4-in-1 HIV drug-combination could be a long-acting treatment with the potential to target residual virus in tissues and improve patient adherence.     

Coformulation of Broadly Neutralizing Antibodies 3BNC117 and PGT121: Analytical Challenges During Preformulation Characterization and Storage Stability Studies

In this study, we investigated analytical challenges associated with the formulation of 2 anti-HIV broadly neutralizing antibodies (bnAbs), 3BNC117 and PGT121, both separately at 100 mg/mL and together at 50 mg/mL each. The bnAb formulations were characterized for relative solubility and conformational stability followed by accelerated and real-time stability studies. Although the bnAbs were stable during 4°C storage, incubation at 40°C differentiated their stability profiles. Specific concentration-dependent aggregation rates at 30°C and 40°C were measured by size exclusion chromatography for the individual bnAbs with the mixture showing intermediate behavior. Interestingly, although the relative ratio of the 2 bnAbs remained constant at 4°C, the ratio of 3BNC117 to PGT121 increased in the dimer that formed during storage at 40°C. A mass spectrometry-based multiattribute method, identified and quantified differences in modifications of the Fab regions for each bnAb within the mixture including clipping, oxidation, deamidation, and isomerization sites. Each bnAb showed slight differences in the levels and sites of lysine residue glycations. Together, these data demonstrate the ability to differentiate degradation products from individual antibodies within the bnAb mixture, and that degradation rates are influenced not only by the individual bnAb concentrations but also by the mixture concentration.     

Intradermal Delivery of a Near-Infrared Photosensitizer Using Dissolving Microneedle Arrays

Nodular basal cell carcinoma is a deep skin lesion and one of the most common cancers. Conventional photodynamic therapy is limited to treatment of superficial skin lesions. The parenteral administration of near-IR preformed photosensitizers suffers from poor selectivity and may result in prolonged skin photosensitivity. Microneedles (MNs) can provide localized drug delivery to skin lesions. Intradermal delivery of the preformed near-IR photosensitizer; 5,10,15,20-tetrakis(2,6-difluoro-3-N-methylsulfamoylphenyl bacteriochlorin (Redaporfin?) using dissolving MN was successful in vitro and in vivo. MN demonstrated complete dissolution 30 min after skin application and showed sufficient mechanical strength to penetrate the skin to a depth of 450 μm. In vitro deposition studies illustrated that the drug was delivered and detected down to 5 mm in skin. In vivo biodistribution studies in athymic nude mice Crl:NU(NCr)-Foxn1^nu showed both fast initial release and localized drug delivery. The MN-treated mice showed a progressive decrease in the fluorescence intensity at the application site over the 7-day experiment period, with the highest and lowest fluorescence intensities measured being 9.2 × 10¹⁰ ± 2.5 × 10¹⁰ and 3.8 × 10⁹ ± 1.6 × 10⁹ p/s, respectively. By day 7, there was some migration of fluorescence away from the site of initial MN application. However, the majority of the body surfaces showed fluorescence levels that were comparable to those seen in the negative control group. This work suggests utility for polymeric MN arrays in minimally invasive intradermal delivery to enhance photodynamic therapy of deep skin lesions.     

Dibucaine in Ionic-Gradient Liposomes: Biophysical,Toxicological, and Activity Characterization

Administration of local anesthetics is one of the most effective pain control techniques for postoperative analgesia. However, anesthetic agents easily diffuse into the injection site, limiting the time of anesthesia. One approach to prolong analgesia is to entrap local anesthetic agents in nanostructured carriers (e.g., liposomes). Here, we report that using an ammonium sulphate gradient was the best strategy to improve the encapsulation (62.6%) of dibucaine (DBC) into liposomes. Light scattering and nanotracking analyses were used to characterize vesicle properties, such as, size, polydispersity, zeta potentials, and number. In vitro kinetic experiments revealed the sustained release of DBC (50% in 7 h) from the liposomes. In addition, in vitro (3T3 cells in culture) and in vivo (zebrafish) toxicity assays revealed that ionic-gradient liposomes were able to reduce DBC cyto/cardiotoxicity and morphological changes in zebrafish larvae. Moreover, the anesthesia time attained after infiltrative administration in mice was longer with encapsulated DBC (27 h) than that with free DBC (11 h), at 320 μM (0.012%), confirming it as a promising long-acting liposome formulation for parenteral drug administration of DBC.     

Health service utilization,unmet healthcare needs,and the potential of telemedicine services among Korean expatriates

Background

With the significant growth of migration and expatriation, facilitated by increased global mobility, the number of Koreans living abroad as of 2016 is approximately 7.4 million (15% of the Korean population). Healthcare utilization or health problems, especially among expatriates in developing countries, have not been well researched despite the various health risks these individuals are exposed to. Consequently, we identified the health utilization patterns and healthcare needs among Korean expatriates in Vietnam, Cambodia, and Uzbekistan.

Methods

This cross-sectional survey examined 429 Korean expatriates living in Vietnam (n?=?208), Cambodia (n?=?60), and Uzbekistan (n?=?161) who had access to the Internet and were living abroad for at least 6?months. A 67-item questionnaire was used, and feedback was received via an online survey program. Stepwise logistic regression analyses were performed to evaluate factors associated with unmet healthcare needs and preferences of certain type of telemedicine.

Results

We found that 45.5% (195/429) of respondents had used medical services in their country of stay. Among those who visited health institutions >?3 times, the most popular choice was general hospitals (39.4%, 15/38); however, they initially visited Korean doctors’ or local doctors’ offices. The most essential criteria for healthcare service facilities was a “skilled professional” (39.3%, 169/429), 42% wanted a health program for chronic disease management, and 30% wanted specialized internal medicine. A substantial number wanted to access telemedicine services and were willing to pay for this service. They were particularly interested in experts’ second opinion (61.5%, 264/429) and quick, 24-h medical consultations (60.8%, 261/429). Having unmet healthcare needs and being younger was strongly associated with all types of telemedicine networks.

Conclusions

Nearly half of the expatriates in developing countries had unmet healthcare needs. Telemedicine is one potential solution to meet these needs, especially in developing countries.

     

A high frequency missense SULT1B1 allelic variant (L145V) selectively expressed in African descendants exhibits altered kinetic properties

1.?Human cytosolic sulfotransferase 1B1 (SULT1B1) sulfates small phenolic compounds and bioactivates polycyclic aromatic hydrocarbons. To date, no SULT1B1 allelic variants have been well-characterized.

2.?While cloning SULT1B1 from human endometrial specimens, an allelic variant resulting in valine instead of leucine at the 145th amino acid position (L145V) was detected. NCBI reported this alteration as the highest frequency SULT1B1 allelic variant.

3.?L145V frequency comprised 9% of 37 mixed-population human patients and was specific to African Americans with an allelic frequency of 25%. Structurally, replacement of leucine with valine potentially destabilizes a conserved helix (α8) that forms the “floor” of both the substrate and PAPS binding domains. This destabilization results in altered kinetic properties including a four-fold decrease in affinity for PAP (3′, 5′-diphosphoadenosine). K_ms for 3′-phosphoadenosine- 5′-phosphosulfate (PAPS) are similar; however, maximal turnover rate of the variant isoform (0.86?pmol/(min*μg)) is slower than wild-type (WT) SULT1B1 (1.26?pmol/(min*μg)). The L145V variant also displays altered kinetics toward small phenolic substrates, including a diminished p-nitrophenol K_m and increased susceptibility to 1-naphthol substrate inhibition.

4.?No significant correlation between genotype and prostate or colorectal cancer was observed in patients; however, the variant isoform could underlie specific pathologies in sub-Saharan African carriers.