Quality attributes of CTVad1, a nanoemulsified adjuvant for phase I clinical trial of SpiN COVID-19 vaccine.

Aim: To develop, characterize and evaluate an oil/water nanoemulsion with squalene (CTVad1) to be approved as an adjuvant for the SpiN COVID-19 vaccine clinical trials. Materials & methods: Critical process parameters (CPPs) of CTVad1 were standardized to meet the critical quality attributes (CQAs) of an adjuvant for human use. CTVad1 and the SpiN-CTVad1 vaccine were submitted to physicochemical, stability, in vitro and in vivo studies. Results & conclusion: All CQAs were met in the CTVad1 production process. SpiN- CTVad1 met CQAs and induced high levels of antibodies and specific cellular responses in in vivo studies. These results represented a critical step in the process developed to meet regulatory requirements for the SpiN COVID-19 vaccine clinical trial.

A new oral self-emulsifying drug delivery system improves the antileishmania efficacy of fexinidazole in vivo.

The aim of this study was to develop, characterize and evaluate the in vivo oral efficacy of self-emulsifying drug delivery systems (SEDDS) containing fexinidazole (FEX) in the experimental treatment of visceral leishmaniasis (VL). The developed FEX-SEDDS formulation presented as a clear, yellowish liquid, with absence of precipitate. The droplet size, polydispersion index and zeta potential after dilution in water (1:200) was of 91 ± 3 nm, 0.242 ± 0.005 and -16.7 ± 0.2, respectively. In the simulated gastric and intestinal media, the FEX-SEDDS had a size of 97 ± 1 and 106 ± 9 nm, respectively. The FEX retention in droplet after SEDDS dilution in simulated gastrointestinal media was almost 100 %. Antileishmanial efficacy studies showed that FEX-SEDDS was the only treatment able to significantly (p < 0.05) reduce the parasite burden in the liver and spleen of animals experimentally infected with Leishmania infantum. Our intestinal permeability data suggest that FEX-SEDDS showed no evidence of injury to the intestinal mucosa. These findings suggest that FEX-SEDDS can be a promising oral alternative for the treatment of VL caused by L. infantum.

pH-sensitive doxorubicin-tocopherol succinate prodrug encapsulated in docosahexaenoic acid-based nanostructured lipid carriers: An effective strategy to improve pharmacokinetics and reduce toxic effects.

Side effects often limit the use of doxorubicin (DOX) in cancer treatment. We have recently developed a nanostructured lipid carrier (NLC) formulation for synergistic chemotherapy, encapsulating DOX and the anticancer adjuvants docosahexaenoic acid (DHA) and α-tocopherol succinate (TS). Hydrophobic ion-pairing with TS allowed a high DOX entrapment in the nanocarrier. In this work, we investigated the pharmacokinetics of this formulation after intravenous administration in mice. The first data obtained led us to propose synthesizing covalent DOX-TS conjugates to increase DOX retention in the NLC. We successfully conjugated DOX to TS via an amide or hydrazone bond. In vitro studies in 4T1 tumor cells indicated low cytotoxicity of the amide derivative, while the hydrazone conjugate was effective in killing cancer cells. We encapsulated the hydrazone derivative in a DHA-based nanocarrier (DOX-hyd-TS/NLC), which had reduced particle size and high drug encapsulation efficiency. The pH-sensitive hydrazone bond allowed controlled DOX release from the NLC, with increased drug release at acidic conditions. In vivo studies revealed that DOX-hyd-TS/NLC had a better pharmacokinetic profile than free DOX and attenuated the short-term cardiotoxic effects caused by DOX, such as QT prolongation and impaired left ventricular systolic function. Moreover, this formulation showed excellent therapeutic performance by reducing tumor growth in 4T1 tumor-bearing mice and decreasing DOX-induced toxicity to the heart and liver, demonstrated by hematologic, biochemical, and histologic analyses. These results indicate that DOX-hyd-TS/NLC may be a promising nanocarrier for breast cancer treatment.

Nanomedicine in Oncocardiology: Contribution and Perspectives of Preclinical Studies.

Cancer and cardiovascular diseases are the leading causes of death and morbidity worldwide. Strikingly, cardiovascular disorders are more common and more severe in cancer patients than in the general population, increasing incidence rates. In this context, it is vital to consider the anticancer efficacy of a treatment and the devastating heart complications it could potentially cause. Oncocardiology has emerged as a promising medical and scientific field addressing these aspects from different angles. Interestingly, nanomedicine appears to have great promise in reducing the cardiotoxicity of anticancer drugs, maintaining or even enhancing their efficacy. Several studies have shown the benefits of nanocarriers, although with some flaws when considering the concept of oncocardiology. Herein, we discuss how preclinical studies should be designed as closely as possible to clinical protocols, considering various parameters intrinsic to the animal models used and the experimental protocols. The sex and age of the animals, the size and location of the tumors, the doses of the nanoformulations administered, and the acute vs. the long-term effects of treatments are essential aspects. We also discuss the perspectives offered by non-invasive imaging techniques to simultaneously assess both the anticancer effects of treatment and its potential impact on the heart. The overall objective is to accelerate the development and validation of nanoformulations through high-quality preclinical studies reproducing the clinical conditions.

Co-delivery of doxorubicin, docosahexaenoic acid, and α-tocopherol succinate by nanostructured lipid carriers has a synergistic effect to enhance antitumor activity and reduce toxicity.

Doxorubicin (DOX) is widely used in cancer treatment, however, its use is often limited due to its side effects. To avoid these shortcomings, the encapsulation of DOX into nanocarriers has been suggested. Herein, we proposed a novel nanostructured lipid carrier (NLC) formulation loading DOX, docosahexaenoic acid (DHA), and α-tocopherol succinate (TS) for cancer treatment. DHA is an omega-3 fatty acid and TS is a vitamin E derivative. It has been proposed that these compounds can enhance the antitumor activity of chemotherapeutics. Thus, we hypothesized that the combination of DOX, DHA, and TS in NLC (NLC-DHA-DOX-TS) could increase antitumor efficacy and also reduce toxicity. NLC-DHA-DOX-TS was prepared using emulsification-ultrasound. DOX was incorporated after preparing the NLC, which prevented its degradation during manufacture. High DOX encapsulation efficiency was obtained due to the ion-pairing with TS. This ion-pairing increases lipophilicity of DOX and reduces its crystallinity, contributing to its encapsulation in the lipid matrix. Controlled DOX release from the NLC was observed in vitro, with increased drug release at the acidic environment. In vitro cell studies indicated that DOX, DHA, and TS have synergistic effects against 4T1 tumor cells. The in vivo study showed that NLC-DHA-DOX-TS exhibited the greatest antitumor efficacy by reducing tumor growth in 4T1 tumor-bearing mice. In addition, this formulation reduced mice mortality, prevented lung metastasis, and decreased DOX-induced toxicity to the heart and liver, which was demonstrated by hematologic, biochemical, and histologic analyses. These results indicate that NLC-DHA-DOX-TS may be a promising carrier for breast cancer treatment.

Cleaning Validation for Residual Estimation of Mometasone Furoate on Stainless-Steel Surface of Pharmaceutical Manufacturing Equipment Using a UHPLC-UV Method.

Cleaning validation is the documented evidence that shows the effectiveness of cleaning procedures for the removal of product residues and other contaminants. The cleaning procedures must be validated and methods to determine trace amounts of drugs have to be considered with special attention. An ultra-high-performance liquid chromatography-ultraviolet (UHPLC-UV) method for the determination of mometasone furoate residues on stainless-steel surfaces was developed and validated in order to control a cleaning procedure. The chromatography separation was achieved on a Waters Acquity UPLC HSS T3 column (50 × 2.1 mm, 1.8 µm) at 40°C using acetonitrile and water (1:1, v/v) as the mobile phase at a flow rate of 0.5 mL/min. The injection volume was 2 µL, and the detection was performed at 254 nm. The swab and rinse procedures were optimized in order to obtain a recovery higher than 90% of mometasone furoate from stainless-steel surfaces, using ethanol as the extraction solvent. The method was validated in the range of 0.2-2.6 µg/mL and showed appropriate selectivity, limit of detection and quantification, linearity, precision, accuracy, and robustness. This method was found to be simple, fast, and sensitive for determination of mometasone furoate residues and, therefore, can be used for cleaning validation analysis.

Stability-indicating HPLC method for determination of amiodarone hydrochloride and its impurities in tablets: a detailed forced degradation study

Resumo Amiodarone hydrochloride is one of the most important drugs used to treat arrhythmias. The USP monograph for amiodarone hydrochloride describes an HPLC method for the quantification of seven impurities, however, this method shows problems that result in unresolved peaks. Moreover, there is no monograph for tablets in this compendium. Thus, a stability indicating HPLC method was developed for the determination of amiodarone, its known impurities and degradation products in tablets. A detailed forced degradation study was performed submitting amiodarone API, tablets and placebo to different stress conditions: acid and alkaline hydrolysis, oxidation, metal ions, heat, humidity, and light. Amiodarone hydrochloride API was susceptible to degradation in all stress conditions. The tablets also showed degradation in all environments, except in acidic condition. The analytes separation and quantification were achieved on an Agilent Zorbax Eclipse XDB-C18 column (100 x 3.0 mm, 3.5 µm). The mobile phase was composed of 50 mM acetate buffer pH 5.5 (A) and a mixture of methanol-acetonitrile (3:4, v/v) (B) in gradient elution. The method was validated in the range of 350-650 µg/mL for assay and 10-24 µg/mL for impurities determination. Therefore, this method can be used both for stability studies and routine quality control analyses.

Stability-indicating HPLC-UV for the determination of 4-bromomethyl-3-nitrobenzoic acid, a bioactive nitrocompound.

The nitroaromatic compound 4-bromomethyl-3-nitrobenzoic acid (ANB) is a promising antitumoral agent whose activity has recently been investigated. Forced degradation studies were conducted on ANB with a high-performance liquid chromatography-ultraviolet assay to establish its stability and selectivity. ANB was subjected to degradation studies under hydrolytic (acid and alkaline), oxidative and light exposition conditions. The compound showed greater lability only in acid and alkaline conditions by forming a major degradation product. The chromatographic separation of ANB and its degradation product was achieved on an octadecylsilane column using a mobile phase of methanol-water (80:20, v/v), pH 4.0 adjusted with formic acid, flow rate of 0.7 mL/min, ultraviolet diode array detection at λ 271 nm, injection volume of 20 µL and temperature of 30°C. The method was validated according to International Conference on Harmonization guidelines with respect to precision (average relative standard deviation 0.67%), accuracy (average 99.97%), linearity (y = 118730x + 12912; coefficient of determination > 0.999), specificity and robustness (p > 0.05). The degradation product formed as a result from the hydrolysis of nitrobenzyl bromide, corresponded to its benzyl alcohol, 4-hydroxymethyl-3-nitrobenzoic acid (ANOH) and was characterized by co-elution with a synthetic ANOH working standard.

Evaluation of antitumor activity and development of solid lipid nanoparticles of metronidazole analogue.

Nitroheterocyclic compounds have received considerable interest as hypoxia-selective cytotoxins (HSC) for cancer treatment. In the present study, we investigated antitumor activity of an iodide analogue of metronidazole, 1-(2-iodoethyl)-2-methyl-5-nitroimidazole (MTZ-I), using Swiss mice bearing solid Ehrlich tumor. MTZ-I showed potent anti-cancer activity at a dose of 40 mg/kg. MTZ-I loaded solid lipid nanoparticles (SLN) were developed as an alternative colloidal carrier system to enhance tumor drug uptake. SLN were characterized for particle size, polydispersity index, zeta potential and entrapment efficiency. In addition, the influence of presence of the cationic lipid stearylamine (STE) on stability of formulation was assessed. The results of DSC study showed that MTZ-I exhibited interaction with STE.