Aerosol delivery and spatiotemporal tissue distribution of hydroxychloroquine in rat lung.

Inhalation enables the delivery of drugs directly to the lung, increasing the retention for prolonged exposure and maximizing the therapeutic index. However, the differential regional lung exposure kinetics and systemic pharmacokinetics are not fully known, and their estimation is critical for pulmonary drug delivery. The study evaluates the pharmacokinetics of hydroxychloroquine in different regions of the respiratory tract for multiple routes of administration. We also evaluated the influence of different inhaled formulations on systemic and lung pharmacokinetics by identifying suitable nebulizers followed by early characterization of emitted aerosol physicochemical properties. The salt- and freebase-based formulations required different nebulizers and generated aerosol with different physicochemical properties. An administration of hydroxychloroquine by different routes resulted in varied systemic and lung pharmacokinetics, with oral administration resulting in low tissue concentrations in all regions of the respiratory tract. A nose-only inhalation exposure resulted in higher and sustained lung concentrations of hydroxychloroquine with a lung parenchyma-to-blood ratio of 386 after 1440 min post-exposure. The concentrations of hydroxychloroquine in different regions of the respiratory tract (i.e., nasal epithelium, larynx, trachea, bronchi, and lung parenchyma) varied over time, indicating different retention kinetics. The spatiotemporal distribution of hydroxychloroquine in the lung is different due to the heterogeneity of cell types, varying blood perfusion rate, clearance mechanisms, and deposition of inhaled aerosol along the respiratory tract. In addition to highlighting the varied lung physiology, these results demonstrate the ability of the lung to retain increased levels of inhaled lysosomotropic drugs. Such findings are critical for the development of future inhalation-based therapeutics, aiming to optimize target site exposure, enable precision medicine, and ultimately enhance clinical outcomes.

Use of Capillary Aerosol Generator in Continuous Production of Controlled Aerosol for Non-Clinical Studies.

The capillary aerosol generator (CAG) is operated with the principal of thermal liquid evaporation through heating of e-liquid in the initial phase, followed by nucleation and condensation regulated through a mixture of airflow to generate aerosols, such as in an electronic cigarette (EC). The CAG is particularly useful in generating aerosols of large volumes in a continuous manner, for instances such as in vivo inhalation toxicology studies, where usage of ECs is not feasible. The thermal effects of generating aerosol from the CAG are similar in terms of temperature applied in an EC, thus allowing investigators to assess the vapors of e-liquids at scale and reproducibility. As the operation of the CAG allows users to control critical parameters such as the flow rate of e-liquid, heating temperatures and dilution air flows, it allows investigators to test various e-liquid formulations in a well-controlled device. Properties, such as aerosol particle size, are demonstrated to be regulated with the air flow rate with respect to the e-liquid flow and e-liquid composition. The CAG, however, is limited in assessing common EC-related issues, such as overheating of its elements. We seek to demonstrate that the CAG can generate aerosol that is reproducible and continuous, by assessing the chemical and physical aerosol characteristics with a chosen e-liquid formulation. The protocol describes the operating parameters of liquid flow rate, dilution air-flow rates and operating procedures needing to optimize the aerosol concentration and particle size required for an in vivo toxicology study. Presenting the representative results from the protocol and discussing the challenges and applications of working with a CAG, we demonstrate that CAG can be used in a reproducible fashion. The technology and protocol, that has been developed from prior work, serve as a foundation for future innovations for laboratory-controlled aerosol generation investigations.

Impact of whole-body versus nose-only inhalation exposure systems on systemic, respiratory, and cardiovascular endpoints in a 2-month cigarette smoke exposure study in the ApoE-/- mouse model.

Cigarette smoking is one major modifiable risk factor in the development and progression of chronic obstructive pulmonary disease and cardiovascular disease. To characterize and compare cigarette smoke (CS)-induced disease endpoints after exposure in either whole-body (WB) or nose-only (NO) exposure systems, we exposed apolipoprotein E-deficient mice to filtered air (Sham) or to the same total particulate matter (TPM) concentration of mainstream smoke from 3R4F reference cigarettes in NO or WB exposure chambers (EC) for 2 months. At matching TPM concentrations, we observed similar concentrations of carbon monoxide, acetaldehyde, and acrolein, but higher concentrations of nicotine and formaldehyde in NOEC than in WBEC. In both exposure systems, CS exposure led to the expected adaptive changes in nasal epithelia, altered lung function, lung inflammation, and pronounced changes in the nasal epithelial transcriptome and lung proteome. Exposure in the NOEC caused generally more severe histopathological changes in the nasal epithelia and a higher stress response as indicated by body weight decrease and lower blood lymphocyte counts compared with WB exposed mice. Erythropoiesis, and increases in total plasma triglyceride levels and atherosclerotic plaque area were observed only in CS-exposed mice in the WBEC group but not in the NOEC group. Although the composition of CS in the breathing zone is not completely comparable in the two exposure systems, the CS-induced respiratory disease endpoints were largely confirmed in both systems, with a higher magnitude of severity after NO exposure. CS-accelerated atherosclerosis and other pro-atherosclerotic factors were only significant in WBEC.

In Vivo Profiling of a Natural Alkaloid, Anatabine, in Rodents: Pharmacokinetics and Anti-Inflammatory Efficacy.

Natural alkaloids, a large class of plant-derived substances, have attracted considerable interest because of their pharmacological activities. In this study, the in vivo pharmacokinetics and anti-inflammatory profile of anatabine, a naturally occurring alkaloid, were characterized in rodents. Anatabine was found to be bioavailable and brain-penetrant following systemic administration. Following intraperitoneal (i.p.) administration (1, 2, and 5 mg/kg), anatabine caused a dose-dependent reduction in carrageenan-induced paw edema in rats; in mice, it inhibited the production of pro-inflammatory cytokines and simultaneously elevated the levels of an anti-inflammatory cytokine in a dose-dependent manner 2 h after lipopolysaccharide challenge. Furthermore, anatabine (∼10 and ∼20 mg/kg/day for 4 weeks; inhalation exposure) had effects in a murine model of multiple sclerosis, reducing neurological deficits and bodyweight loss. Comparative studies of the pharmacokinetics and anti-inflammatory activity of anatabine demonstrated its bioequivalence in rats following i.p. administration and inhalation exposure. This study not only provides the first detailed profile of anatabine pharmacokinetics in rodents but also comprehensively characterizes the anti-inflammatory activities of anatabine in acute and chronic inflammatory models. These findings provide a basis for further characterizing and optimizing the anti-inflammatory properties of anatabine.

E-vapor aerosols do not compromise bone integrity relative to cigarette smoke after 6-month inhalation in an ApoE-/- mouse model.

Cigarette smoke (CS) exposure is one of the leading risk factors for human health. Nicotine-containing inhalable products, such as e-cigarettes, can effectively support tobacco harm reduction approaches. However, there are limited comparative data on the effects of the aerosols generated from electronic vapor products (e-vapor) and CS on bone. Here, we report the effects of e-vapor aerosols and CS on bone morphology, structure, and strength in a 6-month inhalation study. Eight-week-old ApoE-/- mice were exposed to aerosols from three different e-vapor formulations-CARRIER (propylene glycol and vegetable glycerol), BASE (CARRIER and nicotine), TEST (BASE and flavor)-to CS from 3R4F reference cigarettes at matched nicotine concentrations (35 µg/L) or to fresh air (Sham) (N = 10 per group). Tibiae were analyzed for bone morphology by µCT imaging, biomechanics by three-point bending, and by histological analysis. CS inhalation caused a significant decrease in cortical and total bone volume fraction and bone density relative to e-vapor aerosols. Additionally, CS exposure caused a decrease in ultimate load and stiffness. In contrast, bone structural and biomechanical parameters were not significantly affected by e-vapor aerosol or Sham exposure. At the dissection time point, there was no significant difference in body weight or tibia bone weight or length among the groups. Histological findings revealed microcracks in cortical bone areas among all exposed groups compared to Sham control. In conclusion, because of the bone-preserving effect of e-vapor aerosols relative to CS exposure, e-vapor products could potentially constitute less harmful alternatives to cigarettes in situations in which bone health is of importance.

Evaluation of the Tobacco Heating System 2.2. Part 6: 90-day OECD 413 rat inhalation study with systems toxicology endpoints demonstrates reduced exposure effects of a mentholated version compared with mentholated and non-mentholated cigarette smoke.

The toxicity of a mentholated version of the Tobacco Heating System (THS2.2M), a candidate modified risk tobacco product (MRTP), was characterized in a 90-day OECD inhalation study. Differential gene and protein expression analysis of nasal epithelium and lung tissue was also performed to record exposure effects at the molecular level. Rats were exposed to filtered air (sham), to THS2.2M (at 15, 23 and 50 µg nicotine/l), to two mentholated reference cigarettes (MRC) (at 23 µg nicotine/l), or to the 3R4F reference cigarette (at 23 µg nicotine/l). MRCs were designed to meet 3R4F specifications. Test atmosphere analyses demonstrated that aldehydes were reduced by 75%-90% and carbon monoxide by 98% in THS2.2M aerosol compared with MRC smoke; aerosol uptake was confirmed by carboxyhemoglobin and menthol concentrations in blood, and by the quantities of urinary nicotine metabolites. Systemic toxicity and alterations in the respiratory tract were significantly lower in THS2.2M-exposed rats compared with MRC and 3R4F. Pulmonary inflammation and the magnitude of the changes in gene and protein expression were also dramatically lower after THS2.2M exposure compared with MRCs and 3R4F. No menthol-related effects were observed after MRC mainstream smoke-exposure compared with 3R4F.