Exploring Isolated Perfused Rat Lung technology: a pharmacokinetic comparison between powder and nanosuspension

Exploring Isolated Perfused Rat Lung technology: a pharmacokinetic comparison between powder and nanosuspension Introduction The ex vivo isolated perfused lung (IPL) model has emerged as a valuable tool for studying the pharmacokinetics of various compounds administered via different formulations intratracheally [1]. This technique involves the isolation of the lung from the rest of the body while maintaining its physiological conditions through perfusion with an appropriate buffer and excluding the effects of systemic deposition [2]. The use of rodent IPL offers numerous advantages for studying uptake kinetics, including the ability to mimic in vivo conditions while providing greater control over experimental variables [3]. It also allows the construction of lung pharmacokinetic curves reducing the number of animals used and limiting the procedure on live animals, thanks to the sampling of the perfusate at pre-established intervals after administration. This work focuses on a study using the ex vivo rat IPL technique to analyse the pharmacokinetics of a specific-poorly-water soluble compound, administered as a nanosuspension and as powder intratracheally. Through this research, we aim to clarify the influence of the formulation on the pulmonary absorption kinetics, especially related to the device used for administration which impacts the pulmonary deposition of the drug. In fact, the aim of this research is to compare the pharmacokinetic profile of the same compound in the form of nanosuspension and powder obtained with the isolated perfused lung following administration with the Hamilton syringe and Penncentury device, respectively. This analysis therefore allows comparative pharmacokinetic studies to clarify the impact of the different administration methods on the absorption of the drug, due to the different deposition in the lung. Materials and methods The experiments were performed using an isolated perfused lung system (Harvard, Massachusetts, US) for rat lung. A Krebs-Henseleit buffer added of albumin (Sigma Aldrich) and indometacin (Alfasigma) was fluxed at 0.04 ml/g body wt/min and thermostated at 37 °C. Respiratory acts were set at 70 acts/min with the positive pressure in the ventilation pump. After the surgical removal of lungs, heart and trachea from rat, the organs were positioned in the system and buffer was fluxed for 30 minutes before the administration. A micronized powder or a nanosuspension of the same compound were administered intratracheally with PennCentury (PennCentury Inc.) and Hamilton Syringe, respectively, setting 0.6 mg/lung as delivered dose for both formulations. At established timepoints (1,5', 3', 4.5', 6', 7.5', 9', 10.5', 15', 30', 45', 60', 80', 100') after administration the perfusate was collected for one minute. At the end of the experiments the rat lungs and trachea were homogenized to analyse the drug remained in the lung and in the main airways. The samples were analysed by HPLC-MS (Sciex 6500, MA, USA). Results The results of the experiments are presented in the graph below, expressed as drug concentration found in the perfusate during the time. The nanosuspension profile achieves the maximum concentration and then it decreases during the time, while the powder administered by Penncentury shows an oscillating trend without a pick of maximum concentration. The comparison of the two pharmacokinetic profiles highlights the influence of the administration device on the pharmacokinetic profile of the same compound. In fact, the powder certainly presents a non-homogeneous deposition of the drug at the lung level. The intratracheal administration of the powder requires that only a part reaches the lung district, and since the compound under analysis is poorly-water soluble, it is difficult to determine a pharmacokinetic profile with the isolated perfused lung technique, which has shown a fluctuating absorption profile. The nanosuspension, on the other hand, reaches the lung area completely, distributing itself uniformly and showing a pulmonary absorption profile with a trend that follows the pharmacokinetic studies performed with plasma samples. Furthermore, it is likely that the dissolution kinetics of the two formulations is different since it could be influenced by the formulation form (liquid vs. solid) and the particle size of the substance (nanometric vs micrometric). This study highlights the importance of the type of formulation and the device with which pulmonary drugs are administered, which in turn influence deposition at the pulmonary level and therefore the bioavailable amount. It is important to consider these aspects since they influence the results in terms of absorbed and bioavailable concentrations. Conclusion The isolated perfused lung model has proven to be very useful for conducting different types of analysis for pulmonary compounds, in particular pharmacokinetics, building absorption profiles that could potentially mirror those obtained in vivo with plasma samples, with the use of a minimum of 3 animals for all the desired timepoints. The type of device used for the intratracheal administration of the compounds, which depends on the formulation used, influences the deposition of the compound at the lung level and consequently also the dissolved fraction, ready to be absorbed, which is collected by the perfusate. In fact, it was evident that a nanosuspension, compared to the powder, showed a higher bioavailable concentration and a maximum concentration of available drug and then this decreases linearly over time. The powder, on the other hand, probably due to the uneven distribution in the lung district and possibly a slower dissolution profile, presents a non-linear bioavailable drug profile over time. Probably by managing to administer the powder with other devices, which guarantee uniform pulmonary distribution, it would be possible to have a release of drug in the perfusate which after a maximum concentration decreases linearly over time. References 1. Eriksson J. et al. AAPS J. 22, 71 (2020) 2. Cárdenes, N. et al. Sci Rep 11, 490 (2021). 3. McNeil, S. E. Methods Mol Biol. 697, 3-8. (2010).