Reliable, experimentally determined partition coefficient P (logP) for most drugs are often unavailable in the literature. Many values are from in silico predictions and may not accurately reflect drug lipophilicity. In this study, a robust, viable, and resource sparing method to measure logP was developed using reverse phase high performance liquid chromatography (RP-HPLC). The logP of twelve common drugs was measured using calibration curves at pH 6 and 9 that were created using reference standards with well-established logP. The HPLC method reported here can be used for high throughput estimation of logP of commonly used drugs. LogP values here showed general agreement with the other few HPLC-based literature logP values available. Additionally, the HPLC-based logP values found here agreed partially with literature logP values found using other methodologies (±10%). However, there was no strong agreement since there are few experimentally determined literature logP values. This paper shows a facile method to estimate logP without using octanol or computational approaches. This method has excellent promise to provide reliable logP values of commonly used drugs available in literature. A larger pool of reliable logP values of commonly drugs has promise to improve quality of medicinal chemistry and pharmacokinetic (PK) models.
Many stroke survivors suffer long‐term functional disabilities with no effective treatment option available. For that reason, we tested the potential effects of enhancing noradrenergic and serotoninergic systems in combination with delayed voluntary running on recovery after stroke in mice. To accomplish this, we used adult male mice (n=12/group) and divided them into sham, stroke, vehicle, or drug‐treated groups with either selective norepinephrine reuptake inhibitor atomoxetine (0.3, 1 mg/kg, once a day i.p.) or selective serotonin reuptake inhibitor fluoxetine (3, 10 mg/kg, once a day i.p.) starting from day 5 after stroke and continued for 12 days. For evaluation of motor function, we used grid walking and cylinder tests 3 days prior and 3, 7, 14, 28, 42 days after using ischemic stroke model (photothrombosis). At 42 days after stroke, mice brains were collected either after fixing the brain through cardiac perfusion (for infarct size measurements and immunohistochemistry experiments after brain sectioning) or as a fresh tissue from the motor cortex area (for immunoblotting experiments).The results of both motor function tests showed that all stroke‐subjected groups had comparable and substantial motor impairment on day 3 after stroke when compared to their baseline and that voluntary running group did not significantly improve throughout the days after stroke. However, administrating either atomoxetine or fluoxetine along with exercise promoted motor recovery at 42 days after stroke dose‐dependently with no significant changes seen in infarct size among groups (0.5 ± 0.1 mm 3 ). Further, immunoblotting experiment showed no significant changes amongst groups in synaptophysin, an integral membrane glycoprotein of neuronal synaptic vesicles, nor with PSD‐95, an excitatory postsynaptic density scaffolding protein, after normalization to beta actin in the ischemic hemisphere. However, the immunohistochemistry data showed that atomoxetine and fluoxetine treatment significantly decreased the expression of parvalbumin (PV), a calcium binding albumin protein associated with depressing synapses, in the boundaries of medial agranular cortex and medial frontal cortex areas of the ischemic hemisphere when compared to their corresponding area in the healthy hemisphere. Whereas no significant relative changes seen with the expression of PV showed between the healthy and ischemic hemispheres of sham, stroke, and vehicle groups. Our data showed that low duration voluntary running does not facilitate motor recovery independently after ischemic stroke in mice. But it promoted recovery of motor function only when combined with atomoxetine or fluoxetine in a dose‐dependent manner. Our data showed a significant decrease of the inhibitory interneurons PV concomitant with the recovery seen with atomoxetine and fluoxetine treatment. Our ongoing experiments include the evaluation of growth associated protein (GAP‐43) and glutamate receptors expression. Support or Funding Information Research in the Karamyan laboratory is supported by R01NS106879 Enhanced motor recovery of mice treated with atomoxetine or fluoxetine after an experimental stroke in the grid walking test (A) and the cylinder test (B). Figure 1 Atomoxetine and fluoxetine treatment significantly decreased parvalbumin expression in the ischemic hemisphere compared to the healthy hemisphere on 42 days after stroke. Figure 2
This study leverages physiologically based biopharmaceutics modeling (PBBM) to predict the clinical performance of two itraconazole (ITRA) amorphous solid dispersions (ASDs), Sempera® and Tolsura®, under fasted and fed state conditions, exploring the potential of PBBM in predicting formulation-specific food interactions. The ITRA formulations were subjected to extensive in vitro biopharmaceutical testing, including solubility studies and dissolution tests under fasted and fed state conditions, revealing significant differences in dissolution behaviors between Sempera® and Tolsura®. The impact of food and hypochlorhydria on drug absorption was evaluated using a stepwise mechanistic deconvolution-reconvolution PBBM approach, integrating fundamental parameters based on the in vitro data into the final model. Our model not only successfully predicted the effects of acid reducing agents (ARA) and food on the oral absorption of ITRA, but also captured the between-subject variability, demonstrating the utility of this approach in understanding the complex interplay between drug, formulation, and gastrointestinal environment. Most importantly, the PBBM was able to accurately predict the positive impact of food on the absorption of Sempera® and the negative food effect of Tolsura®. The findings highlight the importance of considering formulation characteristics and gastrointestinal physiology, underscoring the potential of PBBM in bioequivalence (BE) assessment of generic formulations under varying physiological conditions, including in the fed state and in hypochlorhydric patients. The successful application of this stepwise and mechanistic PBBM approach suggests a potential pathway for streamlining drug development and may contribute to more informed decision-making for BE assessment.
Abstract Purpose The goal was to assess, for lipophilic drugs, the impact of logP on human volume of distribution at steady-state (VD ss ) predictions, including intermediate fut and Kp values, from six methods: Oie-Tozer, Rodgers-Rowland (tissue-specific Kp and only muscle Kp), GastroPlus, Korzekwa-Nagar, and TCM-New. Method A sensitivity analysis with focus on logP was conducted by keeping pKa and fup constant for each of four drugs, while varying logP. VD ss was also calculated for the specific literature logP values. Error prediction analysis was conducted by analyzing prediction errors by source of logP values, drug, and overall values. Results The Rodgers-Rowland methods were highly sensitive to logP values, followed by GastroPlus and Korzekwa-Nagar. The Oie-Tozer and TCM-New methods were only modestly sensitive to logP. Hence, the relative performance of these methods depended upon the source of logP value. As logP values increased, TCM-New and Oie-Tozer were the most accurate methods. TCM-New was the only method that was accurate regardless of logP value source. Oie-Tozer provided accurate predictions for griseofulvin, posaconazole, and isavuconazole; GastroPlus for itraconazole and isavuconazole; Korzekwa-Nagar for posaconazole; and TCM-New for griseofulvin, posaconazole, and isavuconazole. Both Rodgers-Rowland methods provided inaccurate predictions due to the overprediction of VD ss . Conclusions TCM-New was the most accurate prediction of human VD ss across four drugs and three logP sources, followed by Oie-Tozer. TCM-New showed to be the best method for VD ss prediction of highly lipophilic drugs, suggesting BPR as a favorable surrogate for drug partitioning in the tissues, and which avoids the use of fup.
Abstract Previous studies documented up‐regulation of peptidase neurolysin (Nln) after brain ischemia, however, the significance of Nln function in the post‐stroke brain remained unknown. The aim of this study was to assess the functional role of Nln in the brain after ischemic stroke. Administration of a specific Nln inhibitor Agaricoglyceride A (AgaA) to mice after stroke in a middle cerebral artery occlusion model, dose‐dependently aggravated injury measured by increased infarct and edema volumes, blood–brain barrier disruption, increased levels of interleukin 6 and monocyte chemoattractant protein‐1, neurological and motor deficit 24 h after stroke. In this setting, AgaA resulted in inhibition of Nln in the ischemic hemisphere leading to increased levels of Nln substrates bradykinin, neurotensin, and substance P. AgaA lacked effects on several physiological parameters and appeared non‐toxic to mice. In a reverse approach, we developed an adeno‐associated viral vector (AAV2/5‐CAG‐Nln) to overexpress Nln in the mouse brain. Applicability of AAV2/5‐CAG‐Nln to transduce catalytically active Nln was confirmed in primary neurons and in vivo . Over‐expression of Nln in the mouse brain was also accompanied by decreased levels of its substrates. Two weeks after in vivo transduction of Nln using the AAV vector, mice were subjected to middle cerebral artery occlusion and the same outcome measures were evaluated 72 h later. These experiments revealed that abundance of Nln in the brain protects animals from stroke. This study is the first to document functional significance of Nln in pathophysiology of stroke and provide evidence that Nln is an endogenous mechanism functioning to preserve the brain from ischemic injury. image
The blood-brain barrier (BBB) is a fundamental component of the central nervous system (CNS). Its functional and structural integrity is vital to maintain the homeostasis of the brain microenvironment by controlling the passage of substances and regulating the trafficking of immune cells between the blood and the brain. The BBB is primarily composed of highly specialized microvascular endothelial cells. These cells' special features and physiological properties are acquired and maintained through the concerted effort of hemodynamic and cellular cues from the surrounding environment. This complex multicellular system, comprising endothelial cells, astrocytes, pericytes, and neurons, is known as the neurovascular unit (NVU). The BBB strictly controls the transport of nutrients and metabolites into brain parenchyma through a tightly regulated transport system while limiting the access of potentially harmful substances via efflux transcytosis and metabolic mechanisms. Not surprisingly, a disruption of the BBB has been associated with the onset and/or progression of major neurological disorders. Although the association between disease and BBB disruption is clear, its nature is not always evident, specifically with regard to whether an impaired BBB function results from the pathological condition or whether the BBB damage is the primary pathogenic factor prodromal to the onset of the disease. In either case, repairing the barrier could be a viable option for treating and/or reducing the effects of CNS disorders. In this review, we describe the fundamental structure and function of the BBB in both healthy and altered/diseased conditions. Additionally, we provide an overview of the potential therapeutic targets that could be leveraged to restore the integrity of the BBB concomitant to the treatment of these brain disorders.
In this study, we investigated the potential of two histone deacetylase (HDAC) inhibitors, panobinostat and entinostat, to enhance recovery of motor function after ischemic stroke in CD-1 male mice. Panobinostat, which is a pan-HDAC inhibitor, is an FDA-approved drug for certain cancers, whereas entinostat is a class-I HDAC inhibitor and is widely expected to get approval for clinical use in near future. Stroke was induced by photothrombosis and the drugs were administered intraperitoneally every other day (panobinostat at 10 mg/kg or 30 mg/kg; entinostat at 5 mg/kg or 15 mg/kg) starting from day 5 to day 15 after stroke. The control group received only vehicle following the same regimen. In addition, all drug or vehicle-treated mice exercised for 2 hours (voluntary wheel running) starting from day 9 to day 41 after stroke. Additional control groups included sham-operated animals, and mice which had stroke but were not drug/vehicle-treated or allowed to run. Motor function of the mice was evaluated by blinded investigators using gridwalk and cylinder tests before and after stroke. Acetylation of histone 3 in the peri-infarct region of the brain samples was measured by immunoblotting. Our results indicate that lower dose of both panobinostat and entinostat marginally improved motor function in mice by day 42 after stroke, although it did not reach statistical significance. Notably, this improvement trend was lost with the higher dose of both drugs which also showed some toxicity. No statistical difference was observed in running distance of mice among experimental groups. Likewise, we did not observe statistically significant difference in stroke volumes among the experimental groups. Immunoblotting experiments indicated that both panobinostat and entinostat dose-dependently increased the level of acetylated histone in the cortical, peri-infarct region of drug-treated animals compared to the non-treated groups. In summary, our results indicate that both panobinostat and entinostat do not facilitate improvement of motor function after stroke at the tested doses. However, it is likely that lower doses of these drugs may enhance recovery of motor function after stroke.
