Development Of Phospholipidosis Research Articles

Chromatography approaches for early screening of the phospholipidosis-inducing potential of pharmaceuticals

Drug-induced phospholipidosis (PLD) is an excessive accumulation of polar phospholipids within cells as a result of medical treatment. Even though a direct link between in vitro drug-induced PLD and toxicity in humans has not yet been firmly established, the development of PLD during preclinical testing in animals is a recognized problem in the pharmaceutical industry and can delay or abort the development process. Therefore, it is of interest to investigate the potential PLD risk of candidates at an early stage of the drug discovery process. In this work, a high throughput physicochemical approach, which is based on measuring the retention factors of the test compound within several different separation systems, was developed for screening phospholipidosis-inducing potential (PLIP) of pharmaceuticals. The measured retention factors of 36 drugs were compared with literature data on PLIP risk from three different sources. It is clearly shown that there is a statistical correlation between the chromatographic retention parameters of tested drugs and their PLIP risk. In conclusion, the retention factor (log k AOT ) observed on a docusate sodium salt (AOT) surfactant vesicle electrokinetic chromatography (EKC) system and the logarithm of the volume of distribution (log V d ) calculated from immobilized artificial membrane chromatography at pH 7.4 (CHI IAM 7.4) and HSA binding value (% HSA) can provide primary profile prediction for a large number of drug candidates early in the drug discovery process with minimal resources. The observations are that the higher the value of both log k AOT and log V d , the higher the PLIP risk, and we would recommend this dual approach.

Predicting Phospholipidosis Using Machine Learning

Phospholipidosis is an adverse effect caused by numerous cationic amphiphilic drugs and can affect many cell types. It is characterized by the excess accumulation of phospholipids and is most reliably identified by electron microscopy of cells revealing the presence of lamellar inclusion bodies. The development of phospholipidosis can cause a delay in the drug development process, and the importance of computational approaches to the problem has been well documented. Previous work on predictive methods for phospholipidosis showed that state of the art machine learning methods produced the best results. Here we extend this work by looking at a larger data set mined from the literature. We find that circular fingerprints lead to better models than either E-Dragon descriptors or a combination of the two. We also observe very similar performance in general between Random Forest and Support Vector Machine models.

In vitro assays and biomarkers for drug-induced phospholipidosis

Drug-induced phospholipidosis is the cytoplasmic accumulation of phospholipids as a result of xenobiotic exposure. This accumulation results in a unique histological effect in cells noted as electron-dense lamellar inclusions or whorls in the cytoplasm when observed with transmission electron microscopy. Electron microscopy has been the widely accepted standard for classification of the phospholipidosis effect. Molecules that have been prone to induce such an effect are made up of a lipophilic region and a positively charged region. Phospholipidosis has most commonly been associated with drugs that are cationic, amphiphilic drugs and can occur in a variety of tissues. Although phospholipidosis is not considered adverse in isolation, depending on the tissue affected or the occasional circumstance of concurrent toxicity, phospholipidosis can be perplexing if identified in early drug development. In most circumstances, characterisation of the effect with in vivo studies allows for determination of exposure and the magnitude of the effect. On occasion in drug development, there may be an interest to screen early stage compounds to minimise phospholipidosis. In such circumstances, in silico and in vitro assays can be employed in a strategy with in vivo assessments. In addition, there may be an interest to monitor for the potential development of phospholipidosis in longer-term animal studies. In such cases, biomarker approaches could be used. The challenge in the overall assessment of phospholipidosis remains the question of the possible relevance to any toxicity, and, therefore, any screening approach, while assessing the potential to induce phospholipidosis, must be considered in relation to prediction of findings in vivo. The status of current assays and biomarkers is presented with strategies for screening.

A cell-based approach for the early assessment of the phospholipidogenic potential in pharmaceutical research and drug development.

Phospholipidosis is a term commonly used to indicate a phospholipid storage disorder; in affected cells, phospholipids accumulate in lysosomes that acquire a multilamellar morphological appearance. Cationic amphiphilic drugs (CADs) are suggested to induce phospholipidosis by direct interaction of xenobiotics with intracellular phospholipids or by the action of xenobiotics on the synthesis and metabolism of phospholipids. To date, electron microscopy (EM) represents the most reliable and the preferred method for the demonstration of phospholipidotic cell damage. Nevertheless, EM has a low throughput, it is expensive, and it is not suitable for screening purposes. We discuss here the assessment of the the phospholipidogenic potential of drugs using a cell culture-based model. In this test, intracellular phospholipids of treated U-937 cells (a human monocyte-derived cell line) were measured using the fluorescent probe Nile red. Eleven CADs reported to induce phospholipidosis in vivo and eight nonphospholipidogenic drugs were tested. Results obtained with the U-937 model confirmed the phospholipidogenic potential of drugs tested as described in the literature. Results have also been correlated with data obtained with a physical-chemical model (chromatographic hydrophobicity index measurement). Good correlation was obtained, confirming that the physical-chemical properties of CADs play a crucial role in the development of phospholipidosis. This work demonstrates that the U-937 model is a rapid and sensitive method for the determination of phospholipidosis-mediated cell damage. The specificity and the predictive potency observed make this method suitable for screening purposes in pharmaceutical development.

Amiodarone-induced Phospholipidosis in Rat Alveolar Macrophages

Humans treated with the antiarrhythmic drug amiodarone may develop pulmonary toxicity accompanied by the presence of alveolar macrophages (AM) containing lamellar inclusions. This cellular response is indicative of the development of a drug-induced phospholipidosis. To characterize this response of the AM, Fischer-344 rats were treated with amiodarone, and the macrophages were recovered by pulmonary lavage. The development of phospholipidosis was dose- and time-dependent and was reversible. Daily treatment for 1 wk (5 days/wk) at 150 mg/kg resulted in a 5-fold increase in total phospholipid in the cells. Phospholipid levels were increased only slightly more through 9 wk of treatment. Cells were filled with lamellar inclusions and contained areas of amorphous granular and membranous material. Individual classes of phospholipids were all increased during the development of phospholipidosis. When expressed as mumol/10(7) cells, phosphatidylcholine demonstrated the largest increase. Levels of amiodarone and its major metabolite, desethylamiodarone, increased in AM in parallel with the increase in phospholipid. From 3 days through 9 wk of treatment, the level of desethylamiodarone was always higher than that of amiodarone. Treatment with desethylamiodarone also induced phospholipidosis in AM. Administration of phenobarbital along with amiodarone for 1 wk caused a reduction in the levels of amiodarone, desethylamiodarone, and phospholipid in the cells. The molar ratio of amiodarone to phospholipid was decreased, whereas the molar ratio of desethylamiodarone to phospholipid remained unchanged. Taken together, the results indicate that, along with amiodarone, desethylamiodarone and/or its metabolites may play an important role in the phospholipidosis induced in AM when rats are treated with amiodarone.

Experimental studies on nephrotoxicity of aminoglycosides at low doses: Mechanisms and perspectives

The nephrotoxicity of aminoglycosides is an important consideration in the clinical use of these agents. The underlying biochemical and cytologic mechanisms have, therefore, been the subject of many extensive studies. Few experimental studies, however, have employed doses relevant to those used in clinical practice. This article focuses on data obtained using such a “low-dose” approach. Aminoglycosides accumulate specifically in the lysosomes of proximal tubular cells by a process of adsorptive pinocytosis; in the lysosomes, they induce a phospholipidosis that has been studied in vivo and in vitro (e.g., by computer-aided conformational analysis). A similar phospholipidosis is also observed in humans. The overall nephrotoxicity of aminoglycosides results from a combination of their intrinsic ability to bind to phospholipids and the extent of their renal uptake. The development of phospholipidosis is accompanied by focal necroses, tubular regeneration, and interstitial proliferation, even at low, therapeutic doses of these agents. These changes are related to the nephrotoxic potential of the aminoglycosides. Of all 2-deoxystreptamine-containing aminoglycosides tested so far, and by all criteria, amikacin is associated with the least dramatic alterations. Together, these approaches may provide a rational basis to study and compare the nephrotoxicity of aminoglycosides at clinically relevant dosages. They may also serve for the screening and design of aminoglycosides with lower toxicity than those currently available.