Novel generic UPLC/MS/MS method for high throughput analysis applied to permeability assessment in early Drug Discovery

Mensch, J.; Noppe, M.; Adriaensen, Jef; Melis, A.; Mackie, Claire; Augustijns, Patrick; Brewster, Marcus E.

doi:10.1016/j.jchromb.2006.10.031

Cited by 70 publications

(51 citation statements)

References 17 publications

(17 reference statements)

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“…Predictive multiple reaction monitoring (MRM) is the most sensitive approach in metabolite identification [18,19]. The predictive MRM transitions of most of the metabolic pathways of BF are listed in supplemental online Table 3.…”

Section: Metabolite Identification and Validation In Hihsmentioning

confidence: 99%

Highly Efficient Differentiation of Functional Hepatocytes From Human Induced Pluripotent Stem Cells

Duan

Tschudy-Seney

et al. 2013

Stem Cells Translational Medicine

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Human induced pluripotent stem cells (hiPSCs) hold great potential for use in regenerative medicine, novel drug development, and disease progression/developmental studies. Here, we report highly efficient differentiation of hiPSCs toward a relatively homogeneous population of functional hepatocytes. hiPSC-derived hepatocytes (hiHs) not only showed a high expression of hepatocyte-specific proteins and liver-specific functions, but they also developed a functional biotransformation system including phase I and II metabolizing enzymes and phase III transporters. Nuclear receptors, which are critical for regulating the expression of metabolizing enzymes, were also expressed in hiHs. hiHs also responded to different compounds/inducers of cytochrome P450 as mature hepatocytes do. To follow up on this observation, we analyzed the drug metabolizing capacity of hiHs in real time using a novel ultraperformance liquid chromatography-tandem mass spectrometry. We found that, like freshly isolated primary human hepatocytes, the seven major metabolic pathways of the drug bufuralol were found in hiHs. In addition, transplanted hiHs engrafted, integrated, and proliferated in livers of an immune-deficient mouse model, and secreted human albumin, indicating that hiHs also function in vivo. In conclusion, we have generated a method for the efficient generation of hepatocytes from induced pluripotent stem cells in vitro and in vivo, and it appears that the cells function similarly to primary human hepatocytes, including developing a complete metabolic function. These results represent a significant step toward using patient/disease-specific hepatocytes for cell-based therapeutics as well as for pharmacology and toxicology studies. STEM CELLS TRANS- LATIONAL MEDICINE 2013;2:409 -419

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Section: Metabolite Identification and Validation In Hihsmentioning

confidence: 99%

Highly Efficient Differentiation of Functional Hepatocytes From Human Induced Pluripotent Stem Cells

Duan

Tschudy-Seney

et al. 2013

Stem Cells Translational Medicine

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“…On the one hand, non-UV detectable and very low soluble NPs can be difficult to analyse. For these specific issues, highly sensitive MS/MS detection methods can be adapted in order to analyse the PAMPA compartments using the same approach [52,53]. In this case, however, the analytical method would be compound-dependent and less adapted to a high-throughput and generic screening.…”

mentioning

confidence: 99%

Prediction of the Passive Intestinal Absorption of Medicinal Plant Extract Constituents with the Parallel Artificial Membrane Permeability Assay (PAMPA)

et al. 2016

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At the early drug discovery stage, the high-throughput parallel artificial membrane permeability assay is one of the most frequently used in vitro models to predict transcellular passive absorption. While thousands of new chemical entities have been screened with the parallel artificial membrane permeability assay, in general, permeation properties of natural products have been scarcely evaluated. In this study, the parallel artificial membrane permeability assay through a hexadecane membrane was used to predict the passive intestinal absorption of a representative set of frequently occurring natural products. Since natural products are usually ingested for medicinal use as components of complex extracts in traditional herbal preparations or as phytopharmaceuticals, the applicability of such an assay to study the constituents directly in medicinal crude plant extracts was further investigated. Three representative crude plant extracts with different natural product compositions were chosen for this study. The first extract was composed of furanocoumarins (Angelica archangelica), the second extract included alkaloids (Waltheria indica), and the third extract contained flavonoid glycosides (Pueraria montana var. lobata). For each medicinal plant, the effective passive permeability values Pe (cm/s) of the main natural products of interest were rapidly calculated thanks to a generic ultrahigh-pressure liquid chromatography-UV detection method and because Pe calculations do not require knowing precisely the concentration of each natural product within the extracts. The original parallel artificial membrane permeability assay through a hexadecane membrane was found to keep its predictive power when applied to constituents directly in crude plant extracts provided that higher quantities of the extract were initially loaded in the assay in order to ensure suitable detection of the individual constituents of the extracts. Such an approach is thus valuable for the high-throughput, cost-effective, and early evaluation of passive intestinal absorption of active principles in medicinal plants. In phytochemical studies, obtaining effective passive permeability values of pharmacologically active natural products is important to predict if natural products showing interesting activities in vitro may have a chance to reach their target in vivo.

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“…34,35) The UHPLC system employs sub-2 mm particles which enable very high resolution even at a high flow rate, but small filler particles cause extremely high pressure. Therefore, this system requires special devices which are robust enough to withstand high back-pressure.…”

Section: Mim Vs the Conventional Methodsmentioning

confidence: 99%

Development of a Novel High-Throughput Analytical Methodology, Multiple Injection Method, for Quantitative Analysis in Drug Metabolism and Pharmacokinetic Studies Using Liquid Chromatography with Tandem Mass Spectrometry

Tanaka

Ohkawa

Yasui

2011

Biological & Pharmaceutical Bulletin

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It is the need of the hour to speed up the development of new compounds in the pharmaceutical industry. At the early stages of drug development, many compounds are screened as rapidly as possible to identify the optimal lead compounds. Liquid chromatography-tandem mass spectrometry (LC-MS/MS) is essential for high-throughput quantitative analysis of the compounds. Because LC-MS/MS is more sensitive and selective for biological samples than other methods, it is popular in various fields, including pharmacokinetics and toxicokinetics. High-throughput analysis with LC-MS/MS plays a key role in speeding up drug discovery.In recent years, many developments in high-throughput analytical methods based on quantification by LC-MS/MS are reported to simplify sample preparation and speed up the analysis of the compounds.1-5) Among them, turbulent-flow chromatography, which uses turbulence caused by a high flow rate, is one method to separate the analyte from the biological matrices by the size-exclusion mechanism and hydrophobic interactions. 6,7) This method is advantageous in that it is applicable in a wide range of analyses by employing an online purification devise with column-switching systems. However, the technique requires specialized equipment and disposable cartridges, leading to high cost per sample. Monolithic chromatography is another example of the abovementioned developments. [8][9][10][11] This method employs the use of a monolithic column that consists of a unique stationary phase with high porosity due to the presence of double structures comprising mesopores and macropores, which enable the pressure in the column to be much lower than that in a conventional column of silica gel. Therefore, LC-MS/MS analysis can be carried out at a high flow rate using a monolithic column, speeding up the analysis. However, analysis of polar compounds such as metabolites and endogenous substances cannot be carried out using a monolithic column because the retention capability for these compounds is poorer than that of an octa decyl silyl (ODS) column and the lineup such as the bore diameter and length of monolithic column is not enough. Next, parallel LC-MS/MS method is also an option that could increase the throughput. [12][13][14] This method employs multiple HPLC systems and columns connected in parallel to an MS system, [15][16][17][18] and the eluted analyte from each HPLC system is alternately introduced into the MS system through a switching valve. Hence, the time spent before and after the elution of analytes can be diverted to detect the analytes eluted from other systems, thus considerably speeding up the analysis. However, this method requires specialized equipment and involves preparation of individual calibration curves for each system, even for the same analyte.In this study, we focused on maximizing the throughput of pharmacokinetic studies, and we developed a new method with conventional instruments, which lowers costs and effort. We named this approach multiple injection method (MIM), as multiple compound...

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Novel generic UPLC/MS/MS method for high throughput analysis applied to permeability assessment in early Drug Discovery

Cited by 70 publications

References 17 publications

Highly Efficient Differentiation of Functional Hepatocytes From Human Induced Pluripotent Stem Cells

Highly Efficient Differentiation of Functional Hepatocytes From Human Induced Pluripotent Stem Cells

Prediction of the Passive Intestinal Absorption of Medicinal Plant Extract Constituents with the Parallel Artificial Membrane Permeability Assay (PAMPA)

Development of a Novel High-Throughput Analytical Methodology, Multiple Injection Method, for Quantitative Analysis in Drug Metabolism and Pharmacokinetic Studies Using Liquid Chromatography with Tandem Mass Spectrometry

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