High-throughput respirometric assay identifies predictive toxicophore of mitochondrial injury

Wills, Lauren P.; Beeson, Gyda; Trager, Richard E.; Lindsey, Christopher C.; Beeson, Craig C.; Peterson, Yuri K.; Schnellmann, Rick G.

doi:10.1016/j.taap.2013.06.014

Cited by 19 publications

(15 citation statements)

References 43 publications

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“…With the recognition that cellular OCR measures are useful for in vitro drug testing (Schoonen et al,2009(Schoonen et al, , 2012Wang et al, 2015;Wills et al, 2013), other OCR methods are available in static, closed systems in which oxygen depletion is measured within a well-stirred vessel containing a suspension of cells or isolated mitochondria (Jacobus et al, 1982). Although the precision and sensitivity of such systems is good, they are limited by a short duration of OCR measurement (1-2 h at most), and a low throughput (2 experiments can typically be carried out concomitantly).…”

Section: Comparison Of Our Flow Culture System To Static (Nonflowing)mentioning

confidence: 99%

Quantification of Low-Level Drug Effects Using Real-Time,in vitroMeasurement of Oxygen Consumption Rate

et al. 2015

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There is a general need to detect toxic effects of drugs during preclinical screening. We propose that increased sensitivity of xenobiotics toxicity combined with improved in vitro physiological recapitulation will more accurately assess potentially toxic perturbations of cellular biochemistry that are near in vivo pharmacological exposure levels. Importantly, measurement of such cytopathologies avoids activating mechanisms mediating toxicity at suprapharmacologic levels not relevant to in vivo effects. We present a sensitive method to measure changes in oxygen consumption rate (OCR), a well-established parameter reflecting a potential hazard, in response to exposure to pharmacologic levels of drugs using a flow culture system and state of the art oxygen sensing system. We tested metformin and acetaminophen on rat liver slices to illustrate the method. The features of the method include continuous and very stable measurement of OCR over the course of 48 h in liver slices in a continuous flow chamber with the ability to resolve changes as small as 0.3%/h. Kinetic modeling of metformin inhibition of OCR over a wide range of concentrations revealed both a slow and fast mechanism, where the fast mechanism activated only at concentrations above 0.6 mM. For both drugs, small amounts of inhibition were reversible, but higher decrements were irreversible. Overall the study highlights the advantages of measuring low-level toxicity so as to avoid the common extrapolations made about drug toxicity based on effects of drugs tested at suprapharmacologic levels.

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Section: Comparison Of Our Flow Culture System To Static (Nonflowing)mentioning

confidence: 99%

Quantification of Low-Level Drug Effects Using Real-Time,in vitroMeasurement of Oxygen Consumption Rate

et al. 2015

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“…Statistical methods, such as Fisher’s exact test, can be used to define associations between the mechanisms of mitochondrial toxicity and side effects. In addition to enhanced insight into mitochondrial toxicity, the recent rapid development of machine learning technologies can be used to analyze and predict side effects [ 41 , 42 ] or identify toxicophores that are predictive of mitochondrial toxicity [ 43 ].…”

Section: Discussionmentioning

confidence: 99%

MitoTox: a comprehensive mitochondrial toxicity database

Lin

Huang

et al. 2021

BMC Bioinformatics

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Background Mitochondria play essential roles in regulating cellular functions. Some drug treatments and molecular interventions have been reported to have off-target effects damaging mitochondria and causing severe side effects. The development of a database for the management of mitochondrial toxicity-related molecules and their targets is important for further analyses. Results To correlate chemical, biological and mechanistic information on clinically relevant mitochondria-related toxicity, a comprehensive mitochondrial toxicity database (MitoTox) was developed. MitoTox is an electronic repository that integrates comprehensive information about mitochondria-related toxins and their targets. Information and data related to mitochondrial toxicity originate from various sources, including scientific journals and other electronic databases. These resources were manually verified and extracted into MitoTox. The database currently contains over 1400 small-molecule compounds, 870 mitochondrial targets, and more than 4100 mitochondrial toxin-target associations. Each MitoTox data record contains over 30 fields, including biochemical properties, therapeutic classification, target proteins, toxicological data, mechanistic information, clinical side effects, and references. Conclusions MitoTox provides a fully searchable database with links to references and other databases. Potential applications of MitoTox include toxicity classification, prediction, reference and education. MitoTox is available online at http://www.mitotox.org.

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“…Mitochondrial toxicity is now widely accepted as a common mechanism underlying drug induced organ toxicities (29,31,76), and is being increasingly detected in early stages of drug development using extracellular flux analysis (37,38,42). Mitochondrial toxicity of compounds is often assessed by growing the cells in the presence of high galactose (30,42,57), which forces the cells to use OXPHOS to produce ATP, thereby increasing the cells' sensitivity to the effects of mitochondrial toxicants compared to cells grown in glucose (77,78). However, the glucose-galactose switch does not sensitize all cell-types to mitochondrial cytotoxicity (79).…”

Section: Discussionmentioning

confidence: 99%

Host bioenergetic parameters reveal cytotoxicity of anti-tuberculosis drugs undetected using conventional viability assays

Cumming¹,

Baig²,

Addicott³

et al. 2021

Preprint

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High attrition rates in tuberculosis (TB) drug development have been largely attributed to safety, which is likely due to the use of endpoint assays measuring cell viability to detect drug cytotoxicity. In drug development of cancer, metabolic and neurological disorders, and antibiotics, cytotoxicity is increasingly being assessed using extracellular flux (XF) analysis, which measures cellular bioenergetic metabolism in real-time. Here, we adopt the XF platform to investigate the cytotoxicity of drugs currently used in TB treatment on the bioenergetic metabolism of HepG2 cells, THP-1 macrophages, and human monocyte derived macrophages (hMDM). We found that the XF analysis reveals earlier drug-induced effects on the cells' bioenergetic metabolism prior to cell death, measured by conventional viability assays. Furthermore, each cell type has a distinct response to drug treatment, suggesting that more than one cell type should be considered to examine cytotoxicity in TB drug development. Interestingly, chemically unrelated drugs with different modes of action on Mycobacterium tuberculosis have similar effects on the bioenergetic parameters of the cells, thus, discouraging the prediction of potential cytotoxicity based on chemical structure and mode of action of new chemical entities. The clustering of the drug-induced effects on the hMDM bioenergetic parameters are reflected in the clustering of the effects of the drugs on cytokine production in hMDMs, demonstrating concurrence between the effects of the drugs on the metabolism and functioning of the macrophages. These findings can be used as a benchmark to establish XF analysis as a new tool to assay cytotoxicity in TB drug development.

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High-throughput respirometric assay identifies predictive toxicophore of mitochondrial injury

Cited by 19 publications

References 43 publications

Quantification of Low-Level Drug Effects Using Real-Time,in vitroMeasurement of Oxygen Consumption Rate

Quantification of Low-Level Drug Effects Using Real-Time,in vitroMeasurement of Oxygen Consumption Rate

MitoTox: a comprehensive mitochondrial toxicity database

Host bioenergetic parameters reveal cytotoxicity of anti-tuberculosis drugs undetected using conventional viability assays

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