The Metabolic Fate of isoCombretastatin A-4 in Human Liver Microsomes: Identification, Synthesis and Biological Evaluation of Metabolites

DN203368 ((E)-3-[1-(4-[4-isopropylpiperazine-1-yl]phenyl) 3-methyl-2-phenylbut-1-en-1-yl] phenol) is a 4-hydroxy tamoxifen analog that is a dual inverse agonist of estrogen-related receptor β/γ (ERRβ/γ). ERRγ is an orphan nuclear receptor that plays an important role in development and homeostasis and holds potential as a novel therapeutic target in metabolic diseases such as diabetes mellitus, obesity, and cancer. ERRβ is also one of the orphan nuclear receptors critical for many biological processes, such as development. We investigated the in vitro metabolism of DN203368 by conventional and metabolomic approaches using high-resolution mass spectrometry. The compound (100 μM) was incubated with rat and human liver microsomes in the presence of NADPH. In the metabolomic approach, the m/z value and retention time information obtained from the sample and heat-inactivated control group were statistically evaluated using principal component analysis and orthogonal partial least-squares discriminant analysis. Significant features responsible for group separation were then identified using tandem mass spectra. Seven metabolites of DN203368 were identified in rat liver microsomes and the metabolic pathways include hydroxylation (M1-3), N-oxidation (M4), N-deisopropylation (M5), N,N-dealkylation (M6), and oxidation and dehydrogenation (M7). Only five metabolites (M2, M3, and M5-M7) were detected in human liver microsomes. In the conventional approach using extracted ion monitoring for values of mass increase or decrease by known metabolic reactions, only five metabolites (M1-M5) were found in rat liver microsomes, whereas three metabolites (M2, M3, and M5) were found in human liver microsomes. This study revealed that nontargeted metabolomics combined with high-resolution mass spectrometry and multivariate analysis could be a more efficient tool for drug metabolite identification than the conventional approach. These results might also be useful for understanding the pharmacokinetics and metabolism of DN203368 in animals and humans.

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Roles of cytochrome P450 2A6 in the oxidation of flavone, 4´-hydroxyflavone, and 4´-, 3´-, and 2´-methoxyflavones by human liver microsomes

Xenobiotica ◽

10.1080/00498254.2021.1950866 ◽

2021 ◽

pp. 1-47

Author(s):

Haruna Nagayoshi ◽

Norie Murayama ◽

Shigeo Takenaka ◽

Vitchan Kim ◽

Donghak Kim ◽

...

Keyword(s):

Cytochrome P450 ◽

Human Liver ◽

Liver Microsomes ◽

Human Liver Microsomes ◽

Cytochrome P450 2A6

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Pyrrolidinyl Synthetic Cathinones α-PHP and 4F-α-PVP Metabolite Profiling Using Human Hepatocyte Incubations

International Journal of Molecular Sciences ◽

10.3390/ijms22010230 ◽

2020 ◽

Vol 22 (1) ◽

pp. 230

Author(s):

Jeremy Carlier ◽

Xingxing Diao ◽

Raffaele Giorgetti ◽

Francesco P. Busardò ◽

Marilyn A. Huestis

Keyword(s):

Metabolite Profiling ◽

Human Liver ◽

Liver Microsomes ◽

Drug Market ◽

Metabolite Profile ◽

Illegal Drug ◽

Human Hepatocyte ◽

Synthetic Cathinones ◽

Metabolic Fate ◽

Analytical Standards

For more than ten years, new synthetic cathinones (SCs) mimicking the effects of controlled cocaine-like stimulants have flooded the illegal drug market, causing numerous intoxications and fatalities. There are often no data on the pharmacokinetics of these substances when they first emerge onto the market. However, the detection of SC metabolites is often critical in order to prove consumption in clinical and forensic settings. In this research, the metabolite profile of two pyrrolidinyl SCs, α-pyrrolidinohexaphenone (α-PHP) and 4′′-fluoro-α-pyrrolidinovalerophenone (4F-α-PVP), were characterized to identify optimal intake markers. Experiments were conducted using pooled human hepatocyte incubations followed by liquid chromatography–high-resolution tandem mass spectrometry and data-mining software. We suggest α-PHP dihydroxy-pyrrolidinyl, α-PHP hexanol, α-PHP 2′-keto-pyrrolidinyl-hexanol, and α-PHP 2′-keto-pyrrolidinyl as markers of α-PHP use, and 4F-α-PVP dihydroxy-pyrrolidinyl, 4F-α-PVP hexanol, 4F-α-PVP 2′-keto-pyrrolidinyl-hexanol, and 4F-α-PVP 2′-keto-pyrrolidinyl as markers of 4F-α-PVP use. These results represent the first data available on 4F-α-PVP metabolism. The metabolic fate of α-PHP was previously studied using human liver microsomes and urine samples from α-PHP users. We identified an additional major metabolite (α-PHP dihydroxy-pyrrolidinyl) that might be crucial for documenting exposure to α-PHP. Further experiments with suitable analytical standards, which are yet to be synthesized, and authentic specimens should be conducted to confirm these results.

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