Development, Validation, and Interlaboratory Comparison of an HMG-CoA Reductase Inhibition Assay for Quantitation of Atorvastatin in Plasma Matrices

Shum, Y. Y.; Huang, Naijia; Walter, Gary A.; Black, Ann E.; Sekerke, Catherine; Chang, Tsun; Whitfield, Lloyd R.

doi:10.1097/00007691-199802000-00008

Cited by 22 publications

(16 citation statements)

References 21 publications

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“…The concentration of atorvastatin used in the experiment is comparable with the plasma levels achieved in treated humans. 62 The downstream product of HMGcoA reductase, mevalonate, prevented atorvastatin from inhibiting growth of tumor cells ( Figure 1C). The effect of atorvastatin (10 M) was compared with MYC inactivation in the MYC-induced tumor cells.…”

Section: Atorvastatin Is Effective In Reversing and Preventing Myc-inmentioning

confidence: 97%

Inhibition of HMGcoA reductase by atorvastatin prevents and reverses MYC-induced lymphomagenesis

Shachaf

Perez

Youssef

et al. 2007

Blood

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Statins are a class of drugs that inhibit 3-hydroxy-3-methylglutaryl-coenzyme A (HMGcoA) reductase, a critical enzyme in the mevalonate pathway. Several reports document that statins may prevent different human cancers. However, whether or not statins can prevent cancer is controversial due to discordant results. One possible explanation for these conflicting conclusions is that only some tumors or specific statins may be effective. Here, we demonstrate in an in vivo transgenic model in which atorvastatin reverses and prevents the onset of MYC-induced lymphomagenesis, but fails to reverse or prevent tumorigenesis in the presence of constitutively activated K-Ras (G12D). Using phosphoprotein fluorescence-activated cell sorter (FACS) analysis, atorvastatin treatment was found to result in the inactivation of the Ras and ERK1/2 signaling pathways associated with the dephosphorylation and inactivation of MYC. Correspondingly, tumors with a constitutively activated K-Ras (G12D) did not exhibit dephosphorylation of ERK1/2 and MYC. Atorvastatin's effects on MYC were specific to the inhibition of HMGcoA reductase, as treatment with mevalonate, the product of HMG-CoA reductase activity, abrogated these effects and inhibited the ability of atorvastatin to reverse or suppress tumorigenesis. Also, RNAi directed at HMGcoA reductase was sufficient to abrogate the neoplastic properties of MYC-induced tumors. IntroductionCancer is largely caused by genomic events which result in the activation of oncogenes or the loss of tumor-suppressor genes. 1 The targeted inactivation of oncogenes may be a specific and effective therapy for treating certain malignancies. 2,3 Experimental results in animal models validate the notion that the targeted inactivation of a single oncogene can be sufficient to reverse tumorigenesis. 4 Drugs that target specific proteins have been identified and shown to be effective in the treatment of certain cancers, [5][6][7][8][9][10][11][12][13][14][15][16][17][18][19][20][21] which indicates that some neoplasms are susceptible to therapies that target their molecular causes. However, cancers can escape dependence upon oncogenes, as observed in animal models as well as in human patients. [21][22][23] Thus, the best targets may not be the suspected oncogenes but rather proteins essential to multiple signaling or metabolic pathways required for tumor maintenance.MYC is one of the oncogenes most commonly associated with human cancer. 24 Experimentally, the inactivation of MYC has been shown to induce sustained tumor regression in transgenic mouse models of many different tumors, suggesting that MYC may be a good target for cancer treatment. [25][26][27][28] However, MYC as a transcription factor is not easy to target directly with small molecules. Thus, rather than directly targeting MYC, it might be possible to inactivate MYC through the disruption of upstream regulatory signaling molecules such as those in pathways regulated by Ras and ERK1/2. 29,30 The enzyme 3-hydroxy-3-methylglutaryl-coenzyme A (HMGcoA) r...

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Section: Atorvastatin Is Effective In Reversing and Preventing Myc-inmentioning

confidence: 97%

Inhibition of HMGcoA reductase by atorvastatin prevents and reverses MYC-induced lymphomagenesis

Shachaf

Perez

Youssef

et al. 2007

Blood

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“…HMGCR activity assay was performed as described previously ( 45,46 ). Briefl y, 24 h after HepG2 cells were transfected with pre-miR-185 or control pre-miR, cells were incubated with 5% LPDS overnight to stimulate HMGCR activity.…”

Section: Hmgcr Activity Assaymentioning

confidence: 99%

Identification of miR-185 as a regulator of de novo cholesterol biosynthesis and low density lipoprotein uptake

Yang¹,

Liu²,

Pellicane³

et al. 2014

Journal of Lipid Research

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“…2-Benzyloxyaniline and 4-benzyloxyaniline were used respectively in the preparation of 2-and 4-hydroxy-atorvastatin and the corresponding lactones. D 5 -Aniline was used for the preparation of the D 5 -atorvastatin and its lactone. D 5 -Benzaldehyde was used for the preparation of the D 5 -analogs of 2-and 4-hydroxyatorvastatin and their lactones.…”

Section: Chemicals and Reagentsmentioning

confidence: 99%

“…The chemical structures of the acid and lactone forms of atorvastatin and the two metabolites are shown in Figs 1 and 2. An HMGCoA reductase inhibition assay [3][4][5] and a radioimmunoassay 6,7 have been used to quantitate atorvastatin equivalent concentrations in post-dose plasma samples. Recently, a high-performance liquid chromatography tandem mass spectrometry (LC/MS/MS) method has been reported for the quantitative determination of atorvastatin, 2-hydroxyatorvastatin and 4-hydroxyatorvastatin in human, dog and rat plasma.…”

mentioning

confidence: 99%

Quantitation of the acid and lactone forms of atorvastatin and its biotransformation products in human serum by high-performance liquid chromatography with electrospray tandem mass spectrometry

Jemal

Ouyang

Chen

et al. 1999

Rapid Commun. Mass Spectrom.

141

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A method for simultaneous quantitation of both the acid and lactone forms of atorvastatin, a new synthetic inhibitor of HMG-CoA reductase that is being marketed for the treatment of high serum cholesterol, and both the acid and lactone forms of its two biotransformation products, 2-hydroxyatorvastatin and 4-hydroxyatorvastatin, in human serum (a total of six analytes) by high-performance liquid chromatography with electrospray tandem mass spectrometry was developed and validated. A deuterium labeled analog was used as internal standard for each of the six analytes. Each point of the calibration standard curve, which ranged from 0.5 to 200 ng/mL, contained the six analytes at equal concentrations. Three groups of quality control (QC) samples were used. In the first group, combination QC samples contained all six analytes at equal concentrations. In the second group, acid-only QC samples contained only the acid forms (i.e. three analytes) at equal concentrations. In the third group, lactone-only QC samples contained only the lactone forms (i.e. three analytes) at equal concentrations. After adding the internal standard to 0.5 mL of each standard and the QC sample kept at 4 degrees C, the samples were acidified with sodium acetate buffer (pH 5.0) and then extracted with methyl tert-butyl ether. Detection was by positive ion electrospray tandem mass spectrometry using eight selected reaction monitoring channels. The acid compounds were stable in human serum at room temperature but the lactone compounds were unstable as they hydrolyzed rapidly to their respective acid forms. The conversion of the lactone compounds in both QC and post-dose human serum samples was nearly complete after 24 h at room temperature. The lactone compounds in serum could be stabilized by lowering the working temperature to 4 degrees C or lowering the serum pH to 6.0. The acid-only and the lactone-only QC samples showed that, under the sample processing conditions used, the degree of the hydrolysis of the lactone compounds or the lactonization of the acid compounds during the assay procedure was minimal (< 5%). The intra-day C.V., inter-day C.V. and the deviations from the nominal concentrations for all six analytes were within 15%, demonstrating good precision and accuracy. The required lower limit of quantitation (LLQ) of 0.5 ng/mL was achieved for each analyte.

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Development, Validation, and Interlaboratory Comparison of an HMG-CoA Reductase Inhibition Assay for Quantitation of Atorvastatin in Plasma Matrices

Cited by 22 publications

References 21 publications

Inhibition of HMGcoA reductase by atorvastatin prevents and reverses MYC-induced lymphomagenesis

Inhibition of HMGcoA reductase by atorvastatin prevents and reverses MYC-induced lymphomagenesis

Identification of miR-185 as a regulator of de novo cholesterol biosynthesis and low density lipoprotein uptake

Quantitation of the acid and lactone forms of atorvastatin and its biotransformation products in human serum by high-performance liquid chromatography with electrospray tandem mass spectrometry

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