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Benzylmorpholine Analogs as Selective Inhibitors of Lung Cytochrome P450 2A13 for the Chemoprevention of Lung Cancer in Tobacco Users

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ABSTRACT

Purpose

4-(Methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK), one of the most prevalent and procarcinogenic compounds in tobacco, is bioactivated by respiratory cytochrome P450 (CYP) 2A13, forming DNA adducts and initiating lung cancer. CYP2A13 inhibition offers a novel strategy for chemoprevention of tobacco-associated lung cancer.

Methods

Twenty-four analogs of a 4-benzylmorpholine scaffold identified by high throughput screening were evaluated for binding and inhibition of both functional human CYP2A enzymes, CYP2A13 and the 94%-identical hepatic CYP2A6, whose inhibition is undesirable. Thus, selectivity is a major challenge in compound design.

Results

A key feature resulting in CYP2A13-selective binding and inhibition was substitution at the benzyl ortho position, with three analogs being >25-fold selective for CYP2A13 over CYP2A6.

Conclusions

Two such analogs were negative for genetic and hERG toxicities and metabolically stable in human lung microsomes, but displayed rapid metabolism in human liver and in mouse and rat lung and liver microsomes, likely due to CYP2B-mediated degradation. A specialized knockout mouse mimicking the human lung demonstrates compound persistence in lung and provides an appropriate test model. Compound delivered by inhalation may be effective in the lung but rapidly cleared otherwise, limiting systemic exposure.

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Abbreviations

CYP:

Cytochrome P450

DMSO:

Dimethyl sulfoxide

hERG:

Human ether-a-go-go-related gene

K d :

Equilibrium dissociation constant

K i :

Inhibition constant

NNK:

4-(Methylnitrosamino)-1-(3-pyridyl)-1-butanone

PEITC:

Phenethyl isothiocyanate

POR:

NADPH-cytochrome P450 oxidoreductase

SAR:

Structure activity relationship

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ACKNOWLEDGMENTS AND DISCLOSURES

Thanks are due to Kelin Li, who provided synthetic training to LB. We are grateful for purity analysis by Patrick Porubsky and Ben Neuenswander at The University of Kansas Center for Chemical Methodologies and Library Design. Toxicity studies were undertaken at Cerep. Metabolic stability in microsomes was undertaken at Xenotech, LLC. Mike Wester facilitated metabolic screening with the major human xenobiotic metabolizing CYP enzymes. Transgenic microsomes were a gift from Xinxin Ding at the Wadsworth Center, New York State Department of Health and State University of New York at Albany. This work was supported by the Kansas Masonic Cancer Research Institute, the University of Kansas General Research Fund, NIH grant NIGMS GM076343, and the Institute for Advancing Medical Innovation.

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Authors and Affiliations

  1. Department of Medicinal Chemistry, University of Kansas, 1251 Wescoe Hall Dr., Lawrence, Kansas, 66045, USA

    Linda C. Blake, Jeffrey Aubé & Emily E. Scott

  2. High Throughput Screening Laboratory, University of Kansas, 2034 Becker Drive, Lawrence, Kansas, 66047, USA

    Anuradha Roy

  3. Pharmacokinetics, Dynamics and Metabolism, Pfizer Inc., La Jolla, California, 92121, USA

    David Neul

  4. University of Kansas Specialized Chemistry Center, University of Kansas, 2034 Becker Drive, Lawrence, Kansas, 66047, USA

    Frank J. Schoenen & Jeffrey Aubé

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  1. Linda C. Blake
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  2. Anuradha Roy
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  4. Frank J. Schoenen
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  5. Jeffrey Aubé
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  6. Emily E. Scott
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Correspondence to Emily E. Scott.

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Blake, L.C., Roy, A., Neul, D. et al. Benzylmorpholine Analogs as Selective Inhibitors of Lung Cytochrome P450 2A13 for the Chemoprevention of Lung Cancer in Tobacco Users. Pharm Res 30, 2290–2302 (2013). https://doi.org/10.1007/s11095-013-1054-z

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  • DOI: https://doi.org/10.1007/s11095-013-1054-z

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