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Design of isoform-selective phospholipase D inhibitors that modulate cancer cell invasiveness

Nature Chemical Biology volume 5, pages 108–117 (2009)Cite this article

Abstract

Phospholipase D (PLD) is an essential enzyme responsible for the production of the lipid second messenger phosphatidic acid. Phosphatidic acid participates in both G protein–coupled receptor and receptor tyrosine kinase signal transduction networks. The lack of potent and isoform-selective inhibitors has limited progress in defining the cellular roles of PLD. We used a diversity-oriented synthetic approach and developed a library of PLD inhibitors with considerable pharmacological characterization. Here we report the rigorous evaluation of that library, which contains highly potent inhibitors, including the first isoform-selective PLD inhibitors. Specific members of this series inhibit isoforms with >100-fold selectivity both in vitro and in cells. A subset of inhibitors was shown to block invasiveness in metastatic breast cancer models. These findings demonstrate the power of diversity-oriented synthesis combined with biochemical assays and mass spectrometric lipid profiling of cellular responses to develop the first isoform-selective PLD inhibitors—a new class of antimetastatic agents.

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Figure 1: PLD and phosphatidic acid in the cell.
Figure 2: Design and synthetic strategy for PLD inhibitor development.
Figure 3: In vitro inhibition of recombinant PLD with select compounds.
Figure 4: Direct small-molecule inhibition of PLD.
Figure 5: Development of distinct PLD isoform cell systems.
Figure 6: Small molecules potently inhibit cellular PLD activity.
Figure 7: Inhibition of PLD leads to decreased invasive migration in breast cancer cell lines.

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Acknowledgements

The authors acknowledge outstanding support from A. Goodman (mass spectrometry) and D. Myers (data analysis and statistics) as well as R. Bruntz (Vanderbilt University) for assisting with the generation of the HEK293-gfpPLD2 stable cell line. We thank C. Rouzer for helpful discussions. We gratefully acknowledge partial support for this project from the A.B. Hancock Jr. Memorial Laboratory for Cancer Research and the Vanderbilt Institute for Chemical Biology (to H.A.B. and C.W.L.). P.E.S. was supported in part by a US National Research Service Award Training Fellowship (T32 GM007628). T.L.C. was partially supported by the US National Cancer Institute T32 grant CA09592 and the Breast Cancer Specialized Program of Research Excellence (P50-CA98131 to C.L.A.).

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Author notes

  1. Sarah A Scott and Paige E Selvy: These authors contributed equally to this work.

Authors and Affiliations

  1. Department of Pharmacology, Vanderbilt Institute of Chemical Biology, Vanderbilt-Ingram Comprehensive Cancer Center, Vanderbilt University School of Medicine, 2220 Pierce Avenue South, Nashville, 37232-6600, Tennessee, USA

    Sarah A Scott, Paige E Selvy, Jason R Buck, Hyekyung P Cho, Ashley L Thomas, Michelle D Armstrong, Craig W Lindsley & H Alex Brown

  2. Department of Cancer Biology, Vanderbilt Institute of Chemical Biology, Vanderbilt-Ingram Comprehensive Cancer Center, Vanderbilt University School of Medicine, 2220 Pierce Avenue South, Nashville, 37232-6600, Tennessee, USA

    Tracy L Criswell & Carlos L Arteaga

  3. Department of Medicine, Vanderbilt Institute of Chemical Biology, Vanderbilt-Ingram Comprehensive Cancer Center, Vanderbilt University School of Medicine, 2220 Pierce Avenue South, Nashville, 37232-6600, Tennessee, USA

    Carlos L Arteaga

  4. Department of Chemistry, Vanderbilt Institute of Chemical Biology, Vanderbilt-Ingram Comprehensive Cancer Center, Vanderbilt University School of Medicine, 2220 Pierce Avenue South, Nashville, 37232-6600, Tennessee, USA

    Craig W Lindsley & H Alex Brown

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Contributions

S.A.S. carried out the cell-based inhibitor screening. P.E.S. and M.D.A. carried out in vitro inhibitor assays and protein purification. J.R.B. and A.L.T. performed chemical synthesis and characterization. H.P.C. was responsible for siRNA, RT-PCR and western blots. T.L.C. performed transwell experiments. C.L.A., C.W.L. and H.A.B. were responsible for coordination, planning and data interpretation of different aspects of the project.

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Correspondence to H Alex Brown.

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Scott, S., Selvy, P., Buck, J. et al. Design of isoform-selective phospholipase D inhibitors that modulate cancer cell invasiveness. Nat Chem Biol 5, 108–117 (2009). https://doi.org/10.1038/nchembio.140

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