Dr. Melissa C. Srougi

Research Areas:

  • Experimental targeted cancer chemotherapies
  • Breast cancer
  • DNA damage and repair
  • Redox signaling
  • Transcriptional regulation

Techniques:

  • Mammalian cell cultures
  • Dose-response assays
  • Flow cytometry
  • SDS-PAGE
  • Western blot
  • Immunoprecipitation assays
  • Redox signaling
  • Flourescence microscopy
  • RNA-seq data analysis
  • Molecular cloning and transfection
  • Cornet assays

Project Description:

The long-term goal of our research is to exploit natural products that target breast cancers for the advancement of precision medicine with notable clinical benefit. β-Lapachone (β-lap) is a naturally occurring antitumor quinone currently in Phase I clinical trials that is selectively bioactivated in tumors expressing NAD(P)H:quinone oxidoreductase 1 (NQO1). NQO1 is constitutively overexpressed in a number of solid tumors including breast (~60%) (1-3), non-small cell lung carcinomas (NSCLCs) (>85%), and pancreatic cancers (~85%) compared to associated normal tissues (4-7). Interestingly, breast tumors with mutations in breast cancer susceptibility genes BRCA1/2 also overexpress NQO1. However, the efficacy of β-lap has not been tested in these or other tumors with inherent defects in DNA repair.

NQO1 futile redox cycling of β-lap or IB-DNQ.

Figure 1. NQO1 futile redox cycling of β-lap or IB-DNQ.

NQO1-mediated futile cycle for IB-DNQ is shown.

NQO1 metabolizes β-lap via a two-electron oxidoreduction, resulting in the generation of an unstable hydroquinone form that rapidly reverts back to the parent quinone, causing multiple rounds of redox cycling (Fig. 1). β-Lap-induced reactive oxygen species (ROS) cause DNA base damage and DNA breaks (both single (SSB) and double-stranded (DSB)(8). SSBs and cytosolic Ca2+ release from endoplasmic reticulum pools hyperactive poly(ADP-ribose) polymerase-1 (PARP-1) resulting in cell death due to NAD+ and ATP loss (8-10). Interestingly, sub-lethal doses of β-lap, in combination with DNA damaging agents such as ionizing radiation, result in tumor sensitization (11,12). Recently, a novel more potent derivative of β-lap, isobutyldeoxynyboquinone (IB-DNQ) has been generated, which kills NQO1+ cells in a manner similar to β-lap (13). We hypothesize that sub-lethal doses of β-lap or IB-DNQ will induce low-level DNA damage in NQO1+; BRCA1/2-mutant breast tumors. Existing deficits in DNA repair caused by BRCA1/2 mutations in these cells will, therefore, selectively sensitize them to sub-lethal doses of  β-lap- or IB-DNQ resulting in lethality. Furthermore,  combination treatment with PARP inhibitors (PARPi) will further potentiate β-lap- or IB-DNQ- induced lethality for genotype-driven cytotoxicity, and thereby, circumvent possible resistance mechanisms. Two projects will be pursued to test this hypothesis:

  1. To determine the mechanism of β-lap or IB-DNQ-induced cell death in NQO1+;BRCA1/2-mutant breast cancers.
  1. To elucidate the efficacy of β-lap or IB-DNQ treatment in NQO1+;BRCA1/2-mutant breast cancers with or without PARPi combination treatment.

The findings gleaned from our studies will provide insight into the mechanisms of precision targeting of β-lap or IB-DNQ in NQO1-expressing BRCA1/2 mutant breast tumors to increase tumor response to therapy while decreasing normal tissue toxicity.

References

  1. Siegel, D., and Ross, D. (2000) Immunodetection of NAD(P)H:quinone oxidoreductase 1 (NQO1) in human tissues. Free Radic Biol Med 29, 246-253
  2. Yang, Y., Zhang, Y., Wu, Q., Cui, X., Lin, Z., Liu, S., and Chen, L. (2014) Clinical implications of high NQO1 expression in breast cancers. J Exp Clin Cancer Res 33, 14
  3. Menzel, H. J., Sarmanova, J., Soucek, P., Berberich, R., Grunewald, K., Haun, M., and Kraft, H. G. (2004) Association of NQO1 polymorphism with spontaneous breast cancer in two independent populations. Br J Cancer 90, 1989-1994
  4. Dong, Y., Bey, E. A., Li, L. S., Kabbani, W., Yan, J., Xie, X. J., Hsieh, J. T., Gao, J., and Boothman,
    1. D.A. (2010) Prostate cancer radiosensitization through poly(ADP-Ribose) polymerase-1 hyperactivation. Cancer Res 70, 8088-8096
  5. Dong, Y., Chin, S. F., Blanco, E., Bey, E. A., Kabbani, W., Xie, X. J., Bornmann, W. G., Boothman, D. A., and Gao, J. (2009) Intratumoral delivery of beta-lapachone via polymer implants for prostate cancer therapy. Clin Cancer Res 15, 131-139
  6. Lewis, A. M., Ough, M., Hinkhouse, M. M., Tsao, M. S., Oberley, L. W., and Cullen, J. J. (2005) Targeting NAD(P)H:quinone oxidoreductase (NQO1) in pancreatic cancer. Mol Carcinog 43, 215- 224
  7. Marin, A., Lopez de Cerain, A., Hamilton, E., Lewis, A. D., Martinez-Penuela, J. M., Idoate, M. A., and Bello, J. (1997) DT-diaphorase and cytochrome B5 reductase in human lung and breast tumours. Br J Cancer 76, 923-929
  8. Bentle, M. S., Reinicke, K. E., Bey, E. A., Spitz, D. R., and Boothman, D. A. (2006) Calcium- dependent modulation of poly(ADP-ribose) polymerase-1 alters cellular metabolism and DNA repair. J Biol Chem 281, 33684-33696
  9. Bey, E. A., Bentle, M. S., Reinicke, K. E., Dong, Y., Yang, C. R., Girard, L., Minna, J. D., Bornmann, W. G., Gao, J., and Boothman, D. A. (2007) An NQO1- and PARP-1-mediated cell death pathway induced in non-small-cell lung cancer cells by beta-lapachone. Proc Natl Acad Sci U S A 104, 11832-11837
  10. Tagliarino, C., Pink, J. J., Dubyak, G. R., Nieminen, A. L., and Boothman, D. A. (2001) Calcium is a key signaling molecule in beta-lapachone-mediated cell death. J Biol Chem 276, 19150-19159
  11. Boothman, D. A., Greer, S., and Pardee, A. B. (1987) Potentiation of halogenated pyrimidine radiosensitizers in human carcinoma cells by beta-lapachone (3,4-dihydro-2,2-dimethyl-2H- naphtho[1,2-b]pyran- 5,6-dione), a novel DNA repair inhibitor. Cancer Res 47, 5361-5366
  12. Boothman, D. A., Trask, D. K., and Pardee, A. B. (1989) Inhibition of potentially lethal DNA damage repair in human tumor cells by beta-lapachone, an activator of topoisomerase I. Cancer Res 49, 605-612

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