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Systemic depletion of L-cyst(e)ine with cyst(e)inase increases reactive oxygen species and suppresses tumor growth

Nature Medicine volume 23, pages 120–127 (2017)Cite this article

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Abstract

Cancer cells experience higher oxidative stress from reactive oxygen species (ROS) than do non-malignant cells because of genetic alterations and abnormal growth; as a result, maintenance of the antioxidant glutathione (GSH) is essential for their survival and proliferation1,2,3. Under conditions of elevated ROS, endogenous L-cysteine (L-Cys) production is insufficient for GSH synthesis. This necessitates uptake of L-Cys that is predominantly in its disulfide form, L-cystine (CSSC), via the xCT(−) transporter. We show that administration of an engineered and pharmacologically optimized human cyst(e)inase enzyme mediates sustained depletion of the extracellular L-Cys and CSSC pool in mice and non-human primates. Treatment with this enzyme selectively causes cell cycle arrest and death in cancer cells due to depletion of intracellular GSH and ensuing elevated ROS; yet this treatment results in no apparent toxicities in mice even after months of continuous treatment. Cyst(e)inase suppressed the growth of prostate carcinoma allografts, reduced tumor growth in both prostate and breast cancer xenografts and doubled the median survival time of TCL1-Tg:p53−/− mice, which develop disease resembling human chronic lymphocytic leukemia. It was observed that enzyme-mediated depletion of the serum L-Cys and CSSC pool suppresses the growth of multiple tumors, yet is very well tolerated for prolonged periods, suggesting that cyst(e)inase represents a safe and effective therapeutic modality for inactivating antioxidant cellular responses in a wide range of malignancies4,5.

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Figure 1: Engineered human cyst(e)inase for cancer treatment.
Figure 2: Effect of cyst(e)inase in PCa cells.
Figure 3: Pharmacokinetics, pharmacodynamics and efficacy of cyst(e)inase administration in mice.
Figure 4: In vitro and in vivo efficacy of cyst(e)inase in the TCL1-Tg:p53−/− mouse model and primary CLL cells.

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Acknowledgements

We are grateful to D. Lowe (Aeglea Biotherapeutics), O. Paley, and M. Bonem for assistance with various aspects of this work and data interpretation and to L. Beltran and S. Carbajal for their expert technical assistance. Instrumentation and technical assistance for parts of this work were provided by the Macromolecular Crystallography Facility, with financial support from the College of Natural Sciences, the Office of the Executive Vice President and Provost, and the Institute for Cellular and Molecular Biology at the University of Texas at Austin. Diffraction data was collected at the Advanced Light Source at the Berkeley Center for Structural Biology which is supported in part by the National Institutes of Health, National Institute of General Medical Sciences, and the Howard Hughes Medical Institute. The Advanced Light Source is supported by the Director, Office of Science, Office of Basic Energy Sciences, of the US Department of Energy under Contract No. DE-AC02-05CH11231. This work was supported by funding from: Welch Foundation F-1778 (Y.J.Z.), NIH 1 R01 GM104896 (Y.J.Z.), NIH 1 R01 CA172724 (P.H.), NIH 1 R01 CA154754 (G.G., E.S.), NIH 1 RO1 CA189623 (E.S., J.D. and G.G.), NCI P30 CA54174 (S. Tiziani) and Aeglea Biotherapeutics.

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  1. Shira L Cramer, Achinto Saha and Jinyun Liu: These authors contributed equally to this work.

Authors and Affiliations

  1. Department of Chemical Engineering, University of Texas at Austin, Austin, Texas, USA

    Shira L Cramer, Kendra Triplett, Candice Lamb & George Georgiou

  2. Division of Pharmacology and Toxicology and Dell Pediatric Research Institute, University of Texas at Austin, Austin, Texas, USA

    Achinto Saha & John DiGiovanni

  3. Department of Translational Molecular Pathology, University of Texas MD Anderson Cancer Center, Houston, Texas, USA

    Jinyun Liu & Peng Huang

  4. Department of Nutritional Sciences, University of Texas at Austin, Austin, Texas, USA

    Surendar Tadi & Stefano Tiziani

  5. Department of Molecular Biosciences, University of Texas at Austin, Austin, Texas, USA

    Wupeng Yan, Kendra Triplett, Candice Lamb, Yan Jessie Zhang, George Georgiou & Everett Stone

  6. Aeglea Biotherapeutics, Austin, Texas, USA,

    Susan E Alters & Scott Rowlinson

  7. Department of Leukemia, University of Texas MD Anderson Cancer Center, Houston, Texas, USA

    Michael J Keating

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  1. Shira L Cramer
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Contributions

S.L.C., A.S., J.L., S. Tadi, W.Y., K.T., C.L., and Y.J.Z. performed key experiments; E.S., P.H., J.D. and G.G. conceived and designed the research; S.L.C., A.S., J.L., S. Tadi, S. Tiziani, W.Y., K.T., S.E.A., S.R., Y.J.Z., M.J.K, P.H., J.D., G.G., and E.S. analyzed data; M.J.K. provided critical materials (CLL blood samples); S.L.C., A.S., J.L., G.G., J.D. and E.S. wrote the manuscript.

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Correspondence to George Georgiou or Everett Stone.

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Competing interests

G. Georgiou and E. Stone are inventors on intellectual property related to this work, and G. Georgiou, E. Stone, S. Rowlinson and S. Alters have an equity interest in Aeglea Biotherapeutics, a company pursuing the commercial development of this technology.

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Cramer, S., Saha, A., Liu, J. et al. Systemic depletion of L-cyst(e)ine with cyst(e)inase increases reactive oxygen species and suppresses tumor growth. Nat Med 23, 120–127 (2017). https://doi.org/10.1038/nm.4232

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