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The use of Trichomonas vaginalis purine nucleoside phosphorylase to activate fludarabine in the treatment of solid tumors

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Abstract

Treatment with fludarabine phosphate (9-β-D-arabinofuranosyl-2-F-adenine 5′-phosphate, F-araAMP) leads to regressions and cures of human tumor xenografts that express Escherichia coli purine nucleoside phosphorylase (EcPNP). This occurs despite the fact that fludarabine (F-araA) is a relatively poor substrate for EcPNP, and is cleaved to liberate 2-fluoroadenine at a rate only 0.3% that of the natural E. coli PNP substrate, adenosine. In this study, we investigated a panel of naturally occurring PNPs to identify more efficient enzymes that may be suitable for metabolizing F-araA as part of experimental cancer therapy. We show that Trichomonas vaginalis PNP (TvPNP) cleaves F-araA with a catalytic efficiency 25-fold greater than the prototypic E. coli enzyme. Cellular extracts from human glioma cells (D54) transduced with lentivirus stably expressing TvPNP (D54/TvPNP) were found to cleave F-araA at a rate similar to extracts from D54 cells expressing EcPNP, although much less enzyme was expressed per cell in the TvPNP transduced condition. As a test of safety and efficacy using TvPNP, human head and neck squamous cell carcinoma (FaDu) xenografts expressing TvPNP were studied in nude mice and shown to exhibit robust tumor regressions, albeit with partial weight loss that resolved post-therapy. F-araAMP was also a very effective treatment for mice bearing D54/TvPNP xenografts in which approximately 10% of tumor cells expressed the enzyme, indicating pronounced ability to kill non-transduced tumor cells (high bystander activity). Moreover, F-araAMP demonstrated activity against D54 tumors injected with an E1, E3 deleted adenoviral vector encoding TvPNP. In that setting, despite higher F-araA cleavage activity using TvPNP, tumor responses were similar to those obtained with EcPNP, indicating factors other than F-Ade production may limit regressions of the D54 murine xenograft model. Our results establish that TvPNP is a favorable enzyme for activating F-araA, and support further studies in combination with F-araAMP for difficult-to-treat human cancers.

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Acknowledgments

We thank Jan Tindall for help with preparation and revision of the manuscript. The research reported in this publication was supported in part by the Cancer Animal Models shared resource of Winship Cancer Institute of Emory University and NIH/NCI under award number P30CA138292. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.

Funding

The work was funded by NIH grant CA119170, R01DE026941, 5T32CA160040, and the Georgia Research Alliance.

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Authors and Affiliations

  1. Southern Research Institute, Birmingham, AL, USA

    William B. Parker, Paula W. Allan & William R. Waud

  2. Emory University School of Medicine, Atlanta, GA, USA

    Jeong Hong, Disha Joshi, Regina Rab & Eric J. Sorscher

  3. Emory University Cancer Animal Models Shared Resource, Winship Cancer Center, Atlanta, GA, USA

    Melissa Gilbert-Ross, B. R. Achyut & Turang Behbahani

  4. Department of Chemistry and Chemical Biology, Cornell University, Ithaca, NY, USA

    Steven E. Ealick

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  1. William B. Parker
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  2. Paula W. Allan
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  11. Eric J. Sorscher
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Correspondence to Eric J. Sorscher.

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Conflict of interest

Dr. Parker and Dr. Sorscher have ownership interests in PNP Therapeutics and serve on the Board of Directors for the company, which develops products used in research described by the paper. Drs. Parker and Sorscher are also inventors of technology being evaluated in studies described by this report. The terms of this arrangement for Dr. Sorscher have been reviewed and approved by Emory University in accordance with its conflict of interest policies. Paula Allan and Jeong Hong also have minor equity interest in this company.

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Parker, W.B., Allan, P.W., Waud, W.R. et al. The use of Trichomonas vaginalis purine nucleoside phosphorylase to activate fludarabine in the treatment of solid tumors. Cancer Chemother Pharmacol 85, 573–583 (2020). https://doi.org/10.1007/s00280-019-04018-7

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