First-in-class multifunctional TYMS nonclassical antifolate inhibitor with potent in vivo activity that prolongs survival
Maria V. Guijarro, Patrick C. Kellish, Peter E. Dib, Nicholas G. Paciaroni, Akbar Nawab, Jacob Andring, Lidia Kulemina, Nicholas V. Borrero, Carlos Modenutti, Michael Feely, Elham Nasri, Robert P. Seifert, Xiaoping Luo, Richard L. Bennett, Daniil Shabashvili, Jonathan D. Licht, Robert McKenna, Adrian Roitberg, Robert W. Huigens III, Frederic J. Kaye, Maria Zajac-Kaye
Maria V. Guijarro, Patrick C. Kellish, Peter E. Dib, Nicholas G. Paciaroni, Akbar Nawab, Jacob Andring, Lidia Kulemina, Nicholas V. Borrero, Carlos Modenutti, Michael Feely, Elham Nasri, Robert P. Seifert, Xiaoping Luo, Richard L. Bennett, Daniil Shabashvili, Jonathan D. Licht, Robert McKenna, Adrian Roitberg, Robert W. Huigens III, Frederic J. Kaye, Maria Zajac-Kaye
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Research Article Therapeutics

First-in-class multifunctional TYMS nonclassical antifolate inhibitor with potent in vivo activity that prolongs survival

Abstract

Although thymidylate synthase (TYMS) inhibitors have served as components of chemotherapy regimens, the currently available inhibitors induce TYMS overexpression or alter folate transport/metabolism feedback pathways that tumor cells exploit for drug resistance, limiting overall benefit. Here we report a small molecule TYMS inhibitor that i) exhibited enhanced antitumor activity as compared with current fluoropyrimidines and antifolates without inducing TYMS overexpression, ii) is structurally distinct from classical antifolates, iii) extended survival in both pancreatic xenograft tumor models and an hTS/Ink4a/Arf null genetically engineered mouse tumor model, and iv) is well tolerated with equal efficacy using either intraperitoneal or oral administration. Mechanistically, we verify the compound is a multifunctional nonclassical antifolate, and using a series of analogs, we identify structural features allowing direct TYMS inhibition while maintaining the ability to inhibit dihydrofolate reductase. Collectively, this work identifies nonclassical antifolate inhibitors that optimize inhibition of thymidylate biosynthesis with a favorable safety profile, highlighting the potential for enhanced cancer therapy.

Authors

Maria V. Guijarro, Patrick C. Kellish, Peter E. Dib, Nicholas G. Paciaroni, Akbar Nawab, Jacob Andring, Lidia Kulemina, Nicholas V. Borrero, Carlos Modenutti, Michael Feely, Elham Nasri, Robert P. Seifert, Xiaoping Luo, Richard L. Bennett, Daniil Shabashvili, Jonathan D. Licht, Robert McKenna, Adrian Roitberg, Robert W. Huigens III, Frederic J. Kaye, Maria Zajac-Kaye

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Figure 2

Compound 19-S inhibits tumor growth and progression.

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Compound 19-S inhibits tumor growth and progression. (A) Experimental ti...
(A) Experimental timeline for the subcutaneous (SQ) Luc-PANC-1 cell line–derived tumor model treated with compound 19-S or vehicle control. Compound 19-S (25 mg/kg) or vehicle control treatment cycles were administered by IP injection. For each cycle animals were treated once a day for 5 continuous days, then allowed 2 days’ rest when no treatments were administered. Tumor-bearing mice received a total of 4 treatment cycles before endpoint (n = 4 per cohort). (B) Effect of compound 19-S and vehicle control administered by IP injection on body weight as described in A. (C) Bioluminescence imaging of Luc-PANC-1–derived tumors and quantification of bioluminescence photon flux over time for animals treated with compound 19-S or vehicle control (**P = 0.0023). (D) Luc-PANC-1 tumor volumes for animals treated with compound 19-S or vehicle control (***P < 0.0001). (E) Final excised Luc-PANC-1 tumor weight for animals treated with compound 19-S or vehicle control (**P = 0.0029). (F) Timeline indicating the IP injection of Luc-CM cells to generate a disseminated tumor model and compound 19-S (25 mg/kg) or vehicle control treatment cycles administered by IP injection. For each cycle animals were treated once a day for 5 continuous days, then allowed 2 days’ rest when no treatments were administered. Tumor-bearing animals received 3 treatment cycles before endpoint (n = 8 per cohort). (G) Effect of compound 19-S and vehicle control administered by IP injection on body weight for Luc-CM tumor–bearing NSG mice. (H) Bioluminescence imaging of Luc-CM tumor–bearing NSG mice and quantification of bioluminescence photon flux from the abdominal region of animals treated with compound 19-S (n = 8) or vehicle control (n = 8). Data are expressed as mean ± SEM, ***P < 0.0001. Statistical analysis in C, D, and H was performed using 2-way ANOVA; in E, unpaired 2-tailed Student’s t test with Welch’s correction was used.