Acute myeloid leukemia (AML) with mutations in the tumor suppressor gene TP53 confers a dismal prognosis with 3-year overall survival of <5%. While inhibition of kinases involved in cell cycle regulation induces synthetic lethality in a variety of TP53 mutant cancers, this strategy has not been evaluated in mutant TP53 AML. Previously, we demonstrated that TP-0903 is a novel multikinase inhibitor with low nM activity against AURKA/B, Chk1/2, and other cell cycle regulators. Here, we evaluated the preclinical activity of TP-0903 in TP53 mutant AML cell lines, including a single-cell clone of MV4-11 containing a TP53 mutation (R248W), Kasumi-1 (R248Q), and HL-60 (TP 53 null). TP-0903 inhibited cell viability (IC50, 12–32 nM) and induced apoptosis at 50 nM. By immunoblot, 50 nM TP-0903 upregulated pChk1/2 and pH2AX, suggesting induction of DNA damage. The combination of TP-0903 and decitabine was additive in vitro, and in vivo significantly prolonged median survival compared to single-agent treatments in mice xenografted with HL-60 (vehicle, 46 days; decitabine, 55 days; TP-0903, 63 days; combination, 75 days) or MV4-11 (R248W) (51 days; 62 days; 81 days; 89 days) (p < 0.001). Together, these results provide scientific premise for the clinical evaluation of TP-0903 in combination with decitabine in TP53 mutant AML.

The FASEB JournalVolume 35, Issue S1 PharmacologyFree Access Preclinical Characterization of TP-0903, a Novel Multikinase Inhibitor, in TP53 Mutant Acute Myeloid Leukemia Eric Eisenmann, Eric Eisenmann Divisions of Pharmaceutics and Pharmacology, The Ohio State University, Columbus, OHSearch for more papers by this authorSydney Fobare, Sydney Fobare Division of Hematology, The Ohio State University, Columbus, OHSearch for more papers by this authorKevin Huang, Kevin Huang Division of Pharmaceutics and Pharmacology, The Ohio State University, Columbus, OHSearch for more papers by this authorJae Yoon Jeon, Jae Yoon Jeon Division of Pharmaceutics and Pharmacology, The Ohio State University, Columbus, OHSearch for more papers by this authorRobert Weber, Robert Weber Division of Pharmaceutics and Pharmacology, The Ohio State University, Columbus, OHSearch for more papers by this authorJosie Silvaroli, Josie Silvaroli Division of Pharmaceutics and Pharmacology, The Ohio State University, Columbus, OHSearch for more papers by this authorBill Larsen, Bill Larsen Division of Pharmaceutics and Pharmacology, The Ohio State University, Columbus, OHSearch for more papers by this authorJack Stromatt, Jack Stromatt Division of Pharmaceutics and Pharmacology, The Ohio State University, Columbus, OHSearch for more papers by this authorDaelynn Buelow, Daelynn Buelow Division of Pharmaceutics and Pharmacology, The Ohio State University, Columbus, OHSearch for more papers by this authorShelley Orwick, Shelley Orwick Division of Hematology, The Ohio State University, Columbus, OHSearch for more papers by this authorErin Hertlein, Erin Hertlein Division of Hematology, The Ohio State University, Columbus, OHSearch for more papers by this authorJohn Byrd, John Byrd Division of Hematology, The Ohio State University, Columbus, OHSearch for more papers by this authorSharyn Baker, Sharyn Baker Division of Pharmaceutics and Pharmacology, The Ohio State University, Columbus, OHSearch for more papers by this author Eric Eisenmann, Eric Eisenmann Divisions of Pharmaceutics and Pharmacology, The Ohio State University, Columbus, OHSearch for more papers by this authorSydney Fobare, Sydney Fobare Division of Hematology, The Ohio State University, Columbus, OHSearch for more papers by this authorKevin Huang, Kevin Huang Division of Pharmaceutics and Pharmacology, The Ohio State University, Columbus, OHSearch for more papers by this authorJae Yoon Jeon, Jae Yoon Jeon Division of Pharmaceutics and Pharmacology, The Ohio State University, Columbus, OHSearch for more papers by this authorRobert Weber, Robert Weber Division of Pharmaceutics and Pharmacology, The Ohio State University, Columbus, OHSearch for more papers by this authorJosie Silvaroli, Josie Silvaroli Division of Pharmaceutics and Pharmacology, The Ohio State University, Columbus, OHSearch for more papers by this authorBill Larsen, Bill Larsen Division of Pharmaceutics and Pharmacology, The Ohio State University, Columbus, OHSearch for more papers by this authorJack Stromatt, Jack Stromatt Division of Pharmaceutics and Pharmacology, The Ohio State University, Columbus, OHSearch for more papers by this authorDaelynn Buelow, Daelynn Buelow Division of Pharmaceutics and Pharmacology, The Ohio State University, Columbus, OHSearch for more papers by this authorShelley Orwick, Shelley Orwick Division of Hematology, The Ohio State University, Columbus, OHSearch for more papers by this authorErin Hertlein, Erin Hertlein Division of Hematology, The Ohio State University, Columbus, OHSearch for more papers by this authorJohn Byrd, John Byrd Division of Hematology, The Ohio State University, Columbus, OHSearch for more papers by this authorSharyn Baker, Sharyn Baker Division of Pharmaceutics and Pharmacology, The Ohio State University, Columbus, OHSearch for more papers by this author First published: 14 May 2021 https://doi.org/10.1096/fasebj.2021.35.S1.02109 This work was supported by funds from The Ohio State University Comprehensive Cancer Center Pelotonia Foundation (SDB) and the Pelotonia Fellowship Program (EDE). About ToolsRequest permissionExport citationAdd to favoritesTrack citation ShareShare Give accessShare full text accessShare full-text accessPlease review our Terms and Conditions of Use and check box below to share full-text version of article.I have read and accept the Wiley Online Library Terms and Conditions of UseShareable LinkUse the link below to share a full-text version of this article with your friends and colleagues. Learn more.Copy URL Share a linkShare onFacebookTwitterLinked InRedditWechat Abstract Objective Acute myeloid leukemia (AML) with mutations in the tumor suppressor gene TP53 confers a dismal prognosis with 1-year overall survival of <5%. Effective treatment options are limited and current standard-of-care includes the hypomethylating agents (HMA) decitabine and azacytidine. While inhibition of kinases involved in cell cycle regulation has been shown to induce synthetic lethality in a variety of TP53 mutant cancers, this strategy has not been evaluated in mutant TP53 AML. Previously, we demonstrated that TP-0903 is a novel multikinase inhibitor with low nM activity against AURKA/B, CHEK1/2, and other cell cycle regulators (Jeon JY et al. JCI Insight 2020), thus providing scientific rationale to evaluate TP-0903 activity in TP53 mutant AML. Methods To generate an in vitro model of TP53 mutant AML, we isolated single-cell clones containing mutant (R248W) or wild-type (WT) TP53 from the established MV4-11 AML cell line; regulation of p53 targets (MDM2, p21) following gamma irradiation and inhibition of p-AURKA and p-CHEK1 by TP-0903 were assessed by immunoblot. Using these and additional TP53 mutant AML cell lines (Kasumi-1, HL-60), in vitro efficacy of TP-0903 alone and in combination with HMA was assessed in viability (MTT) and apoptosis (Annexin V) assays. In vivo efficacy studies were conducted in NSG mice following intravenous injection of HL-60 or luciferase-tagged MV4-11/TP53-R248W cells. Mice (5-10 per treatment cohort) were treated with vehicle, TP-0903 (50 mg/kg orally; 5 days on/2 days off), decitabine (0.2-0.4 mg/kg i.p.; 4 days on/10 days off) or the combination. Whole body bioluminescence imaging was performed weekly and median survival was determined. Results Compared to the clone with WT TP53, we observed a lack of MDM2 and p21 induction in MV4-11/TP53-R248W cells following gamma irradiation. In vitro, TP-0903 inhibited pAURKA and pCHEK1 at 50 nM, inhibited cell viability (IC50 values, 12-40nM), and induced apoptosis at 20-50nM. The combination of TP-0903 with HMA was additive to synergistic in all AML cell lines evaluated. In the HL-60 xenograft model, the TP-0903/decitabine combination prolonged median survival (75 days) compared to cohorts of mice treated with TP-0903 (63 days), decitabine (55 days), or vehicle (46 days) (P<0.0001). In the MV4-11/TP53-R248W xenograft model, bioluminescence imaging showed that TP-0903 alone or in combination with decitabine was more effective in suppressing the outgrowth of leukemia cells compared to mice treated with vehicle or decitabine alone (P<0.05); survival analysis is ongoing. Conclusions TP-0903 was effective in all evaluated preclinical models of TP53 mutant AML. Together, these results provide scientific premise for the initiation of a Phase 1b/2 trial of TP-0903 in combination with decitabine in TP53 mutant/complex karyotype AML under the umbrella Beat AML Master Trial. Volume35, IssueS1Special Issue: Experimental Biology 2021 Meeting AbstractsMay 2021 RelatedInformation