American Association for Cancer Research
15357163mct141105-sup-143216_1_supp_2995179_nzyznp.docx (7.59 MB)

Supplementary Materials and Methods, Supplementary Tables 1 through 4, and Supplementary Figures 1 through 9 from Altiratinib Inhibits Tumor Growth, Invasion, Angiogenesis, and Microenvironment-Mediated Drug Resistance via Balanced Inhibition of MET, TIE2, and VEGFR2

Download (7.59 MB)
journal contribution
posted on 2023-04-03, 14:20 authored by Bryan D. Smith, Michael D. Kaufman, Cynthia B. Leary, Benjamin A. Turner, Scott C. Wise, Yu Mi Ahn, R. John Booth, Timothy M. Caldwell, Carol L. Ensinger, Molly M. Hood, Wei-Ping Lu, Tristan W. Patt, William C. Patt, Thomas J. Rutkoski, Thiwanka Samarakoon, Hanumaiah Telikepalli, Lakshminarayana Vogeti, Subha Vogeti, Karen M. Yates, Lawrence Chun, Lance J. Stewart, Michael Clare, Daniel L. Flynn

Supplementary Materials and Methods. Supplementary table 1: XRay crystallography parameters for DP-4157/MET complex. Supplementary table 2: Source and description of kinases. Supplementary table 3: Selectivity of profile of altiratinib inhibition from Reaction Biology 295 kinase panel (fold selectivity versus IC50 value of 2.7 nM MET inhibition). Supplementary table 4: A. Inhibition of MET phosphorylation in the MKN-45 xenograft model after a single oral dose of altiratinib (30 mg/kg); B. Inhibition of MET phosphorylation in the MKN-45 xenograft model after a single oral dose of altiratinib (10 mg/kg). Supplementary figure 1: Co-crystal structures of altiratinib analog DP-4157. Supplementary figure 2: Comparison of altiratinib with other clinical stage MET inhibitors regarding potency versus activation loop mutant forms D1228H, D1228N, Y1230D, Y1230C, and Y1230H. Supplementary figure 3: Determination of off-rate, residency time, and tight-binding inhibitor constant Kd for altiratinib binding to the MET kinase domain. Supplementary figure 4: Michaelis-Menton analysis of altiratinib inhibition of MET with respect to ATP. Supplementary figure 5: Kinome tree interaction profile for altiratinib vs 295 human kinases. Altiratinib was evaluated using Kinase HotspotSM platform from Reaction Biology (Malvern, PA). Supplementary figure 6: Inhibitory potencies for inhibiting HUVEC cell signaling activated by HGF/MET (black), VEGF-A/VEGFR2 (melon), and ANG2/TIE2 (tan). Comparative potencies are shown for altiratinib, E-7050, cabozantinib, and MGCD-265. Supplementary figure 7: Upper panel: inhibition of capillary tube formation by various concentrations of altiratinib. Tube formation was initiated by addition of 200 ng/mL of angiopoietin 2 (ANG2). Lower table: titrated IC50 values for inhibition of capillary tube formation induced by ANG2, HGF, and VEGF-A. Supplementary figure 8: Inhibition of proliferation in the mutant B-RAF melanoma SK-MEL-28 cell line in the absence or presence of HGF or MRC-5 fibroblast conditioned medium. Supplementary figure 9: Ratio of brain: plasma concentrations of altiratinib through 24 hr after administration of a single dose of 5 mg/kg IV to C57/Black mice.



Altiratinib (DCC-2701) was designed based on the rationale of engineering a single therapeutic agent able to address multiple hallmarks of cancer (1). Specifically, altiratinib inhibits not only mechanisms of tumor initiation and progression, but also drug resistance mechanisms in the tumor and microenvironment through balanced inhibition of MET, TIE2 (TEK), and VEGFR2 (KDR) kinases. This profile was achieved by optimizing binding into the switch control pocket of all three kinases, inducing type II inactive conformations. Altiratinib durably inhibits MET, both wild-type and mutated forms, in vitro and in vivo. Through its balanced inhibitory potency versus MET, TIE2, and VEGFR2, altiratinib provides an agent that inhibits three major evasive (re)vascularization and resistance pathways (HGF, ANG, and VEGF) and blocks tumor invasion and metastasis. Altiratinib exhibits properties amenable to oral administration and exhibits substantial blood–brain barrier penetration, an attribute of significance for eventual treatment of brain cancers and brain metastases. Mol Cancer Ther; 14(9); 2023–34. ©2015 AACR.