Preclinical Characterization of XL092, a Novel Receptor Tyrosine Kinase Inhibitor of MET, VEGFR2, AXL, and MER

Abstract

The multi-receptor tyrosine kinase inhibitor XL092 has been developed to inhibit the activity of oncogenic targets, including MET, VEGFR2, and the TAM family of kinases TYRO3, AXL and MER. Presented here is a preclinical evaluation of XL092. XL092 causes a significant decrease in tumor MET and AXL phosphorylation (P < 0.01) in murine Hs 746T xenograft models relative to vehicle, and a 96% inhibition of VEGFR2 phosphorylation in murine lungs. Dose-dependent tumor growth inhibition with XL092 was observed in various murine xenograft models, with dose-dependent tumor regression seen in the NCI-H441 model. Tumor growth inhibition was enhanced with the combination of XL092 with anti-PD-1, anti-programmed death ligand-1 (PD-L1), or anti-CTLA-4 compared with any of these agents alone in the MC38 murine syngeneic model and with anti-PD-1 in the CT26 colorectal cancer survival model. In vivo, XL092 promoted a decrease in the tumor microvasculature and significant increases of peripheral CD4+ T cells and B cells and decreases in myeloid cells versus vehicle. Significant increases in CD8+ T cells were also observed with XL092 plus anti-PD-1 or anti-PD-L1 versus vehicle. In addition, XL092 promoted M2 to M1 repolarization of macrophages in vitro and inhibited primary human macrophage efferocytosis in a dose-dependent manner. In summary, XL092 was shown to have significant antitumor and immunomodulatory activity in animal models both alone and in combination with immune checkpoint inhibitors, supporting its evaluation in clinical trials.

Figure 1.

XL092 inhibits MET, AXL, and VEGFR2 activation in vivo.A, Structure of XL092. pMET/MET levels were evaluated in NCI-H441 (B) and SNU-5 (C) tumors from mice treated with either 3 (red) or 10 (blue) mg/kg XL092 or vehicle (black) orally daily for 14 days (n = 10, pooled for each treatment group). D, pVEGFR2 and VEGFR2 levels were evaluated in hVEGF-treated Hs 746T tumor-bearing mouse lung tissue following single oral doses of 30 or 100 mg/kg XL092 (n = 5, pooled for each treatment group) or vehicle (n = 6, pooled). E, Mean pMET/MET and pAXL/AXL ratios ± SEM were measured in Hs 746T tumor-bearing mice treated with single oral doses of either 30 or 100 mg/kg XL092 or vehicle (n = 6 for each treatment group). Significance values were determined using a nonparametric Mann–Whitney U test with significance levels indicated using **, P < 0.01.

Figure 2.

XL092 exhibits antitumor activity in multiple xenograft models. Tumor-bearing mice NCI-H441 (A), Hs 746T (B), SNU-5 (C), and MDA-MB-231 (D) were dosed with XL092 or vehicle orally once daily as indicated in Table 2. Mean tumor volume ± SEM (n = 10) presented. Significance values represent % TGI levels, and were determined using a nonparametric Mann–Whitney U test, with significance levels indicated using *, P < 0.05; **, P < 0.01; ***, P < 0.001; ****, P < 0.0001.

Figure 3.

Enhanced activity of XL092 plus ICI compared with single agents in MC38 syngeneic tumor model. Study design and tumor growth results in MC38 tumor-bearing mice treated with XL092 alone and in combination with ICI: anti-PD-1 (A), anti-PD-L1 (B), anti-CTLA-4 (C). Symbols represent mean tumor volume ± SEM (n = 10). BIW, twice a week. IP, intraperitoneal. PO, oral. QD, once a day.

Figure 4.

Preliminary evaluation of XL092 effect on tumor angiogenesis in vivo. MC38 tumor-bearing mice were treated with a range of XL092 doses (orally, once a day) for 5 days, and tumors were analyzed for the presence of tumor microvessels by CD31 staining. A, Paraffin-embedded tumor tissue stained with the blood vessel marker CD31 (blue arrows). B, Density score of blood vessels across conditions. Horizontal bars represent mean values for n = 3 tumors per condition.

Figure 5.

Effect of XL092/ICI combination treatment on peripheral immunocytes and CD8 T-cell tumor infiltration. A, The proportion of peripheral leukocytes that were CD4⁺ T cells, CD8⁺ T cells, B cells, and total myeloid cells were determined in MC38 tumor-bearing mice treated for 7 days with XL092 (10 or 30 mg/kg), 5 mg/kg anti-PD-1 antibody, 5 mg/kg anti-PD-L1 antibody, XL092 plus either anti-PD-1 or anti-PD-L1 antibody, or vehicle. Presented are study design and results. Bars represent median with significance determined using a nonparametric Mann–Whitney U test, with significance levels indicated using *, P < 0.05; **, P < 0.01; ***, P < 0.001; ****, P < 0.0001. B, Changes in CD8⁺ T-cell tumor density in MC38 tumor-bearing mice treated for 7 days with 10 mg/kg XL092 (n = 7), 5 mg/kg anti-PD-1 antibody (n = 10), XL092 plus anti-PD-1 antibody (n = 8) or vehicle (n = 12). Bars representing the mean displayed in the graph with arrows pointing to specific tumors representing the mean CD8⁺ T-cell numbers in the figure. IP, intraperitoneal. mpk, mg per kg. PO, oral. QD, once a day.