Development Of a Novel Small Molecule Inhibitor Of The Mer Tyrosine Kinase For Treatment Of Acute Lymphoblastic Leukemia

In children, acute lymphoblastic leukemia (ALL) is currently treated with an intense regimen of chemotherapy yielding cure rates near 85%, yet significant hurdles remain, including decreasing toxicity and improving patient outcome in relapsed or refractory disease. Alternative strategies using available drugs are unlikely to provide significant improvements while more targeted therapies have the potential to reduce the risk of severe therapy-associated toxicities including infertility, organ damage, and secondary malignancy. We have identified Mer, a member of the TAM-family of receptor tyrosine kinases, as a potential therapeutic target in pediatric ALL. Mer is ectopically expressed in pediatric B- and T- cell ALL. Using a genetic approach (shRNA-mediated Mer knockdown), we previously demonstrated anti-leukemia effects in B-ALL and T-ALL models. MerTK inhibition decreased pro-survival signaling and lymphoblast cell survival, inhibited growth in colony forming assays, increased resistance to chemotherapy, and increased leukemia-free survival in animal models. Here we report testing of a novel, first-in-class Mer-selective small molecule inhibitor (Mer TKI) that has effects similar to Mer knock-down in B-ALL pre-clinical models. Our current lead candidate for clinical development mediates potent inhibition of Mer with an IC₅₀ of 0.74 nM. It has ≥10-fold selectivity for Mer over other TAM-family members, has limited off-target activity against other tyrosine kinases (with the exception of FLT3), and inhibits phosphorylation/activation of Mer in 697 B-ALL cells with an IC₅₀ of 2.6 nM. Treatment with this Mer TKI also inhibits downstream signaling through the ERK1/2 MAP kinase and Akt pathways. In mouse models, our Mer TKI is 100% orally bioavailable and has a favorable pharmacokinetic profile with a maximum serum concentration of 1.75 µM after administration of a single 3 mg/kg dose and a half-life of 3.8 hours. Pharmacodynamic studies to assess effective target engagement in murine models demonstrate inhibition of Mer phosphorylation in leukemic blasts in the bone marrow up to 12 hours post-treatment with Mer TKI. The compound is well-tolerated up to a dose of 100 mg/kg administered once daily with the major observed side effects being anemia and reduced white blood cell count. In an orthotopic B-ALL murine xenograft model of minimal residual disease, treatment with Mer TKI significantly inhibited leukemogenesis and increased median survival from 25 days after inoculation with tumor cells to 65 days (p < 0.0001). In a similar model of existent disease in which leukemia was established for 11 days prior to initiation of treatment, median survival increased from 25 to 47 days in response to treatment with Mer TKI (p < 0.0001). In both models, tumor burden measured by bioluminescent imaging was significantly decreased in mice treated with Mer TKI relative to mice treated with vehicle, even after the development of advanced disease in the control animals (0.27 +/- 0.04 x10⁷ photons/second in the minimal residual disease model and 0.98 +/- 0.23 x10⁷ photons/second in the established disease model verses 26.75 +/- 4.54 x10⁷ photons/sec in vehicle-treated mice, p < 0.0001). Its very high potency, relative selectivity, oral bioavailability, and favorable toxicity, pharmacokinetic, and pharmacodynamic profiles along with demonstrated therapeutic efficacy in murine ALL models make this novel Mer TKI an excellent candidate for clinical development.