MEK Inhibition Reduces Vascular Malformations and Gene Dysregulation in NRAS Q61R Human Endothelial Cells

ABSTRACT Background An activating NRAS p .Q61R ( NRAS Q61R ) somatic mutation occurs in the lesions of patients with kaposiform lymphangiomatosis (KLA). Therapies for KLA patients are limited and since the mitogen‐activated protein kinase pathway is hyperactivated by NRAS Q61R , we evaluated the potency of Mitogen‐activated protein kinase kinase (MEK) inhibitors (trametinib, selumetinib, and cobimetinib) on human NRAS Q61R endothelial cells (ECs). RNA sequencing analysis was performed to identify dysregulated genes and those restored by trametinib. Trametinib was tested in a mouse xenograft model of NRAS Q61R ECs. Procedure Doxycycline (Dox)‐inducible NRAS Q61R ECs were treated with trametinib, selumetinib, cobimetinib, or vehicle (control). Cell signaling pathways, proliferation, migration, morphology, and angiopoietin‐2 (ANG‐2) levels were assessed. RNA‐sequencing analysis was performed on Dox‐induced NRAS wild‐type ( NRAS WT ), NRAS Q61R , and trametinib‐treated NRAS Q61R ECs. Selected proteins were verified by immunoblotting. NRAS Q61R ECs were injected into nude mice on Dox‐containing diet to generate xenografts and treated with trametinib or vehicle. Results Trametinib reduced NRAS Q61R ‐activated ERK phosphorylation, cell proliferation, migration, spindling, and ANG‐2 at lower doses than selumetinib or cobimetinib. RNA‐sequencing detected 1315 upregulated and 1773 downregulated genes in NRAS Q61R ECs compared with NRAS WT . Trametinib corrected 19% upregulated and 8% downregulated genes, including elevated Notch ligands (DLL4, JAG1), activated Notch 1, and HES1. Trametinib reduced NRAS Q61R ECs xenograft weights by 46%, vascular area by 63%, and phosphorylated ERK staining. Conclusion Trametinib was the most effective MEK inhibitor, correcting some dysregulated genes in NRAS Q61R ECs, including some in the Notch pathway. Trametinib reduced vessel overgrowth in xenografts of NRAS Q61R ECs. These studies support using trametinib treatment for KLA patients with an NRAS Q61R mutation.