FOXF1 promotes tumor vessel normalization and prevents lung cancer progression through FZD4.

Cancer cells re-program normal lung endothelial cells (EC) into tumor-associated endothelial cells (TEC) that form leaky vessels supporting carcinogenesis. Transcriptional regulators that control the reprogramming of EC into TEC are poorly understood. We identified Forkhead box F1 (FOXF1) as a critical regulator of EC-to-TEC transition. FOXF1 was highly expressed in normal lung vasculature but was decreased in TEC within non-small cell lung cancers (NSCLC). Low FOXF1 correlated with poor overall survival of NSCLC patients. In mice, endothelial-specific deletion of FOXF1 decreased pericyte coverage, increased vessel permeability and hypoxia, and promoted lung tumor growth and metastasis. Endothelial-specific overexpression of FOXF1 normalized tumor vessels and inhibited the progression of lung cancer. FOXF1 deficiency decreased Wnt/ß-catenin signaling in TECs through direct transcriptional activation of Fzd4. Restoring FZD4 expression in FOXF1-deficient TECs through endothelial-specific nanoparticle delivery of Fzd4 cDNA rescued Wnt/ß-catenin signaling in TECs, normalized tumor vessels and inhibited the progression of lung cancer. Altogether, FOXF1 increases tumor vessel stability, and inhibits lung cancer progression by stimulating FZD4/Wnt/ß-catenin signaling in TECs. Nanoparticle delivery of FZD4 cDNA has promise for future therapies in NSCLC.

Epidermal growth factor-like 7 is a novel therapeutic target in mantle cell lymphoma.

Mantle cell lymphoma (MCL) is an aggressive, noncurative, mature B-cell lymphoma, with a median overall survival of 6-7 years. This underlines a need for effective therapeutic strategies to treat MCL better. Epidermal growth factor-like 7 (EGFL7) is a protein secreted by endothelial cells shown to play a critical role in angiogenesis. Our laboratory has previously demonstrated that EGFL7 supports the growth of leukemic blasts in patients with acute myeloid leukemia (AML); however, its role in MCL has not been investigated yet. In this study, we report that EGFL7 messenger RNA (mRNA) is increased in the cells of patients with MCL compared with cells from healthy controls, and patients with high EGFL7 are associated with lower overall survival rates. Furthermore, EGFL7 is increased in the plasma of patients with MCL compared with the plasma from healthy controls. We further show that EGFL7 binds to epidermal growth factor receptor (EGFR) and activates AKT signaling pathway in MCL cells and that blocking EGFL7 in MCL in patient and cell lines decreases cell growth and increases apoptosis in vitro. Finally, anti-EGFL7 treatment inhibits tumor size and prolongs survival in a mouse model of MCL. In conclusion, our study reveals a role for EGFL7 in MCL cell proliferation and highlights EGFL7 inhibition as a promising new treatment for patients with MCL.

Modulating endothelial cells with EGFL7 to diminish aGVHD after allogeneic bone marrow transplantation in mice.

Acute graft-versus-host disease (aGVHD) is the second most common cause of death after allogeneic hematopoietic stem cell transplantation (allo-HSCT), underscoring the need for novel therapies. Based on previous work that endothelial cell dysfunction is present in aGVHD and that epidermal growth factor-like domain 7 (EGFL7) plays a significant role in decreasing inflammation by repressing endothelial cell activation and T-cell migration, we hypothesized that increasing EGFL7 levels after allo-HSCT will diminish the severity of aGVHD. Here, we show that treatment with recombinant EGFL7 (rEGFL7) in 2 different murine models of aGVHD decreases aGVHD severity and improves survival in recipient mice after allogeneic transplantation with respect to controls without affecting graft-versus-leukemia effect. Furthermore, we showed that rEGFL7 treatment results in higher thymocytes, T, B, and dendritic cell counts in recipient mice after allo-HSCT. This study constitutes a proof of concept of the ability of rEGFL7 therapy to reduce GHVD severity and mortality after allo-HSCT.

Loss of FOXM1 in macrophages promotes pulmonary fibrosis by activating p38 MAPK signaling pathway.

Idiopathic pulmonary fibrosis (IPF) is a chronic disease with high mortality and is refractory to treatment. Pulmonary macrophages can both promote and repress fibrosis, however molecular mechanisms regulating macrophage functions during fibrosis remain poorly understood. FOXM1 is a transcription factor and is not expressed in quiescent lungs. Herein, we show that FOXM1 is highly expressed in pulmonary macrophages within fibrotic lungs of IPF patients and mouse fibrotic lungs. Macrophage-specific deletion of Foxm1 in mice (myFoxm1-/-) exacerbated pulmonary fibrosis. Inactivation of FOXM1 in vivo and in vitro increased p38 MAPK signaling in macrophages and decreased DUSP1, a negative regulator of p38 MAPK pathway. FOXM1 directly activated Dusp1 promoter. Overexpression of DUSP1 in FOXM1-deficient macrophages prevented activation of p38 MAPK pathway. Adoptive transfer of wild-type monocytes to myFoxm1-/- mice alleviated bleomycin-induced fibrosis. Altogether, contrary to known pro-fibrotic activities in lung epithelium and fibroblasts, FOXM1 has anti-fibrotic function in macrophages by regulating p38 MAPK.

FOXF1 maintains endothelial barrier function and prevents edema after lung injury.

Multiple signaling pathways, structural proteins, and transcription factors are involved in the regulation of endothelial barrier function. The forkhead protein FOXF1 is a key transcriptional regulator of embryonic lung development, and we used a conditional knockout approach to examine the role of FOXF1 in adult lung homeostasis, injury, and repair. Tamoxifen-regulated deletion of both Foxf1 alleles in endothelial cells of adult mice (Pdgfb-iCreER/Foxf1(-/-)) caused lung inflammation and edema, leading to respiratory insufficiency and death. Deletion of a single Foxf1 allele made heterozygous Pdgfb-iCreER/Foxf1(+/-)mice more susceptible to acute lung injury. FOXF1 abundance was decreased in pulmonary endothelial cells of human patients with acute lung injury. Gene expression analysis of pulmonary endothelial cells with homozygous FOXF1 deletion indicated reduced expression of genes critical for maintenance and regulation of adherens junctions. FOXF1 knockdown in vitro and in vivo disrupted adherens junctions, enhanced lung endothelial permeability, and increased the abundance of the mRNA and protein for sphingosine 1-phosphate receptor 1 (S1PR1), a key regulator of endothelial barrier function. Chromatin immunoprecipitation and luciferase reporter assays demonstrated that FOXF1 directly bound to and induced the transcriptional activity of the S1pr1 promoter. Pharmacological administration of S1P to injured Pdgfb-iCreER/Foxf1(+/-)mice restored endothelial barrier function, decreased lung edema, and improved survival. Thus, FOXF1 promotes normal lung homeostasis and repair, in part, by enhancing endothelial barrier function through activation of the S1P/S1PR1 signaling pathway.