Inhibition of TGF-β in Bone Marrow Endothelial Progenitor Cells Promotes Hematopoietic Recovery in Acute Myeloid Leukemia after Chemotherapy

Background: Chemotherapy is the basic treatment for acute myeloid leukemia (AML), but it impairs bone marrow (BM) cells and leads to myelosuppression. Endothelial progenitor cell (EPCs) plays an essential role in the maintenance and expansion of hematopoietic stem cells (HSCs). Our previous studies have reported that the recovery of dysfunctional EPCs is a prerequisite step in the hematopoietic reconstitution of patients with acute leukemia after allo-HSCT (2016 Blood, 2018 AJH, 2019 Blood Advances). Therefore, it is critical to explore the underlying mechanism of EPC impairment and establish strategies for targeted therapy. It has been reported that transforming growth factor-β (TGF-β) plays an important role in regulating the behavior of HSCs. Furthermore, luspatercept promotes advanced erythrocyte maturation by inhibiting TGF-β, which is used to treat thalassemia and myelodysplastic neoplasms. However, there is no report about the role of TGF-β signaling pathway in regulating functions of BM EPCs, especially in their hematopoietic-supporting ability.

Aims: To investigate the role of TGF-β pathway in regulation of BM EPC function from CR patients (CR EPCs) and its mechanism. Furthermore, to attenuate the dysfunction of EPCs and promote hematopoiesis recovery through targeting TGF-β pathway in EPCs.

Methods: In the prospective nested case-control study, de novo AML patients, CR patients and age-matched healthy control (HC) were collected to evaluate the expression of TGF-β by flow cytometry. To evaluate the functions of BM EPCs, migration, tube formation, apoptosis and reactive oxygen species were performed. The gene of TGF-βRI was knocked down in EPCs from HC (HC EPCs) after treated TGF-β1 in vitro. Moreover, the mechanistic role of TGF-β in BM EPC damage was determined by well-established BM EPC damage models in vitro, AAV-mediated BM EC-specific TGF-βRI overexpression murine model and AML-CR murine model, respectively. To understand the mechanism of the impairment in CR EPCs, RNA-Seq and qPCR were performed.

Results: When compared with HC EPCs, TGF-β signaling was overexpressed in CR EPCs. Although the hematopoietic-supporting ability of AML EPCs was partially recovered after CR, it remained dysfunctional in CR EPCs. Knockdown of TGF-βRI or TGF-β inhibitor rescued the dysfunction of EPCs caused by TGF-β1 in vitro, especially their hematopoiesis-supporting ability. Through BM EC specific TGF-βRI overexpression murine model, we proved that the overexpression of TGF-βRI in ECs induced the EC dysfunction, while inhibition of TGF-β in ECs could restore BM hematopoiesis through repairing EC dysfunction. Moreover, inhibition of TGF-β could repair the damage triggered by chemotherapy in AML EPCs both in vitro and in AML-CR murine model, and promote normal hematopoiesis recovery without promoting malignant hematopoiesis. Mechanistically, our data indicated that the downstream molecules of TGF-βRI, pSmad2/3, and functional genes including adhesion, angiogenesis-suppressor and pro-apoptosis were overexpressed in CR EPCs.

Summary/Conclusion: The current findings firstly reveal the abnormal high expression of TGF-β in CR EPCs, which leads to the dysfunction of EPC and poor hematopoietic reconstitution. Notably, inhibition of TGF-β in CR EPCs recovers trilineage hematopoiesis without promoting AML progression. Collectively, targeting TGF-β in BM EPCs (such as luspatercept) represents a potential therapeutic strategy to promote recovery of trilineage hematopoiesis, not just erythropoiesis, thereby benefiting more cancer patients who suffer from myelosuppression after chemotherapy.