Targeting EBF1-ITGB8 Axis in MPN Bone Marrow Niche

Hematopoiesis occurs in a complex bone marrow (BM) microenvironment that includes mesenchymal stromal cells (MSCs). The integrity and fitness of hematopoietic stem cell (HSC) pool is regulated by cross communication with MSCs and other niche components. In blood malignancies, hematopoietic cells carrying disease-causing mutations corrupt MSCs via cell-cell contact and secreted factors, creating a dysfunctional niche. In turn, the defective niche can negatively affect normal hematopoiesis, but support mutant cells. In one medically important example, such as myelofibrosis (MF), corrupted MSCs cause BM fibrosis that impairs blood cell production leading to extramedullary hematopoiesis, anemia and fatigue. Corrupted MSCs in myelofibrosis produce excessive amounts of extracellular matrix (ECM), contribute to inflammatory milieu in BM, and reduce production of essential hematopoietic supportive factors. While the genetic events that initiate myelofibrosis in HSCs acquiring somatic driver mutations in JAK2, MPL or CALR are well defined, less is understood about the mechanisms responsible for BM fibrosis and niche dysfunction. JAK inhibitors, the current standard of care for MF patients, fail to eradicate the mutant clone and do not alleviate BM fibrosis. Moreover, although allogeneic HSC transplantation is potentially curative, patients often suffer from early transplant-related mortality, from graft failure that is attributed to a fibrotic BM niche. In mouse models, reducing fibrosis by targeting the corrupted MSCs directly and/or inhibiting the aberrant signals that they receive from mutant cells can attenuate MF. Therefore, combined targeting of the mutant blood clone and the defective and pro-fibrotic niche should improve therapy for MF. However, the molecular mechanisms that drive niche dysfunction and MSC fibrotic transformation are poorly understood. Here, we show that transcription factor early B cell factor 1 (EBF1) is a key regulator of MSC function and MF niche by controlling the expression of ECM components, integrins, and proinflammatory cytokines. EBF1 expression is upregulated in MSCs from MF mouse models and MF patients. In co-culture assays, priming of mouse and human EBF1-deficient MSCs towards fibrosis-initiating cells is blunted in response to cells carrying the MPLW515L or JAK2V617F mutations identified in patients. Moreover, transplantation of hematopoietic progenitors expressing MPLW151L mutation into mice with MSC-specific deletion of Ebf1 results in reduced BM fibrosis, fewer megakaryocytes, and reduced expansion of mutant myeloid cells. We identified ITGB8 as an EBF1-regulated gene in the MSCs, which contributes to fibrotic transformation of the BM. Itgb8 deletion renders MSCs resistant to MPL mutant cells, which results in decreased expression of typical fibrotic markers such as Acta2, Col1a1 or S100a8. Most importantly, MF mice treated with ITGB8 neutralizing antibodies exhibit reduced disease burden, as indicated by decreased marrow fibrosis, significantly reduced frequencies of MPL mutant cells, and lower inflammation in the BM. Also, mice with MSC-specific deletion of ITGB8 show significantly lower BM fibrosis and reduced expansion of MF mutant cells upon transplantation with MPLW515L expressing hematopoietic progenitors confirming the phenotype observed with ITGB8 inhibition using neutralizing antibodies. Our data indicate EBF1 and ITGB8 as key mediators of MSC dysfunction and fibrotic transformation and suggest that targeting EBF1-ITGB8 axis in the MF niche may have therapeutic benefits.