Annelise J. Glode

Myelodysplastic syndrome (MDS) is characterized by S100A9-mediated constitutive innate immune activation that leads to the inflammatory death of hematopoietic stem/progenitor cells (HSPCs). This suppressive microenvironment includes the secretion of TGFβ and is phenocopied by the S100A9-transgenic (Tg) mice. However, the role of the different ALK/TGFB receptors in hematopoiesis or in S100A9-mediated inflammation is not understood. Primary low-risk MDS bone marrow mononuclear cells (BM-MNCs) were analyzed in vitro for hematopoietic restoration by an ALK4/ALK5 inhibitor (Vactosertib), an ALK2 inhibitor (TP-0184/Itacnosertib) and an ALK5/TGFBR1-specific inhibitor (TP-6379). Specimens significantly responded to TP-6379 treatment restoring hematopoiesis, with a positive bias for specimens harboring spliceosomal mutations. We previously shown that S100A9 drives phenotypic pathology of MDS, and it has been known to induce and cooperate with TGFβ in this disease. Hence, we treated healthy BM-MNC with recombinant human (rh) S100A9 where TP-6379 rescued the S100A9-reduced hematopoiesis. The effect of TP-6379 was validated in vivo in S100A9-Tg mice. Similar results were obtained with K562 cells, with or without the knock-in (KI) mutation in SF3B1, with strong sensitivity to rhS100A9 that was overcome by TP-6379. Hence, directly targeting TGFBR1/ALK5 with TP-6379 restores healthy hematopoiesis in low-risk MDS preclinical models.

Background: Lenalidomide is an immunomodulatory drug approved in the treatment of autoimmune disease, inflammation, and cancer. Its impact continues to grow due to its diverse spectrum of effects hampered only by toxicities and reduced efficacy. Therefore, development of strategies that enhance function while reducing drawbacks remains a prime goal. Objective and Hypothesis: The mechanisms of action of lenalidomide on the activity of natural killer cells (NK cells) remains understudied yet could be critical for the development of strategies to enhance its efficacy. These cells are critical drivers of anti-tumor immune responses which are often functionally suppressed in malignancies. NK cell and T cell survival and function is driven by the IL-2 family of cytokines (IL-2 or IL-15) and work has shown that lenalidomide potentially works by increasing the secretion of IL-2 by other lymphocytes, such as CD4 ⁺ T helper cells. Thus, we hypothesized that improving NK activity with IL-2 family of cytokines could lead to enhanced lenalidomide-induced responses of these cells. Results: We show that lenalidomide does not affect NK cell viability but reduces their proliferation through cell cycle arrest which could be overcome by exogenous addition of IL-2 family of cytokines. Moreover, lenalidomide induced the secretion of IL-2 on isolated NK cells although it also modulated NK receptor expression, such as NKp46, trough downregulation of PI3K/AKT pathway reduction. This was overcome by exogeneous addition of IL-2 family of cytokines increasing natural cytotoxicity, through higher perforin and granzyme expression. Mechanistically, this increased gene and protein expression occurred through the activation of STAT5 by lenalidomide which was also enhanced through the exogenous addition of IL-2 family of cytokines and modulation of IL-2R subunit changes. Conclusions: These data provide a rationale for the combination of lenalidomide with IL-2 family of cytokines to enhance the effectiveness of NK cells.