EMU-116: An oral CXCR4 antagonist as mobilizer of stem and immune cells in normal, neutropenic and sickle cell mice.

Abstract

Abstract The use of CXCR4 antagonists as cell mobilization agents has resulted in 3 FDA approved drugs for use in hematopoietic stem cell transplantation (Plerixafor, Motixafortide) and for treating neutropenia (Mavorixafor). Given these developments, the need for agents which can be delivered orally is apparent. An oral mobilizer could find uses in different disease conditions including in-patient gene-based therapy for sickle cell cure and chronic neutropenia or that which is induced by chemotherapy. Herein we describe the effects of the novel CXCR4 antagonist EMU-116 on the ability to mobilize stem and immune cells in naïve, neutropenic and sickle cell disease mouse breeds when dosed orally. In DBA mice, single doses of EMU-116 and Plerixafor were conducted to observe short-term effects on the blood and bone marrow compartments. EMU-116 was dosed in both sub-cutaneous (s.c.; 10mg/kg) and oral (p.o.; 30 mg/kg) routes along with Plerixafor (s.c.; 5mg/kg) and the cell types were measured over an 8-hour period using CBC and FACS analysis. EMU-116 showed robust and sustained increases in multiple immune and stem cell types including white blood cells (WBCs), neutrophils, T-cells and LSK+ cells superior to Plerixfor. The LSK and CFU counts correlated well, despite limited amounts of blood that collected at each time point. While Plerixafor peaked at around 2-hours, the responses from EMU-116 were more robust and tended to peak around the 6-hour time point matching the peak plasma levels seen in mouse PK studies with EMU-116. In general, EMU-116 induces “pan-mobilization” of mature hematopoietic subsets with the magnitude of fold changes from baseline ranging from 40-fold (for pDCs) to 2-fold (for eosinophils). It is also notable that EMU-116 induced no significant changes compared to the vehicle control in all the red blood cell indices (RBC no., HCT, MCV, MCH, Hb, etc.). In a mouse model of neutropenia, Swiss mice were used in a model to simulate the effects of chemotherapy on immune cells. The mice were pre-conditioned by dosing with cyclophosphamide (60mg/kg, i.p.q.d.) for 4 days and on day 5 EMU-116 was dosed (30 mg/kg, p.o.q.d.) for an 8-day period. Here, EMU-116 treated mice rebounded in WBCs, neutrophils, lymphocytes, and monocytes 3 days earlier than the control groups. Increases in HSCs were also observed earlier indicating faster recovery in the bone marrow. The responses were robust (5 to 20-fold above baseline) and sustainable in that the height and downward return happened within a 3-day period. In Townes mice (genetically induced sickle cell disease; SS), EMU-116 was studied to determine the feasibility as a stem cell mobilization agent for sickle cell gene therapy. A single time point was used which matched the previous peak effects of EMU-116 (6 hours), The study, involving 6 Townes sickle mice (SS; 3M, 3F), was conducted where EMU-116 was dosed via s.c. (4mg/kg) and oral (30 mg/kg; p.o.) routes and compared to Plerixafor (s.c.; 5mg/kg). The effects were mobilization of multiple cell types, including WBCs and LSK cells where both routes of EMU-116 showed mobilization of more LSK cells without the enhanced leukocytosis observed with Plerixafor. In summary, studies in multiple mouse breeds show that the CXCR4 antagonist EMU-116 serves as a mobilization agent in several settings. In all the studies, mobilization of WBCs and HSCs were observed with no changes in the red blood cell components. In Swiss mice, EMU-116 dosed orally provides enhanced responses to cyclophosphamide indicating a potential use in chemotherapy-induced neutropenia. In sickle cell mice, EMU-116 boosts LSKs when dosed orally and could possibly be utilized for a future in vivo gene therapy setting with superior effects to Plerixafor.