Nonmyeloablative Conditioning Regimen Plus A2AR Agonist Oral Administration Promote Engraftment On Allogeneic Bone Marrow Transplantation Mouse Model

Although non-myelaoblative conditioning regimens have proven successful in obtaining engraftment and reducing the toxicity of allogeneic transplant, improvements are still needed to improve engraftment rates as well as limit the incidence of graft versus host disease.  Methods to improve engraftment without added toxicity are especially desirable in the non malignant setting. Agonists of the Gs-coupled Adenosine A_2A receptor (A_2AR) have been reported to promote T cell tolerance in several in vivo models, particularly in the GVHD setting; however it’s role in engraftment is unclear.  In this study, we examined the ability of an A_2AR agonist to promote engraftment in a murine allogeneic bone marrow transplantation model. B6D2F1/J recipients were conditioned with 300 cGy and transplanted 24 hours later via lateral tail vein infusion with 2×10⁶ T-cell depleted bone marrow cells from C57BL/6J (B6) donor cells (H2^kb into H2^kb/d). Recipients received an A_2AR agonist orally in a chow formulation (1000 mg/kg/day) beginning 2weeks before cell infusion and continuing for 9 weeks post transplantation.  A second cohort also received the A_2AR agonist using osmotic pumps (100 ng/kg/min) placed subcutaneously 48 hours prior to cell infusion, for 28days post transplantation as well as orally via the chow for the same time as the first group.  A non-treatment transplant group was used as a control.

There was no apparent toxicity from the radiation or the drug, and all animals maintained normal blood counts throughout. Engraftment was observed in only the A_2AR agonist treated groups and a 13 fold higher level of engraftment in the mice treated both subcutaneously and orally was observed. This engraftment appeared to be long term as the mice were followed out 5 months post transplant with all mice surviving.

As a possible mechanism of this effect, we measured the level of CXCR4 expression on the donor bone marrow cells (Lin^-) in vitro and observed that A_2AR agonist (10 μM) treatment increased CXCR4 expression 1.6 fold compared to non-treatment on donor cells in vitro. To further investigate the effect of the A_2AR agonist on engraftment, we are now repeating the same type of transplant using an A_2AR Knock out mouse strain as the donor.  We are also assessing the levels of SDF-1 in the serum of the recipients.

These preliminary results demonstrate that A_2AR agonists may improve engraftment by increasing homing of the donor cells without increasing the risk of graft versus host disease and may be a significant addition to transplant regimens.

Acknowledgements: This was supported by the Intramural Research Program in the National Institute of Allergy and Infectious Diseases, NIH.