This project involves the conduct of therapeutic clinical trials for the treatment of inherited immune deficiencies using hematopoietic stem cell transplantation. We previously reported the successful use of non-ablative conditioning to achieve successful long term engraftment and cure of CGD patients using HLA-matched sibling donors as the source of the hematopoietic stem cell graft. One of the problems with this approach was the high rate (30%) of graft failure or very low engraftment. In 2004 we performed a follow up transplant on an X-CGD child previously transplanted by us who had achieved high level donor T cell engraftment but less than 1% long term myeloid engraftment. We demonstrated successful permanent conversion to almost 100% donor chimerism in the lymphoid and myeloid lineages using conditioning with only busulfan at 10 mg/kg. This strongly supports the use of this approach to rescue low engraftment rather than using a fully myelo- and lympho-ablative conditioning regimen for such salvage therapy. We have now opened a clinical trial to treat patients with immunodeficiencies using either a matched related, matched unrelated, or cord blood product and a tolerance inducing conditioning regimen consisting of Campath 1-H and busulfan with sirolimus for graft versus host disease (GvHD) prophylaxis. For patients receiving an unrelated product, total body irradiation is also added to the regimen. To date we have transplanted a total of 35 patients, 31 of whom received an unrelated donor graft. In patients not receiving a stem cell boost we have we have seen very limited GvHD with only one patient having Grade 2 skin GvHD and two patients with Grade 1 GvHD of the gut. We have used cord blood in 3 cases. One was for a patient with P67 deficient CGD and another with the X-linked form, both of whom failed to engraft. With the first patient, the thinking was that the Campath impacted negatively with the ability of the more naive T cells in a cord blood graft preventing engraftment, and the protocol was modified to use ATG in the case of a cord blood product. However with the second graft failure in a CGD patient, we have stopped using cord blood products for these patients. This data is also correlated by results from Duke University where even with a myeloablative regimen, the engraftment rate using cord blood products in CGD patients is between 40 to 50%. The third patient to receive a cord blood graft was transplanted for X-linked Severe Combined Immunodeficiency. He was conditioned with ATG instead of Campath and is currently 6 years out with full engraftment, no evidence of graft versus host disease, and normal immune function. We now have had a total of seven deaths on the protocol. The first patient, who had CGD, died due to renal failure unrelated to transplant and subsequent refusal to continue dialysis. The second patient had leukocyte adhesion deficiency and died due to sepsis, despite evidence of engraftment. This patient had failed a previous transplant, was infected at the time of the transplant, and was also highly alloimmunized from previous granulocyte infusions. The third patient was transplanted for Rag 1 deficiency, tolerated transplant extremely well and engrafted successfully but died due to accidental self-inflicted drug overdose. A fourth patient died from sepsis with CMV and KPC despite full engraftment. The 5th patient had engrafted but then rejected the graft late. He then underwent two more transplants, the final one with a very immunosuppressive regimen and did obtain complete engraftment. Unfortunately he died due to adenovirus infection as a result of the more immunosuppressive regimen and the resulting GvHD something we have not otherwise seen with our protocol regimen emphasizing its lower toxicity profile. The most recent patient developed autoimmune hemolytic anemia unresponsive to standard therapies. As this appeared to be recipient directed against donor erythropoiesis, the decision was made to augment the donor chimerism. The patient developed graft versus host disease and required platelet transfusions for GI bleeding. He then developed transfusion related acute lung injury after a platelet infusion. The last patient developed alveolar hemorrhage and CMV pneumonitis and died as a result. Despite these outcomes, our overall mortality rate has been low relative to bone marrow transplantation in general using unrelated donors. Further we have transplanted a number of patients with ongoing infections including fungal osteomyelitis of the spine and/or meninges and in some cases have used granulocyte infusions during the period of transplant-induced neutropenia with no adverse effects. To date, all patients except one with CGD transplanted with an ongoing infection have done well without any significant morbidity or mortality due to infection progression during the transplant course. Overall our results for the CGD patients in particular are especially promising with an overall survival of 5 out of 30 and an overall engraftment rate of 27 out of 30. In 2013 we have transplanted 5 patients with an additional 3 more patients waiting to undergo transplant before 2014. Inorder to determine the outcomes of transplant for CGD in general, we have become a part of the Primary Immune Deficiency Treatment Consortium and have initiated a collaborative protocol to review the results of transplant done for CGD in North America. The protocol was completed in July and has been submitted to the DAIT and extramural NIAID, and implementation and accrual is anticipated to begin within the next few months. In related laboratory pre-clinical studies, we have been investigating the use of an adenosine A2a receptor agonist to prevent or treat graft versus host disease. Prior studies have shown that agonists specific to this receptor improve outcomes in ischemia models of tissue damage. In collaboration with the investigators at the University of Virginia who have supplied a specific agonist known as ATLe146, we tested this drug in a murine model of graft versus host disease. We have seen benefit in attenuating the onset and severity of GVHD in our F1-parental transplant model and have published this data in the Journal of Leukocyte Biology. Further studies have shown a role for T regulatory cells (Tregs) as part of the mechanism of the drugs effects. We have now established a new CRADA with Lewis and Clark Pharmaceuticals (formally Adenosine Therapeutics/PgxHealth/Forest Labs) to study other formulations of the A2a receptor agonists and have seen similar effects on T regs, both invitro and invivo, by these agonists. This data was published in the Journal of Immunology. In addition, we have found that rapamycin also works to reduce TH17 cells in our model and we are studying combination drug therapies now in our model and have evidence that rapamycin may improve engraftment when used in the context of nonmyeloablative regimens. Moreover, we have started to study these drugs in a colitis model given their benefit in GvHD of the gut. We have been working to establish a murine model of CGD related colitis and will begin testing the agonists comparing them to other immunosuppressive therapies. Finally, we have started working with LNC Pharmaceuticals to apply to the NCATS TRND program to complete the studies needed to obtain FDA approval, which is necessary to begin clinical trials using these compounds in transplant studies.
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