Development and conduct of allogeneic stem cell transplant and autologous stem cell gene therapy for inherited immune deficiencies

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The first part of this project involves the development of conditioning regimens for allogeneic transplantation of patients with primary immunodeficiencies. In 2007 we used busulfan, Campath and low dose TBI and treated 44 patients with CGD, 39 of whom received an unrelated donor (MUD) graft. The results were published in the Journal of Clinical Immunology. (Parta et al. JCI). Included in this cohort were two patients with the P40 form of CGD demonstrating complete reversal of refractory colitis in this unique subset. In follow up we opened a new protocol using a higher cell dose and post-transplant cyclophosphamide to improve engraftment but mitigate the risk of Graft versus Host Disease (GvHD) from the larger graft. We initially enrolled 10 patients with 3 deaths (2 with related donor grafts) due to progressive pulmonary disease and one patient with graft loss who did not receive post-transplant cyclophosphamide. A retrospective evaluation of all patients suggested that an elevated C reactive protein (CRP) prior to the transplant itself was the one common risk factor and the protocol was thus modified to exclude patients who have an elevated CRP. We have subsequently transplanted 26 more patients using unrelated donor grafts and 3 with related donor grafts (about 1 patient a month after a hiatus due to COVID) with good outcomes except for one graft loss despite initial engraftment and a second graft loss in a patient who had rejected a prior graft. Both these patients remain alive and well. The remainder of the patients have all done well with no severe GvHD but two patients expired due to their underlying disease. We also transplanted 2 patients as exemptions to the protocol (1 with an elevated CRP) with one patient expiring due to progression of his underlying infection and the other doing well despite extensive disease and functional quadriplegia prior to transplant. In order to improve outcomes in terms of mixed chimerism as well as to be able to treat patients who would be excluded due to their elevated CRP, we have a new protocol, 0009777 which includes the use of pretreatment with tocilizumab and in high-risk patients, emapalumab. This protocol enrolled its first patient in July of 2022 with the patient engrafting without GvHD, and ongoing resolution of his invasive fungal infection. We have also initiated a protocol using an anti-cKIT antibody to replace the busulfan conditioning in an effort to reduce overall toxicity. We have enrolled three patients to date with two patients having engrafted and the third early in the transplant period. Patients have had less toxicity, particularly mucositis during the transplant course. To further expand eligibility, in 2014 we opened a protocol using haploidentical donors. The 1st patient had an ongoing infection refractory to all standard therapy involving the heart and is now 6 years out with complete resolution of his infection. (J Clin Immunol. 2015 Oct;35(7):675-80). We enrolled a total of 7 patients on this protocol but saw severe GvHD in the last 3 patients. This protocol is now closed and a new protocol is now open to accrual (19-I-0080). This new protocol used both early and late Campath along with busulfan, TBI, and post-transplant cyclophosphamide. The first patient did very well with full engraftment, and no GvHD. The second patient developed significant GvHD, thus the protocol was modified to change the timing of the Campath. The third patient then did well with this modification but the fourth patient developed a rare complication known as ADEM. As such the protocol has been further modified to use bone marrow cells instead of peripheral blood stem cells but has not accrued any patients to date due to COVID. A protocol for X-linked and JAK-3 SCID (20-I-0080) has also been opened for accrual, but no patients have yet been enrolled again due to repercussions of COVID as well as staffing issues. We are also working towards developing antibody based conditioning for atypical SCID patients similar to CGD. One patient with Rag1 was enrolled, but engraftment was poor and the protocol is now on hold. As a member of the Primary Immune Deficiency Treatment Consortium (J Allergy Clin Immunol. 2014 Feb;133(2):335-47) we developed a collaborative protocol (6903) to review the results of transplants done for CGD in North America. We enrolled over 100 transplanted patients and published the results on a subgroup of patients with inflammatory bowel disease (Marsh et al, JCI 2019). We have also submitted the overall data which was accepted for publication in Blood. We have also been involved in a microbiome analysis, (a substudy done in collaboration with Emilia Falcone) with a manuscript being published this year in JACI. We are also finalizing a new CGD related protocol (6908). In the laboratory, utilizing our established murine models of GvHD, we have modified the conditioning regimens to induce graft rejection and/or engraftment syndrome with various cytokines to mimic the inflammatory milieu seen in patients as well as manipulating the graft cell composition to assess any donor graft effects. Post-doctoral fellow Andres Zea-Vera has murine data showing a negative impact of high levels of Il-6 and Interferon gamma suggesting that the high CRP and associated pathways seen in patients is directly responsible for the poor outcomes. This data has been presented the CIS and ASH annual meetings. We have also initiated a collaboration with the CHI to perform RNA sequence analysis of bone marrow samples to further assess the inflammation seen in the hematopoietic niche of CGD patients and its impact on engraftment. The second part of this project involves the use of genetically modified autologous cells for the treatment of patients with XCGD and other immunodeficiencies. We initiated a clinical trial in 2006 protocol 07-I-0017. Based on preclinical data in the rhesus as well as clinical data in a patient, we used busulfan at a dose of 10mg/kg prior to infusion of the genetically modified cells. We treated three patients, the results of which were published in Blood. In 2015 we developed a collaborative study, Protocol 15-I-0008, using a lentiviral vector for XCGD. The 1st NIH patient continues to have marking in the 20-30% range. The 2nd NIH patient is now over 4 years post-transplant with persistent high-level marking of 40%. Our 3rd patient was treated in 2017 and unfortunately developed autoimmune thrombocytopenia, unrelated to the gene therapy, and died of a cerebral hemorrhage. Our most recent patient has more than 70% oxidase cells at his last evaluation, 3 years post gene therapy. A total of 9 patients have been treated at the various sites with 3 patients, having loss of their marking. The results, including patients from a London trial that used the same type of conditioning and vector have been published. (Kohn et al. Nature Medicine 2020.) We are also developing a collaborative study to treat patients with the P47 autosomal recessive form of CGD using a lentiviral vector with submission for the IND planned by the late summer and enrollment to begin early winter of 2023. Future plans will include incorporating transduction enhancers to improve transduction. The protocol has been delayed due to COVID and supply issues. Uimook Choi and Nicole Fama (now at medical school) have also developed vectors for the P67 and P22 forms of CGD which we are hoping to use in the future. Finally, Karissa Bever (now attending graduate school) has worked to further optimize a CARD 9 lentivector originally created by Caroline Kreitzer, a former post doc now at medical school. She has assayed this in a murine model, studies which Hula Bayo, a new post bac will continue including using a fungal challenge model to evaluate efficacy.