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

该项目的第一部分涉及原发性免疫缺陷患者同种异体移植的调理方案的发展。2007年，我们启动了一项使用busulfan、Campath和低剂量TBI的临床方案，治疗了44例CGD患者，其中39例接受了非亲属供体（MUD）移植。研究结果发表在《临床免疫学杂志》上。(Parta等。江森自控)。在该研究的后续研究中，我们在之前的研究结果的基础上建立了一个新的方案模型，使用更高的细胞剂量和移植后环磷酰胺来改善移植，同时减轻较大移植物引起的移植物抗宿主病（GvHD）的风险。我们最初招募了10例患者，其中3例死亡（2例有相关的供体移植物），原因是肺部疾病进展，1例移植物丢失，移植后未接受环磷酰胺治疗。对所有移植患者的回顾性评估表明，移植前C反应蛋白（CRP）升高是一个常见的危险因素，因此修改了方案以排除CRP升高的患者。随后，我们又移植了24名患者使用不相关的供体移植物，3名患者使用相关的供体移植物（因COVID而中断后约1名患者一个月）。大多数患者都表现良好，没有患者发展为严重的GvHD，但有两名患者因其潜在疾病而死亡。我们还移植了2例患者作为方案的豁免（1例CRP升高），其中1例患者因其潜在感染的进展而死亡，另1例患者在移植前尽管存在广泛的疾病和功能性四肢瘫痪，但情况良好。为了改善混合嵌合的结果，以及能够治疗因CRP升高而被排除在外的患者，我们现在已经开放了一个新的方案，0009777，其中包括使用tocilizumab预处理和在高风险患者中使用emapalumab。该方案于7月招募了第一个患者，迄今为止情况良好。我们也有一个正在接受监管审查的方案，它将使用抗ckit抗体来取代我们在所有其他方案中使用的布苏凡，以努力实现使用无化疗调节方案的植入。

The first part of this project involves the development of conditioning regimens for allogeneic transplantation of patients with primary immunodeficiencies. In 2007 we initiated a clinical protocol using 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). In follow up to that study we opened a new protocol modeling on our previous results using a higher cell dose and post-transplant cyclophosphamide to improve engraftment but mitigate the increased 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 transplanted 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 24 more patients using unrelated donor grafts and 3 with related donor grafts (about 1 patient a month after a hiatus due to COVID). The majority of the patients have all done well with no patients developing severe GvHD but two patients have 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 now opened a new protocol, 0009777 which includes the use of pretreatment with tocilizumab and in high risk patients, emapalumab. This protocol enrolled it first patient in July who is doing well to date. We also have a protocol undergoing regulatory review which will use an anti-cKit antibody to replace the busulfan we have been using in all our other protocols in an effort to achieve engraftment using a chemotherapy free conditioning regimen. To further expand eligibility, in 2014 we opened a protocol using haploidentical donors. The 1st patient is now 5 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 with 2 of the patients succumbing to its' complications but the 3rd patient recovered and is now doing well. This protocol was closed and a new protocol was opened to accrual in 2019 (19-I-0080). This protocol initially 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 and we will treat a patient this winter. 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 due to COVID complications. We are also working towards developing antibody based conditioning for atypical SCID patients similar to CGD. 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 are now in the midst of analyzing the data from the overall study with a manuscript planned for publication this late fall/early winter. We have also been involved in a microbiome analysis, (a substudy done in collaboration with Emilia Falcone) with a manuscript also in preparation, and collaborated on a study evaluating granulocyte infusions. (J Clin Immunol. 2022 Apr 21;1-10) We are also finalizing a new CGD related protocol (6908) which will specifically evaluate the autoinflammatory aspects of CGD patients pre and post-transplant. 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 preliminary data showing a negative impact of high levels of Il-6 suggesting that the high CRP seen in patients is directly responsible for the poor outcomes and has an abstract submitted for presentation at the ASH meeting this year. Although significant progress has been made in these efforts, there have been delays due to Covid-19. 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 to treat XCGD patients and an underlying infection, 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. The results in 3 patients were published in Blood. In 2015 we then developed a mutlicenter study, Protocol 15-I-0008, using a lentiviral vector for XCGD. The 2nd patient on the trial was treated at NIH in 2016, and continues to have marking in the 20-30% range more than 3 years post treatment. 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 was treated in 2019 and is doing well with close to 80% oxidase cells, 3 years post gene therapy. The results, including patients from a London trial have been published. (Kohn et al. Nature Medicine 2020.) An addendum protocol to treat pediatric patients with the same vector but using a modified conditioning regimen was approved and the first patient was treated in late November 2021, but ultimately also lost his graft. Thus the protocol is now being further amended. We are also developing a collaborative study to treat patients with the P47 autosomal recessive form of CGD using a lentiviral vector and hope to start enrolling patients by early 2023. (Hum Gene Ther. 2021 Sep;32(17-18):949-958) Future plans will include incorporating transduction enhancers to improve the efficiency of transduction and reduce the amount of vector needed to treat individual patients. The protocol has been delayed due to COVID and supply issues, but should go for FDA review this fall/winter. We are also planning to involve the use of antibody-based conditioning depending on the results in our allogeneic transplant study. Uimook Choi and Nicole Fama (now at medical school) have also developed vectors for the P67 and P22 forms of CGD for future gene therapy studies. Finally, Karissa Bever who started in 2021, has been optimizing a CARD 9 lentivector created by Caroline Kreitzer, (now at medical school). Karissa has now modified this vector for a more clinically relevant version and will be using it in a murine challenge study this fall.