Pai, S-Y

Thrasher, AJ

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UCL Discovery

Journal Pre-proofGene therapy for X-linked severe combined immunodeficiency: historical outcomesand current statusSung-Yun Pai, MD, Adrian J. Thrasher, MB PhD FMedSciPII: S0091-6749(20)30837-XDOI: https://doi.org/10.1016/j.jaci.2020.05.055Reference: YMAI 14632To appear in: Journal of Allergy and Clinical ImmunologyReceived Date: 16 April 2020Revised Date: 21 May 2020Accepted Date: 26 May 2020Please cite this article as: Pai S-Y, Thrasher AJ, Gene therapy for X-linked severe combinedimmunodeficiency: historical outcomes and current status, Journal of Allergy and Clinical Immunology(2020), doi: https://doi.org/10.1016/j.jaci.2020.05.055.This is a PDF file of an article that has undergone enhancements after acceptance, such as the additionof a cover page and metadata, and formatting for readability, but it is not yet the definitive version ofrecord. This version will undergo additional copyediting, typesetting and review before it is publishedin its final form, but we are providing this version to give early visibility of the article. Please note that,during the production process, errors may be discovered which could affect the content, and all legaldisclaimers that apply to the journal pertain.© 2020 Published by Elsevier Inc. on behalf of the American Academy of Allergy, Asthma &Immunology.Gene therapy for X-linked severe combined immunodeficiency: historical outcomes and current status  Sung-Yun Pai MD1 and Adrian J. Thrasher MB PhD FMedSci2  1 Division of Hematology-Oncology, Boston Children’s Hospital and Department of Pediatric Oncology, Dana-Farber Cancer Institute 2 University College London Great Ormond Street Institute of Child Health, and Great Ormond Street Hospital for Children NHS Foundation Trust, London, UK  Correspondence to: Professor Adrian J. Thrasher Great Ormond Street Institute of Child Health University College London 30 Guilford Street WC1N 1EH Email: a.thrasher@ucl.ac.uk Direct Telephone: +44 (0)20 79052660 PA Chris Thalasselis: +44 (0)20 7905 2788 Fax: +44 (0)20 7905 2810  Key words Severe combined immunodeficiency; gene therapy; T cell; B cell  Word count: 1315  Conflict of interest disclosures: S. -Y.P. is employed by Boston Children’s Hospital and has received funding from the National Institutes of Health. AJT is a Co-founder of Orchard Therapeutics https://www.orchard-tx.com/. AJT is supported by funding from The Wellcome Trust (217112/Z/19/Z AJT) and National Institute for Health Research Biomedical Research Centre at Great Ormond Street Hospital for Children NHS Foundation Trust and University College London.  SCID-X1 background and outcomes of standard treatment  Severe combined immunodeficiency disorder (SCID) is a genetically heterogeneous group of disorders characterized by a profound absence of T lymphocyte function, resulting in lack of cellular and humoral immunity. The X-linked form (SCID-X1), accounts for 30-40% of cases, and is caused by defects in the common cytokine receptor γ chain (γc), encoded by the IL2RG gene. Originally identified as a component of the high affinity interleukin-2 receptor, γc is an essential component of the IL-4, -7, -9 -15, and -21 cytokine receptor complexes(1) (Figure 1A). The molecular defect in SCID-X1 results in the complete absence of T cell and natural killer cell development and a defect of terminal B cell maturation and function. The prognosis without treatment is uniformly fatal due to recurrent and opportunistic infection.   Allogeneic hematopoietic stem cell transplantation (allo-HSCT) is curative for the disorder, however outcomes remain suboptimal for certain patients.(2) The Primary Immune Deficiency Treatment Consortium (PIDTC) published two retrospective studies of allo-HSCT outcomes, a study of 240 patients with typical SCID and a larger recent study of 662 patients with typical, leaky or Omenn SCID.(2) Both studies confirmed and refined the factors important for survival such as the presence of active infection, use of donors other than matched siblings, and age at transplant. The latter study for first time reported outcomes that are specific to certain genetic types of SCID, showing that patients with SCID-X1 have superior survival.   The lack of functional T and NK cells in SCID-X1 underlies the ability of many patients to develop a T cell graft without conditioning. Donor-derived HSC and progenitors (HSPC) seed the thymus, and can potentially sustain T cell generation for many years. However, without the use of myelosuppressive conditioning agents, cells other than T and NK cells remain largely of host origin. SCID-X1 patients who do not receive conditioning pre-transplant are therefore less likely to become independent of immunoglobulin substitution or respond to vaccination(2). Recent studies have clearly demonstrated that conditioning promotes more complete immune reconstitution in particular using the alkylating agent busulfan, which can be adjusted with pharmacokinetics to achieve a controlled drug exposure and limit toxicity.  The success of standard allo-HSCT remains limited due to the lack of matched donors for all patients, and importantly the complications of acute and chronic graft-versus-host disease (GVHD). Even in this young age group, the incidence of acute and chronic GVHD is approximately 20-25% and up to 16% respectively(2). Furthermore, use of mismatched donors, particularly haploidentical related donors was associated with greater risk of needing a second allo-HSCT(2). Transduction of autologous HSPC cells with an integrating retroviral vector that expresses a normal copy of the gene causing SCID-X1, would obviate the need to find a matched donor and eliminate the risk of GVHD and graft rejection.   Safety of gene therapy for SCID-X1 has improved with use of self-inactivating vectors  A summary of past and current trials of gene therapy for SCID-X1 is shown in Table 1. Seminal clinical trials were performed in Paris and London targeting high-risk patients in hopes of achieving robust immune reconstitution without the risk of GVHD intrinsic to allo-HSCT. CD34+ cells were purified from bone marrow and transduced with a vector expressing the IL2RG transgene (MFG-γc). Overall, 20 boys underwent gene therapy with rapid T cell reconstitution in 18 patients. Among the 17 survivors, all have been generally free of SCID-related infections.(3, 4)   The MFG-γc vector is a gammaretroviral (γRV) vector with intact long-term repeat regions (LTR) containing strong viral enhancers and promoters. These elements drove strong transgene expression and hence rapid T cell reconstitution, but also were associated with insertional oncogenesis. Of 20 boys treated, 6 (5 in Paris, 1 in London) developed clonal T cell proliferation phenotypically mimicking acute T cell lymphoblastic leukemia, each arising from clones bearing insertion sites near genes with lymphoid oncogenic potential (LMO2, CCND2, BMI1). The latency of leukemogenesis using the MFG-γc vector was long (2-15 years).(5)   One study directly tested the hypothesis that removing the strong viral elements from the LTR would deter overexpression of neighboring genes and reduce or eliminate insertional oncogenesis. The parent MFG-γc vector was modified to remove the strong viral elements from LTR, driving expression of the IL2RG transgene instead by a cellular promoter, elongation factor 1a short (EFS). This self-inactivating γRV vector (SIN-γc) was tested in parallel phase I/II gene therapy trials in United States, France and United Kingdom(6). T cell recovery in patients receiving adequate VCN was robust, and to date, none of the 14 patients in follow-up in the SIN-γc trial have developed leukemia (2.7-9.3 years, median 7.9 years). Interim analysis of 9 patients comparing insertion site pattern in the blood of patients receiving the MFG-γc vector to those receiving the SIN-γc vector demonstrated a strong reduction in clustering near LMO2 and CCND2.(6) The global distribution of insertion sites between the two vectors is indistinguishable with regard to transcription start sites, gene density and epigenetic marks.   Safety is expected to be further improved with the use of self-inactivating lentiviral (SIN-LV) vectors, which have by nature a different distribution of insertion sites and do not favor transcription start sites. Three trials of gene transfer for SCID-X1 using SIN-LV, one in older patients who have failed allogeneic HSCT (NCT 01306019),(7) and two in newly diagnosed infants in US (NCT015132888)(8) and internationally (NCT03311503) have not reported any oncogenesis to date, albeit with short follow-up.   Conditioning improves humoral function after gene therapy for SCID-X1 In trials without chemotherapy conditioning, gene marking was extremely low or absent in B cells and in granulocytes,(3, 4, 6) and in general these patients have failed to mount vaccine specific immunoglobulin. In the absence of conditioning, allo-HSCT and gene therapy promote T cell development due to the ability of the infused HSC and committed progenitors to engraft and persist in the empty thymus of patients with SCID (Figure 1B). However, HSC remain deficient in IL2RG and the progeny of those HSC are therefore unresponsive to γc-dependent cytokines, such as IL-21, which are critical for B cell maturation into antibody-producing plasma cells. Conditioning is required to replace HSC with cells expressing γc that can develop into IL-21-responsive B cells. Supporting this notion, a PIDTC study of 48 SCID-X1 or JAK3 SCID patients post-allo-HSCT showed that chemotherapy conditioning was associated with restoration of in vitro response of B cells to IL-21 and clinical B cell function.(9) Use of busulfan conditioning prior to gene therapy would be expected to promote engraftment of gene-marked HSC and therefore gene-marked B cells capable to responding to T cell help (Figure 1B). The last patient enrolled on the SIN-γc trial received low busulfan conditioning and demonstrated recovery of gene marked B cells and humoral immune function (S-YP, unpublished results). All of the 3 currently open and enrolling SIN-LV trials use low dose busulfan.(7, 8) In a trial of 8 infants, 4/8 discontinued IVIG replacement therapy and 3 of these infants responded to vaccines.(8)   Future Directions While the busulfan dose used in current trials is low, approximately 1/3 of the myeloablative dose, exposing these young infants to alkylating agents is not without consequences. In addition to potential long-term toxicities such as impairment of linear growth, infertility, and school function problems, myelodysplastic changes may occur in the marrow and has been reported after autologous gene therapy for other disorders. Using nontoxic conditioning, for example using depleting antibodies that target HSPC, before gene therapy would further improve the safety profile of this treatment. Finally, all of the current trials use integrating vectors as a “gene addition” approach, which predisposes not only to oncogenesis, but also fails to completely recapitulate natural gene expression patterns. Gene editing on the other hand, could be used to correct a mutation in situ, or insert a working copy of the gene into the natural locus, allowing endogenous control of expression. Based on encouraging pre-clinical proof of concept, gene editing for SCID-X1 may also soon find clinical application.(10) Licensure of these technologies is anticipated to change the standard treatment for SCID-X1.  Acknowledgements S.-Y.P. is supported by the National Institutes of Health (U01-AI125051, U01-AI126612) and receives salary support from Boston Children’s Hospital. AJT is supported by funding from The Wellcome Trust (217112/Z/19/Z) and National Institute for Health Research Biomedical Research Centre at Great Ormond Street Hospital for Children NHS Foundation Trust and University College London.   References Cited  1. Leonard WJ, Lin J-X, O'Shea JJ. The γc Family of Cytokines: Basic Biology to Therapeutic Ramifications. Immunity. 2019;50(4):832-50. 2. Haddad E, Logan BR, Griffith LM, Buckley RH, Parrott RE, Prockop SE, et al. SCID genotype and 6-month posttransplant CD4 count predict survival and immune recovery. Blood. 2018;132(17):1737-49. 3. Hacein-Bey-Abina S, Hauer J, Lim A, Picard C, Wang GP, Berry CC, et al. Efficacy of gene therapy for X-linked severe combined immunodeficiency. N Engl J Med. 2010;363(4):355-64. 4. Gaspar HB, Cooray S, Gilmour KC, Parsley KL, Adams S, Howe SJ, et al. Long-term persistence of a polyclonal T cell repertoire after gene therapy for X-linked severe combined immunodeficiency. Sci Transl Med. 2011;3(97):97ra79. 5. Wu C, Dunbar CE. Stem cell gene therapy: the risks of insertional mutagenesis and approaches to minimize genotoxicity. Front Med. 2011;5(4):356-71. 6. Hacein-Bey-Abina S, Pai S-Y, Gaspar HB, Armant M, Berry CC, Blanche S, et al. A Modified γ-Retrovirus Vector for X-Linked Severe Combined Immunodeficiency. N Engl J Med. 2014;371(15):1407-17. 7. De Ravin SS, Wu X, Moir S, Anaya-O&apos;Brien S, Kwatemaa N, Littel P, et al. Lentiviral hematopoietic stem cell gene therapy for X-linked severe combined immunodeficiency. Sci Transl Med. 2016;8(335):335ra57. 8. Mamcarz E, Zhou S, Lockey T, Abdelsamed H, Cross SJ, Kang G, et al. Lentiviral Gene Therapy Combined with Low-Dose Busulfan in Infants with SCID-X1. N Engl J Med. 2019;380(16):1525-34. 9. Miggelbrink AM, Logan BR, Buckley RH, Parrott RE, Dvorak CC, Kapoor N, et al. B-cell differentiation and IL-21 response in IL2RG/JAK3 SCID patients after hematopoietic stem cell transplantation. Blood. 2018;131(26):2967-77. 10. Schiroli G, Ferrari S, Conway A, Jacob A, Capo V, Albano L, et al. Preclinical modeling highlights the therapeutic potential of hematopoietic stem cell gene editing for correction of SCID-X1. Sci Transl Med. 2017;9(411).   Figure Legends Figure 1A: The role of γc in multiple cytokine receptor complexes and cell type specific functions.  Figure 1B: Immune reconstitution after gene therapy for SCID-X1. Without treatment, T cells fail to develop and B cells do not express IL2RG. Without conditioning, gene marked progenitors seed the thymus and give rise to gene-marked T cells. B cells remain unresponsive to γc-dependent cytokines, remain naïve (light blue), and fail to mature into antibody producing B cells. With conditioning, gene marked HSC continue to seed the thymus with progenitors, sustaining T cell production. Gene marked HSC also develop into gene marked B cells that express γc, respond to IL-21, mature into memory B cells (dark blue), and secrete antibody.  Table: Historical and current trials of gene therapy for SCID-X1  Vector Year activated ClinicalTrials.gov number Sites Conditioning regimen Insertional oncogenesis Target Population Status γRV 1999 n/a Paris, France none Yes Infants Completed  2001 n/a London, UK none Yes Infants Completed γRV 2001 NCT00028236 NIH, USA Low dose busulfan None reported Post-transplant, > 18 months old Completed SIN- γRV 2010 NCT01129544 Boston, Los Angeles, Cincinnati, USA none None reported Infants Active not recruiting 2010 NCT01175239 London, UK none None reported Infants Unknown 2011 NCT01410019 Paris, France none None reported Infants Unknown SIN-LV 2012 NCT01512888 St. Jude, UCSF, Seattle, USA Low dose busulfan None reported Infants Recruiting 2017 NCT03315078 NIH, USA Low dose busulfan None reported Post-transplant, > 2 years old Recruiting SIN-LV 2017 NCT03311503 Boston, Los Angeles, USA Low dose busulfan None reported Infants Recruiting 2018 NCT03601286 London, UK Low dose busulfan None reported Infants Recruiting SIN-LV 2017 NCT0321761 Shenzhen, China Unknown None reported Infants Recruiting SIN-LV 2020 NCT04286815 Chongqing, China Unknown None reported Infants Recruiting  

Gene therapy for X-linked severe combined immunodeficiency: historical outcomes and current status

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Gene therapy for X-linked severe combined immunodeficiency: historical outcomes and current status

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