Review Article | Published:

Gene therapy targeting haematopoietic stem cells for inherited diseases: progress and challenges

Nature Reviews Drug Discovery (2019) | Download Citation

Abstract

Pioneering gene therapy trials have shown that the genetic engineering of haematopoietic stem and progenitor cells can be an alternative to allogeneic transplantation in the treatment of primary immunodeficiencies. Early trials also highlighted the risk of insertional mutagenesis and oncogene transactivation associated with the first generation of gammaretroviral vectors. These events prompted the development of safer, self-inactivating lentiviral or gammaretroviral vectors. These lentiviral vectors have been successfully used to treat over 200 patients with 10 different haematological disorders (including primary immunodeficiencies, haemoglobinopathies and metabolic disorders) and for the generation of chimeric antigen receptor-T cells for cancer therapy. However, several challenges, such as effective reconstitution during inflammation, remain if gene therapy is to be extended to more complex diseases in which haematopoietic stem and progenitor cells can be altered by the disease environment. We discuss the progress made and future challenges for gene therapy and contrast gene therapy with gene-editing strategies.

Access optionsAccess options

Rent or Buy article

Get time limited or full article access on ReadCube.

from$8.99

All prices are NET prices.

Additional information

Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

References

  1. 1.

    Poletti, V. & Mavilio, F. Interactions between retroviruses and the host cell genome. Mol. Ther. Methods Clin. Dev. 8, 31–41 (2018).

  2. 2.

    Hacein-Bey-Abina, S. et al. Efficacy of gene therapy for X-linked severe combined immunodeficiency. N. Engl. J. Med. 363, 355–364 (2010). This study is the first HSC gene therapy trial using a gammaretroviral vector to efficiently treat SCID. This trial also highlights the genotoxicity of the gammaretroviral vector, and it has led to the development of safer SIN vectors.

  3. 3.

    Aiuti, A. et al. Lentiviral hematopoietic stem cell gene therapy in patients with Wiskott-Aldrich syndrome. Science 341, 1233151 (2013). This study is the first trial of HSC gene therapy to treat WAS safely and successfully with a lentiviral vector.

  4. 4.

    Hacein-Bey Abina, S. et al. Outcomes following gene therapy in patients with severe Wiskott-Aldrich syndrome. JAMA 313, 1550–1563 (2015).

  5. 5.

    Cooper, A. R. et al. Cytoreductive conditioning intensity predicts clonal diversity in ADA-SCID retroviral gene therapy patients. Blood 129, 2624–2635 (2017).

  6. 6.

    Thompson, A. A. et al. Gene therapy in patients with transfusion-dependent β-thalassemia. N. Engl. J. Med. 378, 1479–1493 (2018).

  7. 7.

    Ribeil, J.-A. et al. Gene therapy in a patient with sickle cell disease. N. Engl. J. Med. 376, 848–855 (2017). This work shows, for the first time, the correction of the clinical phenotype in a patient with SCD after lentiviral-based gene therapy.

  8. 8.

    Biffi, A. et al. Lentiviral hematopoietic stem cell gene therapy benefits metachromatic leukodystrophy. Science 341, 1233158 (2013).

  9. 9.

    Sessa, M. et al. Lentiviral haemopoietic stem-cell gene therapy in early-onset metachromatic leukodystrophy: an ad-hoc analysis of a non-randomised, open-label, phase 1/2 trial. Lancet 388, 476–487 (2016).

  10. 10.

    Maetzig, T., Galla, M., Baum, C. & Schambach, A. Gammaretroviral vectors: biology, technology and application. Viruses 3, 677–713 (2011).

  11. 11.

    Naldini, L. et al. In vivo gene delivery and stable transduction of nondividing cells by a lentiviral vector. Science 272, 263–267 (1996).

  12. 12.

    Schambach, A., Zychlinski, D., Ehrnstroem, B. & Baum, C. Biosafety features of lentiviral vectors. Hum. Gene Ther. 24, 132–142 (2013).

  13. 13.

    Dunbar, C. E. et al. Gene therapy comes of age. Science 359, eaan4672 (2018).

  14. 14.

    Cavazzana-Calvo, M. et al. Gene therapy of human severe combined immunodeficiency (SCID)-X1 disease. Science 288, 669–672 (2000).

  15. 15.

    Picard, C. et al. Primary immunodeficiency diseases: an update on the classification from the International Union of Immunological Societies Expert Committee for Primary Immunodeficiency 2015. J. Clin. Immunol. 35, 696–726 (2015).

  16. 16.

    Hershfield, M. Adenosine deaminase deficiency. GeneReviews https://www.ncbi.nlm.nih.gov/books/NBK1483/ (updated 16 Mar 2017).

  17. 17.

    Cicalese, M. P. et al. Gene therapy for adenosine deaminase deficiency: a comprehensive evaluation of short- and medium-term safety. Mol. Ther. 26, 917–931 (2018). This study reports the safety and efficacy of gammaretroviral-based gene therapy for ADA (with no abnormal clonal proliferation observed in more than 50 patients worldwide). This ADA gene therapy has received marketing approval from the European Medicines Agency (Strimvelis, from GlaxoSmithKline).

  18. 18.

    Kohn, D. B. et al. Consensus approach for the management of severe combined immune deficiency caused by adenosine deaminase deficiency. J. Allergy Clin. Immunol. https://doi.org/10.1016/j.jaci.2018.08.024 (2018).

  19. 19.

    Carbonaro, D. A. et al. Gene therapy/bone marrow transplantation in ADA-deficient mice: roles of enzyme-replacement therapy and cytoreduction. Blood 120, 3677–3687 (2012).

  20. 20.

    Engel, B. C. et al. Prolonged pancytopenia in a gene therapy patient with ADA-deficient SCID and trisomy 8 mosaicism: a case report. Blood 109, 503–506 (2007).

  21. 21.

    Sauer, A. V. et al. Alterations in the brain adenosine metabolism cause behavioral and neurological impairment in ADA-deficient mice and patients. Sci. Rep. 7, 40136 (2017).

  22. 22.

    Meneghini, V. et al. Pervasive supply of therapeutic lysosomal enzymes in the CNS of normal and Krabbe-affected non-human primates by intracerebral lentiviral gene therapy. EMBO Mol. Med. 8, 489–510 (2016).

  23. 23.

    Biffi, A. Hematopoietic gene therapies for metabolic and neurologic diseases. Hematol. Oncol. Clin. North Am. 31, 869–881 (2017).

  24. 24.

    Pai, S.-Y. et al. Transplantation outcomes for severe combined immunodeficiency, 2000–2009. N. Engl. J. Med. 371, 434–446 (2014).

  25. 25.

    Gennery, A. R. et al. Transplantation of hematopoietic stem cells and long-term survival for primary immunodeficiencies in Europe: entering a new century, do we do better? J. Allergy Clin. Immunol. 126, 602–611 (2010).

  26. 26.

    Gaspar, H. B. et al. Gene therapy of X-linked severe combined immunodeficiency by use of a pseudotyped gammaretroviral vector. Lancet 364, 2181–2187 (2004).

  27. 27.

    Cavazzana, M., Six, E., Lagresle-Peyrou, C., André-Schmutz, I. & Hacein-Bey-Abina, S. Gene therapy for X-linked severe combined immunodeficiency: where do we stand? Hum. Gene Ther. 27, 108–116 (2016).

  28. 28.

    Gaspar, H. B. et al. Long-term persistence of a polyclonal T cell repertoire after gene therapy for X-linked severe combined immunodeficiency. Sci. Transl Med. 3, 97ra79 (2011).

  29. 29.

    Hacein-Bey-Abina, S. et al. Insertional oncogenesis in 4 patients after retrovirus-mediated gene therapy of SCID-X1. J. Clin. Invest. 118, 3132–3142 (2008).

  30. 30.

    Howe, S. J. et al. Insertional mutagenesis combined with acquired somatic mutations causes leukemogenesis following gene therapy of SCID-X1 patients. J. Clin. Invest. 118, 3143–3150 (2008).

  31. 31.

    Hacein-Bey-Abina, S. et al. A modified γ-retrovirus vector for X-linked severe combined immunodeficiency. N. Engl. J. Med. 371, 1407–1417 (2014).

  32. 32.

    Zhou, S. et al. A self-inactivating lentiviral vector for SCID-X1 gene therapy that does not activate LMO2 expression in human T cells. Blood 116, 900–908 (2010).

  33. 33.

    Aker, M. et al. Extended core sequences from the cHS4 insulator are necessary for protecting retroviral vectors from silencing position effects. Hum. Gene Ther. 18, 333–343 (2007).

  34. 34.

    Greene, M. R. et al. Transduction of human CD34+ repopulating cells with a self-inactivating lentiviral vector for SCID-X1 produced at clinical scale by a stable cell line. Hum. Gene Ther. Methods 23, 297–308 (2012).

  35. 35.

    De Ravin, S. S. et al. Lentiviral hematopoietic stem cell gene therapy for X-linked severe combined immunodeficiency. Sci. Transl Med. 8, 335ra57 (2016). The study is the first HSC-based gene therapy trial using a lentiviral vector combined with reduced intensity conditioning for SCID-X1, enabling efficient restoration of the T cell immunity associated with sustained restoration of humoral responses.

  36. 36.

    Thrasher, A. J. et al. Failure of SCID-X1 gene therapy in older patients. Blood 105, 4255–4257 (2005).

  37. 37.

    Mamcarz, E. et al. Interim results from a phase I/II clinical gene therapy study for newly diagnosed infants with X-linked severe combined immunodeficiency using a safety-modified lentiviral vector and targeted reduced exposure to busulfan. Blood 130, 523 (2017).

  38. 38.

    Reimann, C. et al. Human T-lymphoid progenitors generated in a feeder-cell-free DL- 4 culture system promote T cell reconstitution in NOD/SCID/γc(−/−) Mice. Stem Cells 30, 1771–1780 (2012).

  39. 39.

    Simons, L. et al. Generation of adult human T cell progenitors for immunotherapeutic applications. J. Allergy Clin. Immunol. 141, 1491–1494 (2018).

  40. 40.

    Touzot, F. et al. Faster T cell development following gene therapy compared to haplo-identical hematopoietic stem cell transplantation in the treatment of SCID-X1. Blood 125, 3563–3569 (2015).

  41. 41.

    Thrasher, A. J. & Burns, S. O. WASP: a key immunological multitasker. Nat. Rev. Immunol. 10, 182–192 (2010).

  42. 42.

    Candotti, F. Clinical manifestations and pathophysiological mechanisms of the Wiskott-Aldrich syndrome. J. Clin. Immunol. 38, 13–27 (2018).

  43. 43.

    Imai, K. et al. Clinical course of patients with WASP gene mutations. Blood 103, 456–464 (2004).

  44. 44.

    Boztug, K. et al. Stem-cell gene therapy for the Wiskott–Aldrich syndrome. N. Engl. J. Med. 363, 1918–1927 (2010).

  45. 45.

    Braun, C. J. et al. Gene therapy for Wiskott-Aldrich syndrome—long-term efficacy and genotoxicity. Sci. Transl Med. 6, 227ra33 (2014).

  46. 46.

    Chu, J. I. et al. Gene therapy using a self-inactivating lentiviral vector improves clinical and laboratory manifestations of Wiskott-Aldrich syndrome. Blood 126, 260 (2015).

  47. 47.

    Mahlaoui, N. et al. Characteristics and outcome of early-onset, severe forms of Wiskott-Aldrich syndrome. Blood 121, 1510–1516 (2013).

  48. 48.

    Grez, M. et al. Gene therapy of chronic granulomatous disease: the engraftment dilemma. Mol. Ther. 19, 28–35 (2011).

  49. 49.

    Kohn, D. B. et al. Gene therapy for X-linked chronic granulomatous disease [abstract 340]. Mol. Ther. 26(Suppl), 157–158 (2018). This presentation from D. B. Kohn, A. J. Thrasher and colleagues at the ASGCT (American Society of Gene and Cell Therapy) meeting in 2018 reports the first lentiviral-based clinical trials for X-CGD enabling sustained persistence of corrected neutrophils in six patients in this complex, inflammatory environment.

  50. 50.

    Santilli, G. et al. Biochemical correction of X-CGD by a novel chimeric promoter regulating high levels of transgene expression in myeloid cells. Mol. Ther. 19, 122–132 (2011).

  51. 51.

    Brendel, C. et al. Non-clinical efficacy and safety studies on G1XCGD, a lentiviral vector for ex vivo gene therapy of X-linked chronic granulomatous disease. Hum. Gene Ther. Clin. Dev. 29, 69–79 (2018).

  52. 52.

    Weisser, M. et al. Hyperinflammation in patients with chronic granulomatous disease leads to impairment of hematopoietic stem cell functions. J. Allergy Clin. Immunol. 138, 219–228 (2016).

  53. 53.

    de Luca, A. et al. IL-1 receptor blockade restores autophagy and reduces inflammation in chronic granulomatous disease in mice and in humans. Proc. Natl Acad. Sci. USA 111, 3526–3531 (2014).

  54. 54.

    Kovtonyuk, L. V., Fritsch, K., Feng, X., Manz, M. G. & Takizawa, H. Inflamm-aging of hematopoiesis, hematopoietic stem cells, and the bone marrow microenvironment. Front. Immunol. 7, 502 (2016).

  55. 55.

    Takizawa, H., Regoes, R. R., Boddupalli, C. S., Bonhoeffer, S. & Manz, M. G. Dynamic variation in cycling of hematopoietic stem cells in steady state and inflammation. J. Exp. Med. 208, 273–284 (2011).

  56. 56.

    Massberg, S. et al. Immunosurveillance by hematopoietic progenitor cells trafficking through blood, lymph, and peripheral tissues. Cell 131, 994–1008 (2007).

  57. 57.

    Dybedal, I., Bryder, D., Fossum, A., Rusten, L. S. & Jacobsen, S. E. Tumor necrosis factor (TNF)-mediated activation of the p55 TNF receptor negatively regulates maintenance of cycling reconstituting human hematopoietic stem cells. Blood 98, 1782–1791 (2001).

  58. 58.

    Yang, L. et al. IFN-gamma negatively modulates self-renewal of repopulating human hemopoietic stem cells. J. Immunol. 174, 752–757 (2005).

  59. 59.

    Takizawa, H., Boettcher, S. & Manz, M. G. Demand-adapted regulation of early hematopoiesis in infection and inflammation. Blood 119, 2991–3002 (2012).

  60. 60.

    Zhao, J. L. et al. Conversion of danger signals into cytokine signals by hematopoietic stem and progenitor cells for regulation of stress-induced hematopoiesis. Cell Stem Cell 14, 445–459 (2014).

  61. 61.

    Burberry, A. et al. Infection mobilizes hematopoietic stem cells through cooperative NOD-like receptor and toll-like receptor signaling. Cell Host Microbe 15, 779–791 (2014).

  62. 62.

    Nagai, Y. et al. Toll-like receptors on hematopoietic progenitor cells stimulate innate immune system replenishment. Immunity 24, 801–812 (2006).

  63. 63.

    Sioud, M., Fløisand, Y., Forfang, L. & Lund-Johansen, F. Signaling through toll-like Receptor 7/8 induces the differentiation of human bone marrow CD34+progenitor cells along the myeloid lineage. J. Mol. Biol. 364, 945–954 (2006).

  64. 64.

    Sioud, M. & Fløisand, Y. TLR agonists induce the differentiation of human bone marrow CD34+progenitors into CD11c+CD80/86+DC capable of inducing a Th1-type response. Eur. J. Immunol. 37, 2834–2846 (2007).

  65. 65.

    King, K. Y. & Goodell, M. A. Inflammatory modulation of HSCs: viewing the HSC as a foundation for the immune response. Nat. Rev. Immunol. 11, 685–692 (2011).

  66. 66.

    Biffi, A. et al. Correction of metachromatic leukodystrophy in the mouse model by transplantation of genetically modified hematopoietic stem cells. J. Clin. Invest. 113, 1118–1129 (2004).

  67. 67.

    Biffi, A. et al. Gene therapy of metachromatic leukodystrophy reverses neurological damage and deficits in mice. J. Clin. Invest. 116, 3070–3082 (2006).

  68. 68.

    Capotondo, A. et al. Intracerebroventricular delivery of hematopoietic progenitors results in rapid and robust engraftment of microglia-like cells. Sci. Adv. 3, e1701211 (2017).

  69. 69.

    Cartier, N. et al. Hematopoietic stem cell gene therapy with a lentiviral vector in X-linked adrenoleukodystrophy. Science 326, 818–823 (2009).

  70. 70.

    Taher, A. T., Weatherall, D. J. & Cappellini, M. D. Thalassaemia. Lancet 391, 155–167 (2018).

  71. 71.

    Chung, J. H., Bell, A. C. & Felsenfeld, G. Characterization of the chicken beta-globin insulator. Proc. Natl Acad. Sci. USA 94, 575–580 (1997).

  72. 72.

    Cavazzana-Calvo, M. et al. Transfusion independence and HMGA2 activation after gene therapy of human β-thalassaemia. Nature 467, 318–322 (2010). This work shows, for the first time, the amelioration of the clinical phenotype in a patient with β-thalassaemia after lentiviral-based gene therapy.

  73. 73.

    Negre, O. et al. Preclinical evaluation of efficacy and safety of an improved lentiviral vector for the treatment of β-thalassemia and sickle cell disease. Curr. Gene Ther. 15, 64–81 (2015).

  74. 74.

    Miccio, A. et al. In vivo selection of genetically modified erythroblastic progenitors leads to long-term correction of beta-thalassemia. Proc. Natl Acad. Sci. USA 105, 10547–10552 (2008).

  75. 75.

    Marktel, S. et al. Intrabone hematopoietic stem cell gene therapy for adult and pediatric patients affected by transfusion-dependent ß-thalassemia. Nat. Med. https://doi.org/10.1038/s41591-018-0301-6 (2019).

  76. 76.

    Ferrari, G., Cavazzana, M. & Mavilio, F. Gene therapy approaches to hemoglobinopathies. Hematol. Oncol. Clin. North Am. 31, 835–852 (2017).

  77. 77.

    Kato, G. J. et al. Sickle cell disease. Nat. Rev. Dis. Prim. 4, 18010 (2018).

  78. 78.

    Kanter, J. et al. Interim results from a phase 1/2 clinical study of lentiglobin gene therapy for severe sickle cell disease. Blood 128, 1176 (2016).

  79. 79.

    Lagresle-Peyrou, C. et al. Plerixafor enables safe, rapid, efficient mobilization of hematopoietic stem cells in sickle cell disease patients after exchange transfusion. Haematologica 103, 778–786 (2018).

  80. 80.

    Tisdale, J. F. et al. Current results of lentiglobin gene therapy in patients with severe sickle cell disease treated under a refined protocol in the phase 1 Hgb-206 study [abstract 1026]. Presented at the 2018 ASH Annual Meeting in San Diego (2018).

  81. 81.

    Glimm, H., Oh, I. H. & Eaves, C. J. Human hematopoietic stem cells stimulated to proliferate in vitro lose engraftment potential during their S/G(2)/M transit and do not reenter G(0). Blood 96, 4185–4193 (2000).

  82. 82.

    Kallinikou, K. et al. Engraftment defect of cytokine-cultured adult human mobilized CD34 + cells is related to reduced adhesion to bone marrow niche elements. Br. J. Haematol. 158, 778–787 (2012).

  83. 83.

    Larochelle, A. et al. Bone marrow homing and engraftment of human hematopoietic stem and progenitor cells is mediated by a polarized membrane domain. Blood 119, 1848–1855 (2012).

  84. 84.

    Piras, F. et al. Lentiviral vectors escape innate sensing but trigger p53 in human hematopoietic stem and progenitor cells. EMBO Mol. Med. 9, 1198–1211 (2017).

  85. 85.

    Zonari, E. et al. Efficient ex vivo engineering and expansion of highly purified human hematopoietic stem and progenitor cell populations for gene therapy. Stem Cell Rep. 8, 977–990 (2017). This study highlights some significant improvements in HSC culture conditions, which are critical for the development of new and more efficient gene addition and gene-editing therapies.

  86. 86.

    Kiernan, J. et al. Clinical studies of ex vivo expansion to accelerate engraftment after umbilical cord blood transplantation: a systematic review. Transfus. Med. Rev. 31, 173–182 (2017).

  87. 87.

    Kajaste-Rudnitski, A. & Naldini, L. Cellular innate immunity and restriction of viral infection: implications for lentiviral gene therapy in human hematopoietic cells. Hum. Gene Ther. 26, 201–209 (2015).

  88. 88.

    Gao, D. et al. Cyclic GMP-AMP synthase is an innate immune sensor of HIV and other retroviruses. Science 341, 903–906 (2013).

  89. 89.

    Petrillo, C. et al. Cyclosporin A and rapamycin relieve distinct lentiviral restriction blocks in hematopoietic stem and progenitor cells. Mol. Ther. 23, 352–362 (2015).

  90. 90.

    Heffner, G. C. et al. Prostaglandin E2 increases lentiviral vector transduction efficiency of adult human hematopoietic stem and progenitor cells. Mol. Ther. 26, 320–328 (2018).

  91. 91.

    North, T. E. et al. Prostaglandin E2 regulates vertebrate haematopoietic stem cell homeostasis. Nature 447, 1007–1011 (2007).

  92. 92.

    Goessling, W. et al. Prostaglandin E2 enhances human cord blood stem cell xenotransplants and shows long-term safety in preclinical nonhuman primate transplant models. Cell Stem Cell 8, 445–458 (2011).

  93. 93.

    Lewis, G. et al. Staurosporine increases lentiviral vector transduction efficiency of human hematopoietic stem and progenitor cells. Mol. Ther. Methods Clin. Dev. 9, 313–322 (2018).

  94. 94.

    Hauber, I. et al. Improving lentiviral transduction of CD34 + hematopoietic stem and progenitor cells. Hum. Gene Ther. Methods 29, 104–113 (2018).

  95. 95.

    Biasco, L. et al. In vivo tracking of human hematopoiesis reveals patterns of clonal dynamics during early and steady-state reconstitution phases. Cell Stem Cell 19, 107–119 (2016). This is the first report of HSC lineage tracing in a human individual, using integration sites analysis in patients with WAS following gene therapy, providing some new insights on human haematopoiesis.

  96. 96.

    Masiuk, K. E. et al. Improving gene therapy efficiency through the enrichment of human hematopoietic stem cells. Mol. Ther. 25, 2163–2175 (2017).

  97. 97.

    Biasco, L. et al. In vivo tracking of T cells in humans unveils decade-long survival and activity of genetically modified T memory stem cells. Sci. Transl Med. 7, 273ra13 (2015).

  98. 98.

    Scala, S. et al. Dynamics of genetically engineered hematopoietic stem and progenitor cells after autologous transplantation in humans. Nat. Med. 24, 1683–1690 (2018).

  99. 99.

    Lidonnici, M. R. et al. Plerixafor and G-CSF combination mobilizes hematopoietic stem and progenitors cells with a distinct transcriptional profile and a reduced in vivo homing capacity compared to plerixafor alone. Haematologica 102, e120–e124 (2017).

  100. 100.

    Cattoglio, C. et al. High-definition mapping of retroviral integration sites identifies active regulatory elements in human multipotent hematopoietic progenitors. Blood 116, 5507–5517 (2010).

  101. 101.

    Romano, O. et al. Transcriptional, epigenetic and retroviral signatures identify regulatory regions involved in hematopoietic lineage commitment. Sci. Rep. 6, 24724 (2016).

  102. 102.

    Schröder, A. R. W. et al. HIV-1 integration in the human genome favors active genes and local hotspots. Cell 110, 521–529 (2002).

  103. 103.

    Mitchell, R., Chiang, C. Y., Berry, C. & Bushman, F. Global analysis of cellular transcription following infection with an HIV-based vector. Mol. Ther. 8, 674–687 (2003).

  104. 104.

    Wu, X., Li, Y., Crise, B. & Burgess, S. Transcription start regions in the human genome are favored targets for MLV integration. Science 300, 1749–1751 (2003).

  105. 105.

    Narezkina, A. et al. Genome-wide analyses of avian sarcoma virus integration sites. J. Virol. 78, 11656–11663 (2004).

  106. 106.

    Meekings, K. N., Leipzig, J., Bushman, F. D., Taylor, G. P. & Bangham, C. R. M. HTLV-1 integration into transcriptionally active genomic regions is associated with proviral expression and with HAM/TSP. PLOS Pathog. 4, e1000027 (2008).

  107. 107.

    Gillet, N. A. et al. The host genomic environment of the provirus determines the abundance of HTLV-1-infected T cell clones. Blood 117, 3113–3122 (2011).

  108. 108.

    Brady, T. et al. Integration target site selection by a resurrected human endogenous retrovirus. Genes Dev. 23, 633–642 (2009).

  109. 109.

    Mitchell, R. S. et al. Retroviral DNA integration: ASLV, HIV, and MLV show distinct target site preferences. PLOS Biol. 2, E234 (2004).

  110. 110.

    Barr, S. D. et al. HIV integration site selection: targeting in macrophages and the effects of different routes of viral entry. Mol. Ther. 14, 218–225 (2006).

  111. 111.

    Lewinski, M. K. et al. Retroviral DNA integration: viral and cellular determinants of target-site selection. PLOS Pathog. 2, e60 (2006).

  112. 112.

    De Rijck, J. et al. The BET family of proteins targets moloney murine leukemia virus integration near transcription start sites. Cell Rep. 5, 886–894 (2013).

  113. 113.

    Ciuffi, A. et al. A role for LEDGF/p75 in targeting HIV DNA integration. Nat. Med. 11, 1287–1289 (2005).

  114. 114.

    Ciuffi, A., Diamond, T. L., Hwang, Y., Marshall, H. M. & Bushman, F. D. Modulating target site selection during human immunodeficiency virus DNA integration in vitro with an engineered tethering factor. Hum. Gene Ther. 17, 960–967 (2006).

  115. 115.

    Marshall, H. M. et al. Role of PSIP1/LEDGF/p75 in lentiviral infectivity and integration targeting. PLOS ONE 2, e1340 (2007).

  116. 116.

    Gijsbers, R. et al. LEDGF hybrids efficiently retarget lentiviral integration into heterochromatin. Mol. Ther. 18, 552–560 (2010).

  117. 117.

    Shun, M.-C. et al. LEDGF/p75 functions downstream from preintegration complex formation to effect gene-specific HIV-1 integration. Genes Dev. 21, 1767–1778 (2007).

  118. 118.

    Marini, B. et al. Nuclear architecture dictates HIV-1 integration site selection. Nature 521, 227–231 (2015).

  119. 119.

    Lelek, M. et al. Chromatin organization at the nuclear pore favours HIV replication. Nat. Commun. 6, 6483 (2015).

  120. 120.

    Hacein-Bey-Abina, S. et al. Sustained correction of X-linked severe combined immunodeficiency by ex vivo gene therapy. N. Engl. J. Med. 346, 1185–1193 (2002).

  121. 121.

    Six, E. et al. LMO2 associated clonal T cell proliferation 15 years after gamma-retrovirus mediated gene therapy for SCIDX1. Mol. Ther. 25 (Suppl.), 347–348 (2017).

  122. 122.

    Desai, P. et al. Somatic mutations precede acute myeloid leukemia years before diagnosis. Nat. Med. 24, 1015–1023 (2018).

  123. 123.

    Levine, B. L. et al. Gene transfer in humans using a conditionally replicating lentiviral vector. Proc. Natl Acad. Sci. USA 103, 17372–17377 (2006).

  124. 124.

    Wang, G. P. et al. Analysis of lentiviral vector integration in HIV + study subjects receiving autologous infusions of gene modified CD4+T cells. Mol. Ther. 17, 844–850 (2009).

  125. 125.

    Moiani, A. et al. Lentiviral vector integration in the human genome induces alternative splicing and generates aberrant transcripts. J. Clin. Invest. 122, 1653–1666 (2012).

  126. 126.

    Cesana, D. et al. Whole transcriptome characterization of aberrant splicing events induced by lentiviral vector integrations. J. Clin. Invest. 122, 1667–1676 (2012). This paper, together with the copublished paper by Moiani et al., demonstrates that the lentiviral vector can be responsible for the generation of aberrant splicing events. This potential genotoxicity should, therefore, be carefully taken into account by minimizing the number of vector copies per cell and by developing safer vectors by recoding cryptic splicing sites.

  127. 127.

    Cesana, D. et al. Uncovering and dissecting the genotoxicity of self-inactivating lentiviral vectors in vivo. Mol. Ther. 22, 774–785 (2014).

  128. 128.

    Maude, S. L. et al. Chimeric antigen receptor T cells for sustained remissions in leukemia. N. Engl. J. Med. 371, 1507–1517 (2014).

  129. 129.

    Davila, M. L. et al. Efficacy and toxicity management of 19-28z CAR T cell therapy in B cell acute lymphoblastic leukemia. Sci. Transl Med. 6, 224ra25 (2014).

  130. 130.

    Lee, D. W. et al. T cells expressing CD19 chimeric antigen receptors for acute lymphoblastic leukaemia in children and young adults: a phase 1 dose-escalation trial. Lancet 385, 517–528 (2015).

  131. 131.

    Porter, D. L. et al. Chimeric antigen receptor T cells persist and induce sustained remissions in relapsed refractory chronic lymphocytic leukemia. Sci. Transl Med. 7, 303ra139 (2015).

  132. 132.

    Fraietta, J. A. et al. Determinants of response and resistance to CD19 chimeric antigen receptor (CAR) T cell therapy of chronic lymphocytic leukemia. Nat. Med. 24, 563–571 (2018).

  133. 133.

    Wilkins, O., Keeler, A. M. & Flotte, T. R. CAR T-cell therapy: progress and prospects. Hum. Gene Ther. Methods 28, 61–66 (2017).

  134. 134.

    Scholler, J. et al. Decade-long safety and function of retroviral-modified chimeric antigen receptor T cells. Sci. Transl Med. 4, 132ra53 (2012).

  135. 135.

    Fraietta, J. A. et al. Disruption of TET2 promotes the therapeutic efficacy of CD19-targeted T cells. Nature 558, 307–312 (2018).

  136. 136.

    Mohr, F., Döhner, K., Buske, C. & Rawat, V. P. S. TET Genes: new players in DNA demethylation and important determinants for stemness. Exp. Hematol. 39, 272–281 (2011).

  137. 137.

    Grogg, K. L., Miller, R. F. & Dogan, A. HIV infection and lymphoma. J. Clin. Pathol. 60, 1365–1372 (2006).

  138. 138.

    Maldarelli, F. et al. HIV latency. Specific HIV integration sites are linked to clonal expansion and persistence of infected cells. Science 345, 179–183 (2014).

  139. 139.

    Wagner, T. A. et al. HIV latency. Proliferation of cells with HIV integrated into cancer genes contributes to persistent infection. Science 345, 570–573 (2014).

  140. 140.

    Haworth, K. G. et al. HIV infection results in clonal expansions containing integrations within pathogenesis-related biological pathways. JCI Insight 3, 99127 (2018).

  141. 141.

    Bhukhai, K. et al. Ex vivo selection of transduced hematopoietic stem cells for gene therapy of β-hemoglobinopathies. Mol. Ther. 26, 480–495 (2018).

  142. 142.

    Santoni De Sio, F. R. et al. Ectopic FOXP3 expression preserves primitive features of human hematopoietic stem cells while impairing functional T cell differentiation. Sci. Rep. 7, 15820 (2017).

  143. 143.

    Multhaup, M. et al. Cytotoxicity associated with artemis overexpression after lentiviral vector-mediated gene transfer. Hum. Gene Ther. 21, 865–875 (2010).

  144. 144.

    Jiang, Q. et al. Retroviral transduction of IL-7Rα into IL-7Rα−/−bone marrow progenitors: correction of lymphoid deficiency and induction of neutrophilia. Gene Ther. 12, 1761–1768 (2005).

  145. 145.

    Chiriaco, M. et al. Dual-regulated lentiviral vector for gene therapy of X-linked chronic granulomatosis. Mol. Ther. 22, 1472–1483 (2014).

  146. 146.

    Sweeney, C. L. et al. Targeted repair of CYBB in X-CGD iPSCs requires retention of intronic sequences for expression and functional correction. Mol. Ther. 25, 321–330 (2017).

  147. 147.

    Punwani, D. et al. Lentivirus mediated correction of artemis-deficient severe combined immunodeficiency. Hum. Gene Ther. 28, 112–124 (2017).

  148. 148.

    Fernández-Rubio, P., Torres-Rusillo, S. & Molina, I. J. Regulated expression of murine CD40L by a lentiviral vector transcriptionally targeted through its endogenous promoter. J. Gene Med. 17, 219–228 (2015).

  149. 149.

    DeWitt, M. A. et al. Selection-free genome editing of the sickle mutation in human adult hematopoietic stem/progenitor cells. Sci. Transl Med. 8, 360ra134 (2016).

  150. 150.

    Hoban, M. D. et al. CRISPR/Cas9-mediated correction of the sickle mutation in human CD34+cells. Mol. Ther. 24, 1561–1569 (2016).

  151. 151.

    Dever, D. P. et al. CRISPR/Cas9 β-globin gene targeting in human haematopoietic stem cells. Nature 539, 384–389 (2016). In this work, Dever and colleagues show that HSCs genetically modified to integrate a donor template in the β-globin gene can be selected and engrafted in immunodeficient mice.

  152. 152.

    Charlesworth, C. T. et al. Priming human repopulating hematopoietic stem and progenitor cells for Cas9/sgRNA gene targeting. Mol. Ther. Nucleic Acids 12, 89–104 (2018).

  153. 153.

    Schiroli, G. et al. Preclinical modeling highlights the therapeutic potential of hematopoietic stem cell gene editing for correction of SCID-X1. Sci. Transl Med. 9, eaan0820 (2017).

  154. 154.

    De Ravin, S. S. et al. CRISPR-Cas9 gene repair of hematopoietic stem cells from patients with X-linked chronic granulomatous disease. Sci. Transl Med. 9, eaah3480 (2017).

  155. 155.

    Kuo, C. Y. et al. Site-specific gene editing of human hematopoietic stem cells for X-linked hyper-IgM syndrome. Cell Rep. 23, 2606–2616 (2018).

  156. 156.

    Canver, M. C. et al. BCL11A enhancer dissection by Cas9-mediated in situ saturating mutagenesis. Nature 527, 192–197 (2015). In this work, Canver and colleagues show that disruption of a BCL11A enhancer can reduce BCL11A levels and reactivate fetal haemoglobin expression in HSPC-derived erythroblasts.

  157. 157.

    Chang, K. H. et al. Long-term engraftment and fetal globin induction upon BCL11A gene editing in bone-marrow-derived CD34+hematopoietic stem and progenitor cells. Mol. Ther. Methods Clin. Dev. 4, 137–148 (2017).

  158. 158.

    Ye, L. et al. Genome editing using CRISPR-Cas9 to create the HPFH genotype in HSPCs: An approach for treating sickle cell disease and β-thalassemia. Proc. Natl Acad. Sci. USA 113, 10661–10665 (2016).

  159. 159.

    Traxler, E. A. et al. A genome-editing strategy to treat β-hemoglobinopathies that recapitulates a mutation associated with a benign genetic condition. Nat. Med. 22, 987–990 (2016).

  160. 160.

    Antoniani, C. et al. Induction of fetal hemoglobin synthesis by CRISPR/Cas9-mediated editing of the human b-globin locus. Blood 131, 1960–1973 (2018).

  161. 161.

    Kosicki, M., Tomberg, K. & Bradley, A. Repair of double-strand breaks induced by CRISPR–Cas9 leads to large deletions and complex rearrangements. Nat. Biotechnol. 36, 765–771 (2018).

  162. 162.

    Marx, V. Base editing a CRISPR way. Nat. Methods 16, 767–770 (2018).

  163. 163.

    Holtzman, L. & Gersbach, C. A. Editing the epigenome: reshaping the genomic landscape. Annu. Rev. Genomics Hum. Genet. 19, 43–71 (2018).

  164. 164.

    Aiuti, A. et al. Immune reconstitution in ADA-SCID after PBL gene therapy and discontinuation of enzyme replacement. Nat. Med. 8, 423–425 (2002).

  165. 165.

    Candotti, F. et al. Gene therapy for adenosine deaminase-deficient severe combined immune deficiency: clinical comparison of retroviral vectors and treatment plans. Blood 120, 3635–3646 (2012).

  166. 166.

    Shaw, K. L. et al. Clinical efficacy of gene-modified stem cells in adenosine deaminase–deficient immunodeficiency. J. Clin. Invest. 127, 1689–1699 (2017).

  167. 167.

    Stein, S. et al. Genomic instability and myelodysplasia with monosomy 7 consequent to EVI1 activation after gene therapy for chronic granulomatous disease. Nat. Med. 16, 198–204 (2010).

  168. 168.

    Ott, M. G. et al. Correction of X-linked chronic granulomatous disease by gene therapy, augmented by insertional activation of MDS1-EVI1, PRDM16 or SETBP1. Nat. Med. 12, 401–409 (2006).

  169. 169.

    Bianchi, M. et al. Restoration of NET formation by gene therapy in CGD controls aspergillosis. Blood 114, 2619–2622 (2009).

  170. 170.

    Castiello, M. C. et al. B cell reconstitution after lentiviral vector-mediated gene therapy in patients with Wiskott-Aldrich syndrome. J. Allergy Clin. Immunol. 136, 692–702 (2015).

  171. 171.

    Chinen, J. et al. Gene therapy improves immune function in preadolescents with X-linked severe combined immunodeficiency. Blood 110, 67–73 (2007).

  172. 172.

    Scaramuzza, S. et al. Lentiviral hematopoietic stem cells gene therapy for beta-thalassemia: update from the phase I/II TIGET BTHAL trial [abstract 2]. Mol. Ther. 26, 1–2 (2018).

  173. 173.

    Mansilla-Soto, J., Riviere, I., Boulad, F. & Sadelain, M. Cell and gene therapy for the beta-thalassemias: advances and prospects. Hum. Gene Ther. 27, 295–304 (2016).

  174. 174.

    Esrick, E. B. et al. Flipping the switch: initial results of genetic targeting of the fetal to adult globin switch in sickle cell patients [abstract 1023]. Presented at the 2018 ASH Annual Meeting in San Diego (2018).

  175. 175.

    Malik, P. et al. Gene therapy for sickle cell anemia using a modified gamma globin lentivirus vector and reduced intensity conditioning transplant shows promising correction of the disease phenotype [abstract 1021]. Presented at the 2018 ASH Annual Meeting in San Diego (2018).

  176. 176.

    Eichler, F. et al. Hematopoietic stem-cell gene therapy for cerebral adrenoleukodystrophy. N. Engl. J. Med. 377, 1630–1638 (2017).

  177. 177.

    Fraldi, A. et al. Gene therapy for mucopolysaccharidoses: in vivo and ex vivo approaches. Ital. J. Pediatr. 44, 130 (2018).

  178. 178.

    Gentner, B. Towards next-generation gene therapy with ex vivo engineered hematopoietic stem and progenitor cells [abstract INV058]. Hum. Gene Ther. 29, A12 (2018).

  179. 179.

    Adair, J. et al. Lessons learned from two decades of clinical trial experience in gene therapy for Fanconi anemia. Curr. Gene Ther. 16, 338–348 (2017).

Download references

Acknowledgements

The authors thank the researchers whose works were not discussed owing to space limitations. M.C., I.A.-S. and E.S. are grateful to members of the human haematolymphopoiesis laboratory at the Imagine Institute and of the biotherapy department in Necker’s Hospital for their commitment allowing clinical development. A.M. thanks members of her laboratory at the Imagine Institute. F.D.B. thanks members of his laboratory at the University of Pennsylvania School of Medicine. M.C., I.A.-S. and E.S. are supported by grants from the European Research Council (ERC Regenerative Therapy 269037 and Gene for Cure 693762), the EU Seventh Framework Programme (Net4CGD 305011), the EU H2020 research and innovation programme (SCIDNET 666908), the Clinical Research Hospital Programme (PHRC) (Ministry of Health and Social Affairs), Assistance Publique-Hôpitaux de Paris, INSERM, the French National Research Agency under the Investments for the Future programme (ANR-01-A0-IAHU) and Bluebird Bio. A.M. is supported by grants from the Agence nationale de la recherche (ANR-16-CE18-0004 and ANR-10-IAHU-01 ‘Investissements d’avenir’ programme). F.D.B.’s laboratory is supported by grants from the US National Institutes of Health (NIH) (AI 052845–13, AI 082020-05A1, AI 045008–15, U19AI117950-01 and UMIAI126620) and the Penn Center for AIDS Research.

Author information

Affiliations

  1. Biotherapy Department, Necker Children’s Hospital, Assistance Publique-Hôpitaux de Paris, Paris, France

    • Marina Cavazzana
  2. Biotherapy Clinical Investigation Center, Groupe Hospitalier Universitaire Ouest, AP-HP, INSERM, Paris, France

    • Marina Cavazzana
  3. INSERM UMR 1163, Laboratory of Human Lymphohematopoiesis, Paris, France

    • Marina Cavazzana
    • , Isabelle André-Schmutz
    •  & Emmanuelle Six
  4. Paris Descartes–Sorbonne Paris Cité University, Imagine Institute, Paris, France

    • Marina Cavazzana
    • , Annarita Miccio
    • , Isabelle André-Schmutz
    •  & Emmanuelle Six
  5. Department of Microbiology, University of Pennsylvania School of Medicine, Philadelphia, PA, USA

    • Frederic D. Bushman
  6. Laboratory of Chromatin and Gene Regulation During Development,, INSERM UMR1163, Imagine Institute, Paris, France

    • Annarita Miccio

Authors

  1. Search for Marina Cavazzana in:

  2. Search for Frederic D. Bushman in:

  3. Search for Annarita Miccio in:

  4. Search for Isabelle André-Schmutz in:

  5. Search for Emmanuelle Six in:

Competing interests

The authors declare no competing interests.

Corresponding author

Correspondence to Marina Cavazzana.

Glossary

Haematopoietic stem and progenitor cells

(HSPCs). A heterogeneous cell population that can be isolated using the surface marker CD34 and comprises both the most immature haematopoietic stem cells responsible for long-term engraftment and haematopoietic progenitors that have lost self-renewal capacity, are more restricted in term of lineage potential and are responsible for short-term engraftment.

Enzyme replacement therapy

(ERT). A medical treatment aiming to replace a missing protein. In the case of severe combined immunodeficiency caused by adenosine deaminase (ADA) deficiency, pegademase bovine ADA is used.

Patient conditioning

The treatments used to prepare a patient for haematopoietic stem and progenitor cell transplantation. The conditioning regimen may include chemotherapy, monoclonal antibody therapy and radiation. It helps make room in the patient’s bone marrow for new haematopoietic stem cells and to prevent rejection in case of allogeneic transplantation.

Haematopoietic stem cell

(HSC). A cell defined by the capacity to self-renew and the ability to ensure continuous production of all blood lineages for the entire life of an individual.

Myelogram

This bone marrow puncture is a medical test that consists of taking a bone marrow sample from the hip or the sternum. Once the extract has been smeared onto slides, the laboratory analyses the cellular composition of the sample.

Bronchiectasis

A form of chronic lung disease defined as the abnormal irreversible dilatation of the bronchi in which the elastic and muscular tissues are destroyed by acute or chronic inflammation and infection.

BCGitis

Regional lymphadenitis, a severe disseminated disease, following bacillus Calmette–Guérin vaccination.

Ochs score

For Wiskott–Aldrich syndrome, the widely used clinical severity score developed by Ochs (ranging from 1 to 5). A score of 5 is associated with severe disease (autoimmunity, infections, inflammation and/or malignancy).

Vector copy number

(VCN). The average number of integrated therapeutic vector copies per cell in a given population. The VCN can be used to evaluate the transduction and/or correction level in this population.

About this article

Publication history

Published

DOI

https://doi.org/10.1038/s41573-019-0020-9