Haemoglobin Disorders: Gene Therapy

Abstract

The ability to transfer a globin gene into stem cells and achieve regulated expression during erythropoiesis following transplantation of genetically modified autologous cells could provide curative therapy for haemoglobin disorders such as sickle cell anaemia or β‐thalassaemia. Success has been achieved in several severe immunodeficiencies in which there is a selective advantage for gene corrected cells. The development of effective globin lentiviral vectors has resulted in resolution of the manifestations of sickle cell anaemia and β‐thalassaemia in mouse models of these diseases. However, a higher stem cell transduction efficiency must be achieved in humans than has been achieved in large animal models to date before success in treatment of haemoglobinopathies could be expected. Various strategies are being evaluated in animal models to increase the proportion of genetically modified, haematopoietic cells following infusion of transduced stem cells.

Keywords: gene therapy; sickle cell disease; β‐thalassaemia; lentiviral vector

Figure 1.

Red blood cell abnormalities and pathophysiology of β‐thalassaemia and sickle cell disease. (a) and (b) Severe β‐thalassaemia. Normal human red cells contain 30 pg of haemoglobin, which in the adult is composed of two α‐ and two β‐globin chains (α2β2). The two globin chains are synthesized in nearly equal amounts in maturing erythroblasts. Before birth, γ‐globin rather than β‐globin is present in the haemoglobin tetramer (α2γ2). During the perinatal period, there is a switch from γ‐ to β‐globin synthesis in developing red blood cells resulting in the replacement of fetal haemoglobin (α2γ2) with the adult form (α2β2). (a) A blood smear taken from a patient with severe homozygous β‐thalassaemia demonstrates red blood cells that lack adequate haemoglobin. The smear has been stained to depict the insoluble inclusions of α chains which appear as dark areas, often abutting the red cell membrane. (b) Biosynthesis of globin chains by erythroid cells from a patient with severe homozygous β‐thalassaemia is characteristically abnormal with relative excess synthesis of α‐globin and deficient synthesis of non‐α (γ‐ and β)‐globin (CPM=counts per minute). Effective gene therapy could be achieved by increasing the synthesis of β‐globin, the normal adult chain or γ‐globin, which is typically expressed during fetal life. (c) and (d) Sickle cell anaemia. The characteristic sickle shape of red blood cells (c) reflects polymerization of haemoglobin which occurs as a consequence of a single amino acid substitution, valine for glutamic acid, at position 6 of the β‐globin polypeptide chain and is located on the surface of the hemoglobin molecule. Expression of the naturally, antisickling fetal (γ) globin in red cells from patients with sickle cell anaemia reduces sickling propensity as a consequence of the formation of mixed tetramers (α2 βS γ) in red blood cells (d). The mixed tetramers are unable to participate in polymerization, thereby effectively reducing the concentration of sickle haemoglobin (HbS; α2βs2). Effective gene therapy could likely be achieved by introducing a functional γ‐globin gene that expressed at least 20% of the level of βS globin genes. From Persons DA and Nienhuis AW (2000) Gene therapy for the hemoglobin disorders: past, present and future. Commentary. Proceedings of the National Academy of Sciences of the USA97: 5022–5024. Copyright (2000) National Academy of Sciences, USA.

Figure 2.

Currently envisioned protocols for gene transfer into stem cells of patients with sickle cell disease or thalassaemia are likely to have an outcome in which only a small proportion of the total blood forming cells are genetically modified (unshaded forms are unmodified cells, shaded forms are genetically modified cells). Use of a drug‐resistance gene in the gene therapy vector potentially offers a method to enrich the low numbers of genetically corrected stem cells and progeny (top panel, before drug) that are present in the patient following cell infusion. Subsequent drug treatment would kill the majority of unmodified cells (middle panel, drug treatment), allowing the genetically modified population to become predominant during subsequent haematopoietic recovery (bottom panel, haematopoietic recovery). The feasibility of this approach is well documented in murine models.

close

References

Aiuti A, Slavin S, Aker M et al. (2002) Correction of ADA‐SCID by stem cell gene therapy combined with nonmyeloablative conditioning. Science 296: 2410–2413.

Allay JA, Persons DA, Galipeau J et al. (1998) In vivo selection of retrovirally transduced hematopoietic stem cells. Nature Medicine 4: 1136–1143.

An DS, Kung SK, Bonifacino A et al. (2001) Lentivirus vector‐mediated hematopoietic stem cell gene transfer of common gamma‐chain cytokine receptor in rhesus macaques. Journal of Virology 75: 3547–3555.

Bank A (2008) On the road to gene therapy for beta‐thalassemia and sickle cell anemia. Pediatric Hematology and Oncology 25: 1–4.

Blouin MJ, Beauchemin H, Wright A et al. (2000) Genetic correction of sickle cell disease: insights using transgenic mouse models. Nature Medicine 6: 177–182.

Bunn HF (2001) Human hemoglobins: sickle hemoglobin and other mutants. In: Stamatoyannopoulos G, Majerus P, Perlmutter R and Varmus H (eds) Molecular Basis of Blood Diseases, pp. 227–274. Philadelphia, PA: WB Saunders.

Cavazzana‐Calvo M, Hacein‐Bey S, de Saint Basile G et al. (2000) Gene therapy of human severe combined immunodeficiency (SCID)‐X1 disease. Science 288: 669–672.

Dick JE, Magli MC, Huszar D et al. (1985) Introduction of a selectable gene into primitive stem cells capable of long‐term reconstitution of the hemopoietic system of W/Wv mice. Cell 42: 71–79.

Dzierzak EA, Papayannopoulou T and Mulligan RC (1988) Lineage‐specific expression of a human β‐globin gene in murine bone marrow transplant recipients reconstituted with retrovirus‐transduced stem cells. Nature 331: 35–41.

Eglitis MA, Kantoff P, Gilboa E et al. (1985) Gene expression in mice after high efficiency retroviral‐mediated gene transfer. Science 230: 1395–1398.

Fischer A, Abina SH, Thrasher A et al. (2004) LMO2 and gene therapy for severe combined immunodeficiency. New England Journal of Medicine 350(24): 2526–2527.

Gaspar HB, Parsley KL, Howe S et al. (2004) Gene therapy of X‐linked severe combined immunodeficiency by use of a pseudotyped gammaretroviral vector. Lancet 364: 2181–2187.

Grosveld F, Van Assendelft GB, Greaves DR et al. (1987) Position‐independent, high‐level expression of the human β‐globin gene in transgenic mice. Cell 51: 975–985.

Hanawa H, Hargrove PW, Kepes S et al. (2004) Extended beta‐globin locus control region elements promote consistent therapeutic expression of a gamma‐globin lentiviral vector in murine beta‐thalassemia. Blood 104: 2281–2290.

Hargrove PW, Kepes S, Hanawa H et al. (2008) Globin lentiviral vector insertions can perturb the expression of endogenous genes in beta‐thalassemic hematopoietic cells. Molecular Therapy 16: 525–533.

Horn PA, Morris JC, Bukovsky AA et al. (2002) Lentivirus‐mediated gene transfer into hematopoietic repopulating cells in baboons. Gene Therapy 9(21): 1464–1471.

Karlsson S, Bodine DM, Perry L et al. (1988) Expression of the human β‐globin gene following retroviral‐mediated gene transfer into multipotential hematopoietic progenitors of mice. Proceedings of the National Academy of Sciences of the USA 85: 6062–6066.

Lebensberger J and Persons DA (2008) Progress toward safe and effective gene therapy for β‐thalassemia and sickle cell disease. Current Opinion in Drug Discovery and Development 11: 225–322.

Lucas ML, Seidel NE, Porada CD et al. (2005) Improved transduction of human sheep repopulating cells by retrovirus vectors pseudotyped with feline leukemia virus type C or RD114 envelopes. Blood 106: 51–58.

Mann R, Mulligan RC and Baltimore D (1983) Construction of a retrovirus packaging mutant and its use to produce helper‐free defective retrovirus. Cell 33: 153–159.

Marshall HM, Ronen K, Berry C et al. (2007) Role of PSIP1/LEDGF/p75 in lentiviral infectivity and integration targeting. PLoS ONE 2: e1340.

May C, Rivella S, Callegari J et al. (2000) Therapeutic haemoglobin synthesis in beta‐thalassemic mice expressing lentivirus‐encoded human beta‐globin. Nature 406: 82–86.

Montini E, Cesana D, Schmidt M et al. (2006) Hematopoietic stem cell gene transfer in a tumor‐prone mouse model uncovers low genotoxicity of lentiviral vector integration. Nature Biotechnology 24: 687–696.

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

Neff T, Horn PA, Peterson LJ et al. (2003) Methylguanine methyltransferase‐mediated in vivo selection and chemoprotection of allogeneic stem cells in a large‐animal model. Journal of Clinical Investigation 112: 1581–1588.

Nienhuis AW, Dunbar CE and Sorrentino BP (2006) Genotoxicity of retroviral integration in hematopoietic cells. Molecular Therapy 13: 1031–1049.

Ott MG, Schmidt M, Schwarzwaelder K et al. (2006) Correction of X‐linked chronic granulomatous disease by gene therapy, augmented by insertional activation of MDS1‐EVI1, PRDM16 or SETBP1. Nature Medicine 12: 401–409.

Owens CM, Yang PC, Gottinger H and Sodroski J (2003) Human and simian immunodeficiency virus capsid proteins are major viral determinants of early, postentry replication blocks in simian cells. Journal of Virology 77: 726–731.

Pawliuk R, Westermann KA, Fabry ME et al. (2001) Correction of sickle cell disease in transgenic mouse models by gene therapy. Science 294: 2368–2371.

Persons DA, Allay ER, Sabatino DE et al. (2001) Functional requirements for phenotypic correction of murine β‐thalassemia implications for human gene therapy. Blood 97: 3275–3282.

Persons DA, Allay ER, Sawai N et al. (2003a) Successful treatment of murine beta‐thalassemia using in vivo selection of genetically modified, drug‐resistant hematopoietic stem cells. Blood 101: 506–513.

Persons DA, Hargrove PW, Allay ER et al. (2003b) The degree of phenotypic correction of murine beta‐thalassemia intermedia following lentiviral‐mediated transfer of a human gamma‐globin gene is influenced by chromosomal position effects and vector copy number. Blood 101: 2175–2183.

Puthenveetil G, Scholes J, Carbonell D et al. (2004) Successful correction of the human beta‐thalassemia major phenotype using a lentiviral vector. Blood 104: 3445–3453.

Rivella S, May C, Chadburn A et al. (2003) A novel murine model of Cooley anemia and its rescue by lentiviral‐mediated human beta‐globin gene transfer. Blood 101: 2932–2939.

Sadelain M, Wang CH, Antioniou M et al. (1995) Generation of a high‐titer retroviral vector capable of expressing high levels of the human β‐globin gene. Proceedings of the National Academy of Sciences of the USA 92: 6728–6732.

Sawai N, Zhou S, Vanin EF et al. (2001) Protection and in vivo selection of hematopoietic stem cells using temozolomide, O6‐benzylguanine, and an alkyltransferase‐expressing retroviral vector. Molecular Therapy 3: 78–87.

Stremlau M, Owens CM, Perron MJ et al. (2004) The cytoplasmic body component TRIM5alpha restricts HIV‐1 infection in Old World monkeys. Nature 427: 848–853.

Weatherall DJ (2001) The thalassemias. In: Stamatoyannopoulos G, Majerus P, Perlmutter R and Varmus H (eds) Molecular Basis of Blood Diseases, pp. 183–226. Philadelphia, PA: WB Saunders.

Zhang CC, Kaba M, Ge G et al. (2006) Angiopoietin‐like proteins stimulate ex vivo expansion of hematopoietic stem cells. Nature Medicine 12: 240–245.

Further Reading

Arumugam PI, Scholes J, Perelman N et al. (2007) Improved human beta‐globin expression from self‐inactivating lentiviral vectors carrying the chicken hypersensitive site‐4 (cHS4) insulator element. Molecular Therapy 15: 1863–1871.

Gaszner M and Felsenfeld G (2006) Insulators: exploiting transcriptional and epigenetic mechanisms. Nature Reviews. Genetics 7: 703–713.

Goff SP (2004) Retrovirus restriction factors. Molecular Cell 16: 849–859.

Hacein‐Bey‐Abina S, Von Kalle C, Schmidt M et al. (2003) LMO2‐associated clonal T cell proliferation in two patients after gene therapy for SCID‐XI. Science 301: 2526–2527.

McMorrow T, van den Wijngaard A, Wollenschlaeger A et al. (2000) Activation of the beta globin locus by transcription factors and chromatin modifiers. EMBO Journal 19: 4986–4996.

Nienhuis AW (2008) Development of gene therapy for blood disorders. Blood 111: 4431–4444.

Persons DA and Tisdale JF (2004) Gene therapy for hemoglobin disorders. Seminars in Haematology 41: 279–286.

Sorrentino BP and Nienhuis AW (2001) Gene therapy for hematopoietic diseases. In: Stamatoyannopoulos G, Majerus P, Perlmutter R and Varmus H (eds) Molecular Basis of Blood Diseases, pp. 969–1004. Philadelphia, PA: WB Saunders.

Walters MC (2005) Stem cell therapy for sickle cell disease: transplantation and gene therapy. Hematology/American Society of Hematology. Education Program 66–73.

Contact Editor close
Submit a note to the editor about this article by filling in the form below.

* Required Field

How to Cite close
Persons, Derek A, and Nienhuis, Arthur W(Sep 2009) Haemoglobin Disorders: Gene Therapy. In: eLS. John Wiley & Sons Ltd, Chichester. http://www.els.net [doi: 10.1002/9780470015902.a0006041.pub2]