A. Diseases Caused by Mutations of Genes Expressed by Hematopoietic Cells and Their Treatment by Bone Marrow Transplantation
1. Hemoglobinopathies and Gaucher Disease
There are over 500 known structural variants of hemoglobin. Most persons having a variant hemoglobin are asymptomatic or only mildly affected. Three common variants are associated with significant disease. Two, HbS (Sickle Hemoglobin) and HbC, are point mutations in the codon encoding Glu.sup.6 of .beta.-globin and are found in Africans and their descendants. HbC results from the substitution G.fwdarw.A in the first position of codon 6 and HbS results from the substitution A.fwdarw.T in the second position of codon 6, producing Lys.sup.6 and Val.sup.6 respectively. The third common structural variant of hemoglobin associated with disease, HbE, results from the substitution G.fwdarw.A in the first position of codon 26, encoding Glu.sup.26 resulting in a Lys.sup.26 and is found primarily in the Southeast Asian population and their descendants.
Persons heterozygous for HbS, HbC or HbE do not have significant symptoms. However, HbS homozygotes, HbS/HbC heterozygotes, and heterozygotes for HbC or HbE and an allele of .beta.-thalassemia are severely effected and require frequent medical attention.
Thalassemias are a class of hemoglobinopathies wherein there is an imbalance between the rate of synthesis .alpha.-globin and .beta.-globin. The thalassemias are classified according to the mutated gene, .alpha. or .beta., and whether there is a complete, e.g., .alpha..sup..degree. and .beta..sup.20, or partial reduction, e.g., .alpha..sup.+ and .beta..sup.+, of the affected globin.
There are two functional .alpha.-globin genes in the haploid human genome, which are closely linked. The most common causes of .alpha.-thalassemia are deletions or rearrangements of the linked .alpha.-globin genes or of the .alpha.-globin locus control region.
By contrast the common causes of .beta.-thalassemia are point mutations. The most common cause is a G.fwdarw.A substitution at position of 110 of the .beta.-globin gene, which creates a novel splice acceptor site in the first intron (IVS1).
About 100 different types .beta.-thalassemia have been characterized. Of the characterized mutations only 14 are dominant, i.e., cause a significant clinical condition when heterozygous with a wild-type .beta.-globin allele. The remaining mutations lead to a severe clinical condition, termed thalassemia major, when homozygous or when heterozygous with a second .beta.-thalassemia gene. A detailed review of Hemoglobinopathies is found at Weatherall et al. in THE METABOLIC AND MOLECULAR BASIS OF INHERITED DISEASE 7th Ed., Chapter 113 (McGraw-Hill, New York, 1995)
Gaucher Disease is a lysosomal glycolipid storage disease. There are three recognized types. Type 1 Gaucher Disease is most prevalent in Ashkenazic (Eastern European) Jews and their descendants; Type 2 is panethnic and Type 3 is most prevalent in Northern Sweden. The cause of Gaucher Disease is a defect in the enzyme glucocerebrosidase, also termed acid .beta.-glucosidase. Glucocerebrosidase is expressed in the macrophage/monocyte cell-type, which originate from the bone marrow.
There are three different, common mutations that cause Gaucher Disease: an A.fwdarw.G substitution at nucleotide 1226 of the mRNA, which causes a Asn.fwdarw.Ser substitution at residue 370 of glucocerebrosidase (the N370S mutation); a T.fwdarw.C substitution at nucleotide 1448 of the mRNA, which causes a Leu.fwdarw.Pro substitution at residue 444 of glucocerebrosidase (the L444P mutation); and a duplication of a G.sup.84 nucleotide which results in a frameshift mutation (the 84GG mutation). The N370S and 84GG mutations are associated with the Ashkenazic population while the L444P mutation is associated with the Northern Swedish population and with sporadic mutations.
Uncommon, but not rare, mutations in the coding sequence that cause Gaucher Disease have been identified as follows: at 754, T.fwdarw.A; at 1192, C.fwdarw.T; at 1193, G.fwdarw.A; at 1297, G.fwdarw.T; at 1342, G.fwdarw.C; at 1504, C.fwdarw.T; and at 1604, G.fwdarw.A.
The mutations that cause Gaucher Disease are recessive, i.e., an individual with one wild-type glucocerebrosidase allele is asymptomatic. Individuals which are homozygous for the N370S mutation frequently develop only a mild form of the disease late in life. By contrast, individuals homozygous for the L444P mutation have the more severe Type 2 and Type 3 Gaucher Disease. The 84GG mutation results in a product with no enzymatic activity and results in severe clinical disease in the homozygous state. Of the uncommon mutations, the mutations at nucleotides 754, 1192, 1297 and 1342 are associated with severe forms of the disease. A detailed review of Gaucher Disease can be found in Beutler, E. & Grabowski in THE METABOLIC and MOLECULAR BASIS OF INHERITED DISEASE 7th Ed., Chapter 86 (McGraw-Hill, New York, 1995).
2. The Treatment of Diseases Caused by Mutations by Bone Marrow Transplantation
The treatment of severe cases of .beta.-thalassemia and sickle anemia by allogenic bone marrow transplantation from HLA-matched bone marrow having a normal .beta.-globin gene has been reported to have clinical benefits. Giardini, C., et al., 1995, Annu. Rev. Med. 46:319-30(thalassemia); Lucarelli, G., et al., 1991 Hematol. Oncol. Clin. North Am. 5:549 (thalassemia); Kalinyak, K. A., et al., 1995, Am. J. of Hemat. 48:256-61 (sickle cell); Abboud, M. R., et al., 1994, Am. J. of Ped. Hem/Onc 16:86-89 (sickle cell); Kirkpatrick, D. V., et al., 1991, Semin. Hematol. 28:240 (sickle cell). The clinical results indicate that engraftment, when successful, is curative. Engraftment can be persistent; reports of 3 to 8 year follow-up periods without rejection are found. However, there is a significant failure rate due to the development of graft versus host disease and maintenance doses of immunosuppressive drugs such as cyclosporin are needed. There can also be significant difficulties in obtaining HLA-matched bone marrow.
Allogeneic bone marrow transplantation for the treatment of Type 1 Gaucher Disease has yielded substantially equivalent results to those described above for thalassemia. Ringden, O., et. al., 1995, Transplantation 59:864; Chan, K. W., et al., 1994, Bone Marrow Transplantation 14:327. The use of bone marrow transplantation to treat the more severe Types 2 and 3 Gaucher Disease, which have central nervous system involvement, also has been reported to have favorable outcomes. Tsai, P., et al., 1992, Pediatr. Res. 31:503; Svennerholm, L., et al., 1991, Dev. Neurosci. 13:345-51. In contrast to the globin genes, an effective level of expression of glucocerebrosidase can be obtained by use of viral expression vectors, which suggests that autotransplantation of transduced bone marrow can be an effective therapy for Gaucher Disease. Karlsson, S. & Correll, P. H., 1993, Bone Marrow Transplantation 11(Suppl 1):124-7.