Anakinra improves sensory deafness in a Japanese patient with Muckle-Wells syndrome, possibly by inhibiting the cryopyrin inflammasome.

Muckle-Wells syndrome (MWS) is a dominantly inherited autoinflammatory syndrome. Patients with MWS have a mutation in CIAS1, the gene encoding cryopyrin, a component of the inflammasome that regulates the processing of interleukin-1beta (IL-1beta). In this report we describe an 8-year-old Japanese girl with MWS who had symptoms of periodic fever, urticarial rash, conjunctivitis, arthropathy, and sensory deafness. Laboratory analysis of the patient's serum showed abnormally high concentrations of C-reactive protein, serum amyloid A, and IL-1beta, and she had a heterozygous mutation in the CIAS1 gene, with C-to-T transversion at nucleotide position 778, encoding an arginine-to-tryptophan mutation at position 260 (R260W). Mononuclear cells (MNCs) isolated from the patient secreted large amounts of IL-1beta, without stimulation, and were highly sensitive to muramyldipeptide and lipopolysaccharide. After treatment with anakinra, laboratory results normalized, and clinical symptoms, including sensory deafness, disappeared, while MNCs appeared to remain activated. Thus, our case suggests that anakinra possibly affects the cryopyrin inflammasome and markedly improves the clinical and laboratory manifestations of MWS.

Development of gene therapy for hemoglobin disorders.

The hemoglobin disorders, severe beta-thalassemia and sickle cell anemia, are prevalent monogenetic disorders which cause severe morbidity and mortality worldwide. Gene therapy approaches to these disorders envision stem cell targeted gene transfer, autologous transplantation of gene-corrected stem cells, and functional, phenotypically corrective globin gene expression in developing erythroid cells. Lentiviral vector systems potentially appear to afford adequately efficient gene transfer into stem cells and are capable, with appropriate genetic engineering, of transferring a globin gene with the regulatory elements required to achieve high-level, erythroid-specific expression. Herein are results obtained in use of lentiviral vectors to insert a gamma-globin gene into murine stem cells with phenotypic correction of the thalassemia phenotype. Further, we have developed a drug-selection system for genetically modified stem cells based on a mutant form of methylguanine, methyltransferase, which allows selective amplification of genetically modified stem cells with phenotypic correction even in the absence of myeloablation prior to stem cell transplantation. These advances provide essential preclinical data which build toward the development of effective gene therapy for the severe hemoglobin disorders.

Successful treatment of murine beta-thalassemia using in vivo selection of genetically modified, drug-resistant hematopoietic stem cells.

Successful gene therapy of beta-thalassemia will require replacement of the abnormal erythroid compartment with erythropoiesis derived from genetically corrected, autologous hematopoietic stem cells (HSCs). However, currently attainable gene transfer efficiencies into human HSCs are unlikely to yield sufficient numbers of corrected cells for a clinical benefit. Here, using a murine model of beta-thalassemia, we demonstrate for the first time that selective enrichment in vivo of transplanted, drug-resistant HSCs can be used therapeutically and may therefore be a useful approach to overcome limiting gene transfer. We used an oncoretroviral vector to transfer a methylguanine methyltransferase (MGMT) drug-resistance gene into normal bone marrow cells. These cells were transplanted into beta-thalassemic mice given nonmyeloablative pretransplantation conditioning with temozolomide (TMZ) and O6-benzylguanine (BG). A majority of mice receiving 2 additional courses of TMZ/BG demonstrated in vivo selection of the drug-resistant cells and amelioration of anemia, compared with untreated control animals. These results were extended using a novel gamma-globin/MGMT dual gene lentiviral vector. Following drug treatment, normal mice that received transduced cells had an average 67-fold increase in gamma-globin expressing red cells. These studies demonstrate that MGMT-based in vivo selection may be useful to increase genetically corrected cells to therapeutic levels in patients with beta-thalassemia.