Transfer of drug resistance genes to hematopoietic stem cells offers the potential to protect cancer patients from drug-induced myelosuppression and to increase the number of gene-modified cells by in vivo selection. In this study, a retroviral vector expressing both a P140K variant of human O6-methylguanine-DNA methyltransferase (MGMT) and an EGFP reporter gene was evaluated for stem cell protection in a murine transplant model. Mice transplanted with vector-transduced cells showed significant resistance to the myelosuppressive effects of temozolomide (TMZ), an orally administered DNA-methylating drug, and O6-benzylguanine (BG), a drug that depletes cells of wild-type MGMT activity. Following drug treatment, increases in EGFP(+) peripheral blood cells were seen in all peripheral blood lineages, and secondary transplant experiments proved that selection had occurred at the stem cell level. In a second set of experiments in which transduced cells were diluted with unmarked cells, efficient stem cell selection was noted together with progressive marrow protection with repeated treatment courses. Altogether, these results show that P140K MGMT gene transfer can protect stem cells against the toxic effects of TMZ and BG and that this vector/drug system may be useful for clinical myeloprotection and for in vivo selection of transduced stem cells.

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Muckle-Wells syndrome (MWS) is a dominantly inherited autoinflammatory syndrome. Patients withMWS have a mutation in CIAS1, the gene encoding cryopyrin, a component of the inflammasome that regulates the processing of interleukin-1␤ (IL-1␤). 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-1␤, and she had a heterozygous mutation in the CIAS1 gene, with C-to-T transversion at nucleotide position 778, encoding an arginineto-tryptophan mutation at position 260 (R260W). Mononuclear cells (MNCs) isolated from the patient secreted large amounts of IL-1␤, 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.

In the present study, we attempted to clarify the effects of interleukin-6 (IL-6) on the growth and properties of human mast cells using cultured mast cells selectively generated by stem cell factor (SCF) from CD34+ cord blood cells. The addition of IL-6 to cultures containing mast cells resulted in a substantial reduction of the number of progenies grown by SCF in the liquid culture. This IL-6–mediated inhibition of mast cell growth may be due in part to the suppression at the precursor level, according to the results of a clonal cell culture assay. Moreover, a flow cytometric analysis showed that the cultured mast cells grown in the presence of SCF+IL-6 had decreased c-kit expression. The exposure of cultured mast cells to SCF+IL-6 also caused substantial increases in the cell size, frequency of chymase-positive cells, and intracellular histamine level compared with the values obtained with SCF alone. The flow cytometric analysis showed low but significant levels of expression of IL-6 receptor (IL-6R) and gp130 on the cultured mast cells grown with SCF. The addition of either anti–IL-6R antibody or anti-gp130 antibody abrogated the biological functions of IL-6. Although IL-4 exerted an effect similar to that of IL-6 on the cultured mast cells under stimulation with SCF, the results of comparative experiments suggest that the two cytokines use different regulatory mechanisms. Taken together, the present findings suggest that IL-6 modulates SCF-dependent human mast cell development directly via an IL-6R-gp130 system.