Activated STING in a vascular and pulmonary syndrome.

BACKGROUND: The study of autoinflammatory diseases has uncovered mechanisms underlying cytokine dysregulation and inflammation. METHODS: We analyzed the DNA of an index patient with early-onset systemic inflammation, cutaneous vasculopathy, and pulmonary inflammation. We sequenced a candidate gene, TMEM173, encoding the stimulator of interferon genes (STING), in this patient and in five unrelated children with similar clinical phenotypes. Four children were evaluated clinically and immunologically. With the STING ligand cyclic guanosine monophosphate-adenosine monophosphate (cGAMP), we stimulated peripheral-blood mononuclear cells and fibroblasts from patients and controls, as well as commercially obtained endothelial cells, and then assayed transcription of IFNB1, the gene encoding interferon-ß, in the stimulated cells. We analyzed IFNB1 reporter levels in HEK293T cells cotransfected with mutant or nonmutant STING constructs. Mutant STING leads to increased phosphorylation of signal transducer and activator of transcription 1 (STAT1), so we tested the effect of Janus kinase (JAK) inhibitors on STAT1 phosphorylation in lymphocytes from the affected children and controls. RESULTS: We identified three mutations in exon 5 of TMEM173 in the six patients. Elevated transcription of IFNB1 and other gene targets of STING in peripheral-blood mononuclear cells from the patients indicated constitutive activation of the pathway that cannot be further up-regulated with stimulation. On stimulation with cGAMP, fibroblasts from the patients showed increased transcription of IFNB1 but not of the genes encoding interleukin-1 (IL1), interleukin-6 (IL6), or tumor necrosis factor (TNF). HEK293T cells transfected with mutant constructs show elevated IFNB1 reporter levels. STING is expressed in endothelial cells, and exposure of these cells to cGAMP resulted in endothelial activation and apoptosis. Constitutive up-regulation of phosphorylated STAT1 in patients' lymphocytes was reduced by JAK inhibitors. CONCLUSIONS: STING-associated vasculopathy with onset in infancy (SAVI) is an autoinflammatory disease caused by gain-of-function mutations in TMEM173. (Funded by the Intramural Research Program of the National Institute of Arthritis and Musculoskeletal and Skin Diseases; ClinicalTrials.gov number, NCT00059748.).

Suramin inhibits the growth of human rhabdomyosarcoma by interrupting the insulin-like growth factor II autocrine growth loop.

Suramin is a polysulfonated naphthyl-urea with antineoplastic activity that binds various peptide growth factors. Since we previously demonstrated that insulin-like growth factor II (IGF-II) is an autocrine growth factor in human rhabdomyosarcoma (RMS), we studied the effect of suramin on the growth of human RMS cells. Suramin caused a dose-dependent decrease of RMS cell number grown either in 10% fetal bovine serum or in serum-free medium (half-maximal effective dose in mitogenic assays, 1.6 x 10(-4) and 9 x 10(-5) M, respectively). IGF-II and IGF-I added to RMS cells in the presence of suramin reversed the suramin-induced inhibition of cell growth. Since IGF-II exerts its mitogenic effects on RMS cells by binding to the type I receptor, we performed radioreceptor assays using 125I-IGF-I and found that suramin displaced 125I-IGF-I from the type I IGF receptor. There was an excellent correlation between the doses of suramin effective in inhibiting the growth of RMS cells and those that displaced the binding of IGF-I. Our data indicate that suramin exerts its effect on RMS cell growth by interfering with the binding of IGF-II to the type I IGF receptor, thereby interrupting the IGF-II autocrine growth in these cells. Disrupting autonomous growth of RMS may be a promising novel therapeutic approach.

Elimination of clonogenic malignant human T cells using monoclonal antibodies in combination with 2'-deoxycoformycin.

2'Deoxycoformycin (dCF) specifically inhibits adenosine deaminase (ADA) and causes selective cytotoxicity of normal and malignant T cells. In clinical trials, dCF caused rapid lysis of malignant T lymphoblasts. Although dCF has been associated with dose-limiting nonhematopoietic toxicities, myelosuppression has not been observed. Since dCF is relatively nontoxic to hematopoietic stem cells, we tested dCF for utility in the ex vivo purging of malignant T lymphoblasts from remission leukemic bone marrow for autologous bone marrow transplantation. We found that T lymphoblast cell lines were sensitive to dCF (plus deoxyadenosine [dAdo]) under conditions that did not ablate human hematopoietic colony-forming cells. Moreover, combined pharmacologic (dCF plus dAdo) and immunologic (anti-T cell monoclonal antibodies [McAb] plus complement) purging resulted in additive reduction in clonogenic T lymphoblasts. These results provide the basis for a clinical trial of bone marrow transplantation using combined pharmacologic/immunologic purging of T lymphoblasts from patients' harvested autologous marrow.

Specific expression of insulin-like growth factor-II in rhabdomyosarcoma tumor cells.

Insulin-like growth factor II (IGF-II) acts as autocrine growth and motility factor in human rhabdomyosarcoma cell lines, and Northern blot analysis of tumor biopsy specimens from both alveolar and embryonal rhabdomyosarcoma demonstrates high levels of IGF-II mRNA expression. To determine the frequency and site of expression of IGF-II in these tumors, the authors performed in situ hybridization. All tumor specimens examined expressed the gene for IGF-II, and this expression was localized to the tumor cells themselves and not to the surrounding stroma. These data suggest that the IGF-II autocrine loop may be operating not only in vitro but also in vivo.

Cranial irradiation and platelet monoamine oxidase in ALL.

Prophylactic cranial irradiation in acute lymphoblastic leukemia (ALL) has been held responsible for long-term neuropsychological sequelae. This study evaluates the activity of monoamine oxidase (MAO) in children with ALL in first complete remission both before and after cranial irradiation, given to prevent central nervous system involvement. There was a significant decrease (p less than 0.025) in platelet MAO activity shortly after cranial irradiation. MAO activity values were in the normal range in patients investigated 3 months or more after the radiation treatment was completed. The pathogenesis and clinical relevance of this decrease are discussed.

Insulin-like growth factor II overexpression in myoblasts induces phenotypic changes typical of the malignant phenotype.

The objective of this study was to examine the role of insulin-like growth factor II (IGF-II) in the pathogenesis of human rhabdomyosarcomas (RMS). We have demonstrated previously that RMS express high levels of IGF-II mRNA, secrete IGF-II peptide, and express both IGF-I and IGF-II receptors. Moreover, we showed that IGF-II functions as an autocrine growth and motility factor in RMS. Since IGF-II is expressed at high levels in fetal muscle cells and RMS are tumors thought to derive from skeletal myoblasts arrested along the normal myogenic pathway, autocrine production of IGF-II by RMS may be an etiological event in the development of this tumor. We have developed a model system which enabled us to study the effects of endogenous IGF-II overprotection in muscle myoblasts. Human cDNA for pre-prohormone IGF-II was transfected into mouse myoblasts in order to achieve high, constant expression of this growth factor, which is normally down-regulated at the end of the differentiation process. Expression of high IGF-II levels resulted in: (a) an increased proliferative rate; (b) impairment of the ability to differentiate into myoblasts; and (c) acquisition of the capability of anchorage-independent growth. No changes in the expression of IGF-I receptors were noted. We conclude that IGF-II overexpression in muscle myoblasts induces morphological and biological changes typical of the malignant phenotype and represents a fundamental event in the pathogenesis of RMS and possibly of other embryonal tumors.

The insulin-like growth factor II (IGF-II)/mannose 6-phosphate receptor mediates IGF-II-induced motility in human rhabdomyosarcoma cells.

Insulin-like growth factor-II (IGF-II) is an autocrine growth and motility factor for human rhabdomyosarcoma. It interacts with three different receptors: the IGF-I, the IGF-II, and the insulin receptor. A specific function of the IGF-II receptor in mediating IGF-II responses has not been defined. In this report we investigate the mechanism of IGF-II-mediated motility in rhabdomyosarcoma cells. We demonstrate that IGF-II and [Leu27]IGF-II, an analog selective for the IGF-II receptor, stimulate motility at concentrations in which they interact only with their own receptor. An antibody that blocks the IGF-I receptor does not inhibit either peptide activity, while an antibody specific for the IGF-II receptor suppresses the IGF-II-induced motility. This antibody does not interfere with rhabdomyosarcoma cell proliferation. We conclude that in rhabdomyosarcoma cells IGF-II stimulates two different responses mediated by distinct receptors: 1) a mitogenic response through the type I receptor and 2) a motility response through the type II receptor.