Wilhelm Friedrich

Loss of the ClC-7 Chloride Channel Leads to Osteopetrosis in Mice and Man

Chloride channels play important roles in the plasma membrane and in intracellular organelles. Mice deficient for the ubiquitously expressed ClC-7 Cl(-) channel show severe osteopetrosis and retinal degeneration. Although osteoclasts are present in normal numbers, they fail to resorb bone because they cannot acidify the extracellular resorption lacuna. ClC-7 resides in late endosomal and lysosomal compartments. In osteoclasts, it is highly expressed in the ruffled membrane, formed by the fusion of H(+)-ATPase-containing vesicles, that secretes protons into the lacuna. We also identified CLCN7 mutations in a patient with human infantile malignant osteopetrosis. We conclude that ClC-7 provides the chloride conductance required for an efficient proton pumping by the H(+)-ATPase of the osteoclast ruffled membrane.

Long-term survival and transplantation of haemopoietic stem cells for immunodeficiencies: report of the European experience 1968–99

BACKGROUND Transplantation of allogeneic haemopoietic stem cells can cure several primary immunodeficiencies. This European report focuses on the long-term results of such procedures done between 1968 and December, 1999, for primary immunodeficiencies. METHODS The report includes data from 37 centres in 18 countries, which participated in a European registry for stem-cell transplantation in severe combined immuno deficiencies (SCID) and in other immunodeficiency disorders (non-SCID). 1082 transplants in 919 patients were studied (566 in 475 SCID patients, 512 in 444 non-SCID patients; four procedures excluded owing to insufficient data). Minimum follow-up of 6 months was required. FINDINGS In SCID, 3-year survival with sustained engraftment was significantly better after HLA-identical than after mismatched transplantation (77% vs 54%; p=0.002) and survival improved over time. In HLA-mismatched stem-cell transplantation, B(-) SCID had poorer prognosis than B(+) SCID. However, improvement with time occurred in both SCID phenotypes. In non-SCID, 3-year survival after genotypically HLA-matched, phenotypically HLA-matched, HLA-mismatched related, and unrelated-donor transplantation was 71%, 42%, 42%, and 59%, respectively (p=0.0006). Acute graft versus host disease predicted poor prognosis whatever the donor origin except in related HLA-identical transplantation in SCID. INTERPRETATION The improvement in survival over time indicates more effective prevention and treatment of disease-related and procedure-related complications--eg, infections and graft versus host disease. An important factor is better prevention of graft versus host disease in the HLA-non-identical setting by use of more efficient methods of T-cell depletion. For non-SCID, stem-cell transplantation can provide a cure, and grafts from unrelated donors are almost as beneficial as those from genetically HLA-identical relatives.

RAG Mutations in Human B Cell-Negative SCID

Patients with human severe combined immunodeficiency (SCID) can be divided into those with B lymphocytes (B+ SCID) and those without (B− SCID). Although several genetic causes are known for B+ SCID, the etiology of B− SCID has not been defined. Six of 14 B− SCID patients tested were found to carry a mutation of the recombinase activating gene 1 (RAG-1), RAG-2, or both. This mutation resulted in a functional inability to form antigen receptors through genetic recombination and links a defect in one of the site-specific recombination systems to a human disease.

Transplantation of hematopoietic stem cells and long-term survival for primary immunodeficiencies in Europe: Entering a new century, do we do better?

BACKGROUND Hematopoietic stem cell transplantation remains the only treatment for most patients with severe combined immunodeficiencies (SCIDs) or other primary immunodeficiencies (non-SCID PIDs). OBJECTIVE To analyze the long-term outcome of patients with SCID and non-SCID PID from European centers treated between 1968 and 2005. METHODS The product-limit method estimated cumulative survival; the log-rank test compared survival between groups. A Cox proportional-hazard model evaluated the impact of independent predictors on patient survival. RESULTS In patients with SCID, survival with genoidentical donors (n = 25) from 2000 to 2005 was 90%. Survival using a mismatched relative (n = 96) has improved (66%), similar to that using an unrelated donor (n = 46; 69%; P = .005). Transplantation after year 1995, a younger age, B(+) phenotype, genoidentical and phenoidentical donors, absence of respiratory impairment, or viral infection before transplantation were associated with better prognosis on multivariate analysis. For non-SCID PID, in contrast with patients with SCID, we confirm that, in the 2000 to 2005 period, using an unrelated donor (n = 124) gave a 3-year survival rate similar to a genoidentical donor (n = 73), 79% for both. Survival was 76% in phenoidentical transplants (n = 23) and worse in mismatched related donor transplants (n = 47; 46%; P = .016). CONCLUSION This is the largest cohort study of such patients with the longest follow-up. Specific issues arise for different patient groups. Patients with B-SCID have worse survival than other patients with SCID, despite improvements in each group. For non-SCID PID, survival is worse than SCID, although more conditions are now treated. Individual disease categories now need to be analyzed so that disease-specific prognosis may be better understood and the best treatments planned.

European experience of bone-marrow transplantation for severe combined immunodeficiency

The outcome of bone-marrow transplantations (BMT) carried out between 1968 and March 1, 1989, in 183 patients with severe combined immunodeficiency (SCID) was analysed. Recipients of HLA-identical BMTs (70) had a 76% probability of survival (median follow-up 73 months). Of the 32 treated since 1983, 97% have been cured (median follow-up 41 months). This good prognosis was associated with rapid development of T and B cell function. HLA-non-identical, T-cell-depleted, BMT (n = 100) gave significantly lower survival (52%; median follow-up 47 months). Factors associated with poor prognosis were the presence of a lung infection before BMT, absence of a protected environment, and use of female donors for male recipients. Use of a conditioning regimen significantly increased the frequency of sustained engraftment (86% vs 50% for non-conditioned BMT) and resulted in more frequent engraftment of donor B lymphocytes and myeloid cells. Donor B-cell chimerism was strongly associated with the development of normal B-cell function.

BONE-MARROW TRANSPLANTATION FOR IMMUNODEFICIENCIES AND OSTEOPETROSIS: EUROPEAN SURVEY, 1968-1985

In this retrospective analysis of allogeneic bone-marrow transplantation (BMT) carried out between 1969 and 1985 at fourteen European centres in 162 patients with sixteen different types of inherited immunodeficiencies and osteopetrosis, the overall survival with functional grafts was 51.7% (85 patients), with a minimum follow-up of 5 months. In patients with severe combined immunodeficiency HLA-matched (n = 41) and T-cell-depleted HLA-mismatched BMT (n = 46) resulted in 68% and 57% disease-free survival, respectively; after HLA-mismatched transplants, older age (greater than 6 months) and adenosine-deaminase deficiency resulted in poorer survival. Eight other lethal immunodeficiencies, including profound T-cell deficiencies, Wiskott-Aldrich syndrome, Kostmann syndrome, LFA-1/CR 3/p150,95 deficiency, and Chediak-Higashi syndrome as well as malignant osteopetrosis, have been successfully treated by BMT. In this group, survival with functional graft was 47% with HLA-matched and 29% with T-cell-depleted HLA-mismatched BMT. Engraftment failure was the major complication in this group. Poorer prognosis was associated with older patients, profound T-cell deficiencies, and the degree of HLA incompatibility.

Reticular dysgenesis (aleukocytosis) is caused by mutations in the gene encoding mitochondrial adenylate kinase 2

Human severe combined immunodeficiencies (SCID) are phenotypically and genotypically heterogeneous diseases. Reticular dysgenesis is the most severe form of inborn SCID. It is characterized by absence of granulocytes and almost complete deficiency of lymphocytes in peripheral blood, hypoplasia of the thymus and secondary lymphoid organs, and lack of innate and adaptive humoral and cellular immune functions, leading to fatal septicemia within days after birth. In bone marrow of individuals with reticular dysgenesis, myeloid differentiation is blocked at the promyelocytic stage, whereas erythro- and megakaryocytic maturation is generally normal. These features exclude a defect in hematopoietic stem cells but point to a unique aberration of the myelo-lymphoid lineages. The dramatic clinical course of reticular dysgenesis and its unique hematological phenotype have spurred interest in the unknown genetic basis of this syndrome. Here we show that the gene encoding the mitochondrial energy metabolism enzyme adenylate kinase 2 (AK2) is mutated in individuals with reticular dysgenesis. Knockdown of zebrafish ak2 also leads to aberrant leukocyte development, stressing the evolutionarily conserved role of AK2. Our results provide in vivo evidence for AK2 selectivity in leukocyte differentiation. These observations suggest that reticular dysgenesis is the first example of a human immunodeficiency syndrome that is causally linked to energy metabolism and that can therefore be classified as a mitochondriopathy.

Clinical and molecular profile of a new series of patients with immune dysregulation, polyendocrinopathy, enteropathy, X-linked syndrome: Inconsistent correlation between forkhead box protein 3 expression and disease severity

BACKGROUND Immune dysregulation, polyendocrinopathy, enteropathy, X-linked (IPEX) syndrome is an autoimmune genetic disorder caused by mutation of the forkhead box protein 3 gene (FOXP3), a key regulator of immune tolerance. OBJECTIVE We sought to provide clinical and molecular indicators that facilitate the understanding and diagnosis of IPEX syndrome. METHODS In 14 unrelated affected male subjects who were given diagnoses of IPEX syndrome based on FOXP3 gene sequencing, we determined whether particular FOXP3 mutations affected FOXP3 protein expression and correlated the molecular and clinical data. RESULTS Molecular analysis of FOXP3 in the 14 subjects revealed 13 missense and splice-site mutations, including 7 novel mutations. Enteropathy, generally associated with endocrinopathy and eczema, was reported in all patients, particularly in those carrying mutations within FOXP3 functional domains or mutations that altered protein expression. However, similar genotypes did not always result in similar phenotypes in terms of disease presentation and severity. In addition, FOXP3 protein expression did not correlate with disease severity. CONCLUSION Severe autoimmune enteropathy, which is often associated with increased IgE levels and eosinophilia, is the most prominent early manifestation of IPEX syndrome. Nevertheless, the disease course is variable and somewhat unpredictable. Therefore genetic analysis of FOXP3 should always be performed to ensure an accurate diagnosis, and FOXP3 protein expression analysis should not be the only diagnostic tool for IPEX syndrome.

An Immunodeficiency Disease with RAG Mutations and Granulomas

We describe three unrelated girls who had an immunodeficiency disease with granulomas in the skin, mucous membranes, and internal organs. All three girls had severe complications after viral infections, including B-cell lymphoma associated with Epstein-Barr virus (EBV). Other findings were hypogammaglobulinemia, a diminished number of T and B cells, and sparse thymic tissue on ultrasonography. Molecular analysis revealed that the patients were compound heterozygotes for mutations in recombination activating gene 1 or 2 (RAG1 or RAG2). In each case, both parents were heterozygous carriers of a RAG mutation. The mutations were associated with reduced function of RAG in vitro (3 to 30% of normal activity). The parents and one sibling in the three families were healthy.

Long-term outcome and lineage-specific chimerism in 194 patients with Wiskott-Aldrich syndrome treated by hematopoietic cell transplantation in the period 1980-2009: an international collaborative study.

In this retrospective collaborative study, we have analyzed long-term outcome and donor cell engraftment in 194 patients with Wiskott-Aldrich syndrome (WAS) who have been treated by hematopoietic cell transplantation (HCT) in the period 1980- 2009. Overall survival was 84.0% and was even higher (89.1% 5-year survival) for those who received HCT since the year 2000, reflecting recent improvement of outcome after transplantation from mismatched family donors and for patients who received HCT from an unrelated donor at older than 5 years. Patients who went to transplantation in better clinical conditions had a lower rate of post-HCT complications. Retrospective analysis of lineage-specific donor cell engraftment showed that stable full donor chimerism was attained by 72.3% of the patients who survived for at least 1 year after HCT. Mixed chimerism was associated with an increased risk of incomplete reconstitution of lymphocyte count and post-HCT autoimmunity, and myeloid donor cell chimerism < 50% was associated with persistent thrombocytopenia. These observations indicate continuous improvement of outcome after HCT for WAS and may have important implications for the development of novel protocols aiming to obtain full correction of the disease and reduce post-HCT complications.