Susanna M. Müller

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.

Neural Crest Origin of Perivascular Mesenchyme in the Adult Thymus

The endodermal epithelial thymus anlage develops in tight association with neural crest (NC)-derived mesenchyme. This epithelial-NC interaction is crucial for thymus development, but it is not known how NC supports thymus development or whether NC cells or their progeny make any significant contribution to the adult thymus. By nude mouse blastocyst complementation and by cell surface phenotype, we could previously separate thymus stroma into Foxn1-dependent epithelial cells and a Foxn1-independent mesenchymal cell population. These mesenchymal cells expressed vascular endothelial growth factor-A, and contributed to thymus vascularization. These data suggested a physical or functional association with thymic blood vessels, but the origin, location in the thymus, and function of these stromal cells remained unknown. Using a transgenic mouse expressing Cre recombinase in premigratory NC (Sox10-Cre), we have now fate-mapped the majority of these adult mesenchymal cells to a NC origin. NC-derived cells represent tightly vessel-associated pericytes that are sandwiched between endothelium and epithelium along the entire thymus vasculature. The ontogenetic, phenotypic, and positional definition of this distinct perivascular mesenchymal compartment provides a cellular basis for the role of NC in thymus development and possibly maintenance, and might be useful to address properties of the endothelial-epithelial barrier in the adult thymus.

Mutations in the gene for the common gamma chain (γc) in X-linked severe combined immunodeficiency

Abstract X-linked severe combined immunodeficiency (XSCID) consitutes a disorder of the immune system caused by mutations in the gene encoding the common gamma chain (γc), a subunit of the IL-2, IL-4, IL-7, IL-9 and IL-15 receptors, which are necessary for lymphocyte development and function. In this study the IL2RG gene of 31 patients with severe combined immunodeficiency (SCID) was examined by nonradioactive single-strand conformation polymorphism and sequence analysis. Among the 11 patients with XSCID, ten different mutations were identified in the IL2RG gene, including eight novel mutations. Ninety percent of the mothers of the XSCID patients are carriers of the mutated allele. One patient showed low numbers of B-cells, a striking deviation from the classical B-cell-positive and T-cell-negative phenotype.

Allogeneic stem cell transplantation for treatment of immunodeficiency.

Primary immunodeficiencies constitute a group of highly complex congenital disorders commonly characterized by an extremely poor prognosis. Allogeneic hematopoietic stem cell transplantation has the potential to establish a permanently functioning immune system and represents a curative approach in many of these disorders. In this review several aspects of stem cell transplantation are presented, with an emphasis on the mechanism of immune reconstitution in severe combined immunodeficiency diseases. In this disorder transplant modalities vary, and also include transplantation without cytoreductive conditioning. Clinical results are summarized based on recent analysis performed in large patient cohorts, which have shown steady improvements and have led to a marked change in the prognosis of patients with primary immunodeficiencies.

Combined CD34 positive plus CD2 negative selection for effective T-cell depletion as GvHD-prophylaxis in HLA-nonidentical blood progenitor cell transplantation

G-CSF mobilized, T-cell-depleted peripheral blood progenitor cells (PBPC) and T-cell-depleted bone marrow (BM) were given to seven children (6 AL, 1 SCID) to prevent severe graft-versus-host-disease (GvHD) as well as graft rejection after transplantation from HLA-nonidentical parental donors. BM was T-cell-depleted by lectin agglutination and E-rosetting. For T-cell-depletion of the PBPC grafts a combination of CD34+ selection with the Ceprate SC immunoadsorption system and a subsequent depletion of CD2+ cells with immunomagnetic Dynabeads was used. The overall recovery was 0.3 (0.1-1.2)% for nucleated cells, 29 (18-45)% for CD3+ cells, respectively. The purity of CD34+ cells was 87 (68-97)% with a 0.3(0.05-0.7)% residual CD3+ T-cell contamination. In spite of the large T-cell number in the PBPC grafts the combination of CD34 positive and subsequent CD2 negative selection achieved a more than 4 log T-cell depletion and prevents severe GvHD even in HLA-nonidentical transplantation. In addition, if a high dose of progenitor cells ensures stable engraftment, this new approach could increase the possibility of wider use of HLA-mismatched family donors for transplantation.

Expansion of γδ T cells in an infant with severe combined immunodeficiency syndrome after disseminated BCG infection and bone marrow transplantation

γδ T cells typically comprise less than 5% of human blood lymphocytes. In contrast to conventional αβ T cells that recognize peptides presented by HLA, cells of the major human blood γδ T-cell subset recognize nonpeptidic phosphoantigens contained in some bacterial extracts.1 However, the physiologic function of γδ T cells remains unclear. We report the case of an infant with severe combined immunodeficiency (SCID) syndrome and BCG septicemia who showed a striking in vivo expansion of γδ T cells after HLA-compatible bone marrow transplantation (BMT). This unique clinical situation may exemplify a protective role for human γδ T cells in mycobacterial diseases. CASE REPORT After BCG vaccination in the neonatal period, hepatosplenomegaly, fever, and multiple skin nodules developed in a 5-month-old male infant. Skin biopsy revealed acid-fast bacteria‐loaded histiocytes in the absence of infiltrating lymphocytes. The BCG strain Mycobacterium bovis was isolated from the skin, urine, and gastric aspirates, and antimycobacterial therapy was initiated. One maternal half brother of the patient and the brother of his mother had died of septicemia in early childhood. Further laboratory studies, including molecular analysis of the common IL-2 receptor γ-chain, confirmed the diagnosis of X-linked SCID (subtype T‐B+).

Improved lectin agglutination method for T-cell depletion of HLA-mismatched bone marrow grafts in children

For T-cell depletion in HLA-nonidentical bone marrow transplantation of children with malignant diseases, we improved the original lectin/rosetting method described in 1981 by adding anti-CD2/3 coated donor red blood cells to the combination to achieve lectin agglutination in one step. Further improvements in handling led to a shortened and simplified method and better quality of the graft. Five bone marrow grafts prepared with this modified protocol contained a median number of 6 (0-28) x 10(4) T-cells per kg, corresponding to 0.02 (0-0.08)% CD3+ cells and 6 (3.7-10.5) x 10(6) CD34+ cells per kg at a median body-weight of 7 (5-38)kg. The overall recoveries after T-cell depletion were: NC 17 (10-44)%, CD34+ cells 61 (22-100)%, and CFU-GM 55 (29-212)%.

Mobilization and Collection of Allogeneic and Autologous Peripheral Blood Progenitor Cells for Transplantation

Bei 10 Patienten mit Non-Hodgkin-Lymphomen wurden nach Chemotherapie und 5µg G-CSF/kg Korpergewicht und Tag insgesamt 28 Blutstammzellapheresen durchgefuhrt. Im Vergleich hierzu erfolgten bei 8 Normalpersonen mit taglich 5 bzw. 10 µg G-CSF/kg am Tag 5 und 6 insgesamt 16 Apheresen. Bei den Patienten und den Spendern mit 10 µg G-CSF kam es zu einer adaquaten Mobilisation von CD34+ Zellen im peripheren Blut (xxx > 200/µl). Die Apheresedauer betrug im Mittel 175 (143-201) min mit einem Separations-volumen von 8,9 (6,9-11,1) Liter Blut. Die Transplantat-Sollzahlen von 5 × 108 mononuklearen Zellen (MNC), 5 × 106 CD34+ Zellen und 1 × 105 CFU-GM/kg konnten bei Patienten nach Chemotherapie und 5 µg G-CSF wie auch bei Spendern mit der hoheren Dosis von 10 µg G-CSF durch

T-Cell Depletion of Allogeneic Peripheral Blood Progenitor Cells for HLA-Nonidentical Transplantation in Children

Bei Patienten mit malignen Systemerkrankungen ohne HLA-kompatiblen Kno-chenmarkspender wurde neben Knochenmark die zusatzliche Transplantation T-Zell-depletierter peripherer Blutstammzellen von nichtidenten Familienspendern durchgefuhrt. Die primar verwendete Lektin/Rosetten-Methode fuhrte nicht immer zu einer ausreichenden Depletion von T-Lymphozyten der allogenen Blutstamm-zellpraparate. Deshalb wurde eine Kombination aus primarer Selektion von CD34+-Zellen und anschlieβender immunmagnetischer T-Zell-Depletion entwickelt. Dieses neue Verfahren reduzierte bei einem guten Ertrag von Stammzellen die T-Lymphozyten unter 1 × 105 pro kg Korpergewicht. Vier Kinder wurden mit einer Kombination aus Knochenmark und peripheren Stammzellen bei 3 bis 4HLA-Dif-ferenzen transplantiert und zeigten eine rasche hamatopoetische Regeneration ohne schwere Komplikationen einer Spender-gegen-Wirt-Reaktion.