R. Michael Blaese

T Lymphocyte-Directed Gene Therapy for ADA− SCID: Initial Trial Results After 4 Years

In 1990, a clinical trial was started using retroviral-mediated transfer of the adenosine deaminase (ADA) gene into the T cells of two children with severe combined immunodeficiency (ADA− SCID). The number of blood T cells normalized as did many cellular and humoral immune responses. Gene treatment ended after 2 years, but integrated vector and ADA gene expression in T cells persisted. Although many components remain to be perfected, it is concluded here that gene therapy can be a safe and effective addition to treatment for some patients with this severe immunodeficiency disease.

Gene transfer into humans--immunotherapy of patients with advanced melanoma, using tumor-infiltrating lymphocytes modified by retroviral gene transduction.

BACKGROUND AND METHODS Treatment with tumor-infiltrating lymphocytes (TIL) plus interleukin-2 can mediate the regression of metastatic melanoma in approximately half of patients. To optimize this treatment approach and define the in vivo distribution and survival of TIL, we used retroviral-mediated gene transduction to introduce the gene coding for resistance to neomycin into human TIL before their infusion into patients--thus using the new gene as a marker for the infused cells. RESULTS Five patients received the gene-modified TIL. All the patients tolerated the treatment well, and no side effects due to the gene transduction were noted. The presence and expression of the neomycin-resistance gene were demonstrated in TIL from all the patients with Southern blot analysis and enzymatic assay for the neomycin phosphotransferase coded by the bacterial gene. Cells from four of the five patients grew successfully in high concentrations of G418, a neomycin analogue otherwise toxic to eukaryotic cells. With polymerase-chain-reaction analysis, gene-modified cells were consistently found in the circulation of all five patients for three weeks and for as long as two months in two patients. Cells were recovered from tumor deposits as much as 64 days after cell administration. The procedure was safe according to all criteria, including the absence of infections virus in TIL and in the patients. CONCLUSIONS These studies demonstrate the feasibility and safety of using retroviral gene transduction for human gene therapy and have implications for the design of TIL with improved antitumor potency, as well as for the possible use of lymphocytes for the gene therapy of other diseases.

A multiinstitutional survey of the Wiskott-Aldrich syndrome

The Wiskott-Aldrich syndrome is an X-linked primary immunodeficiency originally characterized by the clinical triad of thrombocytopenia, eczema, and immunodeficiency. We collected clinical and laboratory information on 154 unselected patients with Wiskott-Aldrich syndrome to define better the clinical expression of this disorder. The classic triad of thrombocytopenia with small platelets, recurrent otitis media, and eczema was seen in only 27% of the study population; 5% of the study population had only infectious manifestations, and 20% of the study group had only hematologic manifestations before diagnosis. The results of immunologic evaluations varied from one patient to another and the course of the disorder varied tremendously, even within a single kindred. We conclude that many patients with Wiskott-Aldrich syndrome have an atypical presentation and that a panel of diagnostic tests is often required to establish the diagnosis. Two high-risk subgroups were identified in the study population: patients with platelet counts < 10 x 10(9)/L (< 10,000/mm3) at the time of diagnosis were at high risk of bleeding, and patients with autoimmune disorders were at increased risk of having a malignancy.

Persisting Illness and Fatigue in Adults with Evidence of Epstein-Barr Virus Infection

Clinical, serologic, virologic, and immunologic evaluations for 31 adults with chronic illness and fatigue suggested that 23 had persisting Epstein-Barr virus infection. Among these 23 patients, cellular immune mechanisms were generally normal, but 4 had mild immunoglobulin deficiencies. However, 20 patients had abnormal serologic profiles specific for Epstein-Barr virus shown by significantly elevated titers of antibodies to the viral capsid antigen or early antigen, or by a deficiency of late-appearing antibodies. In 11 of 15 patients tested, circulating immune complexes were found. Circulating interferon was not found in 18 patients tested, but the activity of 2-5 oligoadenylate synthetase, an interferon-induced enzyme, was increased in 5 patients studied. Of 19 patients, 18 had persisting suppressor T-cell activity typically found in patients recovering from acute infectious mononucleosis. We believe that the Epstein-Barr virus may be associated with chronic illness in adults.

Gene Therapy for Cancer

Cancer is a complex acquired disease that affects millions of individuals each year. Underlying the pathogenesis of cancer are a variety of molecular genetic abnormalities which can be inherited or environmentally induced and result in unregulated cell proliferation. Conventional treatment strategies used to treat cancer: surgery, chemotherapy, and radiation have been only partially successful and new treatment options are critically needed. Gene therapy is currently being explored experimentally as an alternative or addition to established treatment options for malignant melanoma, leukemia, glioma, and others. The aim of this therapy is the introduction of a gene or genes into cells to provide a new set of permanent or temporary instructions for those cells resulting in the indirect elimination or direct killing of tumor cells. Indirect approaches to the treatment of cancer by gene therapy include the augmentation of the immune system; direct approaches include restoration of the normal function of a mutated tumor suppressor gene or expression of a tumoricidal gene. This review will describes these and other strategies.

Disorders of Suppressor Immunoregulatory Cells in the Pathogenesis of Immunodeficiency and Autoimmunity

A series of suppressor cell systems regulate virtually all immunologic processes. Disorders of these systems have been identified in association with a number of diseases. An abnormal number of activated suppressor T-cells have been seen in some patients with common variable hypogammaglobulinemia and in some with selective IgA deficiency. Suppressor T-cells that inhibit immunoglobulin synthesis also develop in an animal model of immunodeficiency, the agammaglobulinemia of the bursectomized bird. Non-T-cell suppressor cells are a pathogenic factor in the humoral immunodeficiency associated with multiple myeloma. At the other end of the spectrum of immunologic response, a reduction in functional activity of suppressor T-cells has been implicated in the pathogenesis of autoimmune diseases. The disorders of suppressor cells that have been shown in immunodeficiency and autoimmunity are important when developing rational strategies for prevention and therapy of these immunologic disorders.

Defective EBV-Specific Suppressor T-Cell Function in Rheumatoid Arthritis

Several lines of evidence, including high antibody titers to Epstein-Barr virus (EBV)-associated antigens and rapid transformation of B cells into lymphoblastoid cells lines, suggest an association between EBV and rheumatoid arthritis. When lymphocytes from normal immune donors were infected with EBV in culture, they produced an exponentially increasing number of immunoglobulin-secreting cells for eight to 10 days. Thereafter, there was a marked late suppression of their response, mediated by immunoregulatory T cells; by 12 days in culture, this suppression averaged 90 per cent. Lymphocytes from 20 EBV-immune patients with rheumatoid arthritis also responded with increasing production of immunoglobulin-secreting cells, but the late suppression expected in immune donors was absent. Tests of several other T-cell functions in these patients gave normal results, suggesting a more restricted defect in suppressor-T cell function relating specifically to EBV. Since EBV persists in host B cells and thus represents a potential stimulus for immunoglobulin production, this persistence, along with a specific regulatory T-cell defect, may contribute to many of the immune abnormalities underlying rheumatoid arthritis.

X-linked hypogammaglobulinemia and isolated growth hormone deficiency.

We undertook clinical, immunologic, and endocrinologic studies of a family in which two brothers and their two maternal uncles had a similar disorder characterized by hypogammaglobulinemia and isolated growth hormone deficiency. Recurrent sinopulmonary infections were a prominent feature in two patients. All patients had short stature and retarded bone age during childhood, and the adults had delayed onset of puberty. The immunodeficiency was characterized by absent specific antibody production in vivo and impaired immunoglobulin production in vitro. Three of the four patients lacked circulating B lymphocytes, even though tonsils were present in those patients. All patients had deficient growth hormone responses to insulin and arginine or levodopa. These patients have an X-linked recessive disorder, but their immunodeficiency differs from the X-linked immune disorders in the World Health Organization classification; their X-linked pattern of growth hormone deficiency, without other endocrine abnormality, is also unique.

Increased immunoglobulin-secreting cells in the blood of patients with active systemic lupus erythematosus.

Abstract Peripheral blood lymphoid cells actively secreting immunoglobulin G (IgG), immunoglobulin A (IgA) and immunoglobulin M (IgM) were quantitated in 24 patients with systemic lupus erythematosus (SLE) and compared with the frequency of such immunoglobulin secreting cells (IgSC) in normal controls utilizing a reverse hemolytic plaque assay. The geometric mean frequency of IgG-secreting cells in the patients with SLE was 489106 peripheral blood mononuclear cells which was significantly higher (p −5 ) but the highest correlation with disease activity found was the frequency of IgG-secreting cells in the blood of the patient with SLE (r = 0.777, p −6 ). These data demonstrate that SLE is associated with increased numbers of IgG- and IgA-secreting cells in the peripheral blood and that increases in these parameters are closely associated with active clinical disease.

The Wiskott-Aldrich syndrome. A disorder with a possible defect in antigen processing or recognition.

Abstract Immunological investigations were carried out in eleven patients with the Wiskott-Aldrich syndrome. Delayed hypersensitivity responses were absent when tested with five skin-test antigens and dinitrochlorobenzene sensitisation, despite the fact that the patients had mean circulating lymphocyte levels of 2300 per c.mm. and normal in-vitro lymphocyte transformation to phytohaemagglutinin. IgG levels were normal, IgA and IgD levels were elevated, and IgM levels were diminished. Turnover studies with 131 I-IgM demonstrated that the low IgM serum concentration was due to decreased synthesis. Natural antibodies to bloodgroup antigens and five serotypes of Escherichia coli were strikingly diminished. In addition, antibody responses to polysaccharide antigens were almost completely absent. It is, therefore, likely that the low IgM level is secondary to diminished responses to polysaccharide antigens which normally sustain the serum IgM concentration. Antibody responses were also significantly impaired to a variety of bacterial, protein, and viral antigens. However, when antibodies were produced, they were of both 19 S and 7 S types. We conclude that the Wiskott-Aldrich patients have a broad immunological defect involving both humoral and cellular immune responses. Such a broad defect associated with the presence of functional lymphocytes and adequate immunoglobulin levels suggests a disorder of antigen processing or recognition, that is, a disorder of the afferent limb of immunity.