Hugh I. McFarland

Treatment of experimental encephalomyelitis with a novel chimeric fusion protein of myelin basic protein and proteolipid protein.

It has been shown that peripheral T cell tolerance can be induced by systemic antigen administration. We have been interested in using this phenomenon to develop antigen-specific immunotherapies for T cell-mediated autoimmune diseases. In patients with the demyelinating disease multiple sclerosis (MS), multiple potentially autoantigenic epitopes have been identified on the two major proteins of the myelin sheath, myelin basic protein (MBP) and proteolipid protein (PLP). To generate a tolerogenic protein for the therapy of patients with MS, we have produced a protein fusion between the 21.5-kD isoform of MBP (MBP21.5) and a genetically engineered form of PLP (deltaPLP4). In this report, we describe the effects of treatment with this agent (MP4) on clinical disease in a murine model of demyelinating disease, experimental autoimmune encephalomyelitis (EAE). Treatment of SJL/J mice with MP4 after induction of EAE either by active immunization or by adoptive transfer of activated T cells completely prevented subsequent clinical paralysis. Importantly, the administration of MP4 completely suppressed the development of EAE initiated by the cotransfer of both MBP- and PLP-activated T cells. Prevention of clinical disease after the intravenous injection of MP4 was paralleled by the formation of long-lived functional peptide-MHC complexes in vivo, as well as by a significant reduction in both MBP- and PLP-specific T cell proliferative responses. Mice treated with MP4 were resistant to disease when rechallenged with an encephalitogenic PLP peptide emulsified in CFA, indicating that MP4 administration had a prolonged effect in vivo. Administration of MP4 was also found to markedly ameliorate the course of established clinical disease. Finally, MP4 therapy was equally efficacious in mice defective in Fas expression. These results support the conclusion that MP4 protein is highly effective in suppressing disease caused by multiple neuroantigen epitopes in experimentally induced demyelinating disease.

Autocrine Feedback Death and the Regulation of Mature T Lymphocyte Antigen Responses

Antigen-induced T cell death is an important regulatory mechanism in the peripheral immune system. Evidence suggests that this process depends on T cell growth-inducing lymphokines such as IL-2 and occurs in proportion to the degree of T cell receptor occupancy. Strong T cell receptor stimulation leads to the synthesis of death molecules such as Fas ligand and tumor necrosis factor that cause T cell suicide. We propose that T cell death under these circumstances is the culmination of a feedback control mechanism termed propriocidal regulation or autocrine feedback death that regulates the expansion of specific T cell clones under conditions of high lymphokine and antigen load. In a quasi-stochastic system such as the antigen receptor repertoire, feedback information may be essential for the appropriate regulation of peripheral immune responses. Our understanding of this feedback mechanism affords a means to manipulate antigen-specific T cell death in vivo. The application of this approach to the therapy of T cell-medicated immunological diseases is discussed.

Effective Antigen-Specific Immunotherapy in the Marmoset Model of Multiple Sclerosis

Mature T cells initially respond to Ag by activation and expansion, but high and repeated doses of Ag cause programmed cell death and can suppress T cell-mediated diseases in rodents. We evaluated repeated systemic Ag administration in a marmoset model of experimental allergic encephalomyelitis that closely resembles the human disease multiple sclerosis. We found that treatment with MP4, a chimeric, recombinant polypeptide containing human myelin basic protein and human proteolipid protein epitopes, prevented clinical symptoms and did not exacerbate disease. CNS lesions were also reduced as assessed in vivo by magnetic resonance imaging. Thus, specific Ag-directed therapy can be effective and nontoxic in primates.

Amelioration of Autoimmune Reactions by Antigen-Induced Apoptosis of T Cells

We have shown that T cells vigorously cycling in response to growth lymphokines are driven into apoptosis by potent TCR restimulation. This process, termed propriocidal regulation, appears to be a normal feedback inhibitory mechanism to prevent excessive T cell proliferation and lymphokine production. Exposure of T cells to repeated high dose antigen treatments creates the conditions just described by activating T cells, and stimulating the production of growth lymphokines and their receptors. High growth lymphokine levels induced by the large amount of antigen present, stimulate vigorous cycling. The continued presence of high antigen levels subjects the cycling T cells to strong TCR restimulation as they enter the vulnerable S phase, inducing apoptosis in T cells responsive to the administered antigen. Thus, simple, repetitive, intravenous administration of high dose antigen may be used to delete potentially destructive clones of T cells, resulting in a state of peripheral tolerance. This has obvious therapeutic potential in disorders where the elimination of pathogenic T cell clones could be beneficial. We have described in EAE, an animal model for MS, that high dose MBP therapy is effective in preventing CNS pathology and the onset of disease as well as reducing the severity of the clinical symptoms of established EAE. We are currently involved in expanding this approach to other animal models of autoimmunity and graft rejection, as well as refining the immunotherapy in the EAE model with the objective of developing a clinical therapy for human demyelinating disease.

T cell receptor transgenic mice recognizing the immunodominant epitope of the Torpedo californica acetylcholine receptor

Myasthenia gravis (MG) is an autoimmune disease caused by T cell‐dependent antibody‐mediated reduction of acetylcholine receptors (AChR) at the neuromuscular junction. Immunization of animals with Torpedo californica AChR (TAChR) results in an experimental model of MG. We used the variable regions of α and β T cell receptor (TCR) genes recognizing an immunodominant peptide containing amino acids 146–162 from the α subunit of TAChR presented in the context of I‐Ab to generate TCR‐transgenic mice. We found that the transgenic TCR was strongly positively selected and that transgenic T cells proliferated robustly to the immunodominant peptide and TAChR. Unexpectedly, there was a variable paucity of B cells in the blood and spleen from transgenic mice, which averaged about 16% of peripheral blood lymphocytes, compared to 55% in wild‐type B6 mice. Unselected transgenic mice immunized with TAChR exhibited weak anti‐TAChR antibody responses. However, transgenic mice selected to have relatively higher B cell numbers produced anti‐TAChR titers equal to B6 mice and a predominance of Th1‐induced antibody isotypes were observed in certain experiments. The incidence and severity of clinical disease was variable following immunizations. These mice should be useful for studying the pathogenesis and treatment of MG.