Laura R. Tranquill

Phase 1 trial of transforming growth factor beta 2 in chronic progressive MS

Transforming growth factor (TGF)-β2 is a pleiotropic cytokine associated with remissions in multiple sclerosis (MS) and amelioration of allergic encephalomyelitis. We assessed the safety of TGF-β2 in an open-label trial of 11 patients with secondary progressive (SP) MS. Five patients had a reversible decline in the glomerular filtration rate. There was no change in expanded disability status scale or MRI lesions during treatment. Systemic TGF-β2 may be associated with reversible nephrotoxicity, and further investigation of its therapeutic potential in MS should be performed with caution.

Immunodominance of a low-affinity major histocompatibility complex-binding myelin basic protein epitope (residues 111-129) in HLA-DR4 (B1*0401) subjects is associated with a restricted T cell receptor repertoire.

The pathogenesis of multiple sclerosis (MS) is currently ascribed in part to a T cell-mediated process targeting myelin components. The T cell response to one candidate autoantigen, myelin basic protein (MBP), in the context of HLA-DR15Dw2, has been previously studied in detail. However, the characteristics of cellular immunity in the context of other MS-associated HLA-DR haplotypes are scarcely known. MBP-specific T cell lines (TCL) were generated from HLA-DR4 (B1*0401)-positive MS subjects. Out of 275 MBP-specific TCL, 178 (64. 7%) specifically recognized region MBP(111-129), predominantly in the context of DRB1*0401. The major T cell epitope for MBP recognition corresponded to residues MBP(116-123). These TCL expressed disparate profiles of cytokine secretion and cytotoxicity. T cell receptor analysis, on the other hand, revealed a strikingly limited heterogeneity of rearrangements. In contrast to MBP(81-99), which binds with high affinity to HLA-DR15 and is recognized by a diverse T cell repertoire, MBP(111-129) binds weakly to DRB1*0401, suggesting that only high affinity T cell receptors might be able to efficiently engage such unstable MHC/peptide complexes, thus accounting for the T cell receptor restriction we observed. This study provides new insight about MBP recognition and proposes an alternative mechanism for immunodominance of self-antigen T cell epitopes in humans.

Identification of a novel T cell epitope of human proteolipid protein (residues 40-60) recognized by proliferative and cytolytic CD4+ T cells from multiple sclerosis patients

Research into the pathogenesis of multiple sclerosis (MS) has focused on myelin antigens as potential targets of autoimmune attack. Proteolipid protein (PLP) is the most abundant myelin protein comprising more than 50% of central nervous system myelin. Although PLP is a hydrophobic membrane protein which has made it difficult to study, the use of synthetic peptides based on the PLP sequence provides an alternative method for studying the immunological properties of PLP. Using peripheral blood lymphocytes from MS patients, long-term TCL established in the presence of PLP reacted weakly to PLP in proliferation assays; however, these same lines were much more reactive to synthetic peptides of PLP. Thus, we established short-term T cell lines (TCL) from the peripheral blood lymphocytes (PBL) of MS patients in the presence of five separate synthetic PLP peptides. In 6/7 MS patients, proliferative responses were elicited most often to PLP 40-60 compared to four other PLP peptides (PLP 89-106, 103-120, 125-143, and 139-154) (Pelfrey et al., 1993). Interestingly, however, the magnitude of the proliferative response was greatest in response to PLP 89-106. Characterization of PLP 89-106-responsive TCL from several MS patients, indicated that TCL proliferating to the peptide also lysed PLP 89-106 pulsed autologous targets. The majority of cytolytic PLP 89-106 TCL were CD4+ and MHC class II restricted and the predominant restriction elements were those most commonly found in MS patients. These findings suggest that the use of synthetic peptides represents a viable alternative approach to the study of PLP reactivity in humans. We report here that MS PBL recognize several PLP peptides, with the predominant responses to PLP 40-60 and PLP 89-106. Since these cells have both helper (CD4+) and cytolytic capabilities, it is possible that they may play a role in the pathogenesis or progression of MS.

Enhanced T cell responsiveness to citrulline-containing myelin basic protein in multiple sclerosis patients

Myelin basic protein (MBP), a candidate autoantigen in multiple sclerosis (MS), exists in different isoforms and charge isomers generated by differential splicing of exons and by a combination of posttranslational modifications, respectively. These various isoforms and charge isomers of MBP vary in abundance and most likely serve different functions during myelinogenesis and remyelination. The least cationic among the charge isomers of MBP is citrullinated and is referred to as MBP-C8. MBP-C8 is relatively increased in the population of MBP isomers in more developmentally immature myelin and in MS brain tissue. In a previous study, we found that MBP-C8-reactive T cells could be detected in CD4+ T cell lines (TCL) generated with MBP from both MS patients and normal controls. Here, we examined the frequency and peptide specificity of MBPC8-specific TCL generated with MBP-C8 in MS patients and controls. Ten subjects grouped in five sets, each an MS patient and a control, were studied. In all cases, the MS patient had either a higher overall number of MBP-C8-responding lines, responded with greater sensitivity to the MBPC8 antigen or both. Few lines responded to the MBP-C8 peptides but, if they did, they appeared to be specific to the carboxyl-half of the MBP-C8 molecule. Given the large amounts of citrullinated MBP in MS brain tissue, a preferential T cell response to MBP-C8 may be involved in the induction and perpetuation of this disease.

Cytokine gene expression in cells derived from CSF of multiple sclerosis patients.

Cytokines are produced by numerous cell types in the peripheral blood and central nervous system (CNS), and have been implicated in the immunopathogenesis of multiple sclerosis (MS). We examined the relationship between cytokine gene expression of cerebrospinal fluid (CSF) derived cells from MS patients and disease activity as measured by contrast enhanced MRI. There was a significant correlation between the number of CSF cells and the number of contrast enhancing MRI lesions. Cytokine gene expression of TNF-alpha, IFN-gamma, and IL-10 was routinely seen, but IL-4 expression was absent except in two clinically quiescent patients. Trends were observed toward decreased TNF-alpha expression, but increased IL-10 expression, after treatment with IFN-beta1b.

T cell response to two immunodominant proteolipid protein (PLP) peptides in multiple sclerosis patients and healthy controls

Multiple sclerosis (MS) is a demyelinating disease of the central nervous system in which autoimmune T lymphocytes reacting with myelin antigens are believed to play a pathogenic role. Since HLA binding is involved in the selection of T cell responses, we have examined PLP peptide binding to HLA DR2, an HLA allele frequently found in MS patients. Both PLP 40–60 and PLP 89–106 show significant, high affinity binding to HLA DR2. We then tested whether responses to PLP peptides 40–60 and 89–106 are elevated in multiple sclerosis patients compared to matched controls. We also analysed T cell responses to MBP 87–106, which is considered to be the immunodominant region of MBP in humans. Here we demonstrate heterogenous T cell responses to PLP 40–60, PLP 89–106 and MBP 87–106 in both MS patients and controls. The overall number of TCL and the HLA restriction of those TCL did not vary significantly in the two groups. PLP 40–60 specific cytolytic TCL were increased in MS patients, whereas healthy controls had increased percentages of cytolytic TCL responding to PLP 89–106 and MBP 87–106. Although the data presented here shows heterogenous responses in T cell numbers, differences in numbers and specificity of cytolytic cells could be involved in the pathogenesis of autoimmune demyelinating disease.

A novel candidate autoantigen in a multiplex family with multiple sclerosis: prevalence of T-lymphocytes specific for an MBP epitope unique to myelination.

Abstract Although the major isoform of myelin basic protein (MBP) in the healthy adult CNS is the 18.5-kDa protein, other isoforms containing exon 2 encoded protein (21.5 kDa and 20.2 kDa) exist and are expressed primarily during myelin formation. Since remyelination is a prominent feature in MS lesions, we examined the frequencies of T cell lines (TCLs) specific for epitopes within exon 2 encoded MBP (X2MBP), and also within 18.5-kDa MBP, in members of a multiplex family with MS. TCLs specific for X2MBP were as prevalent as TCLs specific for immunodominant epitopes within 18.5-kDa MBP. In addition, while frequencies of TCLs specific for 18.5-kDa MBP were no different between the affected and unaffected, the frequency of X2MBP-specific TCLs correlated with disease.

Human T lymphocytes specific for the immunodominant 83–99 epitope of myelin basic protein: Recognition of golli MBP HOG 7

Myelin basic protein (MBP) is a candidate auto‐antigen in the disease multiple sclerosis. Although MBP was thought to be sequestered behind the blood‐brain barrier, isoforms of MBPs have recently been demonstrated in lymphoid tissues. These isoforms, termed golli MBPs, contain sequences that are shared with “classic” MBP within the CNS. In the present study, we have determined that epitopes within golli MBP isoforms may be recognized by human T lymphocyte clones specific for classic MBP. Ten of 12 T‐cell clones recognized golli MBP. Although 11 clones were specific for the immunodominant 83–99 sequence, the clones differed with respect to human leukocyte antigen (HLA) restriction, T‐helper phenotype, cytolytic activity, and T‐cell receptor usage. Greater responses to classic MBP than to golli MBP suggested a difference in the ability of the two proteins to be processed and to present epitopes therein. These data advance the hypothesis that golli MBP sequences expressed within lymphoid tissues may be recognized by classic MBP‐specific T lymphocytes during central or peripheral tolerance.