First-in-man dose escalation and pharmacokinetic study of CAP7.1, a novel prodrug of etoposide, in adults with refractory solid tumours.

AIM: An open-label, phase I dose-escalation trial was performed in adult patients with various solid cancers to identify the maximum tolerated dose (MTD), to assess the safety, pharmacokinetic profile and anti-tumour activity of the new prodrug CAP7.1. The prodrug is converted to its active moiety etoposide via carboxylesterases in selective cells leading to a better tolerability and higher efficacy in therapeutic resistance cells and children with refractory neuroblastoma. PATIENTS AND METHODS: Eligible adult patients with advanced, refractory, solid malignancies received CAP7.1 as intravenous infusion on 5 consecutive days. Doses were escalated in four cohorts consisting of three to six patients, with a starting dose of 45 mg/m2/day. Treatment cycles were repeated in 21-day intervals in the absence of disease progression and prohibitive toxicity. The safety, pharmacokinetics and efficacy were evaluated, and the MTD and dose-limiting toxicity (DLT) were determined. RESULTS: Nineteen patients were assigned to four CAP7.1 dose cohorts (45, 90, 150 and 200 mg/m2/day). CAP7.1 was well tolerated. Haematotoxicity was observed at the two highest dose levels including three DLTs (two febrile neutropenia and one sepsis) only and were reversible with adequate therapy. No organ toxicity was observed. Non-haematological toxicities (mild-moderate) consist mainly of nausea, fatigue, vomiting, pyrexia and alopecia. One partial response and 11 stable diseases were observed as supporting signs of efficacy. CONCLUSION: MTD of CAP7.1 was reached at the dose of 200 mg/m2. A favourable safety profile and initial anti-tumour efficacy of CAP7.1 in therapeutic refractory tumours warrant further evaluation in clinical studies.

TIRC7 and HLA-DR axis contributes to inflammation in multiple sclerosis.

BACKGROUND AND OBJECTIVE: Interactions between TIRC7 (a novel seven-transmembrane receptor on activated lymphocytes) and its ligand HLA-DR might be involved in the inflammatory process in multiple sclerosis (MS). METHODS: Methods comprised immunohistochemistry and microscopy on archival MS autopsies, proliferation-, cytokine-, and surface-staining assays using peripheral blood lymphocytes (PBLs) from MS patients and an in vitro model. RESULTS: TIRC7 was expressed in brain-infiltrating lymphocytes and strongly correlated with disease activity in MS. TIRC7 expression was reduced in T cells and induced in B cells in PBLs obtained from MS patients. After ex vivo activation, T cell expression of TIRC7 was restored in patients with active MS disease. The interaction of TIRC7(+) T lymphocytes with cells expressing HLA-DR on their surface led to T cell proliferation and activation whereas an anti-TIRC7 mAb preventing interactions with its ligand inhibited proliferation and Th1 and Th17 cytokine expression in T cells obtained from MS patients and in myelin basic protein-specific T cell clone. CONCLUSION: Our findings suggest that TIRC7 is involved in inflammation in MS and anti-TIRC7 mAb can prevent immune activation via selective inhibition of Th1- and Th17-associated cytokine expression. This targeting approach may become a novel treatment option for MS.

MR proton relaxivities of some ions, albumin, and cholesterol added to odontogenic jaw cysts.

The relaxation rates (1/T(1) and 1/T(2)) in cysts have already been analyzed in terms of materials such as albumin, cholesterol, manganese, iron, and copper. However, the relaxivities of these materials have not been determined yet. In this work, five sets containing the ions, albumin, and cholesterol were prepared by addition of increasing concentration of one material to each set. The relaxation times in these sets were measured by MRI, and the relaxation rates were fitted versus concentrations. The slopes of the fits were used as relaxivities. The (r(1), r(2)) values of manganese, iron, and copper in mM(-1) s(-1), and those of albumin and cholesterol in (g/dl)(-1) s(-1) were found to be (32.64, 89.77), (0.31, 1.19), (0.5, 1.479), (0.01, 0.066) and (0.03, 0.458), respectively. The r(2)/r(1) ratio ranged from 2.75 to 15.27. Manganese is an efficient relaxer, but iron and copper are poor ones. Albumin and cholesterol are efficient relaxers for only T(2). The contribution of water associated with native manganese of the cystic fluid to T(1) was 0.268 s(-1), whereas those of water associated with native manganese, albumin, cholesterol, and iron to T(2) were 0.736, 0.185, 0.092, and 0.076 s(-1), respectively. The other contributions were much smaller than 0.076 s(-1). Manganese is most likely the compound altering T(1)-weighted images between different jaw cysts, whereas manganese and albumin are most likely the compounds altering the T(2)-weighted images. Present data suggest that such alterations may be used to separate jaw cysts from other jaw masses. The high r(2)/r(1) suggests that T(2) is a more convenient parameter than T(1) for diagnostic use.

Donor age intensifies the early immune response after transplantation.

Increasing donor age is associated with reduced graft function. We wondered if donor age may not only affect intrinsic function but also alter the immune response of the recipient. Kidneys from young and old F-344 rats (3 vs 18 months) were transplanted into bilaterally nephrectomized young Lewis recipients and compared with age-matched controls (follow-up: 6 months). Renal function and structural changes were assessed serially in both native kidneys and allografts. Host alloreactivity, graft-infiltrating cells, and their inflammatory products were determined at intervals to examine the correlation of immune response and donor age. Functional and structural deterioration had advanced significantly in older allografts compared with age-matched native controls, whereas differences between young allografts and native controls of similar age were only minor. Changes in grafts from elderly rats were associated with a more intense host immune response early post-transplant (up to 1 month) reflected by significantly higher numbers of peripheral T and B cells, increased T-cell alloreactivity and modified cytokine patterns associated with elevated frequencies of intragraft dendritic cells, B cells, and CD31+ cells. By 6 months, recipients of young donor grafts produced comparable or more intense alloantigen-specific immune responses. Older donor grafts elicit a stronger immune response in the early period after transplantation.

Antibody targeting of TIRC7 results in significant therapeutic effects on collagen-induced arthritis in mice.

TIRC7 is a cell surface molecule which is expressed in T and B lymphocytes and negatively regulates their function. Anti-TIRC7 specific monoclonal antibody (mAb) inhibited T cell memory response to recall antigens. Up-regulation of TIRC7 on lymphocytes from joint tissue of patients with Rheumatoid Arthritis (RA) and mice with collagen induced arthritis (CIA) suggested TIRC7 as a novel target to promote anti-inflammatory reaction. Anti-TIRC7 mAb administration significantly inhibited the induction and progression of CIA and the anti-collagen IgG1 and IgG2a antibody response. Combination therapy of anti-TIRC7 mAb and soluble TNF-alpha receptor demonstrated an increased inhibitory effect over the single compounds on CIA. The results demonstrate the therapeutic potential of TIRC7 targeting with mAb in diseases associated with exaggerated T and B cell responses.

Regulation of human intestinal intraepithelial lymphocyte cytolytic function by biliary glycoprotein (CD66a).

Human small intestinal intraepithelial lymphocytes (iIEL) are a unique population of CD8alphabeta+ TCR-alphabeta+ but CD28- T lymphocytes that may function in intestinal epithelial cell immunosurveillance. In an attempt to define novel cell surface molecules involved in iIEL function, we raised several mAbs against activated iIELs derived from the small intestine that recognized an Ag on activated, but not resting, iIELs. Using expression cloning and binding studies with Fc fusion proteins and transfectants, the cognate Ag of these mAbs was identified as the N domain of biliary glycoprotein (CD66a), a carcinoembryonic Ag-related molecule that contains an immune receptor tyrosine-based inhibitory motif. Functionally, these mAbs inhibited the anti-CD3-directed and lymphokine-activated killer activity of the P815 cell line by iIELs derived from the human small intestine. These studies indicate that the expression of biliary glycoprotein on activated human iIELs and, potentially, other mucosal T lymphocytes is involved in the down-regulation of cytolytic function.

Genomic organization of the gene coding for TIRC7, a novel membrane protein essential for T cell activation.

A novel human membrane protein, TIRC7, was recently identified and demonstrated to be essential in T cell activation. Here we report on the genomic organization of the TIRC7 gene, which is composed of 15 exons and spans 7.9 kb. The seven predicted transmembrane-spanning domains of the TIRC7 protein coincide well with exon-intron boundaries. TIRC7 and OC116, a recently described putative subunit of the vacuolar proton pump that was demonstrated to be expressed in an osteoclastoma tumor as well as in a human pancreatic adenocarcinoma cell line, are demonstrated to be alternative transcripts of the same gene. OC116 consists of 20 exons with the last 14 introns and exons being identical with those of TIRC7. The chromosomal locus for both transcripts was identified on chromosome 11q13.4-q13.5. In human alloactivated T lymphocytes, mRNA expression of TIRC7, but not OC116, is demonstrated, indicating that OC116 is not involved in regular T cell proliferation.

The human homolog of Drosophila cornichon protein is differentially expressed in alloactivated T-cells.

To identify novel genes induced in the early stage of T-cell activation, mRNA expression in alloactivated human lymphocytes was examined. Differential display-reverse transcription PCR analysis revealed a 207-bp cDNA fragment which was upregulated 24 h after allostimulation of a human T-cell line. The corresponding complete 1396 bp cDNA, named TGAM77, encodes a predicted 134 amino acid protein which shares 63% homology with the cornichon (cni) protein of Drosophila melanogaster. Upregulation of TGAM77 mRNA in the early phase of T-cell activation was confirmed by Northern blot and RT-PCR analysis of activated human lymphocytes. TGAM77 mRNA is expressed in a variety of human tissues with various expression levels. In analogy to cni which is involved in an epidermal growth factor-like signaling pathway inducing cellular asymmetry in Drosophila oogenesis, TGAM77 might function in similar signaling establishing vectorial re-localization and concentration of signaling events in T-cell activation.

Prevention of acute allograft rejection by antibody targeting of TIRC7, a novel T cell membrane protein.

A novel 75 kDa membrane protein, TIRC7, is described that exhibits a central role in T cell activation in vitro and in vivo. Modulation of TIRC7-mediated signals with specific anti-TIRC7 antibodies in vitro efficiently prevents human T cell proliferation and IL-2 secretion. Moreover, anti-TIRC7 antibodies specifically inhibit type 1 subset specific IFN-gamma expression but spare the type 2 cytokine IL-4. Diminished proliferation but not IFN-gamma secretion is reversible by exogenous rIL-2. An anti-TIRC7 antibody that cross-reacts with the 75 kDa rat homolog exhibits inhibition of rat alloimmune response in vitro and significantly prolongs kidney allograft survival in vivo. Targeting of TIRC7 may provide a novel therapeutic approach for modulation of the immune response.

Differential display cloning of a novel human histone deacetylase (HDAC3) cDNA from PHA-activated immune cells.

The nucleosomal histones can be modified through reversible acetylation by histone acetyltransferases (HATs) and deacetylases (HDACs). HATs induce nucleosomal relaxation and allow DNA-binding by transcriptional activators. HDACs from corepressor complexes which negatively regulate cell growth. However, the HDAC inhibitors butyrate and Trichostatin A block T cell proliferation, suggesting that not all effects of HDACs lead to repression. Using mRNA differential display and 5'RACE we isolated human HDAC3, a novel gene that is upregulated in PHA-activated T cell clones. HDAC3 is homologous to other human HDACs and yeast RPD3. In peripheral blood mononuclear cells (PBMCs), activation by PHA, PMA and alpha-CD3 increased HDAC mRNA but no effect was seen with IFN-gamma, LPS, or IL-4. In contrast, GMCSF downregulated PBMC levels of HDAC3 mRNA. All HDACs were found to be ubiquitously expressed in immune and non-immune tissues. In human myeloid leukemia THP-1 cells, HDAC3 transfection resulted in increased size, aberrant nuclear morphology and cell cycle G2/M cell accumulation. Functional activity of the expressed HDAC3 protein was confirmed in alpha-HDAC3 antibody immunoprecipitates by a histone deacetylase assay. Our study suggests the participation of HDACs in cell cycle progression and activation.

[Immunoglobulin therapy for systemic lupus erythematosus]. / Immunglobulintherapie bei systemischem Lupus erythematodes.

A 25-year-old patient with lupus erythematosus was admitted with myositis and erythema of the skin under chloroquine therapy. After improvement of clinical symptoms with cyclophosphamide and prednisolone he was again progredient with myositis. The changing of therapy to methotrexate showed a hepatotoxic side effect with elevated liver enzymes. Under subsequent therapy with azathioprine and prednisolone he developed leukopenia and sepsis. Because of persistent erythema of the skin under therapy with different immunosuppressives we performed a therapy with high-dose intravenous immunoglobulins. After application of immunoglobulins we observed an improvement of the erythema after 10 days, which was persistent after dose reduction for about 4 months.