R.J.M. ten Berge

CD4 antibody treatment in patients with active Crohn's disease: a phase 1 dose finding study.

BACKGROUND: T cells play an important part in Crohns disease. Immunomodulating therapies that target T cell activation may have clinical effects in Crohns disease. AIM: To investigate the toxicity and potential efficacy of anti-CD4 monoclonal antibody therapy in patients with Crohns disease. PATIENTS AND METHODS: A dose escalating pilot study was conducted in three groups of four patients with intractable Crohns disease, refractory to steroids. They received 70, 210, or 700 mg of cM-T412, a depleting anti-CD4 monoclonal antibody (mAb). RESULTS: The mean reduction in Crohns disease activity index (CDAI) was respectively 25%, 24%, and 36% at four weeks, and 24% and 52% at 10 weeks in the 210 mg and 700 mg groups. There was only a minor effect on endoscopically evaluated disease activity. Side effects were mild to moderate fever with chills and headache. No signs of opportunistic infection were seen. There was a sustained decrease in CD4 count which lasted at least four weeks in the 70 mg group (76.3 (SD 40.6)% of the baseline value), and 10 weeks in both the 210 mg group (80.8 (SD 60.9)%) and the 700 mg group (24.8 (SD 15.4)%). The primary and secondary humoral immune response was not influenced by anti-CD4 mAb treatment. CONCLUSION: This study shows the moderate potential efficacy of treatment of patients with Crohns disease using a depleting chimeric monoclonal anti-CD4 antibody.

Administration of prednisolone in vivo affects the ratio of OKT4/OKT8 and the LDH-isoenzyme pattern of human T lymphocytes.

Oral administration of prednisolone (in single doses of 10, 30, or 60 mg) to healthy volunteers was found to affect the T lymphocytes in the blood with regard to binding of monoclonal antibodies and lactate dehydrogenase isoenzyme pattern. The findings indicate that these effects are dependent on the dose of the drug and the time after the administration of the drug. Prednisolone induces a T lymphocytopenia in the peripheral blood that affects OKT4-positive lymphocytes more than OKT8-positive lymphocytes, resulting in a slight decrease in the ratio OKT4/OKT8. Moreover, the lactate dehydrogenase isoenzyme pattern changes, resulting in a decrease of the H/M ratio of this enzyme. The proliferative responses of peripheral blood lymphocytes are not affected after a single dose of 10 mg. However, after administration of either 30 or 60 mg of prednisolone, the proliferative responses are decreased to a different extent, depending on the stimulus used. In vitro experiments are presented showing that any effect of prednisolone on nonstimulated lymphocytes is reversible. Based on the observed changes in OKT pattern and lactate dehydrogenase isoenzyme profile of the T lymphocytes induced by administration of prednisolone, it is concluded that the drug induces a temporary depletion from the peripheral blood, preferentially of high-reactive T lymphocytes. As a consequence, the peripheral blood compartment is enriched for T lymphocytes with a low H/M ratio of lactate dehydrogenase isoenzymes, known to be less reactive to proliferative stimuli.

Interleukin-6 concentrations in the serum of patients with AIDS-associated Kaposi's sarcoma during treatment with interferon-alpha

Abstract. Interleukin‐6 (IL‐6) levels were determined in the serum of 14 HIV‐1‐infected patients with Kaposis sarcoma. 10 HIV‐1‐infected patients without symptoms, and 10 healthy male subjects. IL‐6 levels were also determined in the serum of the 14 patients with Kaposis sarcoma during treatment with high‐dose human recombinant interferon‐alpha (IFNα). Serum IL‐6 levels were significantly higher in the patients with Kaposis sarcoma than in the HIV‐infected patients without symptoms and the controls. There was no consistent pattern of changes of IL‐6 levels during IFNα treatment. These results support the view that IL‐6 is a cytokine involved in the pathogenesis of AIDS‐associated Kaposis sarcoma, but appear to argue against an effect of IFNα on the production or release of IL‐6 as an important mechanism of action of IFNα.

Interleukin-6 and neopterin in renal transplant recipients: a longitudinal study

Serum and urine interleukin-6 (IL-6) levels and serum neopterin/creatinine ratios were longitudinally studied in 86 renal transplant recipients until 4 months after transplantation. During acute rejection and acute tubular necrosis (ATN), serum and urine IL-6 levels were elevated compared to during stable transplant function (P<0.001). During acute rejection, serum IL-6 levels increased at least 2 days before plasma creatinine started to rise (P<0.05), indicating its early involvement in the rejection process. During cytomegalovirus (CMV) disease, serum, but not urine, IL-6 levels were higher (P<0.01), and serum neopterin/creatinine values were higher than during stable transplant function, ATN, or acute rejection (P<0.01). No significant differences with stable transplant function occurred during cyclosporin A toxicity. Measurement of serum IL-6 provided a sensitivity of 84% and a specificity of 85% for the diagnosis of acute rejection episodes not coinciding with ATN. All cases of CMV disease could be diagnosed by measurement of serum neopterin/creatinine, which provided a specificity of 73%.

Administration of OKT3 as a two-hour infusion attenuates first-dose side effects

BACKGROUND Use of the murine CD3 monoclonal antibody OKT3 is limited by first-dose side effects, which are thought to be caused by the release of inflammatory mediators. Because these processes might be influenced by the speed of administration, we compared a 2-hr OKT3 infusion with the bolus infusion usually applied nowadays. METHODS Eighteen renal allograft recipients were prophylactically treated with OKT3 and randomized to receive the first dose either as a 2-hr infusion or as an intravenous bolus infusion. Clinical side effects score and the occurrence of complement activation, cytokine release, and activation of neutrophils were determined. RESULTS Two-hour infusion of OKT3 completely prevented the occurrence of dyspnea, reduced the incidence of other side effects, and attenuated complement activation. Cytokine release and depletion of peripheral blood lymphocytes were similar in both groups. CONCLUSIONS Thus, complement activation seems to play an additional role in the development of side effects after the first OKT3 dose.

Apoptosis of acinar cells in pancreas allograft rejection

BACKGROUND Recently it has been recognized that apoptosis of target cells may occur during liver and kidney allograft rejection and is probably induced by infiltrating cells. Pancreas rejection is also characterized by a cellular infiltrate, however, the occurrence of apoptosis has not been investigated. We assessed whether pancreas rejection was associated with apoptosis of target cells and an influx of granzyme B (GrB)-positive or CD68-positive cells. METHODS Eighteen pancreas biopsies (10 of 18 with rejection) from 15 patients with a pancreas-kidney transplantation were stained with the in situ end-labeling method for apoptosis, and for CD3, GrB, and CD68. RESULTS Significantly more apoptotic acinar cells were found in biopsies with rejection when compared with biopsies without rejection. No difference was observed between the groups for GrB+ or CD68+ cells. CONCLUSION We conclude that pancreas rejection is associated with apoptosis of acinar cells, but not with an increased influx of GrB+ cells or macrophages.

Analysis of 6-mercaptopurine in human plasma with a high-performance liquid chromatographic method including post-column derivatization and fluorimetric detection.

A relatively simple assay with improved reliability and sensitivity for measuring levels of 6-mercaptopurine in human plasma is presented. After extraction of the compound and the added internal standard with phenyl mercury acetate, samples were separated by ion-pair reversed-phase high-performance liquid chromatography. On-line the analytes were oxidized to fluorescent products and detected in a flow-fluorimeter. The within-day coefficient of variation was 3.8% at a concentration of 25 ng/ml. The lower detection limit was 2 ng/ml when 1.0 ml of plasma was used. Mercaptopurine concentration versus time curves of two subjects after a single oral dose of azathioprine are shown.

Influence of HLA-DRB1* incompatibility on the occurrence of rejection episodes and graft survival in serologically HLA-DR-matched renal transplant combinations

BACKGROUND The aim of the present study was to analyze the effect of HLA-DRB1* mismatches on graft function and graft survival in 92 patients who received serologically HLA-DR split antigen-matched cadaveric renal transplants. METHODS The polymorphic second exon of the HLA-DRB1 alleles was typed using the sequence-specific oligonucleotides technique. RESULTS The results show that in 26 of the 92 analyzed combinations, one or more HLA-DRB1* mismatches were found (28%). The analysis of the occurrence of treatable rejection episodes during the first 3 months after transplantation demonstrated a significantly higher incidence of rejection episodes in the HLA-DRB1*-mismatched group: 18 of 26 (69%) in the HLA-DRB1*-mismatched group against 23 of 66 (35%) in the HLA-DRB1*-matched group (P(uncorr)=0.0033). However, no effect of HLA-DRB1* mismatches on graft survival was found, although in general graft survival in the whole patient group was negatively influenced by the occurrence of rejection episodes during the first 3 months after transplantation (P(uncorr)=0.0008). In contrast, in the HLA-DR4-matched donor-recipient combinations (n=28), the effect of mismatching for the HLA-DRB1*04 alleles seemed to have a pronounced effect not only on the occurrence of rejection episodes but also in the form of diminished graft survival. CONCLUSIONS Thus, this study indicates that the existence of HLA-DRB1* allele mismatches in renal transplant recipients, matched for the serologically defined HLA-DR split antigens, is not harmful for the transplant. The exception is the HLA-DRB1*04 mismatch, which seems to be deleterious for the grafted organ.

Sequestration of labelled granulocytes in the lungs following administration of OKT3 is dose-dependent

In the present study the consequences of administration of low-dose (0.5 mg) OKT3 for respiratory side-effects and pulmonary sequestration of labelled granulocytes are compared with the known effects of 5 mg OKT3. Ten renal transplant patients were studied, of whom five were treated with 0.5 mg OKT3 and five with 5 mg OKT3. None of the patients in the 0.5 mg group and two of the patients in the 5 mg group experienced dyspnoea. Sequestration of labelled granulocytes in the lungs was significantly lower in the patients receiving 0.5 mg OKT3 compared with the patients receiving 5 mg OKT3. The simultaneously occurring peripheral blood granulocytopenia was significantly more severe in the 5 mg group than in the 0.5 mg group. We suppose that this sequestration of circulating granulocytes in the lungs is at least partly mediated by complement activation products. In vitro it is demonstrated that fixation of complement activation products on peripheral blood lymphocytes depends on the concentration of OKT3 present in the culture medium. We conclude that respiratory side-effects shortly following infusion of OKT3 are related to complement-induced pulmonary leucostasis, the degree of which is dependent on the administered dose of OKT3.

Allo‐HLA Cross‐Reactivities of Cytomegalovirus‐, Influenza‐, and Varicella Zoster Virus–Specific Memory T Cells Are Shared by Different Healthy Individuals

Virus‐specific T cells can recognize allogeneic HLA (allo‐HLA) through TCR cross‐reactivity. The allospecificity often differs by individual (private cross‐reactivity) but also can be shared by multiple individuals (public cross‐reactivity); however, only a few examples of the latter have been described. Because these could facilitate alloreactivity prediction in transplantation, we aimed to identify novel public cross‐reactivities of human virus‐specific CD8+ T cells directed against allo‐HLA by assessing their reactivity in mixed‐lymphocyte reactions. Further characterization was done by studying TCR usage with primer‐based DNA sequencing, cytokine production with ELISAs, and cytotoxicity with 51chromium‐release assays. We identified three novel public allo‐HLA cross‐reactivities of human virus‐specific CD8+ T cells. CMV B35/IPS CD8+ T cells cross‐reacted with HLA‐B51 and/or HLA‐B58/B57 (23% of tetramer‐positive individuals), FLU A2/GIL (influenza IMP[58‐66] HLA‐A*02:01/GILGFVFTL) CD8+ T cells with HLA‐B38 (90% of tetramer‐positive individuals), and VZV A2/ALW (varicella zoster virus IE62[593‐601] HLA‐A*02:01/ALWALPHAA) CD8+ T cells with HLA‐B55 (two unrelated individuals). Cross‐reactivity was tested against different cell types including endothelial and epithelial cells. All cross‐reactive T cells expressed a memory phenotype, emphasizing the importance for transplantation. We conclude that public allo‐HLA cross‐reactivity of virus‐specific memory T cells is not uncommon and may create novel opportunities for alloreactivity prediction and risk estimation in transplantation.