Evelyne Racadot

Treatment of acute graft-versus-host disease with methylprednisolone and cyclosporine with or without an anti-interleukin-2 receptor monoclonal antibody : a multicenter phase III study

A double-blind, placebo-controlled trial of BT563, including 13 European centers, was initiated in October 1989 to compare the efficacy of the combination of in vivo anti-CD25 mAb (BT 563), cyclosporine, and steroids versus placebo and CSA-steroids in the treatment of grade II and III acute graft-versus-host disease (GVHD). Sixty-nine patients participated in the study, which excluded non-genotypically identical allogeneic bone marrow transplant recipients. No statistically significant differences were observed, clinically or biologically, between the 2 groups before the onset of the treatment. Treatment responses were scored during and after the 3-week treatment period (mAb or placebo). Efficacy was evaluated on days 4, 10, 20, 30, and 60 or on any day the patients condition was found to be deteriorating. Preceding and systemically untreated GVHD of grade I was observed in 59% of the cases. No statistically clinically significant differences between the 2 groups were observed during or upon completion of treatment in GVHD grade. Nine patients in the placebo group and 6 in the active group were withdrawn of the study. Thirteen of these 15 patients were withdrawn because of failure of GVHD therapy (9 in the placebo group and 4 in the BT563 group). At day 20 after onset of the treatment, the response rate was 63% and 70% for the placebo and BT563 groups, respectively (NS). Probability of survival at 1 year was 59% and 66% (NS) for the placebo and active groups, respectively. In conclusion, despite preliminary promising results in the treatment of steroid-resistant acute GVHD, the role of first-line treatment with an in vivo anti-interleukin-2 receptor mAb remains to be determined.

Serum levels of MMP-3 and cathepsin K in patients with ankylosing spondylitis: Effect of TNFα antagonist therapy ☆

OBJECTIVE To measure serum levels of MMP-3 and cathepsin K in patients with ankylosing spondylitis (AS) and in controls and to look for changes in these variables during TNFalpha antagonist therapy. METHODS We prospectively studied a group of patients who met New York criteria for AS and a group of healthy volunteers. We recorded age, disease duration, main features of the disease, BASDAI, erythrocyte sedimentation rate (ESR), and C-reactive protein (CRP). Serum MMP-3 and cathepsin K were assayed in duplicate using ELISA kits (Quantikine MMP-3, R&D Systems; and Cathepsin K, Biomedica). We also assayed IL-17 (Quantikine IL-17, R&D Systems) and BMP-7 (human BMP-7 DuoSet, R&D Systems). In patients treated with TNFalpha antagonists, the assays were repeated 10 weeks after treatment initiation. The Mann-Whitney test was used for between-group comparisons and the Wilcoxon test for evaluations of changes under treatment. Correlation testing was performed. P-values less than 0.05 were considered significant. RESULTS We studied 23 outpatients with AS and 21 controls, with mean age of 39.9 years and 41.2 years, respectively (NS). Disease duration was 10.1 years (1.3); most patients had axial disease (n=21) and carried HLA-B27 (n=19). At baseline, the mean BASDAI was 44.1 mm (4.1) and the mean CRP level was 22.3 mg/L (4.7). Serum MMP-3 levels were significantly higher in the patients than in the controls (4.71 vs. 2.79 ng/ml, P=0.04); levels were also higher for cathepsin K (6.4 vs. 3.6 pg/ml) and IL-17 (60.4 vs. 32 pg/ml), but the differences were not statistically significant. No difference was noted for BMP-7. The only positive correlation was between the ESR and the CRP level (P=0.0002). Thirteen patients were evaluated 10 weeks into TNFalpha antagonist therapy (adalimumab, n=7; etanercept, n=4; or infliximab, n=2). Serum MMP-3 decreased significantly (P=0.04); significant decreases were also noted for the ESR, CRP, and BASDAI. CONCLUSION MMP-3 is significantly increased in patients with active AS but fails to correlate significantly with conventional variables used to assess disease activity. TNFalpha antagonist therapy induces a significant decrease in MMP-3 levels, together with decreases in conventional variables (ESR, CRP, and BASDAI). MMP-3 may be a biomarker for disease activity in AS but supplies no additional information to the clinician.

Treatment of rheumatoid arthritis with anti CD4 monoclonal antibody. Open study of 25 patients with the B-F5 clone

SummaryTwenty-five defined severe RA patients (pts) (17 F, 8 M) were treated in an open study with a CD4 murine monoclonal antibody (Mab) (B-F5 clone, IgG1). Mabs daily dose was 10 mg (1 pt), 15 mg (2 pts), 20 mg (17 pts), 30 mg (4 pts) and 50 mg (1 pt) for 10 days. Tolerance was fair. Clinical improvement occurred during treatment period or within the first month in all but 2 patients, irrespective of Mab dosage. Improvement duration was variable (1 to 12 months), half of the patients still show signs of improvement at month 4. Biological parameters (CRP) improved parallel to the clinical. At day 180, 25% of the patients showed a reduction of 50% or more of the initial CRP values. There is no modification of RF titers, renal and hepatic parameters. Sequential evaluation showed a decrease of B, TCD3, CD4, CD8 lymphocytes and monocytes two hours after Mab infusion and return to baseline in 20 hours. Xenogenic immunization occurred in 6 patients without influence upon clinical response. These modifications are moderate and transient and do not account for the more prolonged effect in some cases, nor do they offer any prediction of further clinical response.

Graft Failure after T Cell Depleted HLA Identical Allogeneic Bone Marrow Transplantation: Risk Factors in Leukemic Patients

In a retrospective analysis of T cell depleted bone marrow transplantation, we have looked at different parameters in order to determine risk-factors of graft-failure after allogeneic bone marrow transplantation for leukemia. Fifty-one patients with acute leukemia or chronic myeloid leukemia have been analysed. For 33 of them, the pretransplant conditioning regimen consisted of fractionated total body irradiation (TBI) at 12 Gy prior to cyclophosphamide (120 mg/kg). The other patients received various reinforced preparative regimens. T-cell depletion consisted of treating marrow cells with pan-T monoclonal antibodies (CD2+CD3 or CD2-CD5-CD7) followed by complement mediated cytolysis. No post-transplant immunosuppressive prophylaxis was administered except for the first nine patients who received Methotrexate alone. In this group of 51 patients, 12 died within 3 months from graft-related complications and 10 developed graft failure (no engraftment or rejection). Among the possible risk factors associated with this failure, two graft-related parameters appeared significant: the number of CFU-GM progenitors and the number of viable T cells injected with the marrow inoculum. No correlation with graft failure was found with other parameters including diagnosis, disease status at transplant, conditioning regimen, age, sex, and CMV status of donor/host pairs. However, the interpretation must remain cautious because of the relatively small samples in each group.

Soluble interleukin-2 receptors in chronic renal failure.

A depression of the general immune response in uremia is well documented, and hemodialyzed (HD) patients present deficient interleukin-2 (IL2) secretion. Since soluble IL2 receptors (SIL2R) could affect the immune response through interaction with circulating immune cells, we studied the potential relationship between SIL2R concentration and lymphocyte subsets in 44 HD patients. HD patients present lymphopenia, higher CD4/CD8 ratio. CD16 counts and SIL2R concentrations than controls. A significant negative correlation was found between SIL2R concentration and lymphocyte count (p less than 0.01), and between SIL2R concentration and T4/T8 ratio (p less than 0.01). An increase of SIL2R concentration due to abnormal T cell preactivation in HD patients with nonreused cuprophan membranes could perhaps contribute to cell immunity impairment through IL2 binding and inhibition of T cell activation.

Comparative Preclinical Study of Three Bone Marrow Purging Methods Using PCR Evaluation of Residual t(14;18) Lymphoma Cells

The t(14;18) chromosomal translocation occurring in most follicular lymphomas can be exploited by a Bcl2/JH polymerase chain reaction (PCR) to detect residual disease and to monitor the effectiveness of ex-vivo tumor cell immunological purging. We first demonstrated the 10(-5) Bcl2/JH PCR sensitivity with serial dilutions of OCY-LY8 lymphoma cell lines in normal mononuclear cells; and then the specificity and reproductibility of this technique by analysing follicular and non follicular lymphoma samples. With the Bcl2/JH PCR, we tested the efficiency of three marrow purging protocols with an experimentally contaminated bone marrow either treated by three anti-B cell monoclonal antibodies (mAb) followed by three rounds of rabbit complement or two rounds of immunomagnetics beads. Samples obtained after each purging were amplified by Bcl2/JH PCR and hybridized with PFL3 probe. We were able to produce a 2 to 3 log tumor cell reduction after three rounds of complement and a 4 to 5 log reduction after two rounds of beads. This study showed that it is feasible to use the Bcl2/JH PCR technique for residual cell lymphoma detection in patients undergoing intensive chemotherapy or BM transplantation. These results indicate that ex-vivo immunomagnetic BM purging is probably superior to complement mediated lysis for the eradication of B lymphoma cells from the marrow of patients undergoing autologous transplantation.

Soluble CD8, IL-2 receptor, and tumor necrosis factor-alpha levels in steroid-resistant acute graft-versus-host disease : relation with subsequent response to anti-IL-2 receptor monoclonal antibody treatment

Serial determination of soluble CD8 (sCD8), soluble IL-2 receptors (sIL-2R), and tumor necrosis factor-alpha serum levels were performed in bone marrow transplant patients upon initiation, day 0 (D0) and at D10 of an anti-IL-2 receptor (alpha chain) monoclonal antibody (B-B10) in vivo treatment for steroid-resistant grade greater than or equal to 2 acute graft-versus-host disease (aGVHD). D0 and D10 sCD8 serum levels correlated strongly with response to B-B10 treatment (p = .003 and .001, respectively); 76% of the patients with D0 sCD8 levels less than 500 U/ml responded favorably to B-B10 treatment, versus only a 30% response if the sCD8 levels were greater than 500 U/ml (p = .02). Likewise, D0 tumor necrosis factor-alpha levels significantly correlated with subsequent response to B-B10 treatment (p = .03). D0 sIL-2R levels were not significantly different in B-B10-responsive and nonresponsive aGVHD patients. These results suggest that the serial determination of sCD8 and TNF serum levels could provide valuable predictive information as to steroid-resistant aGVHD responsiveness to anti-IL-2R treatment.

Preclinical studies of a panel of 12 monoclonal antibodies in view of bone marrow purging in acute lymphoblastic leukemia

We analysed the optimal conditions for autologous bone marrow purging using complement binding monoclonal antibodies (mAbs). Twelve mAbs belonging to four clusters (CD9, CD10, CD19, CD24), alone or combined were evaluated by using direct cytotoxicity and clonogenic assays. We observed the following data: (1) optimal cytotoxicity was reached with doses of 1-10 micrograms mAbs for 10(7) cells, (2) the concentration of the cell suspension had to be below 3 X 10(7)/ml, (3) combinations of mAbs were more effective than a single mAb treatment, (4) in the case of an IgM isotype, there seems to be a clear dissociation between the amount needed for optimal toxicity and that for antigenic saturation measured by cytofluorometry.

Clinical characteristics and laboratory testing of patients with suspected HIT: a survey on current practice in 11 university hospitals in France.

UNLABELLED We undertook a survey of French university hospital hematological laboratories to ascertain the clinical characteristics of patients with suspected HIT, the laboratory tests performed, and the therapeutic strategy adopted in current practice. METHODS A standardized medical records database for patients with suspected HIT was sent to 19 laboratories. During two months, all consecutive patients for whom a biological test was performed were included. RESULTS 169 patients were included, 27 (16%) patients having a final diagnosis of HIT. At the time HIT was suspected, the heparin duration and the level of thrombocytopenia were similar in HIT- positive and HIT-negative groups. The use of unfractionated heparin, a therapeutic heparin dose regimen and the presence of thrombotic complications were significantly more frequent in HIT-positive patients. When the heparin dose regimen was taken into account, only thrombotic complications under a therapeutic dose regimen were significantly increased in HIT-positive patients. Eighty-six percent of patients presented at least one alternative diagnosis of thrombocytopenia without significant difference between the two groups. Laboratory tests were performed after a mean of 0.3days and mainly consisted of antigen assays. At the time HIT was suspected, heparin was stopped in 56 (33%) patients, being replaced mainly by danaparoid. Only three laboratories declared they usually received all the necessary clinical information to establish the likelihood of HIT. CONCLUSION In current practice in France, the clinical probability of HIT is rarely established, leading to systematic requests for laboratory HIT tests.

Interferon γ, IL2, IL4, IL10 and TNFα Secretions in Multiple Sclerosis Patients Treated with an Anti-CD4 Monoclonal Antibody

In order to better understand the mechanisms of action of a monoclonal anti-CD4/BF5 antibody(mAb), cytokine secretions were studied in 14 multiple sclerosis (MS) patients treated in a phase 1 trial. Secretion patterns of IFNγ, IL2, IL4, IL10 and TNFα by peripheral blood mononuclear cells were studied before (DO) and after (D30) the treatment. We decided to undertake this study because in a previous one we observed no variations in serum levels of TNFα, IFNγ, IL1, IL6. Results showed significant reductions in IFNγ IL2 and TNFα secretions after treatment. The anti-CD4 mAb seemed to act on both Th1- and Th2-cells but with preferential action on Th1-cells. Results on Th2-cells were less obvious even though a significant decrease in IL10 was observed. There was no correlation between any of the immunological markers studied and disease activity. This study demonstrates that pharmacological modifications of the CD4 receptor can induce variations in several cytokine secretion levels. It also stresses the role pl...