Pharmacokinetics and antibody responses to the CD3 antibody otelixizumab used in the treatment of type 1 diabetes.

Otelixizumab is a chimeric CD3 antibody that has been genetically engineered to remove the glycosylation site in the Fc domain. This limits its ability to bind to complement or Fc receptors and reduces the risk of adverse clinical reactions due to cytokine release. In a trial for treatment of type 1 diabetes, a short treatment with otelixizumab resulted in a reduced requirement for insulin lasting at least 18 months. In the course of this trial, the blood concentrations of the antibody were measured by flow cytometry to determine its pharmacokinetic profile. Dose-dependent accumulation of otelixizumab was demonstrated and modeling of the data indicated that the terminal half-life was approximately 1.5 days. Antibody responses to otelixizumab were measured by 2 methods: a bridging enzyme-linked immunosorbent assay and surface plasmon resonance. The surface plasmon resonance method had a greater sensitivity and was able to detect responses in all patients, starting at 8 days after the commencement of therapy. Neutralizing antibodies were detected in a significant proportion of patients by days 22 to 29. Although no adverse clinical effects were associated with these antibody responses and they did not appear to affect the clearance of the drug, they might have important implications for possible retreatment of the patients.

Insulin needs after CD3-antibody therapy in new-onset type 1 diabetes.

BACKGROUND: Type 1 diabetes mellitus is a T-cell-mediated autoimmune disease that leads to a major loss of insulin-secreting beta cells. The further decline of beta-cell function after clinical onset might be prevented by treatment with CD3 monoclonal antibodies, as suggested by the results of a phase 1 study. To provide proof of this therapeutic principle at the metabolic level, we initiated a phase 2 placebo-controlled trial with a humanized antibody, an aglycosylated human IgG1 antibody directed against CD3 (ChAglyCD3). METHODS: In a multicenter study, 80 patients with new-onset type 1 diabetes were randomly assigned to receive placebo or ChAglyCD3 for six consecutive days. Patients were followed for 18 months, during which their daily insulin needs and residual beta-cell function were assessed according to glucose-clamp-induced C-peptide release before and after the administration of glucagon. RESULTS: At 6, 12, and 18 months, residual beta-cell function was better maintained with ChAglyCD3 than with placebo. The insulin dose increased in the placebo group but not in the ChAglyCD3 group. This effect of ChAglyCD3 was most pronounced among patients with initial residual beta-cell function at or above the 50th percentile of the 80 patients. In this subgroup, the mean insulin dose at 18 months was 0.22 IU per kilogram of body weight per day with ChAglyCD3, as compared with 0.61 IU per kilogram with placebo (P<0.001). In this subgroup, 12 of 16 patients who received ChAglyCD3 (75 percent) received minimal doses of insulin (< or =0.25 IU per kilogram per day) as compared with none of the 21 patients who received placebo. Administration of ChAglyCD3 was associated with a moderate "flu-like" syndrome and transient symptoms of Epstein-Barr viral mononucleosis. CONCLUSIONS: Short-term treatment with CD3 antibody preserves residual beta-cell function for at least 18 months in patients with recent-onset type 1 diabetes.

Neutralizing tumor necrosis factor activity leads to remission in patients with refractory noninfectious posterior uveitis.

OBJECTIVE: To evaluate the efficacy and safety of tumor necrosis factor (TNF) inhibition with the p55 TNF receptor fusion protein (TNFr-Ig) for severe sight-threatening noninfectious posterior segment intraocular inflammation. METHODS: Seventeen patients with refractory noninfectious posterior segment intraocular inflammation received TNFr-Ig by intravenous infusion in this nonrandomized, open-label, pilot study. The primary outcome measure was logMAR visual acuity. Secondary outcome measures were binocular indirect ophthalmoscopy score, cystoid macular edema, adverse effects, and vision-related (visual core module 1) and health-related (36-Item Short-Form Health Survey) quality of life. RESULTS: Within 1 month of TNFr-Ig therapy, 9 patients (53%) achieved at least a 2-line improvement in visual acuity, 8 (57%) of 14 patients with vitreous haze before treatment achieved an improvement in binocular indirect ophthalmoscopy score to 0, and macular edema resolved in 5 (56%) of 9 affected patients. Twelve (71%) of the patients achieved complete cessation of intraocular inflammation following TNFr-Ig therapy. A reduction in concomitant immunosuppression was possible for 11 patients (65%) following TNFr-Ig therapy. However, all but 1 patient required continuing adjuvant therapy during the response to TNFr-Ig, which had a median duration of 3 months. Adverse effects included mild infusion reactions in 3 patients and transient lymphocytopenia in 2 patients. CONCLUSION: Therapy with TNFr-Ig was safe and effective for treating patients with sight-threatening noninfectious posterior segment intraocular inflammation resistant to conventional immunotherapy, but adjuvant immunosuppression and repeat infusions would be required to maintain long-term remission.

Blood concentrations of alemtuzumab and antiglobulin responses in patients with chronic lymphocytic leukemia following intravenous or subcutaneous routes of administration.

Alemtuzumab is a humanized anti-CD52 antibody licensed for refractory B-cell chronic lymphocytic leukemia (B-CLL), when given intravenously at 30 mg thrice weekly. However, the intravenous route is associated with infusion-related reactions and is inconvenient. We measured blood concentrations in 30 relapsed patients treated with intravenous alemtuzumab and in 20 patients from a previously untreated group who received similar doses subcutaneously. Highest trough samples in the intravenous group were less than 0.5 microg/mL to 18.3 microg/mL (mean 5.4 microg/mL). The cumulative dose required to reach 1.0 microg/mL was 13 mg to 316 mg (mean 90 mg). Higher blood concentrations correlated with the achievement of better clinical responses and minimal residual disease. The highest measured concentrations in the subcutaneous group were similar (0.6 microg/mL to 24.8 microg/mL, mean 5.4 microg/mL). However, the cumulative dose to reach 1.0 microg/mL was higher: 146 mg to 1106 mg (mean 551 mg). No antiglobulin responses were detected in 30 patients given intravenous alemtuzumab whereas 2 of 32 patients given subcutaneous alemtuzumab made substantial anti-idiotype responses. Thus, subcutaneous alemtuzumab achieved concentrations similar to those for intravenous alemtuzumab, although with slightly higher cumulative doses. Subcutaneous alemtuzumab is more convenient and better tolerated but may be associated with some patients forming anti-alemtuzumab antibodies, particularly those patients who were previously untreated.

Pharmacokinetics of alemtuzumab used for in vivo and in vitro T-cell depletion in allogeneic transplantations: relevance for early adoptive immunotherapy and infectious complications.

Persistence of alemtuzumab at lympholytic concentrations after reduced-intensity conditioning allogeneic stem cell transplantations (RITs) could impair immune reconstitution and reduce donor T-cell-mediated graft-versus-leukemia/lymphoma (GVL) effects, derived from the graft or subsequent adoptive immunotherapy. We have studied the pharmacokinetics of alemtuzumab in 2 different groups: RIT (100 mg alemtuzumab in vivo over 5 days) and myeloablative allografts (20 mg alemtuzumab added in vitro to the stem cells prior to return). Alemtuzumab concentrations in RIT patients were in excess of that required to kill infused donor CD52+ cells at the time of transplantation and remained at potentially lympholytic levels (> 0.1 microg/mL) for approximately 56 days after transplantation, 26 days longer than for the myeloablative group. Total lymphocyte counts were significantly lower in the RIT group persisting beyond 6 months after transplantation (P =.005), and median absolute CD4 counts higher than 200 x 106/L were delayed until 9 months after transplantation.

Phase II trial of subcutaneous anti-CD52 monoclonal antibody alemtuzumab (Campath-1H) as first-line treatment for patients with B-cell chronic lymphocytic leukemia (B-CLL).

This phase II study determined the efficacy and safety of alemtuzumab, a humanized anti-CD52 monoclonal antibody, delivered subcutaneously as first-line therapy, over a prolonged treatment period of 18 weeks in 41 patients with symptomatic B-cell chronic lymphocytic leukemia (B-CLL). Injections were administered subcutaneously 3 times per week, from week 2 to 3 onward. An overall response rate (OR) of 87% (95% CI, 76%-98%; complete remission [CR], 19%; partial remission [PR], 68%) was achieved in 38 evaluable patients (81% of intent-to-treat population). CLL cells were cleared from blood in 95% patients in a median time of 21 days. CR or nodular PR in the bone marrow was achieved in 66% of the patients and most patients achieved this after 18 weeks of treatment. An 87% OR (29% CR) was achieved in the lymph nodes. The median time to treatment failure has not yet been reached (18+ months; range, 8-44+ months). Transient injection site skin reactions were seen in 90% of patients. Rigor, rash, nausea, dyspnea, and hypotension were rare or absent. Transient grade IV neutropenia developed in 21% of the patients. Infections were rare, but 10% patients developed cytomegalovirus (CMV) reactivation. These patients rapidly responded to intravenous ganciclovir. One patient, allergic to cotrimoxazole prophylaxis, developed Pneumocystis carinii pneumonia. Alemtuzumab is highly effective as first-line treatment in patients with B-CLL. Prolonged treatment is important for maximal bone marrow response. Subcutaneous administration induced very few "first-dose" flulike symptoms and may reduce health care costs in comparison with the intravenous infusions.

Pharmacokinetics of CAMPATH-1H: assay development and validation.

CAMPATH-1H is a humanised monoclonal antibody against the CD52 antigen which is being developed for treatment of chronic lymphocytic leukaemia (CLL), autoimmune disease and prevention of transplant rejection. Measurement of antibody serum levels is important for optimising dose regimens but difficult owing to the low concentration compared with normal human IgG. After consideration of various methods, a suitable assay was developed based on indirect immunofluorescence. Test samples were incubated with target cells (HUT-78, a human T cell line) and the CAMPATH-1H was detected by binding of a fluorescent-labelled anti-human Ig using a flow cytometer. Robustness of the assay was demonstrated under a range of experimental conditions. Because of the low affinity of CAMPATH-1H, only a weak signal was seen at low concentrations. The limit of detection was 0.15 microg/ml and the limit of quantitation was 0.25 microg/ml. Since serum samples were diluted at least 1:2, the lowest concentration which can be measured in patient serum was 0.5 microg/ml. The overall precision (coefficient of variation) was +/-13% and the overall accuracy (bias) was +9%. There was a low incidence of false-positive results (<2%) in normal or pre-treatment patient serum. Quantitative recovery was obtained from serum samples spiked with CAMPATH-1H and stored under a variety of conditions, including being treated at 56 degrees C for 30 min and frozen and thawed up to four times. This validated assay is suitable for the measurement of CAMPATH-1H levels in clinical trials and the same principles may be applied to any other cell-binding monoclonal antibody.