ALBERT

Description: Background. In 2005 we analyzed the results of a prospective randomized phase III intergroup trial evaluating the role of rituximab (R) both in remission induction and maintenance treatment of 465 relapsed/resistant follicular lymphoma (FL) patients. Major conclusions were that addition of R to CHOP induction yielded an increased ORR and CR rate, and that R maintenance strongly improved median progression free survival (PFS), both after induction with CHOP and R-CHOP, and overall survival (OS (van Oers et al Blood2006;108:3295). At that time the median follow for the maintenance phase was 33 months. Now we report the long-term outcome of maintenance treatment, with a median follow up of 6 years from start of maintenance. Study design. Patients with stages III or IV FL at initial diagnosis and relapsed after or resistant to a maximum of two non-anthracycline containing systemic chemotherapy regimens, were randomized to remission induction with either 6 cycles of standard CHOP (once every 3 weeks) or CHOP + R (375 mg/m2 at day 1 of each cycle of CHOP). Those with a complete or partial remission after 6 cycles of therapy underwent a second randomization to no further treatment (observation) or maintenance treatment with R (375 mg/m2 once every 3 months) until relapse or for a maximum period of two years. Results. 465 patients were randomized to induction with either CHOP (231) or R-CHOP (234). As reported, CHOP and R-CHOP induction yielded similar partial response rates (57% vs.56%), but significantly different CR rates (16% and 29%; p=0.0001). 334 patients were randomized to either R maintenance treatment (167) or observation (167). R maintenance resulted in a highly significant improvement of PFS: median 3.7 years versus 1.3 years in the observation arm (p

Description: Introduction Achieving a PET-negative metabolic (m)CR with salvage chemotherapy prior to autologous stem cell transplantation (ASCT) in relapsed/refractory Hodgkin lymphoma (R/R HL) is a strong predictive factor for long-term progression-free survival (PFS). In a phase I dose-escalation trial we have shown a favorable toxicity profile of 3 cycles of DHAP in combination with brentuximab vedotin (BV) with a high mCR rate (12/12 patients). A subsequent phase II study in 55 patients has been performed; the 6 patients treated at full dose of BV-DHAP in the phase I part were added to the current analysis. Methods BV was given at a fixed dose of 1.8 mg/kg on day (d) 1 of 3 cycles of DHAP q 3 weeks (dexamethasone 40 mg iv d 1-4, cisplatin 100 mg/m² iv d1 and cytarabine 2x2 g/m² iv d2). Cycle 2 was used for stem cell mobilization; after the 1st and 3rd cycle patients received pegylated G-CSF to prevent prolonged neutropenia and dose delays. The primary endpoint of the study was the mCR rate after 3 cycles of BV-DHAP based on central PET-review. Patients with a partial or complete response proceeded to high dose chemotherapy (BEAM) and ASCT. Results Twenty-three of the 61 patients (29 males; median age 29 yrs, range 19-71) had not achieved a CR to 1st line treatment (37%), which consisted of ABVD (n=45), (escalated) BEACOPP (n=11) or other regimens (n=5). The median time from response to 1st line treatment to relapse was 6 months (range 0-160 months); 38 patients (62%) had either primary refractory disease or early relapse (

Description: Ofatumumab is a unique monoclonal antibody that targets a distinct small loop epitope on the CD20 molecule. Preclinical data show that ofatumumab is active against B-cell lymphoma/chronic lymphocytic leukemia cells with low CD20-antigen density and high expression of complement inhibitory molecules. In a phase 1/2 trial evaluating safety and efficacy of ofatumumab in relapsed or refractory follicular non-Hodgkin lymphoma (FL) grade 1 or 2, 4 dose groups of 10 patients received 4 weekly infusions of 300, 500, 700, or 1000 mg. Patients had a median of 2 prior FL therapies and 13% had elevated lactate dehydrogenase. No safety concerns or maximum tolerated dose was identified. A total of 274 adverse events were reported; 190 were judged related to ofatumumab, most occurring on the first infusion day with Common Terminology Criteria grade 1 or 2. Eight related events were grade 3. Treatment caused immediate and profound B-cell depletion, and 65% of patients reverted to negative BCL2 status. Clinical response rates ranged from 20% to 63%. Median time to progression for all patients/responders was 8.8/32.6 months, and median duration of response was 29.9 months at a median/maximum follow-up of 9.2/38.6 months. Ofatumumab is currently being evaluated in patients with rituximab-refractory FL. This trial was registered at www.clinicaltrials.gov as #NCT00092274.

Description: HuMax-CD20 is a fully human monoclonal IgG1 antibody targeting a unique extracellular epitope of the CD20 molecule on B-cells. HuMax-CD20 stops growth of engrafted B-cell tumors in SCID mouse tumor models more efficiently than Rituximab®, and i.v. infusion of HuMax-CD20 in cynomolgus monkeys has led to profound, long lasting, dose-dependent B-cell depletion. A total of 40 patients with CD20+ relapsed or refractory follicular non-Hodgkin’s lymphoma grade I-II will be enrolled in this open-label, dose-escalating, international, multi-center clinical trial. Cohorts of 10 patients will receive i.v. infusions at doses of either 300, 500, 700 or 1000 mg once weekly for 4 weeks. The patients are followed for 12 months. Patients receive oral acetaminophen and i.v. antihistamin before infusion. In case of adverse events of CTC grade 3 or higher, i.v. glucocorticosteroids are given. The endpoints are CT scan verified tumor response according to the Cheson criteria, B-cell depletion in peripheral blood and lymph nodes, time to next anti-lymphoma treatment, duration of response, BCL2 conversion, pharmacokinetics, and adverse events. Tumor and bone marrow biopsies and CT scans are assessed centrally. The first 17 patients treated with HuMax-CD20 are the subject of this report. Mean age is 60 years. In the 300 mg group all 10 patients have received all 4 infusions. Seven patients have been enrolled in the 500 mg group; three of them have received 4 infusions, two have received 3 infusions, and two patients have received 2 infusions. Baseline B-cell count was in the range of 11-382 x 106 cells per L with a median of 114 x 106. One week after the first infusion the median B-cell count available in 16 patients was 8 x 106 cells per L with a range of 0–19 x 106. In six of the 16 patients no B-cells were detected. B-cell counts measured one week after the 4th infusion are available for 10 patients. Eight patients had no detectable B-cells, one patient had 11 x 106 and one had 34 x 106 cells per L. B-cell counts eight weeks after the 4th infusion are available for two patients. No B-cells were detectable in these two patients. No dose limiting toxicity has been reported with administration of 300 or 500 mg. One serious adverse event assessed as not related to HuMax-CD20 has been reported in the 300 mg group. Infusion related adverse events have primarily been seen during the first infusion of HuMax-CD20. The events have, as expected, predominantly been signs and symptoms of cytokine release, e.g. pruritus, dyspnoea, rigors/chills, nausea, hypotension, urticaria, fatigue, fever and rash. In 15 of the 17 patients, 51 adverse events have been reported. Nine adverse events were CTC grade 3, 16 were grade 2, and 26 events were grade 1. In conclusion, this analysis based on preliminary data for the first 17 patients treated with HuMax-CD20 demonstrated significant depletion of peripheral blood B-cells and a favorable safety profile. An updated report of results for all 40 patients including preliminary tumor response data will be presented.

Description: A promising strategy to control Graft versus Host disease (GvHD) in allogeneic stem cell transplantation is the use of donor T cells, genetically modified to express the Herpes Simplex Thymidine Kinase (HSVtk) gene. The HSVtk+ donor T cells can be selectively eliminated by ganciclovir (GCV) in vivo. Despite the encouraging clinical results limitations have been observed with this system. The viral HSVtk gene is immunogenic, GCV-activity is limited to dividing cells, and aberrant splicing of the HSVtk mRNA occurs. The human CD20 molecule, in combination with the humanized CD20 Ab Rituximab (RTX), was recently proposed as a novel ‘suicide’ system (Introna et al., Hum Gene Ther 2000). CD20 is non-immunogenic and both resting and dividing CD20+ cells can be eliminated in vivo by RTX, which triggers the complement system and recruits effector cells. However, for a safe and effective clinical application a high, stable and homogenous CD20 expression by human T cells is required. We constructed different CD20-encoding retroviral vectors. To enhance CD20 expression we included the WPRE element, which is widely used to increase transgene expression. Chromatin insulators separate differentially regulated loci, shield promoters from neighboring regulatory elements and protect against transcriptional silencing. Thus, insulators may protect against insertional mutagenesis and provide a more homogenous and stable expression of the transgene. We therefore included insulators into the vectors. For this, CEM T cells were transduced with the different CD20-vectors at an MOI of 1 and a mean transduction efficiency of 22.9±5.1% was obtained. Results showed a significantly more homogenous CD20 expression for cells containing the insulators. Also after CD20 magnetic bead selection the CD20+ insulator containing cells had a more homogenous CD20 expression pattern. Surprisingly, the CD20 expression pattern became 2-fold more homogenous after continuous culturing for 3 months. In contrast, 60% of the cells without insulators lost CD20 expression. Inclusion of the WPRE element increased the CD20 expression level. CD20-vectors containing both the insulator and the WPRE element gave the highest, most homogenous and stable CD20 expression in CEM T cells. In addition, we generated individual CEM-CD20 clones with different CD20 expression levels, to analyze the efficacy of killing by RTX in a complement dependent cytotoxicity (CDC), antibody-dependent cellular cytotoxicity (ADCC) and a combination of both assays. The CDC assay showed that RTX-mediated kill depends on the level of CD20 expression. We observed a clear, significant correlation between number of CD20 molecules per cells and the extent of CDC (p

Description: Abstract 716 In autoimmune hemolytic anemia (AIHA) autoantibodies (auto-Ab's) to red blood cells (RBCs) may induce complement activation via the classical pathway of complement resulting in complement deposition on RBCs. RBCs coated with auto-Ab and/or complement (C3b) are removed by extravascular destruction in the spleen and liver. In a minority of cases complement activation proceeds until insertion of the membrane attack complex (MAC) resulting in intravascular hemolysis. The inflammatory response and the anoxia induced by hemolysis significantly contribute to the morbidity and fatality in AIHA. However, the recovery of RBC transfusion in these situations is inadequate since auto-Abs to RBC also react with donor cells leading to rapid destruction of RBC. Activation of classical pathway of complement is critically involved in this process. C1-inhibitor (Cetor) is a plasma-derived inhibitor of the classical pathway of complement with an excellent safety profile. We hypothesized that co-administration of C1-inhibitor concentrate with a RBC transfusion improves recovery of a transfusion by attenuation of the activation of the classical pathway of complement. In order to test this hypothesis we adapted a standard haemolytic assay (CH50). Sera from patients (n=6) with AIHA due to IgM antibodies with reactivity above 30°C induced dose-dependent lysis of bromealin-treated RBC, whereas sera from healthy donors (n=4) did not. Addition of C1-inhibitor concentrate dose-dependently inhibited RBC lysis significantly in this model. Subsequently, by flowcytometry we studied C3 deposition on RBC after incubation with AIHA patient sera or sera from healthy controls. Anti-C5 antibody (eculizumab) was added in order to prevent lysis during analysis. AIHA sera containing eculizumab induced extensive C3 deposition on RBC, which was significantly inhibited by C1-inhibitor concentrate. This inhibition was dose-dependent. Next, we were interested whether these in-vitro effects could also be observed in-vivo. A 65 year old patient suffering from fulminant life threatening hemolysis due to IgM type auto-Abs reactive 〉30°C was admitted to our department. Despite treatment with anti-CD20, vincristine and high-dose methylprednisolone the anaemia did not improve necessitating transfusion of 3 RBC concentrates. However, 2.5 day after transfusion the starting haemoglobin levels was reached again and hemolysis deteriorated. Since the patient became symptomatic again the next transfusion of 3 RBC concentrates was preceded by infusion of 6000 U of C1-inhibitor. Three additional donations of 4000 U, 2000 U and 1000 U of C1-inhibitor concentrate have been administered 12, 24 and 36 hours after the first dose, respectively. The recovery of the transfusion significantly increased and was accompanied by a decrease in C3 deposition on RBC, as evidenced by an attenuation of the direct antiglobulin test. In conclusion we have demonstrated that in-vitro C1-inhibitor concentrate can efficiently inhibit RBC lysis and C3 deposition on RBC induced by sera from patients with AIHA. Moreover we have shown that this approach is also effecitive in-vivo. Therefore, in AIHA administration of C1-inhibitor concentrate might be life-saving by improving recovery of RBC transfusion by inhibition of both intra- and extravascular destruction of recipient and donor RBCs. Disclosures: Off Label Use: C1-inhibitor-inhibition of complement-induced hemolysis.