The tetraspanin CD37 is widely expressed in B-cell malignancies and represents an attractive target for immunotherapy with mAbs. We have chimerized a high-affinity mouse Ab to CD37 and engineered the CH2 domain for improved binding to human

The DNA damage pathway plays a central role in chemoresistance in chronic lymphocytic leukemia (CLL), as indicated by the prognostic impact of TP53 and ATM loss/ mutations. We investigated the function of the p53 axis in primary CLL samples by studying p53 and p21 responses to irradiation by FACS and RT-PCR. We observed a distinct response pattern for most cases with a 17p deletion (n ‫؍‬ 16) or a sole TP53 mutation (n ‫؍‬ 8), but not all cases with a p53 aberration were detected based on a number of different assays used. Samples with a small clone with a TP53 mutation remained undetected in all assays. Only 1 of 123 cases showed high expression of p53, which is suggestive of p53 aberration without proof of mutation of TP53. Samples with an 11q deletion showed a heterogeneous response, with only 13 of 30 showing an abnormal response based on cutoff. Nevertheless, the overall induction of p53 and p21 was impaired, suggesting a gene-dosage effect for ATM in the 11q-deleted samples. The detectability of p53 defects is influenced by clonal heterogeneity and sample purity. Functional assays of p53 defects will detect a small number of cases not detectable by FISH or TP53 mutational analysis. The clinical utility of functional p53 testing will need to be derived from clinical trials. (Blood. 2011;117(5): 1622-1632)

Purpose BAL101553, the prodrug of the microtubule-destabilizer BAL27862, previously showed signs of antitumor activity when administered as a 2-h infusion, but its use was limited by vascular toxicity. We investigated an alternative dosing strategy aimed at improving the safety profile of BAL101553. Methods This multicenter, open-label, Phase 1 dose-escalation study used a 3 + 3 design to determine the maximum tolerated dose (MTD), dose-limiting toxicities (DLTs), pharmacokinetics, and antitumor activity of BAL101553 administered as a 48-h IV infusion on Days 1, 8, and 15 of a 28-day cycle. Patients received oral BAL101553 on Days 15-21 of cycle 2 to assess oral bioavailability. Results BAL101553 was well tolerated at doses up to ≤70 mg/m 2. Three grade 3 DLTs occurred: hypotension (70 mg/m 2), hyponatremia and neutropenia (both 90 mg/m 2). The MTD for 48-h IV BAL101553 was 70 mg/m 2. At this dose level, the AUC for BAL27862 was 8580 ng.h/mL and the C max was 144 ng/ mL. No apparent dose-related effects on blood pressure were observed with 48-h BAL101553 IV infusion. BAL27862 oral bioavailability was >80%. Conclusions Continuous 48-h IV BAL101553 infusion achieved higher exposure of the BAL27862 active metabolite than a 2-h infusion at the RP2D and did not cause vascular toxicity. Clinicaltrials.gov registration: NCT02895360.

2379 Poster Board II-356 Introduction: Anti-CD20 (i.e. rituximab) based immuno-chemotherapies are now considered standard of care in CLL. However, addition of rituximab does not appear to decisively alter the dismal clinical outcome for 17p- / TP53 mutated CLL. Relatively little is known about activity of different anti-CD20 antibodies in genetic subgroups of CLL. Methods: In an effort to assess the activity of the next generation mAb GA101 in vitro, we studied B cell depletion / apoptosis in a set of CLL patients. CLL samples were genetically characterized with respect to genetics (genomic aberrations, TP53 mutation, IGHV mutation status), as well as clinical course and immunophenotype. To study the effect on CLL cells, we assessed GA101, Rituximab and Alemtuzumab by in vitro treatment after ficoll separation with FACS (7-AAD) for viability. In addition, in order to maintain ADCC and to mirror the in vivo situation, we studied the effect of GA101 in a whole blood culture (n=10). Results: With increasing concentrations (0.01-100mg/ml) of GA101 we observed significant cell death as measured by a B cell depletion assay (whole blood) after GA101 exposure (n=10). After 3 hours the maximum effect was observed at a concentration of 10mg/ml with a reduction of CLL cells to 43% of untreated cells. The results after 8h were similar, again showing the profound effect in GA101 treated samples (mean B-cell depletion to 34.6% remaining cells). Interestingly, further increasing the dose to 100mg/ml did not increase the B-cell depletion by the antibody, but appeared to decrease the ability to deplete CLL cells (Fig. 1). In contrast treatment with control antibodies (isotype control, rituximab, alemtuzumab) did not lead to similar B-cell depletion (101%, 69%, and 73% of control cells at 3h). We were particularly interested in any potential effect in cases with TP53 mutation. While the sample number is still low (TP53 mutation / 17p deletion n=3), currently we have no indication of differential response in different genetic subgroups. Conclusion: Compared to the currently most widely used mAb in CLL (Rituximab), GA101 appears more potent at equivalent concentration in depleting CLL cells. Further in vitro studies are currently ongoing to assess this antibody in particular in genetic high-risk, refractory CLL. Disclosures: Zenz: Roche: Honoraria. Klein:Roche: Employment, Equity Ownership, Patents & Royalties. Umana:Roche: Employment, Equity Ownership, Patents & Royalties. Döhner:Roche: Research Funding. Stilgenbauer:Roche, Bayer Schering Pharma: Honoraria, Research Funding.

2460 Introduction: Anti-CD20 (rituximab) based immuno-chemotherapies are considered standard of care in CLL. However, rituximab as single agent has limited activity in CLL. In addition, Rituximab does not appear to decisively alter the clinical outcome for 17p- / TP53 mutated CLL. CD20 is expressed at low levels in CLL and alternative targets for antibody-based treatment have been explored. CD37, a member of the tetraspanin superfamily, is a heavily glycosylated cell surface molecule with four transmembrane domains and two extracellular loops. CD37 is almost exclusively expressed on mature B cells, with highest expression levels on peripheral blood B cells, reduced levels on plasma cells and non- detectable levels on CD 10+ precursor B cells in the bone marrow. Lower level expression of CD37 has also been reported on T cells, granulocytes, and monocytes. Methods: In an effort to assess the activity of two CD37 antibodies in CLL, we studied B cell depletion / apoptosis in a set of CLL patients in vitro. MAb 37.1 is a chimeric IgG1-type of antibody which is Fc-engineered to improve ADCC activity. MAb 37.2 is a humanized version of mAb 37.1. CLL samples were characterized with respect to genetics (genomic aberrations, TP53 mutation, IGHV mutation status), as well as clinical course and immunophenotype. To study the effect on CLL cells, we assessed the CD37 antibodies in parallel with Rituximab and Alemtuzumab in a whole blood culture system (n=21). In addition, the effect of the antibodies was assessed after in vitro treatment in cell culture by FACS (7-AAD+Annexin). Results: With increasing concentrations (0.01-100μg/ml) of mAb 37.1 and mAb 37.2 we found effective CLL cell depletion in the whole blood assay. As shown in figure 1 CLL cell depletion was observed with 1μg/ml mAb 37.1 and 37.2 at 3h and 8h. The comparison of mAb 37.1 and mAb 37.2 showed higher CLL cell depletion with mAb 37.1 (1μg/ml mAb 37.1 3h: 52.2% alive cells vs. mAb 37.2 3h: 71.5%). Maximum CLL cell depletion was observed at 10μg/ml (23.1% alive cells (mAb 37.1) vs. 46.7% alive cells (mAb 37.2)). Incubation with mAb 37.1 and mAb 37.2 consistently led to homotypic aggregation. Both CD37 Abs were significantly more effective in depleting CLL cells than Alemtuzumab (89.4% alive cells at 3h (10μg/ml)(p<0.001)) and Rituximab (94.8% alive cells 3h (10μg/ml)(p<0.001)). After 8h of incubation with mAb 37.1 and mAb 37.2 CLL cell depletion was highly efficacious with 15.4% and 37.1% remaining B-cells. This compared to moderate CLL depletion in this whole blood assay with Campath and Rituximab (85% and 92% remaining CLL cells respectively). We were particularly interested in the effect in cases with TP53 mutation or deletion and fludarabine refractory cases. To this end we compared the CLL cell depletion in this high risk group (n=10) and the remaining patients (n=11). For both antibodies the CLL cell depletion was highly effective in low and high risk groups (i.e. 10μg/ml mAb 37.1 mean remaining CLL cells: 21.3% (8h) high risk; 11.1% low risk; 10μg/ml mAb 37.2 mean remaining CLL cells: 43.1% (8h) high risk; 33.2% low risk). Conclusion: Both CD37 antibodies studied showed highly effective CLL cell depletion irrespective of genetic risk category. Our data suggest that CD37 antibodies could be more effective than CD20 or CD52 antibodies in CLL. CD37 Abs should be investigated in clinical trials. Disclosures: Zenz: Boehringer: Honoraria. Heider: Boehringer Ingelheim: Employment. Reichardt: Boehringer Ingelheim: Employment. Stilgenbauer: Boehringer, Roche, Celgene, GSK,: Honoraria, Research Funding.