ALBERT

Beschreibung: GA101 is a novel monoclonal antibody of IgG1 type which binds with high affinity and selectivity to the extracellular domain of the human CD20 antigen on B cells. In contrast to rituximab which is a chimeric antibody and recognizes a type I epitope, GA101 is humanized and recognizes a type II epitope which is also localized in the extracellular loop of CD20. The recognition of the type II epitope together with a modification of the elbow hinge region results in enhanced direct non-caspase dependent cell death induction, and concomitant reduction in CDC upon binding to CD20. In addition, using GlycoMab technology, the Fc-region of GA101 was glycoengineered to contain bisected, afucosylated carbohydrates. As a result GA101 has increased affinity for the low and high affinity FcγRIIIa receptor expressed on natural killer cells, macrophages and monocytes. Consequently, GA101 mediated a 5–50 fold enhanced induction of effector cell mediated ADCC. In B-cell depletion assays with whole blood from healthy donors, an assay combining all mechanisms of action, GA101 was significantly more potent and efficacious in depleting B cells than rituximab. In preclinical NHL testing these properties translated into superior anti-tumoral efficacy of GA101 in direct comparison to rituximab against a number of aggressive NHL xenograft models. In cynomolgus monkeys the induction of B cell depletion mediated by GA101 and subsequent B cell recovery were investigated. GA101 induced complete, rapid and long-lasting B cell depletion both in peripheral blood and in lymphoid tissue e.g. spleen and lymph nodes. The efficacy of GA101 (10 and 30 mg/kg) at depleting B cells in different lymphoid tissues of cynomolgus monkeys was compared with that of rituximab (10 mg/kg) following 2 i.v. doses administered on days 0 and 7. Notably, GA101 showed statistically superior depletion of total B cells from lymph nodes compared to Rituximab from day 9 to 35 onwards with B cell numbers decreased by over 95%. These results demonstrated that GA101 was more efficacious at depleting B cells from lymph nodes and spleen of cynomolgus monkeys compared to rituximab. Compared to existing antibodies, GA101 constitutes the first type II CD20 antibody engineered for increased ADCC with significantly enhanced efficacy in a variety of preclinical models. Based on these data it is assumed that the combination of the recognition of a type II epitope together with improved ADCC potency might translate into superior efficacy in the clinical treatment of CD20 positive malignant diseases.

Beschreibung: Background & Aim: T-cell bispecific antibodies (TCBs) binding to a target on tumor cells and CD3 on T cells induce potent T-cell mediated killing of cells carrying the target. In contrast to targets like e.g. CD38 or CD138, B-cell maturation antigen (BCMA) is suggested to be only expressed on plasma cells (PCs) and multiple myeloma (MM) PCs. Therefore, a BCMA-TCB should specifically act on these cell types. We report on a new class of BCMA-TCBs designed for effective and convenient therapy of MM. Molecular structure & its rationale (fig. 1A): The new class of BCMA-TCBs are asymmetric two-arm IgG-based human antibodies, bivalent to BCMA, monovalent to CD3 and comprising an engineered Fc. To achieve tumor specificity of the BCMA-TCBs and avoid unspecific T-cell activation, monovalent binding to CD3 was fixed at an affinity of KD= 70 nM, whereas various BCMA-TCBs with binding affinities to BCMA ranging from KD= 50 pM to 10 nM have been investigated (measurement of binding affinities by surface plasmon resonance and flow cytometry (FC)). For long elimination half-life, an Fc was introduced to enable once a week intravenous or subcutaneous administration. Fc is engineered to abolish binding to FcgR and C1q to minimize risk of infusion reactions without interfering with FcRn binding. The BCMA-TCBs can be well-manufactured and have no tendency to aggregation. In vitro profile: Potency of BCMA-TCBs to activate T cells and to induce killing of human MM cell lines was measured in a 24h co-culture assay with human PBMCs and MM cells in a ratio of 10:1. Results: (i) NCI-H929 cells expressing on cell surface up to 100 times more BCMA than primary patient MM cells were killed in presence of BCMA-TCBs with EC50 ranging from 5 to 50 pM, but not in presence of a control-TCB binding to CD3 only. (ii) As next, RPMI-8226 cells were used as target cells because surface BCMA expression was found to be only slightly higher than on primary patient MM cells (specific antigen binding capacity SABC as measured by FC between 1165 and 5461 per RPMI-8226 cell compared to 116 to 4479 per primary patient MM cell). Effective killing of RPMI-8226 MM cells was observed; the killing potency was higher respectively EC50 lower with the BCMA-TCBs having binding affinities below 1 nM (fig. 1B; EC50 from 50 pM to 1000 pM). (iii) Killing of U266 and L363 human MM cell lines was also observed with BCMA-TCBs. (iv) T-cell activation and increased T-cell function go in parallel with lysis of MM target cells as observed by an upregulation of CD69 and CD25 expression, release of granzyme B (〉20 ng/mL at 3 nM vs. 20 pg/mL for control) and proinflammatory cytokines e.g. IFN-g, TNF-a, IL-2, IL-6, and IL-10. (v) At 120h incubation, BCMA-TCBs induced concentration dependant CD4 and CD8 T-cell proliferation as observed by CFSE dilution. A small exploratory study in whole bone marrow (BM) aspirates from MM patients (n=3) suggested a concentration dependent killing of MM PCs induced by BCMA-TCBs in presence of autologous T cells, thus justifying for a much larger trial to investigate one of the BCMA-TCB of this new class to induce killing and T-cell activation/function in MM patient BM aspirates performed by two clinical groups (Abstract also submitted to ASH). In vivo profile: New BCMA-TCBs bind to BCMA as well as CD3 of cynomolgus monkeys (cyno). Single dose pharmacokinetics (PK) and pharmacodynamics of a BCMA-TCB was studied in cyno at 0.003, 0.03, 0.3 mg/kg intravenously. Dose linear PK was observed. Mean serum concentrations in the first 7 days after 0.03 mg/kg were ≈1 to 2 nM, BM aspirates collected at 96h showed also concentrations of ≈1 to 2 nM. Elimination between 24 h and 504 h was found to be first order with a half-life of approximately 6 to 8 days. These data suggest a convenient clinical dosing schedule, e.g. once a week. Peripheral blood T-cell redistribution was observed 24h after dosing. Reduction of PCs could be observed by FC while total B cells and other cell types were unaffected. Summary: Effective and specific killing of MM cells was demonstrated with the BCMA-TCBs. Killing goes in parallel with T-cell activation and increased function. TCB with binding affinity to BCMA below 1 nM have higher potency/lower EC50 than those with affinity above 1 nM. TCBs could be produced with high purity and were stable with no tendency to aggregation. In cynomolgus monkeys, a PK profile has been found, suitable for once a week administration. This new class of BCMA-TCB has promises for clinical development. Disclosures Vu: EngMab AG: Employment, Equity Ownership, Membership on an entity's Board of Directors or advisory committees. Moser:Roche: Employment. Delon:Roche: Employment. Latzko:Roche: Employment. Gianotti:Roche: Employment. Lüoend:Roche: Employment. Friang:Roche: Employment. Murr:Roche: Employment. Duerner:Roche: Employment. Weinzierl:Roche: Employment. Fauti:Roche: Employment. Bacac:Roche: Employment. Ast:Roche: Employment. Freimoser-Grundschober:Roche: Employment. Rodriguez Diaz:Roche: Employment. Zielonka:Roche: Employment. van Puijenbroek:Roche: Employment. Hosse:Roche: Employment. Bruenker:Roche: Employment. Mössner:Roche: Employment. Klein:Roche: Employment. Umaña:Roche: Employment. Strein:EngMab AG: Employment, Equity Ownership, Membership on an entity's Board of Directors or advisory committees; BB Biotech AG: Membership on an entity's Board of Directors or advisory committees; Novimmune SA: Membership on an entity's Board of Directors or advisory committees.

Beschreibung: Despite the recent advancements in treatment options with the introduction of anti-CD20 monoclonal antibody therapy, approximately 50% of patients with non-Hodgkin's lymphomas (NHL) will not sustain a durable response to standard of care (SOC) treatment. Thus, there remains a continuous need for safer and more effective anti-cancer therapies in this indication. T-cell bispecific antibodies (TCBs) represent a new class of disease targeting agents shown to promote the activation of a patient's own T cells to attack and kill cancer cells. CD20 TCB is a new bispecific antibody with IgG-like pharmacokinetic properties whose unique "2:1" structure leads to increased tumor antigen avidity, T cell activation, and tumor cell killing, as compared to other T cell engaging bispecific antibody molecular formats. The molecule comprises two CD20 binding Fabs (derived from the Type II CD20 IgG1 obinutuzumab), one CD3e binding Fab (fused to one of the CD20 Fabs via a short flexible linker), and an engineered, heterodimeric Fc region with completely abolished binding to FcgRs and C1q. In vitro, CD20 TCB was shown to dose-dependently induce tumor lysis with EC50 values in the range of 0.05 - 3.1 pM. The "2:1"format of CD20 TCB was shown to confer superior potency (up to 10 - 1000x) when compared to CD20 TCBs having the conventional "1:1" IgG-based format (i.e., one binding domain for CD20 and one for CD3). CD20 TCB-mediated tumor lysis resulted in T-cell activation, proliferation and cytokine release with up-regulation of programmed death 1 (PD-1)/programmed death ligand 1 (PD-L1) axis upon tumor lysis. CD20 TCB also demonstrated potent ex vivo activity in whole bone marrow aspirate samples of NHL and CLL patients (n=17). Such primary tumor samples preserve the native tumor microenvironment and bear low effector to target cell ratios ranging in this study from 0.02 to 0.8 (average value 0.3). CD20 TCB activity was consistently superior to that of the "1:1" CD20 TCB and demonstrated faster, more profound and more potent B cell depletion with EC50 values ranging from 0.002 to 2.7 nM. In vivo, CD20 TCB displayed potent anti-tumor activity in xenograft models in stem cell humanized mice and induced regression of large, aggressive WSU-DLCL2 lymphoma tumors (0.5 mg/kg, weekly administration). In addition to tumor regression, CD20 TCB treatment led to fast and complete elimination of peripheral blood B cells within 24 h after the first administration (0.05, 0.15 and 0.5 mg/kg, weekly administration) and to a complete elimination of B cells in spleen, bone marrow and lymph nodes after two administrations. B cell depletion was paralleled by transient decrease of T-cell counts in the peripheral blood and by the peak of cytokine release 24 h after the first administration, followed by rapid recovery and return to baseline levels at 72 h post treatment. Tumor growth inhibition mediated by CD20 TCB was accompanied by increase in intra-tumor T-cell infiltration, up-regulation of PD-1 receptor on T cells and PD-L1 in the tumor. Combination studies of CD20 TCB with PD-L1 blocking antibody led to more profound and faster tumor growth inhibition. Taken together, the preclinical data show that CD20 TCB is a novel differentiated CD20-targeting T cell bispecific antibody with promising anti-tumor activity and the ability to modify the tumor microenvironment. CD20 TCB consistently demonstrated superior potency compared to other CD20 TCBs with a conventional "1:1" IgG format. This translated into superior efficacy in vitro, ex-vivo and in vivo, which could not be matched by increasing doses of the "1:1" TCBs. The molecule is now scheduled to start clinical trial by December 2016. Disclosures Bacac: Roche: Employment, Equity Ownership, Patents & Royalties. Umaña:Roche: Employment, Equity Ownership, Patents & Royalties. Herter:Roche: Employment, Patents & Royalties. Colombetti:Roche: Employment. Sam:Roche: Employment. Le Clech:Roche: Employment. Freimoser-Grundschober:Roche: Employment. Richard:Roche: Employment. Nicolini:Roche: Employment. Gerdes:Roche: Employment. Lariviere:Roche: Employment. Neumann:Roche: Employment. Klein:Roche: Employment, Patents & Royalties.

Beschreibung: GA101 is a novel humanized and glycoengineered CD20 antibody that was derived by humanization of the parental B-Ly1 mouse antibody and subsequent glycoengineering leading to the following characteristics: high affinity binding to the CD20 type II epitope; low complement dependent cytotoxicity (CDC) activity; high direct cell death induction; and high antibody-dependent cellular cytotoxicity (ADCC) potency. In vivo studies examined the consequences of GA101 on the growth of aggressive NHL xenograft models in SCID beige mice and the subsequent effects on survival. These studies demonstrated an outstanding dose-dependent anti-tumor activity of GA101. In particular, GA101 mediated superior anti-tumor efficacy in staged s.c. Z138 mantle cell lymphoma (MCL) and SU-DHL4 diffuse large B cell lymphoma (DLBCL) xenograft models in direct comparison to rituximab. Weekly dosing of GA101 at 10 mg/kg (Z138 model) or 30 mg/kg (SU-DHL4) was able to induce complete tumor remission and long-term survival (cures) in all animals. Consistent anti-tumor efficacy against established tumors was also observed in a number of additional NHL models including Raji Burkitt’s lymphoma or OCI-LY18 DLBCL models. In addition GA101 was evaluated in an orthotopic disseminated Z138 model. After i.v. injection of Z138 cells SCID beige mice developed large intraperitoneal lymphoid tumors in the ovary. Several experiments showed that GA101 mediated increased overall and median survival in this model in direct comparison to rituximab. All xenograft studies were performed in SCID beige mice that are incompetent for NK-mediated ADCC. Indeed, control experiments with non-glycoengineered GA101 in s.c. xenograft models suggest that the superiority of GA101 does not depend on enhanced interactions of the engineered Fc region with murine effector cells such as macrophages/monocytes or granulocytes. Thus, the anti-tumor effects of GA101 in s.c. xenograft models can be primarily attributed to non-effector cell mediated activities. Currently, experiments are ongoing to evaluate the efficacy of GA101 in combination with classical chemotherapy regimens and novel targeted agents. First results from these studies will be presented during the 2007 ASH Annual Meeting. In summary it can be assumed that the combination of the recognition of a type II epitope together with improved ADCC potency might translate into superior efficacy in the clinical treatment of CD20 positive malignant diseases.