A New Class of T-Cell Bispecific Antibodies for the Treatment of Multiple Myeloma, Binding to B Cell Maturation Antigen and CD3 and Showing Potent, Specific Antitumor Activity in Myeloma Cells and Long Duration of Action in Cynomolgus Monkeys

Abstract

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.

Bispecific T-Cell Redirection versus Chimeric Antigen Receptor (CAR)-T Cells as Approaches to Kill Cancer Cells

“…These 2:1 (target cell antigen:CD3ε) TRBAs have been dubbed by Roche as “TCBs,” for “T-cell Bispecifics.” There are currently two clinical stage TCBs that incorporate the Fab as an extra appendage, including cibisatamab (aka RG7802, RO6958688, CEA TCB) [159,204], which has two binding arms for CEA and a single binding arm for CD3ε and RG6026 (aka RO7082859) [205], which has two binding arms for CD20 and one for CD3ε [160]. Two additional TCBs, one targeting BCMA [206] and another targeting a carboxyl-terminal fragment of HER2 expressed in about half of HER2-positive tumors [207], have been reported but are not yet in clinical trials.…”

Frequency of expression and generation of T-cell responses against antigens on multiple myeloma cells in patients included in the GMMG-MM5 trial

“…A different approach is (re)directing T-cells towards myeloma cell killing. This can be achieved via T-cell genetic engineering with chimeric antigen receptors or recombinant T-cell receptor, both requiring ex vivo engineering and expansion of patient specific T-cells [ 8 – 11 ], as well as T-cell bispecific antibodies that simultaneously bind a surface target on tumor cells and an associated T-cell receptor chain present on T-cells thereby inducing potent T-cell mediated killing of cells carrying the target [ 12 , 13 ]. A further - and potentially even prophylactic - approach is the development of cancer vaccines generating myeloma-specific immunity selectively targeting malignant cells - with limited toxicity to normal tissues [ 14 – 16 ].…”

The use of CrossMAb technology for the generation of bi- and multispecific antibodies

“…Recently, Vu and colleagues described a novel B cell maturation antigen (BCMA) specific 2+1 TCB based on this platform. 53,54 …”