Targeting B-Cell Maturation Antigen (BCMA) with CC-93269, a 2+1 T Cell Engager, Elicits Significant Apoptosis in Diffuse Large B-Cell Lymphoma Preclinical Models

Abstract

B-cell maturation antigen (BCMA), a member of the TNF receptor superfamily, serves as the cell surface receptor for B-cell activating factor (BAFF). Upon binding of BAFF to BCMA, an intracellular signaling cascade is initiated resulting in upregulation of JNK pathway signaling events and NFkB mediated transcription. The canonical expression pattern of BCMA begins on germinal center B-cells and becomes maximally expressed on mature cells such as plasma cells. Given the high degree of expression of BCMA on multiple myeloma (MM) cells, the malignant counterpart to plasma cells, it has become a target of interest for CAR T and antibody mediated modalities such as antibody drug conjugates and bispecific molecules. Recent clinical data from clinical trials employing BCMA targeted CAR T cells or T cell engager (TCE) antibodies have demonstrated significant responses in heavily pretreated myeloma patients with overall response rates ranging from 70% to 80%. As BCMA is known to be expressed in earlier B-cell lineages, we sought to explore the expression of BCMA in non-Hodgkin lymphoma (NHL) and its sensitivity to CC-93269, a 2+1 TCE currently being clinically investigated in MM. NHL is a heterogeneous collection of lymphomas that can be classified into two major subgroups; aggressive lymphomas of which diffuse large B-cell (DLBCL) is the most prevalent subtype and indolent lymphomas of which follicular lymphoma is the largest constituent. We first sought to directly quantitate cell surface expression of BCMA utilizing a flow cytometry system based on a logarithmic dilution of phycoerythrin beads of a known quantity. In a panel of 43 NHL cell lines, we determined that BCMA expression ranged from 43 to 17,048 molecules per cell (median, 420). An isogenic pair of K562 that is null for BCMA expression and K562 constitutively overexpressing BCMA (K562-BCMA) (15,866 molecules/cell) served as negative and positive controls, respectively. Additionally, the MM cell line H929 was profiled to serve as an additional control with a BCMA expression level of 7,065 molecules/cell. Next, utilizing quantitative PCR we found that relative BCMA mRNA expression in the lymphoma cell lines ranged from 0.001 to 0.17-fold when normalized to the H929 MM cell line. Furthermore, we were able to determine that in the lymphoma cells there is a poor correlation between protein expression (mean fluorescent intensity) and mRNA expression (r2, 0.33). We next examined if there was any correlation between BCMA surface expression and T-cell mediated cytotoxicity after administration of CC-93269 in a co-culture assay. We selected 11 DLBCL cell lines with a surface expression ranging from 45 molecules to 17,000 molecules per cells and incubated them in a co-culture system with a defined 1:5 target:effector ratio with CC-93269 (0-200 ng/ml) for five days. Significant apoptosis as measured by annexin V and ToPro-3 staining of CFSE positive target cells was observed in 10 of the 11 cell lines profiled with an IC50 of 0.1 to 38 ng/ml for CC-93269. As controls, the K562 isogenic pair were also profiled with the K562-BCMA cell line exhibiting an IC50 of 0.5 ng/ml and no activity observed against the parental K562 cell line. Additionally, a bispecific antibody where the two binding domains for BCMA was altered to target HEL (hen egg lysozyme) demonstrated no activity against any of the cell lines profiled at a defined dose of 200 ng/ml. No association between CC-93269 activity and BCMA expression or cell of origin was found. To determine the expression of BCMA in primary DLBCL biopsy samples, we developed a novel monoclonal BCMA immunohistochemistry antibody (clone: G12). The antibody and IHC staining protocol were validated to have good on-target specificity in both cell lines and tissues, including MM and DLBCL biopsies, with a range of stain intensity (1-3+) observed in both the golgi and on the plasma membrane. A proof of concept study on a cohort of 110 commercial DLBCL samples is currently underway. Cumulatively, our data demonstrate that BCMA is expressed on the cell surface of a broad panel of NHL cell lines and in primary DLBCL lymph node biopsies. Additionally, the expression levels of BCMA in these preclinical cell line models was sufficient to elicit significant CC-93269 mediated cytotoxicity. These data highlight the potential for the treatment of DLBCL patients with a 2+1 T-cell engager targeting BCMA. Disclosures Hagner: Celgene Corporation: Employment, Equity Ownership, Patents & Royalties. Waldman:Celgene: Employment, Equity Ownership, Patents & Royalties. Gray:Celgene: Employment, Equity Ownership. Yura:Celgene: Employment, Equity Ownership. Hersey:Celgene: Employment, Equity Ownership. Chan:Celgene: Employment, Equity Ownership. Zhang:Celgene: Employment, Equity Ownership. Boss:Celgene Corporation: Employment, Equity Ownership. Gandhi:Celgene Corporation: Employment, Equity Ownership, Patents & Royalties.

Inflammatory Cells in Diffuse Large B Cell Lymphoma

“…The result is B cell maturation and differentiation into plasma cells [53]. The expression of BCMA has been expressed in B cell lymphoma highlighting its targeting in the potential use for the treatment of DLBCL patients [54][55][56]. Manfroi in a recent work demonstrated that in a significant fraction of DLBCL patients, tumor cells are able to recruit APRIL producing neutrophils through the release of CXCL8 in both GC and non-GC DLBCL subtypes that in turn induce DNA methylation and acetylation, crucial in DLBCL progression [57].…”