Characterization of mechanisms driving CD20 loss in patients with relapsed or refractory large B-cell lymphoma treated with glofitamab.

Abstract

7022 Background: Glofitamab is a CD20xCD3 T-cell engaging bispecific monoclonal antibody that redirects T cells to eliminate malignant B cells in patients (pts) with relapsed or refractory non-Hodgkin Lymphoma. We characterized mechanisms driving CD20 loss in pts from a Phase I/II trial (NP30179) receiving Glofitamab monotherapy for Relapsed/Refractory Large B-cell lymphoma (R/R LBCL). Methods: Pts with LBCL and ≥2 prior therapies received obinutuzumab pretreatment followed by fixed-duration Glofitamab at the approved dose in phase I/II trial NP30179 ( NCT03075696 ) (Dickinson, et al. N Engl J Med 2022). Tumor biopsies were collected prior to treatment (Baseline, BL) in 128 pts, during treatment (tx) or at progression (PD) in 11 pts. The proportion of CD20+ tumor cells was determined by immunohistochemistry (IHC) using a dual CD20+ PAX5+ assay. Expression of MS4A1 , the gene encoding CD20, was measured by RNA-sequencing (RNA-seq) in 105/139 biopsies. MS4A1 mutation profiling was performed by next-generation sequencing on Cell-free circulating tumor DNA (ctDNA) from 133 pts. We subsequently characterized the functional consequences of identified mutations in vitro. Results: CD20 levels evaluated by IHC were high (>75% CD20+ tumor cells) in 110/128 BL biopsies. At BL, CD20 loss (<5% CD20+ tumor cells) was seen in 4/128 (3.1 %) biopsies. For 11 pts with BL and on-tx or at-PD biopsies, 7/11 (63.6%) pts presented CD20 loss on-tx/at-PD and 4/11 (36.4%) did not. Evaluation of gene expression profile showed a good correlation between CD20 gene and protein expression. Among the 7 pts with CD20 loss on-tx/at-PD biopsies, there were 4 biopsies with available gene expression data, and decreased CD20 expression was identified in 2/4. Evaluation of CD20 mutation revealed 11/134 (8.2%) pts harbored 16 MS4A1 mutations at BL or on-tx/at-PD. IHC data were available for 8/11 pts at BL where 2/8 presented CD20 loss (<5% CD20+ tumor cells), and for 1 pt at-PD who presented CD20 loss. 12/16 mutations were not previously reported. We characterized 14 mutations in vitro and subsequently demonstrated that 8 frameshift or deletion mutations lead to truncation of the protein, and 4 missense mutations lead to disruption in the transmembrane domain of CD20. These 12 mutations lead to loss of intracellular and extracellular CD20 expression, and abrogation of Glofitamab-mediated cytotoxicity in vitro. Conclusions: In pts with R/R LBCL treated with Glofitamab, loss of tumor antigen CD20 expression is one resistance mechanism to Glofitamab. Genetic alterations (fs, del or missense mutations) and transcriptional downregulation can contribute to loss of CD20 expression and they were both observed in pts treated with Glofitamab. Acknowledgments: The NCT03075696 study is sponsored by F. Hoffmann-La Roche Ltd.