Intratumor heterogeneity and T cell exhaustion in primary CNS lymphoma

Heming, Michael; Haessner, Svea; Wolbert, Jolien; Lu, I-Na; Li, Xiaolin; Brokinkel, Benjamin; Müther, Michael; Holling, Markus; Stummer, Walter; Thomas, Christian; Schulte-Mecklenbeck, Andreas; de Faria, Flavia; Stoeckius, Marlon; Hailfinger, Stephan; Lenz, Georg; Kerl, Kornelius; Wiendl, Heinz; Meyer Zu Hörste, Gerd; Grauer, Oliver M

Abstract

Background: Primary central nervous system lymphoma (PCNSL) is a rare lymphoma of the central nervous system, usually of diffuse large B cell phenotype. Stereotactic biopsy followed by histopathology is the diagnostic standard. However, limited material is available from CNS biopsies, thus impeding an in-depth characterization of PCNSL. Methods: We performed flow cytometry, single-cell RNA sequencing, and B cell receptor sequencing of PCNSL cells released from biopsy material, blood, and cerebrospinal fluid (CSF), and spatial transcriptomics of biopsy samples. Results: PCNSL-released cells were predominantly activated CD19+CD20+CD38+CD27+ B cells. In single-cell RNA sequencing, PCNSL cells were transcriptionally heterogeneous, forming multiple malignant B cell clusters. Hyperexpanded B cell clones were shared between biopsy- and CSF- but not blood-derived cells. T cells in the tumor microenvironment upregulated immune checkpoint molecules, thereby recognizing immune evasion signals from PCNSL cells. Spatial transcriptomics revealed heterogeneous spatial organization of malignant B cell clusters, mirroring their transcriptional heterogeneity across patients, and pronounced expression of T cell exhaustion markers, co-localizing with a highly malignant B cell cluster. Conclusions: Malignant B cells in PCNSL show transcriptional and spatial intratumor heterogeneity. T cell exhaustion is frequent in the PCNSL microenvironment, co-localizes with malignant cells, and highlights the potential of personalized treatments.