Abstract 4488: Effector B cells and tertiary lymphoid structures predict response to immune checkpoint blockade in solid tumors
Authors: Reddy Sangeetha M. , Helmink Beth , Gao Jianjun , Zhang Shaojun , Yizhak Keren , Sade-Feldman Moshe , Basar Rafet , Blando Jorge , Han Guangchun , Gopalakrishnan Vancheswaran , Zhao Hao , Liu Wenbin , Tawbi Hussein , Amaria Rodabe , Davies Michael , Gershenwald Jeffrey , Burton Elizabeth M. , Allison James , Tetzlaff Michael , Rezvani Katy , Hacohen Nir , Sharma Padmanee , Wang Linghua , Wargo Jennifer 
Published: August 2019

Abstract
Background: Cytotoxic T cell markers, PD-L1, and mutational burden have been identified as biomarkers of response to immune checkpoint blockade (ICB). However, there is a growing appreciation of B cells as biomarkers and mediators of response. We recently conducted a phase II trial of neoadjuvant ICB (NCT02519322) in patients with resectable melanoma, and found increased B cell infiltration in responders (R) versus non-responders (NR).
Methods: We further investigated our B cell findings through whole transcriptomic profiling in samples with high tumor purity and also targeted immune profiling with deconvolution algorithm MCP counter. Spatial organization of the B cells was assessed with singlet and multiplex immunohistochemistry (IHC). Additionally, B cell phenotype was queried through mass cytometry. B cell gene expression signatures were validated in a cohort of renal cell carcinoma (RCC) patients (NCT02210117), and B cell phenotype in a metastatic melanoma cohort, both treated with ICB.
Results: Whole transcriptomic analysis of the neoadjuvant melanoma ICB cohort identified that the most differentially expressed genes by response in baseline (b/l) samples were related to B cells and antibody production (MZB1, JCHAIN, IGLL5, FCRL5, p&lt;0.0001). Targeted profiling via MCP counter confirmed higher B cells by response in all b/l and on-treatment (on-tx) samples (p=0.036 and p=0.038, respectively). This was verified by singlet CD20 IHC stains, which also showed that ICB increased B cells in R (p=0.004) but not NR. The B cells were present in organized tertiary lymphoid structures (TLS) in close proximity to T cells and follicular dendritic cells, and the ratio of area occupied by TLS was higher in R (p=0.037 b/l and 0.002 on-tx). These findings were validated in a RCC cohort, in which B cell lineage scores and TLS marker CXCL13 were higher in R at b/l (p=2.6e-03 and 3.3e-03, respectively). We then investigated the phenotype of the intratumoral B cells in the neoadjuvant melanoma cohort with mass cytometry. Within the tumor microenvironment, we identified naïve, class switched and unswitched memory B cell populations, and plasma cell-like populations (CD27+, IgD-, C38++, CD138-, CD20-). Higher frequencies of class switched memory B cell and plasma-like cell populations were observed in R, whereas NR were characterized by higher naïve B cells. Matching peripheral blood based CyTOF demonstrated non-overlapping clusters of B cell phenotypes. Single cell RNA sequencing in an independent cohort of metastatic melanoma patients also demonstrated class switched, activated B cells in addition to plasma cells, and that activation markers were significantly associated with response to ICB.
Conclusion: Together, these data suggest that B cells, more specifically those with activated effector phenotypes, and TLS predict response to ICB and may also be contributing mechanistically to response.
Citation Format: Sangeetha M. Reddy, Beth Helmink, Jianjun Gao, Shaojun Zhang, Keren Yizhak, Moshe Sade-Feldman, Rafet Basar, Jorge Blando, Guangchun Han, Vancheswaran Gopalakrishnan, Hao Zhao, Wenbin Liu, Hussein Tawbi, Rodabe Amaria, Michael Davies, Jeffrey Gershenwald, Elizabeth M. Burton, James Allison, Michael Tetzlaff, Katy Rezvani, Nir Hacohen, Padmanee Sharma, Linghua Wang, Jennifer Wargo. Effector B cells and tertiary lymphoid structures predict response to immune checkpoint blockade in solid tumors [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 4488.