Abstract 1013: Analyzing infiltrating B cell populations in the tumor microenvironment using single cell transcriptomics
Authors: Taylor Sarah E. , Boutet Stephane , Giangarrá Valeria , Zheng Grace X. , Barrio Alvaro M. , Montesclaros Luz , Lee Josephine , Marrs Samuel , Wu Kevin J. , Ryvkin Paul , Mikkelsen Tarjei , Church Deanna M. 
Published: August 2018

Abstract
Understanding the complex interactions between malignant, stromal and infiltrating lymphocyte cell types within the tumor microenvironment is a critical component of personalized cancer treatments. We describe an approach that couples single-cell transcriptional profiling of tumors with high-resolution receptor profiling of tumor infiltrating lymphocytes (TILs). Using a fully integrated, droplet-based system for 5' single cell RNA sequencing (scRNA-seq), we simultaneously profiled the transcriptome and immune repertoire of the same cells from a primary colorectal cancer (CRC) tumor and a primary non-small cell lung cancer (NSCLC) tumor. Each tumor varied in type and proportion of its cellular components, noticeably in the proportion of TILs. The tumor cells in the CRC were mostly epithelial in nature, with the TILs comprising 21% of the total cells sequenced. This lymphocyte population consisted of both T (5% CD4+, 3% CD8+) and, interestingly, B cells (5% CD19+), with a number of plasma B cells (IGH high, CD138+) also identified. The NSCLC tumor had a high immune cell infiltrate, with lymphocytes comprising 43% of cells (19% T, 23% B, and 5% plasma B cells) indicating a robust adaptive immune response. To examine these cells further, CD45+ cells were subject to scRNA-seq. When compared to the unenriched sample, the enriched cell population had a similar B to T cell ratio with a reduction in the proportion of plasma B cells, likely due to exclusion of mature plasma B cells by the sort (47% T (44% CD4+, 29% CD8+), 42% B, and 2% plasma B cells). To fully characterize the B cell infiltrate we applied targeted 5' scRNA-seq to obtain full length, paired, B cell receptor heavy and light chain sequences. Despite the presence of plasma B cells in the CRC tumor, limited clonal expansion was observed. In the NSCLC tumor, clonal expansion of B cells was observed in the unenriched tumor sample but these clonotypes were not present in the CD45+ population, suggesting the clonally expanded cells are in the mature plasma B cell population. We also applied a targeted scRNA-seq approach to obtain full length paired T cell receptor alpha and beta sequences, however, no clear T cell expansion was observed in either sample. These findings emphasize the importance of examining receptor sequences rather than relying on the presence of B or T cells alone to determine if a robust immune response is being mounted. The presence of tumor-infiltrating B lymphocytes has been linked to a favorable clinical outcome in some types of cancers. Identification of these cells, their subsets and high resolution profiling of their receptor sequences using scRNA-seq allows a novel insight into the adaptive immune response in tumors. This technology will allow better characterization of tumor heterogeneity and the adaptive immune response to the tumor microenvironment and will serve as a foundation for future research into tumor immunology and immunotherapy.
Citation Format: Sarah E. Taylor, Stephane Boutet, Valeria Giangarrá, Grace X. Zheng, Alvaro M. Barrio, Luz Montesclaros, Josephine Lee, Samuel Marrs, Kevin J. Wu, Paul Ryvkin, Tarjei Mikkelsen, Deanna M. Church. Analyzing infiltrating B cell populations in the tumor microenvironment using single cell transcriptomics [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 1013.