Single-cell analysis can define distinct evolution of tumor sites in follicular lymphoma

Blood. 2021 May 27;137(21):2869-2880. doi: 10.1182/blood.2020009855.


Tumor heterogeneity complicates biomarker development and fosters drug resistance in solid malignancies. In lymphoma, our knowledge of site-to-site heterogeneity and its clinical implications is still limited. Here, we profiled 2 nodal, synchronously acquired tumor samples from 10 patients with follicular lymphoma (FL) using single-cell RNA, B-cell receptor (BCR) and T-cell receptor sequencing, and flow cytometry. By following the rapidly mutating tumor immunoglobulin genes, we discovered that BCR subclones were shared between the 2 tumor sites in some patients, but in many patients, the disease had evolved separately with limited tumor cell migration between the sites. Patients exhibiting divergent BCR evolution also exhibited divergent tumor gene-expression and cell-surface protein profiles. While the overall composition of the tumor microenvironment did not differ significantly between sites, we did detect a specific correlation between site-to-site tumor heterogeneity and T follicular helper (Tfh) cell abundance. We further observed enrichment of particular ligand-receptor pairs between tumor and Tfh cells, including CD40 and CD40LG, and a significant correlation between tumor CD40 expression and Tfh proliferation. Our study may explain discordant responses to systemic therapies, underscores the difficulty of capturing a patient's disease with a single biopsy, and furthers our understanding of tumor-immune networks in FL.

Publication types

  • Comparative Study
  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adult
  • Aged
  • Antigens, Neoplasm / biosynthesis
  • Antigens, Neoplasm / genetics
  • Biopsy, Fine-Needle
  • CD40 Antigens / biosynthesis
  • CD40 Antigens / genetics
  • CD40 Ligand / biosynthesis
  • CD40 Ligand / genetics
  • Clonal Evolution / genetics*
  • DNA, Neoplasm / genetics
  • Disease Progression
  • Female
  • Flow Cytometry
  • Gene Rearrangement, B-Lymphocyte, Light Chain
  • Gene Rearrangement, T-Lymphocyte
  • Humans
  • Lymph Nodes / chemistry
  • Lymph Nodes / ultrastructure
  • Lymphocytes, Tumor-Infiltrating / immunology
  • Lymphoma, Follicular / chemistry
  • Lymphoma, Follicular / genetics
  • Lymphoma, Follicular / pathology*
  • Male
  • Middle Aged
  • Neoplasm Proteins / biosynthesis
  • Neoplasm Proteins / genetics
  • Phylogeny
  • RNA, Neoplasm / genetics
  • Sequence Alignment
  • Sequence Homology, Nucleic Acid
  • Single-Cell Analysis*
  • T Follicular Helper Cells / immunology
  • T Follicular Helper Cells / metabolism
  • Transcriptome
  • Tumor Microenvironment


  • Antigens, Neoplasm
  • CD40 Antigens
  • DNA, Neoplasm
  • Neoplasm Proteins
  • RNA, Neoplasm
  • CD40 Ligand