Cell-of-Origin Subtyping of Diffuse Large B-Cell Lymphoma by Using a qPCR-based Gene Expression Assay on Formalin-Fixed Paraffin-Embedded Tissues

Wan-Hui Yan, Xiang-Nan Jiang, Wei-Ge Wang, Yi-Feng Sun, Yi-Xin Wo, Zheng-Zhi Luo, Qing-Hua Xu, Xiao-Yan Zhou, Jun-Ning Cao, Xiao-Nan Hong, Xiao-Qiu Li, Wan-Hui Yan, Xiang-Nan Jiang, Wei-Ge Wang, Yi-Feng Sun, Yi-Xin Wo, Zheng-Zhi Luo, Qing-Hua Xu, Xiao-Yan Zhou, Jun-Ning Cao, Xiao-Nan Hong, Xiao-Qiu Li

Abstract

The well-established cell-of-origin (COO) algorithm categorizes diffuse large B-cell lymphoma (DLBCL) into activated B-cell-like (ABC) and germinal center B-cell-like (GCB) subgroups through gene expression profiling. We aimed to develop and validate a qPCR-based gene expression assay to determine the COO subgroups of DLBCL with formalin-fixed paraffin-embedded (FFPE) tissue. We first established a DLBCL transcriptome database of 1,016 samples retrieved from three published datasets (GSE10846, GSE22470, and GSE31312). With this database, we identified a qPCR-based 32-gene expression signature (DLBCL-COO assay) that is significantly associated with the COO subgroups. The DLBCL-COO assay was further validated in a cohort of 160 Chinese DLBCL patients. Biopsy samples from DLBCL patients with paired FFPE and fresh frozen tissue were collected to assign COO subtypes based on the immunohistochemistry (IHC) algorithm (Han's algorithm), DLBCL-COO assay, and global gene expression profiling with RNA-seq. For 111 paired FFPE and fresh DLBCL samples, the concordance between the IHC, qPCR, and RNA-seq methods was 77.5% and 91.9%, respectively. The DLBCL-COO assay demonstrated a significantly superior concordance of COO determination with the "gold standard" RNA-seq compared with the IHC assignment with Han's algorithm (P = 0.005). Furthermore, the overall survival of GCB patients defined by the DLBCL-COO assay was significantly superior to that of ABC patients (P = 0.023). This effect was not seen when the tumors were classified by the IHC algorithm. The DLBCL-COO assay provides flexibility and accuracy in DLBCL subtype characterization. These findings demonstrated that the DLBCL-COO assay might serve as a useful tool for guiding prognostic and therapeutic options for DLBCL patients.

Keywords: cell-of-origin; diffuse large B-cell lymphoma; formalin-fixed paraffin-embedded tissue; gene expression profiling; immunohistochemistry; quantitative polymerase reaction (PCR).

Copyright © 2020 Yan, Jiang, Wang, Sun, Wo, Luo, Xu, Zhou, Cao, Hong and Li.

Figures

Figure 1
Figure 1
Flow diagram of gene expression signature identification and performance assessment.
Figure 2
Figure 2
Hierarchical clustering analysis of 32 gene expression profiles in 159 diffuse large B-cell lymphoma (DLBCL) specimens. Colored pixels capture the magnitude of the expression for each gene, where shades of red and green represent over-expression and under-expression, respectively. Right: the official symbol of 32 genes; left: a dendrogram of hierarchical clustering of these genes. Bottom: COO subgroups including germinal center B-cell-like (GCB, orange), activated B-cell-like (ABC, purple), unclassified (UNC, blue); top: a dendrogram of hierarchical clustering of all samples.
Figure 3
Figure 3
Patient outcomes according to cell-of-origin (COO) in the validation set. Kaplan–Meier plots of overall survival based on molecular subgroups stratified by (A) immunohistochemistry (IHC) and (B) DLBCL-COO assay. P-values were obtained using the log-rank test. UNC, Unclassified.

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Source: PubMed

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