Dissection of DLBCL Microenvironment Provides a Gene Expression-Based Predictor of Survival Applicable to Formalin-Fixed Paraffin-Embedded Tissue

Abstract

Background

Gene-expression-profiling (GEP) studies recognized a prognostic role for tumor microenvironment (TME) in diffuse large B-cell lymphoma (DLBCL), but the routinely adoption of prognostic stromal signatures remains limited.

Patients and methods

Here, we applied the computational method CIBERSORT to generate a 1,028-gene matrix incorporating signatures of 17 immune and stromal cytotypes. Then, we performed a deconvolution on publicly available GEP data of 482 untreated DLBCLs to reveal associations between clinical outcomes and proportions of putative tumor-infiltrating cell types. 45 genes related to peculiar prognostic cytotypes were selected and their expression digitally quantified by NanoString technology on a validation set of 175 formalin-fixed, paraffin-embedded DLBCLs from two randomized trials. Data from an unsupervised clustering analysis were used to build a model of clustering assignment, whose prognostic value was also assessed on an independent cohort of 40 cases. All tissue samples consisted of pretreatment biopsies of advanced-stage DLBCLs treated by comparable R-CHOP/R-CHOP-like regimens.

Results

In silico analysis demonstrated that higher proportion of myofibroblasts (MF), dendritic cells, and CD4⁺ T-cells correlated with better outcomes and the expression of genes in our panel is associated with a risk of overall and progression-free survival. In a multivariate Cox model, the microenvironment genes retained high prognostic performance independently of the cell-of-origin (COO), and integration of the two prognosticators (COO + TME) improved survival prediction in both validation set and independent cohort. Moreover, the major contribution of MF-related genes to the panel, Gene Set Enrichment Analysis suggested a strong influence of extracellular matrix determinants in DLBCL biology.

Conclusions

Our study identified new prognostic categories of DLBCL, providing an easy-to-apply gene panel that powerfully predicts patients' survival. Moreover, owing to its relationship with specific stromal and immune components, the panel may acquire a predictive relevance in clinical trials exploring new drugs with known impact on TME.