Investigating the prognostic implications of extrachromosomal DNA in diffuse large B-cell lymphoma

Abstract

Although the first-line treatment R-CHOP for Diffuse large B cell lymphoma (DLBCL) is curative, 30-40% of patients eventually relapse. Previous studies in the lab have shown that a unique subpopulation of cells co-expressing MYC, BCL2, but not BCL6 (M+2+6-), and correlates with poor prognosis in DLBCL. It is hypothesized that this is due to a non chromosomal phenomenon, and we hereby propose extrachromosomal DNA (ecDNA) amplification as a potential mechanism leading to the overexpression of MYC and BCL2, thus originating the M+2+6- subpopulation. Here, we have optimized a DNA-FISH staining and image analysis pipeline which was used to detect and quantify ecDNA in DLBCL tissues. Interestingly, we observed that MYC and BCL2 FISH signals were positively correlated with their respective protein expression, suggesting ecDNA as one of the underlying mechanisms for oncogene overexpression. Since the tumour immune microenvironment (TME) plays an important role in treatment response and outcomes, we characterized the TME of DLBCL patients to investigate the potential effects of the M+2+6- subpopulation. To achieve this, we utilized data from a previously performed PhenoCycler-Fusion experiment and a combination of MAPS and QuPath software to accurately predict and classify cell types and marker expressions. We found that the TMEs of patients with dispersed M+2+6- cells was associated with an immunosuppressive microenvironment, which might explain the poor clinical outcomes observed. Our investigation suggests that an ecDNA-driven mechanism drives MYC and BCL2 overexpression in DLBCL, while the spatial distribution of M+2+6– cells influence the TME. These findings offer valuable insights into disease biology.