Shared and Distinct Genetic Features in Human and Canine B-Cell Lymphomas

Introduction

Animal models of human cancers are an important tool for the development and preclinical evaluation of new treatments. Canine B-cell lymphoma (cBCL) is an appealing alternative to murine preclinical models due to its frequent, spontaneous incidence and its clinical and histological similarity to human B-cell non-Hodgkin lymphoma (NHL). The potential utility of cBCL as a veterinary model of human B-cell lymphomas would be bolstered by a more complete understanding of the genetic features found in cBCL.

Methods

To study the genetics of cBCL, we obtained fresh frozen and matched plasma/serum from 86 patients from the Canine Comparative Oncology Genomic Consortium(CCOGC) with 65 confirmed as B-cell lymphomas by immunophenotyping. Tumor DNA was prepared into libraries using the QIAseq FX DNA Library Kit (Qiagen). Plasma and serum DNA was prepared into libraries using the NebNext Ultra II DNA Library Prep Kit. Targeted hybridization enrichment was performed on the libraries using our custom baits and sequencing reads were aligned to canFam3.1 using Geneious and each mutation was visually confirmed. Variants were annotated with Variant Effect Predictor and human-dog pairwise alignments were extracted from Ensembl to identify the orthologous human amino acid for all canine variants.

Results

Our analysis confirmed the previously reported high frequency of mutations in TRAF3 and FBXW7. We also observed mutations in POT1, TP53, and SETD2 at similar frequencies to those reported in previous studies. DDX3X was mutated in 20% of cases, which is substantially higher than previously reported. MYC mutations were also more frequent (13%) than has been previously described in cBCL.

In human lymphomas, MYC is commonly deregulated by translocation to a potent enhancer and these events are often associated with point mutations in MYC that are induced by aberrant somatic hypermutation (aSHM). Interestingly, we identified a more focal pattern of MYC mutations in cBCL that implies they do not result from aSHM and are likely functional. This finding implicates the conserved MYC phosphodegron sequence, a motif commonly mutated among additional aSHM-associated mutations, as the target of bona fide driver mutations in both human and cBCLs. Mutations in FBXW7 primarily affected the substrate recognition domain responsible for MYC degradation. The observation that MYC and FBXW7 mutations did not co-occur in any canine patient is consistent with the notion that FBXW7 mutations operate as an alternative path to MYC stabilization which is not frequently observed in human NHL.

DDX3X was one of the most frequently mutated genes in our cohort (20%). DDX3X mutations are common in human Burkitt lymphoma and, though less abundant in hDLBCL, tend to be observed in samples with MYC translocations. In Burkitt lymphoma, these mutations display a sex-specific pattern, wherein females show mainly missense mutations, while males are affected by loss-of-function mutations. Interestingly, all DDX3X mutations in cBCL are missense variants and are presumed to be dominant acting. This lack of sex difference in DDX3X mutations is an important distinction between human and canine B-cell lymphomas that warrants further exploration.

Conclusions

Our study has revealed key differences in the mutational profiles of canine and human B-cell lymphomas and provides an impetus for enhanced genomic characterization of canine lymphomas as a model for human NHL, particularly in clinical trial settings.