Circulating tumour DNA concentration and genetic classification improve risk stratification in newly diagnosed patients with Diffuse Large B‐Cell Lymphoma

Introduction: Diffuse large B-cell lymphoma (DLBCL) is a genetically heterogeneous disease with poor outcomes for the 40% of patients who relapse or are refractory to upfront therapy, and current prognostic tools are unable to identify many of those patients. Recently, novel prognostic biomarkers like genetic classification and circulating tumor DNA (ctDNA) have been developed. Here we explore the value these biomarkers to improve prognostic stratification in DLBCL. Methods: DNA was extracted from 2 to 5 ml of plasma and formalin-fixed paraffin-embedded (FFPE) tumor tissue obtained at diagnosis from patients with DLBCL treated with R-CHOP-like regimes. Samples were sequenced in an Illumia NovaSeq using a panel of 112 genes that are recurrently mutated in lymphoid neoplasms, and variants were called using a pipeline that follows GATK best practices. Genetic subtype (GS) was determined using LymphGen tool, excluding A53, as copy number variation data was unavailable. ST2, EZB MYC negative and BN2 subtypes were considered favorable, while N1, MCD, EZB MYC positive and other were considered unfavorable. ctDNA levels were reported as haploid genome equivalents per mL of plasma and expressed as a base 10 logarithm (log hGE/mL) Results: We included 46 patients with median age of 64.8 years, 58% advanced stage (III-IV) and 65% germinal center DLBCL by Hans’s algorithm. Somatic mutations were detected in FFPE in all patients and in ctDNA in 40 (87%) patients. Most frequently mutated genes were KMT2D, CREBBP, TP53, ARID1A, MYD88 and CARD11. The sensitivity of ctDNA to detect mutations present in paired FFPE samples was 66% (for all variants) and 74.5% (for variants with >5% allelic frequency in FFPE samples). 67.4% of patients were successfully classified by LymphGen, with a concordance between ctDNA and FFPE of 84.7%. Patients without detectable ctDNA presented localized disease (Ann Arbor stage I) and low DNA concentration in plasma (<15 ng/ml). In our cohort, 24 patients presented a poor R-IPI score with a 3-year progression-free survival (PFS) of 61.5% (Figure 1A). These patients were further stratified by ctDNA levels and GS, and those unfavorable GS or ctDNA concentration >4 log hGE/ml presented worse PFS and OS than without risk factors a 3-year PFS of 34.6% and 90.9%, respectively (Figure 1B). Patients were classified into three groups according IPI score modified by ctDNA levels (Figure 1C), with a 3-year PFS of 94.7%, 81.2% and 18.8%, and a 3-year overall survival (OS) of 100%, 87.5% and 37.5%, respectively (Figure 1D–E). Keywords: Diagnostic and Prognostic Biomarkers, Genomics, Epigenomics, and Other -Omics No conflicts of interests pertinent to the abstract.