Shallow whole-genome sequencing of cell-free DNA for B-cell lymphoma diagnosis and disease monitoring: a standardized approach with underappreciated potential

Background B-cell lymphoma (BCL) has recurrent multiple somatic alterations. Given its cell turnover and physical relation to the cardiovascular system, it’s excellent for cell-free DNA (cfDNA) research through liquid biopsies (LBs). Recent studies focused on targeted sequencing for single nucleotide variation and translocation detection, require targeted expensive panels and are yet to be clinically standardized. The much cheaper shallow whole-genome sequencing (sWGS) for copy number alteration (CNA) detection is operative at hospitals offering noninvasive prenatal testing. Methods 127 BCL patients (39 Hodgkin lymphoma (HL) and 88 non-Hodgkin lymphoma (nHL)) were recruited at UZGent and AZDelta: 258 samples, including 127 baseline LBs; 98 follow-up interim and surveillance LBs from 32 patients; and 33 paired formalin-fixed-paraffin-embedded (FFPE) solid biopsies (SB). cfDNA, analyzed by sWGS (0.5x coverage), was mapped to human and viral reference genomes. The cohort was extended with 60 NIPT (liquid negative control) and 9 benign FFPE (tissue negative control) to statistically determine cancerous CNAs and abnormal viral titers. Results At staging, 36/46 (78.3%) of HL and 75/103 (72.8%) of nHL CNAs emerged from LBs. The remainder had no structural aberrations or insufficient cfDNA fraction. No CNAs were detected in 87% of HL FFPE samples. For DLBCL liquid CNAs were well-represented by their solid equivalent, indicated by an average Pearson correlation of 0.82. To quantify CNA, a statistic was developed that increases with rising deviance from the overall healthy diploid state. This copy-number-profile-abnormality (CPA) score positively and significantly correlates with Ann-Arbor stage (p=0.044), International Prognostic Index (p=0.038) and metabolic tumor volume (p=0.0004). For refractory/relapsed patients CNAs were very similar across different longitudinal moments. No CNAs emerged from patients in remission and CNAs at staging were indicative for histological subtype. Conclusion CNAs are detected frequently and accurately in LBs of BCL, enable histological classification and are constant at relapse. CPA score correlates to disease status