CSF NGS-MRD assay as a prognostic tool for the risk of CNS recurrence in lymphoma. (A) Cumulative incidence of CNS recurrence in the prospective cohort of patients with no known CNS involvement (n = 22), stratified by the result of the CSF NGS-MRD assay at diagnosis; P-value by log-rank test; 2 of 8 NGS-MRD⁺ and 0 of 14 NGS-MRD⁻ patients with CNS relapse. (B) Amount of clonotypic DNA in the CSF (each dot represents a specific DNA sequence; sequences that were undetectable were excluded), measured as copy number per milliliter of acellular fluid, per 10⁶ diploid genomes in the cell pellet fraction, and as the clonotype frequency (in both types of material), according to subsequent CNS relapse status. The bars show medians and interquartile ranges; note logarithmic scale on the left and middle panels; P-values are by+ nonparametric Somers’ D statistic adjusted for within-patient clustering. (C) Lack of correlation between CSF contamination by blood (as measured by red cell count per mm³ of CSF) and abundance of clonotypic DNA in the same CSF sample, measured by copy number per milliliter (of the CSF acellular fluid), copy number per 10⁶ diploid genomes (in the CSF cell pellet), or as clonotype frequency in both fractions; P-values by nonparametric Somers’ D statistic adjusted for within-patient clustering; a linear fit with 95% CI bands is also shown. (D) Clonotypic DNA sequences in patients’ plasma (n = 10), measured as cfDNA copy count per milliliter of plasma, and as clonotype frequency among all B-cell genomes. (E) Lack of correlation between the abundance of clonotypic DNA in paired plasma and CSF samples (n = 10), as measured by copy count per milliliter and clonotype frequency in the acellular CSF. P-values by nonparametric Somers’ D statistic adjusted for within-patient clustering; a linear fit with 95% CI bands is also shown.