Fig. 1

Identification of CLL-specific DNA methylation events using Methyl-COOM. a Schematic outline of the Methyl-COOM pipeline used for the identification of CLL-specific DNA methylation events. Methylome data of six distinct B cell subpopulations, representing different stages of B cell differentiation were used to infer normal B cell differentiation. A linear regression model was applied to model DNA methylation dynamics during normal B cell differentiation (“DNA methylation: B cells”). DNA methylomes of 34 primary CLL samples were used to identify the closest virtual normal B cell (cell-of-origin; COO) based on phylogeny analysis. The linear regression model was then used to infer the DNA methylome of the COO (“DNA methylation: COO”). Next, the DNA methylome of each CLL was compared to the DNA methylome of its COO. CLL-specific aberrant DNA methylation was defined as a significant deviation from the inferred COO methylome (“DNA methylation: CLL-specific”). b Identification of the cell-of-origin in CLL samples using phylogenetic analysis. A phylogenetic tree was generated using a set of linear CpG sites that show dynamic DNA methylation changes during normal B cell differentiation (linear B cell-specific CpGs, 59,326 CpGs). Pairwise Manhattan distances were calculated between DNA methylation profiles of normal B cells and CLL samples at B cell-specific CpGs and were subsequently used to assign the closest normal (virtual) B cell methylome (location of the node on the phylogenetic tree = differentiation stage of the cell-of-origin) to each CLL case. NBCs – naïve B cells; GCFs – germinal center founder B cells; loMBCs – early non class-switched memory B cells; intMBCs – non class-switched memory B cells; sMGZs – splenic marginal zone B cells; hiMBCs – class-switched memory B cells (mature B cells). CLL samples are depicted in orange color. Normal B cells are represented in green. c Summary of CLL-specific DNA methylation events. Top: pie chart displays the frequency of CpGs that are either dynamic (green) or stable (gray) during normal B cell differentiation. Middle: pie charts depict the frequency of CLL-specific DNA methylation events as fractions of the dynamic (classes A and B; left), and stable (classes C and D; right) sites. Bottom: schematic depicting the classification of CLL-specific DNA methylation events. We identified two groups: “sites with epigenetic B cell programming” and “sites without epigenetic B cell programming.” “Sites with epigenetic B cell programming” undergo DNA methylation programming during normal B cell differentiation, encompassing hypomethylation (class A) and hypermethylation events (class B) relative to the DNA methylome of the COO. “Sites without epigenetic B cell programming” are defined as CpG sites without significant DNA methylation changes during normal B cell differentiation and are classified as either hypo- or hypermethylation (classes C and D, respectively). Numbers of CLL-specific DNA methylation events (CLL-specific CpGs) resolved by class are indicated at the bottom