Clinical and transcriptomic characterization of patients with chronic lymphocytic leukemia harboring t(14;19): an ERIC study

Abstract

In chronic lymphocytic leukemia (CLL), the role of complex karyotype (CK) for prognostic stratification remains a topic of debate, and the impact of specific cytogenetic abnormalities is still unclear. This study aims to investigate the clinical and biological features of CLL with t(14;19)(q32;q13) (tCLL) involving the BCL3 gene. Patients with tCLL were younger and more commonly presented unmutated IGHV gene, subset #8 stereotypy, trisomy of chromosome 12, and complex karyotype than other patients without t(14;19) (oCLL). The presence of t(14;19) was associated with a shorter time to treatment and overall survival compared to oCLL. Gene expression analysis revealed a unique transcriptome profile in tCLL, characterized by the upregulation of BCL3 and the activation of B-cell receptor, PI3K-Akt. Conversely, apoptosis-related pathways were suppressed in tCLL. While the BTK gene was upregulated, the BCL2L11 gene, coding for the pro-apoptotic protein BIM, was downregulated. Notably, patients with tCLL were characterized by a trend (p = 0.058) for a longer time to the next treatment with BTK inhibitors (BTKi) compared to those treated with a venetoclax-based (Ven-based) regimen. We underscore the adverse outcomes of tCLL, its distinct molecular features and gene expression patterns. Therefore, our data suggest that identifying tCLL could help tailor therapeutic approaches.

Fig. 1. Kaplan-Meier curves for survival analysis.

The Kaplan-Meier curves of (A) time to first treatment and B overall survival of patients with t(14;19) (n = 101, tCLL) and other karyotyped CLL without t(14;19) (n = 450, oCLL), in patients at Binet A stage at diagnosis (C, D) and harboring unmutated IGHV genes (U-IGHV, E, F).

Fig. 2. Kaplan-Meier of time to next treatment.

The Kaplan-Meier curves of time to the next treatment of patients treated with t(14;19) (tCLL) and other CLL without t(14;19) (oCLL) who received (A) fludarabine-cyclophosphamide-rituximab (FCR) / bendamustine-rituximab (BR) (n = 32 and n = 107, respectively), B venetoclax-based therapy (n = 6 and n = 39) and C BTK inhibitor (n = 14 and n = 180) as frontline therapy.

Fig. 3. Gene expression profiling of tCLL, oCLL, and B cells from RNA-seq.

A Dimensionality reduction by principal component analysis (PCA) performed on gene expression estimates. t(14;19) CLL patients (red crosses), control CLL (yellow triangles), and healthy donor B cell (blue circles) samples are displayed according to the first two principal components (PC1, horizontal axis; PC2, vertical axis) explaining the expression variability of the data (31.6% and 15.3%, respectively). B Volcano plot of the gene expression differences computed between t(14;19) CLL (tCLL) and other CLL without t(14;19) (oCLL). Dots and labels are colored according to the gene expression rank in tCLL (100% for the most expressed, 0% for the least expressed). C Western blotting analysis of Bcl-3 protein in tCLL and oCLL. The histogram in the bottom panel shows the densitometric analysis of Bcl-3/Actin, normalized to normal B cells, expressed as arbitrary units (A.I.). Expression levels, quantified by RNA-seq and RT-qPCR (DDCt method; GAPDH used as reference gene; Mean ± SD shown; A.U., Arbitrary Units) of BTK (D, E) and BCL2L11 (F, G) in patients with tCLL and oCLL. H Significant gene sets resulted from gene set enrichment analysis on the MSigDb hallmarks. Upward and downward arrows represent positive and negative normalized enrichment scores (NES), respectively, colored according to the magnitude of the NES (red indicates high positive, and blue indicates low negative). The arrow size indicates the statistical significance of the enrichment. I Heatmap of differentially expressed genes (rows) between t(14;19) CLL (tCLL), other CLL without t(14;19) (oCLL) samples, and B cells (columns). Pink and green cells represent gene-scaled expression, higher and lower than the gene mean of B cells, respectively. Columns are clustered according to the relative expression; rows are grouped according to differential expression significance, considering the three comparisons oCLL vs. B cells, tCLL vs. B cells, and tCLL vs. oCLL (dots in the panel mid sections indicate the significant comparisons). For each gene set, the enriched pathways are shown on the left side of the panel.

Fig. 4. Kaplan-Meier curves for different therapies in patients with tCLL.

The Kaplan-Meier curve of time to next therapy after different treatments, such as FCR/BR, venetoclax-based therapy (Ven-based), BTKi-based, or other chemo/chemo-immunotherapy (CIT), both as first-line (A) or further-line (B) therapy.