Clonal dynamics towards the development of venetoclax resistance in chronic lymphocytic leukemia

Abstract

Deciphering the evolution of cancer cells under therapeutic pressure is a crucial step to understand the mechanisms that lead to treatment resistance. To this end, we analyzed whole-exome sequencing data of eight chronic lymphocytic leukemia (CLL) patients that developed resistance upon BCL2-inhibition by venetoclax. Here, we report recurrent mutations in BTG1 (2 patients) and homozygous deletions affecting CDKN2A/B (3 patients) that developed during treatment, as well as a mutation in BRAF and a high-level focal amplification of CD274 (PD-L1) that might pinpoint molecular aberrations offering structures for further therapeutic interventions.

Conflict of interest statement

C.D.H, C.-M.W., B.E., K.-A.K., M.H., and L.P.F. received research funding from Hofmann-La Roche. Research support was also provided by AbbVie to P.C., C.-M.W., B.E., K.-A.K., and M.H. P.C., C.-M.W., B.E., K.-A.K., H.C.R., M.H., and L.P.F. obtained consulting and/or speaker’s honoraria from AbbVie. P.C., C.-M.W., B.E., K.-A.K., and M.H. received consulting and/or speaker’s honoraria from Hofmann-La Roche. AbbVie provided travel support to P.C., N.K., and L.P.F. The remaining authors declare no competing financial interests.

Figures

Fig. 1

Patient and their related matched pre-treatment and relapse samples characteristics. a Absolute lymphocyte counts and lymphadenopathy of the patients during venetoclax therapy. Day zero marks the start of the venetoclax treatment. Green lines show the time points of sample collection. Computer tomography (CT) scans for staging were performed at the time points marked by red lines. b Results from whole-exome sequencing are shown, including: number of somatic mutations, sample ploidy, percent of the genome undergoing copy number alterations (blue for losses and red for gains), and cancer-related gene mutations with pronounced clonal dynamics during therapy. Genomic alterations are annotated according to the color panel below the image. Sample type/compartment and the status if a patient has undergone a Richter’s transformation are additionally indicated. Pre-treatment samples (T0) are shown in red. c Giemsa, Ki67, CD3, and CD274 stains from lymph node material of patient C811 after relapse from venetoclax. High protein levels of CD274 are consistent with the genomic amplification of the locus containing CD274. Scale bar, 100 µm

Fig. 2

Heterogeneous clonal evolutions under venetoclax therapy. Phylogenetic trees at the left side of each panel demonstrate the clonal evolution of the reconstructed cell populations for each patient. Highlighted mutations that occurred during tumor evolution are present in all descendent clones. Therefore, mutations in the most common ancestor population (C0) are present in all analyzed samples at a clonal level. The second type trees (right-bottom of each panel) demonstrate the phylogenetic relations of the matched pre-treatment and relapse samples from a patient, as commonly used in other cancer evolution studies,. Clonal composition of the samples (top-right of each panel) provides a link between both types of phylogenetic trees. We inferred diverse evolutionary paths across the patients: a linear evolution (C789), b branching evolution into three lineages (C577), c divergent evolution of two branches (C548), and d convergent evolution (C586). Pre-treatment sample names are displayed in red. Notable gene alterations are shown in the context of the ancestral relation of the clones

Fig. 3

Overexpression of oncogenic BRAF in the OCI-LY19 cell line. a Western blot analysis of BRAF and MCL1 in the BRAFV600E overexpressing OCI-LY19 cell line vs. its empty vector control. b Growth inhibition of BRAFV600E transfected OCI-LY19 cells and the empty vector control is shown as a function of the concentration of venetoclax