Functional characterization of ARID1A mutations in follicular lymphoma

Antoniolli, Martina (2023): Functional characterization of ARID1A mutations in follicular lymphoma. Dissertation, LMU München: Medizinische Fakultät

Abstract

Background: Follicular lymphoma (FL) is one of the most common malignant lymphomas worldwide and the most common form of indolent lymphoma. FL is highly heterogeneous from both the clinical and molecular point of view. It remains a clinical challenge since advanced-stage disease is still considered incurable, and patients ultimately present with relapsed or re-fractory disease. Molecularly, FL is characterized by highly recurrent genetic mutations in genes coding for epigenetic modifiers. ARID1A mutations are among the most frequent mutations in FL (~10-20 % at the time of diagnosis). ARID1A mutations are a component of the prognostic clinic-genetic risk model m7-FLIPI (Pastore, Jurinovic et al. 2015). These mutations are primari-ly disruptive and result in protein haplodeficiency. Functionally, ARID1A is part of a SWI/SNF complex, which controls chromatin accessibility and is involved in numerous processes, includ-ing gene expression. Aim: Functionally characterize ARID1A mutations in representative human FL model systems. Methods: I used established and primary FL-like cell lines that harbor the hallmark t(14;18) translocation with or without heterozygous or homozygous ARID1A mutations (introduced by CRISPR/Cas9) or knock-down (by shRNA). I applied complementary omics approaches (RNA-Seq and ATAC-Seq) and functional assays to untangle the consequences of ARID1A loss in these FL model systems. Results: ARID1A loss profoundly altered gene expression. Across three cell lines, we observed consistent down-regulation of genes involved in cell cycle regulation and apoptosis pathways upon ARID1A loss. In functional experiments, I could show that ARID1A mutant clones are characterized by significantly slower cell proliferation and increased formation of anaphase bridges. Next, I demonstrated that ARID1A loss results in decreased FAS levels and lower sensitivity to FASLG-induced apoptosis. We discovered the underlying molecular mechanism through ad-vanced bioinformatics analyses and functional experiments. Briefly, ARID1A loss does not di-rectly affect FAS expression. Still, it results in reduced DNA accessibility and expression of the co-transcription factor RUNX3, thereby hindering RUNX3-ETS1 cooperativity and ETS1-induced FAS expression, which promotes a functionally and clinically relevant immune-evasive pheno-type. Finally, RNA-Seq analysis indicated that ARID1A loss alters the plasma membrane and cyto-skeleton functions, as well as the overall abundance of ligands and receptors. Ex vivo co-cultures of FL-like cells with T cells suggested impaired immune synapse formation with CD4 T lymphocytes upon ARID1A loss. Discussion and conclusion: Overall, our analyses provide novel insights into the functional consequences of ARID1A mutations in FL, most notably promoting immune evasion. A better understanding of mutation-specific biology, including its impact on interactions within the tumor microenvironment, holds promise for improved patient stratification and the development of personalized treatment approaches.

• Dokumententyp: Dissertationen (Dissertation, LMU München)
• Themengebiete: 500 Naturwissenschaften und Mathematik ; 500 Naturwissenschaften und Mathematik > 570 Biowissenschaften, Biologie
• Fakultäten: Medizinische Fakultät
• Sprache der Hochschulschrift: Englisch
• Datum der mündlichen Prüfung: 4. Oktober 2023
• 1. Bericht­erstatter:in: Weigert, Oliver