Research project
DC2 Identifying Epigenetic determinants in Primary Immunodeficiencies in their Progression to Hematological Malignancies
Some individuals with primary immunodeficiencies (PIDs) are prone to developing lymphoproliferative disorders, including chronic lymphocytic leukemia (CLL). In this project, we aim to decipher the epigenomic and transcriptomic determinants of this group of PID patients, helping us predict the progression from PID to CLL. The project will unveil pathogenic underlying mechanisms and contribute to the development of novel clinical markers.
Keywords: primary immunodeficiencies, hematological malignancies, epigenetics, methylation, single-cell omics
State-of-the-art and Rationale
Individuals with primary immunodeficiencies (PIDs), including common variable immunodeficiency (CVID), are prone to develop lymphoproliferative disorders, including chronic lymphocytic leukemia (CLL). In CLL, the most prevalent leukemia among adults, a monoclonal expansion of mature B cells infiltrates lymphoid tissue, bone marrow, and peripheral blood. CLL can cause hypogammaglobulinemia per se or as a result of therapies targeting B cells, resulting in a secondary immunodeficiency (SID). Recent evidence supports that CLL patients also come from a malignant progression of patients who previously had a PID. Our team has studied for years both the epigenetic and transcriptomic alterations in PIDs. Specifically, we have focused on CVID, the most frequent symptomatic PID. In fact, we were the first to describe significant alterations in DNA methylation in memory B cells and published the first single cell atlas for CVID patients [1,2]. We now hypothesize that patients with CLL with different situations in relation to immunodeficiency may also exhibit different epigenetic and transcriptomic profiles. It is of inherent interest to understand the relationship between these differential profiles in the progression from CVID (PID) to CLL.
Research aim and objectives
Our project aims to obtain bulk and single-cell epigenomic and transcriptomic profiles of B cells and other immune cells in a prospective cohort of patients with CVID and CLL, representing CLL patients without immunodeficiency, CLL patients that have developed SID, CLL patients that progressed from a PID and, finally, CVID patients. We have three objectives: 1) To determine unique alterations, and underlying pathways, that distinguish CLL patients without and with immunodeficiency; 2) To identify pathways and factors and develop markers in CVID patients that anticipate the development of CLL; 3) To model some of the events leading to the transformation from a monogenic form of CVID (mouse) to a B cell leukemia and the impact in the epigenome and transcriptome.
Scientific approach
Cohort development. The project will involve the use of both retrospective and prospective cohorts for CVID and CLL obtained in collaboration with immunologists and hemato-oncologists in hospitals in Barcelona and Madrid. Patients will be fully immunophenotyped and tested for serum free light chain (sFLC) (relevant to infer the existence of an immunodeficiency of primary origin in CLL).
Mouse models. We will use MD4 mice (as a model of CVID), bred as hemizygous MD4 x C57BL/6J, and hemizygous MD4 mice, which will be compared to WT littermate controls, to be crossed with Eμ-TCL1 transgenic mouse as a transgenic mouse that develops a CLL-like disease to study the interplay and acquisition of epigenetic and transcriptomic alteration during development in the B cell lineage. Epigenetic and transcriptomic profiling as indicated below.
Epigenomic profiling, single cell atlasing and proteomic analysis. We will profile the patients’ groups described above. We will use whole genome DNA methylation analysis of the B cell compartment and single-cell transcriptomic profiling of several immune cell types. We will infer cell signaling pathways, transcription factors and cell-cell crosstalk to reconstruct the underlying mechanisms.
Identification of differential epigenomic, transcriptomic and proteomic changes in CLL patients without immunodeficiency, with SID, progressed from PID and CVID. These comparisons will identify hypogammaglobulinemia- and CLL- specific alterations and relate them to clinical features. For validation, we will perform amplicon sequencing, bisulfite pyrosequencing and qRT-PCR.
Modeling. We will perform in vitro modeling using B cells derived from monogenic forms of CVID, obtained through collaboration with clinicians and testing CRIPR-Cas9 libraries for B cell differentiation genes for immunophenotyping & epigenetic analysis.
References:
EpiQMax (Munich, Germany) (secondment 1); Ospedale Sant Raffaele (Milan, Italy) (secondment 2)