Friday, April 4, 2025
4/4/2025
Defining the Regulatory Landscape of Lupus with CVD Comorbidity - PROJECT SUMMARY Systemic lupus erythematosus (SLE) is an incurable autoimmune disease featuring large clinical heterogeneity and multi-organ dysfunction, including cardiovascular diseases (CVD). Understanding this heterogeneity is vital for accurate diagnosis, identifying therapeutic targets, providing personalized treatment, and improving patient care. Lupus has a substantial genetic component, and studying immune cell-specific genetic architecture reveals pathogenic mechanisms and therapeutic targets for SLE. Genome-wide association studies (GWAS) have identified numerous SLE-predisposing loci, typically defined by blocks of correlated single nucleotide polymorphisms (SNPs). GWAS loci are enriched in regulatory elements (REs) and can influence target gene expression through regulatory SNPs (rSNPs). It is challenging but critical to identify the underlying causal variants, target genes, and cell types involved in SLE pathogenesis. Monocyte-derived macrophages are pivotal in atherosclerosis and SLE, as their dysfunction impairs phagocytosis and clearance of dead cells, contributing to both diseases. Yet, the impact of SLE and CVD-associated variants on macrophage biology remains unclear. Our central hypothesis is that some SLE rSNPs alter cis-regulatory elements (cREs) controlling target genes involved in macrophage biology, fueling SLE and CVD. To test this hypothesis, we will perform systematic, high- throughput, unbiased, large-scale studies in macrophages to determine the regulatory effects of 9,931 SNPs within 182 SLE risk loci using induced pluripotent stem cells (iPSCs) from SLE patients and healthy donors. We will differentiate iPSC-derived macrophage progenitors (monocytes) into unpolarized (M0), pro-inflammatory (M1), and anti-inflammatory (regulatory, M2) phenotypes, recapitulating the natural molecular and functional phenotypes of primary macrophages. Through our studies, we aim to identify dysfunctions arising from genetic differences, and uncover regulatory elements and target genes in macrophages disrupted by SLE risk alleles. Aim 1 will utilize the high-throughput technique, “self-transcribing active regulatory region-sequencing” (STARR- seq) to screen thousands of SNP-containing regions for their regulatory potential in macrophages. Aim 2 will employ next-generation (NG)-Capture-C to discover SNP-specific cis interactions with endogenous, cognate target genes in macrophages. Aim 3 will apply CRISPR-Cas9 approaches to uncover the allele-specific functional consequences of the top 10 selected rSNPs on their target gene(s) in iPSC-derived macrophages. Also, to determine the potential association of SLE-associated risk alleles with CVD risk, we will analyze target gene effects in primary monocytes/macrophages from SLE patients with or without atherosclerosis, as well as healthy controls. Through this analysis, we aim to identify genetic factors contributing to both SLE and CVD, gaining valuable insights into their interplay. This enhanced knowledge may inform targeted therapies or drug repurposing strategies to address macrophage dysfunction in treating SLE and CVD.
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