Friday, November 22, 2024
11/22/2024
PROJECT SUMMARY Cystic lymphatic malformations (LMs) are congenital vascular anomalies of the lymphatic system that cause life- threatening complications. LMs most commonly arise in the neck, and affected children are often born with large, disfiguring neck masses that obstruct the airway and cause severe functional deficits. LMs frequently become inflamed and infected, and spread locally by remodeling the extracellular matrix (ECM) of surrounding tissue. This causes recurring episodes of pain and a poor quality of life. These episodes require recurrent antibiotics and steroids and make surgery more difficult. There is no cure for LMs, but patients endure multiple resections, injections, and laser treatments in attempts to control the disease, often for their entire lives. When it was determined that LMs were caused by single nucleotide variations (SNVs) in in the oncogene PIK3CA, we began prescribing two chemotherapeutic medications that target the PI3K/mTOR pathway, which is activated by these genetic changes. While some patients experience reductions in LM size, many receive no benefit and none are ultimately cured despite being subjected to severe side effects. In efforts to develop individualized treatments that are more effective and safer, our laboratory performed the first ever multi-omic analysis of LM endothelial cells (LM-ECs) which are responsible for LM growth. This integration of RNA sequencing, microRNA sequencing, and proteomic mass spectrometry uncovered dysregulation of several pathways that control cell proliferation, inflammation and infection, and ECM remodeling. These pathways are controlled by microRNAs (miRNAs), which fine tune gene expression by silencing protein translation. By delivering miRNA mimics and inhibitors to cells, it is possible to reverse the dysregulation of these pathways, which we have demonstrated by inhibiting miR-21 and reducing LM-EC proliferation in a recent publication. We propose to deliver a topically administered, nanocarrier-conjugated miRNA-21 inhibitor to LMs in an in vivo murine xenograft model (Aim 1). Based on preliminary data, we expect this to reduce the growth of LMs and demonstrate a novel approach to treatment. We will also characterize the mechanisms of the dysregulated miRNA-mRNA-protein pathways we discovered that drive recurrent infection, and will reverse this dysregulation using miRNA mimics and inhibitors in vitro (Aim 2). These include cell surface receptors that bind immune cells (ICAM-1, CD44), and endothelial junction proteins that maintain lymphatic vessel integrity (ANGPT2, JUP, DSP, GJA1). Finally, we discovered that LM-ECs dysregulate two secreted ECM-remodeling enzymes (PLAT/RELN), along with their upstream miRNAs, and that LM-ECs are activating ECM-remodeling in distant cells. By targeting these miRNAs, we will demonstrate restoration of normal RELN/PLAT expression and reduction of paracrine ECM remodeling (Aim 3). The completion of these aims will facilitate the development of a combinational miRNA-based therapy that simultaneously targets the most pathologic mechanisms in LM-ECs. This would change the paradigm in the treatment of all vascular anomalies and other diseases of the lymphatic system.
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