Microrna-155 Trafficking Incites Compartmental Inflammation in the Leukemic Niche

Compartmental bone marrow (BM) inflammation has been linked to acute myeloid leukemia (AML) progression and hematopoietic dysfunction, yet the underlying mechanisms remain unclear. MicroRNAs (miRs) are capable of broadly deregulating cellular gene expression programs through simultaneous, dose-dependent action on a panel of target genes. MicroRNA-155, a widely known proinflammatory miRNA is overexpressed in AML patients, in particular those with Flt3-ITD. We recently observed that miR-155 is highly abundant in AML patient plasma-derived extracellular vesicles (AML-EV) and identified inflammation as one of the most upregulated gene ontology (GO) categories in a proteomic screen of hematopoietic stem and progenitor cells (HSPCs) exposed to AML-EV. Here, we investigate whether AML-derived miR-155 coordinately regulates inflammatory signaling in the leukemic niche.

To better model compartmental inflammation in AML, we first established a highly penetrant, immunocompetent, fully congenic system. Using the monoblastic AML cell line C1498 (CD45.2+), we were able to reliably graft animals without myeloablation, thereby avoiding unintended changes in BM niche function. Across multiple experimental cohorts, we then analyzed the BM plasma secretome and observed dynamic changes in the level of several inflammatory chemokines and cytokines. At sacrifice, flow cytometric analysis of the residual HSPC (CD45.1+) showed concurrent upregulation of key inflammatory response genes (ISG15, STAT1, CXCL10) and evidence of replicative stress (γ-H2AX) in residual HSPC. Consistent with potential trafficking of miR-155 via AML-EV, we next confirmed the enrichment of miR-155 in C1498-EVs, observed gains in miR-155 in BM plasma EVs and demonstrated increased levels in HSPCs following EV uptake. To determine if miR-155 could function as a regulatory hub under inflammatory conditions we utilized four separate target prediction databases (miRDB, TargetScan, miRWalk, and miRanda) to generate miR-155 target gene set. With an initial list of 360 potential gene targets that met a minimum threshold prediction score, we queried six publicly available RNA-Seq datasets to focus on genes belonging to inflammation or DNA damage repair GO groups highly expressed in HSPCs. This yielded a final set of 9 candidate targets (Wee1, E2f2, Rps6ka3, Rps6ka5, Csflr, Mgme1, Rad51, Rad51b, Ercc1). Reasoning that miR-155 target sets are involved in niche inflammation and genotoxicity in AML, these computationally predicted targets will undergo testing and validation for involvement in inflammatory and genotoxic stress. Chronic inflammation in the AML niche contributes to disease persistence and failure of residual hematopoiesis. Our study will determine the role of miR-155 as a regulatory hub for compartmental inflammatory signaling, and simultaneously serve as a potential opportunity for novel therapeutic target discovery in leukemic BM.