Investigating the role of neutrophils in trametinib-induced cardiomyopathy

Background/Introduction Trametinib (Trm) is a highly selective inhibitor of MEK1/2, downstream targets of the RAS signaling pathway. Trm has been used widely for the treatment of BRAF V600E/K-mutant melanoma. Trm is generally well tolerated but can be associated with potentially serious cardiotoxicity in up to 20% of treated patients. However, the specific mechanisms driving Trm-induced cardiomyopathy (TIC) remain under investigation. Our data suggest that the immune system, particularly recruited neutrophils, plays a critical role in the pathogenesis of TIC. Additionally, how current guideline directed medical therapies (GDMT) in heart failure (HF) impact cardiac inflammation in the setting of chemotherapeutic cardiotoxicity have yet to be elucidated. Purpose The goal of this project to determine how neutrophils contribute to TIC. Previous work from our lab demonstrates that Trm treatment causes mitochondrial damage in cardiac myocytes resulting in release of danger associated molecular patterns (DAMPs) that leads to heart failure. This project investigates how DAMPs active the innate immune system resulting in neutrophil influx into cardiac tissue. It also investigates which mechanistic pathways are involved and how metoprolol, a commonly prescribed pillar of heart failure GDMT, can potentially mitigate the pathological effects of neutrophilic influx. Methods/Approach FVB mice were gavaged with Trm 3mg/kg/day for 0-14 days prior to sacrifice. Cardiac function was assessed using murine echocardiography. Heart tissue was analyzed via RNAseq and flow cytometry. In vitro studies used neutrophils isolated from murine bone marrow or peripheral human blood samples. Results/Data Flow cytometry analysis of cardiac leukocytes demonstrates increased neutrophilic influx starting as early as day 3 in treatment and peaking on day 14 which correlates with decreasing cardiac function. In vitro experiments supported by RNAseq data suggest that proinflammatory chemokines like CXCL12, CCR2, and ICAM1 play a role in mediating neutrophil recruitment through macrophage activation and recruitment. Genes associated with neutrophil function and netosis like matrix metalloproteinase-8 are also upregulated in treated mice. Preliminary in vitro studies suggest metoprolol inhibits critical neutrophil functioning including netosis. Conclusion(s) Trm is associated with cardiomyopathy in up to 20% of patients. Understanding the mechanisms driving this cardiotoxicity may help reduce the likelihood of adverse cardiac events associated with future MEK inhibitor therapy used alone or in combination with other agents. Our findings suggest that Trm causes increased neutrophil influx into the heart following treatment due to DAMP release and activation of tissue resident macrophages. Future studies will focus on abrogating this pathologic response and better understanding how beta blockers influence cardiac inflammation and microvascular damage.