Circulating factors following doxorubicin treatment in a tumor-bearing model induce endothelial dysfunction

Abstract

Chemotherapy (CTx) is essential to prevent the growth of tumors and has increased patient survival significantly. However, CTx agents have cardiotoxic side effects that can ultimately lead to heart failure. As a result, cardiovascular (CV) death is now the second leading cause of death in cancer patients. It has been accepted that adverse CV events are driven by a dysfunctional microcirculation, therefore CTx-induced endothelial dysfunction may play a key role in development of myocardial pathologies.Many CTx agents induce mitochondrial DNA (mtDNA) damage as many drugs intercalate into double stranded DNA, promoting the release of cell-free mtDNA (cf-mtDNA) into the circulation. Toll-like receptor 9 (TLR9) on the vascular endothelium detects cf-mtDNA and results in an inflammatory response. Prolonged exposure to CTx agents may enhance the mtDNA damage and stimulate the release of additional cf-mtDNA. To determine whether increased drug exposure exacerbates mtDNA damage, two forms of clinically used CTx were utilized: doxorubicin (Dox) and a lysosomal-derived nanoparticle with encapsulated Dox (Dox-NP), which targets tumor cells more specifically. With the ability of tumor-targeted delivery via Dox-NP, we hypothesize that Dox-NP augments mtDNA damage and the inflammatory response.Male Sprague Dawley rats received rat hepatoma-derived tumor cells into the liver twelve days prior to treatment start. Rats were treated with one weekly Dox or Dox-NP injection (7.5mg/kg·BW) for two weeks (cumulative dose 15mg/kg·BW). Cardiac function was evaluated before and six weeks after the initial dose of CTx. Vessel function was evaluated following euthaniasia six weeks after the initial dose. Plasma was obtained upon euthanasia, and cf-mtDNA levels were quantified via qPCR. NF-kB expression, downstream of TLR9 was determined via western blot.Rat microvessels from both treatment groups showed reduced endothelial-mediated vasodilation. Quantification of cf-mtDNA showed a trend toward increased cf-mtDNA levels in response to both Dox and Dox-NP. Likewise, NF-kB expression was elevated three-fold in comparison to Dox. To confirm whether endothelial dysfunction was the result of the inflammatory TLR9 signaling in vivo, isolated human microvessels were treated with 100nM Dox-NP. Vasodilation was not impaired, suggesting that Dox-NP is not directly harmful to the endothelium.Together, these data indicate that tumor-targeted Dox-NP have little to non-direct impact on the endothelium. Thus, endothelial dysfunction in animals treated with Dox-NP is secondary to its effects on a tumor and may be mediated by tumor-derived cf-mtDNA and TLR9 signaling. Moreover, the present data shows that direct effects of standard Dox on ECs may be only partially responsible for impaired dilator function. Conclusively, this work supports the concepts of a new mechanism of Dox-induced endothelial dysfunction. This work was supported by the NIH, 5 R01 HL133029-0 This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.