Doxorubicin induces measurable vascular toxicity: assessment in a clinical and preclinical study

Abstract

Funding Acknowledgements Type of funding sources: Public Institution(s). Main funding source(s): Fund for Scientific Research (FWO) Flanders INSPIRE project (H2020-MSCA-ITN program) Background The chemotherapeutic doxorubicin (DOX) is frequently used to treat a wide variety of cancers, but the cardiotoxic side effects limits its clinical use in some patients. Additionally, DOX contributes to vascular toxicity, which may be an early manifestation of DOX-associated toxicity. Therefore, there is an interest to evaluate vascular function in patients receiving DOX-based treatment regimens. Purpose We aimed to assess arterial stiffness and endothelial dysfunction as potential markers of vascular toxicity in both DOX-treated cancer patients and a murine model. Methods Female breast cancer patients with a need for adjuvant or neoadjuvant DOX-based treatment were prospectively included. Chemotherapeutic regimen consisted of 12 weekly cycles of taxane treatment, followed by 4 cycles of DOX (and cyclophosphamide) treatment. Vascular function (endothelial function (FMD and RHI), arterial stiffness (Aix and cfPWV) and cardiac function (LVEF, hsTnI, NT-proBNP) were performed at baseline (T1), after completion of taxane treatment (T2) and after completion of DOX treatment (T3). In addition to the clinical study, male C57Bl6/J mice were intraperitoneally injected with 2 mg /kg (low dose) or 4 mg/kg DOX (high dose) once per week for 6 weeks. Arterial stiffness was assessed in vivo by measuring abdominal aorta pulse wave velocity (aaPWV) by high-frequency ultrasound imaging combined with ex vivo vascular function evaluation. Results Twenty patients (mean age 51.2 +/- 10.1 yrs) treated with DOX were included. After DOX treatment (T3 vs T2), left ventricular ejection fraction (LVEF) was decreased, while hsTnI and NT-proBNP levels were increased, indicating cardiotoxicity (Table 1). Likewise, LVEF was reduced in DOX-treated mice after 3 weeks, which persisted until the end of the treatment (6 weeks). In patients, RHI was impaired at T2 and T3 compared to T1, which indicates progressive endothelial dysfunction during treatment (Table 1). Consistent with these findings, DOX treatment in mice resulted in endothelial dysfunction, as evidenced by a lower basal nitric oxide (NO) index and reduced acetylcholine-induced endothelium-dependent vasorelaxation (Figure 1B & 1C). Finally, cfPWV was decreased at T2 and T3 compared to T1, even after correction for BP, suggesting rather an improvement of arterial stiffness (Table 1). Although DOX treatment in mice resulted in a similar trend towards lower aaPWV at the end of treatment (6 weeks), we observed an initial increase in aaPWV after 2 weeks (Figure 1A). Conclusion Collectively, these findings suggest that, apart from cardiotoxicity, DOX treatment results in early and consistent endothelial dysfunction, while evaluation of arterial stiffness may be time-sensitive. Hence, endothelial dysfunction may be a more reliable and sensitive marker than arterial stiffness to evaluate DOX-induced vascular toxicity in patients.

Characterization of systolic and diastolic function, alongside proteomic profiling, in doxorubicin-induced cardiovascular toxicity in mice

“…and organs were collected and weighed for further analysis. For proteomics, murine cardiac tissue from a historical study was used, previously reported in Bosman et al [12]. These mice underwent the same treatment protocol as the animals used in this study.…”

“…A detailed echocardiographic analysis of the DOX murine model has already been performed by Bosman et al, focusing on vascular and LV systolic function. We previously described EC dysfunction occurring early on in DOX-induced CTR-CVT [12]. The present study focuses on diastolic dysfunction and the identification of molecular changes in DOX-induced CTR-CVT in the aforementioned model.…”

“…The Ultimate 3000 Rapid Separation ultra-high performance liquid chromatography (UHPLC) system was coupled to a Q Exactive HF mass spectrometer (Thermo Scientific) with the following settings: Full MS scan between 250-1,250 m/z at a resolution of 120,000 followed by MS/MS scans of the top 15 most intense ions at a resolution of 15,000. A detailed description of the method is published elsewhere [12].…”

“…Two groups of human samples were included in the study: cancer survivors experiencing CTR-CVT (n = 24), and a control group of patients without prior cancer treatment, with preserved LVEF, undergoing cardiac catheterization for non-HF symptoms (n = 5). Of note, this cohort differs from the one previously described [12]. Blood samples were taken at diagnosis of CTR-CVT (77 ± 3 months after completion of cancer treatment) or at diagnosis of cardiomyopathy in the control cohort.…”

“…Recently, we documented vascular toxicity, represented by endothelial cell (EC) dysfunction and arterial stiffness, as a predecessor to the reduced LVEF upon DOX treatment in mice [10,11]. Thereby, EC dysfunction was reversible after stopping DOX treatment [12]. Endothelial dysfunction contributes to vascular contractile alterations and reduced compliance in both heart and aorta, compromising cardiac relaxation and filling, which can ultimately lead to diastolic dysfunction, or heart failure with preserved ejection fraction (HFpEF) [13,14].…”