Circulating Factors Provoke Endothelial Dysfunction in the Human Microcirculation Following Doxorubicin Chemotherapy

Abstract

RationaleAnthracyclines (e.g., doxorubicin) are among the most effective anti‐cancer therapies (CTx), yet their use is limited by cardiotoxic side effects. We sought to characterize the impact of anthracycline‐based CTx on microvascular endothelial function in breast cancer patients and determine if circulating factors are sufficient to induce endothelial dysfunction.MethodsWe assessed microvascular endothelium‐dependent vasodilation and its underlying mechanisms in two groups of breast cancer patients: 1) CTx‐naïve controls and 2) CTx‐treated patients (one month post‐CTx). Adipose samples were obtained at the time of surgical procedures. Freshly‐dissected arterioles were cannulated and pressurized, then flow‐induced production of nitric oxide (NO) and mitochondria‐derived hydrogen peroxide (H2O2) were visualized with fluorescent indicator probes. Next, we assessed whether plasma obtained from breast cancer patients one month following completion of CTx would evoke endothelial dysfunction. Adipose arterioles from healthy donors were exposed to diluted plasma from either CTx patients or healthy controls. Vessels were cannulated and filled with 10% plasma for overnight incubation prior to assessing flow‐mediated dilation (FMD; videomicroscopy). Data are expressed as area under the curve (AUC) for percentage change in diameter across five flow gradients and are shown as mean ± SE for n = 3–9 per condition.ResultsIn the cross‐sectional study, FMD was ~65% lower in vessels from breast cancer patients who had undergone CTx compared to those who were CTx‐naïve (AUC: 23 ± 10 vs. 61 ± 6, P = 0.004). Preliminary results suggest flow‐induced NO production is abolished in vessels from patients one month after CTx (Panel A, fold change in NO: −0.2 ± 0.2 vs. CTx‐naïve group: 11.3 ± 5.6; P = 0.08), whereas mitochondrial H2O2 production is elevated (Panel B, fold change in mtH2O2: 0.52 ± 0.25 vs. −0.51 ± 0.23; P = 0.03). FMD was also ~40% lower in vessels exposed to CTx plasma vs. healthy plasma (Panels C & D, AUC: 45 ± 7 vs. 75 ± 7, P = 0.049). Degradation of hydrogen peroxide (H2O2) with PEG‐catalase further reduced FMD in vessels exposed to CTx plasma (AUC: 22 ± 12), whereas PEG‐catalase did not affect dilation in vessels exposed to healthy plasma (AUC: 62 ± 17). Inhibition of NO synthase (NOS) with L‐NAME restored FMD in vessels exposed to CTx plasma (AUC: 77 ± 6), while L‐NAME had minimal impact on dilation in vessels exposed to healthy plasma (AUC: 60 ± 9). Future studies are needed to determine whether this reflects NOS uncoupling.Summary & ConclusionsMicrovascular endothelial function is impaired in breast cancer patients following doxorubicin chemotherapy. This is accompanied by a loss of NO and elevation of H2O2. Plasma obtained from breast cancer patients one month after cessation of CTx recapitulates these effects in otherwise healthy vessels, which suggest that circulating factors may contribute to endothelial dysfunction following treatment. Our preliminary findings reveal long‐term defects in the microcirculation that may contribute to adverse cardiovascular outcomes associated with doxorubicin chemotherapy.