Sodium-glucose cotransporter 2 inhibitor dapagliflozin prevents ejection fraction reduction, reduces myocardial and renal NF-κB expression and systemic pro-inflammatory biomarkers in models of short-term doxorubicin cardiotoxicity.

Background: Anthracycline-mediated adverse cardiovascular events are among the leading causes of morbidity and mortality in patients with cancer. Sodium-glucose cotransporter 2 inhibitors (SGLT2i) exert multiple cardiometabolic benefits in patients with/without type 2 diabetes, chronic kidney disease, and heart failure with reduced and preserved ejection fraction. We hypothesized that the SGLT2i dapagliflozin administered before and during doxorubicin (DOXO) therapy could prevent cardiac dysfunction and reduce pro-inflammatory pathways in preclinical models. Methods: Cardiomyocytes were exposed to DOXO alone or combined with dapagliflozin (DAPA) at 10 and 100 nM for 24 h; cell viability, iATP, and Ca++ were quantified; lipid peroxidation products (malondialdehyde and 4-hydroxy 2-hexenal), NLRP3, MyD88, and cytokines were also analyzed through selective colorimetric and enzyme-linked immunosorbent assay (ELISA) methods. Female C57Bl/6 mice were treated for 10 days with a saline solution or DOXO (2.17 mg/kg), DAPA (10 mg/kg), or DOXO combined with DAPA. Systemic levels of ferroptosis-related biomarkers, galectin-3, high-sensitivity C-reactive protein (hs-CRP), and pro-inflammatory chemokines (IL-1α, IL-1ß, IL-2, IL-4, IL-6, IL-10, IL-12, IL17-α, IL-18, IFN-γ, TNF-α, G-CSF, and GM-CSF) were quantified. After treatments, immunohistochemical staining of myocardial and renal p65/NF-kB was performed. Results: DAPA exerts cytoprotective, antioxidant, and anti-inflammatory properties in human cardiomyocytes exposed to DOXO by reducing iATP and iCa++ levels, lipid peroxidation, NLRP-3, and MyD88 expression. Pro-inflammatory intracellular cytokines were also reduced. In preclinical models, DAPA prevented the reduction of radial and longitudinal strain and ejection fraction after 10 days of treatment with DOXO. A reduced myocardial expression of NLRP-3 and MyD-88 was seen in the DOXO-DAPA group compared to DOXO mice. Systemic levels of IL-1ß, IL-6, TNF-α, G-CSF, and GM-CSF were significantly reduced after treatment with DAPA. Serum levels of galectine-3 and hs-CRP were strongly enhanced in the DOXO group; on the other hand, their expression was reduced in the DAPA-DOXO group. Troponin-T, B-type natriuretic peptide (BNP), and N-Terminal Pro-BNP (NT-pro-BNP) were strongly reduced in the DOXO-DAPA group, revealing cardioprotective properties of SGLT2i. Mice treated with DOXO and DAPA exhibited reduced myocardial and renal NF-kB expression. Conclusion: The overall picture of the study encourages the use of DAPA in the primary prevention of cardiomyopathies induced by anthracyclines in patients with cancer.

Combinatorial immune checkpoint blockade increases myocardial expression of NLRP-3 and secretion of H-FABP, NT-Pro-BNP, interleukin-1ß and interleukin-6: biochemical implications in cardio-immuno-oncology.

Background: Immune checkpoint blockade in monotherapy or combinatorial regimens with chemotherapy or radiotherapy have become an integral part of oncology in recent years. Monoclonal antibodies against CTLA-4 or PD-1 or PDL-1 are the most studied ICIs in randomized clinical trials, however, more recently, an anti-LAG3 (Lymphocyte activation gene-3) antibody, Relatlimab, has been approved by FDA in combination with Nivolumab for metastatic melanoma therapy. Moreover, Atezolizumab is actually under study in association with Ipilimumab for therapy of metastatic lung cancer. Myocarditis, vasculitis and endothelitis are rarely observed in these patients on monotherapy, however new combination therapies could expose patients to more adverse cardiovascular events. Methods: Human cardiomyocytes co-cultured with human peripheral blood lymphocytes (hPBMCs) were exposed to monotherapy and combinatorial ICIs (PD-L1 and CTLA-4 or PD-1 and LAG-3 blocking agents, at 100 nM) for 48 h. After treatments, cardiac cell lysis and secretion of biomarkers of cardiotoxicity (H-FABP, troponin-T, BNP, NT-Pro-BNP), NLRP3-inflammasome and Interleukin 1 and 6 were determined through colorimetric and enzymatic assays. Mitochondrial functions were studied in cardiomyocyte cell lysates through quantification of intracellular Ca++, ATP content and NADH:ubiquinone oxidoreductase core subunit S1 (Ndufs1) levels. Histone deacetylases type 4 (HDAC-4) protein levels were also determined in cardiomyocyte cell lysates to study potential epigenetic changes induced by immunotherapy regimens. Results: Both combinations of immune checkpoint inhibitors exert more potent cardiotoxic side effects compared to monotherapies against human cardiac cells co-cultured with human lymphocytes. LDH release from cardiac cells was 43% higher in PD-L1/CTLA-4 blocking agents, and 35.7% higher in PD-1/LAG-3 blocking agents compared to monotherapies. HDAC4 and intracellular Ca++ levels were increased, instead ATP content and Ndufs1 were reduced in myocardial cell lysates (p < 0.001 vs. untreated cells). Troponin-T, BNP, NT-Pro-BNP and H-FABP, were also strongly increased in combination therapy compared to monotherapy regimen. NLRP3 expression, IL-6 and IL-1ß levels were also increased by PDL-1/CTLA-4 and PD-1/LAG-3 combined blocking agents compared to untreated cells and monotherapies. Conclusions: Data of the present study, although in vitro, indicate that combinatorial immune checkpoint blockade, induce a pro- inflammatory phenotype, thus indicating that these therapies should be closely monitored by the multidisciplinary team consisting of oncologists, cardiologists and immunologists.

Interleukin-1 blocking agents as promising strategy for prevention of anticancer drug-induced cardiotoxicities: possible implications in cancer patients with COVID-19.

Cytokines in cardiac tissue plays a key role in progression of cardiometabolic diseases and cardiotoxicity induced by several anticancer drugs. Interleukin-1ß is one on the most studied regulator of cancer progression, survival and resistance to anticancer treatments. Recent findings indicate that interleukin1-ß exacerbates myocardial damages in cancer patients treated with chemotherapies and immune check-point inhibitors. Interleukin1-ß blocking agent canakinumab reduces major adverse cardiovascular events and cardiovascular death in recent cardiovascular trials. We focalized on the main biological functions of interleukin1-ß in cancer and cardiovascular diseases, summarizing the main clinical evidence available to date in literature. Especially in the era of SARS-CoV-2 infection, associated to coagulopathies, myocarditis and heart failure, cancer patients have an increased risk of cardiovascular complications compared to general population, therefore, the pharmacological inhibition of interleukin1-ß should be discussed and considered.