Protect and Counter-attack: Nutritional Supplementation with Essential Amino acid Ratios Reduces Doxorubicin–induced Cardiotoxicity in vivo and promote Cancer Cell Death in vitro

Abstract

CommentaryHuman and experimental data show that nutritional supplementation with a specific mixture of essential amino (EAA) improves both mitochondrial function and biogenesis and reduces either ROS formation or apoptotic drive in myocytes [1][2]. Supplementation with EAA is also effective in humans when tested in patients with cardiac dysfunction [3]. Doxorubicin (Doxo), an effective cancer chemotherapy agent, causes severe cardio-toxicity due to mitochondria damage, production of reactive oxygen species (ROS) and cell apoptosis with myofibrillar disarrangement, which in turn induces chronic heart failure. Pathogenesis of toxicity, as reported by Zhang et al. [4] is connected to the binding of Doxo to topoisomeraseIIb , which triggers a complex cascade of events, altering the DNA response and apoptosis pathways and triggering a marked alteration in the transcriptome, that selectively affects oxidative phosphorylation and mitochondrial biogenesis in cardiomyocytes. As a result, Doxoinduced cardiotoxicity limits optimal drug-dose use, reducing its potential therapeutic effects in cancer patients [5].Consequently, we hypothesized that supplementation with EAA would blunt Doxo cardio-toxicity, but we also tested if this would happen without interfering with the Doxo therapeutic effects on cancer cells. We used C57BL/6 mice fed on a normal diet and i.p. injected with a single saline or Doxo-HCl (15 mg/kg, from SigmaAldrich) injection to induce cardiomyopathy [6], with or without oral supplementation of EAA (1.5 g/kg). After 14 days, the heart was collected and morphologically evaluated by optical and electron microscopy. In addition, to test whether EAA supplementation negatively interferes with the Doxo anticancer action, we treated different cancer cell lines (3 × 10 5 cells/well in DMEM medium of HCT116 colon carcinoma; MCF7 mammary gland carcinoma; M14 melanoma), with Doxo (0.7 microg/ml) and with or without EAA (1% w/v). Untreated cancer cell lines (cultured in DMEM only) were used as controls. The osmolarity of the culture medium was the same for all groups. After 24 hours the cells were counted with the Trypan blue dye exclusion assay to distinguish live from dead cells.The in vivo experiments showed that Doxo significantly lowered body weight (22.59 ± 1.33 g vs 30.12 ± 1.4 g. p<0.05) and heart weight (0.12 ± 0.01 g vs 0.17 ± 0.01 g. p<0.05) compared to the control group. Furthermore, Doxo induced morphological alteration of cardiomyocytes with giant and irregularly shaped nuclei with condensed chromatin (Figure 1a A-C). In addition, Doxo impaired cellular distribution, morphology and the number of mitochondria (24 ± 2.8 vs 31.62 ± 5.5 Nmit/100µ2. p<0.05). Interestingly, EAA supplementation of Doxo-treated animals protected normal heart weight (0.16 ± 0.01 g vs 0.12 ± 0.01 g. p<0.05) and preserved cardiomyocytes architecture, mitochondrial morphology and number (28 ± 2.8 vs 24 ± 2.8 Nmit/100µ2. p<0.05) (Figure 1a The in vivo findings were partially surprising because previous studies had shown that EAA nu...

Intake of Special Amino Acids Mixture Leads to Blunted Murine Colon Cancer Growth In Vitro and In Vivo

“…Similarly, in vivo experiments revealed high levels of nuclear chromatin thickening and fragmentation in tumors, along with strong immunostaining for activated caspase-3 in tumors from EAARD-fed mice. This aligns with previous studies showing that EAA-mix has a cell-dependent anti-proliferative and cytotoxic effect, activating autophagy and apoptotic pathways and leading to cancer cell death without affecting non-cancer cells [7,20,26].…”

“…We have previously explored the safety and benefits of food supplementation with a specific free AA mixture rich in EAA (EAA-mix) under various experimental conditions [5][6][7][8][9][10][11][12][13][14]. Some reports have discussed how reducing certain NEAAs (serine and glycine) and consequently increasing EAAs may impact cancer in animal settings [15,16].…”

“…We have previously demonstrated that altering the EAA/NEAA ratios in diets significantly influences mice survival [19]. Moreover, we have shown in vitro that an EAA-mix containing all free EAA in stoichiometric ratio inhibits tumor cell proliferation and survival by inducing both AUT and apoptosis in duplicating cancer cells [20,7]. Therefore, we investigated whether changes in EAA/NEAA ratios in diet also affect in vivo cancer development.…”

Essential Amino Acids-Rich Diet Increases Cardiomyocytes Protection in Doxorubicin-Treated Mice

“…Previous works wherein rodents were fed a diet containing an excess of all EAAs in a stoichiometric ratio have shown an increase in mitochondriogenesis, improved mitochondria function, and reduced oxidative stress in skeletal muscle and heart [ 42 ]. In addition, in a pilot study, we demonstrated that supplementation of EAAs to Doxo-treated animals preserved cardiomyocytes architecture and improved the efficacy of chemotherapy [ 14 , 49 ]. More recently, we showed that a diet rich in EAAs favorably modified cell metabolism in different organs and experimental conditions [ 50 , 51 ], promoting survival [ 36 ].…”

Is the Response of Tumours Dependent on Dietary Input of Some Amino Acids or on the Ratios Among Essential and Non-Essential Amino Acids? All That Glisters is Not Gold

“…The proteasome inhibition exerted by EAA, different from that exerted by drugs like bortezumib or carfilzomib [ 22 ], is peculiarly evident and limited to cancer cells. Furthermore EAA have already been shown to be protective against doxorubicin heart damages [ 23 ], characterized by disregulation of mitochondria and autophagy [ 24 ], and being also efficient in ameliorating heart performance in aged diabetic patients affected by heart failure [ 25 ]. Introduction of serine and ornithine-αKG, in specific ratios with EAA, seems efficient to further implement the effects of altering environmental EAA/NEAA, and this not so predictable result is peculiarly efficient in eliciting activity and control of the endoplasmic reticulum proteases, and specifically by inducing cathepsin-L, mainly involved in triggering autophagy.…”