Abstract B85: Plasmatic levels of arginine, its precursors and acylcarnitines affected by melatonin at specific time of the day
Authors: Junior Rubens Paula , Sonehara Nathália Martins , Zuccari Debora A. P. C. 
Published: May 2019

Abstract
The circadian rhythm regulates almost the whole metabolism modulating the concentration of various circulating substances through the clear and dark variation. Melatonin is the main mediator of this bidimensional flow, although studies have shown that metabolic diseases like cancer have already been associated to the imbalance of this rhythm. Actually, melatonin has several anticancer effects extensively described in the literature and proved by our group. Nowadays, metabolomics analysis has been used in cancer science to provide a comprehensive profile of the metabolic changes seeking to uncover characteristics about the tumor behavior. Studies in this field have discovered a lot of new biochemical pathways involved in the homeostasis and disorders. It is noticeable that important tumor features are regulated by essential molecules involved in bioenergy and cell structure, likewise amino acids and fatty acids. Therefore, the comprehension of specific metabolites that may be modulated by the exogenous melatonin is essential to verify its use as cancer adjuvant treatment. The aim of this study was to evaluate the metabolic profile in breast cancer xenograft model under treatment with melatonin across the circadian variation. We used 160 female Balb/c nude athymic mice separated into four groups: tumor/melatonin, tumor/vehicle, nontumor/melatonin, and nontumor/vehicle. The tumor model was developed by implantation of 3 x 106 triple-negative human breast cancer cells (MDA-MB-231) in the mammary fat pad. All the animals received 100 ul of solution by intraperitoneal injection (IP) for five days a week during 21 days. Treated groups received 40 mg/kg of melatonin and nontreated group only received vehicle solution. Blood sample were collected every 3 hours during 24 hours and the plasma was extracted for liquid chromatography and mass spectrometry (LC-MS) analysis using the Absolute IDQ p180 Kit (BioCrates). Our Orthogonal Partial Least Squares (OPLS) analysis showed evident separation of the metabolic profile not only because of the circadian cycle, but also because of the melatonin treatment. By looking for specific pathways we found that tumor/vehicle group showed significant increases in the plasmatic levels of arginine, citrulline, and ornithine, specifically at 6 am and 9 am (p &lt; 0.001). Arginine metabolism is well known to be strongly involved in the initial phase of tumor growth (Lind, 2004; HU et al., 2016), favoring the synthesis of molecules very important for various biologic processes of normal and tumor cells. Beyond the aminoacids we also found significantly raised levels of medium- and long-chain acylcarnitines, specifically at 3 pm (p &lt; 0.001). Acylcarnitines are mainly responsible for the fatty acid metabolism, which is a fundamental energy source. Synthesis of fatty acids is required for membrane synthesis and therefore for cell growth and proliferation. Our results show that tumor-bearing animals treated with melatonin have shown reduced plasma levels of these metabolites, achieving the same levels of non-tumor bearing animals. In summary, this work reveals a strong relationship between the metabolic profile and the entire circadian variation as well as the fact that exogenous melatonin can regulate the metabolites' concentration. This preclinical trial showed increased plasmatic levels of arginine, its precursors and acylcarnitines at specific time of the day, and the treatment with melatonin is able to decrease the serum levels of these metabolites. Thus our results corroborate the literature which characterizes these metabolites as cancer biomarkers and increasingly allows the use of melatonin as an adjuvant therapy for breast cancer.
Citation Format: Rubens Paula Junior, Nathália Martins Sonehara, Debora A. P. C. Zuccari. Plasmatic levels of arginine, its precursors and acylcarnitines affected by melatonin at specific time of the day [abstract]. In: Proceedings of the AACR International Conference held in cooperation with the Latin American Cooperative Oncology Group (LACOG) on Translational Cancer Medicine; May 4-6, 2017; São Paulo, Brazil. Philadelphia (PA): AACR; Clin Cancer Res 2018;24(1_Suppl):Abstract nr B85.