br b QingYunTang Biotech Beijing Co
b QingYunTang Biotech (Beijing) Co., Ltd., No. 14, Zhonghe Street, Beijing Economic-Technological Development Area, Beijing 100176, PR China *Corresponding author. E-mail address: [email protected](A. Liu).
Seleno-β-lactoglobulin (Se-β-Lg) was synthesized using seleninic acid, an organoselenium compound, and β-lactoglobulin (β-Lg), an important component of milk. Previously, we have studied the effects of Se-β-Lg on hepatocellular carcinoma and gastric cancer cells. In this study, we investigated the antitumor effects of Se-β-Lg and its potential mechanisms of action against human breast cancer Protease Inhibitor Cocktail (MCF-7 and MDA-MB-231). The results showed that the half-maximal inhibitory concentrations (IC50) of Se-β-Lg were 40.84 µg/mL for MCF-7 cells and 46.04 µg/mL for MDA-MB-231 cells at 24 h, while the compound showed no cytotoxicity to normal breast cells. The involvement of reactive oxygen species (ROS) in the activation of the apoptotic signaling pathway by Se-β-Lg was demonstrated by the incubation of cells with 80 µg/mL Se-β-Lg and determination of the rates of apoptosis and intracellular ROS levels after the addition of 10 mM N-acetyl-L-cysteine, a ROS inhibitor. Our findings revealed highly potent anticancer activities of Se-β-Lg against breast cancer cells and suggested that the compound may be used as a chemopreventive agent for breast cancer. Furthermore, we thoroughly elucidated the antitumor mechanism of Se-β-Lg.
Keywords: Seleno-β-lactoglobulin; Breast cancer; Cell apoptosis; Mitochondrial pathway
Breast cancer is one of the most common cancers and includes a heterogeneous group of diseases that affect an increasing number of women annually, with the highest mortality rate (Amaral et al., 2018; Joyce et al., 2017). Among all subtypes of breast cancers, triple-negative breast cancer (TNBC) accounts for about 15−20% of cases and is characterized by the absence of the expression of estrogen receptor, progesterone receptor (PR), and human epidermal growth factor type 2 receptors (Gucalp and Traina, 2016; Lin et al., 2012). Compared with other breast cancer subtypes, TNBC has higher rates of recurrence and metastasis (Liedtke et al., 2008). Treatment options for TNBC patients are limited because targeted therapies and endocrine therapies are ineffective (O'Reilly et al., 2015). Therefore, it is extremely important to find new anti-cancer drugs and therapeutic strategies, with minimal undesirable side effects.
As an essential and unique trace element, selenium plays an important role in health and immune activation as well as cancer treatment (Fernandes and Gandin, 2015). Moreover, Se-containing compounds act as pro-oxidants to maintain the stability of the oxidative balance in the cells, which has aroused extensive interest and attention of researchers (Skalickova et al., 2017). Compared with inorganic Se compounds, organic Se compounds exhibit stronger antitumor activities (Gandin et al., 2018) and lower toxicities (Chen et al., 2013). In our previous studies, L-arginine-seleninic acid (selenoarginine) was synthesized using inorganic selenium dioxide and arginine under vacuum and low-temperature conditions (Zheng et al., 2016). The compound showed significantly higher antioxidant activities than
selenium dioxide did against carbon tetrachloride-induced or alcohol-induced liver damage. β-Lactoglobulin (β-Lg), containing 162 amino acid residues, is one of the most widely studied proteins and the most abundant whey protein in cow milk (Kanakis et al., 2011). Similarly, β-Lg is one of the main allergens in the milk. However, the related study showed that hydrolysate of β-Lg could suppress the immunogenicity and maintain the original function to decrease the sensitization (Pescuma et al., 2019). β-Lg exhibits perfect nutritional, biochemical, and pharmacological properties, such as anti-cancer activities (Brandelli et al., 2015), angiotensin-converting effects (Prasanna and Saraswathi, 2017), and free radical scavenging (Zheng et al., 2016).
Therefore, the process was also applied to compound a novel drug seleno-β-lactoglobulin (Se-β-Lg), which has been proven to exert anti-tumor effects on hepatocellular carcinoma cell lines (Zhao et al., 2018), and gastric cancer cells (Yu et al., 2018). In previous studies, we have found that HepG2 cells were more sensitive than Hep3B cells to the treatment with Se-β-Lg (125, 250, and 500 µg/mL), and p53 was the key factor responsible for the difference in sensitivity. MGC-803 cells treated with Se-β-Lg (100, 200, and 400 µg/mL) underwent mitochondria-mediated apoptosis. However, the anti-tumor effects and the apoptotic pathway of Se-β-Lg in human breast cancer cells have not largely been elucidated.
Apoptosis, or programmed cell death, mainly involves the following pathways: the cell death receptor-mediated external apoptosis pathway, mitochondrial-mediated internal apoptosis pathway, and endoplasmic reticulum pathway (Wang et al., 2016). Induction of the apoptotic pathway depends on the production of reactive oxygen species (ROS), which play an important role in cell apoptosis and, to a certain extent, in cell growth. (Uchihara et al., 2018). The action