In silico and in vitro studies on the anti-cancer activity of andrographolide targeting survivin in human breast cancer stem cells

Abstract

Breast cancer stem cells (BCSCs) express high levels of the anti-apoptotic protein, survivin. This study aimed to discover a natural active compound with anti-cancer properties that targeted survivin in human breast cancer stem cells. From the seven examined compounds, andrographolide was selected as a lead compound through in silico molecular docking with survivin, caspase-9, and caspase-3. We found that the affinity between andrographolide and survivin is higher than that with caspase-9 and caspase-3. Human CD24-/CD44+ BCSCs were treated with andrographolide in vitro for 24 hours. The cytotoxic effect of andrographolide on BCSCs was compared to that on human mesenchymal stem cells (MSCs). The expression of survivin, caspase-9, and caspase-3 mRNA was analyzed using qRT-PCR, while Thr34-phosphorylated survivin and total survivin levels were determined using ELISA and Immunoblotting assay. Annexin-V/PI flow cytometry assays were performed to evaluate the apoptotic activity of andrographolide. Our results demonstrate that the CC50 of andrographolide in BCSCs was 0.32mM, whereas there was no cytotoxic effect in MSCs. Moreover, andrographolide decreased survivin and Thr34-phosphorylated survivin, thus inhibiting survivin activation and increasing survivin mRNA in BCSCs. The apoptotic activity of andrographolide was revealed by the increase of caspase-3 mRNA and protein, as well as the increase in both the early and late phases of apoptosis. In conclusion, andrographolide can be considered an anti-cancer compound that targets BCSCs due to its molecular interactions with survivin, caspase-9, and caspase-3, which induce apoptosis. We suggest that the binding of andrographolide to survivin is a critical aspect of the effect of andrographolide.

Andrographolide: A review of its pharmacology, pharmacokinetics, toxicity and clinical trials and pharmaceutical researches

“…Given that the toxicity and side effects has been acknowledged with the vast majority of chemotherapeutic agents, in the past few decades, researchers have paid increasing attention to the anticancer investigation of natural active ingredients, especially andrographolide (Islam et al, 2018; Malik et al, 2021; Soo et al, 2019). Published projects have validated the multiple anticancer effects, such as its ability to colon cancer (Banerjee et al, 2021; Vukmirovic, Vo, Seymour, Rollo, & Mothersill, 2021), lung cancer (Luo et al, 2021), cervical cancer (Pasha et al, 2021), breast cancer (Wanandi et al, 2020), hepatoma cancer (Shi, Zhang, Zheng, Lu, & Ji, 2017) as well as renal carcinoma (Bi et al, 2020) and so on, which the cytotoxic effects on these cancer cells above were bound up with inhibiting proliferation (Khan, Mahfooz, & Ansari, 2020), migration and invasion (Pearngam, Kumkate, Okada, & Janvilisri, 2019), blocking cell cycle (Khan, Mahfooz, Faisal, Alatar, & Ansari, 2020), inducing apoptosis (Wang et al, 2020), restraining tumor angiogenesis (Kajal et al, 2019), as well as enhancing chemosensitivity (Li et al, 2020; Mao, He, Wang, Wu, & Wei, 2019). The main anticancer mechanisms of andrographolide are depicted in Figure 4.…”

Molecular Docking as a Therapeutic Approach for Targeting Cancer Stem Cell Metabolic Processes

“…In this study, the authors compared 11 popular scoring functions, including four scoring functions of the LigFit module in Cerius2 (LigScore, PLP, PMF, and LUDI), four scoring functions of the CScore module in SYBYL (FScore, G-Score, D-Score, and ChemScore), the scoring function of the AutoDock program, and two stand-alone scoring functions (DrugScore and X-Score) by performing them on 100 protein−ligand complexes to scrutinize their performance and sift the most effective and efficient methods among them. In this regard, (Chan et al, 2013;Hongwiangchan et al, 2021;Jaitak, 2016;Liu et al, 2014;Madhunapantula et al, 2011;Ozsvari et al, 2017;Wanandi et al, 2020).…”

“…Herein, there will be a few examples in the context of some molecular docking usages in metabolic pathways such as 1) mitoketoscins application in targeting metabolic tumor promoters (OXCT1 and ACAT1) in both ketone re-utilization and mitochondrial function ( Ozsvari et al, 2017 ). 2) Survivin protein interaction with andrographolide, which can lead to having an influence on human breast CSCs apoptosis ( Wanandi et al, 2020 ). 3) S-phase kinase-associated protein-2 (Skp2) inhibition process by compound #25 which can result in CSCs survival suppression ( Chan et al, 2013 ).…”

“…CIN-RM can inhibit mTOR pathway. Abbreviations: ATP, Adenosine triphosphate; CIN-RM, Hydroquinone 5-O-cinnamoyl ester of renieramycin M; CSCs, Cancer stem cells; Hh, Hedgehog; mTOR, Mammalian target of rapamycin; Smo, smoothened ( Chan et al, 2013 ; Hongwiangchan et al, 2021 ; Jaitak, 2016 ; Liu et al, 2014 ; Madhunapantula et al, 2011 ; Ozsvari et al, 2017 ; Wanandi et al, 2020 ).…”

Existing and Emerging Breast Cancer Detection Technologies and Its Challenges: A Review

“…The tumor is classified as non-cancerous (benign) or cancerous (malignant). Benign tumors do not invade nearby tissues or spread to other areas of the body(metastasize) [12]. On the other hand, a malignant tumor consists of cancer cells that can invade and kill surrounding tissues and can attack different parts of the body.…”