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Melatonin and andrographolide synergize to inhibit the colospheroid phenotype by targeting Wnt/beta-catenin signaling

Published on Friday, 18 August 2023

Abstract

β-catenin signaling, and angiogenesis are associated with colospheroid (CSC), development. CSCs, spheroids derived from colon cancer cells, are responsible for metastasis, drug resistance, and disease recurrence. Whether dysregulating β-catenin and inhibiting angiogenesis reduce CSC growth is unknown.

In this study, the molecular mechanism of CSC growth inhibition was evaluated using a novel combination of melatonin (MLT) and andrographolide (AGP). These drugs have anticarcinogenic, antioxidant, and antimetastatic properties.

CSCs were obtained from two metastatic colon cancer cell lines (HT29 and HCT-15). The viability and stemness were monitored (FDA propidium iodide staining and immunoblot for CD44, CD133, Nanog, Sox2, and Oct4). The drug combination synergistically diminished stemness via increased reactive oxygen species (ROS) levels, reduced mitochondrial membrane potential and ATP level. MLT + AGP induced cell death by inhibiting β-catenin expression and its downregulatory signals, Cyclin D1, c-Myc. MLT + AGP treated cells exhibited translocation of phospho-β-catenin to the nucleus and dephosphorylated-β-catenin. Downregulation of β-catenin activation and its transcription factors (TCF4 and LEF1) and GTP binding/G-protein related activity were found in the dual therapy.

Angiogenic inhibition is consistent with downregulation of VEGF messenger RNA transcripts (VEGF189), phosphorylated VEGF receptor protein expression, matrigel invasion, and capillary tube inhibition.

In vivo, the intravenous injection of MLT + AGP slowed HT29 metastatic colon cancer.

Histopathology indicated significant reduction in microvascular density and tumor index. Immunohistochemistry for caspase 7, and β-catenin found increased apoptosis and downregulation of β-catenin signals.

The mechanism(s) of decreased colospheroids growth were the inhibition of the Wnt/β-catenin pathway.

Our results provide a rationale for using MLT in combination with AGP for the inhibition of CRCs.

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About this publication.

- The Di Bella Method (A Fixed Part - Melatonin tablets. From 30-40mg/day up to 200mg/day orally in patients with advanced stage of cancer disease and/or patients without respond to traditional treatments);

- Melatonin use in cancer patients have started in 1974, when melatonin prepared according to Prof. Di Bella’s formulation [...]. For 11 days was administered to the patient, admitted to the general medical ward at the Maggiore-Pizzardi Hospital in Bologna, very slowly (over approx. 8 hours) and intravenously administered 1000 mg of melatonin for 11 days. During the course of each day, the patient was intravenously administered 4 saline drips of 500 ml, each containing ten 25 mg bottles of freeze-dried melatonin, lasting 2 hours, totaling 1000 mg per day. No other drug of any kind was administered in order to ascertain the effect of the MLT without interference [...]. From Melatonin with adenosine solubilized in water and stabilized with glycine for oncological treatment - technical preparation, effectivity and clinical findings;

- About Melatonin - In vitro, review and in vivo publications;

- Publication: Melatonin anticancer effects: Review (from Di Bella's Foundation);

- Publication: Key aspects of melatonin physiology: 30 years of research (from Di Bella's Foundation);

- The Di Bella Method (A Fixed Part - Cyclophosphamide 50mg tablets and/or Hydroxyurea 500mg tablets, one or two per day);

- The Di Bella Method (A Fixed Part - Somatostatin, Octreotide, Sandostatin LAR, analogues and/or derivatives);

- Somatostatin in oncology, the overlooked evidences - In vitro, review and in vivo publications;

- Publication, 2018 Jul: Over-Expression of GH/GHR in Breast Cancer and Oncosuppressor Role of Somatostatin as a Physiological Inhibitor (from Di Bella's Foundation);

- Publication, 2018 Sep: The over-expression of GH/GHR in tumour tissues with respect to healthy ones confirms its oncogenic role and the consequent oncosuppressor role of its physiological inhibitor, somatostatin: a review of the literature (from Di Bella's Foundation);

- Publication, 2019 Aug: The Entrapment of Somatostatin in a Lipid Formulation: Retarded Release and Free Radical Reactivity (from Di Bella's Foundation);

- Publication, 2019 Sep: Effects of Somatostatin and Vitamin C on the Fatty Acid Profile of Breast Cancer Cell Membranes (from Di Bella's Foundation);

- Publication, 2019 Sep: Effects of somatostatin, curcumin, and quercetin on the fatty acid profile of breast cancer cell membranes (from Di Bella's Foundation);

- Publication, 2020 Sep: Two neuroendocrine G protein-coupled receptor molecules, somatostatin and melatonin: Physiology of signal transduction and therapeutic perspectives (from Di Bella's Foundation);