Antiproliferative effects of curcuma purpurascens on H103 and its interaction mechanism with human serum albumin / Nurul Jannah Mohd Asngari

Nurul Jannah , Mohd Asngari (2024) Antiproliferative effects of curcuma purpurascens on H103 and its interaction mechanism with human serum albumin / Nurul Jannah Mohd Asngari. PhD thesis, Universiti Malaya.

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Abstract

Curcuma purpurascens (C. purpurascens) commonly known as Temu tis is extensively utilised in Southeast Asian for culinary and therapeutic applications of diverse ailments, including cancer. In the present study, the possible mechanisms underlying the cytotoxic effects of C. purpurascens extracts against H103 human oral squamous carcinoma cells were investigated. The cytotoxic effects of the crude methanol and fractionated extracts (hexane, chloroform, ethyl acetate and water) of C. purpurascens against several human oral cancer cell lines and non-cancer gingival fibroblast cells were conducted using the MTT assay. The C. purpurascens chloroform fractionated extract (CPCE) was selected for further investigations on the induction of apoptosis as it exhibits high cytotoxicity effect against H103 cancer cells. The CPCE was also selected for further investigations on the interactions with human serum albumin (HSA) along with ar-turmerone and odanacatib (ODN) as comparison using spectroscopic, microscopic, and molecular docking approaches. Morphological observations by an inverted phase contrast microscope and double staining assay in the treated H103 cancer cells with the CPCE exhibited typical apoptotic morphology such as cell shrinkage, chromatin condensation and membrane blebbing. Flow cytometry analysis of CPCE-treated H103 cancer cells showed that apoptosis was triggered by the externalization of phosphatidylserine, significant disruption of mitochondrial membrane potential and DNA damage. The activation of caspases-3, -8 and -9 implied the trigger of both extrinsic and intrinsic pathways in the CPCE-treated H103 cancer cells. Furthermore, CPCE was able to induce cell cycle arrest in H103 cancer cell at G2/M phase. Both CPCE and ar-turmerone binding to HSA quenched the protein fluorescence intensity, but only HSA spectrum at 90 μM ar-turmerone yielded a 2 nm blueshift. In contrast, ODN binding to HSA resulted in fluorescence enhancement and 4 nm blueshift. Since CPCE is composed of many compounds, it binds to HSA with a low binding constant (Ka) value of (1.27±0.05) × 102 mg/l. A moderate binding affinity was obtained for ar-turmerone-HSA and ODN-HSA binding, with Ka values of 104 M−1. CPCE and ODN binding to HSA caused the hyperchromic effect and ar-turmerone binding caused the hypochromic effect of the HSA’s absorption mainly near the Trp residue, indicating a static mechanism interaction. Circular dichroism results suggested that the overall secondary and tertiary structures of HSA remained unaltered or slightly altered upon CPCE, ar-turmerone and ODN binding. The presence of CPCE, ar-turmerone and ODN causes aggregation on the HSA’s surface morphology confirming the complex formations. Ar-turmerone and ODN showed competitive behaviour as they preferably bind to site III in subdomain IB of HSA. Their interactions were mainly stabilised by hydrogen bonds, hydrophobic and van der Waals forces. Conclusively, these findings suggest CPCE possesses a potential therapeutic role in treating oral cancer. The characterisation of CPCE, ar-turmerone and ODN binding to HSA provide a better understanding on the interaction mechanism of these ligands in blood circulation and their pharmacokinetics properties that may enhance their solubility, reduce the side effects and help in designing new therapeutic drugs for various diseases, particularly in cancer treatments.

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