- Title
- Design, synthesis and bioactivity screening of small molecules based on natural product scaffolds
- Creator
- Lei, Shuwen
- Relation
- University of Newcastle Research Higher Degree Thesis
- Resource Type
- thesis
- Date
- 2024
- Description
- Research Doctorate - Doctor of Philosophy (PhD)
- Description
- Natural products are valuable sources of biologically active compounds containing diverse structures, such as benzofuran, pyrazole, and purine. Scientists are now leveraging these structures to develop small molecules with medicinal properties, which have demonstrated breakthroughs in areas such as oncology, antimicrobial research, cardiovascular diseases, ophthalmic diseases, and neurodegenerative diseases. This dissertation comprises four sections dedicated to synthesising, screening and biology evaluating bioactive small molecules based on natural product scaffolds. By introducing structural modifications and functional groups, these small molecules are tailored to address recent, yet unresolved, targets such as Kirsten rat sarcoma viral oncogene homolog (KRAS) mutations in anti-tumour research, Sortase A in antimicrobial research, small C-terminal domain phosphatase 1 (Scp1) and Aldehyde Oxidase 1 (AOX1) to reduce oxidative damage and prevent skin cancer. The discovery of small molecule inhibitors targeting KRAS represents a significant breakthrough in cancer therapy. Based on the previous work, we used the structural scaffolds of natural products to design and synthesise a batch of pyrazole derivatives and purine derivatives. Most compounds did not show cytotoxicity on BEAS-2B cells, with some demonstrating moderate activity against KRAS mutation cell lines, exhibiting IC50 values ranging from 2.9 μM to 19.5 μM. Notably, compound XXIX exhibited inhibitory activity against H358, A549, and A427 cell lines, with IC50 values of 4.7 μM, 4.0 μM, and 3.0 μM, respectively, emerged as a promising pan-KRAS inhibitor, potentially serving as a SHP2 inhibitor or impacting the phosphoinositide 3-kinases (PI3K)-protein kinase B (AKT)-mammalian target of rapamycin (mTOR) pathway. Current single mutation-based KRAS inhibitors still face challenges in addressing resistance mechanisms and optimising efficacy across patient populations. These findings underscore the therapeutic potential of pan-KRAS inhibitors in various malignancies. Sortase A (SrtA) is a surface protein virulence factor. It functions as a cysteine transpeptidase found in the majority of gram-positive bacteria, playing a crucial role in anchoring surface proteins to the bacterial cell wall. Deletion of SrtA has been shown to mitigate infections without compromising bacterial viability. In this study, we assessed a series of benzofuranene cyanide derivatives and several compounds demonstrated notable inhibitory effects against SrtA. III-1, III-15, III-34 and V-1 exhibited potent inhibitory effects on biofilm formation, displaying half-maximal inhibitory (IC50) values ranging from 2.1 μM to 54.2 μM. Invasion assays revealed that these four compounds led to a reduction in S. aureus uptake by 293T cells, with decreases ranging from 4% to 24.0%. Moreover, III-15 was found to decrease the level of cell wall-associated protein A by 26.5%. Structure-activity relationship analyses and docking studies indicated that the acrylonitrile moiety of these compounds played a crucial role in enhancing their activity. Protein mass spectrometry showed that acrylonitrile, as a Michael acceptor, effectively inhibited SrtA activity by covalently binding to the thiol group of Cys184. Glioma, a highly aggressive primary brain tumour with limited treatment options, poses a significant challenge. Studies suggest that high levels of the RE-1 silencing transcription factor (REST) protein contribute to brain tumours and neurological diseases. However, REST is not an ideal drug target due to its role as a transcription factor. On the other hand, Scp1 triggers REST activation by dephosphorylating its phosphoserine residues, making it a potential drug target. This study aimed to discover more potent Scp1 inhibitors, for which a series of geniposide and genipin derivatives were investigated. It was found that compound 3-1d can cause a destabilisation of REST and lead to its degradation in a dose-dependent manner. As a result, the expression levels of REST-silenced genes such as DYRK1A, ELAVL1, USP37, CELSR3, CHGB, BDNF, and SCN2 genes were increased. The compound was tested on NODSCID mice with glioma for 14 days without any observed toxicity. It was found that the group treated with compound 3-1d experienced a significant decrease in tumour growth compared to the untreated group. On day 14, the area of luminescence measured in square centimetres was 2.4 times larger in the untreated group than in the treated group. Additionally, the average radiance efficiency (measured in p/s/cm2sr) was 1.9 times greater in the untreated group than in the treated group. Our findings provide the first evidence of the anti-glioma properties of Scp1 inhibitors both in vitro and in vivo. These findings offer hope for the development of more effective treatments for glioma. AOX1 is an attractive target for drug development and therapeutic strategies to combat oxidative damage. Effective AOX1 inhibitors require optimisation through in vitro analyses, which is essential for selecting potential inhibitors for in vivo and clinical investigation. Limited experimental models for drug development pose a challenge to clinical trials. Our study involved the screening of a mini compound library consisting of 287 compounds with different scaffolds. We used the p-DMAC assay to identify 79 compounds that exhibited better inhibition of human AOX1 activity than known AOX1 inhibitor raloxifene. We further narrowed our selection down to 7 active compounds, all of which demonstrated inhibition of both human and mouse liver extracts. Subsequently, we conducted tests to assess their cytotoxicity on human skin cells. After the testing, we selected 2 compounds, along with raloxifene, for further experiments. Through our experiments, we discovered that YX5 and YX128 have remarkable abilities to reduce the increase in 4-HNE levels induced by UVA irradiation and the increase in 8-OHdG levels induced by UVB irradiation. This demonstrated the compounds’ effectiveness in preventing oxidative damage to DNA/RNA and lipids, thereby preventing the occurrence of skin cancer. This dissertation successfully designed, synthesised, and screened small molecules based on natural active compound scaffolds, and conducted their bioactivity studies in the fields of antibacterial, antitumour, and antioxidant therapies. These findings form the basis for advancing research, development, and application of natural product scaffolds in the design and synthesis of small molecules. Through the introduction of different functional groups, they facilitate the development of small-molecule drugs, thereby opening up new avenues for drug discovery and therapeutic strategies.
- Subject
- natural product scaffolds; antitumour; antimicrobial; antioxidation
- Identifier
- http://hdl.handle.net/1959.13/1511241
- Identifier
- uon:56474
- Rights
- This thesis is currently under embargo and will be available from 25.09.2025. Copyright 2024 Shuwen Lei
- Language
- eng
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