Design, synthesis, and biological evaluation of indazole-based PLK4 inhibitors
Cunzheng Fan1, Nian Liu1, Ningyuan Hu1
1Key Laboratory of Structure-Based Drug Design and Discovery, Ministry of Education, School of Pharmaceutical Engineering, Shenyang Pharmaceutical University 103 Wenhua Road, Shenhe District Shenyang 110016 PR China medchemzhao@163.com.
Abstract:
Polo-like kinase 4 (PLK4), a member of the serine/threonine protein kinase family, serves as a central regulator of centriole duplication and plays a critical role in eukaryotic mitosis. Overexpression of PLK4 in several cancer types underscores its potential as a therapeutic target. In our previous studies, compound 28t demonstrated acceptable kinase inhibitory activity but exhibited poor cellular activity. Consequently, 28t was selected as a lead compound for further optimization. Through functional group migration and rational drug design strategies, we conducted structural modifications that ultimately yielded 23 novel indazole-based PLK4 inhibitors. Among these, compound C05 exhibited exceptional kinase inhibitory activity (IC50 < 0.1 nM). At the cellular level, C05 demonstrated potent antiproliferative effects against IMR-32 (neuroblastoma), MCF-7 (breast cancer), and H460 (non-small cell lung cancer) cell lines, with IC50 values of 0.948 μM, 0.979 μM, and 1.679 μM, respectively. Notably, compound C05 demonstrated favorable kinase selectivity towards PLK4 among the 10 kinases tested, achieving an inhibition rate of 87.45%. Further pharmacological experimental studies, including apoptosis induction, cell cycle arrest analysis, and clonogenic formation experiments, revealed that C05 outperformed the positive control LCR-263 in both potency and efficacy. Western blot analysis demonstrated that compound C05 effectively suppressed PLK4 autophosphorylation at 4 μM. Unfortunately, compound C05 demonstrated poor metabolic stability in human liver microsomes (HLMs), exhibiting a short half-life (T 1/2) of 2.69 minutes under standard incubation conditions. Notwithstanding the suboptimal metabolic stability, the compelling biological activity profile of compound C05 warrants further structural refinement.
Insights
Novel indazole-based compounds were developed to inhibit Polo-like kinase 4 (PLK4), a key regulator of cell division and potential cancer target. Compound C05 shows potent anticancer activity and selectivity, despite poor metabolic stability, indicating promise for further drug development.
Area of Science:
- Medicinal Chemistry
- Molecular Biology
- Cancer Research
Background:
- Polo-like kinase 4 (PLK4) is crucial for centriole duplication and mitosis, and its overexpression is linked to various cancers, making it a therapeutic target.
- Previous lead compound 28t showed kinase inhibition but lacked cellular efficacy, necessitating optimization.
Purpose of the Study:
- To design and synthesize novel indazole-based inhibitors targeting PLK4.
- To evaluate the in vitro kinase inhibitory activity, cellular antiproliferative effects, and pharmacological profile of the synthesized compounds.
Main Methods:
- Rational drug design and functional group modifications were employed to synthesize 23 novel indazole derivatives.
- In vitro kinase inhibition assays, cell-based proliferation assays (IMR-32, MCF-7, H460), kinase selectivity profiling, apoptosis induction, cell cycle arrest analysis, and Western blot were performed.
- Metabolic stability was assessed using human liver microsomes (HLMs).
Main Results:
- Compound C05 emerged as a potent PLK4 inhibitor with IC50 < 0.1 nM and demonstrated significant antiproliferative effects against neuroblastoma, breast, and lung cancer cell lines (IC50s ~1 μM).
- C05 exhibited high selectivity for PLK4 over other kinases and outperformed the positive control LCR-263 in cellular assays, including apoptosis induction and cell cycle arrest.
- Western blot confirmed C05's ability to inhibit PLK4 autophosphorylation, but it displayed poor metabolic stability in HLMs (T1/2 = 2.69 min).
Conclusions:
- Compound C05 is a highly potent and selective PLK4 inhibitor with promising anticancer activity.
- Despite its suboptimal metabolic stability, C05's strong biological profile warrants further structural optimization for potential therapeutic applications in cancer treatment.
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