Synthesis and Evaluation of New Pyrazoline Derivatives as Potential Anticancer Agents in HepG-2 Cell Line

Abstract

Cancer is a major public health concern worldwide. Adverse effects of cancer treatments still compromise patients' quality of life. To identify new potential anticancer agents, a series of novel pyrazoline derivatives were synthesized and evaluated for cytotoxic effects on HepG-2 (human liver hepatocellular carcinoma cell line) and primary hepatocytes. Compound structures were confirmed by ¹H-NMR, mass spectrometry, and infrared imaging. An in vitro assay demonstrated that several compounds exerted cytotoxicity in the micromolar range. Benzo[b]thiophen-2-yl-[5-(4-hydroxy-3,5-dimethoxy-phenyl)-3-(2-hydroxy-phenyl)-4,5-dihydo-pyrazol-1-yl]-methanone (b17) was the most effective anticancer agent against HepG-2 cells owing to its notable inhibitory effect on HepG-2 with an IC₅₀ value of 3.57 µM when compared with cisplatin (IC₅₀ = 8.45 µM) and low cytotoxicity against primary hepatocytes. Cell cycle analysis and apoptosis/necrosis evaluation using this compound revealed that b17 notably arrested HepG-2 cells in the G₂/M phase and induced HepG-2 cells apoptosis. Our findings indicate that compound b17 may be a promising anticancer drug candidate.

Keywords: HepG-2 cells; anticancer activity; apoptosis; pyrazoline.

Figure 1

Structures of pyrazoloacridine (PZA) (I); doramapimod (BIRB-796) (II); axitinib (AG013736) (III); pazopanib (GW786034) (IV); tozasertib (VX-680) (V); and 3-(5’-hydroxymethyl-2’-furyl)-1-benzyl indazole (YC-1) (VI).

Scheme 1

General synthesis of compounds b1–19. Reagents and conditions: (i) substituted phenylethanone (0.01 mol), substituted benzaldehyde (0.012 mol), piperidine (1 mL), 160 °C, 20 min or 30% NaOH, ethanol, r.t., 24 h; (ii) ethanol, hydrazine hydrate, reflux, 4 h; (iii) substituted benzoyl chloride or substituted benzenesulfonyl chloride, 80 °C, ethanol, pyridine, 1 h; (iv) ethanol, phenylhydrazine, tetrabutylammonium bromide (TBAB), reflux, 1 h.

Figure 2

Inhibitory effects of compounds b15–17 and cisplatin on HepG-2 cells after 24 h and 48 h.

Figure 3

Effects of compound b17 on cell cycle progression in HepG-2 cells. Cells were treated with b17 at concentrations of 0 µM, 0.9 µM, 2.7 µM, and 4.5 µM for 24 h.

Figure 4

Compound b17 induced apoptosis in HepG-2 cells at concentrations of control, 0.9 µM, 2.7 µM, and 4.5 µM in a 12 h exposure. PI: propidium iodide.