Synthesis and Biological Evaluation of Novel Alkyl Amine Substituted Icariside II Derivatives as Potential Anticancer Agents

Abstract

A series of novel alkyl amine-substituted icariside II (ICA II) derivatives were synthesized by Mannich reactions at the 6-C position (compounds 4a⁻d) and changing the carbon chain length at the 7-OH position (compounds 7a⁻h), and their in vitro antitumor activity towards human breast cancer lines (MCF-7 and MDA-MB-231) and human hepatoma cell lines (HepG2 and HCCLM3-LUC) were evaluated by the MTT assay. Compared with ICA II, most of the twelve derivatives showed good micromole level activity and a preliminary structure-activity relationship (SAR) for the anticancer activity was obtained. Compound 7g showed the most potent inhibitory activity for the four cancer cell lines (13.28 μM for HCCLM3-LUC, 3.96 μM for HepG2, 2.44 μM for MCF-7 and 4.21 μM for MDA-MB-231), which was 2.94, 5.54, 12.56 and 7.72-fold stronger than that of ICA II. The preliminary SAR showed that the introduction of a alkyl amine substituent at 6-C was not favorable for the anticancer activity, while most of the 7-O-alkylamino derivatives exhibited good antitumor activity and the anticancer activity 7-O-alkylamino derivatives were influenced by the alkyl chain length and the different terminal amine substituents. Furthermore, the effects of compound 7g on apoptosis and cell cycle of MCF-7 cells were further investigated, which showed that compound 7g triggered apoptosis and arrested the cell cycle at the G0/G1 phase in MCF-7 cells. Our findings indicate that compound 7g may be a promising anticancer drug candidate lead.

Keywords: anticancer; apoptosis; icariside II; structure-activity relationship.

Figure 1

The structures of icariin (A) and icariside II (B).

Scheme 1

Reagents and conditions: (a) CH₃OH, HCl, reflux, 3 h; (b) CH₃CN, K₂CO₃, reflux, 12 h; (c) CH₃CN, K₂CO₃, reflux, 12 h.

Figure 2

The dose-response curves for MTT assay of compound 7d (A,B) on HCCLM3-LUC cells and HepG2 cells, compound 7g (C,D) on MCF-7 cells and MDA-MB-231 cells.

Figure 3

Fluorescent micrographs of Hoechst 33258 stained nuclear patterns when MCF-7 cells treated with compound 7g at 1–4 μM for 48 h (400×). The red arrows indicate the apoptotic cells.

Figure 4

Effect of compound 7g and ICA II on cell cycle distribution on MCF-7 cells. Cell cycle distribution after treatment with compound 7g and ICA II at 1–4 μM for 48 h.

Figure 5

Compound 7g induced apoptosis in MCF-7 cells. After treatment with compound 7g at different concentrations (1–4 μM), the apoptotic rates of MCF-7 cells were determined by Annexin V-APC and PI double staining using flow cytometry.