New perspective on the metabolism of AD-1 in vivo: Characterization of a series of dammarane-type derivatives with novel metabolic sites and anticancer mechanisms of active oleanane-type metabolites.

20(R)-25-methoxyl-dammarane-3ß,12ß,20-triol (AD-1, CN Patent: 201010107476.7) is a novel derivative of dammarane-type ginsenoside. AD-1 has been shown to inhibit cancer cell proliferation without significant host toxicity in vivo, and has excellent development potential as a new anti-cancer agent. This study was designed systematically to explore the metabolic pathway of ginseng sapogenins. The metabolism of drugs in the body is a complex biotransformation process where drugs are structurally modified to different molecules (metabolites) through various metabolizing enzymes. The compounds responsible for the effects of orally administered ginseng are believed to be metabolites produced in the gastrointestinal tract, so understanding the metabolism of the drug candidate can help to optimize its pharmacokinetics. In this study, faeces samples were collected and extracted after oral administration of AD-1. The 16 metabolites of AD-1 were isolated and identified for the first time with various chromatographic techniques, including semi-preparative high performance liquid chromatography, nuclear magnetic resonance spectroscopy, and mass spectrometry; of these 16 metabolites, 10 were novel compounds. We first discovered the biotransformation of dammarane-type sapogenins into oleanane-type sapogenins in rats and found a series of metabolites that changed, mainly at C-25 and C-29. This study provides new ideas for the metabolic pathway of ginseng sapogenins. The isolated compounds were screened for their effect on the viability and proliferation against cancer cell lines (Human A549, MCF-7, HELA, HO-8901 and U87). The discovery of novel active metabolites 3ß,12ß,21α,22ß-Hydroxy-24-norolean-12-ene (M6) may lead to a new or improved drug candidate. For one, M6 could inhibit the growth of all the tested cancer cells. Among the tested cell lines, M6 exhibited the most remarkable inhibitory effect on ovarian cancer HO-8901 cells, with IC50 value of 2.086 µM. On this basis, we studied the anticancer mechanisms of M6. The results indicated that the pro-apoptotic feature of M6 acts via a mitochondrial pathway. Our results indicated that M6 exhibited a higher inhibitory effect on cancer-cell proliferation than AD-1 by inducing cell apoptosis. Our work provides data for future investigations on the metabolic mechanism of AD-1 in vivo and the potential for future research on developing a new drug.

Synthesis, characterization and cytotoxic activity evaluation of ginsengdiol oxidation and nitrogen hybrid derivatives.

Panaxadiol (PD), a diol-type ginseng saponin, with a dammarane skeleton plays a potential role in the apoptosis of tumor cells. In this study, 28 oxidation and nitrogen hybrid derivatives of PD were synthesized, of which 20 were novel compounds. All the obtained compounds were screened for their cytotoxic activity in six cell lines. As compared with the positive control, some compounds showed better anti-proliferative activities while having much weaker effect on the growth of normal cells. Among them, ring-A fused pyrazoline of PD (1j) displayed impressive cytotoxic activity with IC50 9.62 ± 1.34, 11.65 ± 1.71, and 13.45 ± 1.60 µM against A549, HeLa and 8901 cell lines, respectively. Additionally, compound 2f exhibited the most potent activity with an IC50 value of 8.93 ± 1.11 µM against cell line A549. Therefore, our results indicated that 1j and 2f can be promising lead candidates for further studies.

Synthesis and Anticancer Activity Evaluation of Hydrolyzed Derivatives of Panaxnotoginseng Saponins.

To increase the antitumor activity of ginsenosides and acetylsalicylic acid, acid hydrolysis products of Panaxnotoginseng saponin were used as raw materials to be combined with salicylic acid to obtain ginsenoside salicylic acid derivatives. All derivatives were assessed for anti-cancer activity. A total of 20 target compounds were designed and synthesized. The cytotoxic activity on five cancer cell lines, including human colon cancer (HT-29), gastric cancer (BGC-823), cervical cancer (Hela), human breast cancer (MCF-7), human lung cancer cells (A549), and two normal cancer cell lines (human gastric epithelial cells (GES-1), and human ovarian epithelial cells (IOSE144)) was evaluated following treatment with the compounds. The results showed that all compounds inhibited the growth of cancer cells. Compounds 1a, 3a, 7a, 1b, 2b, 3b and 8b showed strong anticancer activity. For MCF-7 cells, compound 3b showed the strongest inhibitory activity, IC50 = 2.56 ± 0.09 µM. In the cytotoxicity test, all compounds showed low toxicity or no toxicity (IC50 > 100 µM). In addition, a cell cycle distribution assay and wound healing assay demonstrated that compound 3b specifically inhibited MCF-7 proliferation and migration ability. Our results indicate that compound 3b represents a promising compound for further cancer studies.

Non-protein amino acid derivatives of 25-methoxylprotopanaxadiol/25-hydroxyprotopanaxadioland their anti-tumour activity evaluation.

As active components of ginseng, 25-methoxylprotopanaxadiol and 25-hydroxyprotopanaxadiol exhibited an ability to inhibit the growth and proliferation or to induce the differentiation and apoptosis of tumour cells. We modified 25-OCH3-PPD and 25-OH-PPD with non-protein amino acids and a series of derivatives was obtained by chromatographic separation, purification and spectroscopy analysis. Thirteen derivatives of 25-OCH3-PPD (compounds 1-13) and 12 derivatives of 25-OH-PPD (compounds 14-25) were synthesised. The anti-cancer activities of the derivatives were evaluated on HCT-116 and BGC-823 cell lines by MTT assay. Compound 9 and compound 14 exhibited considerable anti-tumour activity for HCT-116 and BGC-823 cell lines, exhibited higher cytotoxic activity than 25-OCH3-PPD and 25-OH-PPD. Therefore, these ginsenoside derivatives could be used as potential lead for the development of a new type of anticancer agent.

Water-soluble derivatives of 25-OCH3-PPD and their anti-proliferative activities.

(20R)-25-Methoxyl-dammarane-3ß,12ß,20-triol (25-OCH3-PPD, AD-1) is a dammarane-type sapogenin showing anti-tumor potential. In the search for new anti-tumor agents with higher potency than our previously identified compound 25-OCH3-PPD, 11 novel sulfamic acid and diacid derivatives that could improve water solubility and contribute to good drug potency and pharmacokinetic profiles were designed and synthesized. Their in vitro anti-tumor activities in MCF-7, A-549, HCT-116, and BGC-823 cell lines and one normal cell line were tested by standard MTT assay. Results showed that compared with compound 25-OCH3-PPD, compounds 1, 4, and 5 exhibited higher cytotoxic activity on almost all cell lines, together with lower toxicity in the normal cell. In particular, compound 1 exhibited the best anti-tumor activity in the in vitro assays. The water solubility of 25-OCH3-PPD and its derivatives was tested and the results showed that the solubility of 25-OCH3-PPD sulfamic acid and diacid derivatives were better than that of 25-OCH3-PPD in water, which may provide valuable data for the research and development of new anti-tumor agents.