Anti-tumor effects and 3D-quantitative structure-activity relationship analysis of bufadienolides from toad venom.

Toad venom (venenum bufonis, also called Chan'su) has been widely used for centuries in China to treat different diseases, especially for cancer. Bufadienolides are mainly responsible for the anti-cancer effects of toad venom. However, systematic chemical composition and cytotoxicity as well as key pharmacophores of these bufadienolides from toad venom have not yet been defined clearly. To enrich the understanding of the diversity of bufadienolides and to find bufadienolides with better activities from toad venom. This study was carried out to isolate chemical constituents, research their anti-tumor effects and mechanisms by MTT assay, flow cytometry and Western blotting, and develop a CoMFA and CoMSIA quantitative structure-activity relationship (QSAR) model for illustrating the vital relationship between the chemical structures and cytotoxicities. Among 47 natural bufadienolides, most of bufadienolides (21 compounds isolated in this study and 26 compounds isolated previously) could significantly inhibit the proliferation of cancer cells, and compounds 1, 8, 12, 18 and 19 showed the most potent inhibitory activity against four types of human tumor cells. Compound 18 induced G2/M cell cycle arrest and apoptosis. Moreover, 3D contour maps generated from CoMFA and CoMSIA identified several pharmacophores of bufadienolides responsible for the anti-tumor activities. Our study might provide reliable information for future structure modification and rational drug design of bufadienolides with anticancer activities in medical chemistry.

Novel Oxazolidinone Antibacterial Analogues with a Substituted Ligustrazine C-ring Unit.

A series of novel oxazolidinone compounds with a substituted ligustrazine C-ring unit and different substituted groups at the C-5 side chain were designed and synthesized using linezolid as a lead and based on a scaffold hopping strategy. Their antibacterial and anti-inflammatory activities were evaluated. The results of in vitro antibacterial assays showed that all fourteen target compounds displayed potent activity against Gram-positive pathogens, particularly 8b, 13b, 14a, 14b, 15a, and 15b. Moreover, 14a and 14b exhibited significant inhibitory activities on the production of inflammatory mediators, including nitric oxide, interleukin-6, and tumor necrosis factor-alpha. Thus, these derivatives could serve as valuable candidates to develop anti-infective agents for the treatment of chronic wounds.

Bipiperidinyl derivatives of 23-hydroxybetulinic acid reverse resistance of HepG2/ADM and MCF-7/ADR cells.

Multidrug resistance (MDR) is a major obstacle to successful chemotherapy for cancer; thus, novel MDR reversers are urgently needed. In the present study, we assessed whether two synthetic derivatives of 23-hydroxybetulinic acid, 3,23-O-diacetyl-17-1,4'-bipiperidinyl betulinic amide (DABB) and 3,23-O-dihydroxy-17-1,4'-bipiperidinyl betulinic amide (DHBB), could reverse MDR induced by ATP-binding cassette (ABC) transporters. Using the MTT assay, we found that DABB and DHBB could enhance the cytotoxicities of ABCB1 substrates doxorubicin, vincristine, and paclitaxel in ABCB1-overexpressing HepG2/ADM and MCF-7/ADR cells, whereas that of a non-ABCB1 substrate cisplatin was unaffected. The ABCB1 substrate accumulation and efflux assay showed that DABB and DHBB not only enhanced the retention of doxorubicin and rhodamine123 but also inhibited the efflux of rhodamine123. Further mechanistic studies by reverse transcription PCR, western blot, and ABCB1 ATPase activity assay indicated that DABB and DHBB suppressed ABCB1 ATPase activity, but did not alter mRNA or protein expression of ABCB1. ABCB1 siRNA pretreatment attenuated the reversal effect of DABB and DHBB, indicating that their reversal effects were partially dependent on ABCB1. Docking analysis also implied that DABB and DHBB bind directly to ABCB1 at a site partly overlapped with that of verapamil. Taken together, our findings suggest that two bipiperidinyl derivatives of 23-hydroxybetulinic acid reverse ABCB1-mediated MDR through modulation of ABCB1 ATPase activity, thereby inhibiting its efflux function in both HepG2/ADM and MCF-7/ADR cells. These findings may contribute toward the development of novel MDR reversers using DABB and DHBB as adjuvant anticancer chemotherapy.

BBA, a derivative of 23-hydroxybetulinic acid, potently reverses ABCB1-mediated drug resistance in vitro and in vivo.

23-O-(1,4'-Bipiperidine-1-carbonyl)betulinic acid (BBA), a synthetic derivative of 23-hydroxybetulinic acid (23-HBA), shows a reversal effect on multidrug resistance (MDR) in our preliminary screening. Overexpression of ATP-binding cassette (ABC) transporters such as ABCB1, ABCG2, and ABCC1 has been reported in recent studies to be a major factor contributing to MDR. Our study results showed that BBA enhanced the cytotoxicity of ABCB1 substrates and increased the accumulation of doxorubicin or rhodamine123 in ABCB1 overexpressing cells, but had no effect on non ABCB1 substrate, such as cisplatin; what's more, BBA slightly reversed ABCG2-mediated resistance to SN-38, but did not affect the ABCC1-mediated MDR. Further studies on the mechanism indicated that BBA did not alter the expression of ABCB1 at mRNA or protein levels, but affected the ABCB1 ATPase activity by stimulating the basal activity at lower concentrations and inhibiting the activity at higher concentrations. In addition, BBA inhibited the verapamil-stimulated ABCB1 ATPase activity and the photolabeling of ABCB1 with [(125)I] iodoarylazidoprazosin in a concentration-dependent manner, indicating that BBA directly interacts with ABCB1. The docking study confirmed this notion that BBA could bind to the drug binding site(s) on ABCB1, but its binding position was only partially overlapping with that of verapamil or iodoarylazidoprazosin. Importantly, BBA increased the inhibitory effect of paclitaxel in ABCB1 overexpressing KB-C2 cell xenografts in nude mice. Taken together, our findings suggest that BBA can reverse ABCB1-mediated MDR by inhibiting its efflux function of ABCB1, which supports the development of BBA as a novel potential MDR reversal agent used in the clinic.