Design and Characterization of a Natural Arf-GEFs Inhibitor Prodrug CHNQD-01255 with Potent Anti-Hepatocellular Carcinoma Efficacy In Vivo.

Brefeldin A (BFA), a well-known natural Arf-GEFs inhibitor, is effective against hepatocellular carcinoma (HCC), while the poor solubility, serious toxicity, and short half-life limit its potential. Herein, distinct corresponding prodrugs of BFA, including esters 1-15, carbonates 16-24 and 30-32, and carbamates 25-29, were synthesized and evaluated. CHNQD-01255 (16) with improved aqueous solubility (15-20 mg/mL) demonstrated favorable pharmacokinetic profiles. It behaved as expected by undergoing rapid conversion to BFA in vivo, and achieved sufficient high plasma exposure, prolonged half-life, as well as the improved bioavailability of BFA (F = 18.96%). Meanwhile, CHNQD-01255 significantly suppressed tumor growth (TGI = 61.0%) at a dose of 45 mg/kg (p.o.) in the xenograft model. Notably, the improved safety profile of CHNQD-01255 (MTD > 750 mg/kg, p.o.) was confirmed to be superior to that of BFA (MTD < 506 mg/kg). Overall, CHNQD-01255 may serve as a safe and effective new anti-HCC prodrug.

New brefeldin A-cinnamic acid ester derivatives as potential antitumor agents: Design, synthesis and biological evaluation.

Hepatocellular carcinoma (HCC) is the most common primary liver malignancy and ranks third in mortality rate worldwide. Brefeldin A (BFA, 1), a natural Arf1 inhibitor, qualifies extremely superior antitumor activity against HCC while its low aqueous solubility, poor bioavailability, and high toxicity have greatly hindered its translation to the clinic. Herein, a series of BFA-cinnamic acid ester derivatives was rationally designed and synthesized via introducing active cinnamic acid and its analogues into the structure of 1. Their in vitro cytotoxic activities on five cancer cell lines, including HepG2, BEL-7402, HeLa, Eca-109 and PANC-1, were evaluated using MTT assay. As expected, favorable cytotoxic activity was observed on majority of the mono-substituted derivatives. Especially, the most potent brefeldin A 4-O-(4)-dimethylaminocinnamate (CHNQD-01269, 33) with improved aqueous solubility, demonstrated the strong cytotoxic activity against HepG2 and BEL-7402 cell lines with IC50 values of 0.29 and 0.84 µM, respectively. More importantly, 33 performed low toxicity on normal liver cell line L-02 with the selectivity index (SI) of 9.69, which was more than 17-fold higher than that of 1. Results from mechanistic studies represented that 33 blocked the cell cycle in the G1 phase, and induced apoptosis via elevating reactive oxygen species (ROS) production and increasing expression of apoptosis-related proteins of HepG2 cells. Docking experiment also suggested 33 a promising Arf1 inhibitor, which was confirmed by the cellular thermal shift assay that 33 displayed a significant effect on the stability of Arf1 protein. Furthermore, 33 possessed high safety profile (MTD >100 mg/kg, ip) and favorable pharmacokinetic properties. Notably, the superior antiproliferative activity was verified in HepG2 tumor-bearing xenograft model in which 33 markedly suppressed the tumor growth (TGI = 46.17%) in nude mice at a dose of 10 mg/kg once a day for 16 d. The present study provided evidence of exploiting this series of highly efficacious derivatives, especially 33, for the treatment of HCC.

Brefeldin A delivery nanomicelles in hepatocellular carcinoma therapy: Characterization, cytotoxic evaluation in vitro, and antitumor efficiency in vivo.

Hepatocellular carcinoma (HCC) is one of the major cancers with high mortality rate. Traditional drugs used in clinic are usually limited by the drug resistance and side effect and novel agents are still needed. Macrolide brefeldin A (BFA) is a well-known lead compound in cancer chemotherapy, however, with poor solubility and instability. In this study, to overcome these disadvantages, BFA was encapsulated in mixed nanomicelles based on TPGS and F127 copolymers (M-BFA). M-BFA was conferred high solubility, colloidal stability, and capability of sustained release of intact BFA. In vitro, M-BFA markedly inhibited the proliferation, induced G0/G1 phase arrest, and caspase-dependent apoptosis in human liver carcinoma HepG2 cells. Moreover, M-BFA also induced autophagic cell death via Akt/mTOR and ERK pathways. In HepG2 tumor-bearing xenograft mice, indocyanine green (ICG) as a fluorescent probe loaded in M-BFA distributed to the tumor tissue rapidly, prolonged the blood circulation, and improved the tumor accumulation capacity. More importantly, M-BFA (10 mg/kg) dramatically delayed the tumor progression and induced extensive necrosis of the tumor tissues. Taken together, the present work suggests that M-BFA has promising potential in HCC therapy.

Semi-Synthesis, Cytotoxic Evaluation, and Structure-Activity Relationships of Brefeldin A Derivatives with Antileukemia Activity.

Brefeldin A (1), a potent cytotoxic natural macrolactone, was produced by the marine fungus Penicillium sp. (HS-N-29) from the medicinal mangrove Acanthus ilicifolius. Series of its ester derivatives 2-16 were designed and semi-synthesized, and their structures were characterized by spectroscopic methods. Their cytotoxic activities were evaluated against human chronic myelogenous leukemia K562 cell line in vitro, and the preliminary structure-activity relationships revealed that the hydroxy group played an important role. Moreover, the monoester derivatives exhibited stronger cytotoxic activity than the diester derivatives. Among them, brefeldin A 7-O-2-chloro-4,5-difluorobenzoate (7) exhibited the strongest inhibitory effect on the proliferation of K562 cells with an IC50 value of 0.84 µM. Further evaluations indicated that 7 induced cell cycle arrest, stimulated cell apoptosis, inhibited phosphorylation of BCR-ABL, and thereby inactivated its downstream AKT signaling pathway. The expression of downstream signaling molecules in the AKT pathway, including mTOR and p70S6K, was also attenuated after 7-treatment in a dose-dependent manner. Furthermore, molecular modeling of 7 docked into 1 binding site of an ARF1-GDP-GEF complex represented well-tolerance. Taken together, 7 had the potential to be served as an effective antileukemia agent or lead compound for further exploration.

Targeting Delivery Nanocarriers for (+)-Terrein to Enhance Its Anticancer Effects.

As a compound from marine fungi, (+)-terrein showed significant anticancer activity. In this study, (+)-terrein was extracted from the marine-derived fungus and showed significant cytotoxicity against cancer cells, especially in A549 cells. To enhance its anticancer effects, redox-responsive nanocarriers based on folic acid-chitosan decorating the mesoporous silica nanoparticles were designed to control (+)-terrein target delivery into cancer cells. (+)-Terrein was loaded in the holes, and folic acid-chitosan worked as a gatekeeper by disulfide linkage controlling (+)-terrein release in the tumor microenvironment. The (+)-terrein drug delivery systems exhibited cytotoxicity toward A549 cells through induction of apoptosis. The apoptosis effect was confirmed by the increase in the expression of cleaved caspase-3, caspase-9, and PARP. Taken together, this work evaluates for the first time the (+)-terrein delivery system and provides a promising nanomedicine platform for (+)-terrein.