[Show abstract][Hide abstract] ABSTRACT: Angelica gigas Nakai (AGN) is a major medicinal herb used in Korea and several other Asian countries. Traditionally, its dried root has been used to treat anemia, pain, infection, and articular rheumatism, most often through boiling in water to prepare the dosage forms. AGN extract or AGN-containing herbal mixtures are sold in the USA and globally as dietary supplements for pain killing, memory enhancement, and post-menopausal symptom relief. Decursin (D) and its isomer decursinol angelate (DA) are the major chemicals in the alcoholic extracts of the root of AGN. The anti-cancer activity of AGN alcoholic extract has been established in a number of animal cancer models, including a transgenic model of prostate carcinogenesis. Cell culture structure-activity studies have uncovered distinct cellular and molecular effects of D and DA vs. their pyranocoumarin core decursinol (DOH) with respect to cancer cells and those associated with their microenvironment. Pharmacokinetic (PK) study by us and others in rodent models indicated that DOH is the major and rapid in vivo first pass liver metabolite of D and DA. Cognizant of metabolic differences among rodents and humans, we carried out a first-in-human PK study of D/DA to inform the translational relevance of efficacy and mechanism studies with rodent models. The combined use of rigorous animal tests and human PK studies can provide stronger scientific rationale to inform design and execution of translational studies to move AGN toward evidence-based herbal medicine.
[Show abstract][Hide abstract] ABSTRACT: The ethanol extract of Angelica gigas Nakai (AGN) root has promising anti-cancer and other bioactivities in rodent models. It is currently believed that the pyranocoumarin isomers decursin (D) and decursinol angelate (DA) contribute to these activities. We and others have documented that D and DA were rapidly converted to decursinol (DOH) in rodents. However, our in vitro metabolism studies suggested that D and DA might be metabolized differently in humans. To test this hypothesis and address a key question for human translatability of animal model studies of D and DA or AGN extract, we conducted a single oral dose human pharmacokinetic study of D and DA delivered through an AGN-based dietary supplement Cogni.Q (purchased from Quality of Life Labs, Purchase, NY) in twenty healthy subjects, i.e., 10 men and 10 women, each consuming 119 mg D and 77 mg DA from 4 vegicaps. Analyses of plasma samples using UHPLC-MS/MS showed mean time to peak concentration (Tmax) of 2.1, 2.4 and 3.3 h and mean peak concentration (Cmax) of 5.3, 48.1 and 2,480 nmol/L for D, DA and DOH, respectively. The terminal elimination half-life (t1/2) for D and DA was similar (17.4 and 19.3 h) and each was much longer than that of DOH (7.4 h). The mean area under the curve (AUC0-48h) for D, DA and DOH was estimated as 37, 335 and 27,579 h∙nmol/L, respectively. Gender-wise, men absorbed the parent compounds faster and took shorter time to reach DOH peak concentration. The human data supported an extensive conversion of D and DA to DOH, even though they metabolized DA slightly slower than rodents. Therefore, the data generated in rodent models concerning anti-cancer efficacy, safety, tissue distribution and pharmacodynamic biomarkers will likely be relevant for human translation.
PLoS ONE 02/2015; 10(2):e0114992. DOI:10.1371/journal.pone.0114992 · 3.23 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: As a potential novel agent for treating pancreatic cancer, methylseleninic acid (MSeA) was evaluated in cell culture and xenograft models. Results showed that MSeA induced G1 cell cycle arrest and apoptosis in a majority of human and mouse pancreatic cancer cell lines, but G2 arrest in human PANC-1 and PANC-28 cell lines. In contrast to our previous finding in human prostate cancer LNCaP cells having a lack of P53 activation by MSeA, induction of G2 arrest in PANC-1 cells was accompanied by increased mutant P53 Ser15 phosphorylation, upregulation of P53-targets P21Cip1 and GADD45 and G2 checkpoint kinase (Chk2) activation, suggestive of DNA damage responses. A rapid inhibition of AKT phosphorylation was followed by reduced mTOR signaling and increased autophagy in PANC-1 cells attenuating caspase-mediated apoptosis execution. Furthermore, daily oral treatment with MSeA (3 mg Se/kg body weight) significantly suppressed growth of subcutaneously inoculated PANC-1 xenograft in SCID mice. Immunohistochemical analyses detected increased p-Ser15 P53, P21Cip1, pS139-H2AX (DNA damage responses), and caspase-3 cleavage and decreased pSer473AKT and Ki67 proliferative index and reduced intratumor vascular density in MSeA-treated xenograft. These results provide impetus for further research of MSeA in the therapy and/or chemoprevention of pancreatic cancer.
Nutrition and Cancer 01/2014; 66(2). DOI:10.1080/01635581.2014.868911 · 2.47 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The aim of this study is to investigate and compare the metabolic rate and profiles of pyranocoumarin isomers decursin and decursinol angelate using liver microsomes from humans and rodents, and to characterize the major metabolites of decursin and decursinol angelate in human liver microsomal incubations using LC-MS/MS. First, we conducted liver microsomal incubations of decursin and decursinol angelate in the presence or absence of NADPH. We found that in the absence of NADPH, decursin was efficiently hydrolyzed to decursinol by hepatic esterase(s), but decursinol angelate was not. In contrast, formation of decursinol from decursinol angelate was mediated mainly by cytochrome P450(s). Second, we measured the metabolic rate of decursin and decursinol angelate in liver S9 fractions from mice and humans. We found that human liver S9 fractions metabolized both decursin and decursinol angelate more slowly than those of the mouse. Third, we characterized the major metabolites of decursin and decursinol angelate from human liver microsomes incubations using HPLC-UV and LC-MS/MS methods and assessed the in vivo metabolites in mouse plasma from a one-dose PK study. Decursin and decursinol angelate have different metabolite profiles. Nine metabolites of decursin and nine metabolites of decursinol angelate were identified in human liver microsome incubations besides decursinol using a hybrid triple quadruple linear ion trap LC-MS/MS system, and many of them were later verified to be also present in plasma samples from rodent PK studies.
[Show abstract][Hide abstract] ABSTRACT: Decursin and decursinol angelate are the major components in the alcoholic extract of the root of Angelica gigas Nakai. Our previous work convincingly demonstrated that both decursin and decursinol angelate were rapidly converted to decursinol in mice after administration by either oral gavage or i. p. injection. In the current study, we compared for the first time the plasma profiles of decursinol, when equal moles of decursin/decursinol angelate or decursinol were given to rats by oral gavage, and investigated the effect of different formulas and other chemicals in Angelica gigas extract on the bioavailability of decursinol. Our results show that gavage of decursinol led to a faster attainment of plasma decursinol peak (Tmax ~ 0.7 h) and much higher peak levels than an equal molar amount administered as decursin/decursinol angelate mixture or as Angelica gigas ethanol extract, resulting in 2-3 fold higher bioavailability as estimated by the area under the curve of the respective regimens (65 012 vs. 27 033 h · ng/mL for decursinol and decursin/decursinol angelate treatment groups, respectively). Compared to a formula based on ethanol-PEG400-Tween80, carboxyl methyl cellulose was a less optimized vehicle. In addition, we detected peak levels of decursin and decursinol angelate in the plasma of rats administered with decursin/decursinol angelate or Angelica gigas extract in the nM range (Tmax ~ 0.5 h) with a newly established sensitive UHPLC-MS/MS method. Furthermore, our data support the liver, instead of intestine, as a major organ site where decursin and decursinol angelate were hydrolyzed to decursinol with a S9 microsomal in vitro metabolism assay. Taken together, our study provided important PK, LC-MS/MS methodology, formulation and metabolism insights in a rodent model for the rational design of in vivo efficacy studies of the corresponding chemicals in the future.
[Show abstract][Hide abstract] ABSTRACT: Tanshinones are a class of abietane diterpene compound isolated from Salvia miltiorrhiza (Danshen or Tanshen in Chinese), a well-known herb in Traditional Chinese Medicine (TCM). Since they were first identified in the 1930s, more than 40 lipophilic tanshinones and structurally related compounds have been isolated from Danshen. In recent decades, numerous studies have been conducted to investigate the isolation, identification, synthesis and pharmacology of tanshinones. In addition to the well-studied cardiovascular activities, tanshinones have been investigated more recently for their anti-cancer activities in vitro and in vivo. In this review, we update the herbal and alternative sources of tanshinones, and the pharmacokinetics of selected tanshinones. We discuss anti-cancer properties and identify critical issues for future research. Whereas previous studies have suggested anti-cancer potential of tanshinones affecting multiple cellular processes and molecular targets in cell culture models, data from in vivo potency assessment experiments in preclinical models vary greatly due to lack of uniformity of solvent vehicles and routes of administration. Chemical modifications and novel formulations had been made to address the poor oral bioavailability of tanshinones. So far, human clinical trials have been far from ideal in their design and execution for the purpose of supporting an anti-cancer indication of tanshinones.
International Journal of Molecular Sciences 12/2012; 13(10):13621-66. DOI:10.3390/ijms131013621 · 2.86 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Previous studies suggested chalcones as antineoplastic drug candidates. We synthesized a new chalcone derivative (E)-3-(4-methoxyphenyl)-2-methyl-1-(3,4,5-trimethoxyphenyl)prop-2-en-1-one, (CHO27) with an up to 1000-fold increased cytotoxic potency relative to its parent compound in cell culture assays. CHO27 at low nanomolar levels, inhibited prostate cancer (PCa) cell growth through cell cycle arrest and caspase-dependent apoptosis. Activation of p53 accounted for, at least in part, the growth inhibition by CHO27 in vitro. Furthermore, i.p. administration of CHO27 suppressed the growth of established PCa 22Rv1 xenograft tumors accompanied with p53 and p21(Cip1) induction. CHO27 may be a lead for development of new therapeutic agents for PCa.
Anticancer research 09/2012; 32(9):3689-98. · 1.87 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Korean Angelica gigas Nakai (AGN) is a major medicinal herb used in Asian countries such as Korea and China. Traditionally, its dried root has been used to treat anemia, pain, infection and articular rheumatism in Korea, most often through boiling in water to prepare the dosage forms. The pyranocoumarin compound decursin and its isomer decursinol angelate (DA) are the major chemical components in the alcoholic extracts of the root of AGN. The in vitro anti-tumor activities of decursin and/or DA against prostate cancer, lung cancer, breast cancer, colon cancer, bladder cancer, sarcoma, myeloma and leukemia have been increasingly reported in the past decade whereas the in vivo efficacy in mouse models was established only for a few organ sites. Preliminary pharmacokinetics study by us and others in rodent models indicated that decursinol (DOH), which has much less in vitro direct anti-cancer activities by itself, is the major and rapid in vivo hydrolysis metabolite of both decursin and DA. Besides decursin, DA and DOH, other chemical components in AGN such as polysaccharides and polyacetylenes have been reported to exert anti-cancer and anti-inflammation activities as well. We systematically reviewed the published literature on the anti-cancer and other bio-activities effects of AGN extract and decursin, DA and DOH, as well as other chemicals identified from AGN. Although a number of areas are identified that merit further investigation, one critical need is first-in-human studies of the pharmacokinetics of decursin/DA to determine whether humans differ from rodents in absorption and metabolism of these compounds.
[Show abstract][Hide abstract] ABSTRACT: Our group previously reported that tanshinone IIA induced apoptosis via a mitochondria dependent pathway in LNCaP prostate cancer cells. In the present study, the roles of androgen receptor (AR) and p53 signaling pathways were investigated in tanshinone IIA-induced G1 arrest in LNCaP cells. Tanshinone IIA significantly inhibited the growth and proliferation of LNCaP cells by colony formation and BrdU incorporation assays, respectively. Tanshinone IIA induced cell cycle arrest at G1 phase and down-regulated cyclin D1, CDK2 and CDK4. Furthermore, tanshinone IIA activated the phosphorylation of p53 at Ser 15 residue and its downstream p21 and p27. Additionally, tanshinone IIA suppressed the expression of AR and prostate specific antigen (PSA). Conversely, silencing p53 using its specific siRNA reversed cyclin D1 expression inhibited by tanshinone IIA. However, knockdown of AR had no effect on the p53/p21/p27 signaling pathway activated by tanshinone IIA in LNCaP cells. In AR siRNA-transfected cells, tanshinone IIA did not cause cell cycle arrest and reduce cyclin D1, implying that AR is essential to induce G1 arrest by tanshinone IIA in LNCaP cells. Taken together, the findings suggest that tanshinone IIA induces G1 arrest via activation of p53 signaling and inhibition of AR in LNCaP cells.
Phytotherapy Research 05/2012; 26(5):669-74. DOI:10.1002/ptr.3616 · 2.40 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: To test whether tanshinones inhibit prostate cancer (PCa) growth at least in part through inhibiting androgen receptor (AR) signaling.
We evaluated cell growth, survival and AR signaling parameters of PCa cells after exposure to tanshinones in in vitro models. We also tested the in vivo inhibitory efficacy of tanshinone IIA (TIIA) against LNCaP xenograft model in athymic nude mice.
For androgen-dependent LNCaP cells, a colony growth assay showed strong inhibitory potency following the order of TIIA≈cryptotanshinone>tanshinone I, being 10-30 folds higher than Casodex (racemic). TIIA inhibited growth of LNCaP cells more than several androgen-independent PCa cell lines. All 3 tested tanshinones were devoid of AR agonist activity under castrate condition. Mechanistically, tanshinones inhibited AR nuclear translocation within 2 h, decreased protein and mRNA abundance of AR and its target prostate-specific antigen within 12 h, and stimulated proteosomal degradation of AR. Oral administration of TIIA (25 mg/kg, once daily) retarded LNCaP xenograft growth and down-regulated tumor AR abundance in athymic nude mice.
AR targeting action of tanshinones was distinct from Casodex and contributed to prostate cancer growth suppression in vitro and in vivo.
Pharmaceutical Research 01/2012; 29(6):1595-608. DOI:10.1007/s11095-012-0670-3 · 3.95 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Androgen receptor (AR) signaling is crucial for the genesis and progression of prostate cancer (PCa). We compared the growth responses of AR(+) LNCaP and LNCaP C4-2 vs. AR(-) DU145 and PC-3 PCa cell lines to galbanic acid (GBA) isolated from the resin of medicinal herb Ferula assafoetida and assessed their connection to AR signaling and cell cycle regulatory pathways. Our results showed that GBA preferentially suppressed AR(+) PCa cell growth than AR(-) PCa cells. GBA induced a caspase-mediated apoptosis that was attenuated by a general caspase inhibitor. Subapoptotic GBA downregulated AR protein in LNCaP cells primarily through promoting its proteasomal degradation, and inhibited AR-dependent transcription without affecting AR nuclear translocation. Whereas docking simulations predicted binding of GBA to the AR ligand binding domain with similarities and differences with the AR antagonist drug bicalutamide (Bic), LNCaP cell culture assays did not detect agonist activity of GBA. GBA and Bic exerted greater than additive inhibitory effect on cell growth when used together. Subapoptotic GBA induced G(1) arrest associated with an inhibition of cyclin/CDK4/6 pathway, especially cyclin D(1) without the causal involvement of cyclin-dependent kinase (CDK) inhibitory proteins P21(Cip1) and P27(Kip1) . In summary, the novelty of GBA as an anti-AR compound resides in the distinction between GBA and Bic with respect to AR protein turnover and a lack of agonist effect. Our observations of anti-AR and cell cycle arrest actions plus the anti-angiogenesis effect reported elsewhere suggest GBA as a multitargeting drug candidate for the prevention and therapy of PCa.
International Journal of Cancer 01/2012; 130(1):200-12. DOI:10.1002/ijc.25993 · 5.01 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The pyranocoumarin compound decursin and its isomer decursinol angelate (DA) are the major hydrophobic phytochemicals in the root of Angelica gigas Nakai (AGN, Korean Angelica), a major traditional medicinal herb. The ethanol extract of AGN and especially the purified decursin and DA have been shown to exhibit antitumor activities by our collaborative team and others. Although decursinol has been identified as a major hydrolysis metabolite of decursin and DA in vivo in previous pharmacokinetic studies with mouse and rat, other recently published results sharply disputed this conclusion. In this study, we set up a practical method for the concurrent analysis of decursin, DA, and decursinol in mouse plasma and tumor tissues by liquid-liquid extraction and HPLC-UV and applied the method to several animal experiments. Plasma or tumor homogenate was extracted directly with ethyl acetate. The extraction efficiency for decursin/DA (quantitated together) and decursinol was between 82-95 % in both mouse plasma and tumor homogenate. The lower limit of quantitation (LLOQ) was approximately 0.25 µg/mL for decursin/DA and 0.2 µg/mL for decursinol in mouse plasma. In a pilot pharmacokinetic study, male C57BL/6 mice were given a single dose of 4.8 mg decursin/DA mixture (~240 mg/kg) per mouse either by oral gavage or intraperitoneal injection. Maximum plasma concentrations for decursin/DA and decursinol were 11.2 and 79.7 µg/mL, respectively, when decursin/DA was administered via intraperitoneal injection, and 0.54 and 14.9 µg/mL via oral gavage. Decursin/DA and decursinol contents in the tumor tissues from nude mouse xenografts correlated very well with those in plasma. Overall, our results confirm the conclusion that the majority of decursin/DA hydrolyze to decursinol in rodent models with a tiny fraction remaining as the intact compounds administered.
[Show abstract][Hide abstract] ABSTRACT: The transgenic adenocarcinoma of mouse prostate (TRAMP) model is by far the most practical transgenic model for preclinical prostate cancer chemoprevention studies. It is critical to characterize the prostate lobe-specificity of lesion lineages to consolidate the advantages of this model and minimize its limitations for chemoprevention studies.
We dissected dorsolateral (DLP), ventral (VP), and anterior prostate (AP) lobes, and macroscopic tumors from 90 male C57BL/6J TRAMP mice at 22-24 weeks of age (WOA) and analyzed lesions by histological, biochemical and proteomic approaches. To determine whether methylseleninic acid (MSeA) led to a deletion of initiated cells, we gave oral MSeA to TRAMP mice from 5 to 23 WOA or from 5 to 15 WOA and analyzed lesions at 23 WOA.
All tumors (n = 18) were T-antigen(+), synaptophysin (SYP)(+), androgen-receptor(-), and E-cadherin(-) poorly differentiated neuroendocrine carcinomas (NE-Ca). They were traceable most frequently to VP (66.7%) and rarely to DLP (11.1%) and AP (5.6%) with an estimated life-time incidence of 1 out of 3 mice. In DLP, epithelial lesions ranged from mild-to-severe atypical hyperplasia, with T-antigen(+), SYP(-), androgen-receptor(+), and E-cadherin(+). Proteomic profiling revealed many molecular differences between VP and DLP. In MSeA experiment, 6 out of 19 (31.5%) mice developed NE-Ca in the control group, only 2 in each MSeA group of 17-18 mice (11.1-11.8%) bore a detectable NE-Ca.
The C57BL/6J TRAMP mouse represents at least two lineages of prostate carcinogenesis. Chemoprevention studies should incorporate this knowledge for efficacy assessment and molecular target validations.
The Prostate 09/2011; 71(13):1429-40. DOI:10.1002/pros.21360 · 3.57 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Targeting androgen receptor (AR) signaling with agents distinct from current antagonist drugs remains a rational approach to the prevention and treatment of prostate cancer (PCa). Our previous studies have shown that decursin and isomer decursinol angelate (DA), isolated from the Korean medicinal herb Angelica gigas Nakai, interrupt AR signaling and possess anti-PCa activities in vitro. In the LNCaP PCa cell model, these pyranoccoumarin compounds exhibit properties distinct from currently used antagonists (e.g., Casodex). However, both are rapidly de-esterified to decursinol, a partial AR agonist. We report here that a synthetic decursin analog, decursinol phenylthiocarbamate (DPTC), has greater in vivo stability than the parent compounds. DPTC-decursinol conversion was undetectable in mice. Furthermore, in LNCaP cells, DPTC decreased prostate specific antigen (PSA) expression, down-regulated AR abundance and mRNA and inhibited AR nuclear translocation. The effect of DPTC on AR and PSA mRNA and protein abundance was also observed in VCaP cells expressing wild type AR. DPTC inhibited growth of both PCa cell lines through G(1) cell cycle arrest and apoptosis, as did decursin and DA. Furthermore, i.p. administration of DPTC for 3 weeks suppressed the expression of AR target genes probasin and Nkx3.1 in mouse prostate glands. Overall, our data suggest that DPTC represents a prototype lead compound for development of in vivo stable and active novel decursin analogs for the prevention or therapy of PCa.
Investigational New Drugs 08/2011; 30(5):1820-9. DOI:10.1007/s10637-011-9738-x · 2.93 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Senescence is a permanent growth arrest and has been implicated as an efficient anti-carcinogenesis mechanism. The purpose of this study was designed to test the hypothesis that penta-1,2,3,4,6-O-galloyl-beta-D-glucose (PGG), a naturally occurring polyphonolic gallotannin compound, might induce this type of permanent growth arrest in cancer cells. Our results show, for the first time, that PGG-induced senescence-like S-phase arrest in HepG2, Huh-7 human hepatoma cells, and SKBr3 human breast cancer cells at sublethal doses, judged by cellular morphological changes, increased senescence-associated β-galactosidase (SA-β-gal) activity, together with loss of proliferative capacity after being released from the treatment. This senescence-like response was mediated by intracellular ROS generation, but was not attributed to p53 Ser15 phosphorylative activation and was uncoupled from the p21cip1 axis, which has been shown to mediate Pten loss-induced cellular senescence or oncogene-driven senescence. The findings of the present study implicate a novel mechanism of PGG action to induce an atypical cellular senescence, adding to its promise as a potential chemopreventive agent.
[Show abstract][Hide abstract] ABSTRACT: The transgenic adenocarcinoma of mouse prostate (TRAMP) is the most widely used transgenic model for prostate cancer chemoprevention studies. Although two lobe-specific lineages of carcinogenesis have been described, the molecular mechanisms are still poorly defined. Here, we concurrently profiled the proteome of dorsal-lateral (DLP) and ventral (VP) prostate lobes of both TRAMP and littermate WT C57BL/6 mice of 18 wk by 2-D LC-MALDI-TOF/TOF with iTRAQ labeling. A total of 483 and 748 proteins were identified at critical false discovery rates of 1 and 5%. In WT mice, 84 proteins were found to have different expression levels between DLP and VP. In TRAMP mice, 118 proteins significantly changed in DLP and/or VP during TRAMP carcinogenesis. Among them, 55 and 36 proteins were uniquely changed in DLP or VP lobe, respectively, and 27 proteins in both DLP and LP lobe. Ingenuity Pathway Analysis was able to segregate proteins changed in two lobes into different pathway networks. In addition to serving as reference for prostate proteomic profiles, our data suggest that different sets of proteins are involved in the carcinogenesis in DLP versus VP in the TRAMP model.
[Show abstract][Hide abstract] ABSTRACT: We have shown that, in contrast to selenomethionine (SeMet) or selenite, methylseleninic acid (MSeA) and Se-methylselenocysteine (MSeC) exert prostate cancer (PCa) inhibitory effect in preclinical models. Here we investigated the prostate proteome signatures of mice treated with each selenium (Se) form for hypothesis generation concerning their potential in vivo molecular targets and cancer risk modification. Nude mice bearing subcutaneous PC-3 xenografts were treated daily with each Se form (3 mg Se/kg) orally for 45 days. Five prostates were pooled from each group. Their proteomes were profiled by LC-MS/MS with iTRAQ labeling. Of the 1,088 proteins identified, 72 were significantly modulated by one or more Se forms. MSeA and MSeC each induced separate sets of tumor suppressor proteins and suppressed different onco-proteins. Proteins induced by selenite and shared with MSeC were related to energy metabolism (e.g., fatty-acid synthase), and those induced by SeMet included vimentin and heat-shock protein-70, favoring cancer growth. While proteome changes induced by MSeA were associated with PCa risk reduction, desirable risk-reducing signatures induced by MSeC were counterbalanced by risk-promoting patterns shared with selenite and SeMet. We propose that the balance of oncogenic vs. suppressor protein patterns in the prostate may impact the direction of PCa risk modification by a given selenium.
Nutrition and Cancer 05/2011; 63(5):778-89. DOI:10.1080/01635581.2011.563029 · 2.47 Impact Factor