Enhancing lathyrane structural diversity and MDR activity by combinatorial modification of lathyrane nucleus and ester side chain: A case study of Euphorbia Factor L1 and Euphorbia Factor L3.

The chemical transformation of lathyrane nucleus through reduction and oxidation reactions using Euphorbia Factor L1 (EFL1) and Euphorbia Factor L1 (EFL3) as examples were investigated, along with a co-modification strategy of lathyrane nucleus and its side ester chain. A total of 38 lathyrane derivatives (5-42) including 34 new compounds were obtained, which greatly enriched the structural diversity of the lathyrane-type diterpenoids. Cytotoxicity against drug-sensitive and drug (adriamycin, ADM) resistant MCF-7 cells showed that 23 out of 38 transformed derivatives possessed obvious cytotoxic activity with IC50 values ranging from 7.0 to 41.1 µM and 3.2 to 45.5 µM, respectively, against both cells, compared to the noncytotoxic EFL1 and EFL3. The multidrug resistance (MDR) reversing activities of these lathyrane derivatives were further evaluated in MCF-7/ADM. Three transformed compounds (reversal fold, RF = 151.33, 62.94 and 47.3 for 27, 37 and 42) showed markedly higher activity than EFL1 (RF = 32.92) and EFL3 (RF = 39.68). Structure-activity relationship study revealed an essential role of C-6/17 and C-12/13 double bonds on lathyrane nucleus for exerting MDR reversal activity. Western blotting analysis showed that 42 could reduce the expression level of P-glycoprotein (P-gp) in MCF-7/ADM cells; however, the most active compound 27 with an unnatural 5/7/7/4 fused-ring diterpenoid skeleton, had no inhibitory effect on P-gp expression.

Mechanism of Quercetin as a Multidrug-resistant Reversing Compound in Oxaliplatin-resistant Gastric-cancer Cell Lines.

Context: Treatment failure due to multidrug resistance (MDR) is a crucial hurdle during chemotherapy. MDR is generally correlated with an upregulation of adenosine triphosphate (ATP)-binding cassette (ABC) transport proteins. Also, aberrant activation of the phosphoinositide 3-kinase (PI3K)/ protein kinase B (Akt) pathway can counteract chemotherapeutic induction. Identification of safe and functioning MDR-reversing compounds is necessary in gastric-cancer therapy. Objective: The study intended to examine the role of Quercetin (Qur) in the mediation of osmotic glycoprotein (P-gp) expression and activity as an ABC transporter in the PI3K/Akt/ P-gp cascade in the oxaliplatin (OxR)-resistant, gastric-cancer cell line KATOIII/OxR. Design: The research team performed a laboratory study. Setting: The study took place at Nantong Haimen People's Hospital. Outcome Measures: The research team: (1) determined the impact of OxR on cell viability after treatment with Qur using trypan blue and "3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide" (MTT) assays; (2) employed a rhodamine 123 (Rh123) assay to detect the activity of P-gp; (3) used quantitative reverse transcription polymerase chain reaction (RT-qPCR) to measure mRNA expression of P-gp; and (4) detected apoptosis using an enzyme-linked immunoassay (ELISA) cell-death assay. Results: Qur: (1) increased the cytotoxicity of OxR; (2) downregulated the expression level and activity of P-gp and reversed MDR through the enhancement of the cytotoxicity of intracellular OxR in KATOIII/OxR cells; and (3) enhanced the apoptosis rate in KATOIII/OxR cells. Conclusions: Qur induced a dramatic reduction in the survival rate of KATOIII/OxR cells and may reverse OxR resistance through a decrease in P-gp expression and activity. These data imply that exposure of KATOIII/OxR cells in the dose-dependent manner to Qur can circumvent MDR by improving the intracellular accumulation of OxR. Qur might provide a new treatment strategy and improve patients' survival after chemotherapy for gastric cancer.

Terpenoids from Euphorbia soongarica and Their Multidrug Resistance Reversal Activity.

Ten new terpenoids, including five diterpenoids (1-5), three nortriterpenoids (6-8), and two triterpenoids (9, 10), and 15 known terpenoids (11-25) were isolated from an acetone extract of Euphorbia soongarica. Sooneuphoramine (1) is the first example of a euphoractine B-type diterpenoid alkaloid, while sooneuphanones A-C (6-8) are rare nortriterpenoids from the Euphorbia genus. The isolated terpenoids were tested for their cytotoxicity and multidrug resistance (MDR) reversal activity, 10 of which showed moderate cytotoxicity against the KB and KBv200 cell lines, while 11 compounds exhibited P-gp modulating potential. The triterpenoid sooneuphanone D (9) possessed a remarkable MDR reversal activity much higher than the positive control, verapamil.

Oroxylin A reverses P-glycoprotein-mediated multidrug resistance of MCF7/ADR cells by G2/M arrest.

Oroxylin A is a naturally occurring monoflavonoid isolated from the root of Scutellaria baicalensis Georgi, which has been used in traditional Chinese medicine for its anti-tumor, anti-inflammatory and anti-bacterial properties. The purpose of this study is to investigate the reversal effect and the fundamental mechanisms of oroxylin A in MCF7/ADR cells. Data indicated that oroxylin A showed strong reversal potency in MCF7/ADR cells and the reversal fold (RF) reached 4.68. After treatment with oroxylin A, MCF7/ADR cells displayed reduced functional activity and expression of MDR1 at both the protein and mRNA levels. Meanwhile, oroxylin A induced cells G2/M arrest in a concentration-dependent manner by increasing the expression of p-Chk2 (Thr68). Moreover, western blot and EMSA assays were used to reveal the inhibition of NF-κB in nucleus and the suppression of NF-κB binding activity by oroxylin A. NSC 109555 ditosylate-Chk2 inhibitor partly dismissed G2/M arrest induced by oroxylin A, reversed the increased trend of p-Chk2 and p-P53 (Ser20), inhibited the decreasing effect of oroxylin A on the expression of P-gp and decreased the reversal fold of 90 µM oroxylin A from 4.68 fold to 1.73 fold. In conclusion, we suggested that oroxylin A reversed MDR by G2/M arrest and the underlying mechanism attributed to the suppression of P-gp expression via Chk2/P53/NF-κB signaling pathway.

Synthesis and biological activities of a 3'-azido analogue of Doxorubicin against drug-resistant cancer cells.

Doxorubicin (DOX), an anthracycline antibiotic, is one of the most active anticancer chemotherapeutic agents. The clinical use of DOX, however, is limited by the dose-dependant P-glycoprotein (P-gp)-mediated resistance. Herein, a 3'-azido analogue of DOX (ADOX) was prepared from daunorubicin (DNR). ADOX exhibited potent antitumor activities in drug-sensitive (MCF-7 and K562) and drug-resistant cell lines (MCF-7/DNR, K562/DOX), respectively. The drug resistance index (DRI) values of ADOX were much lower than that of DOX. The cytotoxicity experiments of ADOX or DOX against K562/DOX, with or without P-gp inhibitor, indicated that ADOX circumvents resistance by abolishing the P-gp recognition. This conclusion was further supported by drug influx/efflux flow cytometry experiments, as well as by molecular docking of ADOX to P-gp. In vivo animal tests, ADOX exhibited higher activity and less toxicity than DOX. The current data warranted ADOX for additional pre-clinical evaluations for new drug development.

Triterpenoid saponins affect the function of P-glycoprotein and reduce the survival of the free-living stages of Heligmosomoides bakeri.

We studied the effect of triterpenoid saponins on the development of free-living stages of Heligmosomoides bakeri, a parasitic nematode of the mouse intestine. We evaluated the effectiveness of oleane-type glucuronides (GlcUAOA) isolated from Calendula officinalis and Beta vulgaris. The rhodamine 123 retention assay was used to detect dysfunctions of the major membrane transporter for xenobiotics, P-glycoprotein (Pgp). Both C. officinals and B. vulgaris GlcUAOA affect the development of the free living stages and function of Pgp in H. bakeri. The GlcUAOA inhibits egg hatching and moulting of larvae and also changes their morphology. These saponin fractions reversed the toxic effect of thiabendazole on the nematode; the function of Pgp was also inhibited.

23-Hydroxybetulinic acid from Pulsatilla chinensis (Bunge) Regel synergizes the antitumor activities of doxorubicin in vitro and in vivo.

UNLABELLED: ETHNOPHARMACOLOGICAL REVELANCE: Pulsatilla chinensis (Bunge)Regel has been used as adjuvant in chemotherapy in traditional Chinese medicine. 23-Hydroxybetulinic acid, an isolated pentacyclic triterpene, is the major active constituent of Pulsatilla chinensis (Bunge) Regel. AIM OF THIS STUDY: To evaluate the combinational anticancer effect of 23-hydroxybetulinic acid and doxorubicin in vitro and in vivo. MATERIALS AND METHODS: The effect of combination treatment with 23-hydroxybetulinic acid and doxorubicin was evaluated with a quantitative combination index method based on the median-effect analysis in various cancer cell lines. And in vivo efficacy of combination chemotherapy was also evaluated using ICR mice bearing sarcoma 180 carcinoma tumors. RESULTS: 23-Hydroxybetulinic acid showed a synergistic cytotoxic effect on multiple cancer cell lines by combined use with doxorubicin. In vivo studies further demonstrated that co-administration of 23-HBA significantly improved the sensitivity of the tumor to doxorubicin through increasing intra-tumor doxorubicin concentration and inhibiting doxorubicin-induced up-regulation of P-gp in tumor. CONCLUSION: These results suggest that the combined therapy with 23-hydroxybetulinic acid and doxorubicin may be a new promising strategy to promote the clinical chemotherapy, which needs further verification.

Effects of quercetin on the pharmacokinetics of Etoposide after oral or intravenous administration of etoposide in rats.

Etoposide [4'-demethylepipodophyllotoxin-9-(4,6-O-ethylidene)-beta-D-glucopyranoside] is a substrate for P-glycoprotein (P-gp) and cytochrome P450 (CYP) 3A. This study was designed to investigate the effects of quercetin (3,5,7,3',4'-pentahydroxyflavanone), a P-gp and CYP3A inhibitor, on the pharmacokinetics of etoposide in rats. Etoposide was administered to rats orally (9 mg/kg) or i.v. (3 mg/kg) without or with quercetin (1, 5 or 15 mg/kg). The plasma concentration of etoposide was determined by high performance liquid chromatography (HPLC) equipped with a fluorescence detector. In the presence of quercetin, the pharmacokinetic parameters of etoposide were significantly altered in the oral group, but not in the i.v. group. The presence of quercetin significantly (5 mg/kg, p<0.05; 15 mg/kg, p<0.01) increased the area under the plasma concentration-time curve (AUC) of orally administered etoposide from 43.0 or 53.2% . The presence of 5 or 15 mg/kg of quercetin significantly (p<0.05) decreased the total body clearance (CL/F) of oral etoposide. Consequently, compared to the control group (8.87%), the presence of quercetin significantly (5 mg/kg, p<0.05; 15 mg/kg, p<0.01) increased the absolute bioavailability (AB) of etoposide to 12.7 or 13.6% . The enhanced oral bioavailability of etoposide by quercetin could mainly be due to inhibition of P-gp-mediated efflux and CYP3A-catalyzed metabolism in the intestine by quercetin. The dosage regimen of etoposide in cancer therapy should take drug interaction into consideration when etoposide is administered with quercetin or dietary supplements containing quercetin.

In vivo evidence for the efflux transport of pentazocine from the brain across the blood-brain barrier using the brain efflux index method.

The efflux transport of pentazocine (PTZ) from the brain across the blood-brain barrier (BBB) was investigated using the Brain Efflux Index method. PTZ was eliminated with the apparent elimination half-life of 13.0 min after microinjection into the parietal cortex area 2 region of the rat brain. The apparent efflux clearance of PTZ across the BBB was 137 microl/min/g brain, which was calculated from the elimination rate constant (5.35 x 10(-2) min(-1) and the distribution volume in the brain (2.56 ml/g brain). The efflux transport of PTZ was decreased in the presence of unlabeled PTZ, suggesting that PTZ is eliminated by a carrier-mediated transport system across the BBB. To characterize the efflux transport of PTZ from the brain in vivo, the effects of several compounds on the efflux transport of PTZ were investigated. P-glycoprotein (P-gp) inhibitors (verapamil and quinidine) reduced the PTZ efflux transport. In addition, the efflux transport of PTZ was inhibited by organic cations such as l-carnitine and tetraethylammonium (TEA), whereas organic anions such as p-aminohippuric acid, probenecid and taurocholate did not affect the PTZ efflux transport. The present results suggest that PTZ is transported from the brain across the BBB via l-carnitine/TEA-sensitive carrier-mediated efflux transport system(s) in addition to P-gp.

Expression of P-glycoprotein and cytochrome p450 1A in intertidal fish (Anoplarchus purpurescens) exposed to environmental contaminants.

Whether P-glycoproteins (P-gps) like those which confer multidrug resistance in tumor cell lines are important in adaptation to chemicals in natural populations of vertebrates exposed to contaminant mixtures is the focus of this study. P-gp expression was examined in the intertidal fish high cockscomb blenny (Anoplarchus purpurescens) exposed to crude oil or pulp mill effluent. The relationship between P-gp expression and cytochrome p450 1A (CYP1A) induction also was investigated. Immunohistochemical (IHC) analysis revealed that levels of P-gp expression in the bile canaliculi were three- to five-fold greater in oil exposed fish than in control fish. Levels of P-gp expression were highly correlated with hepatic CYP1A levels previously measured in these fish. In fish from sites near pulp mills, P-gp expression in freshly caught fish did not correlate with proximity to pulp mills. However, hepatic P-gp expression levels in freshly caught fish were 14-fold higher than in fish from those sites that were depurated in clean water for 6 weeks. CYP1A levels were also elevated in liver of freshly caught as compared with depurated fish. Expression of neither CYP1A nor P-gp was elevated in depurated fish exposed to sediment and food from within the original pulp mill effluent stream. Depurated fish, which were injected with the aryl hydrocarbon receptor (AHR) agonist ss-naphthoflavone (BNF) showed an expected induction of CYP1A but no induction of P-gp. These results suggest that in blennies, unlike CYP1A, P-gp expression is not regulated by the AHR pathway; although P-gp and CYP1A both may be induced by some compounds in petroleum and unidentified xenobiotics at field sites. While our data indicate that CYP1A and P-gp are not coordinately regulated, these proteins may play complementary roles in cellular detoxification. Thus the elevation of P-gp activity may be an important mechanism of multixenobiotic resistance for organisms, such as intertidal fish, which are commonly exposed to anthropogenic contaminants and naturally occurring toxins.