Characterization of the inclusion complex of zerumbone with hydroxypropyl--cyclodextrin

In this paper we investigated the inclusion complexation between zerumbone (ZER) and hydroxylpropyl- -cyclodextrin (HPCD) at four different temperatures: 293–318 ◦K. The thermodynamic parameters (H, S and G) for the formation of the complex were obtained from the van’t Hoff equation. The complex with HPCD was characterized by differential scanning calorimetry (DSC), X-ray diffractometry (XRD), Fourier transform infrared spectroscopy (FT-IR), and molecular modeling using PM6. The solubility of ZER was enhanced >30 fold after complexation. Calculations show that ZER penetrates completely into the cavity of HPCD. The complex retained its cytotoxic activity as shown by in vitro cell survival assay on human cervical cancer (Hela), breast cancer (MCF7 and MDA-MB 231) and human leukemic (CEMss) cell lines. HPCD is, therefore, a suitable encapsular capable of forming thermodynamically stable complex with ZER for save delivery of the compound as an anticancer drug in the future

Liquid chromatography - tandem mass spectroscopic method for the determination Of zerumbone in human plasma and its application to Pharmacokinetics

A rapid, sensitive, specific and selective LC-MS/MS method for the determination of zerumbone (ZER) in human plasma using 2,4-diamino-6-(4-methoxyphenyl)-1,3,5-triazine (DMTZ) as an internal standard (IS) has been developed and validated. ZER was chromatographed on C8 column using a mobile phase of acetonitrile/water (80:20, v/v) at a flow rate of 0.25 ml min(-1) . Quantitation was achieved using ESI+ interface, employing multiple reaction monitoring (MRM) mode at m/z 219 > 81 and 218 > 134 for ZER and IS, respectively. The calibration standards were linear over a range of 5-3000 ng ml(-1) (r(2)=0.9994) with an LLOQ of 5 ng ml(-1) (RSD %; 11.4% and bias%; 9.5%). Intra- and inter-day precision of ZER assay ranged from 0.18 to 3.56% with accuracy (bias) that varied between -5.09 and 4.3%, demonstrating good precision and accuracy. Recoveries of ZER and the IS from human plasma were above 85%. The developed method was validated for the determination of ZER in rat plasma. Linearity, stability of ZER and the ME on rat plasma were discussed. The applicability of the developed method was demonstrated by measuring ZER in rat plasma samples following intravenous and intraperitoneal administration of ZER prepared in hydroxypropyl-β-cyclodextrin (HPβCD) and sodium carboxymethyl cellulose (CMC), respectively, in 20 mg kg(-1) and this study indicated a clear significant difference (p<0.05) in pharmacokinetic parameters of ZER in ZER/HPβCD complex compared with ZER in CMC preparation

In vivo antitrypanosomal activity of Garcinia hombroniana aqueous extract

The anti-Trypanosoma evansi activity of Garcinia hombroniana (seashore mangosteen) leaves aqueous extract was tested on experimentally infected Sprague–Dawley rats. Treatment of infected rats with G. hombroniana extract resulted in a significantly extended post-infection longevity (p < 0.05), compared to the untreated control group. The possible mode of antitrypanosomal effect of the plant extract was also investigated on cultured T. evansi in HMI-9 medium with the addition of 25 µg/ml G. hombroniana aqueous extract. It was observed that the addition of G. hombroniana extract resulted in the inhibition of trypanosomal kinetoplast division, with no significant inhibitory effect on nuclear division. It is concluded from the current study that the aqueous extract of G. hombroniana has a potential antitrypanosomal activity through the inhibition of kinetoplast division, as one of the possible mechanisms of its antitrypanosomal effect. This plant could serve as a possible source of new antitrypanosomal compounds

Antioxidant, Anti-inflammatory and Cytotoxicity of Phaleria macrocarpa (Boerl.) Scheff Fruit

<p>Abstract</p> <p>Background</p> <p><it>Phaleria macrocarpa </it>(Scheff.) Boerl (Thymelaceae) originates from Papua Island, Indonesia and grows in tropical areas. The different parts of the fruit of <it>P. macrocarpa </it>were evaluated for antioxidant, anti-inflammatory, and cytotoxic activities.</p> <p>Methods</p> <p><it>Phaleria macrocarpa </it>fruit were divided into pericarp, mesocarp and seed. All parts of the fruit were reflux extracted with methanol. The antioxidant activity of the extracts were characterized in various <it>in vitro </it>model systems such as FTC, TBA, DPPH radical, reducing power and NO radical. Anti-inflammatory assays were done by using NO production by macrophage RAW 264.7 cell lines induced by LPS/IFN-γ and cytotoxic activities were determined by using several cancer cell lines and one normal cell line</p> <p>Results</p> <p>The results showed that different parts (pericarp, mesocarp, and seed) of <it>Phaleria macrocarpa </it>fruit contain various amount of total phenolic (59.2 ± 0.04, 60.5 ± 0.17, 47.7 ± 1.04 mg gallic acid equivalent/g DW) and flavonoid compounds (161.3 ± 1.58, 131.7 ± 1.66, 35.9 ± 2.47 mg rutin equivalent/g DW). Pericarp and mesocarp showed high antioxidant activities by using DPPH (71.97%, 62.41%), ferric reducing antioxidant power (92.35%, 78.78%) and NO scavenging activity (65.68%, 53.45%). Ferric thiocyanate and thiobarbituric acid tests showed appreciable antioxidant activity in the percentage hydroperoxides inhibitory activity from pericarp and mesocarp in the last day of the assay. Similarly, the pericarp and mesocarp inhibited inducible nitric oxide synthesis with values of 63.4 ± 1.4% and 69.5 ± 1.4% in macrophage RAW 264.7 cell lines induced by LPS/IFN-γ indicating their notable anti-inflammatory potential. Cytotoxic activities against HT-29, MCF-7, HeLa and Chang cell lines were observed in all parts.</p> <p>Conclusions</p> <p>These results indicated the possible application of <it>P. macrocarpa </it>fruit as a source of bioactive compounds, potent as an antioxidant, anti inflammatory and cytotoxic agents.</p

Induction of selective cytotoxicity and apoptosis in human T4-lymphoblastoid cell line (CEMss) by boesenbergin a isolated from boesenbergia rotunda rhizomes involves mitochondrial pathway, activation of caspase 3 and G2/M phase cell cycle arrest

Background Boesenbergia rotunda (Roxb.) Schlecht (family zingiberaceae) is a rhizomatous herb that is distributed from north-eastern India to south-east Asia, especially in Indonesia, Thailand and Malaysia. Previous research has shown that the crude extract of this plant has cytotoxic properties. The current study examines the cytotoxic properties of boesenbergin A isolated from Boesenbergia rotunda. Methods MTT assay was used to check the cytotoxicity of boesenbergin A. The morphological assessment of apoptosis was monitored using normal and fluorescence microscopy. The early and late phase of apoptosis was investigated using annexin V and DNA laddering assays, respectively. The mitochondrial membrane potential (MMP) was assessed by fluorescence microscopy. Human apoptosis proteome profiler assays were performed to investigate the mechanism of cell death. In addition, the protein levels of Bax, Bcl2 and HSP 70 were also analyzed using western blot. Assays of caspase =-3/7, -8 and =-9 were carried out in order to test for induction during treatment. Lastly, cell cycle progression was analyzed using flow cytometry. Results Boesenbergin A was found to have the highest toxicity towards CEMss cancer cells (IC50 = 8 μg/ml). The morphology of CEMss cells after treatment showed evidence of apoptosis that included blebbing and chromatin condensation. The annexin V assay revealed that early apoptosis is induced after treatment. The DNA laddering assay confirmed that DNA fragmentation had occurred during late apoptosis. The cell cycle analysis indicated that boesenbergin A was able to induce G2/M phase arrest in CEMss cells. The activity of caspases -3/7, -8 and -9 was increased after treatment which indicates both intrinsic and extrinsic pathways are induced during apoptosis. The involvement of mitochondria was established by increased mitochondrial membrane potential and up and down regulation of Bcl2 and Bax proteins as well as HSP70. Conclusion In conclusion, the results demonstrated that boesenbergin A induced apoptosis of CEMss cells through Bcl2/Bax signaling pathways with the involvement of caspases and G2/M phase cell cycle arrest. The current findings warrant further research on boesenbergin A as a novel chemotherapeutic agent for leukemia intervention including studies in animal models

31st Annual Meeting and Associated Programs of the Society for Immunotherapy of Cancer (SITC 2016) : part two

Background The immunological escape of tumors represents one of the main ob- stacles to the treatment of malignancies. The blockade of PD-1 or CTLA-4 receptors represented a milestone in the history of immunotherapy. However, immune checkpoint inhibitors seem to be effective in specific cohorts of patients. It has been proposed that their efficacy relies on the presence of an immunological response. Thus, we hypothesized that disruption of the PD-L1/PD-1 axis would synergize with our oncolytic vaccine platform PeptiCRAd. Methods We used murine B16OVA in vivo tumor models and flow cytometry analysis to investigate the immunological background. Results First, we found that high-burden B16OVA tumors were refractory to combination immunotherapy. However, with a more aggressive schedule, tumors with a lower burden were more susceptible to the combination of PeptiCRAd and PD-L1 blockade. The therapy signifi- cantly increased the median survival of mice (Fig. 7). Interestingly, the reduced growth of contralaterally injected B16F10 cells sug- gested the presence of a long lasting immunological memory also against non-targeted antigens. Concerning the functional state of tumor infiltrating lymphocytes (TILs), we found that all the immune therapies would enhance the percentage of activated (PD-1pos TIM- 3neg) T lymphocytes and reduce the amount of exhausted (PD-1pos TIM-3pos) cells compared to placebo. As expected, we found that PeptiCRAd monotherapy could increase the number of antigen spe- cific CD8+ T cells compared to other treatments. However, only the combination with PD-L1 blockade could significantly increase the ra- tio between activated and exhausted pentamer positive cells (p= 0.0058), suggesting that by disrupting the PD-1/PD-L1 axis we could decrease the amount of dysfunctional antigen specific T cells. We ob- served that the anatomical location deeply influenced the state of CD4+ and CD8+ T lymphocytes. In fact, TIM-3 expression was in- creased by 2 fold on TILs compared to splenic and lymphoid T cells. In the CD8+ compartment, the expression of PD-1 on the surface seemed to be restricted to the tumor micro-environment, while CD4 + T cells had a high expression of PD-1 also in lymphoid organs. Interestingly, we found that the levels of PD-1 were significantly higher on CD8+ T cells than on CD4+ T cells into the tumor micro- environment (p < 0.0001). Conclusions In conclusion, we demonstrated that the efficacy of immune check- point inhibitors might be strongly enhanced by their combination with cancer vaccines. PeptiCRAd was able to increase the number of antigen-specific T cells and PD-L1 blockade prevented their exhaus- tion, resulting in long-lasting immunological memory and increased median survival

Antitrypanosomal activities of selected medicinal plants

The aqueous and ethanolic extracts from four medicinal plants namely Acanthus ilicifolius, Aquilaria malaccensis, Goniothalamus tapis, and Maesa ramentacea, were tested in vitro against Trypanosoma evansi, a hemoprotozoan parasite causing surra in a vast variety of domesticated and wild animals. The antitrypanosomal activity was evaluated by calculating median inhibitory concentration (IC50), the concentration that gives half maximal growth of the trypanosomes, of the plant extracts. Goniothalamus tapis ethanolic extract showed the most potent antitrypanosomal activity with an IC50 value of 7.6 ± 1.6 μg ml-1. The other extracts scored IC50 values ranging from 36.2 ± 1.3 μg ml-1 to 314.1 ± 37.1 μg ml-1