Ratna Asmah Susidarti

Curcumin and its analogues (PGV-0 and PGV-1) enhance sensitivity of resistant MCF-7 cells to doxorubicin through inhibition of HER2 and NF-kB activation.

Chemoresistance of breast cancer to doxorubicin is mediated mainly through activation of NF-kB and over expression of HER2. Curcumin and its analogues (PGV-0 and PGV-1) exert cytotoxic effects on T47D breast cancer cells. Suppression of NF-kB activation is suggested to contribute to this activity. The present study aimed to explore the effects of curcumin, PGV-0, and PGV-1 singly and in combination with doxorubicin on MCF-7/Dox cells featuring over-expression of HER2. In MTT assays, curcumin, PGV-0, and PGV-1 showed cytotoxicity effects against MCF-7/Dox with IC50 values of 80 μM, 21 μM, and 82 μM respectively. These compounds increased MCF-7/Dox sensitivity to doxorubicin. Cell cycle distribution analysis exhibited that the combination of curcumin and its analogues with Dox increased sub G-1 cell populations. Curcumin and PGV-1 but not PGV-0 decreased localization of p65 into the nucleus induced by Dox, indicating that activation of NF- kB was inhibited. Molecular docking of curcumin, PGV-0, and PGV-1 demonstrated high affinity to HER2 at ATP binding site. This interaction were directly comparable with those of ATP and lapatinib. These findings suggested that curcumin, PGV-0 and PGV-1 enhance the Dox cytotoxicity to MCF-7 cells through inhibition of HER2 activity and NF-kB activation.

Cytotoxic activity of coumarins from Micromelum minutum

The crude petroleum ether and chloroform extracts of Micromelum minutum (G. Frost.) Wright & Arn (Rutaceae) showed strong cytotoxic activity when tested against a T-lymphoblastic leukemia cell line. Further fractionation of the extracts resulted in the isolation of five new coumarins 3″,4″-dihydrocapnolactone, 2′,3′-epoxyisocapnolactone, 8-hydroxyisocapnolactone-29,39-diol, 8-hydroxy-3″,4″-dihydrocapnolactone-29,39-diol and 8,4″-dihydroxy-3″,4″-dihydrocapnolactone-29,39-diol, and two triterpenes. Some of these compounds were strongly active against T-lymphoblastic leukemia (CEM-SS), promyeolocytic leukemia (HL60), cervical cancer (HeLa) and liver cancer (HepG2) cell lines. 8-Hydroxyisocapnolactone-29,39-diol was found to be the most active with IC50 values of 2.9, 2.5, 6.9, and 5.9 μg/ml, respectively. This was followed by 2′,3′-epoxyisocapnolactone. When evaluated against the normal mouse fibroblast (3T3) cell line, 8-hydroxyisocapnolactone-29,39-diol was found to be inactive, hence it could serve as a valuable lead for further design and synthesis of more active analogues.

A new coumarin and triterpenes from Malaysian Micromelum minutum

A new coumarin, 8,4″-dihydroxy-3″,4″-dihydrocapnolactone-2′,3′-diol (1) and two known triterpenes, 5(6)-gluten-3-one (2) and 5(6)-gluten-3α-ol (3) were isolated from the leaves of Micromelum minutum (Rutaceae) collected from Sepilok, Sabah, Malaysia and their structures were characterized by spectroscopic methods.

Hesperidin increase cytotoxic effect of doxorubicin in MCF-7 cells

Hesperidin, a flavonoid, shows strong cytotoxic effect in several cancer cell lines. The aim of this research was to investigate cytotoxic activities of hesperidin alone and in combination with doxorubicin. Cell viability assay of hesperidin, doxorubicin, and combination treatments were carried out by using MTT assay. Apoptosis assay was done using double staining method using Ethidium Bromide-Acridine Orange. Hesperidin did not show cytotoxic effect but doxorubicin showed cytotoxic effect with IC50 467 nM. Hesperidin (5, 50 and 100 μM) increased cytotoxic effect of doxorubicin compared with doxorubicin alone. The strongest cytotoxic activity was showed by the combination of 200 nM doxorubicin and 100 μM hesperidin. Combination treatment of doxorubicin 200 nM and hesperidin 100 μM induced apoptosis in MCF-7 cells. Hesperidin is potentially to be developed as co-chemotherapeutic agent for breast cancer, while molecular mechanism need to be explored.

Two Active Compounds from Caesalpinia sappan L. in Combination withCisplatin Synergistically Induce Apoptosis and Cell Cycle Arrest on WiDr Cells

Purpose: The aim of this study is to observe the synergistic effect of two active compounds of secang, brazilin and brazilein, combined with cisplatin on WiDr colon cancer cells. Methods: Cytotoxic activities of brazilin (Bi) and brazilein (Be) in single and in combination with cisplatin (Cisp) were examined by MTT assay. Synergistic effect was analyzed by combination index (CI) parameter. Apoptosis and cell cycle profiles were observed by using flow cytometry. Results: The result of MTT assay showed that IC50 value of brazilin and brazilein on WiDr cancer cells were 41 µM and 52 µM respectively. The combination of ½ IC50 of Bi-Cisp reduced cells viability up to 64% and showed synergistic effect with CI value less than 1 (CI = 0.8). The combinations of ½ IC50 of Be-Cisp also reduced cells viability up to 78% and showed synergistic effect (CI=0.65). Combination of Bi-Cisp and Be-Cisp induced apoptosis higher than the single treatments. Further analysis on the cell cycle progression showed that single treatment of ½ IC50 of Be and Bi induced S-phase and G2/M-phase accumulation, while combination of Be-Cisp and Bi-Cisp enhanced S-phase accumulation. Conclusion: Both combination of Bi-Cisp and Be-Cisp showed synergistic effect on WiDr cells through induction of apoptosis and halted the cell cycle progression, thus, WiDr cells growth were significantly reduced.

The Cytotoxic and Antimigratory Activity of Brazilin-Doxorubicin on MCF-7/HER2 Cells

Purpose: Breast cancer cells with overexpression of HER2 are known to be more aggressive, invasive, and resistant to chemotherapeutic agent. Brazilin, the major compound in the Caesalpinia sappan L. (CS) heartwood, has been studied for its anticancer activity. The purpose of this study was to investigate the cytotoxic and antimigratory activity of brazilin (Bi) in combination with doxorubicin (Dox) on MCF-7/HER2 cells. Methods: Cytotoxic activities of Bi individually and in combination with Dox were examined by MTT assay. Synergistic effects were analyzed by combination index (CI). Apoptosis and cell cycle profiles were observed by using flow cytometry. Migrating and invading cells were observed by using a Boyden chamber assay. Levels of MMP2 and MMP9 activity were observed by using a gelatin zymography assay. Levels of HER2, Bcl-2, Rac1, and p120 protein expression were observed by using an immunoblotting assay. Results: The results of the MTT assay showed that Bi inhibited MCF-7/HER2 cell growth in a dose-dependent manner with an IC50 of 54 ± 3.7 µM. Furthermore, the combination of Bi and Dox showed a synergistic effect (CI <1). Flow cytometric analysis of Bi and its combination with Dox showed cellular accumulation in the G2/M phase and induction of apoptosis through suppression of Bcl-2 protein expression. In the Boyden chamber assay, gelatin zymography, and subsequent immunoblotting assay, the combination Bi and Dox inhibited migration, possibly through downregulation of MMP9, MMP2, HER2, Rac1, and p120 protein expression. Conclusion: We conclude that Bi enhanced cytotoxic activity of Dox and inhibited migration of MCF-7/HER2 cells. Therefore, we believe that it has strong potential to be developed for the treatment of metastatic breast cancer with HER2 overexpression.

Isolation and 2,2’-diphenyl-1-picrylhydrazyl radical scavenging activity of active compound from Jujube tree (Zizyphus mauritiana Auct. non Lamk.)

ABSTRACT Increased reactive oxygen species causes cells or tissue damages and is associated with some degenerative diseases, such as coronary heart diseases and cancer. This research is intended to isolate and to identify the antiradical compounds from the bark of Jujube tree (Zizyphus mauritiana Auct. non Lamk.). The methanol extract of the bark was successively fractionated with n-hexane and ethyl acetate to give n-hexane fraction, ethyl acetate fraction, and methanol extract residue, respectively. The extract and fractions were examined for its radical scavenging activity using 2,2’-diphenyl-1-picrylhydrazil radical. The ethyl acetate fraction, having the most active 2,2’-diphenyl-1-picrylhydrazil radical scavenging, was further repeatedly separated using vacuum liquid chromatography, and the purification was carried out using preparative thin layer chromatography (TLC) in order to obtain an active isolate. The purity of the isolate was evaluated using thin layer chromatography method and melting point measurement. The structure of the isolate was identified chemically using various spray reagent and ultraviolet-visible, infrared, infrared, Liquid Chromatography-Mass Spectrometry (LC-MS) and Proton Magnetic Resonance (1H-NMR) spectroscopic methods. The extract, fraction, and sub-fractions were determined for its total contents of phenolic and flavonoid. The results showed that the methanol extract residue contained the highest total phenolic (36.29% wt/wt gallic acid equivalent), while fraction E4 of the ethyl acetate extract contained the highest total flavonoids (21.31% wt/wt quercetin equivalent). The active isolate showed to have 2,2’-diphenyl-1-picrylhydrazil radical scavenging activity with IC50 of 7.58 μ/mL and is identified as trans-p-coumaroyl triterpene. Detail structure of the triterpene needs to be identified further.