Objective: This study aimed to explore Hesperetin (Hst) potency as a co-chemotherapeutics agent combined with Doxorubicin (Dox), particularly cytotoxic and antimetastasis effects toward MCF-7/HER2 cells. Methods: The cytotoxic effects were measured under MTT assay. The flowcytometry analysis was used to examine the cell cycle modulation and apoptosis evidence, while the effect of migration was assayed by scratch wound healing assay. Western blotting and gelatin zymography were carried out to examine the expression level of proteins, HER2, and Rac1. Results: Under MTT assay, Hst and Dox exhibited to decrease cell viability in a dose-dependent manner with the IC 50 value of 377 and 0,8 µM, respectively. The combination of Hst and Dox at the respective doses of 95 and 0,2 µM showed a synergistic effect with the combination index of 0,63. Flow cytometry analysis of Hst-Dox revealed that those compounds caused cell cycle arrest at the G2/M phase and induced apoptosis. Hst also decreased HER2 and Rac1 expression, as shown by western blot. Hst inhibited lamellipodia formation and cell migration, as indicated by microscopic observation and wound healing scratch assay. The antimetastatic activity of Hst was associated with the reduction of Rac1 and MMP9 expression as measured by gelatine zymography assay. Conclusion: These results indicated that the combination of Hst and Dox-induced cell cycle arrest, apoptosis, decreased HER2, Rac1, MMP9 expression, and cell migration. Thus, Hst may have the potential to be developed as a co-chemotherapeutic agent combined with doxorubicin toward HER2 overexpressing breast cancer cells.

Purpose: Genistein, a soy isoflavone, exhibits a biphasic effect on cells proliferation with some different effects between ER-alpha and ER-beta. The objective of this present study is to determine the modulatory effect based on cell cycle progression under genistein treatment in combination with 17-β estradiol (E2) on CHO-K1 cells. Methods: The effect of genistein 0.1-100 µM on cells proliferation was examined by MTT assay. The modulation of genistein and estradiol (E2) on cell cycle and apoptosis were observed by using flowcytometry with PI and PI/AnnexinV staining, respectively. Moreover, the effect of genistein and E2 on senescence cells, and ROS level were determined by senescence-associated β-galactosidase (SA β-gal) staining and by using flowcytometry with 2’, 7’–dichlorofluorescin diacetate (DCFDA) staining, respectively. The expression level of the cell cycle and senescence protein markers were observed by immunoblotting. Results: Single treatment of genistein at physiologically achievable (low) concentration (<2 µM) induced proliferation of CHO-K1 cells while at a pharmacological (high) concentration (50 and 100 µM) suppressed cells proliferation. Interestingly, treatment of genistein at the physiological concentration in combination with E2 for 24, 48 and 72 h decreased cells viability on CHO-K1 cells compared to untreated cells. Further analysis of the cells showed that 50 µM genistein induced G2/M phase accumulation and induced apoptosis. Moreover, genistein induced cell senescence and increased ROS level. Immunoblotting analysis showed the decreasing of ERalpha, Bcl2, and ppRb protein level upon treatment of 1 µM Gen and 1 nM E2. Conclusion: Our results suggest that the cell proliferation inhibitory mechanism of genistein at pharmacological concentration involved the induction of cell senescence, and the elevation of ROS level. Moreover, the decreased of cells proliferation upon treatment of physiological concentration of genistein in combination with E2 may be correlated with the alteration of ER expression.

Secang (C. sappan L.) and awar (F. septica Burm.f) are known of Indonesian traditional medicine that highly consumed throughout centuries in order to cure various diseases. Recently, researchers also concern about its effects as anti-cancer on various cell types. This study was conducted to understand the ethanolic extract of C. sappan L. (ECS) and F. septica Burm.f (EFS) effects on 4T1 cells migration at various concentrations. Firstly, we examine cell proliferation profile with MTT assay under treatment with the extracts and obtained the IC50 value of ECS (20 μg/mL) and EFS (15 μg/mL). Subsequent assay conducted was to examine the cells migration under low concentration resulting in the migration inhibitory effect of both EFS and ECS with different intensity. EFS performed better migration inhibitory effect than ECS. Finally, we conducted gelatin zymography and western blot and revealed that the migration inhibitory effect of EFS may correlate to the lowering of protein expression of MMP9 and Rac-1 after 24 hours of treatment. We conclude that both extracts are potential to be developed as anticancer agent and EFS is more potent for anti-metastasis.

Background: Hesperetin (Hst), a citrus flavanone, is widely distributed among citrus fruits, including lemons. Hst has been shown to possess bioactivity as an antioxidant, anti-inflammatory, anti-allergic, hypolipidemic, vasoprotector, and anticancer agent. This study aimed to identify potential combinations of Hst and the chemotherapeutic agent doxorubicin (Dox) as co-chemotherapy agents against 4T1 murine metastatic breast cancer cells. Results: MTT assay results showed that Hst exhibited cytotoxic effect in 4T1 cells, and its combination with Dox showed a synergistic effect based on the CI value. The combination of Hst and Dox increased G2/M phase cell cycle arrest and apoptosis induction. The combination of Hst and Dox inhibited migration and decreased MMP-9 expression in 4T1 cells. Conclusion: In conclusion, the results of this study show that Hst has potential as a Dox co-chemotherapy against 4T1 cells by inducing G2/M phase cell cycle arrest and apoptosis. More importantly, Hst reduces Dox-induced migration and decreases MMP-9 expression.