Singkome Tima

Inhibitory effect of curcumin on MDR1 gene expression in patient leukemic cells.

When patients with cancers are treated with chemotherapeutic agents a long time, some of the cancer cells develop the multidrug resistance (MDR) phenotype. MDR cancer cells are characterized by the overexpression of multidrug resistance1 (MDR1) gene which encodes P-glycoprotein (Pgp), a surface protein of tumor cells that functions to produce an excessive efflux and thereby an insufficient intracellular concentration of chemotherapeutic agents. A variety of studies have sought potent MDR modulators to decreaseMDR1 gene expression in cancer cells. Our previous study has shown that curcumin exhibits characteristics of a MDR modulator in KB-V1 multidrug-resistant cells. The aim of this study was to further investigate the effect of curcumin onMDR1 gene expression in patient leukemic cells. The leukemic cells were collected from 78 childhood leukemia patients admitted at maharaj Nakorn Chiang Mai Hospital, Chiang Mai, Thailand, in the period from July 2003 to February 2005. There were 61 cases of acute lymphoblastic leukemia (ALL), 14 cases of acute myeloblastic leukemia (AML), and 3 cases of chronic myelocytic leukemia (CML). There were 47 males and 31 females ranging from 1 to 15 years old. Bone marrows were collected. The leukemic cells were separated and cultured in the presence or absence of 10 μM curcumin for 48 hours. MDR1 mRNA levels were determined by RT-PCR. It was found that curcumin reducedMDR1 gene expression in the cells from 33 patients (42%). Curcumin affected theMDR1 gene expression in 5 of 11 relapsed cases (45%), 10 of 26 cases of drug maintenance (38%), 7 of 18 cases of completed treatment (39%), and 11 of 23 cases of new patients (48%). The expression levels ofMDR1 gene in leukemic patient cells as compared to that of KB-V1 cells were classified as low level (1–20%) in 5 of 20 cases (25%), medium level (21–60%) in 14 of 32 cases (44%), and high level (61–100%) in 14 of 20 cases (70%). In summary, curcumin decreased MDR1 mRNA level in patient leukemic cells, especially in high level ofMDR1 gene groups. Thus, curcumin treatment may provide a lead for clinical treatment of leukemia patients in the future.

Stable curcumin-loaded polymeric micellar formulation for enhancing cellular uptake and cytotoxicity to FLT3 overexpressing EoL-1 leukemic cells.

&NA; The present study aims to develop a stable polymeric micellar formulation of curcumin (CM) with improved solubility and stability, and that is suitable for clinical applications in leukemia patients. CM‐loaded polymeric micelles (CM‐micelles) were prepared using poloxamers. The chemical structure of the polymers influenced micellar properties. The best formulation of CM‐micelles, namely CM‐P407, was obtained from poloxamer 407 at drug to polymer ratio of 1:30 and rehydrated with phosphate buffer solution pH 7.4. CM‐P407 exhibited the smallest size of 30.3 ± 1.3 nm and highest entrapment efficiency of 88.4 ± 4.1%. When stored at −80 °C for 60 days, CM‐P407 retained high protection of CM and had no significant size change. In comparison with CM solution in dimethyl sulfoxide (CM‐DMSO), CM kinetic degradation in both formulations followed a pseudo‐first‐order reaction, but the half‐life of CM in CM‐P407 was approx. 200 times longer than in CM‐DMSO. Regarding the activity against FLT3 overexpressing EoL‐1 leukemic cells, CM‐P407 showed higher cytotoxicity than CM‐DMSO. Moreover, intracellular uptake to leukemic cells of CM‐P407 was 2–3 times greater than that of CM‐DMSO. These promising results for CM‐P407 will be further investigated in rodents and in clinical studies for leukemia treatment. Graphical abstract Figure. No caption available.

Inhibitory effect of turmeric curcuminoids on FLT3 expression and cell cycle arrest in the FLT3-overexpressing EoL-1 leukemic cell line.

Leukemia is a hematologic malignancy with a frequent incidence and high mortality rate. Previous studies have shown that the FLT3 gene is overexpressed in leukemic blast cells, especially in acute myeloid leukemia. In this study, a commercially available curcuminoid mixture (1), pure curcumin (2), pure demethoxycurcumin (3), and pure bisdemethoxycurcumin (4) were investigated for their inhibitory effects on cell growth, FLT3 expression, and cell cycle progression in an FLT3-overexpressing EoL-1 leukemic cell line using an MTT assay, Western blotting, and flow cytometry, respectively. The mixture (1) and compounds 2-4 demonstrated cytotoxic effects with IC50 values ranging from 6.5 to 22.5 μM. A significant decrease in FLT3 protein levels was found after curcuminoid treatment with IC20 doses, especially with mixture 1 and compound 2. In addition, mixture 1 and curcumin (2) showed activity on cell cycle arrest at the G0/G1 phase and decreased the FLT3 and STAT5A protein levels in a dose-dependent manner. Compound 2 demonstrated the greatest potential for inhibiting cell growth, cell cycle progression, and FLT3 expression in EoL-1 cells. This investigation has provided new findings regarding the effect of turmeric curcuminoids on FLT3 expression in leukemic cells.

Development and Characterization of FLT3-Specific Curcumin-Loaded Polymeric Micelles as a Drug Delivery System for Treating FLT3-Overexpressing Leukemic Cells

This study aimed at developing a curcumin (CM) nanoparticle targeted to Feline McDonough Sarcoma (FMS)-like tyrosine kinase 3 (FLT3) protein on the surface of leukemic cells and at evaluating their properties, specificity, cytotoxicity, and inhibitory effect on FLT3 protein level in FLT3-overexpressing leukemic cells, EoL-1, and MV-4-11 cells. FLT3-specific peptides were conjugated onto modified poloxamer 407 using the copper-catalyzed azide-alkyne cycloaddition reaction. The thin film hydration method was performed for FLT3-specific CM-loaded polymeric micelles (FLT3-CM-micelles) preparation. Flow cytometry and fluorescence microscopy were used to determine rate of cellular uptake. 3-(4,5-dimethythiazol-2-yl)-2,5-diphenyl tetrazolium bromide assay was used to test the cytotoxicity of the micelles on leukemic cells. FLT3-CM-micelles demonstrated a mean particle size less than 50 nm, high entrapment efficiency, and high rate of CM uptake by leukemic cells. The intracellular CM fluorescence is related to FLT3 protein levels on the leukemic cell surfaces. Moreover, FLT3-CM-micelles demonstrated an excellent cytotoxic effect and decreased FLT3 protein expression in the leukemic cells. The FLT3-CM-micelles could enhance both solubility and cytotoxicity of CM on FLT3-overexpressing leukemic cells. These promising nanoparticles may be used for enhancing antileukemic activity of CM and developed as a targeted drug delivery system in the future.

EFFECT OF THAI SARAPHI FLOWER EXTRACTS ON WT1 AND BCR/ABL PROTEIN EXPRESSION IN LEUKEMIC CELL LINES

Background: Saraphi (Mammea siamensis) is a Thai traditional herb. In this study, the cytotoxic effects of crude ethanolic and fractional extracts including hexane, ethyl acetate, and methanol fractions from M. siamensis flowers were investigated in order to determine their effect on WT1 expression in Molt4 and K562 cells and Bcr/Abl expression in K562 cells. Materials and Methods: The flowers of M. siamensis were extracted using ethanol. The ethanol flower extract was further fractionated with hexane, ethyl acetate, and methanol. Cytotoxic effects were measured by the MTT assay. Bcr/Abl and WT1 protein levels after treatments were determined by Western blotting. The total cell number was determined via the typan blue exclusion method. Results: The hexane fraction showed the strongest cytotoxic activity on Molt4 and K562 cells, with IC50 values of 2.6 and 77.6 μg/ml, respectively. The hexane extract decreased Bcr/Abl protein expression in K562 cells by 74.6% and WT1 protein expressions in Molt4 and K562 cells by 68.4 and 72.1%, respectively. Total cell numbers were decreased by 66.2 and 48.7% in Molt4 and K562 cells, respectively. Mammea E/BB (main active compound) significantly decreased both Bcr/Abl and WTlprotein expressions by 75 and 49.5%, respectively when compared to vehicle control. Conclusion: The hexane fraction from M. siamensis flowers inhibited cell proliferation via the suppression of WT1 expression in Molt4 and K562 cells and Bcr/Abl expression in K562 cells. The active compound may be mammea E/BB. Extracts from M. siamensis flowers show promise as naturally occurring anti-cancer drugs.

Doxorubicin-Loaded Micelle Targeting MUC1: A Potential Therapeutic for MUC1 Triple Negative Breast Cancer Treatment

BACKGROUND Triple negative breast cancer (TNBC) is an aggressive disease associated with poor prognosis and lack of validated targeted therapy. Thus chemotherapy is a main adjuvant treatment for TNBC patients, but it associates with severe toxicities. For a better treatment outcome, we developed an alternative therapeutic, doxorubicin (DOX)-loaded micelles targeting human mucin1 protein (MUC1) that is less toxic, more effective and targeted to TNBC. METHODS From many candidate peptides, QNDRHPR-GGGSK (QND) and HSQLPQV-GGGSK (HSQ) were identified computationally, synthesized and purified using solid phase peptide synthesis and semipreparative HPLC. The peptides showed significant high binding to MUC1 expressing cells using a fluorescent microscope. The peptides were then conjugated on pegylated octadecyl lithocholate copolymer. DOX-encapsulated micelles were formed through self-assembly. MUC1-targeted micelles were characterized using dynamic light scattering (DLS) and Transmission Electron Microscopy (TEM). Drug entrapment efficiency was examined using a microplate reader. Cytotoxicity, binding, and uptake were also investigated. RESULTS Two types of DOX-loaded micelles with different targeting peptides, QND or HSQ, were developed. DOX-loaded micelles were spherical in shape with average particle size around 300-320 nm. Drug entrapment efficiency of untargeted and targeted DOX micelles was about 71-93%. Targeted QND-DOX and HSQ-DOX micelles exhibited significantly higher cytotoxicity compared to free DOX and untargeted DOX micelles on BT549-Luc cells. In addition, significantly greater binding and uptake were observed for QND-DOX and HSQ-DOX micelles on BT549-Luc and T47D cells. CONCLUSION Taken together, these results suggested that QND-DOX and HSQ-DOX micelles have a potential application in the treatment of TNBC-expressing MUC1.