A Research On Effects And Mechanisms Of Chlorogenic Acid Inhibiting Leukemic Cells

Leukemia is the most common malignant neoplasm of hematopoietic system. nowadays,chemical therapy and haemopoietic stem cell transplantation are main measures of leukemia therapy.But , chemical therapy can only gain transient relief and 5-year disease free survival rate of haemopoietic stem cell transplantation is 35% — 50 % .In addition,because of the constraint of stem cell donor,only a few people can receive HSCT . HSCT exsits many chancinesses associated with transplantation and recurrence. For the past many years, hematologists have been searching for medicines to cure leukemia worldwild. In a recent study, a Bcr-Abl kinase inhibitor was identified from an herbal source. The compound was characterized to be chlorogenic acid (Chl). Chlorogenic acid has diverse biologic activities. Chl inhibits Bcr-Abl kinases and induces apoptosis of Bcr-Abl-positive cells including Bcr-Abl-positive primary leukemic cells of CML patients in vitro. Berbamine is a calcium ion channel blocker. In our past study, berbamine exhibits selective growth inhibitory effects on Bcr-Abl-positive CML cell line K562 including Bcr-Abl-positive primary leukemic cells of CML patients in vitro and have little cytotoxicityto normal myeloid element. Accordingly, leukemic cell lines of KG1 , K562 and Jurkat are adopted and affected debita spissitudine chlorogenic acid. We observe the inhibition of cells treated by Chl. We also observe the unification of Chl and berbamine.Materials and methodsMaterialsHuman leukemia cell lines:KGl,K562 and Jurkat cell lines. KGl cell line for AML-M1, K562 cell line for AML-M6, and Jurkat for acute lymphoblastic leukemia (ALL).Methods1. Cell inoculation: We use cell lines on logarithmic growth phase. Inoculum density is 8×10³/well.2. Drug treatment: Na-chlorogenic acid and berbamine treat cell lines on graded concentration. Blank well is negative control. Cells are cultivated in 37°C, 5% CO₂ incubator for 48 hours.3. MTT effect: Adding 200ulMTT per well and cultivating for 4 hours.4. Measuring absorbance: centrifuging and discarding supernatant. Adding 200ul DMSO per well. Agitating for 5 minutes and measuring absorbance at 470nm.5. Calculating inhibition ratio and IC50 .Plotting to observe apoptosis.ResultsNa-chlorogenic acid in —20°C exhibits growth inhibitory effects on Bcr-Abl-positive CML cell line K562 in concentration dependent. IC50 is 24.210ug/ml,but have no inhibitoryeffects on KG1,Jurkat cell lines. Na-chlorogenic acid in 4°C exhibits stronger growth inhibitory effects on K562,and have inhibitory effects on Jurkat. It has no growth inhibitory effects on KG1. IC50(K562) is 13.177 ug/ml, IC50(Jurkat) is 19.640ug/ml.Berbamine induces apoptosis of Bcr-Abl-positive cell line K562 in concentration dependent. IC50 is 6.080ug/ml.The unification of Na-chlorogenic acid and berbamine on K562 is stronger than solo effect of Na-chlorogenic or berbamine.Conclusions1. Na-chlorogenic acid in — 20°C exhibits selective growth inhibitory effects on Bcr-Abl-positive CML cell line K562.2. Na-chlorogenic acid in 4°C exhibits stronger growth inhibitory effects on K562,and have inhibitory effects on Jurkat.3. Chlorogenic acid and berbamine have co-inhibitory effect on K562.