| Objective To better investigate and understand the relationship between the drug-resistance related genes and chemotherapy response, prognosis in acute leukemia (AL), semi-quantitative and real time fluorescent quantitative reverse transcription polymerase chain reaction (RT-PCR) was used to detect the expression of the multidrug resistance gene-1 (mdr-1), multidrug resistance-associated protein gene (mrp), lung resistance-related protein gene (Irp), breast cancer resistance protein gene (bcrp), Glutathione S-transferases (GSTs), DNA Topoisomerase (Topo) and bcl-2 genes from patients with AL, from which experimental data for judging multidrug resistance (MDR) and clinical prognosis would be provided.Method 140 AL patients were divided into three groups: untreated (newly diagnosed) > complete remission (CR) and relapsed patients. In addition, sensitive and resistance groups were also designatedaccording to chemotherapy response. The expression level of the mdr-1 ,mrp, lip, bcrp ,GST a , GST ft, Topo I ,Topo II a JOpo II ?, bcl-2 genes were detected by semi-qantitative RT-PCR in peripheral blood or bone marrow sample from patients. After agarose gel electrophoresis, the relative expression level was calculated as related genes/ 0 2-MG or |3 -actin ratio. We also established more accurate real time fluorescent quantitative RT-PCR with TaqMan (FQ RT-PCR) to detect accurate mRNA copies of bcrp, mdr-1, mrp and lip.Results The results from semi-quantitative RT-PCR showed:I .The relative expression level of bcrp, lip, GST ft , Topo II a and TopoII 3 in AL group were significantly higher than that in normal subjects. GST a , however, was exactly reverse (p<0.05). 2.The expression of bcrp and mdr-1 was significantly higher in relapsed group comparing with untreated group. 3. Resistance group had higher mdr-1 levels than sensitive group. 4.The positive percentage of bcrp, mdr-1 and mrp expression in resistance group were higher than that in sensitive group (p<0.05). 5. Patients who overexpressed bcrp, mdr-1, imp and lip at the same time had a poorer response and lower CR rate. 6.Rank correlation analysis showed that overexpression of mdr-1 and GST a were highly correlated with clinical drug resistance. It was also found that there were significantly negative correlation between GST ft , Topo II a and clinical resistance. 7. Multivariable logistic regression analysis showed that mdr-1had a positive correlation with poor prognosis (p<0.05). 8. With linear correlation analysis, positive correlation were observed between bcrp and mdr-1, mdr-1 and Irp, mdr-1 and GST a , mrp and Irp; negative correlation were observed between mrp and GST n (p<0.05).In order to measure the copies of bcrp, mdr-1, mrp and lip mRNA more accurately, we established FQ RT-PCR. The result showed: 1. Copies of bcrp mRNA in AL group were significantly higher than that in normal one (p<0.05). 2. Copies of mdr-1 mRNA in relapsed groups were significantly higher than that in normal one (p<0.05). 3. Resistance group had significantly higher copies of mdr-1 mRNA than that in sensitive group. 4. Positive correlations were observed between mdr-1 and mrp, mdr-1 and Irp, mrp and Irp. 5. Expression level of mdr-1 and Irp were significantly correlated with clinical resistance through rank correlation analysis.Conclusion Through the study mentioned above, we can draw following conclusions: l.Overexpressions of one or more genes of bcrp, mdr-1, mrp and Irp are associated with MDR in AL. Bcrp, mdr-1, mrp and Irp can be the indicators for judging MDR. 2. To clarify the roles of GSTs and Topo in MDR, other factors or variables should be considered. Neither GSTs nor Topo alone is a good indicator for MDR. 3. In evaluation of MDR the combined detection of multi-genes are better than that of single gene. Occurance of MDR is the consequence of multiplemechanisms. Drug resistance could not be explain by the genes mentioned'above, new MDR mechanism remains to be explored.
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