Modulating multidrug resistance gene in leukaemia cells by short interfering RNA.
BACKGROUND
The multidrug resistance gene, MDR1, is one of the genes responsible for resistance to chemotherapy in the treatment of leukaemia and other cancers. The discovery of RNA interference in mammalian cells has provided a powerful tool to inhibit the expression of this gene. However, very little is known about the transfection of leukaemia cells with short interfering RNA (siRNA) targeted at MDR1. This study aims to evaluate the effectiveness of two chemically-synthesised siRNA in modulating MDR1 gene and inhibiting P-glycoprotein expression in leukaemic cells. We also evaluated two siRNA delivery methods in this study.


METHODS
K562/Adr was transfected with two MDR1-targeted siRNA or negative control siRNA, by using cationic lipid-based transfection reagents or electroporator. Gene expression of MDR1 was quantified by real-time polymerase chain reaction and calculated as a percentage relative to the negative control siRNA. P-glycoprotein expression was evaluated via flow cytometry and drug sensitivity after treatment was assessed by cytotoxicity assays.


RESULTS
The percentage of MDR1 gene knockdown from cells transfected with an electroporator was significantly higher (84.4 percent, p-value is 0.094) compared to cells transfected with cationic lipid-based transfection reagents (52.8 percent). Both siRNA significantly reduced the expression of MDR1 by 84.9 percent (p-value is 0.001) and 86.0 percent (p-value is 0.011), respectively. P-glycoprotein expression was down-regulated and drug sensitivity was increased after treatment with the siRNA.


CONCLUSIONS
This study shows that the two siRNA sequences are capable of modulating MDR1 and P-glycoprotein expressions and increased drug sensitivity. Transfection with an electroporator was superior to chemical transfection for leukaemia cells.
Lim
M N
MN
Haematology Unit, Cancer Research Centre, Institute for Medical Research, Jalan Pahang, Kuala Lumpur 50588, Malaysia. limmn@imr.gov.my
Lau
N S
NS
Chang
K M
KM
Leong
C F
CF
Zakaria
Z
Z
D018435:ATP Binding Cassette Transporter, Subfamily B; D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1 / Q000037:antagonists & inhibitors* / Q000235:genetics; D000970:Antineoplastic Agents / Q000494:pharmacology; D004317:Doxorubicin / Q000494:pharmacology; D019008:Drug Resistance, Neoplasm / Q000187:drug effects / Q000235:genetics; D015972:Gene Expression Regulation, Neoplastic / Q000235:genetics; D019450:Genes, MDR / Q000235:genetics; D006801:Humans; D007938:Leukemia / Q000188:drug therapy* / Q000235:genetics*; D034741:RNA, Small Interfering / Q000008:administration & dosage / Q000235:genetics / Q000494:pharmacology*; D014162:Transfection; D014407:Tumor Cells, Cultured
C513055:ABCB1 protein, human; D018435:ATP Binding Cassette Transporter, Subfamily B; D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D000970:Antineoplastic Agents; D034741:RNA, Small Interfering; D004317:Doxorubicin