Reversal of multidrug resistance by a liposome-MDR1 ribozyme complex.
OBJECTIVE
Multidrug resistance (MDR) is a major obstacle in cancer chemotherapy. We examined whether cationic liposome-mediated transfer of a ribozyme could reverse MDR.


METHODS
A ribozyme which cleaved codon 196 of MDR1 mRNA was constructed from synthetic oligonucleotides. The MDR1 ribozyme was mixed with N-(1-(2,3-dileoyloxy)propyl)-N,N,N-trimethylammonium methyl sulfate (DOTAP) to form a liposomal complex. The complex was used to treat two P-glycoprotein-producing MDR cell lines: MCF-7/R human breast cancer cells resistant to doxorubicin and MOLT-3/TMQ800 human ALL cells resistant to trimetrexate (TMQ). In order to investigate the differential sensitivity of these two cell lines to the liposome-ribozyme complex, cellular pharmacological studies including phase-contrast and confocal microscopic studies were performed.


RESULTS
Treatment with the liposome-ribozyme complex resulted in reversal of vincristine (VCR) resistance in MCF-7/R cells, but not in MOLT-3/TMQ800 cells. In MCF-7/R cells the treatment resulted in decreases in MDR1 mRNA expression and P-glycoprotein production, whereas no changes in these parameters were seen in MOLT-3/TMQ800 cells. Phase-contrast microscopy revealed that in MCF-7/R cells treatment with DOTAP led to the formation of cytoplasmic vacuoles, and treatment with latex beads resulted in the development of a shiny material in the cytoplasm. In contrast, in MOLT-3/TMQ800 cells hardly any morphological changes occurred. Confocal microscopic imaging showed cytoplasmic fluorescence in MCF-7/R cells after treatment with DOTAP/FITC-dextran or FITC-conjugated latex beads. In MOLT-3/TMQ800 cells no fluorescence was detected. Treatment with cytochalasin B abolished fluorescence in MCF-7/R cells after treatment with DOTAP/FITC-dextran or FITC-conjugated latex beads. These studies show that MCF-7/R cells have high endocytotic activity whereas MOLT-3/TMQ800 cells have little activity.


CONCLUSIONS
Endocytotic activity was correlated with the success of cationic liposome-mediated transfer of MDR1 ribozyme. Determination of endocytotic activity of target tumor cells may be predictive of efficacy of liposome-mediated gene transfer.
Masuda
Y
Y
Samuel Bronfman Department of Medicine, Mount Sinai School of Medicine, New York, NY 10029, USA.
Kobayashi
H
H
Holland
J F
JF
Ohnuma
T
T
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1 / Q000235:genetics*; D000970:Antineoplastic Agents / Q000494:pharmacology*; D001943:Breast Neoplasms / Q000188:drug therapy; D018432:Drug Resistance, Multiple / Q000235:genetics*; D004354:Drug Screening Assays, Antitumor; D004705:Endocytosis; D015870:Gene Expression; D018014:Gene Transfer Techniques; D006801:Humans; D008081:Liposomes; D018613:Microscopy, Confocal; D008858:Microscopy, Phase-Contrast; D016337:RNA, Catalytic; D012333:RNA, Messenger; D014407:Tumor Cells, Cultured
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D000970:Antineoplastic Agents; D008081:Liposomes; D016337:RNA, Catalytic; D012333:RNA, Messenger