[Modulation of human small cell lung cancer cell line GLC4/ADR multidrug resistance in the inhibition of multidrug resistance-associated protein and its antisense].
OBJECTIVE
To study the effect of antisense multidrug resistance-associated protein (MRP) RNA on multidrug resistance (MDR) in the human small cell lung cancer (SCLC) cell line GLC4/ADR, in which the overexpression of MRP gene is discovered.


METHODS
In using the plasmid pRC/RSV-MRP1 containing complete ORF of MRP as a template, two antisense recombinants targeting at the 5' and 3' regions were constructed with the application of the polymerase chain reaction (PCR) technique. Lipofectamine was conducted to transduce these antisense MRPs into the GLC4/ADR cells. Three clones (the GLC4/ADR-pcDNA3, MO; GLC4/ADR-MRP-5' region, Ma; GLC4/ADR-MRP-3' region, Mb) of transfectants after the selection of G418 were obtained. The expression of MRP protein was detected by the use of the Western blot and the MTT method for determination of chemosensitivity to ADR was used.


RESULTS
The antisense MRPs was found to be effectively expressed in the clones, being transfected with two different antisense MRPs. The MRP expression in these transfectants were inhibited at the rates of 14.0% (GLC4/ADR Ma) and 83.0% (GLC4/ADR Mb), respectively. Moreover, decrease in the ADR resistance was observed in the Ma and Mb at the rates of 9.5% and 28.4%. Although the intracellular ADR concentration was increased in these transfectants, the proliferation and cell cycle and their early apoptosis induced by the ADR, indicated no difference between the transfectants and parental cells.


CONCLUSIONS
It is possible to obtain the effective expression of the MRP antisense structures for the blocking of the mRNA translation in GLC4/ADR cell line and the fragment complementary to the 3'-region of MRP is more effective for the inhibition of the MRP than that related to the 5'-region of MRP. The antisense RNA may be a useful treatment in combination with the conventional chemotherapy for SCLC, in which the MRP overexpression usually occurs.
Peng
X
X
Departments of Medical Oncology and Cell & Molecular Biology Laboratory, Cancer Institute (Hospital), Chinese Academy of Medical Sciences, Peking Union Medical College, Beijing 100021, China.
Feng
F
F
Zhang
W
W
D018435:ATP Binding Cassette Transporter, Subfamily B / Q000235:genetics*; D000818:Animals; D000903:Antibiotics, Antineoplastic / Q000378:metabolism / Q000494:pharmacology*; D000970:Antineoplastic Agents / Q000378:metabolism / Q000494:pharmacology*; D000972:Antineoplastic Agents, Phytogenic / Q000494:pharmacology; D017209:Apoptosis; D018288:Carcinoma, Small Cell / Q000188:drug therapy / Q000235:genetics*; D002453:Cell Cycle / Q000187:drug effects; D002455:Cell Division; D003001:Cloning, Molecular; D004195:Disease Models, Animal; D004317:Doxorubicin / Q000378:metabolism / Q000494:pharmacology*; D018432:Drug Resistance, Multiple / Q000235:genetics*; D019008:Drug Resistance, Neoplasm / Q000235:genetics*; D005047:Etoposide / Q000494:pharmacology; D015870:Gene Expression; D019450:Genes, MDR; D005818:Genetic Engineering; D006801:Humans; D008175:Lung Neoplasms / Q000188:drug therapy / Q000235:genetics*; D051379:Mice; D008819:Mice, Nude; D016372:RNA, Antisense; D014407:Tumor Cells, Cultured
D018435:ATP Binding Cassette Transporter, Subfamily B; D000903:Antibiotics, Antineoplastic; D000970:Antineoplastic Agents; D000972:Antineoplastic Agents, Phytogenic; D016372:RNA, Antisense; D005047:Etoposide; D004317:Doxorubicin