Down-regulation of GRP78 is associated with the sensitivity of chemotherapy to VP-16 in small cell lung cancer NCI-H446 cells.
BACKGROUND
Chemotherapy resistance remains a major obstacle for the treatment of small cell lung cancer (SCLC). Glucose-regulated protein 78 (GRP78), an endoplasmic reticulum chaperone, plays a critical role in chemotherapy resistance in some cancers. However, whether the suppression of the chaperone can enhance the sensitivity of chemotherapy in SCLC is still unclear.


METHODS
The SCLC NCI-H446 cells were divided into three groups: BAPTA-AM-->A23187-treated group, A23187-treated group and control-group. Immunofluorescence, western blot and RT-PCR were used to assess the expression of GRP78 at both protein and mRNA levels. Cell apoptosis and the cell cycle distributions of the cells were analyzed by flow cytometry in order to evaluate the therapeutic sensitivity to VP-16.


RESULTS
The expression of GRP78 at both protein and mRNA levels in the BAPTA-AM-->A23187-treated cells dramatically decreased as compared to that in both A23187-treated and control groups. After treatment by VP-16, the percentage of apoptotic cells in BAPTA-AM-->A23187-treated cells were: 33.4 +/- 1.01%, 48.2 +/- 1.77%, 53.0 +/- 1.43%, 56.5 +/- 2.13%, respectively, corresponding to the concentrations of BAPTA-AM 10, 15, 25, 40 microM, which was statistically significant high in comparison with the A23187-treated group and untreated-group (7.18 +/- 1.03% and 27.8 +/- 1.45%, respectively, p < 0.05). The results from analysis of cell cycle distribution showed that there was a significantly decreased in G1 phase and a dramatically increased in S phase for the BAPTA-AM-->A23187-treated cells as compared with the untreated cells.


CONCLUSIONS
BAPTA-AM is a strong inhibitor of GRP78 in the NCI-H446 cell line, the down-regulation of GRP78 can significantly increase the sensitivity to VP-16. The suppression of GRP78 may offer a new surrogated therapeutic approach to the clinical management of lung cancer.
Wang
Yingyan
Y
Diagnostics Laboratory Center, Dalian Medical University, Liaoning, PR China. wyydmu@yahoo.com.cn
Wang
Wei
W
Wang
Siyan
S
Wang
Jiarui
J
Shao
Shujuan
S
Wang
Qi
Q
D000972:Antineoplastic Agents, Phytogenic / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects; D015153:Blotting, Western; D000001:Calcimycin / Q000494:pharmacology; D002453:Cell Cycle / Q000187:drug effects; D045744:Cell Line, Tumor; D015536:Down-Regulation; D004533:Egtazic Acid / Q000031:analogs & derivatives / Q000494:pharmacology; D000091342:Endoplasmic Reticulum Chaperone BiP; D005047:Etoposide / Q000494:pharmacology*; D005434:Flow Cytometry; D005455:Fluorescent Antibody Technique; D006360:Heat-Shock Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D008175:Lung Neoplasms / Q000188:drug therapy* / Q000378:metabolism*; D018832:Molecular Chaperones / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D055752:Small Cell Lung Carcinoma / Q000188:drug therapy* / Q000378:metabolism*
D000972:Antineoplastic Agents, Phytogenic; D000091342:Endoplasmic Reticulum Chaperone BiP; C000717247:HSPA5 protein, human; D006360:Heat-Shock Proteins; D018832:Molecular Chaperones; D012333:RNA, Messenger; C070379:1,2-bis(2-aminophenoxy)ethane N,N,N',N'-tetraacetic acid acetoxymethyl ester; D000001:Calcimycin; D004533:Egtazic Acid; D005047:Etoposide