Fractionated irradiation of H69 small-cell lung cancer cells causes stable radiation and drug resistance with increased MRP1, MRP2, and topoisomerase IIalpha expression.
OBJECTIVE
After standard treatment with chemotherapy and radiotherapy, small-cell lung cancer (SCLC) often develops resistance to both treatments. Our aims were to establish if fractionated radiation treatment alone would induce radiation and drug resistance in the H69 SCLC cell line, and to determine the mechanisms of resistance.


METHODS
H69 SCLC cells were treated with fractionated X-rays to an accumulated dose of 37.5 Gy over 8 months to produce the H69/R38 subline. Drug and radiation resistance was determined using the MTT (3,-4,5 dimethylthiazol-2,5 diphenyltetrazolium bromide) cell viability assay. Protein expression was analyzed by Western blot.


RESULTS
The H69/R38 subline was resistant to radiation (2.0 +/- 0.2-fold, p < 0.0001), cisplatin (14 +/- 7-fold, p < 0.001), daunorubicin (6 +/- 3-fold, p < 0.05), and navelbine (1.7 +/- 0.15-fold, p < 0.02). This was associated with increased expression of the multidrug resistance-associated proteins, MRP1 and MRP2, and topoisomerase IIalpha and decreased expression of glutathione-S-transferase pi (GSTpi) and bcl-2 and decreased cisplatin accumulation. Treatment with 4 Gy of X-rays produced a 66% decrease in MRP2 in the H69 cells with no change in the H69/R38 cells. This treatment also caused a 5-fold increase in topoisomerase IIalpha in the H69/R38 cells compared with a 1.5-fold increase in the H69 cells.


CONCLUSIONS
Fractionated radiation alone can lead to the development of stable radiation and drug resistance and an altered response to radiation in SCLC cells.
Henness
Sheridan
S
Bill Walsh Cancer Research Laboratories, Medical Oncology Department, Royal North Shore Hospital, St. Leonards, Australia.
Davey
Mary W
MW
Harvie
Rozelle M
RM
Davey
Ross A
RA
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1 / Q000378:metabolism; D000970:Antineoplastic Agents / Q000493:pharmacokinetics; D018288:Carcinoma, Small Cell / Q000188:drug therapy* / Q000378:metabolism / Q000532:radiotherapy*; D002470:Cell Survival; D002945:Cisplatin / Q000493:pharmacokinetics; D004396:Coloring Agents; D004250:DNA Topoisomerases, Type II / Q000378:metabolism; D019583:Dose Fractionation, Radiation; D019008:Drug Resistance, Neoplasm / Q000235:genetics / Q000528:radiation effects*; D015870:Gene Expression; D005978:Glutathione / Q000032:analysis; D006801:Humans; D008175:Lung Neoplasms / Q000188:drug therapy* / Q000378:metabolism / Q000532:radiotherapy*; D026901:Membrane Transport Proteins; D000089762:Multidrug Resistance-Associated Protein 2; D027425:ATP-Binding Cassette, Sub-Family C Proteins / Q000378:metabolism; D009363:Neoplasm Proteins / Q000378:metabolism; D011836:Radiation Tolerance / Q000235:genetics / Q000528:radiation effects*; D013778:Tetrazolium Salts; D013844:Thiazoles; D014407:Tumor Cells, Cultured / Q000528:radiation effects
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D000970:Antineoplastic Agents; D002945:Cisplatin; D004396:Coloring Agents; D004250:DNA Topoisomerases, Type II; D005978:Glutathione; D026901:Membrane Transport Proteins; D000089762:Multidrug Resistance-Associated Protein 2; D027425:ATP-Binding Cassette, Sub-Family C Proteins; D009363:Neoplasm Proteins; D013778:Tetrazolium Salts; D013844:Thiazoles; C000715087:ABCC2 protein, human; C022616:thiazolyl blue; C421109:multidrug resistance-associated protein 1