CD133+ cells contribute to radioresistance via altered regulation of DNA repair genes in human lung cancer cells.
BACKGROUND
Radioresistance in human tumors has been linked in part to a subset of cells termed cancer stem cells (CSCs). The prominin 1 (CD133) cell surface protein is proposed to be a marker enriching for CSCs. We explore the importance of DNA repair in contributing to radioresistance in CD133+ lung cancer cells.


METHODS
A549 and H1299 lung cancer cell lines were used. Sorted CD133+ cells were exposed to either single 4 Gy or 8 Gy doses and clonogenic survival measured. ϒ-H2AX immunofluorescence and quantitative real time PCR was performed on sorted CD133+ cells both in the absence of IR and after two single 4 Gy doses. Lentiviral shRNA was used to silence repair genes.


RESULTS
A549 but not H1299 cells expand their CD133+ population after single 4 Gy exposure, and isolated A549 CD133+ cells demonstrate IR resistance. This resistance corresponded with enhanced repair of DNA double strand breaks (DSBs) and upregulated expression of DSB repair genes in A549 cells. Prior IR exposure of two single 4 Gy doses resulted in acquired DNA repair upregulation and improved repair proficiency in both A549 and H1299. Finally Exo1 and Rad51 silencing in A549 cells abrogated the CD133+ IR expansion phenotype and induced IR sensitivity in sorted CD133+ cells.


CONCLUSIONS
CD133 identifies a population of cells within specific tumor types containing altered expression of DNA repair genes that are inducible upon exposure to chemotherapy. This altered gene expression contributes to enhanced DSB resolution and the radioresistance phenotype of these cells. We also identify DNA repair genes which may serve as promising therapeutic targets to confer radiosensitivity to CSCs.
Department of Pharmacology, Seidman Cancer Center, Case Western Reserve University and University Hospitals Case Medical Center, Cleveland, United States; Division of Hematology/Oncology, National Center for Regenerative Medicine, Seidman Cancer Center, Case Western Reserve University and University Hospitals Case Medical Center, Cleveland, United States.
Amar
A
Desai
Division of Hematology/Oncology, National Center for Regenerative Medicine, Seidman Cancer Center, Case Western Reserve University and University Hospitals Case Medical Center, Cleveland, United States.
Bryan
B
Webb
Department of Pharmacology, Seidman Cancer Center, Case Western Reserve University and University Hospitals Case Medical Center, Cleveland, United States; Division of Hematology/Oncology, National Center for Regenerative Medicine, Seidman Cancer Center, Case Western Reserve University and University Hospitals Case Medical Center, Cleveland, United States; Case Comprehensive Cancer Center, Seidman Cancer Center, Case Western Reserve University and University Hospitals Case Medical Center, Cleveland, United States. Electronic address: slg5@case.edu.
Stanton L
SL
Gerson
D000071916:AC133 Antigen; D015703:Antigens, CD / Q000032:analysis*; D045744:Cell Line, Tumor; D053903:DNA Breaks, Double-Stranded; D004260:DNA Repair / Q000235:genetics*; D006023:Glycoproteins / Q000032:analysis*; D006801:Humans; D008175:Lung Neoplasms / Q000235:genetics / Q000532:radiotherapy*; D014411:Neoplastic Stem Cells / Q000528:radiation effects*; D010455:Peptides / Q000032:analysis*; D051135:Rad51 Recombinase / Q000502:physiology; D011836:Radiation Tolerance / Q000235:genetics
D000071916:AC133 Antigen; D015703:Antigens, CD; D006023:Glycoproteins; C000606023:PROM1 protein, human; D010455:Peptides; C495153:RAD51 protein, human; D051135:Rad51 Recombinase
CD133; Cancer stem cells; Radiation resistance