Cisplatin sensitivity of testis tumour cells is due to deficiency in interstrand-crosslink repair and low ERCC1-XPF expression.
BACKGROUND
Cisplatin based chemotherapy cures over 80% of metastatic testicular germ cell tumours (TGCT). In contrast, almost all other solid cancers in adults are incurable once they have spread beyond the primary site. Cell lines derived from TGCTs are hypersensitive to cisplatin reflecting the clinical response. Earlier findings suggested that a reduced repair capacity might contribute to the cisplatin hypersensitivity of testis tumour cells (TTC), but the critical DNA damage has not been defined. This study was aimed at investigating the formation and repair of intrastrand and interstrand crosslinks (ICLs) induced by cisplatin in TTC and their contribution to TTC hypersensitivity.


RESULTS
We observed that repair of intrastrand crosslinks is similar in cisplatin sensitive TTC and resistant bladder cancer cells, whereas repair of ICLs was significantly reduced in TTC. γH2AX formation, which serves as a marker of DNA breaks formed in response to ICLs, persisted in cisplatin-treated TTC and correlated with sustained phosphorylation of Chk2 and enhanced PARP-1 cleavage. Expression of the nucleotide excision repair factor ERCC1-XPF, which is implicated in the processing of ICLs, is reduced in TTC. To analyse the causal role of ERCC1-XPF for ICL repair and cisplatin sensitivity, we over-expressed ERCC1-XPF in TTC by transient transfection. Over-expression increased ICL repair and rendered TTC more resistant to cisplatin, which suggests that ERCC1-XPF is rate-limiting for repair of ICLs resulting in the observed cisplatin hypersensitivity of TTC.


CONCLUSIONS
Our data indicate for the first time that the exceptional sensitivity of TTC and, therefore, very likely the curability of TGCT rests on their limited ICL repair due to low level of expression of ERCC1-XPF.
Usanova
Svetlana
S
Institute of Toxicology, Clinical Centre of University of Mainz, Obere Zahlbacher Strasse 67, 55131 Mainz, Germany.
Piée-Staffa
Andrea
A
Sied
Ulrike
U
Thomale
Jürgen
J
Schneider
Astrid
A
Kaina
Bernd
B
Köberle
Beate
B
D000818:Animals; D017209:Apoptosis / Q000235:genetics / Q000502:physiology; D016466:CHO Cells; D045744:Cell Line, Tumor; D000071877:Checkpoint Kinase 1; D064447:Checkpoint Kinase 2; D002945:Cisplatin / Q000627:therapeutic use*; D006224:Cricetinae; D003412:Cricetulus; D004260:DNA Repair / Q000235:genetics*; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D004720:Endonucleases / Q000235:genetics / Q000378:metabolism*; D005434:Flow Cytometry; D006801:Humans; D015151:Immunoblotting; D007150:Immunohistochemistry; D008297:Male; D010766:Phosphorylation / Q000187:drug effects; D000071137:Poly (ADP-Ribose) Polymerase-1; D011065:Poly(ADP-ribose) Polymerases / Q000235:genetics / Q000378:metabolism; D011494:Protein Kinases / Q000235:genetics / Q000378:metabolism; D017346:Protein Serine-Threonine Kinases / Q000235:genetics / Q000378:metabolism; D013736:Testicular Neoplasms / Q000188:drug therapy* / Q000378:metabolism*; D001749:Urinary Bladder Neoplasms / Q000188:drug therapy / Q000378:metabolism
D004268:DNA-Binding Proteins; C095888:xeroderma pigmentosum group F protein; C499320:PARP1 protein, human; D000071137:Poly (ADP-Ribose) Polymerase-1; D011065:Poly(ADP-ribose) Polymerases; D011494:Protein Kinases; D064447:Checkpoint Kinase 2; C578634:CHEK2 protein, human; D000071877:Checkpoint Kinase 1; D017346:Protein Serine-Threonine Kinases; C071447:ERCC1 protein, human; D004720:Endonucleases; D002945:Cisplatin