Synthetic lethality by lentiviral short hairpin RNA silencing of thymidylate kinase and doxorubicin in colon cancer cells regardless of the p53 status.
Intracellular supply of dTTP is a highly regulated process and has been a key target for chemotherapeutic drug development. Thymidylate kinase (TMPK) is the key enzyme for dTTP formation in both de novo and salvage pathways. In this study, we used lentiviral-based small hairpin RNA to silence TMPK expression in p53(+/+) and p53(-/-) HCT-116 colon cancer cells. This approach was sufficient to decrease the dTTP pool gradually without affecting p53 expression and generating cytotoxicity. TMPK knockdown significantly increased doxorubicin sensitivity dramatically in p53-proficient, p53-null HCT-116, and LoVo colon cancer cells. The decrease in the dTTP pool using this approach augmented the DNA damage response and enhanced apoptotic induction after exposure to low-dose doxorubicin, leading to cell death. In contrast, silencing of thymidylate synthase which blocks the de novo pathway was incapable of sensitizing p53-null HCT-116 cells to doxorubicin-induced apoptosis because of the compensation by the salvage pathway. Our results suggest the lentiviral delivery of small hairpin RNA targeting TMPK in combination with a low dose of doxorubicin as a new approach to kill colon cancer cells regardless of p53 status.
Hu
Chun-Mei
CM
Graduate Institute of Biochemistry and Molecular Biology, College of Medicine, National Taiwan University, Taipei, Taiwan, ROC.
Chang
Zee-Fen
ZF
D000911:Antibodies, Monoclonal; D045744:Cell Line, Tumor; D003110:Colonic Neoplasms; D004249:DNA Damage; D004317:Doxorubicin / Q000633:toxicity*; D020868:Gene Silencing; D005822:Genetic Vectors; D006801:Humans; D016086:Lentivirus / Q000235:genetics*; D009703:Nucleoside-Phosphate Kinase / Q000235:genetics*; D016337:RNA, Catalytic / Q000235:genetics*; D034741:RNA, Small Interfering / Q000235:genetics*; D012367:RNA, Viral / Q000235:genetics*; D016159:Tumor Suppressor Protein p53 / Q000037:antagonists & inhibitors* / Q000172:deficiency / Q000235:genetics*
D000911:Antibodies, Monoclonal; D016337:RNA, Catalytic; D034741:RNA, Small Interfering; D012367:RNA, Viral; D016159:Tumor Suppressor Protein p53; D004317:Doxorubicin; D009703:Nucleoside-Phosphate Kinase; C020093:dTMP kinase