Anti-CHK1 (N-terminal) polyclonal antibody

There are few reports describing the role of p53-dependent gene repression in apoptotic cell death. To identify such apoptosis-associated p53 target genes, we used the pro-oxidant plant-derived drug thymoquinone and compared p53+/+ and p53-/- colon cancer cells HCT116. The p53 wild-type (wt) status correlated with more pronounced DNA damage and higher apoptosis after thymoquinone treatment. A significant up-regulation of the survival gene CHEK1 was observed in p53-/- cells in response to thymoquinone due to the lack of transcriptional repression of p53. In p53-/- cells, transfection with p53-wt vector and CHEK1 small interfering RNA treatment decreased CHEKI mRNA and protein levels and restored apoptosis to the levels of the p53+/+ cells. p53-/- cells transplanted to nude mice treated with thymoquinone up-regulated CHEK1 expression and did not undergo apoptosis unlike p53+/+ cells. Immunofluorescence analysis revealed that the apoptosis resistance in p53-/- cells after thymoquinone treatment might be conveyed by shuttling of CHEK1 into the nucleus. We confirmed the in Vivo existence of this CHEK1/p53 link in human colorectal cancer, showing that tumors lacking p53 had higher levels of CHEK1, which was accompanied by poorer apoptosis. CHEKI overexpression was correlated with advanced tumor stages (P = 0.03), proximal tumor localization (P = 0.02), and worse prognosis (1.9-fold risk, univariate Cox regression; Kaplan-Meier, P = 0.04). We suggest that the inhibition of the stress response sensor CHEK1 might contribute to the antineoplastic activity of specific DNA-damaging drugs.

Radiotherapy is the most successful nonsurgical treatment for nasopharyngeal carcinoma (NPC). Despite this, the prognosis remains poor. Although NPCs initially respond well to a full course of radiation, recurrence is frequent. The cancer stem cell (CSC) hypothesis provides a framework for explaining the discrepancy between the response of NPC to therapy and the poor survival rate. In this study, a stem cell-like subpopulation (PKH26+) was identified in NPC cell lines using a label-retention technique. PKH26+ cells were enriched for clonogenicity, sphere formation, side-population cells, and resistance to radiotherapy. Using genomic approaches, we show that the proto-oncogene c-MYC (MYC) regulates radiotolerance through transcriptional activation of CHK1 (CHEK1) and CHK2 (CHEK2) checkpoint kinases through direct binding to the CHK1 and CHK2 promoters. Overexpression of c-MYC in the PKH26+ subpopulation leads to increased expression of CHK1 and CHK2 and subsequent activation of the DNA-damage-checkpoint response, resulting in radioresistance. Furthermore, loss of CHK1 and CHK2 expression reverses radioresistance in PKH26+ (c-MYC high expression) cells in vitro and in vivo. This study elucidates the role of the c-MYC-CHK1/CHK2 axis in regulating DNA-damage-checkpoint responses and stem cell characteristics in the PKH26+ subpopulation. Furthermore, these data reveal a potential therapeutic application in reversal of radioresistance through inhibition of the c-MYC-CHK1/CHK2 pathway. Cancer Res; 73(3); 1219-31. (c) 2012 AACR.