2-Chloroethyl-3-sarcosinamide-1-nitrosourea (SarCNU) exhibits p53-dependent and -independent antiproliferative activity in human nasopharyngeal carcinoma cells in vitro and in vivo.
2-Chloroethyl-3-sarcosinamide-1-nitrosourea (SarCNU) has been used to treat patients with advanced solid tumours. However, the molecular mechanisms are not well understood. In the present study, we report that SarCNU inhibited proliferation of human HK-1 and CNE-2 nasopharyngeal carcinoma (NPC) in vivo and in vitro. In vitro study showed that wild-type p53 HK-1 cells were 3-fold more sensitive to SarCNU than p53 mutant CNE-2 cells. G2/M arrest, reduction in p21(Cip1/Waf1) and inactivation of cellular cdc-2 activity were seen in both SarCNU-treated HK-1 and CNE-2 cells. Upregulation of p53, phosphorylated p53 at Ser15 and biochemical markers for apoptosis, such as cleaved caspase-3, cleaved caspase-7 and cleaved PARP, were observed in SarCNU-treated HK-1 but not CNE-2 cells. The levels of cyclin B1, Wee1 and phosphorylated cdc-2 but not total cdc-2 in HK-1 cells were significantly reduced by SarCNU treatment. In contrast to HK-1 cells, decrease in total cdc-2 but increase in phosphorylated cdc-2 at Tyr15, cyclin B1 and Wee1 was observed in CNE-2 cells treated with SarCNU. Introduction of mutant p53 into HK-1 cells resulted in growth enhancement in vivo and increased resistance to SarCNU-induced apoptosis in vitro. Furthermore, CNE-2 cells transfected with wild-type p53 became susceptible to SarCNU-induced apoptosis in vitro but not their growth rate in vivo. The data indicate that in NPC cells SarCNU-induced apoptosis was p53-dependent while SarCNU-induced G2/M arrest was mediated by altering the levels of cyclin B1-cdc-2 complex and phosphorylation of cdc-2 at Tyr15 resulting in inactivation of cellular cdc-2 activity. Our data suggest a potential use of SarCNU in the treatment of NPC.
Nguyen
T H
TH
Laboratory of Molecular Endocrinology, Division of Cellular and Molecular Research, National Cancer Centre of Singapore, Singapore 169610.
Ong
C K
CK
Wong
E
E
Leong
C T
CT
Panasci
L
L
Huynh
H
H
D000818:Animals; D000970:Antineoplastic Agents / Q000494:pharmacology; D017209:Apoptosis; D015153:Blotting, Western; D016203:CDC2 Protein Kinase / Q000378:metabolism; D002277:Carcinoma / Q000473:pathology*; D002330:Carmustine / Q000031:analogs & derivatives* / Q000494:pharmacology; D053148:Caspase 3; D053179:Caspase 7; D020169:Caspases / Q000378:metabolism; D002453:Cell Cycle; D018797:Cell Cycle Proteins / Q000378:metabolism; D002455:Cell Division; D049109:Cell Proliferation; D002470:Cell Survival; D019926:Cyclin B / Q000378:metabolism; D056744:Cyclin B1; D050759:Cyclin-Dependent Kinase Inhibitor p21 / Q000378:metabolism; D018076:DNA, Complementary / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D016195:G2 Phase; D016158:Genes, p53; D015316:Genetic Therapy / Q000379:methods; D006801:Humans; D047468:Immunoprecipitation; D066298:In Vitro Techniques; D008297:Male; D051379:Mice; D016513:Mice, SCID; D008938:Mitosis; D009154:Mutation; D009303:Nasopharyngeal Neoplasms / Q000473:pathology*; D009687:Nuclear Proteins / Q000378:metabolism; D010766:Phosphorylation; D011505:Protein-Tyrosine Kinases / Q000378:metabolism; D013997:Time Factors; D014162:Transfection; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism*; D014443:Tyrosine / Q000737:chemistry; D015854:Up-Regulation
D000970:Antineoplastic Agents; C534699:CCNB1 protein, human; C493956:CDKN1A protein, human; C534700:Ccnb1 protein, mouse; D018797:Cell Cycle Proteins; D019926:Cyclin B; D056744:Cyclin B1; D050759:Cyclin-Dependent Kinase Inhibitor p21; D018076:DNA, Complementary; D009687:Nuclear Proteins; D016159:Tumor Suppressor Protein p53; D014443:Tyrosine; C044565:2-((((2-chloroethyl)nitrosoamino)carbonyl)amino)propanamide; D011505:Protein-Tyrosine Kinases; C499349:WEE1 protein, human; D016203:CDC2 Protein Kinase; C505407:CASP3 protein, human; C505520:CASP7 protein, human; C505409:Casp3 protein, mouse; C505524:Casp7 protein, mouse; D053148:Caspase 3; D053179:Caspase 7; D020169:Caspases; D002330:Carmustine