RESUMEN
BACKGROUND & OBJECTIVE: Epstein-Barr virus (EBV) is closely related to nasopharyngeal carcinoma (NPC). Many kinds of methods can be used to examine antibodies in NPC patients sera. This study was to screen the dominant epitopes from random peptide libraries (RPLs) displayed on phage using the EBV-related antibodies purified from the sera of NPC patients, and find new antigens at the epitope level. METHODS: The EBV-related antibodies were eluted from the sera of NPC patients using B95-8 cell EBV proteins as antigen, and the phages from 12-mer RPLs were elutriated for 3 rounds with the antibodies. The positive clones were gained by sandwich ELISA, and competitive inhibition assay from the third elution. The positive clones were sequenced, and the peptide coded by the inserted DNA were blasted with the antigen region of EBV proteins. RESULTS: Sixty-four phage clones were randomly picked up from the third eluate,25 positive clones were picked up using sandwich ELISA assay, the positive percentage was 39.06%. Thirteen clones were picked up for competitive ELISA assay,11 clones showed inhibitory phenomena,the inhibitory rates were between 18.09% and 65.94%. Five positive clones with high absorbency value, and high inhibitory rates were selected out, the sequences of peptides displayed on these clones were -A-T-S-H-L-H-V-R-L-P-W-T- (d15, and d18), -G-S-T-H-K-H-N-H-F-N-K-T- (d19), -K-P-I-H-E-H-P-H-R-F-K-S- (e8), -H-T-H-K-I-K-I-P-L-P-I-Q- (e23). These peptide sequences showed similarity with the amino acid sequences located in antigen regions of EBV integral membrane protein (d15, and d18), EBV thymidine kinase (d19), and EBV major capsid protein (e8, and e23). CONCLUSION: EBV-related epitopes could be obtained by screening the phages from RPLs with polyclonal antibodies purified from the sera of NPC patients, which may offer new methods of antigen preparation for sera diagnosis of NPC.
Asunto(s)
Anticuerpos Antineoplásicos/análisis , Anticuerpos Antivirales/análisis , Epítopos/análisis , Herpesvirus Humano 4/inmunología , Neoplasias Nasofaríngeas/inmunología , Secuencia de Aminoácidos , Anticuerpos Antineoplásicos/genética , Anticuerpos Antivirales/genética , Antígenos de Neoplasias/genética , Antígenos de Neoplasias/inmunología , Antígenos Virales/genética , Antígenos Virales/inmunología , Proteínas de la Cápside/genética , Epítopos/genética , Herpesvirus Humano 4/genética , Humanos , Proteínas de la Membrana/genética , Datos de Secuencia Molecular , Neoplasias Nasofaríngeas/genética , Biblioteca de Péptidos , Homología de Secuencia de Aminoácido , Timidina Quinasa/genéticaRESUMEN
BACKGROUND & OBJECTIVE: Recent researches manifested that down-regulation of p21(WAF1) had relationship with carcinogenesis and development in various tumors, but its association with epithelial ovarian cancer (EOC) was not clear. This study was designed to investigate the role of p21(WAF1) in the tumorigenesis and development of EOC and its relationship with p53 and proliferating cell nuclear antigen (PCNA) protein. METHODS: Fifty-five EOC tissues, 32 benign ovarian tumor tissues, and 30 normal ovarian tissues were collected. Reverse transcription polymerase chain reaction (RT-PCR) was applied to determine the p21(WAF1)mRNA expression. Immunohistochemistry was applied to examine the protein expression of p21(WAF1), p53, and PCNA. The relationship between the expression of these markers and the clinicopathological characteristics, prognosis of the patients was analyzed. RESULTS: The positive rates of p21(WAF1)mRNA in EOC, benign ovarian tumor, and normal ovary were 40%, 56.25%, and 73.33%, respectively (P=0.012). The positive rates of p21(WAF1) protein were 36.36%, 56.25%, and 80%, respectively (P=0.001). The positive expression rates of p21(WAF1)mRNA and its protein in EOC were lower than those of the other two groups, while the positive expression rates of p53 and PCNA protein in EOC were higher than those of the other two groups (P< 0.05). Expression of p21(WAF1)mRNA had positive relation to its protein, negative relation to PCNA protein, no relation to p53 protein, while expression of p21(WAF1) protein had negative relation to p53 and PCNA protein in EOC. Low-expression of p21(WAF1) protein was associated with advanced FIGO stage (P=0.032), but not with age, histological type, pathological grade, and remnant tumor (P >0.05). There was no relationship between p21(WAF1)mRNA and former parameters (P >0.05). Univariate analysis showed that the patients with low-expression of p21(WAF1)mRNA and p21(WAF1) protein had poor prognosis (P< 0.05). CONCLUSION: p21(WAF1) is down-regulated in EOC. p21(WAF1) might be able to be used as a marker to predict the prognosis of patients with EOC.