CONSTRUCTION OF EUKARYOTIC EXPRESSION VECTOR FOR HUMAN CCL21 AND CHARACTERIZATION OF ITS CHEMOTACTIC ACTIVITY

Objective： To obtain recombinant human CCL21 with biological activity from eukary0tic expression system for further use in cancer gene therapy. Methods： A fragment of human CCL21 gene was obtained from pSK-hCCL21 plasmid digested by Xho I and BamH I, inserted into the responding sites of eukaryotic expression vector pVAX1, and then transfected into COS-7 cells by electroporation method. The expression of hCCL21 protein was detected by western blotting analysis. The in vitro chemotaxis assay was used to test the chemotactic function of the expression product to lymphocytes. Results： Human CCL21 protein was expressed by transfected COS-7 cells with recombinant plasmid containing hCCL21 gene, and was verified by western blotting. The in vitro chemotaxis assay demonstrated that human CCL21 protein had a potent chemotactic function to lymphocytes. Conclusion： Human CCL21 was successfully and transiently expressed in eukaryotic cells, which lays some foundation for the study of CCL21 gene therapy in murine tumor models.

Ki-67 index,progesterone receptor expression,histologic grade and tumor size in predicting breast cancer recurrence risk:A consecutive cohort study

Background:The 21-gene recurrence score(RS)assay has been recommended by major guidelines for treatment decision in hormone receptor(HR)-positive early breast cancer(EBC).However,the genomic assay is not accessible and affordable worldwide.Alternatively,an increasing number of studies have shown that traditional immunohistochemistry(IHC)can partially or even completely replace the role of the 21-gene genomic assay.Here,we developed and validated a predictive model(IHC3 model)combining the Ki-67 index,progesterone receptor(PR)expression,histologic grade,and tumor size to predict the recurrence risk of HR-positive EBC.Methods:The data from 389 patients(development set)with HR-positive,human epidermal growth factor receptor 2-negative,lymph node non-metastasized invasive breast cancer were used to construct the IHC3 model based on the Surexam®21-gene RS and the TAILORx clinical trial criteria.An additional 146 patients with the same characteristics constituted the validation set.The predictive accuracy of the IHC3 model was compared with that of Orucevic et al.’s nomogram.Invasive diseasefree survival(IDFS)was analyzed in the IHC3 predictive low-recurrence risk(pLR)group and the predictive high-recurrence risk(pHR)group.The Pearson chi-square test,Fisher exact test,and log-rank test were used for analysis.Results:The pLR and pHR group could be easily stratified using the decision tree model without network dependence.The accuracies of the IHC3 model were 86.1%in the development set and 87.7%in the validation set.The predictive accuracy of the IHC3 model and Orucevic et al.’s nomogram for the whole cohort was 86.5%and 86.9%,respectively.After a 52-month of median follow-up,a significant difference was found in IDFS between of the IHC3 pLR and the pHR groups(P=0.001)but not in the IDFS between the low-and high-recurrence risk groups according to the Surexam®21-gene RS and the TAILORx clinical trial criteria(P=0.556)or 21-gene binary RS group(P=0.511).Conclusions:The proposed IHC3 model could reliably predict low and high recurrence risks in most HR-positive EBC patients.This easy-to-use predictive model may be a reliable replacement for the 21-gene genomic assay in patients with EBC who have no access to or cannot afford the 21-gene genomic assay.

Prognostic value of the 21-gene recurrence score in ER-positive,HER2-negative,node-positive breast cancer was similar in nodenegative diseases:a single-center study of 800 patients

Multi-gene assays have emerged as crucial tools for risk stratification in early-stage breast cancer.This study aimed to evaluate the prognostic significance of the 21-gene recurrence score(RS)in Chinese patients with pN0-l,estrogen receptor-positive(ER+),human epidermal growth factor receptor 2-negative(HER2)breast cancer.Among 800 patients recruited between 2009 and 2016,the median RS was 24(0-69),with 27.4%,46.8%,and 25.9%patients classified into low-,intermediate-,and high-risk groups.Cox regression analysis demonstrated that the high-risk category was associated with significantly higher odds of invasive disease-free survival(IDFS)and distant disease-free survival(DDFS)events compared with the low-risk category(IDFS:HR=2.450,95%Cl 1.017-5.902,P=0.046;DDFS:HR=2.829,95%Cl 1.013-7.901,P=0.047).No significant association between RS category and overall survival(OS)was found(intermediate vs.low:HR=1.244,95%Cl 0.292-5.297,P=0.768;high vs.low:HR=2.933,95%Cl 0.759-11.327,P=0.119).RS,as a continuous variable,was a highly significant predictor for IDFS(HR=1.028,95%Cl 1.010-1.047,P=0.002),DDFS(HR=1.030,95%Cl 1.010-1.051,P=0.003),and OS(HR=1.034,95%Cl 1.007-1.063,P=0.014).Our findings suggested that RS may predict IDFS in Chinese patients with ER+/HER2 breast cancer with NO or N1 disease.