1.Distinguishing Non-invasive And Invasive Bladder Urothelial Carcinomas By Molecular Markers Selected From Genomic DNA 2.Overexpression Of HSPC300 Is Associated With The Potential Of Metastasis In Human Non-small Cell Lung Cancer

Chapter?:Distinguishing non-invasive and invasive bladder urothelial carcinomas by molecular markers selected from genomic DNAThe differences between two subtypes of bladder urothelial carcinoma,non-invasive tumor and invasive tumor,are closely associated with the clinical performance,the surgical style and the prognoses.However there is a lack of reliable approaches to preoperative differentiation for the two subtypes of bladder cancer.In this study, array comparative genome hybridization(array CGH) analysis was performed first,to identify the DNA copy number variation(CNV) profiles that specifically related to the non-invasive or invasive bladder cancer,respectively.From these,a set of gain or lose genes/regions could be identified to distinguish the two subtypes of tumor. Based on subsequent validation by quantitative PCR and bioinformatical analysis,a panel of molecular markers was selected for classifying the two subtypes of bladder cancer.The tumor tissue samples derived from 35 patients of bladder cancer(18 non-invasive and 17 invasive) were screened by array CGH on the Agilent Human Genome CGH 44B array.With analyses by CGH Analytics 3.4(Agilent) and R project (Bioconductor),the genome-wide CNV profiles of bladder urothelial carcinoma were obtained.In euchromosomes,the copy number alterations were mainly occurred in 1q,2q,3p,4q,5p,5q,7p,8p,8q,9p,9q,10q,11p,11q,13q,15q,16p,16q,17p,17q and 18q.The highest average frequency of the alterations was observed on Chromosome 9.Fourteen genes/regions with high-level amplification and 11 genes/regions with homozygous deletion,whose average frequency were no less than 20%,were identified.In the invasive tumors,the frequency of aberrancy was significantly higher than that detected in the non-invasive ones,especially in Chromosome 5,8,10,17 and 18.Bioinformatical analysis was performed with the array CGH data of the 35 cases,and 8 genes that could classify the two subtypes of bladder cancer correctly(97.1%) were screened out.These 8 genes were then validated by real time polymerase chain reaction(RT-PCR) in this group of tumor tissues.There was a significant correlation of the results between the array CGH and RT-PCR analyses(p=0.0003).In conclusion,data obtained from a reliable array CGH analysis indicated that there were differences in the genome-wide CNV profiles between the non-invasive and invasive bladder cancers.It was promised to build up a molecular model to distinguish the bladder urothelial carcinomas with various clinical phenotypes,by further investigation with an enlarged sample size.Chapter?Overexpression of HSPC300 is associated with the potential of metastasis in human non-small cell lung cancerOur previous work suggests that HSPC300 is one of most overexpressed genes in a differentially expressed cDNA library derived from human non-small cell lung cancer (NSCLC).In present study,following the expressive status of HSPC300 protein in the tumor tissues of NSCLC was examined,the correlation between expression of the HSPC300 and the tumor metastasis was analyzed.The subsequent in vitro investigation focused on the molecular mechanism that HSPC300 facilitates metastasis of NSCLC.Tissue microarrays derived from NSCLCs were stained immunohistochemically. HSPC300 protein was detected in 82.9%(92 of 111) of the primary tumors and 73.7% (28 of 38) of lymph node metastases,but only in 30.8%(4 of 13) of normal lung or bronchial epithelium tissue samples(p?0.0001).HSPC300 expression was associated with the tumor staging and lymph node invasion(p=0.0035 and p=0.0001, respectively).In vitro,silencing HSPC300 by siRNA could reduce the protrusions and pseudopods formed in NSCLC cells PG that has high potential of metastasis. Moreover,downregulating HSPC300 expression inhibited migration and invasion of the PG cells by Transwell assay.With co-immunoprecipitation and immunofluorescence analyses,it was observed that there was protein-protein interaction between HSPC300 and WAVE1 or WAVE2.HSPC300 affected the proteasome-dependent stability of WAVE1 and WAVE2.However,only knock-down WAVE2,but not WAVE1,resulted in morphological change that is similar to that of silencing HSPC300 in the PG cells.EGF activates F-actin to polymerization.Knock-down HSPC300 or WAVE2 abolished the activation of the polymerization.The current findings suggest that HSPC300 may facilitate metastasis of NSCLC mainly through stabilizing WAVE2,a protein associated with cell migration.