Gene Expression Detection Assay for Cancer Clinical Use

Narrandes, Shavira; Xu, Wayne

Through studying candidate genes and the RNA, DNA, and/or protein molecular alterations in each sample, TMAs can be used in the research of diseases, such as cancer, and for drug target discovery, to determine the therapeutic importance of these genes in a patient, as well as their diagnostic and prognostic significance. In cancers, this allows for the examination of tumor progression and the identification and validation of prognostic factors or genes that can potentially be translated into the clinical setting for therapeutic treatments. For example, TMAs were used in an original experiment to study the amplifications of six genes, as well as the expression of p53 and estrogen receptor (ER), in breast cancers, allowing for the further stratification of the cancers into subgroups. 67 To examine how the molecular changes in bladder cancer patients affect their clinical outcomes, a tissue microarray was constructed using 2317 samples from 1842 patients to study the outcome of the amplification of CCNE and its protein expression. 74 Four independent TMAs were used to analyze the prognostic markers (ER, PR, and p53) in impacting molecular changes and clinical endpoints in 553 breast carcinomas. 0.6 mm samples were taken from one central and three peripheral regions from FFPE samples. Multiple punches per tumor were taken that demonstrated the tumors could be distinguished into three subgroups (positive, negative, and heterogeneous). 12 TMAs constructed using three antibodies on four arrays demonstrated significant associations with tumor-specific survival than large section analyses. A single sample per tumor was sufficient to identify associations between molecular alterations and clinical outcome. Further validation of the established molecular markers is required using larger tissue samples from clinical trials. 75 As many previous studies assessing ER expression using immunohistochemistry have not been reproduced, one study used TMAs to examine the variability in ER expression reporting for breast cancers. 5 independent laboratories constructed TMAs by coring an invasive breast cancer donor block from 29 patients twice. A moderate to high interlaboratory agreement was determined, demonstrating that TMAs are efficient for the identification of ER variability reporting and have the potential to be applied to similar projects. 76 TMAs of 544 clinical samples from varying stages of prostate cancer were used to study the expression of candidate genes discovered by cDNA microarrays of the CWR22 xenograft model. Using 60 prostatic intraepithelial neoplasia, 264 primary tumors, 41 distal metastases, and 134 hormone refractory specimens, with 45 samples of benign prostatic hyperplasia acting as the control, they demonstrated the use of S100P, CRYM, and LMO4 in affecting androgen-independent growth and therapy failure in prostate cancers. Following mRNA in situ hybridization and immunohistochemical analyses of the TMAs, strong correlation was found between the cDNA microarrays and mRNA. Over-expression in hormone-refractory CWR22R xenografts of S100P was seen while CRYM and LMO4 were down-regulated. 77 Another study used TMAs from 1023 samples of prostate tissues to examine the significance of the Polycomb group protein EZH2 in the progression of prostate cancer. Included in the cohort were 400 samples from 23 individuals who passed away from hormone refractory metastatic prostate cancer. Highly expressed EZH2 is mainly seen in the nucleus and i