High-throughput tissue microarray analysis to evaluate genes uncovered by cDNA microarray screening in renal cell carcinoma

Abstract

Many genes and signaling pathways are involved in renal cell carcinoma (RCC) development. However, genetic tumor markers have not gained use in RCC diagnostics and prognosis prediction. Identification and evaluation of new molecular parameters are of utmost importance in cancer research and cancer treatment. Here we present a novel approach to rapidly identify clinically relevant molecular changes in cancer. To identify genes with relevance to RCC, a cDNA array analysis was first performed on 5184 cDNA clones on a filter to screen for genes with differential expression between the renal cancer cell line CRL-1933 and normal kidney tissue. There were 89 differentially expressed genes in the cancer cell line, one of them coding for vimentin, a cytoplasmic intermediate filament. In a second step, a renal cancer tissue microarray containing 532 RCC specimen was used to determine vimentin expression by immunohistochemistry. Vimentin expression was seen frequently in clear cell (51%) and papillary RCC (61%), but rarely in chromophobe RCC (4%) and oncocytomas (12%). Furthermore, vimentin expression was significantly associated with poor patient prognosis (P < 0.007) independent of grade and stage. These results obtained from minute arrayed tumor samples match well with previous findings on vimentin expression in renal tumors. It is concluded that the combination of tumor arrays and cDNA arrays is a powerful approach to rapidly identify and further evaluate genes that play a role in tumor biology.

Figure 1.

cDNA array: examples of overexpressed genes in the renal cancer cell line (CRL-1933) are indicated by arrows. The dot for the vimentin gene is circled.

Figure 2.

A: Overview of the renal tumor array with 532 tissue cylinders (H&E; original magnification, ×4.2). B: Renal tumor array. Morphology of 6 different renal tumors (H&E; original magnification, ×42) C: Two different renal tumor subtypes, papillary RCC (top) and clear cell RCC (bottom). (H&E; original magnification, ×140). D: Immunohistochemistry: clear cell RCC with cytoplasmic vimentin expression (top) and chromophobe RCC without vimentin expression (bottom) (ABC-Elite method, anti-vimentin; original magnification, ×88).

Figure 3.

Overall survival analysis for clear cell RCC with and without vimentin expression (log-rank test).