Microarray identification of diagnostic biomarkers in leukocytes isolated from peripheral blood of mice injected with B16-BL6 melanoma cells

We have evaluated the potential of using leukocyte/immune cell gene expression to identify diagnostic biomarkers that can differentiate between mice injected with B16-BL6 cancer cells and normal mice. Eight mice were injected into the flank with B16-BL6 cells, an aggressive murine melanoma. Tumour growth progressed for approximately 20 days. Blood was collected from these mice as well as litter mates not transplanted with the B16-BL6 cells and cDNA microarray analysis was performed on the samples. The main objectives of this project were to: (1) confirm that microarray analysis could generate reproducible gene expression data among different mice; (2) show that RNA amplification does not skew relative fluorescence of the genes; and, (3) generate a repertoire of differentially expressed host genes between control and tumour bearing mice. Oneway analysis of variance (cancer vs non-cancer mice) was completed to identify genes that were significantly different between tumour-bearing mice and their non-tumourgenic counterparts. The test identified 376 genes out of a total of 15600 genes that were significantly different at the p≤ 0.05 level. Gene expression measured by cDNA microarray analysis weakly correlated with QRT-PCR analysis, performed on a different set of mice, emphasizing the individual variability in their expression levels. However, cDNA microarray analysis was able to show differences in the genes expressed by the leukocytes of cancer bearing compared to normal mice. Many of the differentially expressed genes were shown to be involved in pathways that control cell adhesion or increase sensitivity to cell death. Pathway analysis of the differentially expressed genes identified several genes involved in the Crk-dependent signaling pathway. Further studies must be performed to develop analysis methods which will accommodate for the large amount of variability in the data.