Human 70mer Microarrays and Murine cDNA Microarrays

The methodological developments described above were the prerequisite for the successful generation and application of a new type of microarray, which carries 27,000 human gene specific chemically synthesized 70mer oligonucleotides, each one in replicate (i. e., 54,000 spots in total per glass slide). These 70mer microarrays were successfully used within several studies on epithelial, haematological and cranial tumor series, but also for studies on cell culture models, e.g., for specific questions of apoptosis and organ repulsion:

• Basalioma: an expression profiling study of a series of basalioma (in comparison with healthy skin) identified in particular genes involved cell cycle control, meiosis and DNA replication / chromosomal cycling which were up-regulated in the tumors. The comparison of these basalioma data with the results of an expression profiling study on the multi-stage tumor model of murine skin (see below) identified important common candidate genes and pathways which might be involved in human epidermal carcinogenesis. These candidate genes are currently analyzed in more detail.
• Studies on the molecular effects of a new type of immune suppressor and inhibitor of proliferation were done in a cell culture model system. Relevant candidate genes for the drug effects were identified.
• Within a clinical study on primary breast cancer we try to identify gene expression profiles which predict pathologic complete response to preoperative chemotherapy with Gemcitabine, Epirubicin and Docetaxel. Recently, our identified RNA expression signature of relevant candidate genes is validated in a further study.

In addition, two comprehensive types of cDNA-microarrays, which comprise 20,000 (LION-microarray) and 23,000 (NIA-microarray) different gene specific cDNA fragments of the mouse, were generated for the analysis of the murine transcriptome (see Figure 1). We apply these microarrays in studies on murine tumor models. A major effort was the study of tumor progression in murine skin (see Figure 2): Chemically induced mouse carcinogenesis represents the most extensively utilized animal model to unravel the multistage nature of tumour development and to design novel therapeutic concepts of human epithelial neoplasia. We combined this tumour model with comprehensive gene expression analysis and could identify a large set of novel tumour-associated genes that have not been associated with epithelial skin cancer development yet. According to current knowledge about the multistage nature of skin carcinogenesis, functional annotation of our expression data revealed specific signatures for the distinct tumour stages. Moreover, self-organizing map clustering was performed to expose different kinetics of gene expression and co-regulation during skin cancer initiation and progression. Expression data of selected genes were confirmed by reverse transcription PCR, quantitative real-time PCR and in situ hybridisation analysis on mouse tumour sections. The expression of a couple of genes identified in our screen correlated with tumour formation of keratinocyte cell lines in vivo. Finally, we detected high mRNA levels of corresponding genes in human skin tumour specimens demonstrating that tumour-associated genes identified in the murine model of skin carcinogenesis might be of great relevance for the understanding of human epithelial malignancies as well.