The basic pathological research for the establishment of the diagnoses of bone tumors and soft tissue tumors using DNA microarray.

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In this study, we reevaluated about 100 cases of bone tumors and soft tissue tumors whether they had been histologically diagnosed correctly or not. We performed chromosomal analysis and molecular analysis of chromosomal translocations about some cases of diem. For example, a chimeric gene : EWS-ATF1 was identified in all of 8 cases of clear cell sarcoma by RT-PCR. On the basis of the results, we performed the basic experiment for construction of microarray to detect chimeric genes of bone tumors and soft tissue tumors. At first, we examined synovial sarcoma (SS) cases, that were divided into only two subtypes and of which breaking points were simple. We would have independently detected SYT-SSX1, SYT-SSX2 in each SS cases by multiplex RT-PCR method. Moreover, we made a microarray that had three genes: SYT, SSX1, SSX2 and hybridized cDNA fragments amplified by PCR. On the array, the cDNA fragments reacted with the gene arrangement as same as the specific chimeric gene, but didn't reacted with the gene arrangements as same as other chimeric genes. Additionally, we examined EWS-related chimeric genes about EWS-related tunors by both conventional RT-PCR method and multiplex RT-PCR method. As a result, the results by conventional RT-PCR method were as same as the results by multiplex RT-PGR method in many cases. But we couldn't identify target chimeric genes in some cases. And we also had non-specific reactions in some cases. We speculated that these results were due to number of cycle of PCR and reaction temperature. Thus we found, that it is difficult to set up the best condition of the multiplex RT-PCR. But we also found that these results were important and basic data to reach the foundation of the genetic diagnosis of the bone tumors and soft tissue tumors that had chimeric genes, using mnicroarray method.