Comparative amplification analysis of human glioma tissue and glioma derived fragment spheroids using reverse chromosome painting (RCP).

Tumor fragment spheroids (TFS) represent an organotypic in vitro model with preserved cytoarchitecture and matrix components of the native tumor in situ. In order to determine whether DNA amplifications within gliomas remain stable in spheroid culture, tumor fragment spheroids were established from 15 human gliomas including 14 glioblastoma and one anaplastic astrocytoma. Native tumor tissue, monolayers as well as TFS were evaluated for DNA amplification using reverse chromosome painting (RCP). A modified protocol for DNA isolation from TFS was established. Amplifications in the original tumor tissue were found on chromosomes 12q13-15, tel, 4q12-13 and 11p12-13, an amplification on 11p12-13 is reported for the first time. By RCP we could demonstrate that amplified domains on 12q13-15 and 4q12-13 in three tumors were maintained in TFS whereas the amplification on 11p12-13 could not be confirmed in TFS. In monolayer cultures, all amplifications which were detected in primary culture were lost until passage 5. The results of this first comparative study of DNA amplification in glioma by analyzing native tumor tissue and tumor fragment spheroids enables us to conclude that TFS seems to be a promising in vitro model for the study of DNA amplification under cell culture conditions.