A proteomic and genomic approach to the determination of targets associated with diagnosis and development of metastasis in pediatric osteosarcoma

Resumen

Osteosarcoma is the most frequent bone tumour in childhood and adolescence. Two main factors limit survival of patients: chemo-resistance and the development of metastasis (usually to lung or bone), which make conventional treatment ineffective. Therefore, a principal objective of current research in this field is to establish ways to determine, early in the disease, whether a patient will respond or not to treatment with conventional cytostatics and whether a patient will develop metastases or not. The general objective of this study is to establish the comparative genomic and proteomic profiles of tumor and normal osteoblasts isolated from the same patient and to use these profiles to elucidate molecular hallmarks of chemo-resistant and metastatic osteosarcomas. The study focuses on high grade osteosarcomas in pediatric patients and uses a novel approach: isolation and characterization of cell populations from untreated primary tumours and their paired normal bone tissues. By means of 2D-DIGE, proteomic profiles were obtained and compared with a view to discovering common molecular pathways and investigating the genetic mechanisms of childhood osteosarcoma. Some of the proteins identified were validated through a tissue microarray (TMA) that had been developed in our laboratory. The TMA contained samples of normal, primary, metastatic and local recurrence tissues from osteosarcoma patients treated in the hospital to which our laboratory is attached. In addition, the U133A chip ® was used to search for genes involved in altered molecular pathways relevant to the carcinogenesis and metastasis of osteosarcoma. Genes thus identified were subsequently validated by real-time PCR. Conclusion: comparative genomic and proteomic studies followed by validation in high-performance systems is a useful approach to the detection of targets involved in pathologies, such as childhood osteosarcoma, that are infrequent, difficult to access, and associated with small samples.