Gene profiling in soft tissue sarcoma: predictive value of EGFR in sarcoma tumour progression and survival

Abstract

Despite improvements in the clinical management of soft tissue sarcomas (STS), 50%of patients will die of metastatic disease that is largely unresponsive to conventionalchemotherapeutic agents.The aims of this study were to identify genes and pathways that are dysregulated inprogressive and metastatic STS. In addition to this, cell lines from fresh tumours wereinitiated and established, thus increasing the repository of cell lines available forfunctional studies. Recent advances in the understanding of the molecular biology ofSTS have thus far not resulted in the use of molecular markers for clinicalprognostication. Identifying novel genes and pathways will lead to moleculardiagnostic methods to better stratify prognostic groups and could identify cellulartargets for more efficacious treatments.Gene expression profiling of sarcoma cell lines of increasing metastatic potentialrevealed over-expression of genes involved in the epidermal growth factor (EGF) andtransforming growth factor beta (TGFb) pathways. Factors involved in invasion andmetastasis such as integrins and MMPs were over-expressed in the cell lines with highermetastatic potential. The developmental Notch pathway and cell cycle regulators werealso dysregulated. NDRG1 was significantly over-expressed in the high grade sarcomacell line, a novel finding in sarcomas. The expression of EGFR, NDRG1 and othergenes from the above pathways was validated using quantitative RT-PCR in real time(qRT-PCR).A tissue microarray (TMA) comprising STS of varying tumour grades was constructedfor high throughput assessment of target proteins. EGFR, its activated form and itssignal transducers were investigated using immunohistochemistry (IHC). ActivatedEGFR (HR 2.228, p < 0.001) and phosphorylated Akt (HR 2.032, p = 0.003) were foundto be independent predictors of overall survival and both correlated with tumour grade.Of the several STS cultures initiated and maintained, two of these cell lines were fullycharacterised in terms of cytogenetics, telomerase and alternate lengthening of5telomeres (ALT) status, KIT and TP53 mutation and the expression of certainbiomarkers using both qRT-PCR and IHC.In summary, transcript profiling identified several potential biomarkers of tumourprogression and metastasis in STS. Crucially, activated EGFR and pAkt were found in acohort of STS samples to correlate with clinical outcome, identifying them as potentialdiagnostic and therapeutic targets in the treatment of STS. Activated EGFR can be usedas a diagnostic marker for patient selection, as well as for target effect monitoring.Furthermore, the cell lines established in this project will serve as valuable tools infuture preclinical studies