Insulin growth factor-2 binding protein 3 (IGF2BP3) is a glioblastoma-specific marker that activates phosphatidylinositol 3-kinase/mitogen-activated protein kinase (PI3K/MAPK) pathways by modulating IGF-2.

Glioblastoma is the most common and malignant form of primary astrocytoma. Upon investigation of the insulin-like growth factor (IGF) pathway, we found the IGF2BP3/IMP3 transcript and protein to be up-regulated in GBMs but not in lower grade astrocytomas (p < 0.0001). IMP3 is an RNA binding protein known to bind to the 5'-untranslated region of IGF-2 mRNA, thereby activating its translation. Overexpression- and knockdown-based studies establish a role for IMP3 in promoting proliferation, anchorage-independent growth, invasion, and chemoresistance. IMP3 overexpressing B16F10 cells also showed increased tumor growth, angiogenesis, and metastasis, resulting in poor survival in a mouse model. Additionally, the infiltrating front, perivascular, and subpial regions in a majority of the GBMs stained positive for IMP3. Furthermore, two different murine glioma models were used to substantiate the above findings. In agreement with the translation activation functions of IMP3, we also found increased IGF-2 protein in the GBM tumor samples without a corresponding increase in its transcript levels. Also, in vitro IMP3 overexpression/knockdown modulated the IGF-2 protein levels without altering its transcript levels. Additionally, IGF-2 neutralization and supplementation studies established that the proproliferative effects of IMP3 were indeed mediated through IGF-2. Concordantly, PI3K and MAPK, the downstream effectors of IGF-2, are activated by IMP3 and are found to be essential for IMP3-induced cell proliferation. Thus, we have identified IMP3 as a GBM-specific proproliferative and proinvasive marker acting through IGF-2 resulting in the activation of oncogenic PI3K and MAPK pathways.

STAT-1 expression is regulated by IGFBP-3 in malignant glioma cells and is a strong predictor of poor survival in patients with glioblastoma.

OBJECT: Insulin-like growth factor binding proteins (IGFBPs) have been implicated in the pathogenesis of glioma. In a previous study the authors demonstrated that IGFBP-3 is a novel glioblastoma biomarker associated with poor survival. Since signal transducer and activator of transcription 1 (STAT-1) has been shown to be regulated by IGFBP-3 during chondrogenesis and is a prosurvival and radioresistant molecule in different tumors, the aim in the present study was to explore the functional significance of IGFBP-3 in malignant glioma cells, to determine if STAT-1 is indeed regulated by IGFBP-3, and to study the potential of STAT-1 as a biomarker in glioblastoma. METHODS: The functional significance of IGFBP-3 was investigated using the short hairpin (sh)RNA gene knockdown approach on U251MG cells. STAT-1 regulation by IGFBP-3 was tested on U251MG and U87MG cells by shRNA gene knockdown and exogenous treatment with recombinant IGFBP-3 protein. Subsequently, the expression of STAT-1 was analyzed with real-time reverse transcription-polymerase chain reaction (RT-PCR) and immunohistochemistry (IHC) in glioblastoma and control brain tissues. Survival analyses were done on a uniformly treated prospective cohort of adults with newly diagnosed glioblastoma (136 patients) using Kaplan-Meier and Cox regression models. RESULTS: IGFBP-3 knockdown significantly impaired proliferation, motility, migration, and invasive capacity of U251MG cells in vitro (p < 0.005). Exogenous overexpression of IGFBP-3 in U251MG and U87MG cells demonstrated STAT-1 regulation. The mean transcript levels (by real-time RT-PCR) and the mean labeling index of STAT-1 (by IHC) were significantly higher in glioblastoma than in control brain tissues (p = 0.0239 and p < 0.001, respectively). Multivariate survival analysis revealed that STAT-1 protein expression (HR 1.015, p = 0.033, 95% CI 1.001-1.029) along with patient age (HR 1.025, p = 0.005, 95% CI 1.008-1.042) were significant predictors of shorter survival in patients with glioblastoma. CONCLUSIONS: IGFBP-3 influences tumor cell proliferation, migration, and invasion and regulates STAT-1 expression in malignant glioma cells. STAT-1 is overexpressed in human glioblastoma tissues and emerges as a novel prognostic biomarker.