Effects of IGF-1R Nuclear Localization in Glioblastoma Cells

CLEMENT, FLORENCIA; MARIANA GUTIERREZ; MARTÍN, AYELEN; IGNACIO BERGADÁ; VENARA MARCELA; PENNISI PATRICIA A

Atenas

Congreso; 57th Annual Meeting of the European Society for Paediatric Endocrinology (ESPE); 2018

ESPE

Background: CNS tumors are the most frequent solid tumors in children. In paediatric gliomas, IGF-1R nuclear localization was significantly associated with both high grade tumours and increased risk of death, suggesting that nuclear IGF-1R localization may contribute to an aggressive phenotype. Aim: To characterize the impact of IGF-1R overexpression and nuclear localization in glioma cells.Methods: U87Mg glioblastoma cells were transfected with pEGFP-IGF1R to generate stably transfected cells. Lys1025-1100-1120 were targeted by mutagenesis to avoid IGF-1R nuclear translocation. Proliferation assays were carried out during 5 days, and wounding assay for 18h with/without IGF-1. Protein extracts were obtained from whole lysates or after subcellular fractionation and processed by western blot (WB). Gene expression was quantified by rqPCR. IGF-1R/IR inhibitor OSI906 was used at 0.5uM. Male nude mice were injected for in vivo experiments (1,5e6cells/flank/mice).Results: stably transfected cells with 5(B4U87) or 50(B2U87) times basal expression of IGF-1R showed higher tumor incidence and shorter latency time compared to parental cells when injected in nude mice. In vitro, an increase in pAKT, pERK and pMAPK38 was observed after IGF-1 stimulation.Nuclear localization of IGF-1R was verified by IF and WB (8h IGF-1 stimulation). These effects were blocked by 1h preincubation with OSI906. Although CyclinD1 levels were slightly increased in B2U87 and B4U87 cells in response to 24h IGF-1 stimulation, proliferation and apoptosis were not different from parental cells. Wounding assay showed an increased motility in both B2U87 and B4U87 compared to parental cells. Moreover, GLUT-1 expression showed a two-fold increase after 24h IGF-1 stimulation that was abrogated by OSI906. Cells expressing GFP-IGF-1R1025x-1100x-1120x showed no IGF-1R nuclear localization and a lower increase in GLUT-1 expression upon IGF-1 stimulation.Conclusion: IGF-1R overexpression and nuclear localization may contribute to an aggressive phenotype of glioblastoma by increasing motility and metabolism of tumor cells rather than increasing its proliferation.