engleski

EFFECTS OF dnMAML PEPTIDE ON GLIOBLASTOMA CELLS

Introduction Today cancer is one of the most researched diseases in the world. Advances in therapy have been great but remain insufficient. GBMs account for about 70% of the newly diagnosed malignant brain tumors. Current standard treatment includes “the cancer triad”: maximal surgical resection (if possible), radiation therapy with concomitant chemotherapy. In spite of all progress and changes in therapy recurrence rate for GBMs is extremely high with about 90% of the tumors recurring in the original site. Notch signaling pathway is one of the few highly conserved pathways involved in development and homeostasis in multicellular organisms. Consisting of transmembrane receptor and ligands on the neighboring cells it provides means for cell-to-cell communication. Signal transduction occurs in several steps finally releasing NICD that translocates to the nucleus and activates gene expression. Additionally, participants in the pathway can interact with other cellular control mechanisms making Notch effects very variable and context dependent. In cancer Notch has a dual role. In majority of cases activation of Notch is an oncogenic trigger. Some type of Notch activation was found in 80% of GBMs underlining importance of Notch in formation and recurrence of GBMs. Although Notch aberrations are mostly oncogenic in several cancer types it notch activation acts as a tumor suppressor. This puts a question on how to treat Notch without unwanted side effects. Materials and Methods Protein based Notch inhibitor dnMAML is actually a portion of MAML1 co-activator protein and is shown to be efficient in blocking Notch function in T-ALL. Unfortunately as with all peptides problem is efficient delivery into the cells and in the case of GBMs over the BBB. ELP is a thermally reactive polypeptide that can be actively and passively targeted to the tumor site by localized application of heat and using the EPR effect. Entry into the cells is enhanced by the addition of a CPP in this case SynB1, which can also carry the cargo over the BBB. Results SynB1-ELP-dnMAML inhibits growth of the two tested GBM cell lines, D54 and U251 by inducing apoptosis and cell cycle arrest. Samples show 20% to 80% cells dead proportional to the concentration of protein used and presence of hyperthermia. As presumed hyperthermia increases overall entry of the polypeptide into the cells and the resulting effect. Both canonical and non-canonical signaling is inhibited as can be seen from expression levels of Hes-1 and Hey-L and protein levels of pAkt and MAPK. p53 protein that interacts directly with the N-terminal domain of MAML1 present in dnMAML is also affected. Levels of wt p53 in D54 cells are maintained while levels of mutant p53 in U251 cells are significantly reduced. Conclusion Results confirm that dnMAML is effective Notch inhibitor and that combining it with ELP gives effective means of actively targeting and increasing inhibition only in a small area thus avoiding the dangers of systemic Notch inhibition. In GBMs this is good alternative approach that can overcome difficulties posed by poor delivery across the blood-brain barrier.

Glioblastoma; Notch; dnMaml; targeted therapy; protein inhibitor

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