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Glioblastoma (GBM; WHO grade IV) is an aggressive disease with a low 5-year survival rate. Resistance to therapy is common which renders current modalities largely ineffective. We identified TRIB1, a Ser/Thr pseudokinase through a patient derived reverse translational approach as a potential driver of resistance mechanisms in GBM. TRIB1 functions as a scaffold to initiate Ubiquitin Proteasome System-mediated degradation of its substrates and has also been reported to activate the MAPK and Akt pathways in various cell types. Several studies show that TRIB1 contributes to chemotherapy resistance in various cancers. Therefore, we evaluated the role of TRIB1 in radiation therapy resistance routinely encountered in GBM.

Cells were irradiated using the X-Rad cabinet system. Site-directed mutagenesis was performed to substitute specific amino acid (W>A). Patient derived primary cell lines stably overexpressing wild type and mutant (W337A) TRIB1 transgenes were generated by antibiotic selection. Cells were injected intracranially in nude mice to develop tumors. Changes in tumor volume and overall survival (OS) were determined. Co-immunoprecipitation was performed to evaluate protein-protein interactions. Protein levels were detected by western blot.

TRIB1 mRNA and protein levels increased after radiation treatment in GBM cell lines. We observed that TRIB1 overexpression in patient derived primary GBM (PDX) cell lines increased their viability after RT treatment. However, upon overexpression of mutant (W337A) TRIB1, this effect was reversed. We further observed a decrease in apoptosis of TRIB1 overexpressing PDX cells after RT treatment in vitro. Mice bearing TRIB1 transgene overexpressing tumors had the highest tumor volume and lowest OS whereas mice bearing mutant TRIB1 tumors had the highest OS and lower tumor volume at the end of experiment. Both p53 and HDAC1 formed complex(s) with TRIB1 in primary GBM cells lines, as revealed by co-immunoprecipitation. Further, western blot revealed that overexpression of TRIB1 decreased p21 and MDM2 levels in these cells.

Increased expression of TRIB1 in GBM cells contributes to radiation resistance by increasing the cell viability in vitro and promoting tumor growth in vivo. Mechanistic studies revealed that TRIB1 forms a complex with p53 and HDAC1 in primary GBM cell lines consequently inhibiting p53 function by facilitating its degradation through the E3-ligase COP1 or deacetylation through HDAC1, thereby decreasing radiation induced cell death. Taken together TRIB1 can be considered a potential therapeutic target in GBM.