P21 Activated Kinase Pak1 as a Therapeutic Target In Astrocytomas

Abstract

Immunohistochemistry and western blotting were performed to identify the Pak1 and p Pak1 protein levels in fresh Astrocytoma samples and normal brain samples Kaplan-Meir analysis was plotted by using the survival data follow up of the patients Protein phosphorylation status of Pak1 protein was observed by performing invitro kinase assay to check the activated Pak1 level by using PAKtide which is a specific substrate for Pak1 Later knock out clones were generated by using Pak1 CRISPR cas9 plasmid on two cells lines BMG1 and U373 And functional assays such as proliferation assay and Clonogenic assay have been done to evaluate the effect of Pak1 protein on cellular functions We also studied the effect of Pak1 KO on cell signalling molecules such as MAPK and cyclin D and identified the role of Pak1 in proliferation of astrocytoma cell lines under and#947;irradiation This study provides molecular evidence signifying the function of PAK1 and its phosphorylation status in the progression of astrocytomas to more aggressive phenotypes of astrocytoma In the current study to identify Pak1 as a potent clinical molecular marker in astrocytoma we compared the expression levels of Pak1 in various grades of astrocytomas When tumor samples were immunostained for Pak1 we observed the lowest mean Q score in grade I that increased with subsequent progression to Grade IV Kaplan Meier survival analysis of data from the study cohort showed that gradeI cases manifested a better prognosis the rate of fall of survival for grade II tumors was highest and Grade III and IV tumors exhibited the lowest overall survival In summary this report concludes that higher Pak1 expression speculates a lower survival rate across various grades in astrocytoma patients newline Further the in vitro kinase assays performed utilizing immunoprecipitated Pak1 phosphorylated the Pak1 substrate PAKtide more in astrocytoma tissues compared to normal brain tissues obtained from autopsy samples that explained the activation status of Pak1 in astrocytoma cancers Furthermore our s