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Glioblastoma patient-derived cell-based phenotypic drug screening and identification of possible action mechanisms through proteomic analysis.

ABSTRACT: Because glioblastoma (GBM) exhibits high heterogeneity, it is desirable to use patient-derived cells from the first stage of screening for GBM drug discovery. Here, we describe a protocol to culture patient-derived GBM cells on the extracellular matrix-coated plates to allow high-throughput screening. Further, we detail approaches to identify the mechanism of action (MOA) of the selected effective drug through proteomics. This protocol will be useful for researchers interested in drug screening and the MOA of drugs. For complete details on the use and execution of this protocol, please refer to Nam et al. (2021).

SUBMITTER: Kim YE

PROVIDER: S-EPMC8476855 | biostudies-literature | 2021 Dec

REPOSITORIES: biostudies-literature

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Glioblastoma patient-derived cell-based phenotypic drug screening and identification of possible action mechanisms through proteomic analysis.

Kim Young Eun YE Kim Hyun Young HY Jung Daeyoung D Kang Dukjin D Nam Do-Hyun DH Nam Hye Jin HJ Cho Heeyeong H

STAR protocols 20210924 4

Because glioblastoma (GBM) exhibits high heterogeneity, it is desirable to use patient-derived cells from the first stage of screening for GBM drug discovery. Here, we describe a protocol to culture patient-derived GBM cells on the extracellular matrix-coated plates to allow high-throughput screening. Further, we detail approaches to identify the mechanism of action (MOA) of the selected effective drug through proteomics. This protocol will be useful for researchers interested in drug screening ...[more]

PMID: 34611628

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Project description:The development of drugs to inhibit glioblastoma (GBM) growth requires reliable pre-clinical models. To date, proteomic level validation of widely used patient-derived glioblastoma xenografts (PDGX) has not been performed. In the present study, we characterized 20 PDGX models according to subtype classification based on The Cancer Genome Atlas criteria, TP53, PTEN, IDH 1/2, and TERT promoter genetic analysis, EGFR amplification status, and examined their proteomic profiles against those of their parent tumors. The 20 PDGXs belonged to three of four The Cancer Genome Atlas subtypes: eight classical, eight mesenchymal, and four proneural; none neural. Amplification of EGFR gene was observed in 9 of 20 xenografts, and of these, 3 harbored the EGFRvIII mutation. We then performed proteomic profiling of PDGX, analyzing expression/activity of several proteins including EGFR. Levels of EGFR phosphorylated at Y1068 vary considerably between PDGX samples, and this pattern was also seen in primary GBM. Partitioning of 20 PDGX into high (n = 5) and low (n = 15) groups identified a panel of proteins associated with high EGFR activity. Thus, PDGX with high EGFR activity represent an excellent pre-clinical model to develop therapies for a subset of GBM patients whose tumors are characterized by high EGFR activity. Further, the proteins found to be associated with high EGFR activity can be monitored to assess the effectiveness of targeting EGFR. The development of drugs to inhibit glioblastoma (GBM) growth requires reliable pre-clinical models. We validated proteomic profiles using patient-derived glioblastoma xenografts (PDGX), characterizing 20 PDGX models according to subtype classification based on The Cancer Genome Atlas (TCGA) criteria, TP53, PTEN, IDH 1/2, and TERT promoter genetic analysis, EGFR amplification status, and examined their proteomic profiles against those of their parent tumors. Proteins found to be associated with high EGFR activity represent potential biomarkers for GBM monitoring.

Dataset Information

Glioblastoma patient-derived cell-based phenotypic drug screening and identification of possible action mechanisms through proteomic analysis.

Publications

Glioblastoma patient-derived cell-based phenotypic drug screening and identification of possible action mechanisms through proteomic analysis.

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