RBBP4-p300 axis modulates expression of genes essential for cell survival and is a potential target for therapy in glioblastoma

Neuro Oncol. 2022 Aug 1;24(8):1261-1272. doi: 10.1093/neuonc/noac051.

Abstract

Background: RBBP4 activates transcription by histone acetylation, but the partner histone acetyltransferases are unknown. Thus, we investigated the hypothesis that RBBP4 interacts with p300 in a complex in glioblastoma (GBM).

Methods: shRNA silencing of RBBP4 or p300 and RNAseq was used to identify genes co-regulated by RBBP4 and p300 in GBM43 patient-derived xenograft (PDX). RBBP4/p300 complex was demonstrated using proximity ligation assay (PLA) and ChIPseq delineated histone H3 acetylation and RBBP4/p300 complex binding in promoters/enhancers. Temozolomide (TMZ)-induced DNA double strand breaks (DSBs) were evaluated by γ-H2AX and proliferation by CyQuant and live cell monitoring assays. In vivo efficacy was based on survival of mice with orthotopic tumors.

Results: shRBBP4 and shp300 downregulated 4768 genes among which 1485 (31%) were commonly downregulated by both shRNAs, while upregulated genes were 2484, including 863 (35%) common genes. The pro-survival genes were the top-ranked among the downregulated genes, including C-MYC. RBBP4/p300 complex was demonstrated in the nucleus, and shRBBP4 or shp300 significantly sensitized GBM cells to TMZ compared to the control shNT in vitro (P < .05). Moreover, TMZ significantly prolonged the survival of mice bearing GBM22-shRBBP4 orthotopic tumors compared with control shNT tumors (median shNT survival 52 days vs. median shRBBP4 319 days; P = .001). CREB-binding protein (CBP)/p300 inhibitor CPI-1612 suppressed H3K27Ac and RBBP4/p300 complex target proteins, including C-MYC, and synergistically sensitized TMZ in vitro. Pharmacodynamic evaluation confirmed brain penetration by CPI-1612 supporting further investigation to evaluate efficacy to sensitize TMZ.

Conclusions: RBBP4/p300 complex is present in GBM cells and is a potential therapeutic target.

Keywords: RBB4/p300 complex; glioblastoma; histone acetylation; temozolomide.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, N.I.H., Extramural

MeSH terms

  • Acetylation
  • Animals
  • Brain Neoplasms* / drug therapy
  • Brain Neoplasms* / genetics
  • Brain Neoplasms* / pathology
  • Cell Line, Tumor
  • Cell Survival
  • Drug Resistance, Neoplasm
  • E1A-Associated p300 Protein* / genetics
  • E1A-Associated p300 Protein* / metabolism
  • Glioblastoma* / drug therapy
  • Glioblastoma* / genetics
  • Glioblastoma* / metabolism
  • Humans
  • Mice
  • Promoter Regions, Genetic
  • Retinoblastoma-Binding Protein 4* / genetics
  • Retinoblastoma-Binding Protein 4* / metabolism
  • Temozolomide / pharmacology
  • Temozolomide / therapeutic use
  • Xenograft Model Antitumor Assays

Substances

  • RBBP4 protein, human
  • Retinoblastoma-Binding Protein 4
  • E1A-Associated p300 Protein
  • EP300 protein, human
  • Temozolomide