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Small molecule NSC1892 targets the CUL4A/4B-DDB1 interactions and causes impairment of CRL4DCAF4 E3 ligases to inhibit colorectal cancer cell growth.

ABSTRACT: Cullin 4A and 4B (CUL4A and 4B) function as oncogenes in colorectal cancer (CRC) cells. Both of them conservatively associate with DNA damage-binding protein 1 (DDB1) and DDB1-CUL4-associated factor 4 (DCAF4) to form Cullin-RING E3 ligases known as CRL4DCAF4, which specifically ubiquitinate and degrade tumor suppressor ST7 (suppression of tumorigenicity 7). Knockdown either CUL4A/4B or DDB1 significantly inhibits tumor cell growth in vitro and in vivo. Thus, targeting these CRL4DCAF4 components and their interactions may be an effective strategy for the therapy of CRC. In this study, we developed an in vitro AlphaScreen assay to identify small molecules targeting the CUL4A-DDB1 interaction. We obtained a compound NSC1892, which strongly disrupted the CUL4A-DDB1 interaction (IC50 = 1.8 ?M). Oncogenic phenotype analyses indicated that NSC1892 showed significant cytotoxicity to decrease cell proliferation, colony formation and invasion in CRC cells. Biochemical analyses demonstrated that NSC1892 treatment did not change CUL4A and CUL4B protein levels, but caused the degradation of DDB1, thereby leading to the impaired assembly of CRL4DCAF4 E3 ligases and resulting in the accumulation of ST7. The administration of NSC1892 in mice also significantly inhibited tumor growth through degrading DDB1 and accumulating ST7. Interestingly, NSC1892 also showed promising cytotoxicity to decrease the growth of other CUL4A- or CUL4B-overexpressing tumor cells such as SKOV3 ovarian cells and Saos2 osteosarcoma cells. Our results provide a new avenue for the development of a therapeutic compound targeting tumors through disrupting the CUL4-DDB1 interaction.

SUBMITTER: Yang C

PROVIDER: S-EPMC7053343 | biostudies-literature | 2020

REPOSITORIES: biostudies-literature

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Small molecule NSC1892 targets the CUL4A/4B-DDB1 interactions and causes impairment of CRL4DCAF4 E3 ligases to inhibit colorectal cancer cell growth.

Yang Chunmei C Wu Jing J He Hongbo H Liu Hong H

International journal of biological sciences 20200204 6

Cullin 4A and 4B (CUL4A and 4B) function as oncogenes in colorectal cancer (CRC) cells. Both of them conservatively associate with DNA damage-binding protein 1 (DDB1) and DDB1-CUL4-associated factor 4 (DCAF4) to form Cullin-RING E3 ligases known as CRL4DCAF4, which specifically ubiquitinate and degrade tumor suppressor ST7 (suppression of tumorigenicity 7). Knockdown either CUL4A/4B or DDB1 significantly inhibits tumor cell growth in vitro and in vivo. Thus ...[more]

PMID: 32140073

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Project description:HIV-1 Vpr-mediated G2 cell cycle arrest is dependent on the interaction of Vpr with an E3 ubiquitin ligase that contains damage-specific DNA binding protein 1 (DDB1), Cullin 4A (Cul4A), DDB1 and Cul4-associated factor 1 (DCAF1), and Rbx1. Vpr is thought to associate directly with DCAF1 in the E3 ubiquitin ligase complex although the exact interaction pattern of the proteins in the complex is not completely defined. The Vpr of SIVagm induces G2 arrest of cognate African Green Monkey (AGM) cells but not human cells. The molecular mechanism by which SIVagm Vpr exhibits its species-specific function remained unknown.Physical interaction of proteins in the E3 ubiquitin ligase complex was assessed by co-immunoprecipitation followed by western blotting. In addition, co-localization of the proteins in cells was investigated by confocal microscopy. The cell cycle was analyzed by propidium iodide staining and flow cytometry. DNA damage response elicited by Vpr was evaluated by detecting phosphorylation of H2AX, a marker for DNA damage response.We show that RNAi knock-down of DCAF1 prevented the co-immunoprecipitation of DDB1 with HIV-1 Vpr while DDB1 knock-down did not influence the binding of Vpr to DCAF1. HIV-1 Vpr mutants with a L64P or a R90K mutation maintained the ability to associate with DCAF1 but did not appear to be in a complex with DDB1. SIVagm Vpr associated with AGM DCAF1 and DDB1 while, in human cells, it binds to human DCAF1 but hardly binds to human DDB1, resulting in the reduced activation of H2AX.The identification of Vpr mutants which associate with DCAF1 but only poorly with DDB1 suggests that DCAF1 is necessary but the simple binding of Vpr to DCAF1 is not sufficient for the Vpr association with DDB1-containing E3 ligase complex. Vpr may interact both with DCAF1 and DDB1 in the E3 ligase complex. Alternatively, the interaction of Vpr and DCAF1 may induce a conformational change in DCAF1 or Vpr that promotes the interaction with DDB1. The ability of SIVagm Vpr to associate with DDB1, but not DCAF1, can explain the species-specificity of SIVagm Vpr-mediated G2 arrest.

Dataset Information

Small molecule NSC1892 targets the CUL4A/4B-DDB1 interactions and causes impairment of CRL4DCAF4 E3 ligases to inhibit colorectal cancer cell growth.

Publications

Small molecule NSC1892 targets the CUL4A/4B-DDB1 interactions and causes impairment of CRL4<sup>DCAF4</sup> E3 ligases to inhibit colorectal cancer cell growth.

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