Drug Repositioning Inferred from E2F1-Coregulator Interactions Studies for the Prevention and Treatment of Metastatic Cancers

Abstract

Metastasis management remains a long-standing challenge. High abundance of E2F1 triggers tumor progression by developing protein-protein interactions (PPI) with coregulators that enhance its potential to activate a network of prometastatic transcriptional targets. Methods: To identify E2F1-coregulators, we integrated high-throughput Co-immunoprecipitation (IP)/mass spectometry, GST-pull-down assays, and structure modeling. Potential inhibitors of PPI discovered were found by bioinformatics-based pharmacophore modeling, and transcriptome profiling was conducted to screen for coregulated downstream targets. Expression and target gene regulation was validated using qRT-PCR, immunoblotting, chromatin IP, and luciferase assays. Finally, the impact of the E2F1-coregulator complex and its inhibiting drug on metastasis was investigated in vitro in different cancer entities and two mouse metastasis models. Results: We unveiled that E2F1 forms coactivator complexes with metastasis-associated protein 1 (MTA1) which, in turn, is directly upregulated by E2F1. The E2F1:MTA1 complex potentiates hyaluronan synthase 2 (HAS2) expression, increases hyaluronan production and promotes cell motility. Disruption of this prometastatic E2F1:MTA1 interaction reduces hyaluronan synthesis and infiltration of tumor-associated macrophages in the tumor microenvironment, thereby suppressing metastasis. We further demonstrate that E2F1:MTA1 assembly is abrogated by small-molecule, FDA-approved drugs. Treatment of E2F1/MTA1-positive, highly aggressive, circulating melanoma cells and orthotopic pancreatic tumors with argatroban prevents metastasis and cancer relapses in vivo through perturbation of the E2F1:MTA1/HAS2 axis. Conclusion: Our results propose argatroban as an innovative, E2F-coregulator-based, antimetastatic drug. Cancer patients with the infaust E2F1/MTA1/HAS2 signature will likely benefit from drug repositioning.

LncRNA-SLC16A1-AS1 induces metabolic reprogramming during Bladder Cancer progression as target and co-activator of E2F1

“…E2F1 promotes the invasiveness of urothelial bladder carcinoma 3 , 4 , 9 , 10 . To study the role of this transcription factor in bladder cancer progression, we utilized a human tissue culture model consisting of various cell lines that differ in their invasive potential according to the endogenous E2F1 expression status.…”

“…One of the main mechanisms is that they act as transcriptional regulators 61 . Notably, it was recently shown that E2F1 controls the expression of PCGs whose products in turn, physically associate with E2F1 to enhance transcription of prometastatic targets 6 , 9 , 72 . With this in mind, we hypothesized that this could be also the case for the nucleus-localized lncRNA-SLC16A1-AS1, which may interact with E2F1 to fine-tune transcription of E2F1-regulated targets.…”

“…The corresponding peroxidase-labeled secondary antibodies were detected using ECL Western blot reagents (Amersham Biosciences). Immunofluorescence assays were carried out as described in Goody et al 9 using MCT1 (H-1) as primary and fluorescence-labeled anti-mouse A-488 as secondary antibody (Dianova, 715-5454-150).…”

“…RT-4, RT-112 and T24 cells were transiently transduced with adenoviral vector Ad.ER-E2F1 1 at MOI 100. Cells stably containing E2F1 protein fused to the hormone-binding domain of the estrogen receptor (ER) were produced as described 9 and grown in medium containing 2 µg/ml puromycin (Sigma Chemical Co., St. Louis, MO). The ER-E2F1 fusion protein was activated by addition of 0.5 µM 4-hydroxytamoxifen (4-OHT).…”

“…ChIP was performed as described 9 . Protein-DNA complexes were immunoprecipitated using anti-E2F1 antibody (KH-95) or control IgG (Abcam).…”