Targeting CIC::DUX4 sarcoma with Minnelide in a dual recombinase–initiated genetically engineered mouse model
MaKenna R. Browne, Axel V. Silver, Risha Banerjee, Brendan C. Dickson, Benigno Aquino, Kristianne M. Oristian, Jonathon Himes, Peter G. Hendrickson, David G. Kirsch
MaKenna R. Browne, Axel V. Silver, Risha Banerjee, Brendan C. Dickson, Benigno Aquino, Kristianne M. Oristian, Jonathon Himes, Peter G. Hendrickson, David G. Kirsch
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Research Article Cell biology Oncology

Targeting CIC::DUX4 sarcoma with Minnelide in a dual recombinase–initiated genetically engineered mouse model

Abstract

CIC::DUX4 sarcoma (CDS) is a lethal cancer driven by a fusion between the tumor suppressor capicua (CIC) and the pioneer transcription factor double homeobox 4 (DUX4). We previously generated 3 genetically engineered mouse models (GEMMs) of CDS with CIC::DUX4 regulated by loxP-STOP-loxP cassettes, however, mice from all 3 models developed spontaneous tumors without Cre recombinase. Here, we established a next-generation GEMM of CDS (dual-flex [dFLEx] CDS) that used a dual recombinase (Cre plus the thermostable mutant of FLP recombinase FLPE) FLEx-switch design to activate CIC::DUX4 expression and initiate sarcomagenesis in a spatially and temporally controlled manner. Because CIC::DUX4 drives sarcoma development by activating an oncogenic transcriptional program, we performed a drug screen on human-derived CDS cell lines using a library of compounds that modulate transcription. This screen identified Minnelide, an inhibitor of RNA polymerase II–mediated transcription, as a selective inhibitor of CDS. Mechanistically, Minnelide acted through xeroderma pigmentosum type B to alter phosphorylation of RPB1, the largest subunit of RNA polymerase II. Subsequently, RPB1 underwent degradation leading to apoptosis of CDS cells. Minnelide demonstrated in vivo efficacy in dFLEx CDS GEMMs and in human CDS xenografts. As Minnelide has already been demonstrated to be safe in clinical trials, these findings identify Minnelide as a potential therapeutic option to test in patients with CDS.

Authors

MaKenna R. Browne, Axel V. Silver, Risha Banerjee, Brendan C. Dickson, Benigno Aquino, Kristianne M. Oristian, Jonathon Himes, Peter G. Hendrickson, David G. Kirsch

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Figure 5

Minnelide induces cell death independent of transcriptional inhibition.

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Minnelide induces cell death independent of transcriptional inhibition. ...
(A) Schematic for the Minnelide time course in Kitra-SRS human CDS cells. (B) Venn diagrams representing transcriptional changes during early, mid, and late Minnelide treatment times. (C) Volcano plots demonstrating transcriptional changes following Minnelide treatment as a function of time. (D) FLICK assays on human ECD1 and X1C1 cells demonstrating that the onset of cell death was 22.54 hours and 18.65 hours after the start of Minnelide treatment, respectively. (E) EU incorporation 24, 48, and 72 hours after Minnelide treatment to assess changes in nascent transcription in ECD1 and X1C1 cells. (F) PTBP1 immunofluorescence on human CDS#2 cells after 48 hours Minnelide treatment. U2OS osteosarcoma cells were used as a positive control. Scale bars: 15 μm. (G) Quantification of cells with cytoplasmic PTBP1. A total of 24–25 regions per condition were quantified, including regions sampled from 5 independent experiments. *P < 0.05, by 2-tailed exact Mann-Whitney U test. Data are presented as the mean ± SEM.