Targeted brachyury degradation disrupts a highly specific autoregulatory program controlling chordoma cell identity

Sheppard, Hadley E.; Dall’Agnese, Alessandra; Park, Woojun D; Shamim, Md; Dubrulle, Julien; Johnson, Hannah; Stossi, Fabio; Cogswell, Patricia C.; Sommer, Josh; Levy, Joan; Sharifnia, Tanaz; Wawer, Mathias; Nabet, Behnam; Gray, Nathanael S.; Clemons, Paul A.; Schreiber, Stuart L.; Workman, Paul; Young, Richard A.; Lin, Charles Y.

doi:10.1016/j.xcrm.2020.100188

Cited by 23 publications

(34 citation statements)

References 86 publications

(99 reference statements)

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“…CH22) [79] with up to 24 h THZ1 treatment and a study on THZ1 treatment of B-cell acute lymphocytic leukemia cells (Nalm6) [80]. Realignment of corresponding samples confirmed linear splicing upon THZ1 treatment in all cell lines apart from DU145 and showed that splicing rates increased with the duration and dosage of THZ1 treatment (Fig 6A and 6B and S22A Fig).…”

Section: Plos Onementioning

confidence: 78%

“…Experiments identified with the Snaptron search cover prostate cancer (VCaP, LNCaP and DU145 cells) [ 42 ], bladder cancer (HCV-29 cells) [ 74 ], esophageal squamous cell carcinoma (TE7 and KYSE510 cells) [ 75 ], nasopharyngeal carcinoma (C666-1, HK1 and HNE1 cells) [ 76 ], pancreatic ductal adenocarcinoma (BxPC3, MiaPaCa-2 and PANC1 cells) [ 77 ] and chordoma (UM-Chor1 cells) [ 78 ]. Further literature search identified a second study of chordoma cells (UM-Chor1 and CH22) [ 79 ] with up to 24 h THZ1 treatment and a study on THZ1 treatment of B-cell acute lymphocytic leukemia cells (Nalm6) [ 80 ]. Realignment of corresponding samples confirmed linear splicing upon THZ1 treatment in all cell lines apart from DU145 and showed that splicing rates increased with the duration and dosage of THZ1 treatment ( Fig 6A and 6B and S22A Fig ).…”

Section: Resultsmentioning

confidence: 99%

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HSV-1 and influenza infection induce linear and circular splicing of the long NEAT1 isoform

Friedl

Djaković²,

Kluge³

et al. 2022

PLoS ONE

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The herpes simplex virus 1 (HSV-1) virion host shut-off (vhs) protein cleaves both cellular and viral mRNAs by a translation-initiation-dependent mechanism, which should spare circular RNAs (circRNAs). Here, we show that vhs-mediated degradation of linear mRNAs leads to an enrichment of circRNAs relative to linear mRNAs during HSV-1 infection. This was also observed in influenza A virus (IAV) infection, likely due to degradation of linear host mRNAs mediated by the IAV PA-X protein and cap-snatching RNA-dependent RNA polymerase. For most circRNAs, enrichment was not due to increased circRNA synthesis but due to a general loss of linear RNAs. In contrast, biogenesis of a circRNA originating from the long isoform (NEAT1_2) of the nuclear paraspeckle assembly transcript 1 (NEAT1) was induced both in HSV-1 infection–in a vhs-independent manner–and in IAV infection. This was associated with induction of novel linear splicing of NEAT1_2 both within and downstream of the circRNA. NEAT1_2 forms a scaffold for paraspeckles, nuclear bodies located in the interchromatin space, must likely remain unspliced for paraspeckle assembly and is up-regulated in HSV-1 and IAV infection. We show that NEAT1_2 splicing and up-regulation can be induced by ectopic co-expression of the HSV-1 immediate-early proteins ICP22 and ICP27, potentially linking increased expression and splicing of NEAT1_2. To identify other conditions with NEAT1_2 splicing, we performed a large-scale screen of published RNA-seq data. This uncovered both induction of NEAT1_2 splicing and poly(A) read-through similar to HSV-1 and IAV infection in cancer cells upon inhibition or knockdown of CDK7 or the MED1 subunit of the Mediator complex phosphorylated by CDK7. In summary, our study reveals induction of novel circular and linear NEAT1_2 splicing isoforms as a common characteristic of HSV-1 and IAV infection and highlights a potential role of CDK7 in HSV-1 or IAV infection.

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Section: Plos Onementioning

confidence: 78%

Section: Resultsmentioning

confidence: 99%

HSV-1 and influenza infection induce linear and circular splicing of the long NEAT1 isoform

Friedl

Djaković²,

Kluge³

et al. 2022

PLoS ONE

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show abstract

“…Identified experiments cover prostate cancer (VCaP, LNCaP and DU145 cells) (Rasool et al 2019), bladder cancer (HCV-29 cells) (Zarei et al 2019), esophageal squamous cell carcinoma (TE7 and KYSE510 cells) (Jiang et al 2017), nasopharyngeal carcinoma (C666-1, HK1 and HNE1 cells) (Yuan et al 2017), pancreatic ductal adenocarcinoma (BxPC3, MiaPaCa-2 and PANC1 cells) (Lu et al 2019) and chordoma (UM-Chor1 cells) (Sharifnia et al 2019). Further literature search identified a second study of chordoma cells (UM-Chor1 and CH22) (Sheppard et al 2021) with up to 24 h THZ1 treatment and a study on THZ1 treatment of B-cell acute lymphocytic leukemia cells (Nalm6) (Abudureheman et al 2021). Realignment of corresponding samples confirmed linear splicing in all cell lines apart from DU145 and showed that induction increased with the duration and dosage of THZ1 treatment (Fig.…”

Section: Cdk7 Inhibition Induces Neat1_2 Splicing and Disrupts Transcription Termination In Cancer Cell Linesmentioning

confidence: 99%

“…nasopharyngeal carcinoma (C666-1, HK1 and HNE1 cells) [69], pancreatic ductal adenocarcinoma (BxPC3, MiaPaCa-2 and PANC1 cells) [70] and chordoma (UM-Chor1 cells) [71]. Further literature search identified a second study of chordoma cells (UM-Chor1 and CH22) [72] with up to 24 h THZ1 treatment and a study on THZ1 treatment of B-cell acute lymphocytic leukemia cells (Nalm6) [73]. Realignment directly phosphorylates MED1 at Threonine 1457 [38], which together with a phosphorylation at Threonine 1032 promotes MED1 association with the Mediator complex [75].…”

Section: Cdk7 Inhibition Induces Neat1_2 Splicing and Disrupts Transc...mentioning

confidence: 99%

HSV-1 and influenza infection induce linear and circular splicing of the long NEAT1 isoform

Friedl

Djaković

Kluge

et al. 2021

Preprint

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The herpes simplex virus 1 (HSV-1) virion host shut-off (vhs) protein cleaves both cellular and viral mRNAs but not circular RNAs (circRNAs) without an internal ribosome entry site. Here, we show that vhs activity leads to an accumulation of circRNAs relative to linear mRNAs during HSV-1 infection. Strikingly, we found that circular splicing of the long isoform (NEAT1_2) of the nuclear paraspeckle assembly transcript 1 (NEAT1) was massively induced during HSV-1 infection in a vhs-independent manner while NEAT1_2 was still bound to the chromatin. This was associated with induction of linear splicing of NEAT1_2 both within and downstream of the circRNA. NEAT1_2 splicing was absent in uninfected cells but can be induced by ectopic co-expression of the HSV-1 immediate-early proteins ICP22 and ICP27. Interestingly, NEAT1_2 circular and linear splicing was also up-regulated in influenza infection but absent in stress conditions, which disrupt transcription termination similar to but not in the same manner as HSV-1 and influenza infection. Finally, large-scale analysis of published RNA-seq data uncovered induction of NEAT1_2 splicing in cancer cells upon inhibition or knockdown of cyclin-dependent kinase 7 (CDK7) or the MED1 subunit of the Mediator complex phosphorylated by CDK7. Interestingly, CDK7 inhibition also disrupted transcription termination, highlighting a possible link between disruption of transcription termination and NEAT1_2 splicing.

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“…20 Brachyury is an essential transcriptional regulator of chordoma identity and is related to super-enhancer functionality, thus representing a therapeutic target of special interest for chordoma. 21 22 Overexpression of brachyury messenger RNA was also found in gastrointestinal, bladder, kidney, ovarian, uterine, and testicular carcinomas, as well as in cell lines of lung, colon, and prostate cancers. 6 In contrast, brachyury messenger RNA was notably absent in the vast majority of normal tissues tested, making brachyury an attractive target for the development of novel anticancer therapies.…”

Section: Introductionmentioning

confidence: 99%

Phase 1 open-label trial of intravenous administration of MVA-BN-brachyury-TRICOM vaccine in patients with advanced cancer

DeMaria

Lee-Wisdom

Donahue

et al. 2021

J Immunother Cancer

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BackgroundMVA-BN-brachyury-TRICOM is a recombinant vector-based therapeutic cancer vaccine designed to induce an immune response against brachyury. Brachyury, a transcription factor overexpressed in advanced cancers, has been associated with treatment resistance, epithelial-to-mesenchymal transition, and metastatic potential. MVA-BN-brachyury-TRICOM has demonstrated immunogenicity and safety in previous clinical trials of subcutaneously administered vaccine. Preclinical studies have suggested that intravenous administration of therapeutic vaccines can induce superior CD8+ T cell responses, higher levels of systemic cytokine release, and stronger natural killer cell activation and proliferation. This is the first-in-human study of the intravenous administration of MVA-BN-brachyury-TRICOM.MethodsBetween January 2020 and March 2021, 13 patients were treated on a phase 1, open-label, 3+3 design, dose-escalation study at the National Institutes of Health Clinical Center. The study population was adults with advanced solid tumors and was enriched for chordoma, a rare sarcoma of the notochord that overexpresses brachyury. Vaccine was administered intravenously at three DLs on days 1, 22, and 43. Blood samples were taken to assess drug pharmacokinetics and immune activation. Imaging was conducted at baseline, 1 month, and 3 months post-treatment. The primary endpoint was safety and tolerability as determined by the frequency of dose-limiting toxicities; a secondary endpoint was determination of the recommended phase 2 dose.ResultsNo dose-limiting toxicities were observed and no serious adverse events were attributed to the vaccine. Vaccine-related toxicities were consistent with class profile (ie, influenza-like symptoms). Cytokine release syndrome up to grade 2 was observed with no adverse outcomes. Dose-effect trend was observed for fever, chills/rigor, and hypotension. Efficacy analysis of objective response rate per RECIST 1.1 at the end of study showed one patient with a partial response, four with stable disease, and eight with progressive disease. Three patients with stable disease experienced clinical benefit in the form of improvement in pain. Immune correlatives showed T cell activation against brachyury and other tumor-associated cascade antigens.ConclusionsIntravenous administration of MVA-BN-brachyury-TRICOM vaccine was safe and tolerable. Maximum tolerated dose was not reached. The maximum administered dose was 109 infectious units every 3 weeks for three doses. This dose was selected as the recommended phase 2 dose.Trial registration numberNCT04134312.

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Targeted brachyury degradation disrupts a highly specific autoregulatory program controlling chordoma cell identity

Cited by 23 publications

References 86 publications

HSV-1 and influenza infection induce linear and circular splicing of the long NEAT1 isoform

HSV-1 and influenza infection induce linear and circular splicing of the long NEAT1 isoform

HSV-1 and influenza infection induce linear and circular splicing of the long NEAT1 isoform

Phase 1 open-label trial of intravenous administration of MVA-BN-brachyury-TRICOM vaccine in patients with advanced cancer

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