Abstract B56: A CRISPR/Cas9 domain screen identifies a small motif in the PAX3-FOXO1 transactivation domain relevant for tumor maintenance in alveolar rhabdomyosarcoma
Bibliographic record
Abstract
Abstract Fusion transcription factors are the main drivers of tumorigenesis in many pediatric leukemias and sarcomas. One example is PAX3-FOXO1 (P3F), the pathognomonic marker of alveolar rhabdomyosarcoma (aRMS). Since these tumors are often also characterized by a quiet mutational landscape and a paucity of druggable oncogenes, the fusion proteins often remain as the only relevant drug target. However, transcription factors are challenging targets for small-molecule inhibitors, since they lack an enzymatic activity and, apart from the DNA binding domains, are usually intrinsically disordered. Identification of druggable structures in such proteins is therefore of great clinical interest. Our goal of this study was to identify new elements important for P3F activity. Towards this aim, we made use of the CRISPR/Cas9 technology and performed a saturated screen for identification of important functional domains in P3F in the endogenous context. This revealed that both DNA binding domains are indispensable, as expected for a transcription factor. Importantly, however, the screen also identified a 40-amino-acid-long domain at the C-terminus of the FOXO1 part to be strictly required for the function of the protein. Using a series of mutations in this domain in reporter assays, we identified a single cysteine whose mutation leads to more than 50% reduction in transcriptional activity. In addition, cells efficiently differentiated and stopped proliferating when this Cys was mutated in aRMS cells. These results suggest that functional interference with this domain could be a promising approach for aRMS therapy. Citation Format: Marco Wachtel, Katharina Benischke, Beat W. Schäfer. A CRISPR/Cas9 domain screen identifies a small motif in the PAX3-FOXO1 transactivation domain relevant for tumor maintenance in alveolar rhabdomyosarcoma [abstract]. In: Proceedings of the AACR Special Conference on the Advances in Pediatric Cancer Research; 2019 Sep 17-20; Montreal, QC, Canada. Philadelphia (PA): AACR; Cancer Res 2020;80(14 Suppl):Abstract nr B56.
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How this classification was reachedexpand
Full frame machine prediction
Teacher imitationNot calibrated prevalence, not ground truth. Human validation pending. The Gemma side is a direct model label for every work in the frame, read from the title-only record. The Codex side is a classifier learned from the 10,348 direct Codex labels and calibrated to design-weighted sample rates; fields without enough sample support carry no Codex call. Candidate is the union of the two sides; consensus is their intersection. These outputs are machine_predicted_unvalidated and are not human labels.
Distilled classifier scores by category (both heads)
| Category | Codex | Gemma |
|---|---|---|
| Metaresearch | 0.000 | 0.000 |
| Meta-epidemiology (narrow) | 0.000 | 0.000 |
| Meta-epidemiology (broad) | 0.000 | 0.000 |
| Bibliometrics | 0.000 | 0.000 |
| Science and technology studies | 0.000 | 0.000 |
| Scholarly communication | 0.000 | 0.000 |
| Open science | 0.000 | 0.000 |
| Research integrity | 0.000 | 0.000 |
| Insufficient payload (model declined to judge) | 0.003 | 0.001 |
Machine scores (provisional)
The two teacher heads of the student model, read on this work. A score orders the frame for review; it never asserts a category, and the validation status ships verbatim with every row.
Baseline scores from an immature model (maturity gate not passed, 7 training rounds). Scores rank; they never assert a category.
score_only:v0-immature-baseline · verbatim from the scoring run: score_only means the number may rank works, and no category label ships from itClassification
machine, unvalidatedMachine predicted; a candidate call from one source (direct Gemma or distilled Codex), not a consensus.
How this classification was reached, model by model and score by score, is at the end of the page under "How this classification was reached".