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Updated: Jul 18, 2025
Author Spotlight: Modeling Brain Tumors In Vivo Using Electroporation-Based Delivery of Plasmid DNA Representing Patient Mutation Signatures
Published on: June 23, 2023
Modeling human brain rhabdoid tumor by inactivating tumor suppressor genes in induced pluripotent stem cells
Timothy Hua1, Yu Xue1, Drishty B Sarker1
1Department of Chemistry and Biochemistry, Florida State University, Tallahassee, FL, 32306-4390, USA.
Abstract:
Atypical teratoid/rhabdoid tumor (ATRT) is a rare childhood malignancy that originates in the central nervous system. Over ninety-five percent of ATRT patients have biallelic inactivation of the tumor suppressor gene SMARCB1. ATRT has no standard treatment, and a major limiting factor in therapeutic development is the lack of reliable ATRT models. We employed CRISPR/Cas9 gene-editing technology to knock out SMARCB1 and TP53 genes in human episomal induced pluripotent stem cells (Epi-iPSCs), followed by brief neural induction, to generate an ATRT-like model. The dual knockout Epi-iPSCs retained their stemness with the capacity to differentiate into three germ layers. High expression of OCT4 and NANOG in neurally induced knockout spheroids was comparable to that in two ATRT cell lines. Beta-catenin protein expression was higher in SMARCB1-deficient cells and spheroids than in normal Epi-iPSC-derived spheroids. Nucleophosmin, Osteopontin, and Ki-67 proteins were also expressed by the SMARCB1-deficient spheroids. In summary, the tumor model resembled embryonal features of ATRT and expressed ATRT biomarkers at mRNA and protein levels. Ribociclib, PTC-209, and the combination of clofilium tosylate and pazopanib decreased the viability of the ATRT-like cells. This disease modeling scheme may enable the establishment of individualized tumor models with patient-specific mutations and facilitate high-throughput drug testing.
Insights
Researchers developed a novel atypical teratoid/rhabdoid tumor (ATRT) model using CRISPR gene editing to target SMARCB1. This model aids in understanding ATRT and testing potential therapies for this rare childhood brain cancer.
Area of Science:
- Neuro-oncology
- Developmental Biology
- Cancer Genetics
Background:
- Atypical teratoid/rhabdoid tumor (ATRT) is a rare pediatric central nervous system malignancy.
- Most ATRT cases involve biallelic inactivation of the SMARCB1 tumor suppressor gene.
- Lack of reliable ATRT models hinders therapeutic development.
Purpose of the Study:
- To generate a novel in vitro model of ATRT using gene-editing technology.
- To characterize the generated model for its resemblance to ATRT.
- To evaluate potential therapeutic agents against the ATRT model.
Main Methods:
- CRISPR/Cas9 gene editing was used to knock out SMARCB1 and TP53 in human episomal induced pluripotent stem cells (Epi-iPSCs).
- Neura l induction was performed to generate ATRT-like spheroids.
- Gene and protein expression of ATRT markers were analyzed.
- Cell viability assays were conducted using various drug treatments.
Main Results:
- The dual knockout Epi-iPSCs maintained stemness and differentiated into three germ layers.
- Neurally induced spheroids exhibited high OCT4 and NANOG expression, comparable to ATRT cell lines.
- SMARCB1-deficient cells and spheroids showed increased beta-catenin expression and expressed ATRT biomarkers (Nucleophosmin, Osteopontin, Ki-67).
- Ribociclib, PTC-209, and a clofilium tosylate/pazopanib combination reduced ATRT-like cell viability.
Conclusions:
- The developed model recapitulates embryonal features and biomarkers of ATRT at the mRNA and protein levels.
- This ATRT model facilitates the study of disease mechanisms and biomarker expression.
- The model shows promise for high-throughput drug screening and the development of individualized ATRT models.
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