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| Status |
Public on Jan 18, 2023 |
| Title |
Preclinical optimization of a GPC2-targeting CAR T cell therapy for neuroblastoma |
| Organisms |
Homo sapiens; Mus musculus |
| Experiment type |
Expression profiling by high throughput sequencing
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| Summary |
Background: Although most patients with newly diagnosed high-risk neuroblastoma (NB) achieve remission after initial therapy, more than 50% experience late relapses caused by minimal residual disease (MRD) and succumb to their cancer. Therapy strategies to target MRD may benefit these children. We developed a new chimeric antigen receptor (CAR) targeting glypican (GPC)2 and conducted iterative preclinical engineering of the CAR structure to maximize its anti-tumor efficacy before clinical translation. Methods: We evaluated different GPC2-CAR constructs by measuring the CAR activity against several NB cell lines in vitro. NOD-SCID mice engrafted with human NB cell lines or orthotopic patient-derived xenograft (PDX) and treated with human CAR T cells served as in vivo models. Mechanistic studies were performed using single-cell RNA-sequencing. Results: Applying stringent in vitro assays and orthotopic in vivo NB models, we demonstrated that our single-chain variable fragment, CT3, integrated into a CAR backbone with a CD28 hinge, CD28 transmembrane, and 4-1BB co-stimulatory domain elicits the best preclinical anti-NB activity compared to other tested CAR constructs. This enhanced activity was associated with an enrichment of CD8+ effector T cells in the tumor-microenvironment and upregulation of several effector molecules such as GNLY, GZMB, ZNF683, and HMGN2. Finally, we also showed that the CT3.28H.BBζ CAR was more potent than a recently clinically tested GD2-targeted CAR to control NB in vivo. Conclusion: Given the robust preclinical activity of CT3.28H.BBζ, these promising results warrant further clinical testing in children with NB.
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| Overall design |
2 donors, 3 GPC2-CAR constructs and one untransduced control. We manufactured the cells and performed single-cell RNA-seq after cell manufacturing. We then injected the cells into tumor-bearing mice and harvested tumors to perform the same analysis on cells from the TME.
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| Contributor(s) |
Sun M, Cao Y, Thiele CJ, Nguyen R |
| Citation(s) |
36631162 |
| Submission date |
Sep 03, 2022 |
| Last update date |
Jan 18, 2023 |
| Contact name |
Yingying Cao |
| E-mail(s) |
yingying.cao@nih.gov
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| Organization name |
National Institutes of Health
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| Street address |
9000 Rockville pike
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| City |
Bethesda |
| ZIP/Postal code |
20892 |
| Country |
USA |
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| Platforms (2) |
| GPL25526 |
Illumina NovaSeq 6000 (Homo sapiens; Mus musculus) |
| GPL30173 |
NextSeq 2000 (Homo sapiens) |
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| Samples (12)
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| Relations |
| BioProject |
PRJNA876577 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSE212658_RAW.tar |
351.4 Mb |
(http)(custom) |
TAR (of H5) |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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