Preclinical optimization of a GPC2-targeting CAR T cell therapy for neuroblastoma
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ABSTRACT:
PROVIDER: PRJNA876577 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR21413145_1.fastq.gz | Fastqsanger.gz | |||
| SRR21413145_2.fastq.gz | Fastqsanger.gz | |||
| SRR21413146_1.fastq.gz | Fastqsanger.gz | |||
| SRR21413146_2.fastq.gz | Fastqsanger.gz | |||
| SRR21413147_1.fastq.gz | Fastqsanger.gz |
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Project description: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.
2023-01-18 | GSE212658 | GEO
Project description:BackgroundAlthough 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. Therapeutic strategies to target MRD may benefit these children. We developed a new chimeric antigen receptor (CAR) targeting glypican-2 (GPC2) and conducted iterative preclinical engineering of the CAR structure to maximize its anti-tumor efficacy before clinical translation.MethodsWe evaluated different GPC2-CAR constructs by measuring the CAR activity in vitro. NOD-SCID mice engrafted orthotopically with human NB cell lines or patient-derived xenografts and treated with human CAR T cells served as in vivo models. Mechanistic studies were performed using single-cell RNA-sequencing.ResultsApplying stringent in vitro assays and orthotopic in vivo NB models, we demonstrated that our single-chain variable fragment, CT3, integrated into a CAR vector with a CD28 hinge, CD28 transmembrane, and 4-1BB co-stimulatory domain (CT3.28H.BBζ) elicits the best preclinical anti-NB activity compared with 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 we developed was more potent than a recently clinically tested GD2-targeted CAR to control NB growth in vivo.ConclusionGiven the robust preclinical activity of CT3.28H.BBζ, these results form a promising basis for further clinical testing in children with NB.
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Project description:Here we characterize and optimize both systems to increase their utility for preclinical studies. We show that TH-MYCN mice develop tumors in the paraspinal ganglia, but not in the adrenal, with cellular and gene expression patterns similar to human NB. In addition, we present a new ultrasound guided, non-invasive orthotopic xenograft method. This injection technique is rapid, provides accurate targeting of the injected cells and leads to efficient engraftment. We also demonstrate that tumors can be detected, monitored and quantified prior to visualization using ultrasound, MRI and bioluminescence. Finally we develop and test a “standard of care” chemotherapy regimen. This protocol, which is based on current treatments for neuroblastoma, provides a baseline for comparison of new therapeutic agents. A total of 17 samples are provided reprsenting TH-MYCN tumors derived from mice
2011-03-17 | E-GEOD-27516 | biostudies-arrayexpress
Project description:Chimeric antigen receptor T cells (CAR-T) therapy has not yet been fully explored in solid tumors. Human epidermal growth factor receptor-2(HER2) is widely expressed in cancers. Investigators have developed anti-HER2 CAR-modified T cells and validated the efficiency targeting HER2-positive cancer in preclinical studies. This study is aimed to confirm its adverse effects including cytokine storm response and any other adverse effects. In addition, CAR-T cells persistence, tumor elimination and disease status after treatment will be evaluated.
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Project description:Optimization of CAR T cell signaling by incorporation of null mutations
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Project description:An obstacle with continued clinical development of CAR T cells is the limited understanding of CAR T cell biology and its mechanisms of anti-tumor immunity. We and others have shown that CARs with a CD28 co-stimulatory domain drive high levels of T cell activation that also lead to exhaustion and shortened persistence. This led us to hypothesize that by incorporating null mutations of CD28 subdomains (YMNM, PRRP, or PYAP) we could optimize CAR T cell signaling and reduce exhaustion. In vivo we found that mice given CAR T cells with a PYAP CD28 endodomain had a significant survival advantage. We also observed that mutant CAR T cells had significantly less expression of PD1 and were increased in the bone marrow and spleen. In addition, these mutant CAR T cells had optimized signaling resulting in a reduction of exhaustion related transcription factors and genes. Our results demonstrate that CAR T cells with a mutant CD28 endodomain have better survival and decreased exhaustion and is the result of decreased CAR dependent NFAT/NR4A1 transcription factors. This work allows for development of enhanced CAR T cell therapies by optimizing CAR T cell signaling.
2020-12-16 | GSE142506 | GEO
Project description:Diffuse intrinsic pontine glioma (DIPG) and other H3K27M-mutated diffuse midline gliomas (DMGs) are universally lethal central nervous system (CNS) tumors that occur most commonly in children and young adults1. Average life expectancy is ten months from diagnosis, and 5-year survival is less than 1%2. Palliative radiotherapy is the only established treatment3, and neither cytotoxic nor targeted pharmacological approaches have demonstrated anti-tumor responses or improved prognosis to date3,4. We previously discovered that the disialoganglioside GD2 is highly and uniformly expressed on H3K27M+ DMG cells and demonstrated that intravenously (IV) administered GD2.4-1BB.z chimeric antigen receptor (CAR) T-cells eradicated established DMGs in patient-derived orthotopic murine models5, thereby providing the rationale for a first-in-human/first-in-child Phase 1 clinical trial (NCT04196413). Because CAR T-cell-induced inflammation and edema of the brainstem can result in obstructive hydrocephalus, increased intracranial pressure, and dangerous tissue shifts, a number of neurocritical care precautions were incorporated in the clinical trial design and management plan. Here we present the clinical experience from the first four patients with H3K27M+ DMG treated with GD2-CAR T-cells at dose level 1 (DL1; 1e6 GD2-CAR T-cells/kg administered IV). Patients who exhibited clinical benefit were eligible for subsequent administrations of GD2-CAR T-cells. Given preclinical evidence for increased CAR T-cell potency6, and the potential for diminished immunogenicity with locoregional administration, second doses were administered intracerebroventricularly (ICV) through an Ommaya catheter to three patients. As predicted from preclinical models, toxicity was largely related to the neuroanatomical location of the tumors and was reversible with intensive supportive care. Although GD2 is expressed at low levels in normal neural tissue, no evidence of on-target, off-tumor toxicity was observed. Three of four patients exhibited clinical and radiographic improvement, underscoring the promise of this approach for H3K27M+ DMG therapy. Correlative studies of serum and CSF revealed marked proinflammatory cytokine production following GD2 CAR T cell administration and single cell transcriptomic analysis of 65,598 single cells from CAR T cell products and patient CSF has begun to reveal differences that correlate with the heterogeneity between subjects and routes of administration.
2022-02-07 | GSE186802 | GEO
Project description:Neuroblastoma (NB) is the most common extra-cranial solid tumor in children and remains deadly in patients with high-risk disease. Single chimeric antigen receptor T-cell therapies (CAR-T) for solid tumor have shown poor efficacy in clinical trials due, in part, to T cell exhaustion and uneven expression of target antigens in tumors. Here, we attempted to target Glypican-2(GPC2) and CD276(B7-H3), both highly but heterogeneously expressed on NB cell surface, to overcome these challenges in single CAR T- therapies. First, to identify optimal binders for CAR T- cell targeting each antigen, we made individual CAR constructs using multiple binders against these antigens and utilized a multimodal approach including simultaneous protein and transcriptome measurement at single cell level to characterize each individual CAR T- cell in a pooled strategy. Combined with cell expansion and cytotoxicity assays, we identified the most effective CAR construct for each target.
2022-03-18 | GSE181437 | GEO
Project description:Here we characterize and optimize both systems to increase their utility for preclinical studies. We show that TH-MYCN mice develop tumors in the paraspinal ganglia, but not in the adrenal, with cellular and gene expression patterns similar to human NB. In addition, we present a new ultrasound guided, non-invasive orthotopic xenograft method. This injection technique is rapid, provides accurate targeting of the injected cells and leads to efficient engraftment. We also demonstrate that tumors can be detected, monitored and quantified prior to visualization using ultrasound, MRI and bioluminescence. Finally we develop and test a “standard of care” chemotherapy regimen. This protocol, which is based on current treatments for neuroblastoma, provides a baseline for comparison of new therapeutic agents.
2011-03-17 | GSE27516 | GEO