A5000823642- CAR-T cell therapy research
- CRISPR and Genetic Engineering
- Transgenic Plants and Applications
- vaccines and immunoinformatics approaches
- Immune Cell Function and Interaction
- Monoclonal and Polyclonal Antibodies Research
- Advanced biosensing and bioanalysis techniques
- Silicon Carbide Semiconductor Technologies
- Viral Infectious Diseases and Gene Expression in Insects
- T-cell and B-cell Immunology
- Nanowire Synthesis and Applications
- Acute Lymphoblastic Leukemia research
- Cancer Immunotherapy and Biomarkers
- Virus-based gene therapy research
- Advancements in Semiconductor Devices and Circuit Design
- Immunotherapy and Immune Responses
- Biosimilars and Bioanalytical Methods
Washington University in St. Louis
2021-2024
While chimeric antigen receptor (CAR) T cells targeting CD19 can cure a subset of patients with B cell malignancies, most treated will not achieve durable remission. Identification the mechanisms leading to failure is essential broadening efficacy this promising platform. Several studies have demonstrated that disruption genes and transcripts lead disease relapse after initial response; however, few other tumor-intrinsic drivers CAR been reported. Here we identify expression Golgi-resident...
Chimeric antigen receptor (CAR)-based therapies have pioneered synthetic cellular immunity but remain limited in their long-term efficacy. Emerging data suggest that dysregulated CAR-driven T-cell activation causes dysfunction and therapeutic failure. To re-engage the precision of endogenous response, we designed MHC-independent receptors (miTCR) by linking antibody variable domains to constant chains. Using predictive modeling, observed this standard "cut paste" approach protein design...
<title>Abstract</title> Nearly all chimeric antigen receptors (CARs) initiate intracellular signaling in the absence of antigen, referred to as “tonic signaling”. Tonic CARs containing CD28 costimulatory domain has been shown drive T cell exhaustion; contrast, we previously found that tonic 41BB-containing enhances function. Using a panel targeting B CD22, identified 41BB activates BACH2, transcriptional regulator directs stem and memory programs. Overexpression BACH2 prevented exhaustion...
Several pre-clinical models reveal that chronic chimeric antigen receptor (CAR) stimulation drives a dysfunctional state mimics in vivo failure. In this protocol, we describe steps to induce T cell dysfunction by persistent and long-term of CAR-engineered cells using antigen-expressing cancer suspension cultures. We first described validated method for manufacturing CAR cells, followed detailed strategy evaluate these during the process stimulation. For complete details on use execution...
CD19-targeted immunotherapies that stimulate a cytotoxic T-cell response have revolutionized the management of B-cell malignancies, specifically relapsed or refractory acute lymphoblastic leukemia (R/ R ALL).Blinatumomab is an anti-CD19 bispecific engager approved for treatment patients with R/R ALL. 1 KTE-X19 autologous chimeric antigen receptor (CAR) therapy under investigation in ZUMA-3 (XXX) phase 1/2 clinical trial (registered at www.clinicaltrials.gov2][3][4][5] Several distinct...
Abstract Chimeric antigen receptor (CAR)-based therapies have pioneered synthetic cellular immunity against cancer, however remain limited in their scope and long-term efficacy. Emerging data suggest that dysregulated CAR-driven T cell activation causes dysfunction therapeutic failure. To re-engage the endogenous response, we designed hybrid MHC-independent receptors (miTCRs) by linking antibody variable domains to TCR constant domains. While functional, observed stark differences...
<div>Abstract<p>Genome editing technologies have seen remarkable progress in recent years, enabling precise regulation of exogenous and endogenous genes. These advances been extensively applied to the engineering human T lymphocytes, leading development practice changing therapies for patients with cancer promise synthetic immune cell a variety nonmalignant diseases. Many distinct conceptual technical approaches used edit T-cell genomes, however targeted assessments which...
<p>A, Expansion of T cells in which stimulation cultures did or not contain IL7/IL15. B, CD4 and CD8 composition C, memory cell products at the conclusion manufacturing when include IL7/IL15.</p>
<p>A, Representative ow cytometry plots of transduction efficiency various synthetic receptors in control (unedited) and TRAC+TRBCKO T cells when editing is performed pre-stimulation. B, Stability CRISPR over-time performing post-stimulation editing.</p>
<p>A, Representative ow cytometry plots of transduction efficiency various synthetic receptors in control (unedited) and TRAC+TRBCKO T cells when editing is performed pre-stimulation. B, Stability CRISPR over-time performing post-stimulation editing.</p>
<p>A, Expansion of T cells in which stimulation cultures did or not contain IL7/IL15. B, CD4 and CD8 composition C, memory cell products at the conclusion manufacturing when include IL7/IL15.</p>
<div>Abstract<p>Genome editing technologies have seen remarkable progress in recent years, enabling precise regulation of exogenous and endogenous genes. These advances been extensively applied to the engineering human T lymphocytes, leading development practice changing therapies for patients with cancer promise synthetic immune cell a variety nonmalignant diseases. Many distinct conceptual technical approaches used edit T-cell genomes, however targeted assessments which...
<p>Supplementary Figure 6 | a-b, Cytotoxicity of a, miTCR1 and b, miTCR2 variants against Nalm6. c-d, Long-term flow cytometry-based cytotoxicity lead c, d, variants. e, Quantification cytokines in supernatants 24h after co-culture engineered T cells with f, Expression TCRC chains αCD22 miTCR1, CD22 CD19 Jurkat cells. g, Induction reporter NFAT-GFP upon exposure to CD19+CD22+ Nalm6 Screening studies a-b performed using two independent donors; representative data from three one donor....
<p>Supplementary Figure 7 | a, Overlay of predicted mut031 α chain variable region and resolved FMC63 structure. Modeling performed using AlphaFold2. b, Memory lineages human T cells engineered to express miTCR1 WT or mut035 at the conclusion manufacturing cultures. c-d, Proportion that were early lineage (naïve central memory) c, throughout 6 day stimulation cultures d, one after clearance Nalm6. e, Expansion expressing WT, 19/BBζ CAR from two donors. b-d, data donor.</p>
<div>Abstract<p>Chimeric antigen receptor (CAR)–based therapies have pioneered synthetic cellular immunity but remain limited in their long-term efficacy. Emerging data suggest that dysregulated CAR-driven T-cell activation causes dysfunction and therapeutic failure. To re-engage the precision of endogenous response, we designed MHC-independent receptors (miTCR) by linking antibody variable domains to constant chains. Using predictive modeling, observed this standard “cut paste”...
<p>Supplementary Figure 5 | a-b, Expression of TCR constant chains on the surface a, Jurkat and b, human T cells transduced with miTCR variants. c-d, Comparison expression in miTCR1 miTCR2 variant engineered c, Jurkats d, cells. e, Change based presence or absence PD insertion miTCRα chain. f, NFAT g, NFκB reporter expressing 1G4 variants response to Nalm6. h, Expansion upon exposure cumulative data from three independent donors. b,e, representative Screening studies h performed using...
<p>Supplementary Figure 1 | a-b, Expression of NFAT and NFκB in reporter Jurkat cells expressing CARs or miTCRs response to a, CD19+ Nalm6 CD19- Molm14. b, Representative flow cytometry plot TCR constant chain expression after dual TRAC TRBC disruption. c, Summary data surface disruption n=8 individual donors. Analysis using two-sided Student’s t-test.</p>
<p>Supplementary Figure 2 | a-d, Expression of a, TCR constant chains and mCherry transduction marker, b, FMC63 c, d, CD3ε in primary human T cells engineered with CARs or miTCRs. e, Comparison surface expression miTCR1 miTCR2 n=6 independent donors. f, Median fluorescence intensity (MFI) APC on that bound soluble CD19-APC conjugate. Representative five Analysis using Student’s t-test two-way ANOVA.</p>
<p>Supplementary Figure 8 | a, Cytotoxicity, as measured by fold change in Nalm6 growth during co-culture, and b, expansion of human T cells expressing transgenic TCR 1G4, 19/BBζ or 19/28ζ CAR against Nalm6WT Nalm6triple. c-d, Endogenous expression 41BBL, CD80 CD86 c, Raji d, K562 cell lines. e, Growth rate engineered f-g, Cytotoxicity mut035 over time f, acute stimulation g, re-stimulation. were initially co-cultured with re-stimulated Nalm6WT; Nalm641BBL Nalm6WT. h, Percent either...
<p>Supplementary Figure 4 | Names and amino acid changes of miTCR variant panel. PD insertions are highlighted in blue.</p>
<p>Supplementary Figure 3 | a, Predicted structures of miTCR1 and miTCR2 α β chains overlaid on resolved TCR chains. b, Overlay five highest-ranking predictions for variable-constant chain interface. c, Resolved native V-C interface structure predicted WT modified d, WT, chains.</p>