CAR-T Characterization Panel

 

Measure eight essential components of CAR-T biology with 780 human genes.  Ideally suited for new CAR-T development and QC, the CAR-T Characterization Gene Expression Panel can help streamline the CAR-T manufacturing workflow and potentially reduce vein to vein time. Confidently profile CAR-T products for research and manufacturing applications with a robust, automated, and reproducible assay.  Empower your knowledge of CAR-T with the most up-to-date biology developed with experts in the field. 

  • Optimize CAR-T method development
  • Create manufacturing acceptance criteria
  • Measure metabolic fitness and persistence
  • Monitor post-infusion exhaustion and toxicity
CAR-T characterization advanced analysis modules

CAR-T Therapy Workflow

Understanding each step of the CAR-T workflow is critical to ensuring quality and efficacy of the final CAR-T product.  The nCounter CAR-T Characterization Panel can be used throughout development and manufacturing as a standardized panel of genes for optimizing methods, developing manufacturing acceptance criteria as well as understanding the host influences beyond manufacturing.

CAR-T workflow

In addition to the standard nSolver™ Analysis, genes included in the CAR-T Characterization panel are organized and linked to various advanced analysis modules to allow for efficient exploration of the eight essential aspects of CAR-T biology.

 

Advanced Analysis Modules available for CAR-T Characterization:

  • TCR Diversity Score (coming soon)
  • Normalization
  • Quality Control
  • Pathway Analysis
  • Cell Profiling
  • Differential Expression
  • Gene Set Analysis
  • Built-in compatibility for Panel-Plus and Protein analysis

Essential CAR-T Biology Description Pathways and Processes
Phenotype Multiple subtypes of T-cells and the phenotypic changes that accompany them can be distinguished via the cytokines and pathways that maintain, promote, and modulate their activity. Notch, Wnt signaling, Tfh, TGF-beta, Th1, Th17, Th2, Th9, Treg, Innate-like T-cells, Vitamin A (RA) Signaling
Cell Types Other cell types can contaminate the population of active CAR-T cells. Similarly, measurement of B-cell populations may aid in assessment of whole blood post-infusion measurement of tumor burden in B-cell lymphomas. Immune cell profiling
TCR Diversity The TCR diversity of CAR-T cells can provide information on the number of clones present after leukapheresis, manufacturing, and infusion. TCR Content
Activation T-cell activation is primarily mediated by antigens presented to the TCR complex and modulated by costimulatory molecules. Downstream of the TCR, multiple pathways induce transcriptional changes that lead to the production of chemokines and cytokines. Cell surface receptors signal a change in phenotype. Chemokine Signaling, Costimulatory Molecules, Interleukin Signaling, TCR signaling, JAK-STAT, MAPK and PI3K Signaling, Myc targets, NFAT, Antigen processing & presentation, T-cell activation markers
Metabolism Fundamental changes in T-cell metabolism are induced upon activation to enable rapid cell division and thus expansion of relevant clones. These changes can be observed across basic metabolic pathways, including carbohydrate and fatty acid metabolism.

Glycolysis, Mitochondrial biogenesis, Fatty Acid Metabolism

Glutamine metabolism, Circadian Clock, One-carbon metabolism, Oxidative phosphorylation, mTOR, Cell Cycle, Autophagy
Persistence Ongoing presence of a T-cell and cytotoxic T-cell population can be measured by via cell type profiling. By measuring molecules involved in T-cell migration we can assess the ability of T-cells to home in on their target antigens. T-cell migration, T-cell cell type profiling
Exhaustion T-cell exhaustion can be induced by costimulatory molecules, other cell-cell interactions, and cell death via apoptosis. T-cell exhaustion markers, Apoptosis, Interactions with Non-Lymphoid Cells, Costimulatory Molecules
Toxicity Toxicity is correlated with certain cytokine and chemokine profiles. These signals can induce a pro-inflammatory environment in various tissues and lead to off-target toxicities of CAR-T treatment.

NK cell cytotoxicity, NKT Receptors, NF-kB, Type I interferon signaling

Type II interferon signaling, Interleukin signaling, Chemokine signaling

 

Chain Type Constant Chains Variable Chains
Alpha  TRAC

TRAV_
45 probes, 46 genes
TRAV8-2 and 8-4 covered by 1 probe

Beta TRBC1/2

TRBV_
46 probes, 48 genes
TRBV6-3 and 6-2 covered by 1 probe
TRBV12-4 and 12-3 covered by 1 probe

Gamma TRGC1, TRCG2

TRGV_
5 probes, 6 genes
TRGV3 and 5 covered by 1 probe

Delta TRDC

TRDV_
3 probes, 3 genes

Immune Cell Markers

CD3D/E/G, CD4, CD8A/B, PTPRC (CD45), CD45R0, CD45RA, SELL (CD62L), CCR7, CD28, CD40LG, IL2RA (CD25), NCR1 (NKp46)

 

Cell Type Description Panel Genes
B Cells B cells are the primary mediators of the humoral immune response, bearing antigen-specific B cell receptors and producing antibodies that can enable the immune system to respond to a broad variety of antigens. B cells can also function as MHC class II antigen presenting cells to stimulate T cell immunity. BLK, CD19, CD20, TNFRSF17
T Cells T-cells mediate cell-based immunity by recognizing primarily peptide antigens displayed on MHC class I or class II and either producing cytokines or directly killing the presenting cell. CD3D, CD3E, CD3G, CD6, SH2D1A
TH1 CD4+ T cell subset that produces IL2 and Interferon-gamma to promote cellular immunity by acting on CD8+ T Cells, NK Cells and Macrophages. TBX21
Regulatory T Cells (Tregs) CD4+ T Cells that suppress effector B and T Cells and play a central role in suppression of the immune response and tolerance to self-antigens. FOXP3
CD8+ T Cells A subset of T cells that are capable of binding cognate-antigen expressing cells via class I MHC and directly lysing them via perforin and granzymes. CD8A, CD8B
Exhausted CD8+ T Cells T-cells overstimulated by antigen can develop an "exhausted" phenotype, in which they are no longer effective in targeting antigen-bearing cells. CD244, EOMES, CD223
Cytotoxic Cells All cells capable of cytotoxic activity, which can include T, NKT, and NK-cells. CTSW, GNLY, GZMA, GZMB, GZMH, CD161, CD94, KLRK1, PRF1
Dendritic Cells Professional antigen presenting cells that internalize, process, and present antigens to lymphocytes via MHC class I and class II along with costimulatory signals to initiate cellular immune responses. CCL13, CD209, HSD11B1
Macrophages Pluripotent cells with critical roles in initiating innate and adaptive immune responses, phagocytosing abnormal cells, and regulating wound healing and tissue repair. CD163, CD68, CD84
Mast Cells Mast cells release histamine containing granules and other signals in order to promote inflammation and regulate allergic responses. MS4A2, TPSAB1
Neutrophils Neutrophils are highly abundant cells that respond early to sites of infection or inflammation, phagocytose cellular debris, and promote downstream immunity. CSF3R, CD16, S100A12
Natural Killer (NK) Cells Cytotoxic cells of the innate immune system that are a significant source of interferon-gamma and are capable of directly killing targeted cells via detection of a loss in MHC surface expression. NKP46, XCL2
NK CD56dim cells The amount of CD56 present on an NK cell is indicative of its age and differentiation state; CD56 dim cells are mature NK cells, more commonly found in peripheral blood than secondary lymphoid tissues, and have the greatest cytolytic activity. IL21R, KIR2DL3, KIR3DL1, KIR3DL2

 

CAR-T Characterization Product Bulletin

CAR-T Characterization Panel Gene List (Human) - coming soon

Feature Specifications
Number of Targets 780 (Human), including internal reference genes
Sample Input - Standard (No amplification required) 25-300 ng
Sample Input - Low Input As little as 1 ng with nCounter Low Input Kit (sold separately)
Sample Type(s) PBMC, FFPE-derived RNA, total RNA, fragmented RNA, cell lysate, sorted cells, whole blood/plasma, synovial fluid
Customizable Add up to 30 unique genes with Panel-Plus and up to 10 custom protein targets
Time to Results Approximately 24 hours
Data Analysis nSolver™ Analysis Software (RUO)

 

Click here to view publications on CAR-T

Selected Panel References:

  1. Fraietta, JA et al. Disruption of TET2 Promotes the Therapeutic Efficacy of CD19-Targeted Cells. Nature. 2018;558(7709):307-12.
  2. Fraietta JA et al. Determinants of Response and Resistance to CD19 Chimeric Antigen Receptor (CAR) T Cell Therapy of Chronic Lymphocytic Leukemia. Nature Medicine. 2018;24(5):563-71.
  3. Raud B et al. Fatty Acid Metabolism in CD8+ T Cell Memory: Challenging Current Concepts. Immunological Reviews. 2018;283(1):213-31.
  4. Vargas TR and Apetoh L. The Secrets of T Cell Polarization. In: Zitvogel L and Kroemer G. (eds) Oncoimmunology. 2018:69-95.
  5. Xiao, X et al. Mucosal-Associated Invariant T Cells: New Insights into Antigen Recognition and Activation. Frontiers in Immunology. 2017;8:1540.
  6. Simone MD et al. Transcriptional Landscape of Human Tissue Lymphocytes Unveils Uniqueness of Tumor Infiltrating T Regulatory Cells. Immunity. 2016;45(5):1135-47.
  7. Buck, MD et al. T Cell Metabolism Drives Immunity. Journal of Experimental Medicine. 2015;212;(9):1345-60.
  8. Eberl G et al. Innate Lymphoid Cells: A New Paradigm in Immunology. Science. 2015;348(6237).
  9. Best JA et al. Transcriptional Insights into the CD8+ T Cell Response to Infection and Memory T Cell Formation. Nature Immunology. 2013;14(4):404-12.
  10. Chtanova T et al. Identification of T Cell-Restricted Genes, and Signatures for Different T Cell Responses, Using a Comprehensive Collection of Microarray Datasets. Journal of Immunology. 2005;175(12):7837-47.
Product Product Description Quantity Catalog Number
nCounter Human CAR-T Characterization Panel Includes 780 genes;10 internal reference genes for data normalization 12 Reactions XT-CSO-CART1-12
nCounter Human PanCancer IO 360 + Panel Standard Includes 770 genes; 20 internal reference genes for data normalization. Panel standard includes a pool of synthetic oligonucleotides that correspond to the target sequences in the panel 12 Reactions XT-CSPS-HIO360-12
nCounter Human PanCancer Immune Profiling Panel Includes 770 genes; 40 internal reference genes for data normalization 12 Reactions XT-CSO-HIP1-12

 

For Research Use Only. Not for use in diagnostic procedures.