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Latest News: IL-4 Drives Exhaustion of CD8+ CART Cells
2025-01-16
On September 12, 2024, Carli M. Stewart et al. published an article in Nature Communications on IL-4 drives exhaust of CD8+CART cells, mainly for CAR-T cell therapy. Due to the involvement of CAR-T cell failure, although the success rate of treating blood cancer is high, the duration is short, and most patients experience recurrence within one year Cell exhaustion is the main cause of CAR-T cell failure, caused by chronic stimulation through T cell receptors (TCR) in CD8+T cells or CAR in CAR-T cells. Phenotypically, it leads to upregulation of various inhibitory receptors, such as programmed cell death protein 1 (PD-1), T cell immunoglobulin and mucin domain protein 3 (TIM-3), cytotoxic T lymphocyte associated protein 4 (CTLA-4), and lymphocyte activation gene 3 (LAG-3). Functionally, it slows down cell proliferation, reduces effector cell capacity, and causes metabolic changes. Although genetic engineering optimization has been studied, the mechanism and potential therapeutic targets have not been elucidated yet. Studies have shown that CD28 Co stimulation is more likely to cause CAR-T cell failure. Researchers mainly studied cell models targeting CD19 antigens containing the CD28 co stimulatory domain, and identified IL-4 as a key regulatory factor in CAR-T cell failure. Subsequently, IL-4 neutralization was proposed as a strategy to enhance the anti-tumor activity of CAR-T cells through in vivo and in vitro validation. The combination of IL-4mAb and CAR-T cell therapy can reduce T cell exhaustion and enhance anti-tumor activity.
Detailed content introductionINTRODUCTION
CAR-T cell therapy
CAR T cell therapy (chimeric antigen receptor T cell therapy) is an immunotherapy used to treat certain types of cancer, especially leukemia and lymphoma, by genetically engineering one's own T cells. Collect immune related T cells from the blood and genetically engineer them to express chimeric antigen receptors (CARs) on their surface. CAR consists of two parts: an antigen recognition domain (which can recognize antigens on the surface of cancer cells) and a signaling domain (which promotes T cell proliferation when T cells bind to targets). CAR-T cell therapy can be used for precise targeted therapy to reduce damage to normal cells.
Fig 1: CAR-T Cell therapy. (Figure source: CAR T-cell Therapy: A New Era in Cancer Immunotherapy.)
CD28
CD28 can be expressed on T cells and has a stimulating effect on T cell activation and survival. Generally, CD28 is expressed on about 50% of CD8+T cells and over 80% of CD4+T cells in humans. CD28 is highly homologous to CTLA4 and competes for CD80 and CD86 antibodies, making it a drug target. Binding to anti-CD28 can lead to the release of IL-2 and IL-4. We offer products related to CD28 targets:
| Catalog Number | Product Name | Size |
|---|---|---|
| ALP64650 | ALP64650-Recombinant Human CD28 Protein, Fc Tag | 50ug,100ug,500ug |
| ALP64644 | ALP64644-Recombinant Mouse CD28 Protein, Fc Tag, His Tag | 50ug,100ug,500ug |
| ALP64560 | ALP64560-Recombinant Human CD28 Protein, mFc Tag | 50ug,100ug,500ug |
| ALP64537 | ALP64537-Recombinant Human CD28 Protein, His Tag | 50ug,100ug,500ug |
| ALP64506 | ALP64506-Recombinant Human Biotinylated CD28 Protein, Fc Tag, Avi Tag | 50ug,100ug,500ug |
| ADT1043 | ADT1043-Anti-CD28 Biosimilar– Anti-CD28(Tp44) mAb – Research Grade | 1mg,5mg |
| ADT1374 | ADT1374-Lulizumab Biosimilar-Anti-CD28 mAb – Research Grade | 1mg,5mg |
IL-4
The interleukin 4 (IL-4, IL-4) is a cytokine, which can stimulate the differentiation of B cells, the proliferation of B cells and T cells, and is an immune regulator in the immune response. It can also reduce inflammation, promote wound repair and healing. According to research, IL-4 can also promote the mitosis, dedifferentiation and metastasis of rhabdomyosarcoma. IL-4 related diseases include: allergic diseases (asthma or allergic reaction), can promote tumor growth (breast cancer, prostate cancer, lung cancer, etc.), affect the nervous system, HIV diseases, etc. According to research, the level of IL-4 will change in these diseases.
The technology involvedTECHNOLOGY
CRISPR Cas9
CRISPR Cas9 refers to clustered regularly spaced short palindromic repeat sequences (CRISPR), a system for efficient gene editing guided by a segment of RNA for Cas9 nucleases. It mainly involves two basic components: a guide RNA that matches the target DNA and Cas9 (CRISPR related protein 9) - a nuclease that can cleave two strands of DNA. The combination of guide RNA and Cas9 forms a single strand, called synthetic one-way guide RNA (sgRNA). The protospacer adjacent motif (PAM) is located at the 3 'end of the guide RNA for easy cleavage. After cleavage, two DNA repair pathways are selected (non homologous end connection, usually leading to DNA damage). Random insertion/deletion, or homologous directed repair, where homologous DNA fragments are used as repair templates, is one of the methods used for repair, CRISPR/Cas9 It can be used to treat genetic diseases caused by single gene mutations such as cystic fibrosis (CF) and infectious diseases such as HIV. During the treatment process, attention should be paid to the mop effect and targeted delivery mode of CRISPR/Cas9.
Fig 2: The CRISPR/Cas9 system. (Figure source: What is CRISPR/Cas9?)
RNA Sequencing
RNA sequencing (RNA Seq) is a new generation sequencing method that can reveal the presence and quantity of RNA in biological samples, as well as analyze the types of RNA (total RNA, small RNA (such as miRNA), tRNA, and ribosomal RNA). The sequencing process includes: (1) library preparation: RNA isolation, isolation and extraction from tissues, adding DNA enzymes to RNA to reduce the original DNA in tissues, reverse transcribing RNA into cDNA, using enzymes, ultrasound treatment and other methods to fragment cDNA, and quantifying sequencing analysis. It can also include sequencing types such as Complex DNA sequencing (cDNA Seq), Small RNA/non coding RNA sequencing, Direct RNA sequencing, Single molecule real-time RNA sequencing, Single cell RNA sequencing (scRNA Seq), etc.
Fig 3: Typical RNA-Seq experimental workflow. (Figure source: Informatics for RNA Sequencing: A Web Resource for Analysis on the Cloud.)
Flow Cytometry
Flow cytometry physically separates and purifies cells based on the optical properties of the cell surface. It has a collection system that uses parameters including cell size, morphology, and protein expression to detect cells, and then uses droplet sorting technology for cell sorting and recovery for cell culture, operation, research, and other experiments. The operation process: the sample is injected into a sheath fluid flow, which carries cells through a vibrating nozzle to generate droplets containing individual cells. A charged ring is placed at the separation location, and opposite charges are captured on the droplets to form charged droplets. The charged droplets fall into an electrostatic deflection system and are transferred to a container. After droplet separation, the droplet flow returns to neutrality. Flow cytometry can be applied to cell sorting, Cell counting, determination of cell characteristics and functions, biomarker detection, and other aspects.
Fig 4: Cell Sorting Using Flow Cytometry and Droplet Technology. (Figure source: Development of an in vitro model system for studying the interaction of Equus caballus IgE with its high- affinity FcεRI receptor (PhD thesis))
ReferenceREFERENCE
[1] Stewart, C.M., Siegler, E.L., Sakemura, R.L. et al. IL-4 drives exhaustion of CD8+ CART cells. Nat Commun 15, 7921 (2024). https://doi.org/10.1038/s41467-024-51978-3
[2] Androulla N. Miliotou, Lefkothea C. Papadopoulou, CAR T-cell Therapy: A New Era in Cancer Immunotherapy, Current Pharmaceutical Biotechnology; Volume 19, Issue 1, Year 2018, . DOI: 10.2174/1389201019666180418095526
[3] Ran, F., Hsu, P., Wright, J. et al. Genome engineering using the CRISPR-Cas9 system. Nat Protoc 8, 2281–2308 (2013).
[4] https://doi.org/10.1038/nprot.2013.143
[5] Redman M, King A, Watson C, King D. What is CRISPR/Cas9? Arch Dis Child Educ Pract Ed. 2016 Aug;101(4):213-5. doi: 10.1136/archdischild-2016-310459. Epub 2016 Apr 8. PMID: 27059283; PMCID: PMC4975809.
[6] https://en.wikipedia.org/wiki/RNA-Seq
[7] Malachi Griffith, Jason R. Walker, Nicholas C. Spies, Benjamin J. Ainscough, Obi L. Griffith - http://journals.plos.org/ploscompbiol/article?id=10.1371/journal.pcbi.1004393, CC BY 2.5, https://commons.wikimedia.org/w/index.php?curid=53055894
[8] Development of an in vitro model system for studying the interaction of Equus caballus IgE with its high- affinity FcεRI receptor (PhD thesis), The University of Sheffield, CC BY-SA 3.0, https://commons.wikimedia.org/w/index.php?curid=18139883