CAR-NK: Human's new weapon against tumor

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Click on the picture and sign up for the conference on November 6, 2021/YiMike News eMedClub News/--Human natural killer cells (NK) account for all circulating lymphocytes 15%
of
NK cells were discovered in the 1970s and are mainly related to killing infected microorganisms and malignantly transformed allogeneic and autologous cells
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NK cells exhibit anti-tumor cytotoxicity without prior sensitization and production of cytokines and chemokines that regulate various immune responses
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NK cells are innate lymphocytes with cytotoxic effects and are involved in initiating adaptive immune responses
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Once activated, NK cells secrete pro-inflammatory cytokines and trigger perforin/granzyme-induced lysis of target cells
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They can precisely attack and eliminate abnormal cells, and at the same time can regulate the body's immune status and immune function, indicating that NK cells have the potential to act as "anti-cancer guardians"
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▲ NK cell-mediated tumor cytotoxicity mechanism Chimeric antigen receptor (CAR) is a receptor protein that gives immune cells new capabilities to target specific antigen proteins
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Its extracellular antigen binding domain is usually a single-chain antibody (scFv), which can recognize specific antigens on the surface of tumor cells
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In recent years, the application of chimeric antigen receptor modified T cells (CAR-T) in the treatment of hematological tumors has been successful and has become a clinical hot spot in tumor immunotherapy
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However, their wide application is limited by inherent risks, such as graft-versus-host disease (GvHD) and long-term infections that limit the production of CAR-T cells
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In addition, most CAR-T cell therapy studies use autologous T cells, mainly to avoid the risk of GVHD (graft versus host disease) caused by autologous cells recognizing allogeneic T cells
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However, studies have shown that injecting a single source of cord blood NK cells into many patients with hematological tumors and solid malignancies will not cause GVHD
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The safety of NK cells and their anti-tumor potential make them a promising cell type for CAR technology, which can redirect their cytotoxic potential to specific targets
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This gives CAR-NK cell therapy the potential to become a universal product
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These products may be safer than CAR-T cell therapy
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At present, the research of CAR-NK cell immunotherapy has become the direction of choice for more and more researchers
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Recommended reading: Merger of two Korean companies: Owning the CAR-NK favored by Merck and T cell products that have been on the market for 15 years Yimai Meng explosives financing Artiva raises US$120 million in financing to promote allogeneic NK and CAR-NK cell therapies The potential advantages of developing CAR-NK products.
Up to now, 5 CAR-T products have been approved by the US FDA for the treatment of blood cancers
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In June 2021, China also ushered in the first approved CAR-T product
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However, CAR-T products in clinical applications have challenges such as long production cycles, cytokine storms, and neurotoxic side effects, which limit their wide range of applications.

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In addition, CAR-T products have not yet made a breakthrough in the field of solid tumors
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CAR-NK products developed based on NK cells are considered to have the potential to solve the above challenges
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▲ The production process of CAR-NK cells (picture source: Reference 1) According to a review published in the "Lancet" sub-Journal EBioMedicine, CAR-NK cell therapy products have the following potential advantages: better safety First of all, CAR-NK cell immunotherapy has better safety than CAR-T cell immunotherapy
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One of the main side effects of CAR-T cell immunotherapy is the targeting effect due to the continuous existence of CAR-T cells
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On the contrary, CAR-NK cells have a short lifespan and hardly produce targeting effects
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Secondly, allogeneic NK cell infusion is well tolerated and will not cause GVHD and obvious toxicity
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Therefore, NK cells are a more adaptable CAR carrier, not just autologous cells
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In addition, CAR-NK products are also expected to reduce the cytokine release syndrome (CRS) and neurotoxic side effects that occur in CAR-T therapy.
This is because the types of cytokines produced by NK cells are very different from those produced by T lymphocytes.
Big difference
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CAR-T cells usually induce cytokine storm by secreting pro-inflammatory cytokines such as TNF-α, IL-1 and IL-6
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The activated NK cells produce IFN-γ and granulocyte-macrophage colony stimulating factor (GM-CSF)
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Still have natural anti-tumor cell toxicity.
Second, CAR-NK cells still have natural anti-tumor cell toxicity.
In addition to recognizing tumor surface antigens by single-chain antibodies to inhibit cancer cells, NK cells can also pass multiple receptors.
Recognize various ligands to inhibit cancer cells, such as natural cytotoxic receptors (NKp46, NKp44 and NKp30), NKG2D and DNAM-1 (CD226)
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These NK cell receptors generally recognize stress-inducing ligands expressed on tumor cells under the stress of immune cells or long-term treatment
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In addition, NK cells can eliminate tumor cells through the CD16-mediated ADCC mechanism
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Therefore, CAR-NK cells can effectively eliminate tumor antigen-positive cancer cells or cancer cells expressing NK cell receptor ligands through various mechanisms of action that are CAR-dependent and NK cell receptor-dependent
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Clinical trials have shown that CAR-T cells cannot eliminate highly heterogeneous cancer cells, but CAR-NK cells can effectively kill residual tumor cells that may change their phenotype after long-term treatment
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Has the potential to become a "universal" product Third, CAR-NK therapy has the potential to become a "universal" product
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Since NK cells are very abundant in clinical samples, they can be produced from peripheral blood (PB), umbilical cord blood (UCB), human embryonic stem cells (HESC), induced pluripotent stem cells (IPSC) and even NK-92 cell lines, plus Allogeneic NK cells are safer, so CAR-NK cells are expected to provide a "universal" product to solve the problems that plague the current long production cycle and individualized needs of CAR-T products
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Current status of CAR-NK cell therapy The CAR-NK products designed and developed based on the CAR-T strategy are designed to target different tumors so that NK cells can specifically kill tumor cells, and different designs are designed for hematomas and solid tumors.
CAR-NK cell products have been developed for different hematomas, such as NK92-CD19-CD3ζ for B-line malignant tumors and CD138-CAR-NK-92 for multiple myeloma
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▲Pre-clinical studies of CAR-NK cells in hematoma.
In addition, different CAR-NK products have been developed for different solid tumors, such as EGFR-CD28-CD3ζ for glioblastoma and NKG2D-DAP10-CD3ζ for bones Carcinoma, hepatocellular carcinoma (HCC), pancreatic cancer and breast cancer, etc.
, these differently modified CAR-NK cells have shown good effects in anti-tumor
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▲ Pre-clinical studies of CAR-NK cells in solid tumors According to the information published on the ClinicalTrials.
gov website, there are currently many clinical trials of CAR-NK products in progress, most of which are in phase 1/2 clinical trials
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From the target point of view, and CAR-T is similar to the majority of CAR-NK cell therapy products targeted are common malignant blood cancer targets, such as CD19, CD22, BCMA, CD33 and so on
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Some CAR-NK targeted product is a solid tumor-associated antigens, such as PSMA, mesothelin, ROBO1, NKG2D the like
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The potential advantages of CAR-NK products have been initially verified in preclinical studies and some clinical studies
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A study published in the New England Journal of Medicine (NEJM) in 2020 showed that 11 cancer patients received a CAR-NK product that targets CD19, and 7 patients experienced complete remission (CR), and all There are no common side effects such as cytokine release syndrome (CRS) and neurotoxicity in the treatment of CAR-T products
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"Nature-Medical" also listed this progress as one of the ten notable developments in the field of biomedicine in 2020
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▲ Part of the clinical trials of CAR-NK in the treatment of solid tumors.
CAR-NK therapy attempts to break through the challenge of solid tumors.
One of the main obstacles to the challenge of CAR treatment is the lack of curative effect on solid tumors
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This is due to poor tumor perfusion, TAA heterogeneity, and the immunosuppressive tumor microenvironment (TME) associated with solid tumors
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▲ NK cell dysfunction in the tumor microenvironment (TME) (picture source: reference 1) CAR-NK therapy attempts to solve these problems, and some clinical trials have been applied to the treatment of solid malignant tumors
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The recent preclinical and clinical studies support the view that CAR-NK cell therapy can exert expected effects in human hematology and solid malignant tumors, which means that they have a wide range of clinical applications
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This view emphasizes the development of CAR-NK cell therapy as an innovative strategy in the context of cancer immunotherapy, and briefly outlines the latest findings of CAR-NK cell therapy in human malignancies, with an emphasis on preclinical research
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EGF receptor (EGFR) is a well-known tyrosine kinase receptor and is still at the forefront of tumor targeted therapy
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In addition, the report shows that EGFR-specific CAR-NK-92 cells and primary NK cells exhibit significant cytotoxicity and IFN-in vitro exposure to breast cancer cell lines MDA-MB-231 and MDA-MB-468.
γ produces and MCF-7, and EGFR specific CAR-NK-92 shows a better therapeutic effect compared with primary NK cells
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In a xenograft model of breast tumors, intratumoral injection of EGFR-CAR-NK-92 cells or oncolytic herpes simplex virus 1 (oHSV-1) can prevent tumor development, and at the same time, it can be combined with EGFR-CAR-NK-92 to treat cells and oHSV-1, leading to more pronounced elimination of tumor cells and prolonged survival time of transplanted mice
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In addition, administration of EGFR-CAR-NK-92 cells resulted in the regression of triple negative breast cancer (TNBC) in mouse models of breast cancer cell line-derived xenografts (CLDX) and patient-derived xenografts (PDX)
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Similarly, in an orthotopic glioblastoma (GB) xenograft mouse model, intracranial injection of EGFR-CAR-NK-92 cells strongly inhibited tumor development, thereby prolonging the overall survival rate of tumor-bearing mice , Introducing redirected NK cells into an effective treatment strategy
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Other studies have shown that further genetic modification of these EGFR-CAR-NK cells with the chemokine receptor CXCR4 can increase the chemotaxis of U87-MG glioblastoma cells produced by stromal cell-derived factor 1 (SDF1) in vitro.
Compared with EGFR-CAR-NK cell therapy, it can promote the survival of xenotransplanted mice
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In contrast, the specific targeting of folate receptor α (αFR) overexpressed in 90% of ovarian cancers, by using αFR-specific CAR-NK-92 cells, resulted in αFR+ tumor cells in vitro and in mouse xenografts.
Selective lysis of ovarian cancer model
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In addition, comparative analysis shows that the third generation αFR-CAR-NK-92 cells are superior to the first and second generation CARs in terms of effective cytotoxicity to αFR+ tumor cells
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In addition, mesothelin-specific CAR-NK cells can selectively eliminate mesothelin-positive ovarian cancer cells (OVCAR-3 and SK-OV-3), and they have an effect on mesothelin-negative cells (SK-HEP- 1) No response
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More importantly, redirected NK cells can effectively eradicate ovarian cancer cells in both subcutaneous and intraperitoneal tumor models, which indicates that besides αFR, mesothelin may be a viable target for targeted therapy of ovarian cancer
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Recently, other research results indicate that the co-expression of chemokine receptors CXCR1 and CAR may provide an innovative strategy to improve the therapeutic effect of NK cells in human ovarian tumors by enhancing tumor infiltration of effector immune cells
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For example, in a xenotransplantation model, NK cells that have been genetically modified to express CXCR1 and a CAR targeting NKG2D ligand have a better ability to migrate tumor cells in vivo
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In addition, the cytotoxicity exerted by CAR-NK cells is not negatively affected by the expression of the CXCR1 transgene, and the improved tumor trafficking of the administered cells leads to an enhanced anti-tumor response in xenografts
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Compared with parental NK cells, ganglioside GD2-specific CAR-NK cells induce significantly higher cytotoxicity to GD2+neuroblastoma (NB) cells
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In addition to the significant transfer of GD2-CAR-NK cells to the 3D tumor spheres, resulting in a decrease in growth rate, as demonstrated by live cell imaging analysis, GD2-CAR-NK treatment inhibited tumor development in mice
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In addition, further analysis showed that redirection of NK cells and histone deacetylase inhibitor (HDACi) or programmed cell death protein 1 (PD-1)/programmed cell death ligand 1 (PD-L1) block Combination therapy of agents may be more effective in anti-cancer function
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CAR-NK cells are currently considered to be a reliable immunotherapy modality for the treatment of hepatocellular carcinoma (HCC)
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Therefore, glypican-3 (GPC3)-specific CAR-NK-92 cells co-cultured with GPC3+HCC cells cause significant in vitro cytotoxicity and cytokine release, and are also used in HCC xenografts with high and low GPC3 expression Stimulates the level of anti-tumor effects in the body, but not in the absence of GPC3 expression
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Correspondingly, in GPC3+HCC xenografts, tumor development is eliminated and tumor cell apoptosis is enhanced
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Regarding the ability of TGF-β to inhibit the function of NK cells, the researchers genetically engineered NK-92 cells to express a chimeric receptor, in which the extracellular and transmembrane domains of the TGF-β type II receptor are the same as those of NKG2D.
The inner domain is accompanied by a TN chimeric receptor
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They found that NK-92 cells expressing TN have significant resistance to inhibitory signal transduction induced by TGF-β, and also stimulated higher killing ability and IFN-γ production of HCC cells in vitro
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In addition, TN-expressing NK-92 cells significantly metastasized to TGF-β-expressing malignant tumors, and inhibited tumor progression in the HCC xenograft mouse model, indicating that TN chimeric receptors can be used to improve the anti-tumor efficacy of NK cells.
Therapy
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Summary NK cells have unique anti-tumor effects, cytotoxicity not restricted by MHC, cytokine production and immune memory functions, making them a key role in the innate and adaptive immune response system
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Compared with CAR-T cells, CAR-NK cells have their own unique advantages, but they still face some challenges
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These challenges include improving cell proliferation, making the activation of cytotoxicity more effective, and finally finding the best way to rebuild NK cells
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I believe that solving these problems, based on the excellent anti-tumor pedigree of NK cells, is very likely to bring new breakthroughs in tumor treatment under the arm of CAR modification
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It is believed that CAR-NK cell therapy may lead to revolutionary progress in tumor immunotherapy
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Reference materials: 1.
https://onlinelibrary.
wiley.
com/doi/10.
1111/cas.
149932.
https://zhuanlan.
zhihu.
com/p/3079308623.
https://