CAR-macrophages: A new candidate for solid tumor immunotherapy

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CAR-macrophages: A new candidate for solid tumor immunotherapy

CAR-macrophages: A new candidate for solid tumor immunotherapy.  CAR-T cell therapy is a type of cellular immunotherapy, which refers to the transfer of genetic material with specific antigen recognition domains and T cell activation signals to T cells through gene cloning technology, so that the T cells and tumor cells can have specific antigens on the surface Combined with the direct activation method.

CAR-T cells have been successfully used to treat malignant tumors of the circulatory system, such as B cell-derived malignancies. After successful CAR-T treatment, people applied CAR strategy to NK cells and developed CAR-NK cell therapy similar to CAR-T treatment.

The two main cell types of the mononuclear phagocyte system (MPS) are monocytes and monocyte-derived macrophages, both of which have a phagocytic function. MPS selectively devours external targets such as microorganisms, however, macrophages cannot recognize and attack tumors. Even in tumor tissues, macrophages will not play a role in phagocytosis or antigen presentation, but will transform into immunosuppressive M2 type, preventing the immune system from clearing tumor cells.

In addition, in addition to the positive signal of macrophage phagocytosis, there are opposite signals to prevent macrophages from phagocytosis of target tumor cells, such as CD47. These signals have become a hot research area since they were discovered.

In view of the current success of CAR-T cell therapy and the development potential of CAR-NK cells, researchers have shown great interest in the development of CAR macrophages (CAR-M) for tumor immunotherapy. The emergence of CAR-M has opened up new possibilities for the treatment of solid tumors: using specific CARs to modify human macrophages to improve the phagocytic activity and antigen presentation of macrophages to tumors.

Research progress of CAR-M

At present, many attempts have been made to use CAR-M to treat cancer. Researchers have designed chimeric antigen receptor phagocytic cells (CAR-P), which can guide macrophages to phagocytize specific target cells.

Studies have shown that CAR-P expressing the intracellular domain of MegflO or FcRv can promote the phagocytosis of the target antigen. CAR-PMegf10 can specifically trigger the phagocytosis of targeted ligands, and initiate phagocytosis through the local signal cascade of tyrosine phosphorylation. TCR-CD3ζ may promote the phagocytosis of CAR-P by recruiting syk kinase.

Whole-cell phagocytosis is relatively rare, and target cells are gnawed more frequently, indicating that the interaction between CAR-P macrophages and target cells is not sufficient to trigger direct whole-cell phagocytosis. Studies have found that the PI3K signal plays an important role in the endocytosis of large targets and can promote the phagocytosis of macrophages. The researchers linked the pI3K p85 subunit with CAR-P-FcRv to form a “tandem” CAR (CAR-Ptandem). CAR-Ptandem has good whole-cell phagocytosis.

Researchers at the University of Pennsylvania used an anti-HER2 CAR-M containing the intracellular domain of CD3-ζ. In two heterotopic mouse models of solid tumor transplantation, a single injection of anti-HER2-CAR-M can reduce tumor burden and prolong the survival time of mice. It was also found in the humanized mouse model that HER2-CAR-M can transform M2 macrophages into M1 macrophages, induce the inflammatory tumor microenvironment, and enhance the anti-tumor cytotoxicity of T cells. In addition, it was also found that HER2-CAR-M may produce epitope spread, which provides a new idea for avoiding tumor immune escape.

Zhang et al. used induced pluripotent stem cells (iPSCs) to express the CAR structure and differentiate into macrophages, named CAR-iMac. Studies have shown that iPSCs expressing CAR can differentiate into macrophage-like cells. In the absence of antigen, CAR-iMac is closer to the M2 polarization state. However, in the presence of specific antigens such as CD19, CAR-mediated signals promote the phagocytosis of CAR-iMac and lead to the conversion of CAR-iMac to the pro-inflammatory M1 type. Macrophages derived from iPSC may become an important cell source for myeloid tumor immunotherapy.

An important reason why CAR-T is ineffective in treating solid tumors is that it is difficult for T cells to enter the tumor tissue. This is because the physical barrier formed by the extracellular matrix (ECM) of solid tumors prevents T cells from entering the tumor tissue. ECM is produced by highly ordered fiber molecules, glycoproteins and other macromolecules. Its synthesis and degradation are mainly regulated by matrix metalloproteinases (MMPs) and tissue inhibitors of metalloproteinases (TIMPs). Macrophages are an important source of MMPs. .

The researchers designed a CAR-147 structure, which consists of a single-chain antibody targeting human HER2, the hinge region of IghG1, and the transmembrane and intracellular regions of the mouse CD147 molecule. After CAR-147 is co-cultured with HER2+ human breast cancer cells, it induces the expression of multiple MMPs in CAR-147 macrophages, which proves that CAR-147 can specifically recognize HER2 antigen and effectively activate the expression of MMPs in macrophages. CAR-147 macrophages do not inhibit tumor cell proliferation in vitro, but intravenous injection of CAR-147 macrophages can significantly inhibit tumor growth in 4T1 breast cancer mouse models. At the same time, it was found that the proportion of T cells in tumors treated with CAR-147 macrophages was much higher than that in tumors treated with control macrophages, indicating that CAR-147 macrophages can destroy tumor extracellular matrix and promote T cell infiltration .

The clinical application of CAR-M in the treatment of solid tumors

Until November 2020, two clinical trials based on the CAR-M strategy have been approved by the FDA. The first is CT-0508, a drug candidate from CARISMA Therapeutics, which uses anti-HER2 CAR-M to treat patients with relapsed/refractory HER2 overexpression tumors (Phase I clinical trial). The other is Maxyte’s MCY-M11, which uses mRNA to transfect PBMC to express CAR (including CAR-M) targeting mesothelin to treat patients with relapsed/refractory ovarian cancer and peritoneal mesothelioma. It is currently recruiting volunteers Conduct Phase I clinical trials.

Regarding the clinical transformation of CAR-M, there are several aspects that need attention: First, the safety and effectiveness of CAR-M are the basis of the therapy. Although it has been verified by animal experiments, the effect of CAR-M in humans is Safety and effectiveness still need to be verified; secondly, reliable cell source and expansion are necessary conditions for the clinical application of CAR-M, which can be prepared from PBMC or iPSCs. In addition, unlike T cells, macrophages have a lower risk of GVHD, which means that products can be produced in advance for patients to use on demand. Third, there is another problem that must be considered. At present, CAR-M mostly uses viral transfection methods, which may induce insertion mutations. CRISPR/Cas9 provides new possibilities to solve this problem, and it only takes one week to complete Editing of CAR-T gene.

Advantages and challenges of CAR-M

Similar to CAR-T and CAR-NK cells, CAR-M cells are composed of extracellular signaling domains, transmembrane domains, and intracellular activation signaling domains that recognize specific tumor antigens. At present, the research on the extracellular signal domain mainly focuses on several common tumor targets, such as CD19 and HER2.

Unlike CAR-T cells, CAR-M cells have the following three advantages.

1) Due to the physical barrier formed by the matrix around the tumor cells, T cells cannot enter the tumor environment, while macrophages can significantly infiltrate the tumor environment. TAM plays an important role in tumor invasion, metastasis, immunosuppression and angiogenesis. CAR-M can reduce the ratio of TAM, affect the cell phenotype of TAM, and have a positive effect on the treatment of tumors.

2) In addition to the effect of phagocytosing tumor cells, CAR-M also has the effect of promoting antigen presentation and enhancing T cell killing.

3) Compared with CAR-T, CAR-M has limited circulation time and less non-tumor targeted toxicity.

Although CAR-M has great potential to become a powerful tumor immunotherapy method, many problems need to be overcome to achieve the desired effect. The first is the limitation of the number of cells: macrophages will not proliferate whether they are injected in vitro or in vivo. Patients can only receive a limited number of macrophages, which may affect the treatment effect.

The second is related to the migration characteristics of macrophages in the body. After injection, the exogenous macrophages pass through the lungs, and then most of them stay in the liver, which is not conducive to cancer treatment. The third is the complex tumor microenvironment. Although CAR-M has achieved good results in mouse models, the actual tumor microenvironment of humans is much more complicated than that of animal models.

Finally, due to the high degree of heterogeneity of tumor cells, the expression of the target antigen may not be sufficient. This problem has always been very prominent in CAR-T treatment. Clinical studies have found that most tumor cells removed by CAR-T cells have high levels of target antigen expression. It is foreseeable that this will also become a major obstacle to the development of CAR-M therapy.

Summary

CAR-M therapy has shown its effective anti-tumor ability in animal experiments. Compared with CAR-T and CAR-NK, CAR-M as a new cellular immunotherapy has its unique advantages, but there are also many shortcomings that need to be overcome. Therefore, care should be taken to maximize the effectiveness and safety of CAR-M in future clinical treatment. We have reason to believe that with the further development of science and technology, CAR-M will soon show curative effects on patients, adding further help to tumor immunotherapy!

(source:internet, reference only)

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