Research status and challenges of CAR-macrophages
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Research status and challenges of CAR-macrophages
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Research status and challenges of CAR-macrophages.
CAR-T cell therapy is a kind of cellular immunotherapy, which refers to the transfer of genetic material with specific antigen recognition domains and T cell activation signals into T cells through gene cloning technology, so that T cells can interact with specific antigens on the surface of tumor cells Incorporate direct activation methods. CAR-T cells have been successfully used to treat circulatory malignancies, such as B cell-derived malignancies.
After the success of CAR-T therapy, people applied the CAR strategy to NK cells and developed a CAR-NK cell therapy similar to CAR-T therapy.
The two main cell types of the mononuclear phagocyte system ( MPS ) are monocytes and monocyte-derived macrophages, both of which have phagocytic functions.
MPs selectively engulf external targets such as microorganisms, whereas macrophages are unable to recognize and attack tumors.
Even within tumor tissue, macrophages do not play a role in phagocytosis or antigen presentation, but switch to the immunosuppressive M2 type, preventing the immune system from clearing tumor cells.
Furthermore, in addition to the positive signal for macrophage phagocytosis, there are opposite signals preventing macrophage phagocytosis of target tumor cells, such as CD47. These signals have been a hot area of research since their discovery.
In view of the current success of CAR-T cell therapy and the development potential of CAR-NK cells, researchers have generated 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: modifying human macrophages with specific CARs to improve the phagocytic activity and antigen presentation of macrophages to tumors.
Research progress of CAR-M
There have been multiple attempts to use CAR-M to treat cancer. The researchers engineered chimeric antigen receptor phagocytes ( CAR-P ), which guide macrophages to engulf specific target cells.
Studies have shown that CAR-P expressing the intracellular domain of Megf10 or FcRv can promote the phagocytosis of target antigens.
CAR-PMegf10 can specifically trigger the phagocytosis of targeting ligands and initiate phagocytosis through a local signaling cascade of tyrosine phosphorylation, and TCR-CD3ζ may promote the phagocytosis of CAR-P by recruiting syk kinase.
Whole-cell phagocytosis was relatively rare, while biting target cells was more frequent, suggesting that the interaction between CAR-P macrophages and target cells was not sufficient to trigger direct whole-cell phagocytosis.
It has been found that PI3K signaling plays an important role in the endocytosis of large targets and can promote the phagocytosis of macrophages.
The researchers linked the pI3K p85 subunit to CAR-P-FcRv to form a “tandem” CAR ( CAR-Ptandem ). CAR-Ptandem has a good whole-cell phagocytosis function.
Researchers at the University of Pennsylvania used an anti-HER2 CAR-M containing the CD3-ζ intracellular domain.
In two heterotopic mouse models transplanted with solid tumors, a single injection of anti-HER2-CAR-M reduced tumor burden and prolonged mouse survival.
It was also found in a humanized mouse model that HER2-CAR-M can convert M2 macrophages into M1 macrophages, induce an 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 diffusion, which provides a new idea for avoiding tumor immune escape.
Zhang et al. used induced pluripotent stem cells ( iPSCs ) to express CAR constructs and differentiate into macrophages, named CAR-iMac.
Studies have shown that CAR-expressing iPSCs can differentiate into macrophage-like cells, and 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 signaling promoted the phagocytosis of CAR-iMacs and resulted in the conversion of CAR-iMacs to the pro-inflammatory M1 phenotype.
iPSC-derived macrophages may become an important cell source for immunotherapy of myeloid tumors.
An important reason why CAR-T is ineffective in treating solid tumors is that it is difficult for T cells to enter tumor tissue.
This is because the physical barrier formed by the extracellular matrix ( ECM ) of solid tumors prevents T cells from entering tumor tissue.
ECM is produced by highly ordered fiber molecules, glycoproteins and other macromolecules, and its synthesis and degradation are mainly regulated by matrix metalloproteinases (MMPs ) and tissue inhibitors of metalloproteinases ( TIMPs ), and macrophages are an important source of MMPs .
The researchers designed a CAR-147 structure consisting of a single-chain antibody targeting human HER2, the hinge region of IgG1, and the transmembrane and intracellular regions of the mouse CD147 molecule.
After co-cultured with HER2+ human breast cancer cells, CAR-147 induced the expression of multiple MMPs in CAR-147 macrophages, proving that CAR-147 can specifically recognize HER2 antigen and effectively activate the expression of MMPs in macrophages.
CAR-147 macrophages did not inhibit tumor cell proliferation in vitro, but intravenous injection of CAR-147 macrophages significantly inhibited tumor growth in a mouse model of 4T1 breast cancer.
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 of tumors treated with control macrophages, indicating that CAR-147 macrophages can destroy the extracellular matrix of tumors and promote the infiltration of T cells into tumors .
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 drug candidate CT-0508 from CARISMA Therapeutics, which treats patients with relapsed/refractory HER2-overexpressing tumors with an anti-HER2 CAR-M (Phase I clinical trial ) .
The other is Maxyte’s MCY-M11, which uses mRNA to transfect PBMCs to express mesothelin-targeted CARs ( including CAR-M ) for the treatment of patients with relapsed/refractory ovarian cancer and peritoneal mesothelioma, and 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 this therapy.
Although it has been verified by animal experiments, the effect of CAR-M in humans Safety and efficacy still need to be verified; secondly, reliable cell source and expansion are necessary conditions for the clinical application of CAR-M, and CAR-M can be prepared from PBMC or iPSCs.
Furthermore, unlike T cells, macrophages have a lower risk of developing GVHD, which means that products can be manufactured ahead of time for patients to use on demand.
Third, there is another problem that must be considered. At present, CAR-M mostly uses virus transfection methods, which may induce insertion mutations. CRISPR/Cas9 provides a new possibility to solve this problem, and it only takes one week to complete Editing of CAR-T genes.
Advantages and challenges of CAR-M
Similar to CAR-T and CAR-NK cells, CAR-M cells consist of an extracellular signaling domain that recognizes a specific tumor antigen, a transmembrane region, and an intracellular activation signal region.
Currently, studies on extracellular signaling domains are mainly focused 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 stroma around tumor cells, T cells cannot enter the tumor environment, while macrophages can obviously infiltrate into the tumor environment. TAMs play important roles 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 function of phagocytosis of tumor cells, CAR-M also has the function of promoting antigen presentation and enhancing T cell killing.
3) Compared with CAR-T, CAR-M has limited circulation time and less non-tumor targeting 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 do not proliferate either in vitro or after injection in vivo. Patients can only receive a limited number of macrophages, which may affect the efficacy of the treatment.
The second is related to the migration characteristics of macrophages in vivo.
After injection, exogenous macrophages pass through the lungs, and most of them stay in the liver, which is not conducive to the treatment of cancer.
The third is the complex tumor microenvironment. Although CAR-M has achieved good results in mouse models, the actual tumor microenvironment in humans is much more complex than that in animal models.
Finally, due to the high heterogeneity of tumor cells, the expression of the target antigen may be insufficient. This problem has been very prominent in CAR-T therapy.
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.
CAR-M therapy has shown its effective anti-tumor ability in animal experiments.
Compared with CAR-T and CAR-NK, CAR-M has its unique advantages as a new cellular immunotherapy, but there are also many shortcomings that need to be overcome.
Therefore, attention should be paid to maximizing 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 effect in patients, adding another boost to tumor immunotherapy!
1. CAR-macrophage: A new immunotherapy candidate against solid tumors. Biomed Pharmacother. 2021 Jul;139:111605.
Research status and challenges of CAR-macrophages
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