1 Various Approaches to Tumor Treatment
Cancer has become a recognized global health challenge, imposing a significant social and economic burden. In 2022, there were 20 million new cases of cancer worldwide and 9.7 million deaths [1]. Generally, the occurrence of cancer is often accompanied by abnormal activation of oncogenes and persistent silencing of tumor suppressor genes [2].
Currently, cancer treatments include several approaches: traditional methods such as chemotherapy, and emerging therapies like immunotherapy and targeted therapy. Traditional treatments encompass radiation therapy, chemotherapy, and surgical resection, which were the primary strategies for combating cancer in its early stages. However, these methods are often insufficient for completely eliminating the disease, leading to high recurrence rates, drug resistance, and limited specificity with side effects [3,4]. Targeted therapy, often based on antibodies, specifically attacks cancer cells that overexpress certain antigen molecules, such as PD-1 (programmed cell death 1) [5] and PDL-1 (programmed cell death-ligand 1) [6]. Immunotherapy, often based on exogenous cells or exogenous cell factors, enhances the body’s own immune system to effectively kill tumor cells, with examples including engineered CAR-T (Chimeric Antigen Receptor T-Cell) cell therapy [7]. In recent years, immunotherapy has matured and started to be used in clinical practice. Since therapeutic agents often originate from endogenous substances in the body, immunotherapy is characterized by effective tumor cell killing, low recurrence rates, high specificity, and minimal side effects [8]. Moreover, immunotherapy has shown high recovery rates, up to 92% in patients with end-stage acute lymphoblastic leukemia, which has drawn considerable attention [7,9,10]. Additionally, emerging molecular biology technologies have been applied to enhance these therapies, with CRISPR (Clustered Regularly Interspaced Short Palindromic Repeats)/Cas9 being a prominent example [11,12]. Therefore, this paper will discuss the research progress on CRISPR/Cas9-mediated gene editing combined with CAR-T tumor immunotherapy, known as the “gene scissors.”
2 Principles and Development of CAR-T Cell Therapy
2.1 Principles
CAR (Chimeric Antigen Receptor) is an engineered synthetic receptor designed to guide lymphocytes (most commonly T cells) to recognize and eliminate cells expressing specific target antigens. CAR-T cell therapy is a revolutionary cancer treatment method that has been shown to produce highly effective and durable clinical responses [13]. A notable milestone in this field is the unprecedented success of anti-CD19 (cluster of differentiation 19) CAR-T cell therapy in treating B-cell malignancies, which received approval from the U.S. Food and Drug Administration (FDA) in March 2017 [14,15].
The CAR binds to cell surface-expressed target antigens, leading to strong T-cell activation and a robust anti-tumor response, independent of MHC (major histocompatibility complex) receptors [16]. The critical component of CAR-T cells is the modular synthetic receptor CARs, which consists of four main parts [17]: (1) the extracellular target antigen-binding domain, which provides specificity for the target antigen; (2) the hinge region, which provides flexibility to overcome steric hindrance and aids in the adjustment of length to allow the antigen-binding domain to access the target epitope; (3) the transmembrane domain, which anchors the CAR in the T-cell membrane; (4) one or more intracellular signaling domains, which trigger endogenous immune responses in the T cells.
2.2 Development
CAR-T cell therapy has undergone continuous development and iteration, with five generations to date. First-generation CARs involve only one intracellular signaling domain, CD3ζ, while second-generation CARs include an additional co-stimulatory molecule along with CD3ζ. Third-generation CARs contain another co-stimulatory domain. The most advanced fourth-generation CAR-T cells can effectively stimulate downstream transcription factors to trigger cytokine release upon CAR detection of tumor-associated antigens (TAA) [18]. Recently, with the advancement of the CRISPR system, applying CRISPR technology to CAR-T cells to establish fifth-generation CAR-T cells has garnered widespread attention. For example, using CRISPR/Cas9 to knock out negative regulators of T-cell persistence and effector function (such as PD-1 [19], CTLA-4 [20] (cytotoxic T-lymphocyte associated protein-4), and LAG-3 [21]) has the potential to significantly enhance the therapeutic efficacy of CAR-T cells [22].
3 Challenges Facing CAR-T Cell Therapy
3.1 Antigen Escape
One of the most challenging limitations of CAR-T cell therapy is the resistance of tumors to monoclonal antigen-targeted CAR-T constructs. Although initially, single-antigen targeted CAR-T cells can provide high response rates, a significant portion of patients undergoing CAR-T cell therapy [23] exhibit malignant cells that show partial or complete loss of target antigen expression, a phenomenon known as antigen escape [21,24]. For instance, despite 70% of relapsed and 90% of refractory acute lymphoblastic leukemia (ALL) showing durable responses to CD19-targeted CAR-T cell therapy, recent follow-up data indicate a common disease resistance mechanism, with 30-70% of relapsed disease patients experiencing downregulation or loss of CD19 antigen after treatment [25].
3.2 Treatment-Related Side Effects
Despite CAR-T cell therapy being a revolutionary cancer treatment tool, its high toxicity and mortality rates have impeded its adoption as a first-line treatment. The selection of target antigens can result in associated toxicity; ideally, these antigens should possess characteristics such as high coverage, stability, and specificity [26]. Tumor antigens can be classified based on their expression patterns into tumor-specific antigens (TSAs) and tumor-associated antigens (TAAs). TAAs are present not only in tumors but also in normal tissues, leading to toxicity to normal cells when CAR-T cell therapy is applied. However, due to the lack of TSAs, most current CAR-T cell therapies for solid tumors target TAAs. The limited specificity of TAAs makes off-target effects inevitable, sometimes resulting in severe adverse effects that can even be fatal [27].
In addition to the increased likelihood of off-target effects due to the lack of TSAs, in actual treatments, CAR-T cells that recognize target antigens can trigger activation-induced cytokine release or stimulate immune cells to release inflammatory cytokines. This can lead to cytokine release syndrome (CRS), neurotoxicity, and various other adverse reactions [28]. To date, the potential toxicity of CAR-T cell therapy has been most extensively characterized in patients receiving the first FDA-approved CAR-T cell therapy targeting CD19 [29]. Even in clinical trials with the highest response rates, severe, life-threatening adverse reactions occur in patients [30]. Specifically, in patients with acute lymphoblastic leukemia/lymphoblastic lymphoma (LBL) receiving CAR-T cell therapy, almost all patients exhibit at least some mild toxicities, while 23-46% of patients show severe physiological cytokine production and substantial in vivo T-cell expansion [31]. Systemic cytokine release and severe immune cell cross-activation at these toxicity levels can lead to the following toxicities in some patients: 1. CRS [32]; 2. Hemophagocytic lymphohistiocytosis (HLH) and/or macrophage activation syndrome (MAS), a severe high-inflammatory syndrome characterized by CRS, accompanied by elevated serum ferritin, hemophagocytosis, renal failure, elevated liver enzymes, splenomegaly, pulmonary edema, and/or reduced NK cell activity; and immune effector cell-associated neurotoxicity syndrome (ICANS), characterized by elevated cerebrospinal fluid cytokine levels and disruption of the blood-brain barrier [33].
3.3 Immune-Suppressive Microenvironment in Solid Tumors and T-Cell Exhaustion
The complex immune-suppressive network within the tumor microenvironment (TME) is referred to as the tumor immune-suppressive microenvironment, composed of various immune cells, secretions, and inhibitory signals that collectively promote tumor initiation and progression [32]. In the TME, suppressive immune cells, including tumor-associated macrophages (TAMs), regulatory T cells (Tregs), and myeloid-derived suppressor cells (MDSCs), inhibit the proliferation and effective anti-tumor responses of CAR-T cells [34]. These immune cells and tumor cells promote the production of immune-suppressive cytokines such as TGF-β (transforming growth factor-β), IL-10 (interleukin 10), and IL-4 (interleukin 4), accelerating the exhaustion of T cells and CAR-T cells. Due to increased expression of PD-1 (programmed cell death 1) on both the TME and CAR-T cells, the binding of PD-1 on CAR-T cells to PD-L1 (programmed cell death ligand 1) on tumor cells initiates inhibitory signals, leading to impaired CAR-T cell function [35] and promoting immune evasion by tumor cells [36].
3.4 Inefficient Migration and Infiltration of CAR-T Cells in Solid Tumors
In solid tumors, CAR-T cell therapy often yields suboptimal results due to physical barriers and an immune-suppressive TME [37]. Physical barriers, such as abnormal vascular systems, dense extracellular matrix (ECM), and interstitial fluid pressure, obstruct the infiltration and migration of CAR-T cells within solid tumors [38]. Additionally, the presence of immune-suppressive cells, regulatory cytokines, and inhibitory molecules within the TME further limits the expansion and efficacy of CAR-T cells [39].
4 Maturation and Development of the Gene Editing Technology “CRISPR/Cas9”
CRISPR has evolved from an obscure bacterial genomic element to a widely used gene editing platform over several decades. In the late 1980s, researchers first noticed some unusual bacterial elements, where repetitive units were interspersed with non-repetitive spacer sequences [40]. By the mid-2000s, scientists realized that these repetitive units—later known as CRISPR—are part of the bacterial immune system: bacteria integrate phage DNA sequences into the spacer regions to acquire resistance [41]. Protospacer adjacent motifs (PAMs) were discovered as the conserved sequences adjacent to the spacer regions required for target DNA binding [42]. The various components of the CRISPR system were isolated and further defined: CRISPR-associated (Cas) enzymes cleave DNA, CRISPR RNA (crRNA) guides Cas to the DNA target site, and trans-activating CRISPR RNA (tracrRNA) binds with crRNA, forming a protein-RNA-Cas complex [43]. In 2012, two studies demonstrated that CRISPR could programmatically target desired DNA sequences in prokaryotic cells [44]; soon after, there were reports of gene editing in eukaryotic cells [45], solidifying CRISPR’s status as a versatile gene editing tool.
CRISPR systems are classified based on Cas nucleases. The most commonly used gene editing system is CRISPR/Cas9, followed by CRISPR/Cas12a. Cas9 recognizes a PAM sequence of NGG and cuts the target DNA through a blunt double-strand break upstream of the PAM [46].
CRISPR components can be introduced into target cells via transfection or transduction. Studies have shown that high-efficiency gene editing can be achieved using lentiviral transduction of CRISPR/Cas9 [47]. However, stable integration of the Cas9 gene into the host genome may increase off-target editing and trigger unwanted immune responses. Therefore, transient expression of Cas9 protein is advantageous, especially in clinical settings. CRISPR components can be introduced as DNA, RNA, or ribonucleoprotein (RNP) pre-complexes of RNA and Cas protein. RNPs require additional incubation steps but avoid the need for transcription or translation [48]. CRISPR components are typically introduced by electroporation or liposome-mediated infection. In electroporation, a pulsed electric current creates transient pores in the cell membrane, allowing charged RNA/Cas molecules to enter the cytoplasm. Electroporation is a rapid and efficient method for CRISPR gene editing but can be toxic to cells [49]. An alternative method is liposome-mediated infection, which uses cationic lipid particles to complex with CRISPR molecules and enter cells via endocytosis. Liposome-mediated infection has lower toxicity than electroporation but lower transfection efficiency [50].
Identifying target genes that regulate T cell function and fate is a key step in applying CRISPR-Cas9. CRISPR library screening technology is a biological tool based on the CRISPR-Cas9 system used for high-throughput gene function studies. Whole-genome CRISPR-Cas9 screening can unbiasedly identify key factors in cancer cell proliferation, drug resistance, and metastasis [51]. Additionally, CRISPR screening combined with Cas9 gene editing technology can identify essential transcription factors (TFs) for T cell differentiation and functional maintenance, revealing the role of immune suppression and metabolic signaling in shaping T cell fate [52]. Furthermore, it can provide insights into checkpoint regulation of human T cell cytokine production [53] and assist in designing more effective anti-cancer and anti-infection T cells.
5 Improvements and Applications of CRISPR/Cas9 in CAR-T Cell Therapy
5.1 Universal CAR-T Cells (Allogeneic CAR-T Cell Preparation Techniques)
The clinical development of novel CAR-T cell therapies is often hindered by the low yield and poor functionality of autologous peripheral blood T cells, especially in elderly patients and those with extensive pre-treatment. Consequently, the development of allogeneic CAR-T cells has become a major focus [54]. Recently, there has been increasing interest in using non-autologous T cells for treatment. This approach involves generating universal tumor-specific T cells from T cells of healthy donors, which can be used in any patient without the need for human leukocyte antigen (HLA) matching. This strategy could reduce costs, accelerate treatment, and provide T cell products for patients who have experienced lymphopenia or severe cancer, thus lacking sufficient healthy T cells for treatment [55].
A major challenge in allogeneic products is inducing graft-versus-host disease (GVHD). The mechanism of GVHD involves the recognition of “non-self” HLA molecules by the endogenous donor T cell receptors, leading to an immune response [54]. Allogeneic CAR-T cells can be derived from HLA-matched hematopoietic stem cell transplant donors, though this is less common [56]. Therefore, the focus often shifts to gene-edited cells that can be used in non-HLA-matched recipients. The theoretical basis for using these cells in non-HLA-matched subjects is to combine the CAR-T cell manufacturing process with TCR (T cell receptor) knockout. By targeting the TRAC (T cell receptor alpha constant) locus for gene knockout and using CAR as the knock-in target, CAR expression can be driven under the control of the endogenous TRAC promoter, similar to physiological TCR expression [57]. Simultaneously, knocking out the endogenous TCR eliminates the potential for GVHD and enables the production of allogeneic T cell products. This method has shown anti-leukemia efficacy in children and adults with relapsed acute lymphoblastic leukemia (ALL) [58].
5.2 Improving T Cell Dysfunction
Several factors often collectively hinder the long-term remission of CAR-T cell therapy, including issues with autologous CAR-T cell manufacturing, limited CAR-T cell expansion and/or persistence, and various intrinsic and extrinsic resistance mechanisms of T cells. To enhance the survival and functionality of CAR-T cells, various strategies have been employed, such as optimizing costimulatory domains [59], preventing tonic signaling [60], inhibiting the NR4A (Nuclear receptor 4A) transcription factor [61], blocking TOX/TOX2 (Thymocyte selection-associated high mobility group box protein), and overexpressing typical activator protein-1 (AP-1) factor c-Jun [62], but with limited success. Recent studies have shown that intermittent “rest” periods for CAR-T cells can alleviate exhaustion and enhance anti-tumor efficacy under the influence of epigenetic reprogramming. Therefore, using CRISPR/Cas9 technology to perform functional disruption or modulation of genes and epigenetic targets that enhance T cell exhaustion could potentially prevent or even reverse CAR-T cell dysfunction [20]. Targeting inhibitory receptors, transcription factors, and other mediators of CAR-T cell dysfunction through gene editing may rejuvenate the infused cell products. Utilizing CRISPR/Cas9 to remove negative regulatory factors for T cell persistence and functional efficacy (such as PD-1, CTLA-4, and LAG-3) could indeed be the first notable intervention point [63]. In addition to inhibitory receptors, disrupting apoptosis-related factor (FAS) receptor/Fas ligand interactions has been shown to reduce activation-induced cell death and enhance the anti-tumor function of CAR-T cells in vivo [64]. Finally, CRISPR/Cas9-mediated targeting of CAR-T cell diacylglycerol kinase (DGK) metabolism may confer resistance to immunosuppressive mediators in the tumor microenvironment (TME), such as TGFβ [65].
5.3 Enhancing CAR-T Cell Cytotoxicity and Reducing Immunogenicity
During the activation and expansion of CAR-T cells, the CRISPR-Cas9 system combined with transgene knock-in methods has been utilized to modulate cytokine signaling, thereby enhancing anti-tumor activity, improving T cell persistence, and reducing toxicity. Specifically, CRISPR/Cas9-based gene editing, combined with viral or non-viral DNA delivery, can achieve simultaneous biallelic or sequential gene targeting by designing T cells with expression cassettes at specific loci [66]. This technology allows the insertion of cytokine-encoding DNA cassettes into target genomic loci, placing these genes under the control of specific promoters to control expression timing. For example, IL-15 can be inserted into the IL-13 gene locus, thereby placing IL-15 expression under the control of the endogenous IL-13 promoter, which is highly active during T cell activation. This creates an inducible T cell-specific IL-15 activation switch [67]. Additionally, using CRISPR-Cas9 editing technology to remove genes encoding cytokines such as granulocyte-macrophage colony stimulating factor (GM-CSF) and IL-6 [68], which are associated with neurotoxicity and cytokine release syndrome (CRS), may produce cell products with optimal efficacy and persistence while reducing adverse events related to abnormal cytokine production. Therefore, compared to traditional CD19 CAR-T cells, GM-CSF gene knockout CAR-T cells can maintain normal functionality, enhance in vivo anti-tumor activity, and effectively improve overall survival [54]. Gene knockout or ablation of the IL-6 gene may also improve CRS-like toxicity in leukemia mouse models [69].
5.4 CAR-T Cells in Solid Tumor Treatment
In the solid tumor microenvironment, persistent antigen stimulation leads to T cell exhaustion [70]. CRISPR-Cas9 can insert co-stimulatory molecules or disrupt the inhibitory molecule PD-1, thereby enhancing T cell proliferation and anti-tumor activity. In solid tumors, interaction between PD-1 and its ligands causes T cell exhaustion and inhibits CAR-T cell efficacy. Disrupting PD-1 expression in CAR-T cells via CRISPR-Cas9 can enhance their anti-tumor activity.
Additionally, disrupting both TCR and PD-1 can significantly improve CAR-T cell efficacy and reduce autoimmune responses [71]. Overexpression of certain genes, such as runx3, can accelerate T cell migration to tumor sites, which is also applicable to CAR-T therapy [72]. Modifying CAR-T cells with migration markers such as IGAT4, CXCR3, and CXCR1 can improve T cell infiltration [73].
Another approach to enhance CAR-T cells is to enforce the expression of anti-inflammatory cytokines. In melanoma, CAR-T cells can kill tumor-infiltrating lymphocytes that fail to respond to treatment under IL-2 signaling. Overexpression of the p40 subunit of IL-23 in CAR-T cells can enhance their anti-tumor activity in pancreatic cancer models, showing strong expansion capabilities and reduced cell apoptosis, with no significant side effects [74].
5.5 CRISPR Screening of T Cells
T cell therapy has shown impressive results in combating cancer, viruses, and inflammatory diseases. However, their fate and function are largely dependent on the microenvironment. Whole-genome CRISPR-Cas9 screening helps identify key regulators of T cell fate and function. It is also a valuable gene editing tool for determining cell fate, function, and differentiation [75]. CRISPR-based screening technology has driven major discoveries in cell biology and virus-host interactions [76]. This technology has been applied to both in vivo and in vitro T cell lines and primary cells, including whole-genome, metabolic, and transcriptomic screenings. CRISPR-Cas9 screening focused on T cells can be applied to various diseases, not limited to blood and solid cancers/infectious diseases, inflammation, and resistance factors in immunotherapy. It also helps identify pathways that lead to drug resistance in anti-tumor processes [77] and facilitates large-scale exploration of phenotype-related genes [78].
6 Bottlenecks and Prospects
Although the CRISPR-Cas9 system is a highly promising gene editing tool for CAR-T cell therapy, there are some limitations in its clinical application [79]. Firstly, off-target effects are a significant concern. The non-targeting of Cas9 is influenced by DNA topology, transcription, and replication processes, and there are currently no precise tools to predict off-target events. The gold standard for detecting off-target effects is targeted deep sequencing. Secondly, CRISPR-Cas9 is not suitable for all genes and cell types; some cells are difficult to edit, and certain gene regions present greater challenges.
Additionally, occasional chromosomal losses also impact clinical applications [80]. Despite CRISPR-Cas9 being used in Phase I clinical trials for various cancers, enhancing T cell therapies, Cas9-induced chromosomal deletions can impair T cell survival and proliferation. CRISPR-Cas9 targeting TCR chain genes can cause chromosomal truncations, leading to carcinogenic risks and cell death. Chromosomal structural variations can persist in the host for weeks or even months, threatening genomic integrity. Efficient CRISPR-Cas9 toolkits have eliminated chromosomal translocations and viral vector integrations in mouse models [81].
CRISPR-Cas9 genome editing allows T cells to better adapt to specific microenvironments, offering opportunities for advanced T cell therapies. The successful application of CRISPR-Cas9 technology has also driven the development of related tools, such as genome editing tools, gene expression regulation tools (CRISPRa, CRISPRi), and precision editing technologies. These developments will continually address the challenges of combining CRISPR/Cas9 with CAR-T tumor immunotherapy [82].
For example, combining high-throughput techniques, CRISPR-Cas9 screening, single-cell sequencing, and bioinformatics analysis is crucial for in vivo gene editing and screening. Modular pooled gene editing screening platforms (ModPoKI) enable rapid combinations of different gene editing setups, identifying new gene combinations that extend T cell lifespan and enhance anti-cancer efficacy [83]. Research has shown that TFAP4 can improve the adaptability of long-term stimulated CAR-T cells. Non-viral knockout of the BATF-TFAP4 combination significantly enhances engineered T cell capabilities. Combining single-cell sequencing with direct open reading frame capture can examine nearly 12,000 full-length genes driving TCR-induced proliferation, identifying key drivers of T cell secretion of pro-inflammatory cytokines, and providing opportunities for clinical transplantation [84].
7 Conclusion
Overall, given the complexity of diseases and the diversity of genetic backgrounds, using CRISPR-Cas9 technology to personalize treatments by modifying specific genes and cells can significantly improve therapeutic outcomes and enhance patients’ quality of life. However, there are challenges to address, including the high cost of such therapies, the confirmation of long-term efficacy, and the ethical issues associated with gene editing technologies in humans. Further efforts are needed to ensure that more patients benefit from CAR-T cell therapy.
References
[1]. SUNG H, FERLAY J, SIEGEL R L, et al.. Global Cancer Statistics 2020: GLOBOCAN Estimates of Incidence and Mortality Worldwide for 36 Cancers in 185 Countries[J/OL]. CA: A Cancer Journal for Clinicians, 2021, 71(3): 209-249. DOI:10.3322/caac.21660.
[2]. GANESH K, MASSAGUÉ J. Targeting metastatic cancer[J/OL]. Nature medicine, 2021, 27(1): 34-44. DOI:10.1038/s41591-020-01195-4.
[3]. ARRUEBO M, VILABOA N, SÁEZ-GUTIERREZ B, et al.. Assessment of the evolution of cancer treatment therapies[J/OL]. Cancers, 2011, 3(3): 3279-3330. DOI:10.3390/cancers3033279.
[4]. SHAPIRA A, LIVNEY Y D, BROXTERMAN H J, et al.. Nanomedicine for targeted cancer therapy: Towards the overcoming of drug resistance[J/OL]. Drug Resistance Updates, 2011, 14(3): 150-163. DOI:10.1016/j.drup.2011.01.003.
[5]. XU S, OLENYUK B Z, OKAMOTO C T, et al.. Targeting receptor-mediated endocytotic pathways with nanoparticles: rationale and advances[J/OL]. Advanced drug delivery reviews, 2013, 65(1): 121-138. DOI:10.1016/j.addr.2012.09.041.
[6]. SHARKEY R M, GOLDENBERG D M. Targeted Therapy of Cancer: New Prospects for Antibodies and Immunoconjugates[J/OL]. CA: A Cancer Journal for Clinicians, 2006, 56(4): 226-243. DOI:10.3322/canjclin.56.4.226.
[7]. MCCUNE J S. Rapid Advances in Immunotherapy to Treat Cancer[J/OL]. Clinical Pharmacology & Therapeutics, 2018, 103(4): 540-544. DOI:10.1002/cpt.985.
[8]. LETAI A. Functional precision cancer medicine—moving beyond pure genomics[J/OL]. Nature Medicine, 2017, 23(9): 1028-1035. DOI:10.1038/nm.4389.
[9]. ROSENBERG S A, RESTIFO N P, YANG J C, et al.. Adoptive cell transfer: a clinical path to effective cancer immunotherapy[J/OL]. Nature reviews. Cancer, 2008, 8(4): 299-308. DOI:10.1038/nrc2355.
[10]. KARN V, SANDHYA S, HSU W, et al.. CRISPR/Cas9 system in breast cancer therapy: advancement, limitations and future scope[J/OL]. Cancer cell international, 2022, 22(1): 234-234. DOI:10.1186/s12935-022-02654-3.
[11]. CHEN T, WANG M, CHEN Y, et al.. Current challenges and therapeutic advances of CAR-T cell therapy for solid tumors[J/OL]. Cancer Cell International, 2024, 24(1): 133. DOI:10.1186/s12935-024-03315-3.
[12]. RAZEGHIAN E, NASUTION M K M, RAHMAN H S, et al.. A deep insight into CRISPR/Cas9 application in CAR-T cell-based tumor immunotherapies[J/OL]. Stem Cell Research & Therapy, 2021, 12(1): 428. DOI:10.1186/s13287-021-02510-7.
[13]. JUNE C H, O’CONNOR R S, KAWALEKAR O U, et al.. CAR T cell immunotherapy for human cancer[J/OL]. Science, 2018, 359(6382): 1361-1365. DOI:10.1126/science.aar6711.
[14]. NEELAPU S S, LOCKE F L, BARTLETT N L, et al.. Axicabtagene Ciloleucel CAR T-Cell Therapy in Refractory Large B-Cell Lymphoma[J/OL]. New England Journal of Medicine, 2017, 377(26): 2531-2544. DOI:10.1056/NEJMoa1707447.
[15]. MAUDE S L, LAETSCH T W, BUECHNER J, et al.. Tisagenlecleucel in Children and Young Adults with B-Cell Lymphoblastic Leukemia[J/OL]. New England Journal of Medicine, 2018, 378(5): 439-448. DOI:10.1056/NEJMoa1709866.
[16]. SADELAIN M, BRENTJENS R, RIVIÈRE I. The Basic Principles of Chimeric Antigen Receptor Design[J/OL]. Cancer Discovery, 2013, 3(4): 388-398. DOI:10.1158/2159-8290.CD-12-0548.
[17]. ABATE-DAGA D, LAGISETTY K H, TRAN E, et al.. A Novel Chimeric Antigen Receptor Against Prostate Stem Cell Antigen Mediates Tumor Destruction in a Humanized Mouse Model of Pancreatic Cancer[J/OL]. Human Gene Therapy, 2014, 25(12): 1003-1012. DOI:10.1089/hum.2013.209.
[18]. MACKAY M, AFSHINNEKOO E, RUB J, et al.. The therapeutic landscape for cells engineered with chimeric antigen receptors[J/OL]. Nature Biotechnology, 2020, 38(2): 233-244. DOI:10.1038/s41587-019-0329-2.
[19]. LIU X, ZHANG Y, CHENG C, et al.. CRISPR-Cas9-mediated multiplex gene editing in CAR-T cells[J/OL]. Cell Research, 2017, 27(1): 154-157. DOI:10.1038/cr.2016.142.
[20]. ZHANG Y, ZHANG X, CHENG C, et al.. CRISPR-Cas9 mediated LAG-3 disruption in CAR-T cells[J/OL]. Frontiers of Medicine, 2017, 11(4): 554-562. DOI:10.1007/s11684-017-0543-6.
[21]. BRUDNO J N, MARIC I, HARTMAN S D, et al.. T Cells Genetically Modified to Express an Anti–B-Cell Maturation Antigen Chimeric Antigen Receptor Cause Remissions of Poor-Prognosis Relapsed Multiple Myeloma[J/OL]. Journal of Clinical Oncology, 2018, 36(22): 2267-2280. DOI:10.1200/JCO.2018.77.8084.
[22]. REN J, LIU X, FANG C, et al.. Multiplex Genome Editing to Generate Universal CAR T Cells Resistant to PD1 Inhibition[J/OL]. Clinical Cancer Research, 2017, 23(9): 2255-2266. DOI:10.1158/1078-0432.CCR-16-1300.
[23]. GREEN D J, PONT M, SATHER B D, et al.. Fully Human Bcma Targeted Chimeric Antigen Receptor T Cells Administered in a Defined Composition Demonstrate Potency at Low Doses in Advanced Stage High Risk Multiple Myeloma[J/OL]. Blood, 2018, 132(Supplement 1): 1011-1011. DOI:10.1182/blood-2018-99-117729.
[24]. HEGE K M, BERGSLAND E K, FISHER G A, et al.. Safety, tumor trafficking and immunogenicity of chimeric antigen receptor (CAR)-T cells specific for TAG-72 in colorectal cancer[J/OL]. Journal for ImmunoTherapy of Cancer, 2017, 5(1): 22. DOI:10.1186/s40425-017-0222-9.
[25]. MAJZNER R G, MACKALL C L. Tumor Antigen Escape from CAR T-cell Therapy[J/OL]. Cancer Discovery, 2018, 8(10): 1219-1226. DOI:10.1158/2159-8290.CD-18-0442.
[26]. JIANG M, LI Q, XU B. Spotlight on ideal target antigens and resistance in antibody-drug conjugates: Strategies for competitive advancement[J/OL]. Drug Resistance Updates, 2024, 75: 101086. DOI:10.1016/j.drup.2024.101086.
[27]. MURAD J P, KOZLOWSKA A K, LEE H J, et al.. Effective Targeting of TAG72+ Peritoneal Ovarian Tumors via Regional Delivery of CAR-Engineered T Cells[J/OL]. Frontiers in Immunology, 2018, 9: 2268. DOI:10.3389/fimmu.2018.02268.
[28]. XIAO X, HUANG S, CHEN S, et al.. Mechanisms of cytokine release syndrome and neurotoxicity of CAR T-cell therapy and associated prevention and management strategies[J/OL]. Journal of Experimental & Clinical Cancer Research, 2021, 40(1): 367. DOI:10.1186/s13046-021-02148-6.
[29]. ROEX G, TIMMERS M, WOUTERS K, et al.. Safety and clinical efficacy of BCMA CAR-T-cell therapy in multiple myeloma[J/OL]. Journal of Hematology & Oncology, 2020, 13(1): 164. DOI:10.1186/s13045-020-01001-1.
[30]. MAUDE S L, LAETSCH T W, BUECHNER J, et al.. Tisagenlecleucel in Children and Young Adults with B-Cell Lymphoblastic Leukemia[J/OL]. New England Journal of Medicine, 2018, 378(5): 439-448. DOI:10.1056/NEJMoa1709866.
[31]. FREY N V, PORTER D L. Cytokine release syndrome with novel therapeutics for acute lymphoblastic leukemia[J/OL]. Hematology, 2016, 2016(1): 567-572. DOI:10.1182/asheducation-2016.1.567.
[32]. LIU Z, ZHOU Z, DANG Q, et al.. Immunosuppression in tumor immune microenvironment and its optimization from CAR-T cell therapy[J/OL]. Theranostics, 2022, 12(14): 6273-6290. DOI:10.7150/thno.76854.
[33]. SANTOMASSO B D, PARK J H, SALLOUM D, et al.. Clinical and Biological Correlates of Neurotoxicity Associated with CAR T-cell Therapy in Patients with B-cell Acute Lymphoblastic Leukemia[J/OL]. Cancer Discovery, 2018, 8(8): 958-971. DOI:10.1158/2159-8290.CD-17-1319.
[34]. GOBERT M, TREILLEUX I, BENDRISS-VERMARE N, et al.. Regulatory T Cells Recruited through CCL22/CCR4 Are Selectively Activated in Lymphoid Infiltrates Surrounding Primary Breast Tumors and Lead to an Adverse Clinical Outcome[J/OL]. Cancer Research, 2009, 69(5): 2000-2009. DOI:10.1158/0008-5472.CAN-08-2360.
[35]. BARDHAN K, ANAGNOSTOU T, BOUSSIOTIS V A. The PD1:PD-L1/2 Pathway from Discovery to Clinical Implementation[J/OL]. Frontiers in Immunology, 2016, 7[2024-05-12]. http://journal.frontiersin.org/article/10.3389/fimmu.2016.00550/full. DOI:10.3389/fimmu.2016.00550.
[36]. MOTZ G T, COUKOS G. Deciphering and Reversing Tumor Immune Suppression[J/OL]. Immunity, 2013, 39(1): 61-73. DOI:10.1016/j.immuni.2013.07.005.
[37]. NEWICK K, O’BRIEN S, MOON E, et al.. CAR T Cell Therapy for Solid Tumors[J/OL]. Annual Review of Medicine, 2017, 68(1): 139-152. DOI:10.1146/annurev-med-062315-120245.
[38]. MA S, LI X, WANG X, et al.. Current Progress in CAR-T Cell Therapy for Solid Tumors[J/OL]. International Journal of Biological Sciences, 2019, 15(12): 2548-2560. DOI:10.7150/ijbs.34213.
[39]. CARUANA I, SAVOLDO B, HOYOS V, et al.. Heparanase promotes tumor infiltration and antitumor activity of CAR-redirected T lymphocytes[J/OL]. Nature Medicine, 2015, 21(5): 524-529. DOI:10.1038/nm.3833.
[40]. ISHINO Y, SHINAGAWA H, MAKINO K, et al.. Nucleotide sequence of the iap gene, responsible for alkaline phosphatase isozyme conversion in Escherichia coli, and identification of the gene product[J/OL]. Journal of Bacteriology, 1987, 169(12): 5429-5433. DOI:10.1128/jb.169.12.5429-5433.1987.
[41]. MOJICA F J M, SORIA E. Intervening Sequences of Regularly Spaced Prokaryotic Repeats Derive from Foreign Genetic Elements[J/OL]. Journal of Molecular Evolution, 2005, 60(2): 174-182. DOI:10.1007/s00239-004-0046-3.
[42]. BROUNS S J J, JORE M M, LUNDGREN M, et al.. Small CRISPR RNAs Guide Antiviral Defense in Prokaryotes[J/OL]. Science, 2008, 321(5891): 960-964. DOI:10.1126/science.1159689.
[43]. DELTCHEVA E, CHYLINSKI K, SHARMA C M, et al.. CRISPR RNA maturation by trans-encoded small RNA and host factor RNase III[J/OL]. Nature, 2011, 471(7340): 602-607. DOI:10.1038/nature09886.
[44]. GASIUNAS G, BARRANGOU R, HORVATH P, et al.. Cas9–crRNA ribonucleoprotein complex mediates specific DNA cleavage for adaptive immunity in bacteria[J/OL]. Proceedings of the National Academy of Sciences, 2012, 109(39)[2024-05-12]. https://pnas.org/doi/full/10.1073/pnas.1208507109. DOI:10.1073/pnas.1208507109.
[45]. JINEK M, EAST A, CHENG A, et al.. RNA-programmed genome editing in human cells[J/OL]. eLife, 2013, 2: e00471. DOI:10.7554/eLife.00471.
[46]. JINEK M, CHYLINSKI K, FONFARA I, et al.. A Programmable Dual-RNA–Guided DNA Endonuclease in Adaptive Bacterial Immunity[J/OL]. Science, 2012, 337(6096): 816-821. DOI:10.1126/science.1225829.
[47]. COX M J, KUHLMANN C J, STERNER R M, et al.. Improved Anti-Tumor Response of Chimeric Antigen Receptor T Cell (CART) Therapy after GM-CSF Inhibition Is Mechanistically Supported By a Novel Direct Interaction of GM-CSF with Activated Carts[J/OL]. Blood, 2019, 134(Supplement_1): 3868-3868. DOI:10.1182/blood-2019-129349.
[48]. KOURANOVA E, FORBES K, ZHAO G, et al.. CRISPRs for Optimal Targeting: Delivery of CRISPR Components as DNA, RNA, and Protein into Cultured Cells and Single-Cell Embryos[J/OL]. Human Gene Therapy, 2016, 27(6): 464-475. DOI:10.1089/hum.2016.009.
[49]. FAJRIAL A K, HE Q Q, WIRUSANTI N I, et al.. A review of emerging physical transfection methods for CRISPR/Cas9-mediated gene editing[J/OL]. Theranostics, 2020, 10(12): 5532-5549. DOI:10.7150/thno.43465.
[50]. YU X, LIANG X, XIE H, et al.. Improved delivery of Cas9 protein/gRNA complexes using lipofectamine CRISPRMAX[J/OL]. Biotechnology Letters, 2016, 38(6): 919-929. DOI:10.1007/s10529-016-2064-9.
[51]. VAGHARI-TABARI M, HASSANPOUR P, SADEGHSOLTANI F, et al.. CRISPR/Cas9 gene editing: a new approach for overcoming drug resistance in cancer[J/OL]. Cellular & Molecular Biology Letters, 2022, 27(1): 49. DOI:10.1186/s11658-022-00348-2.
[52]. REINA-CAMPOS M, HEEG M, KENNEWICK K, et al.. Metabolic programs of T cell tissue residency empower tumour immunity[J/OL]. Nature, 2023, 621(7977): 179-187. DOI:10.1038/s41586-023-06483-w.
[53]. VILLA M, STANCZAK M A, PEARCE E L. How to make a better T cell: in vivo CRISPR screens have some answers[J/OL]. Cell, 2021, 184(5): 1135-1136. DOI:10.1016/j.cell.2021.02.003.
[54]. BONIFANT C L, JACKSON H J, BRENTJENS R J, et al.. Toxicity and management in CAR T-cell therapy[J/OL]. Molecular Therapy - Oncolytics, 2016, 3: 16011. DOI:10.1038/mto.2016.11.
[55]. AJINA A, MAHER J. Strategies to Address Chimeric Antigen Receptor Tonic Signaling[J/OL]. Molecular Cancer Therapeutics, 2018, 17(9): 1795-1815. DOI:10.1158/1535-7163.MCT-17-1097.
[56]. FRIGAULT M J, LEE J, BASIL M C, et al.. Identification of Chimeric Antigen Receptors That Mediate Constitutive or Inducible Proliferation of T Cells[J/OL]. Cancer Immunology Research, 2015, 3(4): 356-367. DOI:10.1158/2326-6066.CIR-14-0186.
[57]. EYQUEM J, MANSILLA-SOTO J, GIAVRIDIS T, et al.. Targeting a CAR to the TRAC locus with CRISPR/Cas9 enhances tumour rejection[J/OL]. Nature, 2017, 543(7643): 113-117. DOI:10.1038/nature21405.
[58]. BENJAMIN R, GRAHAM C, YALLOP D, et al.. Genome-edited, donor-derived allogeneic anti-CD19 chimeric antigen receptor T cells in paediatric and adult B-cell acute lymphoblastic leukaemia: results of two phase 1 studies[J/OL]. The Lancet, 2020, 396(10266): 1885-1894. DOI:10.1016/S0140-6736(20)32334-5.
[59]. KAWALEKAR O U, O’CONNOR R S, FRAIETTA J A, et al.. Distinct Signaling of Coreceptors Regulates Specific Metabolism Pathways and Impacts Memory Development in CAR T Cells[J/OL]. Immunity, 2016, 44(2): 380-390. DOI:10.1016/j.immuni.2016.01.021.
[60]. LONG A H, HASO W M, SHERN J F, et al.. 4-1BB costimulation ameliorates T cell exhaustion induced by tonic signaling of chimeric antigen receptors[J/OL]. Nature Medicine, 2015, 21(6): 581-590. DOI:10.1038/nm.3838.
[61]. CHEN J, LÓPEZ-MOYADO I F, SEO H, et al.. NR4A transcription factors limit CAR T cell function in solid tumours[J/OL]. Nature, 2019, 567(7749): 530-534. DOI:10.1038/s41586-019-0985-x.
[62]. LIANG C, HUANG S, ZHAO Y, et al.. TOX as a potential target for immunotherapy in lymphocytic malignancies[J/OL]. Biomarker Research, 2021, 9(1): 20. DOI:10.1186/s40364-021-00275-y.
[63]. ZHANG W, SHI L, ZHAO Z, et al.. Disruption of CTLA-4 expression on peripheral blood CD8 + T cell enhances anti-tumor efficacy in bladder cancer[J/OL]. Cancer Chemotherapy and Pharmacology, 2019, 83(5): 911-920. DOI:10.1007/s00280-019-03800-x.
[64]. REN J, ZHANG X, LIU X, et al.. A versatile system for rapid multiplex genome-edited CAR T cell generation[J/OL]. Oncotarget, 2017, 8(10): 17002-17011. DOI:10.18632/oncotarget.15218.
[65]. JUNG I Y, KIM Y Y, YU H S, et al.. CRISPR/Cas9-Mediated Knockout of DGK Improves Antitumor Activities of Human T Cells[J/OL]. Cancer Research, 2018, 78(16): 4692-4703. DOI:10.1158/0008-5472.CAN-18-0030.
[66]. MACLEOD D T, ANTONY J, MARTIN A J, et al.. Integration of a CD19 CAR into the TCR Alpha Chain Locus Streamlines Production of Allogeneic Gene-Edited CAR T Cells[J/OL]. Molecular Therapy, 2017, 25(4): 949-961. DOI:10.1016/j.ymthe.2017.02.005.
[67]. ODÉ Z, CONDORI J, PETERSON N, et al.. CRISPR-Mediated Non-Viral Site-Specific Gene Integration and Expression in T Cells: Protocol and Application for T-Cell Therapy[J/OL]. Cancers, 2020, 12(6): 1704. DOI:10.3390/cancers12061704.
[68]. STERNER R M, SAKEMURA R, COX M J, et al.. GM-CSF inhibition reduces cytokine release syndrome and neuroinflammation but enhances CAR-T cell function in xenografts[J/OL]. Blood, 2019, 133(7): 697-709. DOI:10.1182/blood-2018-10-881722.
[69]. KANG L, TANG X, ZHANG J, et al.. Interleukin-6-knockdown of chimeric antigen receptor-modified T cells significantly reduces IL-6 release from monocytes[J/OL]. Experimental Hematology & Oncology, 2020, 9(1): 11. DOI:10.1186/s40164-020-00166-2.
[70]. SULLIVAN P M, REED S J, KALIA V, et al.. Solid Tumor Microenvironment Can Harbor and Support Functional Properties of Memory T Cells[J/OL]. Frontiers in Immunology, 2021, 12: 706150. DOI:10.3389/fimmu.2021.706150.
[71]. WANG Z, LI N, FENG K, et al.. Phase I study of CAR-T cells with PD-1 and TCR disruption in mesothelin-positive solid tumors[J/OL]. Cellular & Molecular Immunology, 2021, 18(9): 2188-2198. DOI:10.1038/s41423-021-00749-x.
[72]. TANG J, SHENG J, ZHANG Q, et al.. Runx3-overexpression cooperates with ex vivo AKT inhibition to generate receptor-engineered T cells with better persistence, tumor-residency, and antitumor ability[J/OL]. Journal for ImmunoTherapy of Cancer, 2023, 11(2): e006119. DOI:10.1136/jitc-2022-006119.
[73]. FORSBERG E M V, LINDBERG M F, JESPERSEN H, et al.. HER2 CAR-T Cells Eradicate Uveal Melanoma and T-cell Therapy–Resistant Human Melanoma in IL2 Transgenic NOD/SCID IL2 Receptor Knockout Mice[J/OL]. Cancer Research, 2019, 79(5): 899-904. DOI:10.1158/0008-5472.CAN-18-3158.
[74]. MA X, SHOU P, SMITH C, et al.. Interleukin-23 engineering improves CAR T cell function in solid tumors[J/OL]. Nature Biotechnology, 2020, 38(4): 448-459. DOI:10.1038/s41587-019-0398-2.
[75]. FREITAS K A, BELK J A, SOTILLO E, et al.. Enhanced T cell effector activity by targeting the Mediator kinase module[J/OL]. Science, 2022, 378(6620): eabn5647. DOI:10.1126/science.abn5647.
[76]. BAILEY A L, DIAMOND M S. A Crisp(r) New Perspective on SARS-CoV-2 Biology[J/OL]. Cell, 2021, 184(1): 15-17. DOI:10.1016/j.cell.2020.12.003.
[77]. MENNUNI M, FILOGRANA R, FELSER A, et al.. Metabolic resistance to the inhibition of mitochondrial transcription revealed by CRISPR‐Cas9 screen[J/OL]. EMBO reports, 2022, 23(1): e53054. DOI:10.15252/embr.202153054.
[78]. LONG L, WEI J, LIM S A, et al.. CRISPR screens unveil signal hubs for nutrient licensing of T cell immunity[J/OL]. Nature, 2021, 600(7888): 308-313. DOI:10.1038/s41586-021-04109-7.
[79]. NEWTON M D, LOSITO M, SMITH Q M, et al.. Negative DNA supercoiling induces genome-wide Cas9 off-target activity[J/OL]. Molecular Cell, 2023, 83(19): 3533-3545.e5. DOI:10.1016/j.molcel.2023.09.008.
[80]. SONG N, CHU Y, LI S, et al.. Cascade dynamic assembly/disassembly of DNA nanoframework enabling the controlled delivery of CRISPR-Cas9 system[J/OL]. Science Advances, 2023, 9(35): eadi3602. DOI:10.1126/sciadv.adi3602.
[81]. YIN J, FANG K, GAO Y, et al.. Safeguarding genome integrity during gene-editing therapy in a mouse model of age-related macular degeneration[J/OL]. Nature Communications, 2022, 13(1): 7867. DOI:10.1038/s41467-022-35640-4.
[82]. LI T, YANG Y, QI H, et al.. CRISPR/Cas9 therapeutics: progress and prospects[J/OL]. Signal Transduction and Targeted Therapy, 2023, 8(1): 1-23. DOI:10.1038/s41392-023-01309-7.
[83]. CRISPR–Cas9 applications in T cells and adoptive T cell therapies | Cellular & Molecular Biology Letters | Full Text[EB/OL]. [2024-05-27]. https://cmbl.biomedcentral.com/articles/10.1186/s11658-024-00561-1.
[84]. LEGUT M, GAJIC Z, GUARINO M, et al.. A genome-scale screen for synthetic drivers of T cell proliferation[J/OL]. Nature, 2022, 603(7902): 728-735. DOI:10.1038/s41586-022-04494-7.
Cite this article
Hu,W.;Jiang,X. (2024). Research Progress on CRISPR/Cas9-Mediated Gene Editing Combined with CAR-T Tumor Immunotherapy. Theoretical and Natural Science,59,1-12.
Data availability
The datasets used and/or analyzed during the current study will be available from the authors upon reasonable request.
Disclaimer/Publisher's Note
The statements, opinions and data contained in all publications are solely those of the individual author(s) and contributor(s) and not of EWA Publishing and/or the editor(s). EWA Publishing and/or the editor(s) disclaim responsibility for any injury to people or property resulting from any ideas, methods, instructions or products referred to in the content.
About volume
Volume title: Proceedings of the 4th International Conference on Biological Engineering and Medical Science
© 2024 by the author(s). Licensee EWA Publishing, Oxford, UK. This article is an open access article distributed under the terms and
conditions of the Creative Commons Attribution (CC BY) license. Authors who
publish this series agree to the following terms:
1. Authors retain copyright and grant the series right of first publication with the work simultaneously licensed under a Creative Commons
Attribution License that allows others to share the work with an acknowledgment of the work's authorship and initial publication in this
series.
2. Authors are able to enter into separate, additional contractual arrangements for the non-exclusive distribution of the series's published
version of the work (e.g., post it to an institutional repository or publish it in a book), with an acknowledgment of its initial
publication in this series.
3. Authors are permitted and encouraged to post their work online (e.g., in institutional repositories or on their website) prior to and
during the submission process, as it can lead to productive exchanges, as well as earlier and greater citation of published work (See
Open access policy for details).
References
[1]. SUNG H, FERLAY J, SIEGEL R L, et al.. Global Cancer Statistics 2020: GLOBOCAN Estimates of Incidence and Mortality Worldwide for 36 Cancers in 185 Countries[J/OL]. CA: A Cancer Journal for Clinicians, 2021, 71(3): 209-249. DOI:10.3322/caac.21660.
[2]. GANESH K, MASSAGUÉ J. Targeting metastatic cancer[J/OL]. Nature medicine, 2021, 27(1): 34-44. DOI:10.1038/s41591-020-01195-4.
[3]. ARRUEBO M, VILABOA N, SÁEZ-GUTIERREZ B, et al.. Assessment of the evolution of cancer treatment therapies[J/OL]. Cancers, 2011, 3(3): 3279-3330. DOI:10.3390/cancers3033279.
[4]. SHAPIRA A, LIVNEY Y D, BROXTERMAN H J, et al.. Nanomedicine for targeted cancer therapy: Towards the overcoming of drug resistance[J/OL]. Drug Resistance Updates, 2011, 14(3): 150-163. DOI:10.1016/j.drup.2011.01.003.
[5]. XU S, OLENYUK B Z, OKAMOTO C T, et al.. Targeting receptor-mediated endocytotic pathways with nanoparticles: rationale and advances[J/OL]. Advanced drug delivery reviews, 2013, 65(1): 121-138. DOI:10.1016/j.addr.2012.09.041.
[6]. SHARKEY R M, GOLDENBERG D M. Targeted Therapy of Cancer: New Prospects for Antibodies and Immunoconjugates[J/OL]. CA: A Cancer Journal for Clinicians, 2006, 56(4): 226-243. DOI:10.3322/canjclin.56.4.226.
[7]. MCCUNE J S. Rapid Advances in Immunotherapy to Treat Cancer[J/OL]. Clinical Pharmacology & Therapeutics, 2018, 103(4): 540-544. DOI:10.1002/cpt.985.
[8]. LETAI A. Functional precision cancer medicine—moving beyond pure genomics[J/OL]. Nature Medicine, 2017, 23(9): 1028-1035. DOI:10.1038/nm.4389.
[9]. ROSENBERG S A, RESTIFO N P, YANG J C, et al.. Adoptive cell transfer: a clinical path to effective cancer immunotherapy[J/OL]. Nature reviews. Cancer, 2008, 8(4): 299-308. DOI:10.1038/nrc2355.
[10]. KARN V, SANDHYA S, HSU W, et al.. CRISPR/Cas9 system in breast cancer therapy: advancement, limitations and future scope[J/OL]. Cancer cell international, 2022, 22(1): 234-234. DOI:10.1186/s12935-022-02654-3.
[11]. CHEN T, WANG M, CHEN Y, et al.. Current challenges and therapeutic advances of CAR-T cell therapy for solid tumors[J/OL]. Cancer Cell International, 2024, 24(1): 133. DOI:10.1186/s12935-024-03315-3.
[12]. RAZEGHIAN E, NASUTION M K M, RAHMAN H S, et al.. A deep insight into CRISPR/Cas9 application in CAR-T cell-based tumor immunotherapies[J/OL]. Stem Cell Research & Therapy, 2021, 12(1): 428. DOI:10.1186/s13287-021-02510-7.
[13]. JUNE C H, O’CONNOR R S, KAWALEKAR O U, et al.. CAR T cell immunotherapy for human cancer[J/OL]. Science, 2018, 359(6382): 1361-1365. DOI:10.1126/science.aar6711.
[14]. NEELAPU S S, LOCKE F L, BARTLETT N L, et al.. Axicabtagene Ciloleucel CAR T-Cell Therapy in Refractory Large B-Cell Lymphoma[J/OL]. New England Journal of Medicine, 2017, 377(26): 2531-2544. DOI:10.1056/NEJMoa1707447.
[15]. MAUDE S L, LAETSCH T W, BUECHNER J, et al.. Tisagenlecleucel in Children and Young Adults with B-Cell Lymphoblastic Leukemia[J/OL]. New England Journal of Medicine, 2018, 378(5): 439-448. DOI:10.1056/NEJMoa1709866.
[16]. SADELAIN M, BRENTJENS R, RIVIÈRE I. The Basic Principles of Chimeric Antigen Receptor Design[J/OL]. Cancer Discovery, 2013, 3(4): 388-398. DOI:10.1158/2159-8290.CD-12-0548.
[17]. ABATE-DAGA D, LAGISETTY K H, TRAN E, et al.. A Novel Chimeric Antigen Receptor Against Prostate Stem Cell Antigen Mediates Tumor Destruction in a Humanized Mouse Model of Pancreatic Cancer[J/OL]. Human Gene Therapy, 2014, 25(12): 1003-1012. DOI:10.1089/hum.2013.209.
[18]. MACKAY M, AFSHINNEKOO E, RUB J, et al.. The therapeutic landscape for cells engineered with chimeric antigen receptors[J/OL]. Nature Biotechnology, 2020, 38(2): 233-244. DOI:10.1038/s41587-019-0329-2.
[19]. LIU X, ZHANG Y, CHENG C, et al.. CRISPR-Cas9-mediated multiplex gene editing in CAR-T cells[J/OL]. Cell Research, 2017, 27(1): 154-157. DOI:10.1038/cr.2016.142.
[20]. ZHANG Y, ZHANG X, CHENG C, et al.. CRISPR-Cas9 mediated LAG-3 disruption in CAR-T cells[J/OL]. Frontiers of Medicine, 2017, 11(4): 554-562. DOI:10.1007/s11684-017-0543-6.
[21]. BRUDNO J N, MARIC I, HARTMAN S D, et al.. T Cells Genetically Modified to Express an Anti–B-Cell Maturation Antigen Chimeric Antigen Receptor Cause Remissions of Poor-Prognosis Relapsed Multiple Myeloma[J/OL]. Journal of Clinical Oncology, 2018, 36(22): 2267-2280. DOI:10.1200/JCO.2018.77.8084.
[22]. REN J, LIU X, FANG C, et al.. Multiplex Genome Editing to Generate Universal CAR T Cells Resistant to PD1 Inhibition[J/OL]. Clinical Cancer Research, 2017, 23(9): 2255-2266. DOI:10.1158/1078-0432.CCR-16-1300.
[23]. GREEN D J, PONT M, SATHER B D, et al.. Fully Human Bcma Targeted Chimeric Antigen Receptor T Cells Administered in a Defined Composition Demonstrate Potency at Low Doses in Advanced Stage High Risk Multiple Myeloma[J/OL]. Blood, 2018, 132(Supplement 1): 1011-1011. DOI:10.1182/blood-2018-99-117729.
[24]. HEGE K M, BERGSLAND E K, FISHER G A, et al.. Safety, tumor trafficking and immunogenicity of chimeric antigen receptor (CAR)-T cells specific for TAG-72 in colorectal cancer[J/OL]. Journal for ImmunoTherapy of Cancer, 2017, 5(1): 22. DOI:10.1186/s40425-017-0222-9.
[25]. MAJZNER R G, MACKALL C L. Tumor Antigen Escape from CAR T-cell Therapy[J/OL]. Cancer Discovery, 2018, 8(10): 1219-1226. DOI:10.1158/2159-8290.CD-18-0442.
[26]. JIANG M, LI Q, XU B. Spotlight on ideal target antigens and resistance in antibody-drug conjugates: Strategies for competitive advancement[J/OL]. Drug Resistance Updates, 2024, 75: 101086. DOI:10.1016/j.drup.2024.101086.
[27]. MURAD J P, KOZLOWSKA A K, LEE H J, et al.. Effective Targeting of TAG72+ Peritoneal Ovarian Tumors via Regional Delivery of CAR-Engineered T Cells[J/OL]. Frontiers in Immunology, 2018, 9: 2268. DOI:10.3389/fimmu.2018.02268.
[28]. XIAO X, HUANG S, CHEN S, et al.. Mechanisms of cytokine release syndrome and neurotoxicity of CAR T-cell therapy and associated prevention and management strategies[J/OL]. Journal of Experimental & Clinical Cancer Research, 2021, 40(1): 367. DOI:10.1186/s13046-021-02148-6.
[29]. ROEX G, TIMMERS M, WOUTERS K, et al.. Safety and clinical efficacy of BCMA CAR-T-cell therapy in multiple myeloma[J/OL]. Journal of Hematology & Oncology, 2020, 13(1): 164. DOI:10.1186/s13045-020-01001-1.
[30]. MAUDE S L, LAETSCH T W, BUECHNER J, et al.. Tisagenlecleucel in Children and Young Adults with B-Cell Lymphoblastic Leukemia[J/OL]. New England Journal of Medicine, 2018, 378(5): 439-448. DOI:10.1056/NEJMoa1709866.
[31]. FREY N V, PORTER D L. Cytokine release syndrome with novel therapeutics for acute lymphoblastic leukemia[J/OL]. Hematology, 2016, 2016(1): 567-572. DOI:10.1182/asheducation-2016.1.567.
[32]. LIU Z, ZHOU Z, DANG Q, et al.. Immunosuppression in tumor immune microenvironment and its optimization from CAR-T cell therapy[J/OL]. Theranostics, 2022, 12(14): 6273-6290. DOI:10.7150/thno.76854.
[33]. SANTOMASSO B D, PARK J H, SALLOUM D, et al.. Clinical and Biological Correlates of Neurotoxicity Associated with CAR T-cell Therapy in Patients with B-cell Acute Lymphoblastic Leukemia[J/OL]. Cancer Discovery, 2018, 8(8): 958-971. DOI:10.1158/2159-8290.CD-17-1319.
[34]. GOBERT M, TREILLEUX I, BENDRISS-VERMARE N, et al.. Regulatory T Cells Recruited through CCL22/CCR4 Are Selectively Activated in Lymphoid Infiltrates Surrounding Primary Breast Tumors and Lead to an Adverse Clinical Outcome[J/OL]. Cancer Research, 2009, 69(5): 2000-2009. DOI:10.1158/0008-5472.CAN-08-2360.
[35]. BARDHAN K, ANAGNOSTOU T, BOUSSIOTIS V A. The PD1:PD-L1/2 Pathway from Discovery to Clinical Implementation[J/OL]. Frontiers in Immunology, 2016, 7[2024-05-12]. http://journal.frontiersin.org/article/10.3389/fimmu.2016.00550/full. DOI:10.3389/fimmu.2016.00550.
[36]. MOTZ G T, COUKOS G. Deciphering and Reversing Tumor Immune Suppression[J/OL]. Immunity, 2013, 39(1): 61-73. DOI:10.1016/j.immuni.2013.07.005.
[37]. NEWICK K, O’BRIEN S, MOON E, et al.. CAR T Cell Therapy for Solid Tumors[J/OL]. Annual Review of Medicine, 2017, 68(1): 139-152. DOI:10.1146/annurev-med-062315-120245.
[38]. MA S, LI X, WANG X, et al.. Current Progress in CAR-T Cell Therapy for Solid Tumors[J/OL]. International Journal of Biological Sciences, 2019, 15(12): 2548-2560. DOI:10.7150/ijbs.34213.
[39]. CARUANA I, SAVOLDO B, HOYOS V, et al.. Heparanase promotes tumor infiltration and antitumor activity of CAR-redirected T lymphocytes[J/OL]. Nature Medicine, 2015, 21(5): 524-529. DOI:10.1038/nm.3833.
[40]. ISHINO Y, SHINAGAWA H, MAKINO K, et al.. Nucleotide sequence of the iap gene, responsible for alkaline phosphatase isozyme conversion in Escherichia coli, and identification of the gene product[J/OL]. Journal of Bacteriology, 1987, 169(12): 5429-5433. DOI:10.1128/jb.169.12.5429-5433.1987.
[41]. MOJICA F J M, SORIA E. Intervening Sequences of Regularly Spaced Prokaryotic Repeats Derive from Foreign Genetic Elements[J/OL]. Journal of Molecular Evolution, 2005, 60(2): 174-182. DOI:10.1007/s00239-004-0046-3.
[42]. BROUNS S J J, JORE M M, LUNDGREN M, et al.. Small CRISPR RNAs Guide Antiviral Defense in Prokaryotes[J/OL]. Science, 2008, 321(5891): 960-964. DOI:10.1126/science.1159689.
[43]. DELTCHEVA E, CHYLINSKI K, SHARMA C M, et al.. CRISPR RNA maturation by trans-encoded small RNA and host factor RNase III[J/OL]. Nature, 2011, 471(7340): 602-607. DOI:10.1038/nature09886.
[44]. GASIUNAS G, BARRANGOU R, HORVATH P, et al.. Cas9–crRNA ribonucleoprotein complex mediates specific DNA cleavage for adaptive immunity in bacteria[J/OL]. Proceedings of the National Academy of Sciences, 2012, 109(39)[2024-05-12]. https://pnas.org/doi/full/10.1073/pnas.1208507109. DOI:10.1073/pnas.1208507109.
[45]. JINEK M, EAST A, CHENG A, et al.. RNA-programmed genome editing in human cells[J/OL]. eLife, 2013, 2: e00471. DOI:10.7554/eLife.00471.
[46]. JINEK M, CHYLINSKI K, FONFARA I, et al.. A Programmable Dual-RNA–Guided DNA Endonuclease in Adaptive Bacterial Immunity[J/OL]. Science, 2012, 337(6096): 816-821. DOI:10.1126/science.1225829.
[47]. COX M J, KUHLMANN C J, STERNER R M, et al.. Improved Anti-Tumor Response of Chimeric Antigen Receptor T Cell (CART) Therapy after GM-CSF Inhibition Is Mechanistically Supported By a Novel Direct Interaction of GM-CSF with Activated Carts[J/OL]. Blood, 2019, 134(Supplement_1): 3868-3868. DOI:10.1182/blood-2019-129349.
[48]. KOURANOVA E, FORBES K, ZHAO G, et al.. CRISPRs for Optimal Targeting: Delivery of CRISPR Components as DNA, RNA, and Protein into Cultured Cells and Single-Cell Embryos[J/OL]. Human Gene Therapy, 2016, 27(6): 464-475. DOI:10.1089/hum.2016.009.
[49]. FAJRIAL A K, HE Q Q, WIRUSANTI N I, et al.. A review of emerging physical transfection methods for CRISPR/Cas9-mediated gene editing[J/OL]. Theranostics, 2020, 10(12): 5532-5549. DOI:10.7150/thno.43465.
[50]. YU X, LIANG X, XIE H, et al.. Improved delivery of Cas9 protein/gRNA complexes using lipofectamine CRISPRMAX[J/OL]. Biotechnology Letters, 2016, 38(6): 919-929. DOI:10.1007/s10529-016-2064-9.
[51]. VAGHARI-TABARI M, HASSANPOUR P, SADEGHSOLTANI F, et al.. CRISPR/Cas9 gene editing: a new approach for overcoming drug resistance in cancer[J/OL]. Cellular & Molecular Biology Letters, 2022, 27(1): 49. DOI:10.1186/s11658-022-00348-2.
[52]. REINA-CAMPOS M, HEEG M, KENNEWICK K, et al.. Metabolic programs of T cell tissue residency empower tumour immunity[J/OL]. Nature, 2023, 621(7977): 179-187. DOI:10.1038/s41586-023-06483-w.
[53]. VILLA M, STANCZAK M A, PEARCE E L. How to make a better T cell: in vivo CRISPR screens have some answers[J/OL]. Cell, 2021, 184(5): 1135-1136. DOI:10.1016/j.cell.2021.02.003.
[54]. BONIFANT C L, JACKSON H J, BRENTJENS R J, et al.. Toxicity and management in CAR T-cell therapy[J/OL]. Molecular Therapy - Oncolytics, 2016, 3: 16011. DOI:10.1038/mto.2016.11.
[55]. AJINA A, MAHER J. Strategies to Address Chimeric Antigen Receptor Tonic Signaling[J/OL]. Molecular Cancer Therapeutics, 2018, 17(9): 1795-1815. DOI:10.1158/1535-7163.MCT-17-1097.
[56]. FRIGAULT M J, LEE J, BASIL M C, et al.. Identification of Chimeric Antigen Receptors That Mediate Constitutive or Inducible Proliferation of T Cells[J/OL]. Cancer Immunology Research, 2015, 3(4): 356-367. DOI:10.1158/2326-6066.CIR-14-0186.
[57]. EYQUEM J, MANSILLA-SOTO J, GIAVRIDIS T, et al.. Targeting a CAR to the TRAC locus with CRISPR/Cas9 enhances tumour rejection[J/OL]. Nature, 2017, 543(7643): 113-117. DOI:10.1038/nature21405.
[58]. BENJAMIN R, GRAHAM C, YALLOP D, et al.. Genome-edited, donor-derived allogeneic anti-CD19 chimeric antigen receptor T cells in paediatric and adult B-cell acute lymphoblastic leukaemia: results of two phase 1 studies[J/OL]. The Lancet, 2020, 396(10266): 1885-1894. DOI:10.1016/S0140-6736(20)32334-5.
[59]. KAWALEKAR O U, O’CONNOR R S, FRAIETTA J A, et al.. Distinct Signaling of Coreceptors Regulates Specific Metabolism Pathways and Impacts Memory Development in CAR T Cells[J/OL]. Immunity, 2016, 44(2): 380-390. DOI:10.1016/j.immuni.2016.01.021.
[60]. LONG A H, HASO W M, SHERN J F, et al.. 4-1BB costimulation ameliorates T cell exhaustion induced by tonic signaling of chimeric antigen receptors[J/OL]. Nature Medicine, 2015, 21(6): 581-590. DOI:10.1038/nm.3838.
[61]. CHEN J, LÓPEZ-MOYADO I F, SEO H, et al.. NR4A transcription factors limit CAR T cell function in solid tumours[J/OL]. Nature, 2019, 567(7749): 530-534. DOI:10.1038/s41586-019-0985-x.
[62]. LIANG C, HUANG S, ZHAO Y, et al.. TOX as a potential target for immunotherapy in lymphocytic malignancies[J/OL]. Biomarker Research, 2021, 9(1): 20. DOI:10.1186/s40364-021-00275-y.
[63]. ZHANG W, SHI L, ZHAO Z, et al.. Disruption of CTLA-4 expression on peripheral blood CD8 + T cell enhances anti-tumor efficacy in bladder cancer[J/OL]. Cancer Chemotherapy and Pharmacology, 2019, 83(5): 911-920. DOI:10.1007/s00280-019-03800-x.
[64]. REN J, ZHANG X, LIU X, et al.. A versatile system for rapid multiplex genome-edited CAR T cell generation[J/OL]. Oncotarget, 2017, 8(10): 17002-17011. DOI:10.18632/oncotarget.15218.
[65]. JUNG I Y, KIM Y Y, YU H S, et al.. CRISPR/Cas9-Mediated Knockout of DGK Improves Antitumor Activities of Human T Cells[J/OL]. Cancer Research, 2018, 78(16): 4692-4703. DOI:10.1158/0008-5472.CAN-18-0030.
[66]. MACLEOD D T, ANTONY J, MARTIN A J, et al.. Integration of a CD19 CAR into the TCR Alpha Chain Locus Streamlines Production of Allogeneic Gene-Edited CAR T Cells[J/OL]. Molecular Therapy, 2017, 25(4): 949-961. DOI:10.1016/j.ymthe.2017.02.005.
[67]. ODÉ Z, CONDORI J, PETERSON N, et al.. CRISPR-Mediated Non-Viral Site-Specific Gene Integration and Expression in T Cells: Protocol and Application for T-Cell Therapy[J/OL]. Cancers, 2020, 12(6): 1704. DOI:10.3390/cancers12061704.
[68]. STERNER R M, SAKEMURA R, COX M J, et al.. GM-CSF inhibition reduces cytokine release syndrome and neuroinflammation but enhances CAR-T cell function in xenografts[J/OL]. Blood, 2019, 133(7): 697-709. DOI:10.1182/blood-2018-10-881722.
[69]. KANG L, TANG X, ZHANG J, et al.. Interleukin-6-knockdown of chimeric antigen receptor-modified T cells significantly reduces IL-6 release from monocytes[J/OL]. Experimental Hematology & Oncology, 2020, 9(1): 11. DOI:10.1186/s40164-020-00166-2.
[70]. SULLIVAN P M, REED S J, KALIA V, et al.. Solid Tumor Microenvironment Can Harbor and Support Functional Properties of Memory T Cells[J/OL]. Frontiers in Immunology, 2021, 12: 706150. DOI:10.3389/fimmu.2021.706150.
[71]. WANG Z, LI N, FENG K, et al.. Phase I study of CAR-T cells with PD-1 and TCR disruption in mesothelin-positive solid tumors[J/OL]. Cellular & Molecular Immunology, 2021, 18(9): 2188-2198. DOI:10.1038/s41423-021-00749-x.
[72]. TANG J, SHENG J, ZHANG Q, et al.. Runx3-overexpression cooperates with ex vivo AKT inhibition to generate receptor-engineered T cells with better persistence, tumor-residency, and antitumor ability[J/OL]. Journal for ImmunoTherapy of Cancer, 2023, 11(2): e006119. DOI:10.1136/jitc-2022-006119.
[73]. FORSBERG E M V, LINDBERG M F, JESPERSEN H, et al.. HER2 CAR-T Cells Eradicate Uveal Melanoma and T-cell Therapy–Resistant Human Melanoma in IL2 Transgenic NOD/SCID IL2 Receptor Knockout Mice[J/OL]. Cancer Research, 2019, 79(5): 899-904. DOI:10.1158/0008-5472.CAN-18-3158.
[74]. MA X, SHOU P, SMITH C, et al.. Interleukin-23 engineering improves CAR T cell function in solid tumors[J/OL]. Nature Biotechnology, 2020, 38(4): 448-459. DOI:10.1038/s41587-019-0398-2.
[75]. FREITAS K A, BELK J A, SOTILLO E, et al.. Enhanced T cell effector activity by targeting the Mediator kinase module[J/OL]. Science, 2022, 378(6620): eabn5647. DOI:10.1126/science.abn5647.
[76]. BAILEY A L, DIAMOND M S. A Crisp(r) New Perspective on SARS-CoV-2 Biology[J/OL]. Cell, 2021, 184(1): 15-17. DOI:10.1016/j.cell.2020.12.003.
[77]. MENNUNI M, FILOGRANA R, FELSER A, et al.. Metabolic resistance to the inhibition of mitochondrial transcription revealed by CRISPR‐Cas9 screen[J/OL]. EMBO reports, 2022, 23(1): e53054. DOI:10.15252/embr.202153054.
[78]. LONG L, WEI J, LIM S A, et al.. CRISPR screens unveil signal hubs for nutrient licensing of T cell immunity[J/OL]. Nature, 2021, 600(7888): 308-313. DOI:10.1038/s41586-021-04109-7.
[79]. NEWTON M D, LOSITO M, SMITH Q M, et al.. Negative DNA supercoiling induces genome-wide Cas9 off-target activity[J/OL]. Molecular Cell, 2023, 83(19): 3533-3545.e5. DOI:10.1016/j.molcel.2023.09.008.
[80]. SONG N, CHU Y, LI S, et al.. Cascade dynamic assembly/disassembly of DNA nanoframework enabling the controlled delivery of CRISPR-Cas9 system[J/OL]. Science Advances, 2023, 9(35): eadi3602. DOI:10.1126/sciadv.adi3602.
[81]. YIN J, FANG K, GAO Y, et al.. Safeguarding genome integrity during gene-editing therapy in a mouse model of age-related macular degeneration[J/OL]. Nature Communications, 2022, 13(1): 7867. DOI:10.1038/s41467-022-35640-4.
[82]. LI T, YANG Y, QI H, et al.. CRISPR/Cas9 therapeutics: progress and prospects[J/OL]. Signal Transduction and Targeted Therapy, 2023, 8(1): 1-23. DOI:10.1038/s41392-023-01309-7.
[83]. CRISPR–Cas9 applications in T cells and adoptive T cell therapies | Cellular & Molecular Biology Letters | Full Text[EB/OL]. [2024-05-27]. https://cmbl.biomedcentral.com/articles/10.1186/s11658-024-00561-1.
[84]. LEGUT M, GAJIC Z, GUARINO M, et al.. A genome-scale screen for synthetic drivers of T cell proliferation[J/OL]. Nature, 2022, 603(7902): 728-735. DOI:10.1038/s41586-022-04494-7.