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ROR1 Chimeric Antigen Receptor (CAR): A Comprehensive Guide and Our Service & Product Introduction With the rapid development of cancer immunotherapy, ROR1, as an oncofetal antigen with high specificity in tumor cells, has become one of the most promising targets in the field of CAR therapy. In recent years, the number of research papers and clinical trials on ROR1 CAR has increased significantly, and major biotech companies and research institutions have invested heavily in the research and development of ROR1 CAR-based therapies. The combination of ROR1 CAR with other immunotherapies (e.g., oncolytic viruses, immune checkpoint inhibitors) and the development of CAR-NK cells have become important trends in the industry, which are expected to overcome the limitations of traditional CAR-T therapy and improve the therapeutic effect of ROR1-positive tumors. In addition, the development of non-viral vector delivery technology and the optimization of CAR structure have also promoted the clinical transformation of ROR1 CAR therapy. With the continuous progress of technology, ROR1 CAR therapy is expected to become a new standard treatment for ROR1-positive tumors, bringing new hope to patients. Our ROR1 CAR Expression Plasmid Vector Products and Custom Services RGBiotech provides a full range of ROR1 CAR expression plasmid vectors, covering different generations of ROR1 CAR, and offer professional custom vector construction services to meet the diverse needs of researchers in basic research, and preclinical trials. Our products and services are characterized by high quality, high efficiency, and strong adaptability, providing reliable support for ROR1 CAR-related research.
1) 1st Generation: Contains only the CD3ζ signaling domain, enabling basic T cell activation and cytotoxicity, suitable for preliminary research on ROR1 CAR function. 2) 2nd Generation: Adds one costimulatory domain (e.g., CD28, 4-1BB) based on the first generation, enhancing T cell proliferation, survival, and cytotoxicity. Common constructs include ROR1-4-1BB/CD3ζ. 3) 3rd Generation: Incorporates two costimulatory domains (e.g., CD28+4-1BB, ICOS+4-1BB, ICOS+OX40) to further improve T cell persistence and anti-tumor activity. Novel tandem intracellular signaling domains (ICDs) of the third generation, such as CD28 and 4-1BB ICD followed by CD3ζ, have shown impressive preliminary results in research. 4) 4th Generation (CAR-Treg/ARM CAR): Integrates cytokine-encoding sequences (e.g., IL-12, IL-15) or suicide gene sequences, enabling regulated T cell activation and reducing off-target toxicity, suitable for advanced preclinical research. 5) 5th Generation (CAR-NK/CAR-Mφ): Optimized for natural killer (NK) cells or macrophages, enhancing the anti-tumor activity of innate immune cells. For example, anti-ROR1 CAR-NK cells have shown significant targeting efficacy against neuroblastoma. Product Features 1) Multiple Generations Covered: We provide ROR1 CAR expression plasmids of first to fifth generations, each with unique structural characteristics and functional advantages to meet different research needs.2) Diverse Vector Backbones: We offer a variety of vector backbones, including non-viral vectors (plasmid vectors, transposon vectors) and viral vectors (lentiviral vectors, retroviral vectors, AAV vectors), to adapt to different delivery methods and cell types. For example, third-generation ROR1 CAR constructs are often cloned into lentiviral vectors for efficient transduction of T cells. 3) Optimized Promoters: Equipped with high-efficiency promoters to ensure strong and stable expression of ROR1 CAR, including CMV promoter (strong constitutive expression), EF1α promoter (stable expression in eukaryotic cells, low silencing rate), PGK promoter (moderate expression, suitable for long-term culture), and tissue-specific promoters (optional for custom services), meeting different expression requirements. 4) Multiple Fluorescent Markers: Optional fluorescent markers include EGFP (green fluorescence), mCherry (red fluorescence), facilitating the detection of CAR expression efficiency by flow cytometry, fluorescence microscopy, and other methods, and enabling real-time monitoring of transfected cells. 5) Diverse Antibiotic Selection Markers: Provides common antibiotic selection markers such as Puromycin, Neomycin, Hygromycin, and Blasticidin, facilitating the screening and purification of positive clones in mammalian cells, improving the efficiency of vector construction and cell delivery. 6) Strict Quality Control: Each batch of vectors undergoes strict quality testing, ensuring that the product meets the experimental requirements and avoiding experimental failures caused by vector quality problems. 7) Cost-Effective: We provide high-quality vectors at competitive prices, and offer bulk purchase discounts, reducing the research cost of researchers and enterprises. 8) Fast Delivery: ensuring the progress of the research project. Product Applications
1) Basic Research: Used to study the function of ROR1 CAR, the mechanism of anti-tumor activity, the interaction between CAR and ROR1, and the optimization of CAR structure. For example, evaluating the cytotoxicity, cytokine secretion, and persistence of different generations of ROR1 CAR-T cells in vitro. Custom Vector Construction Services In addition to stock ROR1 CAR expression plasmid vectors, we also provide professional custom vector construction services to meet the personalized needs of researchers. Our custom service team consists of experienced molecular biologists who have in-depth research on ROR1 CAR technology and vector construction, and can provide professional technical advice and solutions to help customers solve problems encountered in the research process.1) Custom ROR1 CAR Structure: According to the customer's research needs, construct ROR1 CAR with specific scFv, costimulatory domains, signaling domains, and other elements. 2) Custom Vector Backbone: Select the appropriate vector backbone (non-viral/viral) according to the customer's delivery method and cell type. 3) Custom Markers: Add specific fluorescent markers, antibiotic selection markers, or reporter genes (e.g.,GFP, luciferase) according to the customer's experimental needs, facilitating the detection and monitoring of CAR expression. 4) One-Stop Service: Provide a one-stop service from vector design, construction, and quality control to delivery, with professional technical support throughout the process, ensuring that the custom vector meets the customer's requirements. Introduction of ROR1 ROR1, officially named Receptor Tyrosine Kinase-Like Orphan Receptor 1, is encoded by the ROR1 gene located on human chromosome 1p31.3 (NCBI Gene ID: 4919). The gene consists of 12 exons and undergoes alternative splicing to produce multiple transcript variants, which encode different protein isoforms. As a member of the ROR subfamily of cell surface receptors, ROR1 is evolutionarily conserved and plays important roles in embryonic development and disease progression. ROR1 is a glycosylated type I membrane protein with a typical modular structure, including four main domains: (1) Extracellular domain: Contains an immunoglobulin (Ig)-like domain, two cysteine-rich domains (CRDs), and a kringle domain, which are involved in ligand binding and protein-protein interactions; (2) Transmembrane domain: A hydrophobic segment that anchors the protein to the cell membrane; (3) Intracellular domain: Possesses a pseudokinase domain (lacking catalytic activity) and a carboxy-terminal tail, which mediates signal transduction by interacting with downstream signaling molecules, such as those in the non-canonical Wnt signaling pathway. Additionally, ROR1 CAR constructs often utilize an IgG4 hinge region to enhance structural flexibility and stability of the CAR receptor. ROR1 plays a crucial role in early embryonic development, regulating processes such as neurite growth in the central nervous system, cell proliferation, differentiation, migration, and apoptosis. It functions as a key mediator of the non-canonical Wnt signaling pathway, participating in the regulation of embryonic tissue patterning and organogenesis. In adult tissues, ROR1 expression is significantly downregulated, maintaining low levels in only a few tissues. However, in tumor cells, ROR1 is abnormally overexpressed, where it promotes tumor cell survival, proliferation, invasion, and metastasis by activating downstream signaling pathways (e.g., PI3K/Akt, MAPK) and regulating the epithelial-mesenchymal transition (EMT) process. Recent studies have also shown that ROR1 forms a positive feedback loop with STAT3 to facilitate cartilage degeneration in osteoarthritis, and serves as a receptor for IGFBP5 to promote glioblastoma invasion. ROR1 exhibits a typical oncofetal expression pattern: it is highly expressed during early embryonic development but expressed at very low levels in normal adult tissues. According to The Human Protein Atlas and related studies, low levels of ROR1 can be detected in normal adult tissues such as adipose tissue, lung, endometrium, kidney, and pancreas, but it is almost undetectable in major normal tissues such as the liver, spleen, and mature B cells. In contrast, ROR1 is abnormally overexpressed in a variety of tumor tissues, making it an ideal target for tumor-specific immunotherapy. The abnormal overexpression of ROR1 is closely associated with the occurrence and development of multiple tumors, including hematological malignancies and solid tumors. Introduction of ROR1 Chimeric Antigen Receptor (CAR) ROR1 CAR is a chimeric antigen receptor that specifically recognizes ROR1, which is constructed by fusing the extracellular ROR1-specific scFv, hinge region, transmembrane domain, and intracellular signaling domain(s) into a single gene, and expressed in immune cells (e.g., T cells, NK cells) through plasmid vector transfection or viral transduction. After ROR1 CAR-expressing immune cells recognize ROR1-positive tumor cells, the intracellular signaling domains are activated, triggering the proliferation, activation, and cytotoxicity of immune cells, thereby specifically killing tumor cells. The structure of ROR1 CAR is continuously optimized, with the development of multiple generations, each with improved anti-tumor activity and safety. Current Research Progress of ROR1 CAR In recent years, ROR1 CAR has become a research hotspot in the field of cancer immunotherapy, and significant progress has been made in basic research and preclinical trials, with some products entering clinical trials.1) Hematological Malignancies: In preclinical studies, ROR1 CAR-T cells have shown strong cytotoxicity against B-CLL, MCL, and other hematological malignancies, and can effectively eliminate tumor cells in animal models. For example, ROR1 CAR-T cells can specifically recognize primary B-CLL and MCL cells, including rare chemotherapy-resistant tumor cells, without targeting mature normal B cells. Clinical trials are currently underway to evaluate the safety and efficacy of ROR1 CAR-T cells in patients with relapsed/refractory hematological malignancies. 2) Solid Tumors: ROR1 CAR-T/NK cells have shown promising anti-tumor effects in preclinical studies on solid tumors such as TNBC, ovarian cancer, and neuroblastoma. Lyell Immunopharma reported initial clinical data from a Phase 1 trial of LYL797, a ROR1-targeted CAR-T cell product candidate enhanced with anti-exhaustion technology, showing dose-dependent anti-tumor activity in relapsed/refractory TNBC, with an objective response rate (ORR) of 40% and clinical benefit rate (CBR) of 60% at the highest dose evaluated. Additionally, the combination of anti-ROR1 CAR-NK cells with an IL-21-secreting oncolytic virus significantly extended the survival of human neuroblastoma xenografted mice compared to controls. 3) CAR Structure Optimization: Researchers are constantly optimizing the structure of ROR1 CAR to improve its anti-tumor activity and persistence. For example, third-generation ROR1 CAR constructs with ICOS/OX40 or CD28/4-1BB tandem co-stimulation domains have shown increased IFN-γ secretion, long-term cytotoxicity, and enhanced T cell memory phenotype compared to second-generation CARs. The use of anti-exhaustion technology in ROR1 CAR-T cells has also been shown to improve T cell expansion and tumor infiltration in solid tumors. Approved Drugs of ROR1 CAR As of May 2026, there are no ROR1 CAR-based drugs officially approved by the FDA, EMA, NMPA, or other regulatory authorities. However, multiple ROR1 CAR-T/NK cell products are in clinical trials (Phase 1/2), showing good safety and efficacy prospects.1) LYL797 (Lyell Immunopharma): A ROR1-targeted CAR-T cell product candidate enhanced with proprietary anti-exhaustion technology. Phase 1 trial data showed dose-dependent anti-tumor activity in relapsed/refractory TNBC, with no significant safety signals in patients without lung involvement. The company is expanding its development into other ROR1-positive tumor types, including ovarian cancer, endometrial cancer, multiple myeloma, and CLL, and has completed an IND submission for LYL119, its next-generation ROR1-targeted CAR-T cell product candidate. 2) Other Clinical Candidates: Several biotech companies and research institutions are conducting clinical trials of ROR1 CAR-T cells for the treatment of B-CLL, MCL, and solid tumors, with preliminary data showing promising anti-tumor activity and manageable safety profiles. It is expected that with the progress of clinical trials, the first ROR1 CAR-based drug will be approved for marketing soon, bringing new treatment options for patients with ROR1-positive tumors. Research Hotspots of ROR1 CAR 1) Optimization of CAR Structure: Focus on the optimization of scFv (improving affinity and specificity), co-stimulation domains (screening more effective co-stimulation combinations, such as ICOS/OX40), and signaling domains, to improve the anti-tumor activity, persistence, and safety of ROR1 CAR-expressing cells. Novel tandem co-stimulation domains are gathering significant interest based on impressive preliminary results.2) Combined Therapy Strategies: Exploring the combination of ROR1 CAR therapy with other treatment methods, such as chemotherapy, radiotherapy, immune checkpoint inhibitors (PD-1/PD-L1 inhibitors), oncolytic viruses, and small molecule inhibitors, to enhance the anti-tumor effect and overcome tumor resistance. For example, the combination of anti-ROR1 CAR-NK cells with an IL-21-secreting oncolytic virus has shown synergistic anti-tumor effects in neuroblastoma models. 3) CAR-NK/CAR-Mφ Therapy: Compared with CAR-T cells, CAR-NK cells have the advantages of no graft-versus-host disease (GVHD), fast anti-tumor effect, and wide applicability. Researchers are focusing on the development of ROR1 CAR-NK cells, optimizing their preparation process and enhancing their persistence and anti-tumor activity. CAR-macrophage (CAR-Mφ) therapy is also a new research direction, which can enhance the phagocytosis and anti-tumor activity of macrophages against ROR1-positive tumors. 4) Overcoming Tumor Microenvironment (TME) Suppression: The immunosuppressive TME is one of the main reasons for the poor efficacy of CAR therapy in solid tumors. Researchers are exploring methods to modify ROR1 CAR-expressing cells to resist the immunosuppressive TME, such as expressing cytokines (IL-12, IL-15) or immune checkpoint inhibitors, to improve the survival and anti-tumor activity of CAR cells in the TME. 5) Off-Target Toxicity Reduction: Developing strategies to reduce the off-target toxicity of ROR1 CAR, such as using conditional CAR (e.g., split CAR, inducible CAR), or selecting scFv with higher specificity to avoid damage to normal cells expressing low levels of ROR1. Research Difficulties & Challenges of ROR1 CAR Despite the significant progress made in ROR1 CAR research, there are still many difficulties and challenges that need to be solved.1) Tumor Heterogeneity: ROR1 expression in tumor tissues is heterogeneous, and some tumor cells may have low or no ROR1 expression, leading to the escape of tumor cells from CAR cell killing and affecting the therapeutic effect. How to overcome tumor heterogeneity and improve the coverage of ROR1 CAR therapy is a major challenge. 2) Immunosuppressive Tumor Microenvironment: The TME of solid tumors contains a variety of immunosuppressive cells (e.g., Treg cells, myeloid-derived suppressor cells) and immunosuppressive factors (e.g., IL-10, TGF-β), which can inhibit the activation and proliferation of ROR1 CAR-expressing cells, reducing their anti-tumor activity. Overcoming the immunosuppressive TME is crucial for improving the efficacy of ROR1 CAR therapy in solid tumors. 3) CAR Cell Exhaustion: Long-term stimulation of CAR cells by tumor antigens can lead to cell exhaustion, characterized by decreased proliferation, cytotoxicity, and cytokine secretion, which affects the long-term anti-tumor effect of CAR therapy. Developing strategies to prevent or reverse CAR cell exhaustion, such as anti-exhaustion technology, is a key research direction. Lyell Immunopharma’s LYL797 uses proprietary anti-exhaustion technology to enhance CAR-T cell function and persistence. 4) Off-Target Toxicity: Although ROR1 is mainly expressed in tumor cells, low levels of ROR1 are still present in some normal tissues (e.g., adipose tissue, kidney), which may lead to off-target toxicity of ROR1 CAR, causing damage to normal cells and tissues. How to reduce off-target toxicity while ensuring anti-tumor activity is a major safety challenge. 5) Preparation and Standardization of CAR Cells: The preparation process of ROR1 CAR-T/NK cells is complex, involving cell isolation, transfection/transduction, expansion, and cryopreservation. The lack of standardized preparation processes and quality control standards may affect the stability and reproducibility of CAR cell products, which is a major obstacle to the clinical transformation of ROR1 CAR therapy. 6) Delivery Efficiency of Vector: For non-viral vector delivery, the transfection efficiency is relatively low, and the expression of CAR is temporary; for viral vector delivery, there are potential safety risks (e.g., insertional mutagenesis) and high production costs. Improving the delivery efficiency and safety of vectors is crucial for the development of ROR1 CAR therapy. Frequently Asked Questions (FAQs) Q: What is the difference between different generations of ROR1 CAR? Which generation should I choose for my research? Q: Can ROR1 CAR be used for both hematological malignancies and solid tumors? Q: What vector backbones do you provide for ROR1 CAR expression? How to choose the appropriate backbone? Q: What promoters are used in your ROR1 CAR plasmid vectors? What are their advantages? Q: How to detect the expression efficiency of ROR1 CAR after vector transfection/transduction? Q: What is the shelf life of your ROR1 CAR plasmid vectors? How to store them? References 1. Liu D, et al. ROR1 is upregulated in endometrial cancer and represents a novel therapeutic target. Sci Rep, 2020 Aug 17. PMID: 32807831.2. Zhang S, et al. Ovarian cancer stem cells express ROR1, which can be targeted for anti-cancer-stem-cell therapy. Proc Natl Acad Sci U S A, 2014 Dec 2. PMID: 25411317. 3. de Propris MS, et al. ROR1 is an accurate and reliable marker of minimal residual disease in chronic lymphocytic leukaemia. Br J Haematol, 2020 Sep. PMID: 32579248. 4. Frontiers in Oncology. ICOS and OX40 tandem co-stimulation enhances CAR T-cell cytotoxicity and promotes T-cell persistence phenotype. 2023. DOI: 10.3389/fonc.2023.1200914. 5. Lyell Immunopharma, Inc. Lyell Immunopharma Reports Dose-dependent Clinical Activity from Phase 1 Trial of LYL797, a ROR1-targeted CAR-T Cell Product Candidate Enhanced with its Proprietary Anti-exhaustion Technology. 2024 Jun 26. 6. PMC. Combinatorial immunotherapy with anti-ROR1 CAR NK cells and an IL-21 secreting oncolytic virus against neuroblastoma. PMID: 38434442. 7. National Center for Biotechnology Information. The B-cell tumor–associated antigen ROR1 can be targeted with T cells modified to express a ROR1-specific chimeric antigen receptor. PMID: 20080691. 8. Wang XY, et al. LncRNA ROR1-AS1 promotes colon cancer cell proliferation by suppressing the expression of DUSP5/CDKN1A. Eur Rev Med Pharmacol Sci, 2020 Feb. PMID: 32096171. |
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