In recent years, CAR-T cell therapy has achieved remarkable clinical outcomes in hematologic malignancies. However, the preparation of conventional CAR-T/NK cells still faces multiple bottlenecks: electroporation delivery of Cas9-gRNA ribonucleoprotein (RNP) is not only costly and operationally cumbersome but also prone to cell damage; viral vectors carry safety concerns regarding random genomic integration. Concurrently, significant batch-to-batch variability and high quality control hurdles continue to impede the widespread clinical application of cell therapy products.

Recently, the core R&D team of Shenzhen Cell Valley, in collaboration with Beijing University of Chinese Medicine and Peking University Shenzhen Hospital, published a breakthrough research paper in the internationally renowned immunology journal Frontiers in Immunology (Impact Factor: 7.0). The Cell Valley team successfully established an engineered virus-like particle (eVLP) R&D platform with scalable manufacturing capacity and GMP-compliance, providing a novel technological solution for the industrial-scale preparation of next-generation gene-edited CAR-T/NK cells.

Technological Breakthrough:

From "Four-Plasmid Transient Transfection" to "Stable Production with Engineered Cell Lines"

Traditional eVLP preparation relies on transient transfection of four plasmids—a cumbersome process with unstable yields, making it difficult to meet the demands of clinical-grade scaled production. The research team innovatively integrated three core modules—Gag-Pol, Gag-Cas9, and the BaEV envelope protein—into a monoclonal HEK293T cell line, constructing a precursor cell line. On this basis, a self-inactivating (SIN) retroviral vector was used to introduce a CD7-targeting gRNA, yielding a dedicated high-yield eVLP-producing cell line. Compared to the transient transfection system, this stable cell line platform offers better batch-to-batch consistency, more robust process stability, and greater suitability for large-scale industrial production.

Core Advantages:

High Efficiency, Safety, and Flexibility

In functional validation, CD7-eVLPs produced using this platform achieved up to 93.16% gene knockout efficiency in Jurkat cells, and stable knockout efficiency exceeding 80% in primary T cells. Following monoclonal screening, the optimal clone 10A3 produced eVLPs that achieved over 90% CD7 knockout efficiency in T cells, with post-editing cell viability maintained above 79%.

In terms of safety, the study confirmed that the SIN design effectively prevented the LTR-gRNA element from being re-packaged into eVLPs. The GFP positivity rate in transduced target cells was only 0.23%, far below the positive threshold, eliminating the risk of genomic integration.

Significance of the Results:

Unlocking the "Last Mile" of Industrializing Gene-Edited Cell Therapies

The significance of this study lies not only in achieving technical validation of CD7 gene knockout and the preparation of CD7 CAR-T cells, but also in establishing a replicable, scalable, GMPoriented engineered eVLP platform. Compared to conventional multi-plasmid transient transfection and electroporation-based gene editing processes, this platform—centered on stable production cell lines—enables sustainable, scalable preparation of target-specific eVLP vectors. It helps reduce batch-to-batch variability, simplify manufacturing workflows, lower dependence on serum and reagent consumables, and avoid the cellular damage caused by electroporation, offering a novel solution for integrating gene editing technologies into standardized, scaled production systems. The results demonstrate that CD7 eVLPs produced on this platform achieve efficient and stable gene knockout while preserving cell viability, proliferative capacity, and antitumor function, supporting the scale-up preparation of gene-edited CAR-T cells.

This achievement further reinforces the Cell Valley Group's foundational technological capabilities at the intersection of viral vectors, gene editing, and cell therapy. Leveraging its modular design, the platform can be rapidly adapted to different gene targets by simply replacing the gRNA, and can be further integrated with AAV donor templates, multi-target eVLP combinations, and base editing or prime editing technologies, providing greater flexibility for the development of universal CAR-T, CAR-NK, and other next-generation gene-edited cell products.

Moreover, this platform advances eVLP technology from the laboratory stage toward engineering, industrialization, and clinical-grade manufacturing. It is expected to shorten the development cycle for novel target vectors and gene-edited cell products while improving production consistency and quality controllability. Going forward, the Cell Valley Group will leverage its existing cell therapy, viral vector, and GMP production systems to continuously optimize the eVLP platform's processes, quality standards, and product pipeline translation. It will also provide gene-editing vector development and cell product preparation services to hospitals, research institutions, and biopharmaceutical companies, accelerating the construction of an integrated technology platform covering "tool vectors – gene editing – cell manufacturing – clinical translation."

The publication of this achievement marks a new phased breakthrough for the Cell Valley Group in next-generation core technologies for cell and gene therapy. It also provides critical support for lowering the manufacturing barriers of geneedited cell therapies, promoting the scalable application of advanced therapeutic technologies, and enhancing the independent technological capabilities of China's CGT industry.