Virus-Like Particles Unlock the CRISPR Toolbox in Myeloid Immune Cells, Revealing a Potency Switch for CAR Macrophages

Myeloid immune cells can engulf pathogens, present antigens, and profoundly shape the tumor microenvironment, yet they have long been difficult targets for genetic engineering. Conventional viral vectors or electroporation often require trade-offs among efficiency, toxicity, and impaired cellular function. A study published in *Nature Biotechnology* has now established a delivery toolkit using virus-like particles, enabling primary human monocytes, macrophages, and dendritic cells to undergo multiple types of CRISPR manipulation while preserving viability and innate immune responses as much as possible.

Virus-like particles harness viruses’ ability to enter cells but do not carry a complete viral genome capable of self-replication. The research team used them to deliver editing components including Cas9, achieving gene knockout, base editing, and epigenetic silencing. The significance of this strategy lies not only in modifying a single gene, but also in bringing functional genomics tools—previously used more often in readily cultured cell lines—into primary myeloid cells that more closely reflect human biology.

To conduct pooled large-scale screening, the researchers further developed the SLICeVLP system: a VPX-containing lentiviral vector first delivers guide RNA, followed by engineered virus-like particles supplying Cas9. This division of labor allows different genetic perturbations to be compared within the same batch of cells. Publicly available data also show that the study encompassed single-cell RNA sequencing, CRISPR interference, and macrophage Perturb-seq, and screened CD14-positive monocytes under untreated and lipopolysaccharide-stimulated conditions; raw sequencing data and processed files have been deposited in public databases.

The screening results brought TNFAIP3 into focus. The A20 protein encoded by this gene is an important brake on inflammatory signaling; after TNFAIP3 was removed, macrophages exhibited a stronger pro-inflammatory state and were less readily driven toward suppressive polarization. When this modification was introduced into chimeric antigen receptor macrophages that recognize HER2, the researchers observed greater tumor-cell-killing capacity, suggesting that functional screening may directly identify complementary genetic modifications for cell therapies.

These findings still cannot be equated with a new cancer therapy. TNFAIP3 limits immune activation, and although releasing this brake may enhance attack potency, it may also cause excessive inflammation, loss of control over cellular state, or tissue damage; the information currently available is insufficient to determine its persistence in animals, safety range, or actual therapeutic benefit. Differences in editing efficiency and immune responses among donors, off-target effects, and consistency in large-scale manufacturing also require systematic validation.

The longer-term value may lie in establishing a bridge from gene-function discovery to cellular engineering. The research team has demonstrated multiple editing modalities and pooled screening in primary myeloid cells; if the approach can eventually be expanded to genome-wide scale and the results reproduced in more complex tumor models and clinical-grade manufacturing processes, the platform may help researchers identify more quickly which immune brakes are worth releasing and which modifications would instead create unacceptable risks.