Assembling Genes Evolution Never Made Boosts CAR-T Antitumor Activity

A study has found that assembling genes in large numbers through “structural design,” rather than artificial intelligence, can boost the antitumor activity of CAR-T cells, which are prone to exhaustion.

A research team led by Ansuman T. Satpathy at Stanford University in the United States developed DESynR, a technique for creating synthetic transcription factors by recombining structural domains derived from natural transcription factors, and used it to enhance CAR-T cell antitumor function. The findings were published in the international journal Cell. The paper was published on the 19th (local time).

The team combined DNA-binding domains (DBDs), transcriptional activation domains and protein-interaction domains from the AP-1 transcription factor family as “parts.” In the process, they created 13,824 barcode-labeled synthetic AP-1 genes and introduced them into human CAR-T cells to screen for function.

As a result, many DESynR AP-1 transcription factors showed better antitumor performance than natural AP-1 factors, including JUN and BATF, in vitro and in vivo. These variants reprogrammed gene-expression programs in T cells by introducing domains from transcription factors that are expressed at very low levels in T cells.

In particular, some variants borrowed regulatory modules from non-T-cell types, such as neurons and epithelial cells, to simultaneously optimize T-cell exhaustion, effector function, cytotoxicity, stem cell properties and persistence.

The team also applied the technique to the ETS and FOX transcription factor families, obtaining synthetic variants with enhanced function and confirming the versatility of the DESynR approach.

However, the findings were validated mainly in vitro and in xenograft mouse models, and long-term safety in humans, immunogenicity and off-target effects require further evaluation. In addition, large-scale screening of 13,824 variants relied on lentiviral libraries and barcode sequencing, leaving manufacturing complexity and quality control as challenges for clinical application.

The team said, “Rather than modifying existing genes, this approach starts with a ‘parts bin’ of protein domains to design genes that did not exist in evolution, representing a new paradigm,” adding that it “opens up the possibility of rationally designing next-generation T-cell therapies.”