Anti-FAP mRNA CAR T cells(University of Pennsylvania/Capstan Therapeutics)

Hepatic fibrosis is a key predictor of mortality in liver disease, driven by fibrogenic hepatic stellate cells (HSCs). Targeting these fibrogenic cells may therefore offer a therapeutic approach for hepatic fibrosis. We previously showed that in vivo–generated chimeric antigen receptor (CAR) T cells targeting fibroblast activation protein alpha (FAP) reduced murine cardiac fibrosis. Here, we explored the antifibrotic potential of this in vivo–generated anti-FAP CAR T cell therapy in metabolic dysfunction–associated steatohepatitis (MASH), a highly prevalent disease with no approved antifibrotic therapies. We first established that FAP expression in both human and murine MASH is specific to HSCs. We then used flow cytometry, Sirius Red morphometry, digital pathology analysis, and single nuclear RNA sequencing to assess the impact of anti-FAP CAR T cell therapy on murine MASH. Anti-CD5–targeted lipid nanoparticles carrying anti-FAPCAR messenger RNA transiently generated activated anti-FAP CAR T cells, which substantially reduced fibrosis by depleting profibrogenic HSCs. They also modulated immune cells, endothelial cells, and hepatocytes in a non–cell autonomous manner to mitigate inflammation and restore hepatic homeostasis. These findings highlight the potential of in vivo CAR T therapy to attenuate a highly morbid and pervasive liver disease, not only by directly reducing fibrosis but also through indirect effects on other cell types.

Despite the success of chimeric antigen receptor (CAR) T-cell therapy against hematologic malignancies, successful targeting of solid tumors with CAR T cells has been limited by a lack of durable responses and reports of toxicities. Our understanding of the limited therapeutic efficacy in solid tumors could be improved with quantitative tools that allow characterization of CAR T–targeted antigens in tumors and accurate monitoring of response.

We used a radiolabeled FAP inhibitor (FAPI) [18F]AlF-FAPI-74 probe to complement ongoing efforts to develop and optimize FAP CAR T cells. The selectivity of the radiotracer for FAP was characterized in vitro, and its ability to monitor changes in FAP expression was evaluated using rodent models of lung cancer.

[18F]AlF-FAPI-74 showed selective retention in FAP+ cells in vitro, with effective blocking of the uptake in presence of unlabeled FAPI. In vivo, [18F]AlF-FAPI-74 was able to detect FAP expression on tumor cells as well as FAP+ stromal cells in the tumor microenvironment with a high target-to-background ratio. We further demonstrated the utility of the tracer to monitor changes in FAP expression following FAP CAR T-cell therapy, and the PET imaging findings showed a robust correlation with ex vivo analyses.

This noninvasive imaging approach to interrogate the tumor microenvironment represents an innovative pairing of a diagnostic PET probe with solid tumor CAR T-cell therapy and has the potential to serve as a predictive and pharmacodynamic response biomarker for FAP as well as other stroma-targeted therapies. A PET imaging approach targeting FAP expressed on activated fibroblasts of the tumor stroma has the potential to predict and monitor therapeutic response to FAP-targeted CAR T-cell therapy.See related commentary by Weber et al., p. 5241