Preclinical study of FAP-specific CAR-T cells in treating myocardial fibrosis
M Guo, X Xu, Y Cai, H Zhu, M Shi, J Liu, W Wu, G Wang, D PanAbstract
Background
Myocardial fibrosis (MF) is a common pathological process underlying various cardiac diseases and is generally considered irreversible once established. Following progression to heart failure, approximately 40%–60% of patients die within five years of diagnosis. Therefore, effective inhibition of MF is of great significance for the improvement of patient prognosis. Currently, no effective treatments for MF are available in clinical practice. Some studies have demonstrated that chimeric antigen receptor T (CAR-T) cell therapy can effectively reverse MF in mouse models, offering a promising new therapeutic strategy.
Purpose
In this study, we generated CAR-T cells targeting fibroblast activation protein (FAP), a surface marker highly expressed on activated cardiac fibroblasts. We further evaluated the therapeutic efficacy of these FAP-specific CAR-T cells in a mouse model of MF, aiming to provide a foundation for their potential clinical translation.
Methods
MF was induced in male NCG mice (6–8 weeks old) via continuous subcutaneous injection of isoproterenol (ISO, 5 mg/day) for either 14 or 28 days. Following successful model establishment, FAP CAR-T cells (5×10⁶ cells/mouse) expressing fluorescence were administered via tail vein injection. In vivo imaging was performed on days 1, 3, 7, 14, 21, and 28 post-injection to monitor CAR-T cell proliferation and biodistribution. Blood samples were collected before treatment and on days 1, 7, and 14 for measurement of brain natriuretic peptide (BNP). Flow cytometry assessed CAR expression in peripheral immune cell subsets. On day 42 post-treatment, heart, liver, spleen, kidney, skin, and brain were harvested for histological analysis. HE staining evaluated histopathology, while Masson's trichrome staining assessed fibrosis severity. Collagen volume fraction (CVF) was quantified using ImageJ and compared among groups.
Results
Continuous ISO injection effectively induced MF. In FAP CAR-T-treated mice, in vivo imaging revealed progressively enhanced fluorescence signals over time. Flow cytometry revealed long-term persistence of CAR-T cells in the peripheral blood, indicating successful activation and proliferation. Compared with control mice, both the 14-day and 28-day model groups treated with CAR-T cells exhibited significantly reduced serum BNP levels (P < 0.01). Histological analysis by Masson staining corroborated this finding, demonstrating markedly attenuated myocardial fibrosis in these treatment groups, with no pathological alterations observed in other organs. Quantitatively, the CVF was significantly lower in both the 14-day (P < 0.001) and 28-day (P < 0.001) modeling groups than in the control group.
Conclusions
FAP-specific CAR-T cells effectively attenuate myocardial fibrosis in a preclinical mouse model, providing robust preclinical evidence for their translational potential. These findings support the further development of this strategy for MF therapy.