Targeting Cardiac Fibrosis with a Vaccine Against Fibroblast Activation Protein

Cardiac fibrosis accompanies most heart diseases¹. While fibrosis is vital for cardiac repair and structural integrity following myocardial infarction, fibrosis also occurs in the myocardial interstitium in systemic hypertension, diabetes, obesity, and aging, where it contributes to ventricular dysfunction^1,2. Indeed, the extent of interstitial fibrosis often correlates with increased morbidity and mortality¹. These observations suggest that inhibiting fibrosis may have therapeutic benefits across the spectrum of heart disease.

Myocardial fibrosis is driven by the excessive deposition of extracellular matrix proteins by myofibroblasts. Genetic deletion of myofibroblasts using diphtheria toxin-mediated ablation of periostin-expressing cells decreased cardiac fibrosis and improved function following chronic angiotensin II (AngII) exposure or myocardial infarction^3,4. Building on these observations, Aghajanian and colleagues developed chimeric antigen receptor (CAR) T cells targeting myofibroblasts expressing fibroblast activation protein (FAP)⁵. FAP is a transmembrane serine protease expressed at low levels in normal adult tissues⁶ that is selectively and robustly up-regulated by cardiac fibroblasts in myocardial infarction and cardiomyopathy^5,7. FAP CAR T cells decreased interstitial fibrosis and restored function in the AngII/phenylephrine (PE) model of hypertensive cardiac injury⁵. Although the authors observed no evidence of cardiotoxicity, immune activation, or systemic effects in this model, CAR T cell therapies are associated with inflammatory toxicities and off-target effects⁸. High costs and lengthy production times also hamper the broad adoption of CAR T cell therapy⁹.

Seeking to overcome these limitations, Yoshida and colleagues, in this issue of Circulation Research, demonstrate the efficacy of an FAP peptide vaccine in the AngII/PE cardiomyopathy model¹⁰. The authors examined the immunogenicity of three FAP peptides conjugated to a keyhole limpet hemocyanin (KLH) carrier protein administered with the adjuvant CpG K3. Vaccinated male C57Bl/6J mice, treated at 10 and 12 weeks of age, produced anti-FAP antibodies within 4 weeks, with levels sustained for at least 12 weeks. Importantly, T cell activation was not observed in splenocytes from immunized mice.

The investigators then administered the FAP vaccine to male C57Bl/6J mice at 8, 10, and 12 weeks of age, prior to the implanting of AngII/PE osmotic minipumps to induce hypertensive cardiomyopathy. Compared to AngII/PE and KLH controls, FAP-vaccinated animals showed improved survival, reduced heart weight, and decreased interstitial fibrosis. However, no changes in cardiac functional parameters were observed. The relatively mild severity of cardiomyopathy in their model¹¹ may have limited the their ability to detect these beneficial effects of the FAP vaccine. In a separate myocardial infarction model, FAP vaccination reduced fibrosis relative to unvaccinated mice, although it was no different from KLH-vaccinated controls. Additionally, FAP vaccination did not improve survival or cardiac function in this model. Mechanistically, serum from vaccinated animals triggered both antibody-dependent cell-mediated cytotoxicity and complement-dependent cytotoxicity in HEK cells overexpressing FAP protein.

The FAP vaccine was well-tolerated. Pro-inflammatory cytokine levels (IFNγ, IL-6, and IL-10) in serum were not elevated. While IgG immune complexes were observed in the heart, lung, and kidney, these were not associated with immune cell infiltrates. AngII/PE-induced pulmonary fibrosis was likely reduced, and no histologic abnormalities were observed in glomeruli. Similarly, examination of FAP-expressing non-cardiac tissues revealed no evidence of necrosis. In a wound healing assay, FAP-vaccinated animals had delayed initiation of wound closure, but ultimately healed in the same time frame as unvaccinated controls.

Overall, the authors achieved their goal of developing a functional anti-FAP vaccine that reduced cardiac fibrosis in both the AngII/PE and myocardial infarction models. Although the FAP vaccine was slightly less effective at reducing fibrosis and enhancing functional recovery compared to FAP CAR T cell therapy, CAR T cell therapies carry higher risks of serious toxicities, including cytokine release syndrome and immune effector cell-associated neurotoxicity syndrome⁸. Furthermore, some patients may not be suitable candidates for the lymphodepletion that may be necessary for effective CAR T cell expansion and cytotoxicity¹². In this context, a vaccine-based therapy offers an appealing alternative that is safer, more cost-effective, and easier to produce than CAR T cell therapies.

Regardless of the treatment approach, anti-fibrotic therapies face several significant barriers to clinical use in heart disease¹. The reversibility of cardiac fibrosis remains unclear, though clinical studies suggest that early-stage disease with less fibrosis may be more reversible than advanced disease¹. In this study, mice were vaccinated four weeks prior to model induction. When the FAP vaccine was administered simultaneously with AngII/PE, no reduction in cardiac fibrosis was observed. In contrast, FAP CAR T cells successfully reduced fibrosis when given one week after AngII/PE⁵. Earlier treatment is likely to improve patient outcomes, yet it may also increase the risk of adverse events by disrupting the protective role of myofibroblasts after myocardial injury. For instance, myofibroblast-ablated mice had significantly higher mortality (80%) compared to controls (25%) following experimental myocardial infarction owing to ventricular wall rupture⁴. Both CAR T cells and vaccines have durable effects over months to years and would be administered to patients with risk factors for myocardial infarction. Finding a balance between therapeutic benefit and risk may be particularly challenging with these anti-fibrotic strategies. Some risks might be mitigated new technological innovations, such as T cell-targeted lipid nanoparticles that deliver mRNA for FAP CAR T cell induction in vivo without lymphodepletion¹³.

The development of an FAP-targeted vaccine for cardiac fibrosis has promising implications that extend beyond heart disease. FAP is not only upregulated in cardiac fibrosis but also plays a significant role in the fibrotic processes of other organs, including the lungs, liver, and kidneys. For example, selective elimination of FAP-expressing hepatic stellate cells has been shown to decrease liver fibrosis in mouse models¹⁴, highlighting the potential of FAP-targeted therapies to address fibrotic liver disease. However, the role of FAP-expressing cells in fibrosis is complex and tissue-specific; in pulmonary fibrosis, depleting FAP-expressing cells exacerbates bleomycin-induced fibrosis¹⁵. This underscores the need for a nuanced approach when targeting FAP, as its function varies by tissue type and disease context.

In summary, the development of an FAP-targeted vaccine represents a promising, innovative approach to managing cardiac fibrosis, with the potential to address the limitations of existing CAR T cell therapies. While the vaccine demonstrated efficacy in preclinical models, further research is needed to evaluate its long-term safety, optimal timing, and effectiveness in more advanced models of heart disease. As anti-fibrotic therapies advance toward clinical application, balancing efficacy with safety and finding the ideal window for intervention will be key to translating these therapies from bench to bedside. If successfully implemented, FAP-targeted vaccination could become a valuable tool in the therapeutic arsenal for preventing and managing heart disease, bringing us one step closer to more precise and accessible treatments for cardiac fibrosis.