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Targeting cardiac fibrosis with engineered T cells

An Author Correction to this article was published on 14 November 2019

This article has been updated

Abstract

Fibrosis is observed in nearly every form of myocardial disease1. Upon injury, cardiac fibroblasts in the heart begin to remodel the myocardium by depositing excess extracellular matrix, resulting in increased stiffness and reduced compliance of the tissue. Excessive cardiac fibrosis is an important factor in the progression of various forms of cardiac disease and heart failure2. However, clinical interventions and therapies that target fibrosis remain limited3. Here we demonstrate the efficacy of redirected T cell immunotherapy to specifically target pathological cardiac fibrosis in mice. We find that cardiac fibroblasts that express a xenogeneic antigen can be effectively targeted and ablated by adoptive transfer of antigen-specific CD8+ T cells. Through expression analysis of the gene signatures of cardiac fibroblasts obtained from healthy and diseased human hearts, we identify an endogenous target of cardiac fibroblasts—fibroblast activation protein. Adoptive transfer of T cells that express a chimeric antigen receptor against fibroblast activation protein results in a significant reduction in cardiac fibrosis and restoration of function after injury in mice. These results provide proof-of-principle for the development of immunotherapeutic drugs for the treatment of cardiac disease.

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Fig. 1: Redirected T cells can ablate cardiac fibroblasts.
Fig. 2: Human cardiac fibroblast targets in disease.
Fig. 3: FAP CAR T cells can target cardiac fibrosis.

Data availability

All data are available from the corresponding author upon request.

Change history

  • 14 November 2019

    An Amendment to this paper has been published and can be accessed via a link at the top of the paper.

References

  1. 1.

    Travers, J. G., Kamal, F. A., Robbins, J., Yutzey, K. E. & Blaxall, B. C. Cardiac fibrosis: the fibroblast awakens. Circ. Res. 118, 1021–1040 (2016).

    CAS  PubMed  PubMed Central  Google Scholar 

  2. 2.

    Kong, P., Christia, P. & Frangogiannis, N. G. The pathogenesis of cardiac fibrosis. Cell. Mol. Life Sci. 71, 549–574 (2014).

    CAS  PubMed  Google Scholar 

  3. 3.

    Fang, L., Murphy, A. J. & Dart, A. M. A clinical perspective of anti-fibrotic therapies for cardiovascular disease. Front. Pharmacol. 8, 186 (2017).

    PubMed  PubMed Central  Google Scholar 

  4. 4.

    Ottaviano, F. G. & Yee, K. O. Communication signals between cardiac fibroblasts and cardiac myocytes. J. Cardiovasc. Pharmacol. 57, 513–521 (2011).

    CAS  PubMed  Google Scholar 

  5. 5.

    Fan, Z. & Guan, J. Antifibrotic therapies to control cardiac fibrosis. Biomater. Res. 20, 13 (2016).

    PubMed  PubMed Central  Google Scholar 

  6. 6.

    Lam, C. S. P., Voors, A. A., de Boer, R. A., Solomon, S. D. & van Veldhuisen, D. J. Heart failure with preserved ejection fraction: from mechanisms to therapies. Eur. Heart J. 39, 2780–2792 (2018).

    CAS  PubMed  Google Scholar 

  7. 7.

    Kaur, H. et al. Targeted ablation of periostin-expressing activated fibroblasts prevents adverse cardiac remodeling in mice. Circ. Res. 118, 1906–1917 (2016).

    CAS  PubMed  Google Scholar 

  8. 8.

    Kanisicak, O. et al. Genetic lineage tracing defines myofibroblast origin and function in the injured heart. Nat. Commun. 7, 12260 (2016).

    ADS  CAS  PubMed  PubMed Central  Google Scholar 

  9. 9.

    Schmitt, T. M., Ragnarsson, G. B. & Greenberg, P. D. T cell receptor gene therapy for cancer. Hum. Gene Ther. 20, 1240–1248 (2009).

    CAS  PubMed  PubMed Central  Google Scholar 

  10. 10.

    June, C. H., O’Connor, R. S., Kawalekar, O. U., Ghassemi, S. & Milone, M. C. CAR T cell immunotherapy for human cancer. Science 359, 1361–1365 (2018).

    ADS  CAS  Google Scholar 

  11. 11.

    Mullard, A. FDA approves first CAR T therapy. Nat. Rev. Drug Discov. 16, 669 (2017).

    PubMed  Google Scholar 

  12. 12.

    Ghobadi, A. Chimeric antigen receptor T cell therapy for non-Hodgkin lymphoma. Curr. Res. Transl. Med. 66, 43–49 (2018).

    PubMed  PubMed Central  Google Scholar 

  13. 13.

    June, C. H. & Sadelain, M. Chimeric antigen receptor therapy. N. Engl. J. Med. 379, 64–73 (2018).

    CAS  PubMed  Google Scholar 

  14. 14.

    Lim, W. A. & June, C. H. The principles of engineering immune cells to treat cancer. Cell 168, 724–740 (2017).

    CAS  PubMed  PubMed Central  Google Scholar 

  15. 15.

    Sandhu, U. et al. Strict control of transgene expression in a mouse model for sensitive biological applications based on RMCE compatible ES cells. Nucleic Acids Res. 39, e1 (2011).

    CAS  PubMed  Google Scholar 

  16. 16.

    Cebula, M. et al. An inducible transgenic mouse model for immune mediated hepatitis showing clearance of antigen expressing hepatocytes by CD8+ T cells. PLoS ONE 8, e68720 (2013).

    ADS  CAS  PubMed  PubMed Central  Google Scholar 

  17. 17.

    Hogquist, K. A. et al. T cell receptor antagonist peptides induce positive selection. Cell 76, 17–27 (1994).

    CAS  PubMed  Google Scholar 

  18. 18.

    Clarke, S. R. et al. Characterization of the ovalbumin-specific TCR transgenic line OT-I: MHC elements for positive and negative selection. Immunol. Cell Biol. 78, 110–117 (2000).

    CAS  PubMed  Google Scholar 

  19. 19.

    Ivey, M. J. & Tallquist, M. D. Defining the cardiac fibroblast. Circ. J. 80, 2269–2276 (2016).

    CAS  PubMed  PubMed Central  Google Scholar 

  20. 20.

    Tallquist, M. D. & Molkentin, J. D. Redefining the identity of cardiac fibroblasts. Nat. Rev. Cardiol. 14, 484–491 (2017).

    PubMed  PubMed Central  Google Scholar 

  21. 21.

    Scanlan, M. J. et al. Molecular cloning of fibroblast activation protein α, a member of the serine protease family selectively expressed in stromal fibroblasts of epithelial cancers. Proc. Natl Acad. Sci. USA 91, 5657–5661 (1994).

    ADS  CAS  PubMed  Google Scholar 

  22. 22.

    Rettig, W. J. et al. Cell-surface glycoproteins of human sarcomas: differential expression in normal and malignant tissues and cultured cells. Proc. Natl Acad. Sci. USA 85, 3110–3114 (1988).

    ADS  CAS  PubMed  Google Scholar 

  23. 23.

    Niedermeyer, J. et al. Mouse fibroblast activation protein: molecular cloning, alternative splicing and expression in the reactive stroma of epithelial cancers. Int. J. Cancer 71, 383–389 (1997).

    CAS  PubMed  Google Scholar 

  24. 24.

    Tillmanns, J. et al. Fibroblast activation protein α expression identifies activated fibroblasts after myocardial infarction. J. Mol. Cell. Cardiol. 87, 194–203 (2015).

    CAS  PubMed  Google Scholar 

  25. 25.

    Wang, L. C. et al. Targeting fibroblast activation protein in tumor stroma with chimeric antigen receptor T cells can inhibit tumor growth and augment host immunity without severe toxicity. Cancer Immunol. Res. 2, 154–166 (2014).

    CAS  PubMed  Google Scholar 

  26. 26.

    Kakarla, S. et al. Antitumor effects of chimeric receptor engineered human T cells directed to tumor stroma. Mol. Ther. 21, 1611–1620 (2013).

    CAS  PubMed  PubMed Central  Google Scholar 

  27. 27.

    Lo, A. et al. Tumor-promoting desmoplasia is disrupted by depleting FAP-expressing stromal cells. Cancer Res. 75, 2800–2810 (2015).

    CAS  PubMed  PubMed Central  Google Scholar 

  28. 28.

    Schuberth, P. C. et al. Treatment of malignant pleural mesothelioma by fibroblast activation protein-specific re-directed T cells. J. Transl. Med. 11, 187 (2013).

    CAS  PubMed  PubMed Central  Google Scholar 

  29. 29.

    Petrausch, U. et al. Re-directed T cells for the treatment of fibroblast activation protein (FAP)-positive malignant pleural mesothelioma (FAPME-1). BMC Cancer 12, 615 (2012).

    CAS  PubMed  PubMed Central  Google Scholar 

  30. 30.

    Govindaraju, P., Todd, L., Shetye, S., Monslow, J. & Puré, E. CD44-dependent inflammation, fibrogenesis, and collagenolysis regulates extracellular matrix remodeling and tensile strength during cutaneous wound healing. Matrix Biol. 75–76, 314–330 (2019).

    PubMed  Google Scholar 

  31. 31.

    Croft, A. P. et al. Distinct fibroblast subsets drive inflammation and damage in arthritis. Nature 570, 246–251 (2019).

    ADS  CAS  PubMed  Google Scholar 

  32. 32.

    Fischbach, M. A., Bluestone, J. A. & Lim, W. A. Cell-based therapeutics: the next pillar of medicine. Sci. Transl. Med. 5, 179ps7 (2013).

    PubMed  PubMed Central  Google Scholar 

  33. 33.

    Tran, E. et al. Immune targeting of fibroblast activation protein triggers recognition of multipotent bone marrow stromal cells and cachexia. J. Exp. Med. 210, 1125–1135 (2013).

    CAS  PubMed  PubMed Central  Google Scholar 

  34. 34.

    Cameron, B. J. et al. Identification of a titin-derived HLA-A1-presented peptide as a cross-reactive target for engineered MAGE A3-directed T cells. Sci. Transl. Med. 5, 197ra103 (2013).

    PubMed  PubMed Central  Google Scholar 

  35. 35.

    Linette, G. P. et al. Cardiovascular toxicity and titin cross-reactivity of affinity-enhanced T cells in myeloma and melanoma. Blood 122, 863–871 (2013).

    CAS  PubMed  PubMed Central  Google Scholar 

  36. 36.

    Sun, S., Hao, H., Yang, G., Zhang, Y. & Fu, Y. Immunotherapy with CAR-modified T Cells: toxicities and overcoming strategies. J. Immunol. Res. 2018, 2386187 (2018).

    PubMed  PubMed Central  Google Scholar 

  37. 37.

    Ochel, A. et al. Effective intrahepatic CD8+ T-cell immune responses are induced by low but not high numbers of antigen-expressing hepatocytes. Cell. Mol. Immunol. 13, 805–815 (2016).

    CAS  PubMed  Google Scholar 

  38. 38.

    Mourkioti, F. et al. Role of telomere dysfunction in cardiac failure in Duchenne muscular dystrophy. Nat. Cell Biol. 15, 895–904 (2013).

    CAS  PubMed  PubMed Central  Google Scholar 

  39. 39.

    Newick, K. et al. Augmentation of CAR T-cell trafficking and antitumor efficacy by blocking protein kinase A localization. Cancer Immunol. Res. 4, 541–551 (2016).

    CAS  PubMed  PubMed Central  Google Scholar 

  40. 40.

    Dipla, K., Mattiello, J. A., Jeevanandam, V., Houser, S. R. & Margulies, K. B. Myocyte recovery after mechanical circulatory support in humans with end-stage heart failure. Circulation 97, 2316–2322 (1998).

    CAS  PubMed  Google Scholar 

  41. 41.

    Chen, C. Y. et al. Suppression of detyrosinated microtubules improves cardiomyocyte function in human heart failure. Nat. Med. 24, 1225–1233 (2018).

    CAS  PubMed  Google Scholar 

  42. 42.

    Dobin, A. et al. STAR: ultrafast universal RNA-seq aligner. Bioinformatics 29, 15–21 (2013).

    CAS  PubMed  PubMed Central  Google Scholar 

  43. 43.

    Law, C. W., Chen, Y., Shi, W. & Smyth, G. K. voom: precision weights unlock linear model analysis tools for RNA-seq read counts. Genome Biol. 15, R29 (2014).

    PubMed  PubMed Central  Google Scholar 

  44. 44.

    Love, M. I., Huber, W. & Anders, S. Moderated estimation of fold change and dispersion for RNA-seq data with DESeq2. Genome Biol. 15, 550 (2014).

    PubMed  PubMed Central  Google Scholar 

  45. 45.

    Benjamini, Y. & Hochberg, Y. Controlling the false discovery rate: a practical and powerful approach to multiple testing. J. R. Stat. Soc. B 57, 289–300 (1995).

    MathSciNet  MATH  Google Scholar 

Download references

Acknowledgements

We thank J. Molkentin for sharing the PostnMCM mice with us, A. Stout for help with imaging, D. Martinez for help with image analysis, M. Scherrer-Crosbie for guidance on the echocardiogram procedure and interpretation, F. Mourkioti for sharing the mdx/mTRKO mice with us, E. Radaelli for help with immunohistochemistry and the University of Pennsylvania Diabetes Research Center (DRC) for the use of the RIA Biomarker Core (P30-DK19525). This research was supported by NIH R35 HL140018, T32 HL007843-22, F31 HL147416, the Burroughs Wellcome Fund to R.J., the Cotswold Foundation and the W. W. Smith Endowed Chair to J.A.E.

Author information

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Authors

Contributions

H.A. and J.A.E. conceived the project and designed experiments. H.A., T.K., J.G.R., A.S.H., M.S.L., L.L., J.M., A.L., W.H., T.W., K.B., R.A.L.S., N.A.B., K.M., C.-A.A. and C.L.S. performed experiments and interpreted data. M.P.M. performed bioinformatic analysis and interpretation. J.S., R.J., D.W., C.H.J., K.B.M., E.P. and S.M.A. contributed reagents, analysis and interpretation. H.A. and J.A.E. wrote the manuscript. J.A.E. supervised all aspects of the research.

Corresponding author

Correspondence to Jonathan A. Epstein.

Ethics declarations

Competing interests

C.H.J., E.P. and S.M.A. are inventors (University of Pennsylvania, Wistar Institute) on a patent for a FAP CAR (US Utility Patent 9,365,641 issued 14 June 2016, WIPO Patent Application PCT/US2013/062717). H.A. and J.A.E. are inventors (University of Pennsylvania) on a patent for the use of CAR T therapy in heart disease (US Provisional Patent Application 62/563,323 filed 26 September 2017, WIPO Patent Application PCT/US2018/052605). C.H.J. is a scientific founder and has equity in Tmunity Therapeutics, a biotech dedicated to developing engineered T cells for therapy of cancer, infections and autoimmunity, reports grants from Novartis and Tmunity Therapeutics and is on the scientific advisory boards of Immune Design, Viracta Therapeutics, Carisma Therapeutics and Cabaletta Bio.

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Publisher’s note: Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Peer review information Nature thanks Jeffery D. Molkentin, Richard T. Lee and the other, anonymous, reviewer(s) for their contribution to the peer review of this work.

Extended data figures and tables

Extended Data Fig. 1 Cardiac fibrosis and hypertrophy.

a, Picro-Sirius red staining for cardiac fibrosis (red) in a coronal section of the heart of a PostnMCM;RosaOVA mouse treated with AngII/PE and tamoxifen for 1 week (left). High-powered field of the left ventricular free wall (right). Representative image of three biologically independent mice, showing similar results. Scale bars, 100 μm. b, Control and experimental hearts were measured (weight (mg)) and images captured. Representative images are shown. c, Quantification of heart weight to body weight (HW/BW) ratio of indicated genotypes and conditions. Data are mean ± s.e.m. ****P < 0.0001 (one-way ANOVA between groups, P < 0.0001; post hoc test for multiple comparisons, Tukey’s test; n = 10, 7, 6, 8 biologically independent mice from left to right).

Source data

Extended Data Fig. 2 Markers of activated cardiac fibroblasts in human disease.

a, Fold change and P values of cardiac-fibroblast-specific gene expression of heart samples from patients with HCM and DCM compared with samples from non-failing hearts. n = 122 (non-failing), 27 (HCM) and 89 (DCM). Differential gene-expression analyses were performed using a linear model. b, Immunohistochemistry co-staining of cardiac troponin (red) and FAP (green; left) and vimentin (green; right) in adjacent sections from the left ventricle of a patient with DCM. FAP and vimentin are expressed in the same fibroblasts (arrowheads). Representative images of two independent experiments, showing similar results. Scale bars, 100 μm.

Extended Data Fig. 3 FAP is expression in mouse cardiac fibroblasts after injury.

a, Immunohistochemistry co-staining of cardiac troponin (red) and FAP (green; left) and vimentin (green; right) in adjacent sections from the left ventricle of a mouse treated with AngII/PE for 2 weeks. FAP and vimentin are expressed in the same fibroblasts (arrowheads). Representative images from two independent experiments, showing similar results (n = 7 biologically independent mice). b, Immunohistochemistry of FAP (green) in various organs or tissues after 1 week of AngII/PE treatment. Representative image of n = 3 biologically independent mice, showing similar results. c, Masson’s trichrome stain for fibrosis (blue; top, centre) and immunohistochemistry of FAP (green; bottom) in coronal heart sections of a wild-type mouse 2 weeks after continuous AngII/PE treatment. Staining and immunohistochemistry were performed on adjacent sections. Bottom insets, higher magnification of left ventricular free wall. Representative image of two independent experiments, showing similar results (n = 7 biologically independent mice). d, Immunohistochemistry of FAP (green) in mouse models of cardiac injury. DMD, Duchenne’s muscular dystrophy (mdx/mTRKO G2 mice); MI, myocardial infarction; TAC, transverse aortic constriction. Scale bars, 100 μm.

Extended Data Fig. 4 FAP CAR T cells infiltrate the heart and reduce cardiac fibrosis.

a, Immunohistochemistry of FAP (red) and GFP (green) on the left ventricular free wall of mouse heart coronal sections. Wild-type C57BL/6 mice were treated with (right) or without (left) AngII/PE for 1 week, injected with FAP–GFP CAR T cells and euthanized 1 day later. FAP–GFP CAR T cells co-localize with FAP-expressing cells (arrowheads, bottom). bd, Picro-Sirius red staining of hearts from 21 individual mice (1–21) treated for 4 weeks with saline (b), AngII/PE (c) or AngII/PE and FAP CAR T cells (d) to assess fibrosis (red). Representative images of two independent experiments, showing similar results. Scale bars, 100 μm.

Extended Data Fig. 5 Echocardiography after injury and treatment.

Results from echocardiogram analyses of C57BL/6 mice treated for 4 weeks with saline, AngII/PE or AngII/PE and FAP CAR T cells (n = 12, 10 and 7 biologically independent mice, respectively). Data are mean ± s.e.m. CO, cardiac output; E/E′, ratio of mitral peak velocity of early filling (E) to early diastolic mitral annular velocity (E′); EDV, end diastolic volume; ESV, end systolic volume; FS, fractional shortening; HR, heart rate; IVSd, interventricular septal end diastole; IVSs, interventricular septal end systole; LVIDd, left ventricular internal diameter end diastole; LVIDs, left ventricular internal diameter end systole; LVLd, left ventricular endocardial length (diastole); LVLs, left ventricular endocardial length (systole); LVAEpid, left ventricular epicardial area (diastole); LVAENDd, left ventricular endocardial area (diastole); LVAENDs, left ventricular endocardial area (systole); MV E, early ventricular filling velocity; SV, stroke volume.

Source data

Extended Data Fig. 6 FAP CAR T treatment does not affect perivascular fibrosis or other organs.

a, Masson’s trichrome stain (blue; left, centre) and FAP immunohistochemistry (green; right) on adjacent heart coronal sections 1 week after commencement of continuous AngII/PE treatment. FAP expression is present in interstitial, but not perivascular, fibroblasts (white arrowheads). Image is centred on the vessel shown in Fig. 2c. Representative images of two independent experiments, showing similar results. b, Picro-Sirius red staining of perivascular fibrosis (black arrowheads, red) in heart coronal sections from mice treated for 4 weeks with saline, AngII/PE or AngII/PE and FAP CAR T cells. Representative images of two independent experiments with similar results. c, H&E staining of various tissue sections from mice treated for 4 weeks with saline, AngII/PE or AngII/PE and FAP CAR T cells. Representative images of two independent experiments, showing similar results. Scale bars, 100 μm.

Extended Data Fig. 7 Long-term serum cytokine levels after FAP CAR T cell treatment.

Serum cytokine levels in mice treated with either AngII/PE or AngII/PE and FAP CAR T cells over 12 weeks. FAP CAR T cells were injected at 1 and 2 weeks as indicated. Levels were assessed at 10 days, 2 weeks, 4 weeks and 12 weeks. Basal levels were determined by the average cytokine levels of three untreated mice. INFγ and IL-4 were below the limit of detection in all conditions. *P = 0.019, #P = 0.035, P = 0.045; two-tailed unpaired Student’s t-test; n = 3 biologically independent mice per condition.

Source data

Extended Data Fig. 8 Cardiotoxicity, inflammation and immune assessments after FAP CAR T cell transfer.

a, Volcano plot showing the differential expression of genes known to be modified in cardiotoxicity in the hearts of mice treated with either AngII/PE and FAP CAR T cells or a saline control for 4 weeks. Statistically significant changes are marked to indicate whether genes are expected to increase (orange) or decrease (blue) in the setting of cardiotoxicity. b, Differential expression of the same conditions in a at 8 weeks. a, b, Two-sided Welch’s t-test; n = 3 biologically independent mice per condition. c, Volcano plot showing the differential expression of 1,659 immune- and inflammation-related genes from hearts of mice treated with AngII/PE and FAP CAR T cells or AngII/PE for 4 weeks. In total, 22 genes were differentially expressed between the conditions. n = 3 mice per condition. d, Photomicrographs and quantification (mean ± s.e.m.) of immune cell (arrowheads) residency of the left ventricle at 4 weeks after either AngII/PE or AngII/PE and FAP CAR T cell treatment. Two-tailed unpaired Student’s t-test; n = 3 or 4 biologically independent mice, respectively. Scale bars, 100 μm.

Source data

Extended Data Fig. 9 Assessment of safety and toxicity following FAP CAR T cell transfer.

a, Kaplan–Meier survival curve of mice treated with either AngII/PE or AngII/PE and FAP CAR T cells for 12 weeks. b, Body weight measurements at 12 weeks. Two-tailed unpaired Student’s t-test. c, H&E of sections of heart and organs or tissues at 12 weeks from a mouse treated with AngII/PE and FAP CAR T cells. Representative images of n = 3 independent mice, showing similar results. Scale bars, 100 μm. d, Photomicrographs and H&E sections of a healing wound over 8 days in mice treated with either FAP CAR or control T cells immediately and 3 days after wounding. Scale bars, 1 mm (wounds) and 250 μm (H&E sections). e, f, Quantification of wound area (e) and measurements of body weight (f). g, Serum levels of amylase at day 8 to test pancreatic toxicity. Data are mean ± s.e.m.

Source data

Supplementary information

Supplementary Table 1

List of published genes expressed at high levels in cardiac fibroblasts with their corresponding tissue specificity and subcellular localization.

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Aghajanian, H., Kimura, T., Rurik, J.G. et al. Targeting cardiac fibrosis with engineered T cells. Nature 573, 430–433 (2019). https://doi.org/10.1038/s41586-019-1546-z

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