Thymosin α1 represents a potential potent single-molecule-based therapy for cystic fibrosis

An Author Correction to this article was published on 22 June 2018

A Publisher Correction to this article was published on 22 June 2018

This article has been updated


Cystic fibrosis (CF) is caused by mutations in the gene encoding the cystic fibrosis transmembrane conductance regulator (CFTR) that compromise its chloride channel activity. The most common mutation, p.Phe508del, results in the production of a misfolded CFTR protein, which has residual channel activity but is prematurely degraded. Because of the inherent complexity of the pathogenetic mechanisms involved in CF, which include impaired chloride permeability and persistent lung inflammation, a multidrug approach is required for efficacious CF therapy. To date, no individual drug with pleiotropic beneficial effects is available for CF. Here we report on the ability of thymosin alpha 1 (Tα1)—a naturally occurring polypeptide with an excellent safety profile in the clinic when used as an adjuvant or an immunotherapeutic agent—to rectify the multiple tissue defects in mice with CF as well as in cells from subjects with the p.Phe508del mutation. Tα1 displayed two combined properties that favorably opposed CF symptomatology: it reduced inflammation and increased CFTR maturation, stability and activity. By virtue of this two-pronged action, Tα1 has strong potential to be an efficacious single-molecule-based therapeutic agent for CF.

Access options

Rent or Buy article

Get time limited or full article access on ReadCube.


All prices are NET prices.

Figure 1: Tα1 limits the inflammatory response in cystic fibrosis via IDO1.
Figure 2: Tα1 increases cell surface expression and stability of p.Phe508del-CFTR.
Figure 3: Tα1 rescues p.Phe508del-CFTR by promoting USP36 deubiquitination.
Figure 4: Tα1 rescues p.Phe508del-CFTR functional activity.
Figure 5: Tα1 rescues p.Phe508del-CFTR activity in CftrF508del mice.
Figure 6: Tα1 rescues p.Phe508del-CFTR activity in CF cells and human bronchial epithelial cells from subjects with CF.

Change history

  • 22 June 2018

    In the version of this article originally published, some labels in Fig. 1f are incorrect. The "β-actin" labels on the second and fourth rows of blots should instead be "β-tubulin". The error has been corrected in the HTML and PDF versions of this article.

  • 22 June 2018

    In the version of this article originally published, the amino acid sequence for Tα1 described in the Online Methods is incorrect. The sequence is described as "Ac-SDAAVDTSSEITTJDLKEKKEVVEEAEN-OH". It should be "Ac-SDAAVDTSSEITTKDLKEKKEVVEEAEN-OH". The error has been corrected in the HTML and PDF versions of this article.


  1. 1

    Rowe, S.M., Miller, S. & Sorscher, E.J. Cystic fibrosis. N. Engl. J. Med. 352, 1992–2001 (2005).

    Article  CAS  Google Scholar 

  2. 2

    Lukacs, G.L. & Verkman, A.S. CFTR: folding, misfolding and correcting the ΔF508 conformational defect. Trends Mol. Med. 18, 81–91 (2012).

    Article  CAS  Google Scholar 

  3. 3

    Okiyoneda, T. et al. Peripheral protein quality control removes unfolded CFTR from the plasma membrane. Science 329, 805–810 (2010).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  4. 4

    Pedemonte, N. et al. Small-molecule correctors of defective ΔF508-CFTR cellular processing identified by high-throughput screening. J. Clin. Invest. 115, 2564–2571 (2005).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  5. 5

    Galietta, L.J. Managing the underlying cause of cystic fibrosis: a future role for potentiators and correctors. Paediatr. Drugs 15, 393–402 (2013).

    Article  Google Scholar 

  6. 6

    Wainwright, C.E., Elborn, J.S. & Ramsey, B.W. Lumacaftor–Ivacaftor in patients with cystic fibrosis homozygous for Phe508del CFTR. N. Engl. J. Med. 373, 1783–1784 (2015).

    Article  CAS  Google Scholar 

  7. 7

    Quon, B.S. & Rowe, S.M. New and emerging targeted therapies for cystic fibrosis. Br. Med. J. 352, i859 (2016).

    Article  CAS  Google Scholar 

  8. 8

    Hegde, R.N. et al. Unravelling druggable signalling networks that control F508del-CFTR proteostasis. eLife 4, e10365 (2015).

    Article  PubMed  PubMed Central  Google Scholar 

  9. 9

    Tosco, A. et al. A novel treatment of cystic fibrosis acting on-target: cysteamine plus epigallocatechin gallate for the autophagy-dependent rescue of class II–mutated CFTR. Cell Death Differ. 23, 1380–1393 (2016).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  10. 10

    Villella, V.R. et al. Disease-relevant proteostasis regulation of cystic fibrosis transmembrane conductance regulator. Cell Death Differ. 20, 1101–1115 (2013).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  11. 11

    Cantin, A.M., Hartl, D., Konstan, M.W. & Chmiel, J.F. Inflammation in cystic fibrosis lung disease: pathogenesis and therapy. J. Cyst. Fibros. 14, 419–430 (2015).

    Article  CAS  Google Scholar 

  12. 12

    Rubin, B.K. Cystic fibrosis: myths, mistakes, and dogma. Paediatr. Respir. Rev. 15, 113–116 (2014).

    PubMed  Google Scholar 

  13. 13

    Cohen, T.S. & Prince, A. Cystic fibrosis: a mucosal immunodeficiency syndrome. Nat. Med. 18, 509–519 (2012).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  14. 14

    Hoffman, L.R. & Ramsey, B.W. Cystic fibrosis therapeutics: the road ahead. Chest 143, 207–213 (2013).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  15. 15

    de Benedictis, F.M. & Bush, A. Corticosteroids in respiratory diseases in children. Am. J. Respir. Crit. Care Med. 185, 12–23 (2012).

    Article  CAS  Google Scholar 

  16. 16

    Devor, D.C. & Schultz, B.D. Ibuprofen inhibits cystic fibrosis transmembrane conductance regulator-mediated Cl- secretion. J. Clin. Invest. 102, 679–687 (1998).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  17. 17

    Goldstein, A.L. & Goldstein, A.L. From lab to bedside: emerging clinical applications of thymosin α1. Expert Opin. Biol. Ther. 9, 593–608 (2009).

    Article  CAS  Google Scholar 

  18. 18

    Romani, L. et al. Thymosin α1 activates dendritic cell tryptophan catabolism and establishes a regulatory environment for balance of inflammation and tolerance. Blood 108, 2265–2274 (2006).

    Article  CAS  Google Scholar 

  19. 19

    Mandaliti, W. et al. New studies about the insertion mechanism of thymosin α1 in negative regions of model membranes as starting point of the bioactivity. Amino Acids 48, 1231–1239 (2016).

    Article  CAS  Google Scholar 

  20. 20

    Tuthill, C.V. & King, R.S. Thymosin α1—a peptide immune modulator with a broad range of clinical applications. Clin. Exp. Pharmacol. 3, 133 (2013).

    Google Scholar 

  21. 21

    Puccetti, P. & Grohmann, U. IDO and regulatory T cells: a role for reverse signalling and non-canonical NF-κB activation. Nat. Rev. Immunol. 7, 817–823 (2007).

    Article  CAS  Google Scholar 

  22. 22

    Iannitti, R.G. et al. Th17/Treg imbalance in murine cystic fibrosis is linked to indoleamine 2,3-dioxygenase deficiency but corrected by kynurenines. Am. J. Respir. Crit. Care Med. 187, 609–620 (2013).

    Article  CAS  Google Scholar 

  23. 23

    Latz, E. et al. TLR9 signals after translocating from the ER to CpG DNA in the lysosome. Nat. Immunol. 5, 190–198 (2004).

    Article  CAS  Google Scholar 

  24. 24

    Bruscia, E. et al. Isolation of CF cell lines corrected at ΔF508-CFTR locus by SFHR-mediated targeting. Gene Ther. 9, 683–685 (2002).

    Article  CAS  Google Scholar 

  25. 25

    King, J., Brunel, S.F. & Warris, A. Aspergillus infections in cystic fibrosis. J. Infect. 72 (Suppl. 1), S50–S55 (2016).

    Article  Google Scholar 

  26. 26

    King, R.S. & Tuthill, C. Evaluation of thymosin α1 in nonclinical models of the immune-suppressing indications melanoma and sepsis. Expert Opin. Biol. Ther. 15 (Suppl. 1), S41–S49 (2015).

    Article  CAS  Google Scholar 

  27. 27

    Ancell, C.D., Phipps, J. & Young, L. Thymosin α-1. Am. J. Health Syst. Pharm. 58, 879–885; quiz 886–878 (2001).

    Article  CAS  Google Scholar 

  28. 28

    Iannitti, R.G. et al. IL-1 receptor antagonist ameliorates inflammasome-dependent inflammation in murine and human cystic fibrosis. Nat. Commun. 7, 10791 (2016).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  29. 29

    Snouwaert, J.N. et al. An animal model for cystic fibrosis made by gene targeting. Science 257, 1083–1088 (1992).

    Article  CAS  Google Scholar 

  30. 30

    Stoltz, D.A., Meyerholz, D.K. & Welsh, M.J. Origins of cystic fibrosis lung disease. N. Engl. J. Med. 372, 351–362 (2015).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  31. 31

    McGaha, T.L. IDO–GCN2 and autophagy in inflammation. Oncotarget 6, 21771–21772 (2015).

    Article  PubMed  PubMed Central  Google Scholar 

  32. 32

    Luciani, A. et al. Defective CFTR induces aggresome formation and lung inflammation in cystic fibrosis through ROS-mediated autophagy inhibition. Nat. Cell Biol. 12, 863–875 (2010).

    Article  CAS  Google Scholar 

  33. 33

    Pica, F. et al. Serum thymosin α1 levels in patients with chronic inflammatory autoimmune diseases. Clin. Exp. Immunol. 186, 39–45 (2016).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  34. 34

    Denning, G.M. et al. Processing of mutant cystic fibrosis transmembrane conductance regulator is temperature-sensitive. Nature 358, 761–764 (1992).

    Article  CAS  Google Scholar 

  35. 35

    Bomberger, J.M., Barnaby, R.L. & Stanton, B.A. The deubiquitinating enzyme USP10 regulates the post-endocytic sorting of cystic fibrosis transmembrane conductance regulator in airway epithelial cells. J. Biol. Chem. 284, 18778–18789 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  36. 36

    Gentzsch, M. et al. Endocytic trafficking routes of wild type and ΔF508 cystic fibrosis transmembrane conductance regulator. Mol. Biol. Cell 15, 2684–2696 (2004).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  37. 37

    Sarandeses, C.S., Covelo, G., Díaz-Jullien, C. & Freire, M. Prothymosin α is processed to thymosin α1 and thymosin αl11 by a lysosomal asparaginyl endopeptidase. J. Biol. Chem. 278, 13286–13293 (2003).

    Article  CAS  Google Scholar 

  38. 38

    Heard, A., Thompson, J., Carver, J., Bakey, M. & Wang, X.R. Targeting molecular chaperones for the treatment of cystic fibrosis: is it a viable approach? Curr. Drug Targets 16, 958–964 (2015).

    Article  CAS  Google Scholar 

  39. 39

    Millard, S.M. & Wood, S.A. Riding the DUBway: regulation of protein trafficking by deubiquitylating enzymes. J. Cell Biol. 173, 463–468 (2006).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  40. 40

    Taillebourg, E. et al. The deubiquitinating enzyme USP36 controls selective autophagy activation by ubiquitinated proteins. Autophagy 8, 767–779 (2012).

    Article  CAS  Google Scholar 

  41. 41

    Hassink, G.C. et al. The ER-resident ubiquitin-specific protease 19 participates in the UPR and rescues ERAD substrates. EMBO Rep. 10, 755–761 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  42. 42

    Kucera, A. et al. Spatiotemporal resolution of Rab9 and CI-MPR dynamics in the endocytic pathway. Traffic 17, 211–229 (2016).

    Article  CAS  Google Scholar 

  43. 43

    Lamark, T. & Johansen, T. Autophagy: links with the proteasome. Curr. Opin. Cell Biol. 22, 192–198 (2010).

    Article  CAS  Google Scholar 

  44. 44

    Zhang, L. et al. CFTR delivery to 25% of surface epithelial cells restores normal rates of mucus transport to human cystic fibrosis airway epithelium. PLoS Biol. 7, e1000155 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  45. 45

    Van Goor, F. et al. Correction of the F508del-CFTR protein processing defect in vitro by the investigational drug VX-809. Proc. Natl. Acad. Sci. USA 108, 18843–18848 (2011).

    Article  CAS  Google Scholar 

  46. 46

    Yu, H. et al. Ivacaftor potentiation of multiple CFTR channels with gating mutations. J. Cyst. Fibros. 11, 237–245 (2012).

    Article  CAS  Google Scholar 

  47. 47

    Van Goor, F., Yu, H., Burton, B. & Hoffman, B.J. Effect of ivacaftor on CFTR forms with missense mutations associated with defects in protein processing or function. J. Cyst. Fibros. 13, 29–36 (2014).

    Article  CAS  Google Scholar 

  48. 48

    Caputo, A. et al. TMEM16A, a membrane protein associated with calcium-dependent chloride channel activity. Science 322, 590–594 (2008).

    Article  CAS  Google Scholar 

  49. 49

    Sala-Rabanal, M., Yurtsever, Z., Nichols, C.G. & Brett, T.J. Secreted CLCA1 modulates TMEM16A to activate Ca2+-dependent chloride currents in human cells. eLife 4 (2015).

  50. 50

    Dalakas, M.C., Engel, W.K., McClure, J.E., Goldstein, A.L. & Askanas, V. Immunocytochemical localization of thymosin-α1 in thymic epithelial cells of normal and myasthenia gravis patients and in thymic cultures. J. Neurol. Sci. 50, 239–247 (1981).

    Article  CAS  Google Scholar 

  51. 51

    Collawn, J.F. & Matalon, S. CFTR and lung homeostasis. Am. J. Physiol. Lung Cell. Mol. Physiol. 307, L917–L923 (2014).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  52. 52

    Yuk, J.M. & Jo, E.K. Crosstalk between autophagy and inflammasomes. Mol. Cells 36, 393–399 (2013).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  53. 53

    Soares, M.P., Gozzelino, R. & Weis, S. Tissue damage control in disease tolerance. Trends Immunol. 35, 483–494 (2014).

    Article  CAS  Google Scholar 

  54. 54

    Darrah, R.J. et al. Early pulmonary disease manifestations in cystic fibrosis mice. J. Cyst. Fibros. 15, 736–744 (2016).

    Article  PubMed  PubMed Central  Google Scholar 

  55. 55

    van der Doef, H.P. et al. Association of the CLCA1 p.S357N variant with meconium ileus in European patients with cystic fibrosis. J. Pediatr. Gastroenterol. Nutr. 50, 347–349 (2010).

    Article  CAS  Google Scholar 

  56. 56

    Young, F.D. et al. Amelioration of cystic fibrosis intestinal mucous disease in mice by restoration of mCLCA3. Gastroenterology 133, 1928–1937 (2007).

    Article  CAS  Google Scholar 

  57. 57

    Clarke, L.L. et al. Relationship of a non–cystic fibrosis transmembrane conductance regulator–mediated chloride conductance to organ-level disease in Cftr−/− mice. Proc. Natl. Acad. Sci. USA 91, 479–483 (1994).

    Article  CAS  Google Scholar 

  58. 58

    Boyle, M.P. et al. A CFTR corrector (lumacaftor) and a CFTR potentiator (ivacaftor) for treatment of patients with cystic fibrosis who have a Phe508del CFTR mutation: a phase 2 randomised controlled trial. Lancet Respir. Med. 2, 527–538 (2014).

    Article  CAS  Google Scholar 

  59. 59

    Fajac, I. & De Boeck, K. New horizons for cystic fibrosis treatment. Pharmacol. Ther. 170, 205–211 (2017).

    Article  CAS  Google Scholar 

  60. 60

    Pilewski, J.M., Donaldson, S.H., Cooke, J. & Lekstrom-Himes, J. Phase 2 studies reveal additive effects of VX-661, an investigational CFTR corrector, and ivacaftor, a CFTR potentiator, in patients who carry the ΔF508-CFTR mutation. Pediatr. Pulmonol. 49, 157–159 (2014).

    Google Scholar 

  61. 61

    van Doorninck, J.H. et al. A mouse model for the cystic fibrosis ΔF508 mutation. EMBO J. 14, 4403–4411 (1995).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  62. 62

    De Stefano, D. et al. Restoration of CFTR function in patients with cystic fibrosis carrying the F508del-CFTR mutation. Autophagy 10, 2053–2074 (2014).

    Article  CAS  Google Scholar 

  63. 63

    de Luca, A. et al. Non-hematopoietic cells contribute to protective tolerance to Aspergillus fumigatus via a TRIF pathway converging on IDO. Cell. Mol. Immunol. 7, 459–470 (2010).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  64. 64

    Loffing, J., Moyer, B.D., McCoy, D. & Stanton, B.A. Exocytosis is not involved in activation of Cl secretion via CFTR in Calu-3 airway epithelial cells. Am. J. Physiol. 275, C913–C920 (1998).

    Article  CAS  Google Scholar 

  65. 65

    Pallotta, M.T. et al. Indoleamine 2,3-dioxygenase is a signaling protein in long-term tolerance by dendritic cells. Nat. Immunol. 12, 870–878 (2011).

    Article  CAS  Google Scholar 

  66. 66

    Schägger, H. Tricine–SDS–PAGE. Nat. Protoc. 1, 16–22 (2006).

    Article  CAS  Google Scholar 

  67. 67

    Sowa, M.E., Bennett, E.J., Gygi, S.P. & Harper, J.W. Defining the human deubiquitinating enzyme interaction landscape. Cell 138, 389–403 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  68. 68

    De Luca, A. et al. CD4+ T cell vaccination overcomes defective cross-presentation of fungal antigens in a mouse model of chronic granulomatous disease. J. Clin. Invest. 122, 1816–1831 (2012).

    Article  CAS  Google Scholar 

  69. 69

    Munkonge, F. et al. Measurement of halide efflux from cultured and primary airway epithelial cells using fluorescence indicators. J. Cyst. Fibros. 3 (Suppl. 2), 171–176 (2004).

    Article  CAS  Google Scholar 

Download references


We thank the primary cell culture service offered from the Italian Cystic Fibrosis Research Foundation for kindly providing us with the HBE cells. We thank B. Scholte (Erasmus Medical Center Rotterdam), who provided Cftrtm1EUR mice (F508del mice, European Economic Community European Coordination Action for Research in Cystic Fibrosis program EU FP6 SHMCT-2005-018932). We thank G. Teti (University of Messina, Italy) for providing us with the TLR9-GFP-transfected HEK293 cells. This study was supported by the Specific Targeted Research Project FunMeta (ERC-2011-AdG-293714 to L.R.). M.P. gratefully acknowledges a fellowship from the Italian Cystic Fibrosis Research Foundation.

Author information




V.O., R.G.I. and M. Pariano performed most immunoblotting and immunofluorescence experiments; R.G.I., M.B., S.M. and E.F. performed murine in vivo experiments; M.C.D'A., L.S. and M. Pessia performed electrophysiology experiments; F.F. and M.T.P. performed TLR9 colocalization experiments; M.M.B. and G.S. performed transfection experiments; V.R.V. performed Ussing chamber experiments; and A.L.G., L.M., G.K., M. Pessia, P.P., E.G. and L.R. designed the experiments, analyzed the data and wrote the paper.

Corresponding author

Correspondence to Luigina Romani.

Ethics declarations

Competing interests

A patent application by L.R. and E.G. is pending (filing date, 9 February 2016, RM2015A000056 and 102015000053089).

Supplementary information

Supplementary Figures

Supplementary Figures 1–10. (PDF 2285 kb)

Supplementary Data

Uncropped immunoblots. (PDF 1451 kb)

Rights and permissions

Reprints and Permissions

About this article

Verify currency and authenticity via CrossMark

Cite this article

Romani, L., Oikonomou, V., Moretti, S. et al. Thymosin α1 represents a potential potent single-molecule-based therapy for cystic fibrosis. Nat Med 23, 590–600 (2017).

Download citation

Further reading


Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing