Skip to main content

Advertisement

SpringerLink
Log in

Radiomitigation and Tissue Repair Activity of Systemically Administered Therapeutic Peptide TP508 Is Enhanced by PEGylation

  • Research Article
  • Published:
The AAPS Journal Aims and scope Submit manuscript

Abstract

TP508 is a synthetically derived tissue repair peptide that has previously demonstrated safety and potential efficacy in phase I/II clinical trials for the treatment of diabetic foot ulcers. Recent studies show that a single injection of TP508 administered 24 h after irradiation significantly increases survival and delays mortality in murine models of acute radiation mortality. Thus, TP508 is being developed as a potential nuclear countermeasure. Because of the short plasma half-life of TP508, we hypothesize that increasing the peptide bioavailability would increase TP508 efficacy or reduce the dosage required for therapeutic effects. We, therefore, evaluated the covalent attachment of various sizes of polyethylene glycol to TP508 at either its N-terminus or at an internal cysteine. A size-dependent increase in TP508 plasma half-life due to PEGylation was observed in blood samples from male CD-1 mice using fluorescently labeled TP508 and PEGylated TP508 derivatives. Biological activity of PEGylated TP508 derivatives was evaluated using a combination of biologically relevant assays for wound closure, angiogenesis, and DNA repair. PEG5k-TP508 enhanced wound closure after irradiation and enhanced angiogenic sprouting in murine aortic ring segments relative to equimolar dosages of TP508 without enhancing circulating half-life. PEG30k-TP508 extended the plasma half-life by approximately 19-fold while also showing enhanced biological activity. Intermediate-sized PEGylated TP508 derivatives had enhanced plasma half-life but were not active in vivo. Thus, increased half-life does not necessarily correlate with increased biological activity. Nevertheless, these results identify two candidates, PEG5k-TP508 and PEG30k-TP508, for potential development as second-generation TP508 injectable drugs.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6
Fig. 7
Fig. 8
Fig. 9

Similar content being viewed by others

References

  1. Glenn KC, Frost GH, Bergmann JS, Carney DH. Synthetic peptides bind to high-affinity thrombin receptors and modulate thrombin mitogenesis. Pept Res. 1988;1(2):65–73.

    CAS  PubMed  Google Scholar 

  2. Norfleet AM, Bergmann JS, Carney DH. Thrombin peptide, TP508, stimulates angiogenic responses in animal models of dermal wound healing, in chick chorioallantoic membranes, and in cultured human aortic and microvascular endothelial cells. Gen Pharmacol. 2000;35(5):249–54.

    Article  CAS  PubMed  Google Scholar 

  3. Vartanian KB, Chen HY, Kennedy J, Beck SK, Ryaby JT, Wang H, et al. The non-proteolytically active thrombin peptide TP508 stimulates angiogenic sprouting. J Cell Physiol. 2006;206(1):175–80.

    Article  CAS  PubMed  Google Scholar 

  4. Wang Y, Wan C, Szöke G, Ryaby JT, Li G. Local injection of thrombin-related peptide (TP508) in PPF/PLGA microparticles-enhanced bone formation during distraction osteogenesis. J Orthop Res. 2008;26(4):539–46.

    Article  CAS  PubMed  Google Scholar 

  5. Olszewska-Pazdrak B, Hart-Vantassell A, Carney DH. Thrombin peptide TP508 stimulates rapid nitric oxide production in human endothelial cells. J Vasc Res. 2010;47(3):203–13.

    Article  CAS  PubMed  Google Scholar 

  6. Freyberg S, Song YH, Muehlberg F, Alt E. Thrombin peptide (TP508) promotes adipose tissue-derived stem cell proliferation via PI3 kinase/Akt pathway. J Vasc Res. 2009;46(2):98–102.

    Article  CAS  PubMed  Google Scholar 

  7. Pernia SD, Berry DL, Redin WR, Carney DH. A synthetic peptide representing the thrombin receptor-binding domain enhances wound closure in vivo. SAAS Bull Biochem Biotechnol. 1990;3:8–12.

    CAS  PubMed  Google Scholar 

  8. Norfleet AM, Huang Y, Sower LE, Redin WR, Fritz RR, Carney DH. Thrombin peptide TP508 accelerates closure of dermal excisions in animal tissue with surgically induced ischemia. Wound Repair Regen. 2000;8(6):517–29.

    Article  CAS  PubMed  Google Scholar 

  9. Ryaby JT, Sheller MR, Levine BP, Bramlet DG, Ladd AL, Carney DH. Thrombin peptide TP508 stimulates cellular events leading to angiogenesis, revascularization, and repair of dermal and musculoskeletal tissues. J Bone Joint Surg Am. 2006;88 Suppl 3:132–9.

    PubMed  Google Scholar 

  10. Carney DH, Olszewska-Pazdrak B. Could rusalatide acetate be the future drug of choice for diabetic foot ulcers and fracture repair? Expert Opin Pharmacother. 2008;9(15):2717–26.

    Article  CAS  PubMed  Google Scholar 

  11. Wang H, Li X, Tomin E, Doty SB, Lane JM, Carney DH, et al. Thrombin peptide (TP508) promotes fracture repair by up-regulating inflammatory mediators, early growth factors, and increasing angiogenesis. J Orthop Res. 2005;23(3):671–9.

    Article  CAS  PubMed  Google Scholar 

  12. Hanratty BM, Ryaby JT, Pan XH, Li G. Thrombin related peptide TP508 promoted fracture repair in a mouse high energy fracture model. J Orthop Surg Res. 2009;4:1.

    Article  PubMed  PubMed Central  Google Scholar 

  13. Li X, Wang H, Touma E, Qi Y, Rousseau E, Quigg RJ, et al. TP508 accelerates fracture repair by promoting cell growth over cell death. Biochem Biophys Res Commun. 2007;364(1):187–93.

    Article  CAS  PubMed  Google Scholar 

  14. Oyamada S, Osipov R, Bianchi C, Robich MP, Feng J, Liu Y, et al. Effect of dimerized thrombin fragment TP508 on acute myocardial ischemia reperfusion injury in hypercholesterolemic swine. J Pharmacol Exp Ther. 2010;334(2):449–59.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  15. Chu LM, Osipov RM, Robich MP, Feng J, Sheller MR, Sellke FW. Effect of thrombin fragment (TP508) on myocardial ischemia reperfusion injury in a model of type 1 diabetes mellitus. Circulation. 2010;122(11 Suppl):S162–9.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  16. Osipov RM, Bianchi C, Clements RT, Feng J, Liu Y, Xu SH, et al. Thrombin fragment (TP508) decreases myocardial infarction and apoptosis after ischemia reperfusion injury. Ann Thorac Surg. 2009;87(3):786–93.

    Article  PubMed  Google Scholar 

  17. Fossum TW, Olszewska-Pazdrak B, Mertens MM, Makarski LA, Miller MW, Hein TW, et al. TP508 (Chrysalin) reverses endothelial dysfunction and increases perfusion and myocardial function in hearts with chronic ischemia. J Cardiovasc Pharmacol Ther. 2008;13(3):214–25.

    Article  CAS  PubMed  Google Scholar 

  18. Papareddy P, Rydengård V, Pasupuleti M, Walse B, Mörgelin M, Chalupka A, et al. Proteolysis of human thrombin generates novel host defense peptides. PLoS Pathog. 2010;6(4), e1000857.

    Article  PubMed  PubMed Central  Google Scholar 

  19. Fife C, Mader JT, Stone J, Brill L, Satterfield K, Norfleet A, et al. Thrombin peptide Chrysalin stimulates healing of diabetic foot ulcers in a placebo-controlled phase I/II study. Wound Repair Regen. 2007;15(1):23–34.

    Article  PubMed  Google Scholar 

  20. Kantara C, Moya SM, Houchen CW, Umar S, Ullrich RL, Singh P, et al. Novel regenerative peptide TP508 mitigates radiation-induced gastrointestinal damage by activating stem cells and preserving crypt integrity. Lab Investig. 2015;95(11):1222–33.

  21. Olszewska-Pazdrak B, McVicar SD, Rayavara K, Moya SM, Kantara C, Gammarano C, et al. Nuclear countermeasure activity of TP508 linked to restoration of endothelial function and acceleration of DNA repair. Radiat Res. 2016.

  22. Prasanna PG, Narayanan D, Hallett K, Bernhard EJ, Ahmed MM, Evans G, et al. Radioprotectors and radiomitigators for improving radiation therapy: the small business innovation research (SBIR) gateway for accelerating clinical translation. Radiat Res. 2015;184(3):235–48.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  23. Singh VK, Newman VL, Romaine PL, Wise SY, Seed TM. Radiation countermeasure agents: an update (2011–2014). Expert Opin Ther Pat. 2014;24(11):1229–55.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  24. Li P, Roller PP. Cyclization strategies in peptide derived drug design. Curr Top Med Chem. 2002;2(3):325–41.

    Article  CAS  PubMed  Google Scholar 

  25. Conibear AC, Chaousis S, Durek T, Rosengren KJ, Craik DJ, Schroeder CI. Approaches to the stabilization of bioactive epitopes by grafting and peptide cyclization. Biopolymers. 2016;106(1):89–100.

    Article  CAS  PubMed  Google Scholar 

  26. Berguig GY, Convertine AJ, Frayo S, Kern HB, Procko E, Roy D, et al. Intracellular delivery system for antibody-peptide drug conjugates. Mol Ther. 2015;23(5):907–17.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  27. Gentilucci L. New trends in the development of opioid peptide analogues as advanced remedies for pain relief. Curr Top Med Chem. 2004;4(1):19–38.

    Article  CAS  PubMed  Google Scholar 

  28. Caliceti P, Veronese FM, Jonak Z. Immunogenic and tolerogenic properties of monomethoxypoly(ethylene glycol) conjugated proteins. Farmaco. 1999;54(7):430–7.

    Article  CAS  PubMed  Google Scholar 

  29. Lee SH, Lee S, Youn YS, Na DH, Chae SY, Byun Y, et al. Synthesis, characterization, and pharmacokinetic studies of PEGylated glucagon-like peptide-1. Bioconjug Chem. 2005;16(2):377–82.

    Article  PubMed  Google Scholar 

  30. Meyers FJ, Paradise C, Scudder SA, Goodman G, Konrad M. A phase I study including pharmacokinetics of polyethylene glycol conjugated interleukin-2. Clin Pharmacol Ther. 1991;49(3):307–13.

    Article  CAS  PubMed  Google Scholar 

  31. Thanou M, Duncan R. Polymer-protein and polymer-drug conjugates in cancer therapy. Curr Opin Investig Drugs. 2003;4(6):701–9.

    CAS  PubMed  Google Scholar 

  32. Carpenter AE, Jones TR, Lamprecht MR, Clarke C, Kang IH, Friman O, et al. CellProfiler: image analysis software for identifying and quantifying cell phenotypes. Genome Biol. 2006;7(10):R100.

    Article  PubMed  PubMed Central  Google Scholar 

  33. González JE, Lee M, Barquinero JF, Valente M, Roch-Lefèvre S, García O. Quantitative image analysis of gamma-H2AX foci induced by ionizing radiation applying open source programs. Anal Quant Cytol Histol. 2012;34(2):66–71.

    PubMed  Google Scholar 

  34. Yamaoka T, Tabata Y, Ikada Y. Distribution and tissue uptake of poly(ethylene glycol) with different molecular weights after intravenous administration to mice. J Pharm Sci. 1994;83(4):601–6.

    Article  CAS  PubMed  Google Scholar 

  35. Baker M, Robinson SD, Lechertier T, Barber PR, Tavora B, D’Amico G, et al. Use of the mouse aortic ring assay to study angiogenesis. Nat Protoc. 2012;7(1):89–104.

    Article  CAS  Google Scholar 

  36. Rogakou EP, Pilch DR, Orr AH, Ivanova VS, Bonner WM. DNA double-stranded breaks induce histone H2AX phosphorylation on serine 139. J Biol Chem. 1998;273(10):5858–68.

    Article  CAS  PubMed  Google Scholar 

  37. Osipov RM, Robich MP, Feng J, Clements RT, Liu Y, Glazer HP, et al. Effect of thrombin fragment (TP508) on myocardial ischemia-reperfusion injury in hypercholesterolemic pigs. J Appl Physiol. 2009;106(6):1993–2001.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  38. Kantara C, Moya SM, Houchen CW, Umar S, Ullrich RL, Singh P, et al. Novel regenerative peptide TP508 mitigates radiation-induced gastrointestinal damage by activating stem cells and preserving crypt integrity. Lab Investig. 2015;95(11):1222–33.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  39. Caliceti P, Veronese FM. Pharmacokinetic and biodistribution properties of poly(ethylene glycol)-protein conjugates. Adv Drug Deliv Rev. 2003;55(10):1261–77.

    Article  CAS  PubMed  Google Scholar 

  40. Harris JM, Chess RB. Effect of pegylation on pharmaceuticals. Nat Rev Drug Discov. 2003;2(3):214–21.

    Article  CAS  PubMed  Google Scholar 

  41. Wang J, Boerma M, Fu Q, Hauer-Jensen M. Radiation responses in skin and connective tissues: effect on wound healing and surgical outcome. Hernia. 2006;10(6):502–6.

    Article  PubMed  Google Scholar 

  42. Rudolph R, Vande Berg J, Schneider JA, Fisher JC, Poolman WL. Slowed growth of cultured fibroblasts from human radiation wounds. Plast Reconstr Surg. 1988;82(4):669–77.

    Article  CAS  PubMed  Google Scholar 

  43. Qu JF, Cheng TM, Xu LS, Shi CM, Ran XZ. Effects of total body irradiation injury on the participation of dermal fibroblasts in tissue repair. Sheng Li Xue Bao. 2002;54(5):395–9.

    PubMed  Google Scholar 

  44. Denham JW, Hauer-Jensen M. The radiotherapeutic injury—a complex ‘wound’. Radiother Oncol. 2002;63(2):129–45.

    Article  PubMed  Google Scholar 

  45. Tibbs MK. Wound healing following radiation therapy: a review. Radiother Oncol. 1997;42(2):99–106.

    Article  CAS  PubMed  Google Scholar 

Download references

Acknowledgments

The authors would like to acknowledge Dr. Gerald M. Fuller (University of Alabama) and Dr. Laurie M. Sower (Chrysalis BioTherapeutics, Inc.) for their encouragement during this project and editorial assistance; Dr. Kimberly Burckart and Dr. Barbara Olszewska-Pazdrak, members of the Carney Laboratory, and Dr. Bradford Loucas, at the University of Texas Medical Branch, for assistance with animal studies. This work was supported by NIH/NIAID Grant 5R44AI086135.

Author information

Authors and Affiliations

  1. Department of Biochemistry and Molecular Biology, The University of Texas Medical Branch, 301 University Blvd., Galveston, Texas, 77555, USA

    Scott D. McVicar & Darrell H. Carney

  2. Chrysalis BioTherapeutics, Inc., Galveston, Texas, USA

    Kempaiah Rayavara & Darrell H. Carney

Authors
  1. Scott D. McVicar
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Kempaiah Rayavara
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Darrell H. Carney
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Scott D. McVicar.

Ethics declarations

Conflict of Interest

Chrysalis BioTherapeutics has licensed worldwide exclusive rights to TP508 from The University of Texas Medical Branch. DHC and KR receive compensation from Chrysalis BioTherapeutics or have stock or stock options in the company. Potential conflicts of interest are managed by the University of Texas Medical Branch Conflicts of Interest and Commitment Committee.

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

McVicar, S.D., Rayavara, K. & Carney, D.H. Radiomitigation and Tissue Repair Activity of Systemically Administered Therapeutic Peptide TP508 Is Enhanced by PEGylation. AAPS J 19, 743–753 (2017). https://doi.org/10.1208/s12248-016-0043-7

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1208/s12248-016-0043-7

KEY WORDS

Search

Navigation