Pharmaceutical Research

, Volume 34, Issue 12, pp 2698–2709 | Cite as

Targeted Metabolomics Identifies Pharmacodynamic Biomarkers for BIO 300 Mitigation of Radiation-Induced Lung Injury

  • Jace W. Jones
  • Isabel L. Jackson
  • Zeljko Vujaskovic
  • Michael D. Kaytor
  • Maureen A. KaneEmail author
Research Paper



Biomarkers serve a number of purposes during drug development including defining the natural history of injury/disease, serving as a secondary endpoint or trigger for intervention, and/or aiding in the selection of an effective dose in humans. BIO 300 is a patent-protected pharmaceutical formulation of nanoparticles of synthetic genistein being developed by Humanetics Corporation. The primary goal of this metabolomic discovery experiment was to identify biomarkers that correlate with radiation-induced lung injury and BIO 300 efficacy for mitigating tissue damage based upon the primary endpoint of survival.


High-throughput targeted metabolomics of lung tissue from male C57L/J mice exposed to 12.5 Gy whole thorax lung irradiation, treated daily with 400 mg/kg BIO 300 for either 2 weeks or 6 weeks starting 24 h post radiation exposure, were assayed at 180 d post-radiation to identify potential biomarkers.


A panel of lung metabolites that are responsive to radiation and able to distinguish an efficacious treatment schedule of BIO 300 from a non-efficacious treatment schedule in terms of 180 d survival were identified.


These metabolites represent potential biomarkers that could be further validated for use in drug development of BIO 300 and in the translation of dose from animal to human.


biomarkers genistein lung injury metabolomics radiation 



Acute radiation syndrome


Collision-induced dissociation


Delayed effects of acute radiation exposure


Federal Drug Administration


False discovery rate


Flow injection analysis


Hematoxylin and eosin


High definition mass spectrometry


High-performance liquid chromatography


Liquid chromatography


Lethal dose


Medical countermeasure


Multiple reaction monitoring






Diacyl glycerophosphatidylcholine


Principal component analysis


Ether glycerophosphatidylcholine






Partial least squares-discriminate analysis


Polyunsaturated fatty acid


Standard error of the mean


Saturated fatty acid




Total ion chromatogram


Ultra performance liquid chromatography


Extracted ion chromatogram


Whole thorax lung irradiation

Supplementary material

11095_2017_2200_MOESM1_ESM.docx (2.6 mb)
ESM 1 (DOCX 2622 kb)


  1. 1.
    Dorr H, Meineke V. Acute radiation syndrome caused by accidental radiation exposure - therapeutic principles. BMC med. 2011;9:126.CrossRefPubMedPubMedCentralGoogle Scholar
  2. 2.
    MacVittie TJ, Farese AM, Jackson W III. The hematopoietic syndrome of the acute radiation syndrome in rhesus macaques: a systematic review of the lethal dose response relationship. Health Phys. 2015;109(5):342–66.CrossRefPubMedGoogle Scholar
  3. 3.
    Van Dyk J, Keane TJ, Kan S, Rider WD, Fryer CJ. Radiation pneumonitis following large single dose irradiation: a re-evaluation based on absolute dose to lung. Int J Radiat Oncol Biol Phys. 1981;7(4):461–7.CrossRefPubMedGoogle Scholar
  4. 4.
    Mah K, Van Dyk J. Quantitative measurement of changes in human lung density following irradiation. Radiother Oncol. 1988;11(2):169–79.CrossRefPubMedGoogle Scholar
  5. 5.
    Day RM, Barshishat-Kupper M, Mog SR, McCart EA, Prasanna PG, Davis TA, et al. Genistein protects against biomarkers of delayed lung sequelae in mice surviving high-dose total body irradiation. J Radiat res. 2008;49(4):361–72.CrossRefPubMedPubMedCentralGoogle Scholar
  6. 6.
    Calveley VL, Jelveh S, Langan A, Mahmood J, Yeung IW, Van Dyk J, et al. Genistein can mitigate the effect of radiation on rat lung tissue. Radiat res. 2010;173(5):602–11.CrossRefPubMedPubMedCentralGoogle Scholar
  7. 7.
    Yang Z, Kulkarni K, Zhu W, Hu M. Bioavailability and pharmacokinetics of genistein: mechanistic studies on its ADME. Anti Cancer Agents med Chem. 2012;12(10):1264–80.CrossRefGoogle Scholar
  8. 8.
    Ha CT, Li XH, Fu D, Xiao M, Landauer MR. Genistein nanoparticles protect mouse hematopoietic system and prevent proinflammatory factors after gamma irradiation. Radiat res. 2013;180(3):316–25.CrossRefPubMedGoogle Scholar
  9. 9.
    Jackson IL, Zodda A, Gurung G, Pavlovic R, Kaytor MD, Kuskowski MA, Vujaskovic Z. BIO 300, a nanosuspension of Genistein, mitigates pneumonitis/fibrosis following high dose radiation exposure in the C57L/J murine model. Br J Pharmacol 2017. (in review).Google Scholar
  10. 10.
    Jackson IL, Xu P, Hadley C, Katz BP, McGurk R, Down JD, et al. A preclinical rodent model of radiation-induced lung injury for medical countermeasure screening in accordance with the FDA animal rule. Health Phys. 2012;103(4):463–73.CrossRefPubMedPubMedCentralGoogle Scholar
  11. 11.
    Product development under the animal rule: Guidance for industry. U.S. Department of Health and Human Services, Food and Drug Administration (FDA), Center for Drug Evaluation and Research (CDER), Center for Biologics Evaluation and Research (CBER). 2015. Available from:
  12. 12.
    Guidance for industry and FDA staff: Qualification process for drug development tools. U.S. Department of Health and Human Services, Food and Drug Administration (FDA), Center for Drug Evaluation and Research (CDER), Center for Biologics Evaluation and Research (CBER). 2014. Available from:
  13. 13.
    Biomarkers Definitions Working G. Biomarkers and surrogate endpoints: preferred definitions and conceptual framework. Clin Pharmacol Ther. 2001;69(3):89–95.CrossRefGoogle Scholar
  14. 14.
    Jones JW, Carter CL, Li F, Yu J, Pierzchalski K, Jackson IL, et al. Ultraperformance convergence chromatography-high resolution tandem mass spectrometry for lipid biomarker profiling and identification. Biomed Chromatogr. 2017;31(3):e3822. doi: 10.1002/bmc.3822.
  15. 15.
    Tyburski JB, Patterson AD, Krausz KW, Slavik J, Fornace AJ Jr, Gonzalez FJ, et al. Radiation metabolomics. 1. Identification of minimally invasive urine biomarkers for gamma-radiation exposure in mice. Radiat res. 2008;170(1):1–14.CrossRefPubMedPubMedCentralGoogle Scholar
  16. 16.
    Ossetrova NI, Sandgren DJ, Blakely WF. Protein biomarkers for enhancement of radiation dose and injury assessment in nonhuman primate total-body irradiation model. Radiat Prot Dosim. 2014;159(1–4):61–76.CrossRefGoogle Scholar
  17. 17.
    Jones JW, Scott AJ, Tudor G, Xu PT, Jackson IL, Vujaskovic Z, et al. Identification and quantitation of biomarkers for radiation-induced injury via mass spectrometry. Health Phys. 2014;106(1):106–19.CrossRefPubMedGoogle Scholar
  18. 18.
    Biomarkers Used as Outcomes in Development of FDA-Approved Therapeutics (October 2007–December 2015) [10/18/2016]. Available from:
  19. 19.
    Mukherjee D, Coates PJ, Lorimore SA, Wright EG. Responses to ionizing radiation mediated by inflammatory mechanisms. J Pathol. 2014;232(3):289–99.CrossRefPubMedGoogle Scholar
  20. 20.
    Fleckenstein K, Zgonjanin L, Chen L, Rabbani Z, Jackson IL, Thrasher B, et al. Temporal onset of hypoxia and oxidative stress after pulmonary irradiation. Int J Radiat Oncol Biol Phys. 2007;68(1):196–204.CrossRefPubMedPubMedCentralGoogle Scholar
  21. 21.
    Carter CL, Jones JW, Farese AM, MacVittie TJ, Kane MA. Inflation-fixation method for Lipidomic mapping of lung biopsies by matrix assisted laser desorption/ionization-mass spectrometry imaging. Anal Chem. 2016;88(9):4788–94.CrossRefPubMedGoogle Scholar
  22. 22.
    Xia J, Wishart DS. Using MetaboAnalyst 3.0 for Comprehensive Metabolomics Data Analysis. Curr Protoc Bioinformatics. 2016;55(14.10):1–14.Google Scholar
  23. 23.
    Castro-Perez J, Roddy TP, Nibbering NM, Shah V, McLaren DG, Previs S, et al. Localization of fatty acyl and double bond positions in phosphatidylcholines using a dual stage CID fragmentation coupled with ion mobility mass spectrometry. J am Soc Mass Spectrom. 2011;22(9):1552–67.CrossRefPubMedPubMedCentralGoogle Scholar
  24. 24.
    Lisa M, Cifkova E, Holcapek M. Lipidomic profiling of biological tissues using off-line two-dimensional high-performance liquid chromatography-mass spectrometry. J Chromatogr a. 2011;1218(31):5146–56.CrossRefPubMedGoogle Scholar
  25. 25.
    Houjou T, Yamatani K, Nakanishi H, Imagawa M, Shimizu T, Taguchi R. Rapid and selective identification of molecular species in phosphatidylcholine and sphingomyelin by conditional neutral loss scanning and MS3. Rapid Commun Mass Spectrom. 2004;18(24):3123–30.CrossRefPubMedGoogle Scholar
  26. 26.
    Pannkuk EL, Fornace AJ Jr, Laiakis EC. Metabolomic applications in radiation biodosimetry: exploring radiation effects through small molecules. Int J Radiat Biol. 2017;12:1–26.Google Scholar
  27. 27.
    Brunelli L, Caiola E, Marabese M, Broggini M, Pastorelli R. Comparative metabolomics profiling of isogenic KRAS wild type and mutant NSCLC cells in vitro and in vivo. Sci Report. 2016;6:28398.CrossRefGoogle Scholar
  28. 28.
    Schnackenberg LK, Pence L, Vijay V, Moland CL, George N, Cao Z, et al. Early metabolomics changes in heart and plasma during chronic doxorubicin treatment in B6C3F1 mice. J Appl Toxicol. 2016;36(11):1486–95.CrossRefPubMedGoogle Scholar
  29. 29.
    Qiu Y, Zhou B, Su M, Baxter S, Zheng X, Zhao X, et al. Mass spectrometry-based quantitative metabolomics revealed a distinct lipid profile in breast cancer patients. Int J Mol Sci. 2013;14(4):8047–61.CrossRefPubMedPubMedCentralGoogle Scholar
  30. 30.
    Conlon TM, Bartel J, Ballweg K, Gunter S, Prehn C, Krumsiek J, et al. Metabolomics screening identifies reduced L-carnitine to be associated with progressive emphysema. Clin Sci (Lond.). 2016;130(4):273–87.CrossRefGoogle Scholar
  31. 31.
    Carter CL, Jones JW, Barrow K, Kieta K, Taylor-Howell C, Kearney S, et al. A MALDI-MSI approach to the characterization of radiation-induced lung injury and medical countermeasure development. Health Phys. 2015;109(5):466–78.CrossRefPubMedPubMedCentralGoogle Scholar
  32. 32.
    Wu G. Amino acids: metabolism, functions, and nutrition. Amino Acids. 2009;37(1):1–17.CrossRefPubMedGoogle Scholar
  33. 33.
    Orlando GF, Wolf G, Engelmann M. Role of neuronal nitric oxide synthase in the regulation of the neuroendocrine stress response in rodents: insights from mutant mice. Amino Acids. 2008;35(1):17–27.CrossRefPubMedGoogle Scholar
  34. 34.
    Wu G, Bazer FW, Davis TA, Kim SW, Li P, Marc Rhoads J, et al. Arginine metabolism and nutrition in growth, health and disease. Amino Acids. 2009;37(1):153–68.CrossRefPubMedGoogle Scholar
  35. 35.
    Chen PE, Geballe MT, Stansfeld PJ, Johnston AR, Yuan H, Jacob AL, et al. Structural features of the glutamate binding site in recombinant NR1/NR2A N-methyl-D-aspartate receptors determined by site-directed mutagenesis and molecular modeling. Mol Pharmacol. 2005;67(5):1470–84.CrossRefPubMedGoogle Scholar
  36. 36.
    Said SI, Berisha HI, Pakbaz H. Excitotoxicity in the lung: N-methyl-D-aspartate-induced, nitric oxide-dependent, pulmonary edema is attenuated by vasoactive intestinal peptide and by inhibitors of poly(ADP-ribose) polymerase. Proc Natl Acad Sci U S a. 1996;93(10):4688–92.CrossRefPubMedPubMedCentralGoogle Scholar
  37. 37.
    Li Y, Liu Y, Peng X, Liu W, Zhao F, Feng D, et al. NMDA receptor antagonist attenuates bleomycin-induced acute lung injury. PLoS One. 2015;10(5):e0125873.CrossRefPubMedPubMedCentralGoogle Scholar
  38. 38.
    da Cunha AA, Pauli V, Saciura VC, Pires MG, Constantino LC, de Souza B, et al. N-methyl-D-aspartate glutamate receptor blockade attenuates lung injury associated with experimental sepsis. Chest. 2010;137(2):297–302.CrossRefPubMedGoogle Scholar
  39. 39.
    da Cunha AA, Nunes FB, Lunardelli A, Pauli V, Amaral RH, de Oliveira LM, et al. Treatment with N-methyl-D-aspartate receptor antagonist (MK-801) protects against oxidative stress in lipopolysaccharide-induced acute lung injury in the rat. Int Immunopharmacol. 2011;11(6):706–11.CrossRefPubMedGoogle Scholar
  40. 40.
    Tang F, Yue S, Luo Z, Feng D, Wang M, Qian C, et al. Role of N-methyl-D-aspartate receptor in hyperoxia-induced lung injury. Pediatr Pulmonol. 2005;40(5):437–44.CrossRefPubMedGoogle Scholar
  41. 41.
    Engelmann B, Brautigam C, Thiery J. Plasmalogen phospholipids as potential protectors against lipid peroxidation of low density lipoproteins. Biochem Biophys res Commun. 1994;204(3):1235–42.CrossRefPubMedGoogle Scholar
  42. 42.
    Harwood JL. Lung surfactant. Prog Lipid res. 1987;26(3):211–56.CrossRefPubMedGoogle Scholar
  43. 43.
    Griese M. Pulmonary surfactant in health and human lung diseases: state of the art. Eur Respir J. 1999;13(6):1455–76.CrossRefPubMedGoogle Scholar
  44. 44.
    Finkelstein JN. Physiologic and toxicologic responses of alveolar type II cells. Toxicology. 1990;60(1–2):41–52.CrossRefPubMedGoogle Scholar
  45. 45.
    Agrawal A, Kale RK. Radiation induced peroxidative damage: mechanism and significance. Indian J Exp Biol. 2001;39(4):291–309.PubMedGoogle Scholar
  46. 46.
    Catala A. Lipid peroxidation of membrane phospholipids generates hydroxy-alkenals and oxidized phospholipids active in physiological and/or pathological conditions. Chem Phys Lipids. 2009;157(1):1–11.CrossRefPubMedGoogle Scholar
  47. 47.
    Braverman NE, Moser AB. Functions of plasmalogen lipids in health and disease. Biochim Biophys Acta. 2012;1822(9):1442–52.CrossRefPubMedGoogle Scholar
  48. 48.
    Niaudet C, Bonnaud S, Guillonneau M, Gouard S, Gaugler MH, Dutoit S, et al. Plasma membrane reorganization links acid sphingomyelinase/ceramide to p38 MAPK pathways in endothelial cells apoptosis. Cell Signal. 2017;33:10–21.CrossRefPubMedGoogle Scholar
  49. 49.
    Corre I, Guillonneau M, Paris F. Membrane signaling induced by high doses of ionizing radiation in the endothelial compartment. Relevance in radiation toxicity. Int J Mol Sci. 2013;14(11):22678–96.CrossRefPubMedPubMedCentralGoogle Scholar
  50. 50.
    Kolesnick R, Fuks Z. Radiation and ceramide-induced apoptosis. Oncogene. 2003;22(37):5897–906.CrossRefPubMedGoogle Scholar

Copyright information

© Springer Science+Business Media New York 2017

Authors and Affiliations

  • Jace W. Jones
    • 1
  • Isabel L. Jackson
    • 2
  • Zeljko Vujaskovic
    • 2
  • Michael D. Kaytor
    • 3
  • Maureen A. Kane
    • 1
    Email author
  1. 1.School of Pharmacy, Department of Pharmaceutical SciencesUniversity of MarylandBaltimoreUSA
  2. 2.School of Medicine, Division of Translational Radiation Sciences Department of Radiation OncologyUniversity of MarylandBaltimoreUSA
  3. 3.Humanetics CorporationEdinaUSA

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