Cancer Chemotherapy and Pharmacology

, Volume 65, Issue 4, pp 791–801 | Cite as

A phase I trial of PR-104, a nitrogen mustard prodrug activated by both hypoxia and aldo-keto reductase 1C3, in patients with solid tumors

  • Michael B. JamesonEmail author
  • Danny Rischin
  • Mark Pegram
  • John Gutheil
  • Adam V. Patterson
  • William A. Denny
  • William R. Wilson
Clinical Trial Report



PR-104 is a “pre-prodrug” designed to be activated to a dinitrobenzamide nitrogen mustard cytotoxin by nitroreduction in hypoxic regions of tumors. This study was conducted to establish the maximum tolerated dose (MTD), dose-limiting toxicity (DLT), safety, and pharmacokinetics (PK) of PR-104 in patients with advanced solid tumors.


Patients with solid tumors refractory or not amenable to conventional treatment were evaluated in a dose-escalation trial of PR-104 administered as a 1-h intravenous (IV) infusion every 3 weeks. The plasma PK of PR-104 and its primary metabolite, PR-104A, were evaluated.


Twenty-seven patients received a median of two cycles of PR-104 in doses ranging from 135 to 1,400 mg/m2. The MTD of PR-104 as a single-dose infusion every 3 weeks was established as 1,100 mg/m2. One of six patients treated at 1,100 mg/m2 experienced DLT of grade 3 fatigue. Above the MTD, the DLTs at 1,400 mg/m2 were febrile neutropenia and infection with normal absolute neutrophil count. No objective responses were observed, although reductions in tumor size were observed in patients treated at doses ≥550 mg/m2. The plasma PK of PR-104 demonstrated rapid conversion to PR-104A, with approximately dose-linear PK of both species.


PR-104 was well tolerated at a dose of 1,100 mg/m2 administered as an IV infusion every 3 weeks. The area under the PR-104A plasma concentration–time curve at this dose exceeded that required for activity in human tumor cell cultures and xenograft models. The recommended dose of PR-104 as a single agent for phase II trials is 1,100 mg/m2 and further trials are underway.


PR-104 Hypoxia Prodrug Dinitrobenzamide nitrogen mustard Phase I 



We thank the patients who participated in this trial and the research staff for their assistance in patient care and their dedication to clinical trials. We also thank Kashyap Patel and Prof. Nick Holford for advice on pharmacokinetic analysis, and Terri Melink for assistance with preparation of the manuscript. Financial support for this trial was provided by Proacta, Inc.

Conflicts of interest statement

No financial conflict exists for authors M.B.J., D.R., and M.P. The following authors have indicated a potential conflict of interest: J.G. is employed by Proacta Inc. with stock ownership; A.V.P is a consultant to Proacta Inc.; and W.A.D. and W.R.W. are consultants/played an advisory role to Proacta Inc. and have stock ownership and received research funding.


  1. 1.
    Brown JM, Giaccia AJ (1998) The unique physiology of solid tumors: opportunities (and problems) for cancer therapy. Cancer Res 58:1408–1416PubMedGoogle Scholar
  2. 2.
    Wouters BG, Weppler SA, Koritzinsky M, Landuyt W, Nuyts S, Theys J, Chiu RK, Lambin P (2002) Hypoxia as a target for combined modality treatments. Eur J Cancer 38:240–257CrossRefPubMedGoogle Scholar
  3. 3.
    Brown JM, Wilson WR (2004) Exploiting tumor hypoxia in cancer treatment. Nature Rev Cancer 4:437–447CrossRefGoogle Scholar
  4. 4.
    Tannock IF (1968) The relation between cell proliferation and the vascular system in a transplanted mouse mammary tumour. Br J Cancer 22:258–273PubMedGoogle Scholar
  5. 5.
    Minchinton AI, Tannock IF (2006) Drug penetration in solid tumours. Nat Rev Cancer 6:583–592CrossRefPubMedGoogle Scholar
  6. 6.
    Hicks KO, Pruijn FB, Secomb TW, Hay MP, Hsu R, Brown JM, Denny WA, Dewhirst MW, Wilson WR (2006) Use of three-dimensional tissue cultures to model extravascular transport and predict in vivo activity of hypoxia-targeted anticancer drugs. J Natl Cancer Inst 98:1118–1128PubMedCrossRefGoogle Scholar
  7. 7.
    Graeber TG, Osmanian C, Jacks T, Housman DE, Koch CJ, Lowe SW, Giaccia AJ (1996) Hypoxia-mediated selection of cells with diminished apoptotic potential in solid tumours. Nature 379:88–91CrossRefPubMedGoogle Scholar
  8. 8.
    Gray LH (1957) Oxygenation in radiotherapy. I. Radiobiological considerations. Br J Radiol 30:403–406CrossRefPubMedGoogle Scholar
  9. 9.
    Thomlinson RH, Gray LH (1955) The histological structure of some human lung cancers and possible implications for radiotherapy. Br J Cancer 9:539–549PubMedGoogle Scholar
  10. 10.
    Teicher BA, Lazo JS, Sartorelli AC (1981) Classification of antineoplastic agents by their selective toxicities toward oxygenated and hypoxic tumor cells. Cancer Res 41:73–81PubMedGoogle Scholar
  11. 11.
    Comerford KM, Wallace TJ, Karhausen J, Louis NA, Montalto MC, Colgan SP (2002) Hypoxia-inducible factor-1-dependent regulation of the multidrug resistance (MDR1) gene. Cancer Res 62:3387–3394PubMedGoogle Scholar
  12. 12.
    Wartenberg M, Ling FC, Muschen M, Klein F, Acker H, Gassmann M, Petrat K, Putz V, Hescheler J, Sauer H (2003) Regulation of the multidrug resistance transporter P-glycoprotein in multicellular tumor spheroids by hypoxia-inducible factor (HIF-1) and reactive oxygen species. FASEB J 17:503–505PubMedGoogle Scholar
  13. 13.
    Raspaglio G, Filippetti F, Prislei S, Penci R, De Maria I, Cicchillitti L, Mozzetti S, Scambia G, Ferlini C (2008) Hypoxia induces class III beta-tubulin gene expression by HIF-1alpha binding to its 3′ flanking region. Gene 409:100–108CrossRefPubMedGoogle Scholar
  14. 14.
    Liu L, Sun L, Zhang H, Li Z, Ning X, Shi Y, Guo C, Han S, Wu K, Fan D (2009) Hypoxia-mediated up-regulation of MGr1-Ag/37LRP in gastric cancers occurs via hypoxia-inducible-factor 1-dependent mechanism and contributes to drug resistance. Int J Cancer 124:1707–1715CrossRefPubMedGoogle Scholar
  15. 15.
    Brizel DM, Scully SP, Harrelson JM, Layfield LJ, Bean JM, Prosnitz LR, Dewhirst MW (1996) Tumor oxygenation predicts for the likelihood of distant metastases in human soft tissue sarcoma. Cancer Res 56:941–943PubMedGoogle Scholar
  16. 16.
    Fyles A, Milosevic M, Hedley D, Pintilie M, Levin W, Manchul L, Hill RP (2002) Tumor hypoxia has independent predictor impact only in patients with node-negative cervix cancer. J Clin Oncol 20:680–687CrossRefPubMedGoogle Scholar
  17. 17.
    Harris AL (2002) Hypoxia—a key regulatory factor in tumour growth. Nature Rev Cancer 2:38–47CrossRefGoogle Scholar
  18. 18.
    Nordsmark M, Alsner J, Keller J, Nielsen OS, Jensen OM, Horsman MR, Overgaard J (2001) Hypoxia in human soft tissue sarcomas: adverse impact on survival and no association with p53 mutations. Br J Cancer 84:1070–1075CrossRefPubMedGoogle Scholar
  19. 19.
    Subarsky P, Hill RP (2003) The hypoxic tumour microenvironment and metastatic progression. Clin Exp Metastasis 20:237–250CrossRefPubMedGoogle Scholar
  20. 20.
    McKeown SR, Cowen RL, Williams KJ (2007) Bioreductive drugs: from concept to clinic. Clin Oncol 19:427–442CrossRefGoogle Scholar
  21. 21.
    Plumb JA, Gerritsen M, Milroy R, Thomson P, Workman P (1994) Relative importance of DT-diaphorase and hypoxia in the bioactivation of EO9 by human lung tumor cell lines. Int J Radiat Oncol Biol Phys 29:295–299PubMedGoogle Scholar
  22. 22.
    Haffty BG, Wilson LD, Son YH, Cho EI, Papac RJ, Fischer DB, Rockwell S, Sartorelli AC, Ross DA, Sasaki CT, Fischer JJ (2005) Concurrent chemo-radiotherapy with mitomycin C compared with porfiromycin in squamous cell cancer of the head and neck: final results of a randomized clinical trial. Int J Radiat Oncol Biol Phys 61:119–128PubMedGoogle Scholar
  23. 23.
    van der Heijden AG, Moonen PM, Cornel EB, Vergunst H, de Reijke TM, van Boven E, Barten EJ, Puri R, van Kalken CK, Witjes JA (2006) Phase II marker lesion study with intravesical instillation of apaziquone for superficial bladder cancer: toxicity and marker response. J Urol 176:1349–1353CrossRefPubMedGoogle Scholar
  24. 24.
    Sharp SY, Kelland LR, Valenti MR, Brunton LA, Hobbs S, Workman P (2000) Establishment of an isogenic human colon tumor model for NQO1 gene expression: application to investigate the role of DT-diaphorase in bioreductive drug activation in vitro and in vivo. Mol Pharmacol 58:1146–1155PubMedGoogle Scholar
  25. 25.
    Knox RJ, Friedlos F, Marchbank T, Roberts JJ (1991) Bioactivation of CB 1954: reaction of the active 4-hydroxylamino derivative with thioesters to form the ultimate DNA–DNA interstrand crosslinking species. Biochem Pharmacol 42:1691–1697CrossRefPubMedGoogle Scholar
  26. 26.
    Celli CM, Tran N, Knox R, Jaiswal AK (2006) NRH:quinone oxidoreductase 2 (NQO2) catalyzes metabolic activation of quinones and anti-tumor drugs. Biochem Pharmacol 72:366–376CrossRefPubMedGoogle Scholar
  27. 27.
    Chung-Faye G, Palmer D, Anderson D, Clark J, Downes M, Baddeley J, Hussain S, Murray PI, Searle P, Seymour L, Harris PA, Ferry D, Kerr DJ (2001) Virus-directed, enzyme prodrug therapy with nitroimidazole reductase: a phase I and pharmacokinetic study of its prodrug, CB1954. Clin Cancer Res 7:2662–2668PubMedGoogle Scholar
  28. 28.
    Palmer DH, Mautner V, Mirza D, Oliff S, Gerritsen W, van dS, Jr, Hubscher S, Reynolds G, Bonney S, Rajaratnam R, Hull D, Horne M, Ellis J, Mountain A, Hill S, Harris PA, Searle PF, Young LS, James ND, Kerr DJ (2004) Virus-directed enzyme prodrug therapy: intratumoral administration of a replication-deficient adenovirus encoding nitroreductase to patients with resectable liver cancer. J Clin Oncol 22:1546–1552Google Scholar
  29. 29.
    Searle PF, Chen MJ, Hu L, Race PR, Lovering AL, Grove JI, Guise C, Jaberipour M, James ND, Mautner V, Young LS, Kerr KJ, Mountain A, White SA, Hyde EI (2004) Nitroreductase: a prodrug-activating enzyme for cancer gene therapy. Clin Exp Pharmacol Physiol 31:811–816CrossRefPubMedGoogle Scholar
  30. 30.
    von Pawel J, von Roemeling R, Gatzemeier U, Boyer M, Elisson LO, Clark P, Talbot D, Rey A, Butler TW, Hirsh V, Olver I, Bergman B, Ayoub J, Richardson G, Dunlop D, Arcenas A, Vescio R, Viallet J, Treat J (2000) Tirapazamine plus cisplatin versus cisplatin in advanced non-small-cell lung cancer: a report of the international CATAPULT I study group. Cisplatin and tirapazamine in subjects with advanced previously untreated non-small-cell lung tumors. J Clin Oncol 18:1351–1359Google Scholar
  31. 31.
    Shepherd F, Koschel G, von Pawel J, Gatzmeier U, van Zandwiyk N, Woll P, van Klavren R, Krasko P, Desimone P, Nicolson M, Pieters W, Bigelow R, Rey A, Biallet J, Loh E (2000) Comparison of Tirazone (tirapazamine) and cisplatin vs. etoposide and cisplatin in advanced non-small cell lung cancer (NSCLC): final results of the international phase III CATAPULT II Trial. Lung Cancer 29 (suppl 1):28, abstract 87Google Scholar
  32. 32.
    Williamson SK, Crowley JJ, Lara PN Jr, McCoy J, Lau DH, Tucker RW, Mills GM, Gandara DR (2005) Phase III trial of paclitaxel plus carboplatin with or without tirapazamine in advanced non-small-cell lung cancer: Southwest Oncology Group Trial S0003. J Clin Oncol 23:9097–9104CrossRefPubMedGoogle Scholar
  33. 33.
    Rischin D, Peters L, O’Sullivan B, Giralt J, Yuen K, Trotti A, Bernier J, Bourhis J, Henke M, Fisher R (2008) Phase III study of tirapazamine, cisplatin and radiation versus cisplatin and radiation for advanced squamous cell carcinoma of the head and neck. J Clin Oncol 26:Abstract LBA6008Google Scholar
  34. 34.
    Rischin D, Hicks RJ, Fisher R, Binns D, Corry J, Porceddu S, Peters LJ, Trans-Tasman Radiation Oncology Group (2006) Prognostic significance of [18F]-misonidazole positron emission tomography-detected tumor hypoxia in patients with advanced head and neck cancer randomly assigned to chemoradiation with or without tirapazamine: a substudy of Trans-Tasman Radiation Oncology Group Study 98.02. J Clin Oncol 24:2098–2104CrossRefPubMedGoogle Scholar
  35. 35.
    Wilson WR, Hicks KO, Pullen SM, Ferry DM, Helsby NA, Patterson AV (2007) Bystander effects of bioreductive drugs: potential for exploiting pathological tumor hypoxia with dinitrobenzamide mustards. Radiat Res 167:625–636CrossRefPubMedGoogle Scholar
  36. 36.
    Koch CJ (1993) Unusual oxygen concentration dependence of toxicity of SR-4233, a hypoxic cell toxin. Cancer Res 53:3992–3997PubMedGoogle Scholar
  37. 37.
    Hicks KO, Siim BG, Pruijn FB, Wilson WR (2004) Oxygen dependence of the metabolic activation and cytotoxicity of tirapazamine: implications for extravascular transport and activity in tumors. Radiat Res 161:656–666CrossRefPubMedGoogle Scholar
  38. 38.
    Lee AE, Wilson WR (2000) Hypoxia-dependent retinal toxicity of bioreductive anticancer prodrugs in mice. Toxicol Appl Pharmacol 163:50–59CrossRefPubMedGoogle Scholar
  39. 39.
    Parmar K, Mauch P, Vergilio J, Sackstein R, Down JD (2007) Distribution of hematopoietic stem cells in the bone marrow according to regional hypoxia. Proc Natl Acad Sci USA 104:5431–5436CrossRefPubMedGoogle Scholar
  40. 40.
    Helsby NA, Wheeler SJ, Pruijn FB, Palmer BD, Yang S, Denny WA, Wilson WR (2003) Effect of nitroreduction on the alkylating reactivity and cytotoxicity of the 2, 4-dinitrobenzamide-5-aziridine CB 1954 and the corresponding nitrogen mustard SN 23862: distinct mechanisms of bioreductive activation. Chem Res Toxicol 16:469–478CrossRefPubMedGoogle Scholar
  41. 41.
    Patterson AV, Ferry DM, Edmunds SJ, Gu Y, Singleton RS, Patel K, Pullen SM, Syddall SP, Atwell GJ, Yang S, Denny WA, Wilson WR (2007) Mechanism of action and preclinical antitumor activity of the novel hypoxia-activated DNA crosslinking agent PR-104. Clin Cancer Res 13:3922–3932CrossRefPubMedGoogle Scholar
  42. 42.
    Hicks KO, Myint H, Patterson AV, Pruijn FB, Siim BG, Patel K, Wilson WR (2007) Oxygen dependence and extravascular transport of hypoxia-activated prodrugs: comparison of the dinitrobenzamide mustard PR-104A and tirapazamine. Int J Radiat Oncol Biol Phys 69:560–571PubMedGoogle Scholar
  43. 43.
    Singleton RS, Guise CP, Ferry DM, Pullen SM, Dorie MJ, Brown JM, Patterson AV, Wilson WR (2009) DNA crosslinks in human tumor cells exposed to the prodrug PR-104A: relationships to hypoxia, bioreductive metabolism and cytotoxicity. Cancer Res 69:3884–3891CrossRefPubMedGoogle Scholar
  44. 44.
    Patel K, Lewiston D, Gu Y, Hicks KO, Wilson WR (2007) Analysis of the hypoxia-activated dinitrobenzamide mustard phosphate prodrug PR-104 and its alcohol metabolite PR-104A in plasma and tissues by liquid chromatography–mass spectrometry. J Chromatogr B Analyt Technol Biomed Life Sci 856:302–311CrossRefPubMedGoogle Scholar
  45. 45.
    Guise CP, Wang A, Thiel A, Bridewell D, Wilson WR, Patterson AV (2007) Identification of human reductases that activate the dinitrobenzamide mustard prodrug PR-104A: a role for NADPH:cytochrome P450 oxidoreductase under hypoxia. Biochem Pharmacol 74:810–820CrossRefPubMedGoogle Scholar
  46. 46.
    Gu Y, Patterson AV, Atwell GJ, Chernikova SB, Brown JM, Thompson LH, Wilson WR (2009) Roles of DNA repair and reductase activity in the cytotoxicity of the hypoxia-activated dinitrobenzamide mustard PR-104A. Mol Cancer Ther 8:1714–1723CrossRefPubMedGoogle Scholar
  47. 47.
    Liu SC, Ahn GO, Kioi M, Dorie MJ, Patterson AV, Brown JM (2008) Optimised Clostridium-directed enzyme prodrug therapy improves the antitumor activity of the novel DNA crosslinking agent PR-104. Cancer Res 68:7995–8003CrossRefPubMedGoogle Scholar
  48. 48.
    Guise CP, Abbattista M, Singleton RS, Holford SD, Connolly J, Dachs GU, Fox SB, Pollock R, Harvey J, Guilford P, Doñate F, Wilson WR, Patterson AV (2008) The bioreductive prodrug PR-104 is activated under aerobic conditions by human aldo-keto reductase 1C3. Cancer Res (in press)Google Scholar
  49. 49.
    Penning TM, Drury JE (2007) Human aldo-keto reductases: function, gene regulation, and single nucleotide polymorphisms. Arch Biochem Biophys 464:241–250CrossRefPubMedGoogle Scholar
  50. 50.
    Wako K, Kawasaki T, Yamana K, Suzuki K, Jiang S, Umezu H, Nishiyama T, Takahashi K, Hamakubo T, Kodama T, Naito M (2008) Expression of androgen receptor through androgen-converting enzymes is associated with biological aggressiveness in prostate cancer. J Clin Pathol 61:448–454CrossRefPubMedGoogle Scholar
  51. 51.
    Ito K, Utsunomiya H, Suzuki T, Saitou S, Akahira J, Okamura K, Yaegashi N, Sasano H (2006) 17Beta-hydroxysteroid dehydrogenases in human endometrium and its disorders (Review). Mol Cell Endocrinol 248:136–140CrossRefPubMedGoogle Scholar
  52. 52.
    Sakurai M, Oishi K, Watanabe K (2005) Localization of cyclooxygenases-1 and -2, and prostaglandin F synthase in human kidney and renal cell carcinoma. Biochem Biophys Res Commun 338:82–86CrossRefPubMedGoogle Scholar
  53. 53.
    Therasse P, Arbuck SG, Eisenhauer EA, Wanders J, Kaplan RS, Rubinstein L, Verweij J, van Glabbeke M, Van Oosterom AT, Christian MC, Gwyther SG (2000) New guidelines to evaluate the response to treatment in solid tumors. J Natl Cancer Inst 92:205–216CrossRefPubMedGoogle Scholar
  54. 54.
    Atwell GJ, Denny WA (2007) Synthesis of 3H- and 2H4-labelled versions of the hypoxia-activated pre-prodrug 2-[(2-bromoethyl)-2, 4-dinitro-6-[[[2-(phosphonooxy)ethyl]amino]carbonyl]anilino]ethyl methanesulfonate (PR-104). J Labelled Comp Radiopharm 50:7–12CrossRefGoogle Scholar
  55. 55.
    Tannock IF, Lee CM, Tunggal JK, Cowan DS, Egorin MJ (2002) Limited penetration of anticancer drugs through tumor tissue: a potential cause of resistance of solid tumors to chemotherapy. Clin Cancer Res 8:878–884PubMedGoogle Scholar
  56. 56.
    Huxham LA, Kyle AH, Baker JHE, Nykilchuk LK, Minchinton AI (2004) Microregional effects of gemcitabine in HCT-116 xenografts. Cancer Res 63:6537–6541CrossRefGoogle Scholar
  57. 57.
    Birtwistle J, Hayden RE, Khanim FL, Green RM, Pearce C, Davies NJ, Wake N, Schrewe H, Ride JP, Chipman JK, Bunce CM (2009) The aldo-keto reductase AKR1C3 contributes to 7, 12-dimethylbenz(a)anthracene-3, 4-dihydrodiol mediated oxidative DNA damage in myeloid cells: implications for leukemogenesis. Mutat Res 662:67–74PubMedGoogle Scholar
  58. 58.
    Koukourakis MI, Bentzen SM, Giatromanolaki A, Wilson GD, Daley FM, Saunders MI, Dische S, Sivridis E, Harris AL (2006) Endogenous markers of two separate hypoxia response pathways (hypoxia inducible factor 2 alpha and carbonic anhydrase 9) are associated with radiotherapy failure in head and neck cancer patients recruited in the CHART randomized trial. J Clin Oncol 24:727–735CrossRefPubMedGoogle Scholar

Copyright information

© Springer-Verlag 2009

Authors and Affiliations

  • Michael B. Jameson
    • 1
    Email author
  • Danny Rischin
    • 2
  • Mark Pegram
    • 3
  • John Gutheil
    • 4
  • Adam V. Patterson
    • 5
  • William A. Denny
    • 5
  • William R. Wilson
    • 5
  1. 1.Regional Cancer CentreWaikato HospitalHamiltonNew Zealand
  2. 2.Peter MacCallum Cancer CentreMelbourneAustralia
  3. 3.University of California, Los AngelesLos AngelesUSA
  4. 4.Proacta Inc.San DiegoUSA
  5. 5.Auckland Cancer Society Research CentreThe University of AucklandAucklandNew Zealand

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