Cancer Chemotherapy and Pharmacology

, Volume 64, Issue 2, pp 253–261

Enhancement of the in vivo antitumor activity of clofarabine by 1-β-d-[4-thio-arabinofuranosyl]-cytosine

  • William B. Parker
  • Sue C. Shaddix
  • Karen S. Gilbert
  • Rodney V. Shepherd
  • William R. Waud
Original Article



Clofarabine increases the activation of 1-β-d-arabinofuranosyl cytosine (araC) in tumor cells, and combination of these two drugs has been shown to result in good clinical activity against various hematologic malignancies. 1-β-d-[4-thio-arabinofuranosyl] cytosine (T-araC) is a new cytosine analog that has exhibited excellent activity against a broad spectrum of human solid tumors and leukemia/lymphoma xenografts in mice and is currently being evaluated in patients as a new drug for the treatment of cancer. Since T-araC has a vastly superior preclinical efficacy profile in comparison to araC, we have initiated studies to determine the potential value of clofarabine/T-araC combination therapy.


In vitro studies have been conducted to determine the effect of clofarabine on the metabolism of T-araC, and in vivo studies have been conducted to determine the effect of the clofarabine/T-araC combination on five human tumor xenografts in mice.


Initial studies with various tumor cells in culture indicated that a 2-h incubation with clofarabine enhanced the metabolism of T-araC 24 h after its removal by threefold in three tumor cell types (HCT-116 colon, K562 leukemia, and RL lymphoma) and by 1.5-fold in two other tumor cell types (MDA-MB-435 breast (melanoma), and HL-60 leukemia). Pretreatment with clofarabine resulted in a slight decrease in metabolism of T-araC in RPMI-8226 myeloma cells (65% of control) and inhibited metabolism of T-araC in CCRF-CEM leukemia cells by 90%. In vivo combination studies were conducted with various human tumor xenografts to determine whether or not the modulations observed in vitro were reflective of the in vivo situation. Clofarabine and T-araC were administered on alternate days for five treatments each (q2dx5) with the administration of T-araC 24 h after each clofarabine treatment. Combination treatment of HCT-116, K562, HL-60, or RL tumors with clofarabine and T-araC resulted in dramatically superior anti-tumor activity than treatment with either agent alone, whereas this combination resulted in antagonism in CCRF-CEM tumors. The in vivo antitumor activity of clofarabine plus T-araC against HCT-116 tumors was much better than the activity seen with clofarabine plus araC.


These studies provide a rationale for clinical trials using this combination in the treatment of acute leukemias as well as solid tumors and suggest that this combination would exhibit greater antitumor activity than that of clofarabine plus araC.


Clofarabine Arabinofuranosyl cytosine 1-β-d-[4-Thio-arabinofuranosyl]-cytosine Anti-solid tumor activity Synergistic antitumor activity 


  1. 1.
    Bonate PL, Arthaud L, Cantrell WR Jr, Stephenson K, Secrist JA III, Weitman S (2006) Discovery and development of clofarabine: a nucleoside analogue for treating cancer. Nat Rev Drug Discov 5:855–863PubMedCrossRefGoogle Scholar
  2. 2.
    Faderl S, Gandhi V, Keating MJ, Jeha S, Plunkett W, Kantarjian HM (2005) The role of clofarabine in hematologic and solid malignancies—development of a next-generation nucleoside analog. Cancer 103:1985–1995PubMedCrossRefGoogle Scholar
  3. 3.
    Kantarjian HM, Jeha S, Gandhi V, Wess M, Faderl S (2007) Clofarabine: past, present, and future. Leuk Lymphoma 48:1922–1930PubMedCrossRefGoogle Scholar
  4. 4.
    Cooper T, Ayres M, Nowak B, Gandhi V (2005) Biochemical modulation of cytarabine triphosphate by clofarabine. Cancer Chemother Pharmacol 55:361–368PubMedCrossRefGoogle Scholar
  5. 5.
    Faderl S, Gandhi V, O’Brien S, Bonate P, Cortes J, Estey E, Beran M, Wierda W, Garcia-Manero G, Ferrajoli A, Estrov Z, Giles FJ, Du M, Kwari M, Keating M, Plunkett W, Kantarjian H (2005) Results of a phase 1–2 study of clofarabine in combination with cytarabine (ara-C) in relapsed and refractory acute leukemias. Blood 105:940–947PubMedCrossRefGoogle Scholar
  6. 6.
    Faderl S, Verstovsek S, Cortes J, Ravandi F, Beran M, Garcia-Manero G, Ferrajoli A, Estrov Z, O’Brien S, Koller C, Giles FJ, Wierda W, Kwari M, Kantarjian HM (2006) Clofarabine and cytarabine combination as induction therapy for acute myeloid leukemia (AML) in patients 50 years of age or older. Blood 108:45–51PubMedCrossRefGoogle Scholar
  7. 7.
    Gidwani P, Ramesh KH, Liu Y, Kolb EA (2008) The combination of clofarabine and cytarabine in pediatric relapsed acute lymphoblastic leukemia: a case report. Chemotherapy 54:120–124PubMedCrossRefGoogle Scholar
  8. 8.
    Tiwari KN, Shortnacy-Fowler AT, Cappellacci L, Parker WB, Waud WR, Montgomery JA, Secrist JA III (2000) Synthesis of 4′-thio-β-d-arabinofuranosyl-cytosine (4′-thio-ara-C) and comparison of its anticancer activity with that of ara-C. Nucleosides Nucleotides Nucleic Acids 19:329–340PubMedCrossRefGoogle Scholar
  9. 9.
    Waud WR, Gilbert KS, Shepherd RV, Montgomery JA, Secrit JA III (2003) Preclinical antitumor activity of 4′-thio-beta-d-arabinofuranosylcytosine (4′-thio-ara-C). Cancer Chemother Pharmacol 51:422–426PubMedGoogle Scholar
  10. 10.
    Waud WR, Shepherd RV, Gilbert KS, Tiwari KN, Secrist JA III (2004) Precinical antitumor activity of 4′-thio-β-d-arabinofuranosylcytosine (4′-thio-ara-C, OSI-7836) in human leukemia and lymphoma xenograft models. Proc Am Assoc Cancer Res 45:714Google Scholar
  11. 11.
    Parker WB, Shaddix SC, Rose LM, Waud WR, Shewach DS, Tiwari KN, Secrist JA III (2000) Metabolism of 4′-thio-β-d-arabinofuranosylcytosine in CEM cells. Biochem Pharmacol 60:1925–1932PubMedCrossRefGoogle Scholar
  12. 12.
    Richardson F, Black C, Richardson K, Franks A, Wells E, Karimi S, Sennello G, Hart K, Meyer D, Emerson D, Brown E, LeRay J, Nilsson C, Tomkinson B, Bendele R (2005) Incorporation of OSI-7836 into DNA of Calu-6 and H460 xenograft tumors. Cancer Chemother Pharmacol 55:213–221PubMedCrossRefGoogle Scholar
  13. 13.
    Clarke ML, Damaraju VL, Zhang J, Mowles D, Tackaberry T, Lang T, Smith KM, Young JD, Tomkinson B, Cass CE (2006) The role of human nucleoside transporters in cellular uptake of 4′-thio-beta-d-arabinofuranosylcytosine and beta-d-arabinosylcytosine. Mol Pharmacol 70:303–310PubMedGoogle Scholar
  14. 14.
    Richardson KA, Vega TP, Richardson FC, Moore CL, Rohloff JC, Tomkinson B, Bendele RA, Kuchta RD (2004) Polymerization of the triphosphates of AraC, 2′,2′-difluorodeoxycytidine (dFdC) and OSI-7836 (T-araC) by human DNA polymerase alpha and DNA primase. Biochem Pharmacol 68:2337–2346PubMedCrossRefGoogle Scholar
  15. 15.
    Someya H, Shaddix SC, Tiwari KN, Secrist JA III, Parker WB (2003) Phosphorylation of 4′-thio-β-d-arabinofuranosylcytosine and its analogs by human deoxycytidine kinase. J Pharmacol Exp Ther 304:1314–1322PubMedCrossRefGoogle Scholar
  16. 16.
    Someya H, Waud WR, Parker WB (2006) Long intracellular retention of 4′-thio-arabinofuranosylcytosine 5′-triphosphate as a critical factor for the anti-solid tumor activity of 4′-thio-arabinofuranosylcytosine. Cancer Chemother Pharmacol 57:772–780PubMedCrossRefGoogle Scholar
  17. 17.
    Thottassery JV, Westbrook L, Someya H, Parker WB (2006) c-Abl-independent p73 stabilization during gemcitabine- or 4′-thio-beta-d-arabinofuranosylcytosine-induced apoptosis in wild-type and p53-null colorectal cancer cells. Mol Cancer Ther 5:400–410PubMedCrossRefGoogle Scholar
  18. 18.
    Goss G, Siu LL, Gauthier I, Chen EX, Oza AM, Goel R, Maroun J, Powers J, Walsh W, Maclean M, Drolet DW, Rusk J, Seymour LK, Investigational New Drug Program of the National Cancer Institute of Canada Clinical Trials Group (2006) A phase I, first in man study of OSI-7836 in patients with advanced refractory solid tumors: IND.147, a study of the Investigational New Drug Program of the National Cancer Institute of Canada Clinical Trials Group. Cancer Chemother Pharmacol 58:703–710PubMedCrossRefGoogle Scholar
  19. 19.
    Lee CP, de Jonge MJ, O’Donnell AE, Schothorst KL, Hanwell J, Chick JB, Brooimans RA, Adams LM, Drolet DW, de Bono JS, Kaye SB, Judson IR, Verweij J (2006) A phase I study of a new nucleoside analogue, OSI-7836, using two administration schedules in patients with advanced solid malignancies. Clin Cancer Res 12:2841–2848PubMedCrossRefGoogle Scholar
  20. 20.
    Dykes DJ, Abbott BJ, Mayo JG, Harrison SD Jr, Laster WR, Simpson-Herren L, Griswold DP Jr (1992) Development of human tumor xenograft models for in vivo evaluation of new antitumor drugs. Contrib Oncol Basel Karger 42:1–22Google Scholar
  21. 21.
    Gandhi V, Kantarjian H, Faderl S, Bonate P, Du M, Ayres M, Rios MB, Keating MJ, Plunkett W (2003) Pharmacokinetics and pharmacodynamics of plasma clofarabine and cellular clofarabine triphosphate in patients with acute leukemias. Clin Cancer Res 9:6335–6342PubMedGoogle Scholar
  22. 22.
    Waud WR, Schmid SM, Montgomery JA, Secrist JA III (2000) Preclinical antitumor activity of 2-chloro-9-(2-deoxy-2-fluoro-beta-d-arabinofuranosyl) adenine (Cl–F-ara-A). Nucleosides Nucleotides Nucleic Acids 19:447–460PubMedCrossRefGoogle Scholar
  23. 23.
    Parker WB, Shaddix SC, Chang CH, White EL, Rose LM, Brockman RW, Shortnancy AT, Montgomery JA, Secrist JA III, Bennett LL Jr (1991) Effects of 2-chloro-9-(2-deoxy-2-fluoro-β-d-arabinofuranosyl) adenine on K562 cellular metabolism and the inhibition of human ribonucleotide reductase and DNA polymerases by its 5-triphosphate. Cancer Res 51:2386–2394PubMedGoogle Scholar
  24. 24.
    Xie C, Plunkett W (1995) Metabolism and actions of 2-chloro-9-(2-deoxy-2-fluoro-beta-d-arabinofuranosyl)-adenine in human lymphoblastoid cells. Cancer Res 55:2847–2852PubMedGoogle Scholar
  25. 25.
    Parker WB, Shaddix SC, Rose LM, Shewach DS, Hertel LW, Secrist JA III, Montgomery JA, Bennett LL Jr (1999) Comparison of the mechanism of cytotoxicity of 2-chloro-9-(2-deoxy-2-fluoro-β-d-arabinofuranosyl) adenine, 2-chloro-9-(2-deoxy-2-fluoro-β-d-ribofuranosyl) adenine, and 2-chloro-9-(2-deoxy-2,2-difluoro-β-d-ribofuranosyl) adenine in CEM cells. Mol Pharmacol 55:515–520PubMedGoogle Scholar
  26. 26.
    Spasokoukotskaja T, Sasvári-Székely M, Keszler G, Albertioni F, Eriksson S, Staub M (1999) Treatment of normal and malignant cells with nucleoside analogues and etoposide enhances deoxycytidine kinase activity. Eur J Cancer 35:1862–1867PubMedCrossRefGoogle Scholar
  27. 27.
    Csapó Z, Sasvári-Székely M, Spasokoukotskaja T, Talianidis I, Eriksson S, Staub M (2001) Activation of deoxycytidine kinase by inhibition of DNA synthesis in human lymphocytes. Biochem Pharmacol 61:191–197PubMedCrossRefGoogle Scholar
  28. 28.
    Keszler G, Spasokoukotskaja T, Sasvári-Székely M, Eriksson S, Staub M (2006) Deoxycytidine kinase is reversibly phosphorylated in normal human lymphocytes. Nucleosides Nucleotides Nucleic Acids 25:1147–1151PubMedCrossRefGoogle Scholar
  29. 29.
    Qian M, Wang X, Shanmuganathan K, Chu CK, Gallo JM (1994) Pharmacokinetics of the anticancer agent 2-chloro-9-(2-deoxy-2-fluoro-beta-d-arabinofuranosyl) adenine in rats. Cancer Chemother Pharmacol 33:484–488PubMedGoogle Scholar

Copyright information

© Springer-Verlag 2008

Authors and Affiliations

  • William B. Parker
    • 1
  • Sue C. Shaddix
    • 1
  • Karen S. Gilbert
    • 1
  • Rodney V. Shepherd
    • 1
  • William R. Waud
    • 1
  1. 1.Southern Research InstituteBirminghamUSA

Personalised recommendations