Download PDF
Arzneimittelforschung 2011; 61(4): 252-259
DOI: 10.1055/s-0031-1296196
Antibiotics · Antimycotics · Antiparasitics · Antiviral Drugs · Chemotherapeutics · Cytostatics
Editio Cantor Verlag Aulendorf (Germany)

In vitro and in vivo chemosensitizing activity of LFM-A13, a dual-function inhibitor of Bruton's tyrosine kinase and polo-like kinases, against human leukemic B-cell precursors

Fatih Uckun
1   Division of Hematology-Oncology, Department of Pediatrics, Childrens Center for Cancer and Blood Diseases, University of Southern California Keck School of Medicine, Los Angeles, CA, USA
2   Developmental Therapeutics Program, Childrens Center for Cancer and Blood Diseases, Childrens Hospital Los Angeles MS#57, Los Angeles, CA, USA
3   Developmental Therapeutics Program, USC Comprehensive Norris Cancer Center, Los Angeles, CA, USA
,
Ilker Dibirdik
2   Developmental Therapeutics Program, Childrens Center for Cancer and Blood Diseases, Childrens Hospital Los Angeles MS#57, Los Angeles, CA, USA
,
Aniee Sarkissian
2   Developmental Therapeutics Program, Childrens Center for Cancer and Blood Diseases, Childrens Hospital Los Angeles MS#57, Los Angeles, CA, USA
,
Sanjive Qazi
4   Department of Biology and Bioinformatics Program, Gustavus Adolphus College, St. Peter, MN, USA
› Author Affiliations
Further Information

Publication History

Publication Date:
27 November 2011 (online)

    Permissions and Reprints

Abstract

The present study documents the chemosensitizing anti-leukemic activity of the leflunomide metabolite (LFM) analog, LFM-A13, a dual-function inhibitor of Bruton’s tyrosine kinase (BTK) and Pololike kinases (PLK), against human leukemic B-cell precursors. The results in 135 xenografted NOD/SCID mice regarding the anti-leukemic activity of GMP-grade LFM-A13, obtained with only 4-days of LFM-A13 therapy at nontoxic dose levels corresponding to 1 – 20 % of its NOAEL (no observable advserse effect level), alone or in combination with the standard chemotherapy drug vincristine, demonstrate the potential of LFM-A13 as a new anti-leukemic drug candidate. A11 82 LFM-A13-treated mice, including those receiving a combination of vincristine + LFM-A13 at the highest dose level of LFM-A13, tolerated their treatments well without weight loss, diarrhea, lethargy/paralysis, other signs of morbidity, or mortality.

The present study provides preclinical proof-of-principle for the development of LFM-A13 as a new chemosensitizing and apoptosis-promoting anti-leukemic agent and lends support to the hypothesis that the chemoresistance of relapsed B-cell precursor acute lymphoblastic leukemia (BCP-ALL) can be overcome by using LFM-A13 in combination with chemotherapy. Also presented are the results of a comprehensive meta-analysis of the overexpression of genes for LFM-A13 targeted kinases and their downstream effector molecules in B-lineage lymphoid malignancies utilizing the Oncomine database.

Key words

Anti-leukemic drug - Bruton's tyrosine kinase - inhibitor - LFM-A13 - Polo-like kinases inhibitor - Vincristine - Xenografted NOD/SCID mice
 

  • References

  • 1 Trigg ME, Gaynon P, Uckun FM. Acute lymphoblastic leukemia in children. In: Cancer Medicine Fourth Edition. James F Holland, Emile III Fry, Robert C Jr. Bast, Donald W Kufe, Donald L Morton, Ralph R Weichselbaum, editors. Chapter 164. 1996. p. 2945-2960
    Google Scholar
  • 2 Gaynon PS. Childhood acute lymphoblastic leukemia and relapse. Br J Haematol. 2005; 131: 579-87
    CrossrefPubMedGoogle Scholar
  • 3 Bailey LC, Lange BJ, Rheinhold SR, Bunin NJ. Bone marrow relapse in paediatric acute lymphoblastic leukemia. Lancet Oncol. 2008; 9: 873-83
    CrossrefPubMedGoogle Scholar
  • 4 Roy A, Cargill A, Love S, Moorman AV, Stoneham S, Lim A et al. Outcome after first relapse in childhood acute lymphoblastic leukemia - lessons from the United Kingdom R2 trial. Br J Haematol. 2005; 130: 67-75
    CrossrefPubMedGoogle Scholar
  • 5 Uckun FM, Morar S, Qazi S. Vinorelbine-based salvage chemotherapy for therapy-refractory aggressive leukemias. Br J Haematol. 2006; 135: 500-8
    CrossrefPubMedGoogle Scholar
  • 6 Malempati S, Gaynon PS, Sather H, La MK, Stork LC. Outcome after relapse among children with standard-risk acute lymphoblastic leukemia: Children's Oncology Group study CCG-1952. J Clin Oncol. 2007; 25: 5800-7
    CrossrefPubMedGoogle Scholar
  • 7 Gaynon PS, Harris RE, Altman AJ, Bostrom BC, Breneman JC, Hawks R et al. Bone marrow transplantation versus prolonged intensive chemotherapy for children with acute lymphoblastic leukemia and an initial bone marrow relapse within 12 months of the completion of primary therapy: Children's Oncology Group Study CCG-1941. J Clin Oncol. 2006; 24: 3150-6
    CrossrefPubMedGoogle Scholar
  • 8 Raetz EA, Borowitz MJ, Devidas M, Linda SB, Hunger SP, Winick NJ et al. Reinduction platform for children with first marrow relapse of acute lymphoblastic leukemia: A Children's Oncology Group Study. J. Clin. Oncol. 2008; 26: 3971-3978
    CrossrefPubMedGoogle Scholar
  • 9 Chessells JM, Veys P, Kempski H, Henley P, Leiper A, Webb D et al. Long-term follow-up of relapsed childhood acute lymphoblastic leukaemia. Br J Haematol. 2003; 123: 396-405
    CrossrefPubMedGoogle Scholar
  • 10 Einsiedel HG, von Stackeiberg A, Hartmann R, Fengler R, Schrappe M, Janka-Schaub G et al. Long-term outcome in children with relapsed ALL by risk-stratified salvage therapy: results of trial acute lymphoblastic leukemia-relapse study of the Berlin-Frankfurt-Munster Group 87. J Clin Oncol. 2005; 23: 7942-50
    CrossrefPubMedGoogle Scholar
  • 11 Uckun FM, Tibbies HE, Vassilev AO. Bruton's tyrosine kinase as a new therapeutic target. Anti-Cancer Agents Med Chem. 2007; 7: 624-32
    CrossrefPubMedGoogle Scholar
  • 12 Kurosaki T, Hikida M. Tyrosine kinases and their substrates in B lymphocytes. Immunol Rev. 2009; 228: 132-48
    CrossrefPubMedGoogle Scholar
  • 13 Qiu Y, Kung HJ. Signaling network of the Btk family kinases. Oncogene. 2000; 19: 5651-61
    CrossrefPubMedGoogle Scholar
  • 14 Vassilev A, Ozer Z, Navara C, Mahajan S, Uckun FM. Bruton's tyrosine kinase as an inhibitor of the Fas/CD95 death-inducing signaling complex. J Biol Chem. 1999; 274: 1646-56
    CrossrefPubMedGoogle Scholar
  • 15 Mahajan S, Vassilev A, Sun N, Ozer Z, Mao C, Uckun FM. Transcription factor STAT5A is a substrate of Bruton's tyrosine kinase in B cells. J Biol Chem. 2001; 276: 31216-28
    CrossrefPubMedGoogle Scholar
  • 16 Glassford J, Soeiro I, Skarell SM, Banerji L, Holman M, Klaus GG et al. BCR targets Cyclin D2 via Btk and the p85alpha subunit of PI3-K to induce cell cycle progression in primary mouse B cells. Oncogene. 2003; 22: 2248-59
    CrossrefPubMedGoogle Scholar
  • 17 Bajpai UD, Zhang K, Teutsch M, Sen R, Wortis HH. Bruton's tyrosine kinase links the B cell receptor to nuclear factor kappaB activation. J Exp Med. 2000; 191: 1735-44
    CrossrefPubMedGoogle Scholar
  • 18 Petro JB, Khan WN. Phospholipase C-gamma 2 couples Bruton's tyrosine kinase to the NF-kappaB signaling pathway in B lymphocytes. J Biol Chem. 2001; 276: 1715-9
    CrossrefPubMedGoogle Scholar
  • 19 Uckun FM. Clinical potential of targeting Bruton's tyrosine kinase [Review]. Int Rev Immunol. 2008; 27: 43-69
    CrossrefPubMedGoogle Scholar
  • 20 Mahajan S, Ghosh S, Sudbeck EA, Zheng Y, Downs S, Hupke M et al. Rational design and synthesis of a novel anti-leukemic agent targeting Bruton's tyrosine kinase (BTK), LFM-A13 [alpha-cyano-beta-hydroxy-beta-methyl-N-(2,5-dibromophenyl)propenamide]. J Biol Chem. 1999; 274: 9587-99
    CrossrefPubMedGoogle Scholar
  • 21 Mao C, Zhou M, Uckun FM. Crystal structure of Bruton's tyrosine kinase domain suggests a novel pathway for activation and provides insights into the molecular basis of X-linked agammaglobulinemia. J Biol Chem. 2001; 276: 41435-43
    CrossrefPubMedGoogle Scholar
  • 22 Uckun FM. Chemosensitizing anti-cancer activity of LFM-A13, a leflunomide metabolite analog targeting polo-like kinases. Cell Cycle. 2007; 6: 3021-6
    CrossrefPubMedGoogle Scholar
  • 23 Uckun FM, Dibirdik I, Qazi S, Vassilev A, Ma H, Mao C et al. Anti-breast cancer activity of LFM-A13, a potent inhibitor of Polo-like kinase (PLK). Bioorg Med Chem. 2007; 15: 800-14
    CrossrefPubMedGoogle Scholar
  • 24 Uckun FM, Qazi S, Ma H, Tuel-Ahlgren L, Ozer Z. STAT3 is a substrate of SYK tyrosine kinase in B-lineage leukemia/lymphoma cells exposed to oxidative stress. Proc Natl Acad Sci. USA. 2010; 107: 2902-7
    CrossrefPubMedGoogle Scholar
  • 25 Kim YJ, Sekiya F, Poulin B, Bae YS, Rhee SG. Mechanism of B-cell receptor-induced phosphorylation and activation of phospholipase C-gamma2. Mol Cell Biol. 2004; 24: 9986-99
    CrossrefPubMedGoogle Scholar
  • 26 Heinonen JE, Smith CI, Nore BF. Silencing of Bruton's tyrosine kinase (Btk) using short interfering RNA duplexes (siRNA). FEBS Lett. 2002; 527: 274-8
    CrossrefPubMedGoogle Scholar
  • 27 Feldhahn N, Rio P, Soh BN, Liedtke S, Sprangers M, Klein F et al. Deficiency of Bruton's tyrosine kinase in B cell precursor leukemia cells. Proc Natl Acad Sci U S A. 2005; 102: 13266-71
    CrossrefPubMedGoogle Scholar
  • 28 Contri A, Brunati AM, Trantin L, Cabrelle A, Miorin M, Cesaro L et al. Chronic lymphocytic leukemia B cells contain anomalous Lyn tyrosine kinase, a putative contribution to defective apoptosis. J Clin Invest. 2005; 115: 369-78
    PubMedGoogle Scholar
  • 29 Fernandes MJ, Lachance G, Pare G, Rollet-Labelle E, Naccache PH. Signaling through CD 16b in human neutrophils involves the Tec family of tyrosine kinases. J Leukoc Biol. 2005; 78: 524-32
    CrossrefPubMedGoogle Scholar
  • 30 Tu T, Thotala D, Geng L, Hallahan DE, Willey CD. Bone marrow X-kinase-mediated signal transduction in irradiated vascular endothelium. Cancer Res. 2008; 68: 2861-9
    CrossrefPubMedGoogle Scholar
  • 31 Martino A, Holmes JH, Lord JD, Moon JJ, Nelson BH. Stat5 and Spl regulate transcription of the Cyclin D2 gene in response to IL-2. J Immunol. 2001; 166: 1723-9
    PubMedGoogle Scholar
  • 32 Fernandez-de MattosS, Essafi A, Soeiro I, Pietersen AM, Birkenkamp KU, Edwards CS et al. Fox03a and BCR-ABL regulate Cyclin D2 transcription through a STAT5/BCL6-dependent mechanism. Mol Cell Biol. 2004; 24: 10058-71
    CrossrefPubMedGoogle Scholar
  • 33 Uckun FM, Ozer Z, Qazi S, Tuel-Ahlgren L, Mao C. Polo-like kinase 1 (PLK1) as a molecular target to overcome Syk-mediated resistance of B-lineage acute lymphoblastic leukemia cells to oxidative stress. Br J Haematol. 2010; 148: 714-25
    CrossrefPubMedGoogle Scholar
  • 34 Uckun FM, Zheng Y, Cetkovic-Cvrtje M, Vassilev A, Lisowski E, Waurzyniak B. In vivo pharmacokinetic features, toxicity profile, and chemosensitizing activity of alpha-cyano-beta-hydroxy-beta-methyl-N-(2,5-dibromophenyl)propen-amide (LFM-A13), a novel anti-leukemic agent targeting Bruton's tyrosine kinase. Clin Cancer Res. 2002; 8: 1224-33
    PubMedGoogle Scholar
  • 35 Uckun FM, Tibbies H, Venkatachalam TK, DuMez D, Erbeck D. Preclinical toxicity and pharmacokinetics of the Bruton's tyrosine kinase-targeting anti-leukemic drug candidate, a-cyano-β-hydroxy-β-methyl-N-(2,5-dibromophenyl) propen-amide(LFM-A13). Arzneimittelforschung. 2007; 57 (1) 31-46
    Article in Thieme ConnectPubMedGoogle Scholar
  • 36 DuMez D, Venkatachalam TK, Uckun FM. Large-scale synthesis of GMP grade α-cyano-β-hydroxy-b-methyl-N-(2,5-dibromophenyl) propenamide (LFM-A13), a new anticancer drug candidate. Arzneimittelforschung. 2007; 57 (3) 155-63
    Article in Thieme ConnectPubMedGoogle Scholar
  • 37 Uckun FM, Ek RO, Jan ST, Chen CL, Qazi S. Targeting SYK kinase-dependent anti-apoptotic resistance pathway in B-lineage acute lymphoblastic leukemia (ALL) cells with a potent SYK inhibitory pentapeptide mimic. Br J Haematol. 2010; 149: 508-17
    CrossrefPubMedGoogle Scholar
  • 38 Rhodes DR, Kalyana-Sundaram S, Mahavisno V, Varambally R, Yu J, Briggs BB et al. Oncomine 3.0:genes, pathways, and networks in a collection of 18,000 cancer gene expression profiles. Neoplasia. 2007; 9: 166-80
    CrossrefPubMedGoogle Scholar
  • 39 Uckun FM, Qazi S. Bruton's Tyrosine kinase as a molecular target in treatment of leukemias and lymphomas as well as inflammatory disorders and autoimmunity. Expert Opin Ther Patents. 2010; 20: 1457-70
    CrossrefPubMedGoogle Scholar
  • 40 Goodman PA, Wood CM, Vassilev AO, Ma C, Uckun FM. Defective expression of Bruton's tyrosine kinase in acute lymphoblastic leukemia. LeukLymphoma. 2003; 44: 1011-8
    PubMedGoogle Scholar
  • 41 Katz FE, Lovering RC, Bradley LA, Rigley KP, Brown D, Cotter F et al. Expression of the X-linked agammaglobulinemia gene, btk in B-cell acute lymphoblastic leukemia. Leukemia. 1994; 8: 574-7
    PubMedGoogle Scholar
  • 42 Trader P. Tyrosine kinases as targets in cancer therapy -successes and failures. Expert Opin Ther Targets. 2003; 7: 215-34
    CrossrefPubMedGoogle Scholar
  • 43 Pan Z. Bruton's tyrosine kinase as a drug discovery target. Drug News Perspect. 2008; 21: 357-62
    CrossrefPubMedGoogle Scholar
  • 44 Hantschel O, Rix U, Schmidt U, Bürckstümmer T, Kneidinger M, Schütze G et al. The Btk tyrosine kinase is a major target of the Bcr-Abl inhibitor dasatinib. PNAS. 2007; 104: 13283-8
    CrossrefPubMedGoogle Scholar
  • 45 Ritis K, Speletas M, Tsironidou V, Pardali E, Kanariou M, Moschese V et al. Absence of Bruton's tyrosine kinase (Btk) mutations in patients with acute myeloid leukaemia. Br J Haematol. 1998; 102: 1241-8
    CrossrefPubMedGoogle Scholar
  • 46 Davis RE, Ngo VN, Lenz G, Tolar P, Young RM, Romesser PB et al. Chronic active B cell receptor signaling in diffuse large B cell lymphoma. Nature. 2010; 463: 88-92
    CrossrefPubMedGoogle Scholar
  • 47 Kaukonen J, Lahtinen I, Laine S, Alitalo K, Palotie A. BMX tyrosine kinase gene is expressed in granulocytes and myeloid leukaemias. Br J Haematol. 1996; 94: 455-60
    PubMedGoogle Scholar
  • 48 Tu T, Thotala D, Geng L, Hallahan DE, Willey CD. Bone marrow X kinase-mediated signal transduction in irradiated vascular endothelium. Cancer Res. 2008; 68: 2861-9
    CrossrefPubMedGoogle Scholar
  • 49 Redondo PC, Ben-Amor N, Salido GM, Bartegi A, Pariente JA, Rosado JA. Ca2+-independent activation of Bruton's tyrosine kinase is required for store-mediated Ca2+ entry in human platelets. Cell Signal. 2005; 17: 1011-21
    CrossrefPubMedGoogle Scholar
  • 50 Redondo PC, Harper MT, Rosado JA, Sage SO. A role for co-filin in the activation of store-operated calcium entry by de novo conformational coupling in human platelets. Blood. 2006; 107: 973-9
    PubMedGoogle Scholar
  • 51 Bouaziz A, Amor NB, Woodard GE, Zibidi H, Lopez JJ, Bartegi A et al. Tyrosine phosphorylation/dephosphorylation balance is involved in thrombin-evoked microtubular reorganisation in human platelets. Thromb Haemost. 2007; 98: 375-84
    PubMedGoogle Scholar
  • 52 Crosby D, Poole AW. Interaction of Bruton's tyrosine kinase and protein kinase Ctheta in platelets. Cross-talk between tyrosine and serine/threonine kinases. J Biol Chem. 2002; 277: 9958-65
    CrossrefPubMedGoogle Scholar
  • 53 Uckun FM, Vassilev A, Bartell S, Zheng Y, Mahajan S, Tibbies HE. The Anti-leukemic Bruton's tyrosine kinase inhibitor α-cyano-β-hydroxy-β-methyl-N-(2,5-dibromophenyl) propenamide (LFM-A13) prevents fatal thromboembolism. Leuk Lymphoma. 2003; 44: 1569-77
    CrossrefPubMedGoogle Scholar
  • 54 Tibbies HE, Samuel P, Erbeck D, Mahajan S, Uckun FM. In vivo toxicity and antithrombotic profile of the oral formulation of the anti-leukemic agent, LFM-A13-F. Arzneimittelforschung. 2004; 54 (6) 330-5
    Article in Thieme ConnectPubMedGoogle Scholar
  • 55 Mitchell LG. A prospective cohort study determining the prevalence of thrombotic events in children with acute lymphoblastic leukemia and a central venous line who are treated with L-asparaginase. Cancer. 2003; 97: 508-13
    CrossrefPubMedGoogle Scholar
  • 56 Gomes MP, Deitcher SR. Diagnosis of venous thromboembolic disease in cancer patients. Oncology (Williston Park) 2003; 17: 126-35 139; discussion 139–44
    PubMedGoogle Scholar