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The development of imatinib as a therapeutic agent for chronic myeloid leukemia

Michael Deininger, Elisabeth Buchdunger and Brian J. Druker

Article Figures & Data

Figures

  • Figure 1.

    Lead optimization. Development of imatinib from a 2-phenylaminopyrimidine backbone (shown in white). (A) Activity in cellular assays was improved by introduction of a 3′ pyridyl group (yellow) at the 3′ position of the pyrimidine. (B) Activity against tyrosine kinases was further enhanced by addition of a benzamide group (orange) to the phenyl ring. (C) Attachment of a “flag-methyl” group (green) ortho to the diaminophenyl ring strongly reduced activity against PKC. (D) Addition of an N-methylpiperazine (purple) increased water solubility and oral bioavailability.

  • Figure 2.

    Establishment of imatinib selectivity. (A) Imatinib (CGP57148) at a concentration of 10 μM does not inhibit the growth of 32D cells (32D + IL-3, ○; 32D + IL-3 + imatinib, ▵). (B) Imatinib at 1 μM inhibits the growth of 32D cells expressing BCR-ABL (32Dp210); the effect is partially reversible with IL-3 (32Dp210 alone, ▴; + imatinib, ○; + imatinib + IL-3, □). (C) Imatinib does not affect the growth of 32D cells transformed by SRC (32Dv-SRC alone, □; + 10 μM imatinib, ○). Reproduced from Druker et al18 with permission from Nature, copyright 1996. O.D. indicates optical density. Error bars indicate mean ± standard deviation.

  • Figure 3.

    In vivo antitumor activity of imatinib. (A) Mice injected with 32D cells expressing BCR-ABL were treated with graded doses of intraperitoneal imatinib from days 11 to 18 (○, placebo; ♦, 2.5 mg/kg; ○, 10 mg/kg; ▵, 25 mg/kg; ▪, 60 mg/kg). There was a significant difference in tumor size between controls and mice treated with at least 10 mg/kg. (B) No effect was seen in mice injected with 32D cells expressing v-SRC. Reproduced from Druker et al18 with permission from Nature, copyright 1996. Error bars indicate mean ± standard deviation.

  • Figure 4.

    Rates of complete hematologic remission, complete cytogenetic remission, and molecular remission (defined as RT-PCR negativity) in the phase 2 and 3 trials with imatinib monotherapy. Blue bars indicate complete hematologic remission (CHR); green bars, complete cytogenetic remission (CCyR); red bars, molecular remission (MoR). M-BC indicates myeloid blast crisis; and AP, accelerated phase.

  • Figure 5.

    Ribbon representation of ABL in complex with imatinib and ID180970. Shown is (A) the conformation of Abl (blue) in complex with imatinib (orange), with the A-loop (magenta) in a “closed” conformation, and (B) the Abl conformation (green) in the PD180970 (red) complex with the A-loop (magenta) in an “open” conformation. The P-loop (yellow) folds down over the inhibitor in both cases. Figure prepared by Sandra W. Cowan-Jacob based on reported structures.85,88

  • Figure 6.

    Frequency of BCR-ABL mutations detected in clinical specimens (n = 177). Mutations cluster in 4 distinct regions of the kinase domain. Mutations of the P-loop (amino acids 248-255, green) are most common, followed by mutations of T315 (red), which forms a hydrogen bond with imatinib. M351 (turquoise) interacts with the SH2 domain and participates in autoregulation of kinase activity. The fourth cluster (magenta) encompasses the A-loop (amino acids 379-398).

  • Figure 7.

    Structures of alternate Abl/Src kinase inhibitors

  • Figure 8.

    Potential mechanisms underlying disease persistence (molecular refractoriness). (A) Kinase domain mutations that confer moderate resistance to imatinib. (B) BCR-ABL levels may be particularly high in the most primitive leukemic stem cells. (C) Inadequate intracellular levels of imatinib as a result of PGP expression. (D) Physiologic growth factor signaling or (E) integrin signals may maintain viability even with BCR-ABL kinase activity completely inhibited. (F) Quiescent (dormant) cells may be protected against imatinib. VLA-5 indicates very late activation antigen-5.

Tables

  • Table 1.

    Inhibition of protein kinases by imatinib

    EnzymeSubstrate phosphorylation IC50 [μM]Cellular tyrosine phosphorylation IC50 [μM]
    c-ABL 0.2; 0.025* ND
    ν-ABL 0.038 0.1-0.3
    P210BCR-ABL 0.025* 0.25
    P185BCR-ABL 0.025* 0.25
    TEL-ABL ND 0.35
    PDGF-R α and β 0.38 (PDGF-Rβ) 0.1
    Tel-PDGF-R ND 0.15
    c-KIT 0.41 0.1
    FLT-3 > 10 > 10
    Btk > 10 ND
    c-FMS and v-FMS ND > 10
    c-SRC > 100 ND
    v-SRC ND > 10
    c-LYN > 100 ND
    c-FGR > 100 ND
    LCK 9.0 ND
    SYK (TPK-IIB) > 100 ND
    JAK-2 > 100* > 100
    EGF-R > 100 > 100
    Insulin receptor > 10 > 100
    IGF-IR > 10 > 100
    FGF-R1 31.2 ND
    VEGF-R2 (KDR) 10.7 ND
    VEGF-R1 (FLT-1) 19.5 ND
    VEGF-R3 (FLT-4) 5.7 ND
    TIE-2 (TEK) > 50 ND
    c-MET > 100 ND
    PKA > 500 ND
    PPK > 500 ND
    PKCα, β1, β2, γ, δ, ϵ, ζ, or η > 100 ND
    Protein kinase CK-1, CK-2 > 100 ND
    PKB > 10 ND
    P38 > 10 ND
    PDK1 > 10 ND
    c-Raf-1 0.97 ND
    CDC2/cyclin B > 100 ND
    • Imatinib concentrations causing a 50% reduction in kinase activity (IC50) are given.

      PDGF-R indicates platelet-derived growth-factor receptor; ND, not done; Btk, Bruton tyrosine kinase; TPK, tyrosine-protein kinase; EGF-R, epidermal growth-factor receptor; IGF-IR, insulin-like growth factor receptor I, FGF-R1, fibroblast growth factor receptor 1; VEGF-R, vascular endothelial growth factor receptor; PKA, cAMP-dependent protein kinase; PPK, phosphorylase kinase; PKC, protein kinase C; CK, casein kinase; PKB, protein kinase B (also known as Akt); and PKD1, 3-phosphoinoside-dependent protein kinase 1.

    • * IC50 was determined in immunocomplex assays.

  • Table 2.

    Responses to imatinib in blast-crisis and accelerated phases of CML

    Overall hematologic response/CHR, %Sustained hematologic responses, % (> 4 wk)MCR, %CCR, %Median survival, mo
    Myeloid blast crisis, n = 229 52/15 31 16 7 6.8
    Ph+ ALL*, n = 56 59/22 27 NA NA 4.9
    Accelerated phase, % (n = 181) 82/53 69 24 17 Not reached
    Chronic phase after failure of IFN therapy 95% sustained CHR 95% sustained CHR 60 41 Not reached
    • MCR indicates major cytogenetic response; CCR, complete cytogenetic response; NA, not available; and CHR, complete hematologic response.

    • * Also lymphoid blast crisis of CML.

  • Table 3.

    Responses to imatinib versus IFN plus cytarabine in newly diagnosed patients with CML in chronic phase

    CHR, %MCR, %CCR, %Progression-free survival, 14 mo
    Imatinib, n = 553 95.3 85.2 73.8 92.1
    IFN + cytarabine, n = 553 55.5 22.1 8.5 73.5
    P .001 .001 .001 .001
    • Median duration of follow-up equaled 19 months. CHR indicates complete hematologic response; MCR, major cytogenetic response; and CCR, complete cytogenetic response.

  • Table 4.

    Frequency and in vitro sensitivity of clinically isolated BCR-ABL KD mutants to imatinib in various assays

    IC50, μM
    Wild-type ABL numbering according to ABL exon Ia/IbExchangePatients, total = 177%BCR-ABL phosphorylation in cellsABL autophosphorylation (in vitro kinase assay)Cell proliferation
    M244/M263 V 3 1.69 NR* 0.03, 0.64, 1 1, 1.6
    L248/L267 V 1 0.56 NR* NR NR
    G250/G269 E 6 3.39 7.4 to > 10 1.6 4.5 to > 20
    G250/G269 A 1 0.56 NR NR NR
    Q252/Q271 H 8 4.52 2.9, 10.4 NR 2.8 to 9.3
    Q252/Q271 R 1 0.56 10.4 NR NR
    Y253/Y272 F 6 3.39 5.9, 21.4 72 3 to 8.9
    Y253/Y272 H 12 6.78 21.4 150 17.7, > 33
    E255/E274 K 34 19.21 27.9 > 200 15 to 33
    E255/E274 V 4 2.26 7.3 3.5, > 200 7.7, > 33
    D276/D295 G 1 0.56 NR NR NR
    F311/F330 L 1 0.56 NR* 0.031, 1 0.7 to 2.1
    T315/T334 I 37 20.9 > 10 > 10 5.2 to > 10
    T315/T334 N 1 0.56 NR NR NR
    F317/F336 L 7 3.95 0.8 to 11.8 0.25, 0.38, 8.3 1.3 to 13
    M343/M362 T 1 0.56 NR NR NR
    M351/M370 T 27 15.25 0.6 to 3.9 0.26, < 1 0.9 to 7.3
    E355/E374 G 5 2.82 2.8, 10 0.02, 0.8 2.3, 4
    F359/F378 A 1 0.56 NR NR NR
    F359/F378 V 7 3.95 NR 0.05, 1.8 1.4, 2.8
    V379/V398 I 1 0.56 NR 0.03, 1.1 1, 2
    F382/F401 L 1 0.56 NR NR NR
    L387/L406 M 1 0.56 NR 0.06, 2.1 1.1, 2.2
    H396/H415 P 1 0.56 NR 0.34, 0.87 1.4 to 4.3
    H396/H415 R 6 3.39 NR 0.22, 7.3 5.4, 11
    S417/S436 Y 1 0.56 NR NR 1.8
    E459/E478 K 1 0.56 NR NR NR
    F486/F505 S 1 0.56 1.1 0.67 1.4, 9.1
    • The values are given as fold changes over wild type. The data are compiled from several studies.83,84,89-98

      NR indicates not reported.

    • * Immunoblots were published but IC50 values were not given.94

    • L267R had greater than 33-fold increased resistance in proliferation assays.

    • D276V had greater than 2-fold increased resistance in proliferation assays.

  • Table 5.

    Alternate ABL kinase inhibitors with activity against KD mutants

    IC50 cell proliferation, nM
    CompoundClassWild-type BCR-ABLE255KM351TH396PT3151Reference no.
    PD180970 Pyridopyrimidine 25 140 45 15 840 99
    SKI DV-M016 Pyridopyrimidine ∼11 ∼30 NR ∼10 > 500 119
    AP23848 Trisubstituted purine 14 94 24 8 9050 120
    BMS-354825 2-amino-thiazole-5-carboxamide 0.087-1.000 Low nanomolar range Low nanomolar range Low nanomolar range > 1000 121,122
    • NR indicates not reported.

  • Table 6.

    Examples of active agents targeting signaling pathways downstream of BCR-ABL

    TargetCompoundActivity in imatinib-resistant cellsClinical trialsReference no.
    Farnesyl transferase Lonafarnib Tipifamib Yes Yes 136-139
    MEK1 PD98059 Yes No 140
    RAF-1 BAY43-9006 Yes Yes 141
    PI3 kinase LY294002 Wortmannin Yes No 142
    mTOR Rapamycin Yes Yes 143,144
    Cyclin-dependent kinases Favopiridol Yes Yes 145