RESUMEN
A t(9;22) chromosomal translocation which forms the chimeric tyrosine kinase breakpoint cluster region (BCR)Abelson murine leukemia viral oncogene homolog 1 (ABL) is a key mechanism underlying the pathogenesis of chronic myelogenous leukemia (CML). Pharmacological inhibition of BCRABL with imatinib (Gleevec) has been reported as an effective targeted therapy; however, mutations (including the kinase domain of ABL) suppress the efficacy of inhibitors. PF114, a derivative of the third generation BCRABL inhibitor ponatinib, demonstrated a high inhibitory activity against wild-type and mutant BCRABL forms, such as the clinically important T315I. Furthermore, PF114 exhibited preferential kinase selectivity, safety, notable pharmacokinetic properties and therapeutic efficacy in a murine model. Investigation into the mechanisms of CML cell death revealed an exceptional potency of PF114 (at low nanomolar concentrations) for the CMLderived K562 cell line, whereas leukemia cell lines that lack the chimeric tyrosine kinase were markedly more refractory. The molecular ordering of events mechanistically associated with K562 cell death included the dephosphorylation of CrkL adaptor protein followed by inhibition of ERK1/2 and Akt, G1 arrest, a decrease of phosphorylated Bcl2associated death promoter, Bcl2like protein 11, BH3 interactingdomain death agonist, Bclextra large and Bcl2 family apoptosis regulator, and reduced mitochondrial transmembrane potential. Increased Annexin V reactivity, activation of caspases and poly(ADPribose)polymerase cleavage were proposed to lead to internucleosomal DNA fragmentation. Thus, PF114 may be a potent inducer of apoptosis in CML cells. Nevertheless, activation of STAT3 phosphorylation in response to PF114 may permit cell rescue; thus, a combination of BCRABL and STAT3 inhibitors should be considered for improved therapeutic outcome. Collectively, the targeted killing of BCRABLpositive cells, along with other beneficial properties, such as in vivo characteristics, suggests PF114 as a potential candidate for analysis in clinical trials with CML patients.
Asunto(s)
Proteínas de Fusión bcr-abl/antagonistas & inhibidores , Leucemia Mielógena Crónica BCR-ABL Positiva/tratamiento farmacológico , Piridinas/administración & dosificación , Triazoles/administración & dosificación , Animales , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Proteínas de Fusión bcr-abl/genética , Células HL-60 , Humanos , Células K562 , Leucemia Mielógena Crónica BCR-ABL Positiva/genética , Ratones , Mutación , Fosforilación/efectos de los fármacos , Piridinas/farmacología , Factor de Transcripción STAT3/metabolismo , Triazoles/farmacología , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Anthraquinones and their analogues, in particular heteroarene-fused anthracendiones, are prospective scaffolds for new compounds with improved antitumor characteristics. We herein report the use of a 'scaffold hopping' approach for the replacement of the core structure in the previously discovered hit compound naphtho[2,3-f]indole-5,10-dione 2 with an alternative anthra[2,3-b]furan-5,10-dione scaffold. Among 13 newly synthesized derivatives the majority of 4,11-dihydroxy-2-methyl-5,10-dioxoanthra[2,3-b]furan-3-carboxamides demonstrated a high antiproliferative potency against a panel of wild type and drug resistant tumor cell lines, a property superior over the reference drug doxorubicin or lead naphtho[2,3-f]indole-5,10-dione 2. At low micromolar concentrations the selected derivative of (R)-3-aminopyrrolidine 3c and its stereoisomer (S)-3-aminopyrrolidine 3d caused an apoptotic cell death preceded by an arrest in the G2/M phase. Studies of intracellular targets showed that 3c and 3d formed stable intercalative complexes with the duplex DNA as determined by spectral analysis and molecular docking. Both 3c and 3d attenuated topoisomerase 1 and 2 mediated unwinding of the supercoiled DNA via a mechanism different from conventional DNA-enzyme tertiary complex formation. Furthermore, 3d decreased the activity of selected human protein kinases in vitro, indicating multiple targeting by the new chemotype. Finally, 3d demonstrated an antitumor activity in a model of murine intraperitoneally transplanted P388 leukemia, achieving the increase of animal life span up to 262% at tolerable doses. Altogether, the 'scaffold hopping' demonstrated its productivity for obtaining new perspective antitumor drug candidates.
Asunto(s)
Antineoplásicos/química , Antineoplásicos/farmacología , Furanos/química , Furanos/farmacología , Inhibidores de Topoisomerasa/química , Inhibidores de Topoisomerasa/farmacología , Animales , Antracenos/química , Antracenos/farmacología , Antracenos/uso terapéutico , Antineoplásicos/uso terapéutico , Línea Celular Tumoral , Resistencia a Múltiples Medicamentos , Resistencia a Antineoplásicos , Ensayos de Selección de Medicamentos Antitumorales , Furanos/uso terapéutico , Humanos , Leucemia/tratamiento farmacológico , Leucemia/metabolismo , Ratones , Neoplasias/tratamiento farmacológico , Neoplasias/metabolismo , Inhibidores de Proteínas Quinasas/química , Inhibidores de Proteínas Quinasas/farmacología , Inhibidores de Proteínas Quinasas/uso terapéutico , Relación Estructura-Actividad , Inhibidores de Topoisomerasa/uso terapéuticoRESUMEN
The concept of immunological surveillance implies that immunogenic variants of tumor cells arising in the organism can be recognized by the immune system. Tumor progression is provided by somatic evolution of tumor cells under the pressure of the immune system. The loss of MHC Class I molecules on the surface of tumor cells is one of the most known outcomes of immune selection. This study developed a model of immune selection based on the immune response of TCR 1d1 single ß-chain transgenic B10.D2(R101) (K(d)I(d)D(b)) mice to allogeneic EL4 (H-2(b)) thymoma cells. In wild-type B10.D2(R101) mice, immunization with EL4 cells induced a vigorous CTL response targeted to the H-2K(b) molecule and results in full rejection of the tumor cells. In contrast, transgenic mice developed a compromised proliferative response in mixed-lymphocyte response assays and were unable to reject transplanted allogeneic EL4 cells. During the immune response to EL4 cells, CD8(+) T-lymphocytes with endogenous ß-chains accumulated predominantly in the spleen of transgenic mice and only a small part of the T-lymphocytes expressing transgenic ß-chains became CD8(+)CD44(+)CD62L(-) effectors. Then, instead of a full elimination of tumor cells as in wild-type mice, a reproducible prolonged equilibrium phase and subsequent escape was observed in transgenic mice that resulted in death of 90% of the mice in 40-60 days after grafting. Prolonged exposure of tumor cells to the pressure of the immune system in transgenic mice in vivo resulted in a stable loss of H-2K(b) molecules on the EL4 cell surface. Genetic manipulation of the T-lymphocyte repertoire was sufficient to reproduce the classic pattern of interactions between tumor cells and the immune system, usually observed in reliable syngeneic models of anti-tumor immunity. This newly-developed model could be used in further studies of immunoregulatory circuits common for transplantational and anti-tumor immune responses.