RESUMEN
BACKGROUND: Defects in programmed cell death, or apoptosis, are a hallmark of cancer. The anti-apoptotic B-cell lymphoma 2 (BCL-2) family proteins, including BCL-2, BCL-X(L), and MCL-1 have been characterized as key survival factors in multiple cancer types. Because cancer types with BCL2 and MCL1 amplification are more prone to inhibition of their respectively encoded proteins, we hypothesized that cancers with a significant frequency of BCL2L1 amplification would have greater dependency on BCL-X(L) for survival. METHODS: To identify tumor subtypes that have significant frequency of BCL2L1 amplification, we performed data mining using The Cancer Genome Atlas (TCGA) database. We then assessed the dependency on BCL-X(L) in a panel of cell lines using a selective and potent BCL-X(L) inhibitor, A-1155463, and BCL2L1 siRNA. Mechanistic studies on the role of BCL-X(L) were further undertaken via a variety of genetic manipulations. RESULTS: We identified colorectal cancer as having the highest frequency of BCL2L1 amplification across all tumor types examined. Colorectal cancer cell lines with BCL2L1 copy number >3 were more sensitive to A-1155463. Consistently, cell lines with high expression of BCL-XL and NOXA, a pro-apoptotic protein that antagonizes MCL-1 activity were sensitive to A-1155463. Silencing the expression of BCL-X(L) via siRNA killed the cell lines that were sensitive to A-1155463 while having little effect on lines that were resistant. Furthermore, silencing the expression of MCL-1 in resistant cell lines conferred sensitivity to A-1155463, whereas silencing NOXA abrogated sensitivity. CONCLUSIONS: This work demonstrates the utility of characterizing frequent genomic alterations to identify cancer survival genes. In addition, these studies demonstrate the utility of the highly potent and selective compound A-1155463 for investigating the role of BCL-X(L) in mediating the survival of specific tumor types, and indicate that BCL-X(L) inhibition could be an effective treatment for colorectal tumors with high BCL-X(L) and NOXA expression.
Asunto(s)
Neoplasias Colorrectales/genética , Genómica , Proteína bcl-X/genética , Benzotiazoles/farmacología , Línea Celular Tumoral , Supervivencia Celular/efectos de los fármacos , Supervivencia Celular/genética , Análisis por Conglomerados , Neoplasias Colorrectales/metabolismo , Neoplasias Colorrectales/patología , Expresión Génica , Perfilación de la Expresión Génica , Humanos , Isoquinolinas/farmacología , Proteínas Proto-Oncogénicas c-bcl-2/genética , Proteínas Proto-Oncogénicas c-bcl-2/metabolismo , Proteína bcl-X/antagonistas & inhibidores , Proteína bcl-X/metabolismoRESUMEN
BACKGROUND: Evasion of apoptosis is a hallmark of cancer cells. One mechanism to deregulate the apoptotic pathway is by upregulation of the anti-apoptotic Bcl-2 family members. Navitoclax (ABT-263) is a Bcl-2/Bcl-xL inhibitor that restores the ability of cancer cells to undergo apoptosis. METHODS: In this study we performed a high-throughput screen with 640 FDA-approved drugs to identify potential therapeutic combinations with navitoclax in a non-small cell lung cancer (NSCLC) cell line. RESULTS: Other than a panel of cancer compounds such as doxorubicin, camptothecin, and docetaxel, four antihelminthic compounds (benzimidazoles) potentiated navitoclax activity. Treatment with benzimidazoles led to induction of the pro-apoptotic protein Noxa at the mRNA and protein level. Noxa binds and antagonizes antiapoptotic protein Mcl-1. siRNA-mediated knock-down of Noxa completely rescued benzimidazole-potentiated navitoclax activity. In addition, inhibiting caspase 3 and 9 partially rescued benzimidazole-potentiated navitoclax activity. CONCLUSIONS: We have identified compounds and mechanisms which potentiate navitoclax activity in lung cancer cell lines. Further validation of the benzimidazole-potentiated navitoclax effect in vivo is required to evaluate the potential for translating this observation into clinical benefit.
RESUMEN
Immunotherapy of cancer has made tremendous progress in recent years, as demonstrated by the remarkable clinical responses obtained from adoptive cell transfer (ACT) of patient-derived tumor infiltrating lymphocytes, chimeric antigen receptor (CAR)-modified T cells (CAR-T) and T cell receptor (TCR)-engineered T cells (TCR-T). TCR-T uses specific TCRS optimized for tumor engagement and can recognize epitopes derived from both cell-surface and intracellular targets, including tumor-associated antigens, cancer germline antigens, viral oncoproteins, and tumor-specific neoantigens (neoAgs) that are largely sequestered in the cytoplasm and nucleus of tumor cells. Moreover, as TCRS are naturally developed for sensitive antigen detection, they are able to recognize epitopes at far lower concentrations than required for CAR-T activation. Therefore, TCR-T holds great promise for the treatment of human cancers. In this focused review, we summarize basic, translational, and clinical insights into the challenges and opportunities of TCR-T. We review emerging strategies used in current ACT, point out limitations, and propose possible solutions. We highlight the importance of targeting tumor-specific neoAgs and outline a strategy of combining neoAg vaccines, checkpoint blockade therapy, and adoptive transfer of neoAg-specific TCR-T to produce a truly tumor-specific therapy, which is able to penetrate into solid tumors and resist the immunosuppressive tumor microenvironment. We believe such a combination approach should lead to a significant improvement in cancer immunotherapies, especially for solid tumors, and may provide a general strategy for the eradication of multiple cancers.
Asunto(s)
Neoplasias , Receptores Quiméricos de Antígenos , Epítopos , Humanos , Inmunoterapia Adoptiva , Neoplasias/terapia , Receptores de Antígenos de Linfocitos T/genética , Receptores Quiméricos de Antígenos/genética , Linfocitos T , Microambiente TumoralRESUMEN
TRAIL can activate cell surface death receptors, resulting in potent tumor cell death via induction of the extrinsic apoptosis pathway. Eftozanermin alfa (ABBV-621) is a second generation TRAIL receptor agonist engineered as an IgG1-Fc mutant backbone linked to two sets of trimeric native single-chain TRAIL receptor binding domain monomers. This hexavalent agonistic fusion protein binds to the death-inducing DR4 and DR5 receptors with nanomolar affinity to drive on-target biological activity with enhanced caspase-8 aggregation and death-inducing signaling complex formation independent of FcγR-mediated cross-linking, and without clinical signs or pathologic evidence of toxicity in nonrodent species. ABBV-621 induced cell death in approximately 36% (45/126) of solid cancer cell lines in vitro at subnanomolar concentrations. An in vivo patient-derived xenograft (PDX) screen of ABBV-621 activity across 15 different tumor indications resulted in an overall response (OR) of 29% (47/162). Although DR4 (TNFSFR10A) and/or DR5 (TNFSFR10B) expression levels did not predict the level of response to ABBV-621 activity in vivo, KRAS mutations were associated with elevated TNFSFR10A and TNFSFR10B and were enriched in ABBV-621-responsive colorectal carcinoma PDX models. To build upon the OR of ABBV-621 monotherapy in colorectal cancer (45%; 10/22) and pancreatic cancer (35%; 7/20), we subsequently demonstrated that inherent resistance to ABBV-621 treatment could be overcome in combination with chemotherapeutics or with selective inhibitors of BCL-XL. In summary, these data provide a preclinical rationale for the ongoing phase 1 clinical trial (NCT03082209) evaluating the activity of ABBV-621 in patients with cancer. SIGNIFICANCE: This study describes the activity of a hexavalent TRAIL-receptor agonistic fusion protein in preclinical models of solid tumors that mechanistically distinguishes this molecular entity from other TRAIL-based therapeutics.
Asunto(s)
Neoplasias Colorrectales/tratamiento farmacológico , Factor IX/farmacología , Fragmentos Fc de Inmunoglobulinas/farmacología , Neoplasias Pancreáticas/tratamiento farmacológico , Receptores del Ligando Inductor de Apoptosis Relacionado con TNF/metabolismo , Proteínas Recombinantes de Fusión/farmacología , Ligando Inductor de Apoptosis Relacionado con TNF/metabolismo , Animales , Antineoplásicos/farmacología , Apoptosis , Proliferación Celular , Neoplasias Colorrectales/genética , Neoplasias Colorrectales/metabolismo , Neoplasias Colorrectales/patología , Femenino , Humanos , Ratones , Ratones Endogámicos NOD , Ratones SCID , Neoplasias Pancreáticas/genética , Neoplasias Pancreáticas/metabolismo , Neoplasias Pancreáticas/patología , Receptores del Ligando Inductor de Apoptosis Relacionado con TNF/genética , Ligando Inductor de Apoptosis Relacionado con TNF/genética , Células Tumorales Cultivadas , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
BACKGROUND: The Wilms' tumor antigen (WT1) is an attractive target for immunotherapy of leukemia. In the past, we isolated and characterized the specificity and function of a WT1-specific T-cell receptor. The goal of this translational study was to develop a safe and efficient WT1-T-cell receptor retroviral vector for an adoptive immunotherapy trial with engineered T cells. DESIGN AND METHODS: We generated a panel of retroviral constructs containing unmodified or codon-optimized WT1-T-cell receptor alpha and beta genes, linked via internal ribosome entry sites or 2A sequences, with or without an additional inter-chain disulfide bond in the T-cell receptor constant domains. These constructs were functionally analyzed in vitro, and the best one was tested in an autologous primary leukemia model in vivo. RESULTS: We identified a WT1-T-cell receptor construct that showed optimal tetramer staining, antigen-specific cytokine production and killing activity when introduced into primary human T cells. Fresh CD34(+) cells purified from a patient with leukemia were engrafted into NOD/SCID mice, followed by adoptive immunotherapy with patient's autologous T cells transduced with the WT1-T-cell receptor. This therapeutic treatment evidently decreased leukemia engraftment in mice and resulted in a substantial improvement of leukemia-free survival. CONCLUSIONS: This is the first report that patient's T cells, engineered to express the WT1-T-cell receptor, can eliminate autologous leukemia progenitor cells in an in vivo model. This study provides a firm basis for the planned WT1-T-cell receptor gene therapy trial in leukemia patients.
Asunto(s)
Antígenos de Neoplasias/inmunología , Crisis Blástica/inmunología , Ingeniería Genética/métodos , Leucemia/inmunología , Receptores de Antígenos de Linfocitos T/genética , Receptores de Antígenos de Linfocitos T/uso terapéutico , Linfocitos T/patología , Tumor de Wilms/patología , Adulto , Animales , Antígenos de Neoplasias/genética , Antígenos de Neoplasias/metabolismo , Crisis Blástica/genética , Crisis Blástica/terapia , Terapia Genética/métodos , Vectores Genéticos/biosíntesis , Vectores Genéticos/química , Virus de la Hepatitis B de la Marmota/genética , Humanos , Células Jurkat , Leucemia/genética , Leucemia/terapia , Ratones , Ratones Endogámicos NOD , Ratones SCID , Receptores de Antígenos de Linfocitos T/fisiología , Linfocitos T/inmunología , Linfocitos T/metabolismo , Trasplante Autólogo/inmunología , Tumor de Wilms/inmunología , Tumor de Wilms/terapiaRESUMEN
MCL-1 is one of the most frequently amplified genes in cancer, facilitating tumor initiation and maintenance and enabling resistance to anti-tumorigenic agents including the BCL-2 selective inhibitor venetoclax. The expression of MCL-1 is maintained via P-TEFb-mediated transcription, where the kinase CDK9 is a critical component. Consequently, we developed a series of potent small-molecule inhibitors of CDK9, exemplified by the orally active A-1592668, with CDK selectivity profiles that are distinct from related molecules that have been extensively studied clinically. Short-term treatment with A-1592668 rapidly downregulates RNA pol-II (Ser 2) phosphorylation resulting in the loss of MCL-1 protein and apoptosis in MCL-1-dependent hematologic tumor cell lines. This cell death could be attenuated by either inhibiting caspases or overexpressing BCL-2 protein. Synergistic cell killing was also observed between A-1592668 or the related analog A-1467729, and venetoclax in a number of hematologic cell lines and primary NHL patient samples. Importantly, the CDK9 inhibitor plus venetoclax combination was well tolerated in vivo and demonstrated efficacy superior to either agent alone in mouse models of lymphoma and AML. These data indicate that CDK9 inhibitors could be highly efficacious in tumors that depend on MCL-1 for survival or when used in combination with venetoclax in malignancies dependent on MCL-1 and BCL-2.
Asunto(s)
Antineoplásicos/farmacología , Quinasa 9 Dependiente de la Ciclina/antagonistas & inhibidores , Neoplasias Hematológicas , Inhibidores de Proteínas Quinasas/farmacología , Animales , Apoptosis/efectos de los fármacos , Compuestos Bicíclicos Heterocíclicos con Puentes/farmacología , Línea Celular Tumoral , Supervivencia Celular/efectos de los fármacos , Sinergismo Farmacológico , Humanos , Ratones , Sulfonamidas/farmacología , Células Tumorales Cultivadas , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Herein we describe the discovery of A-1331852, a first-in-class orally active BCL-XL inhibitor that selectively and potently induces apoptosis in BCL-XL-dependent tumor cells. This molecule was generated by re-engineering our previously reported BCL-XL inhibitor A-1155463 using structure-based drug design. Key design elements included rigidification of the A-1155463 pharmacophore and introduction of sp3-rich moieties capable of generating highly productive interactions within the key P4 pocket of BCL-XL. A-1331852 has since been used as a critical tool molecule for further exploring BCL-2 family protein biology, while also representing an attractive entry into a drug discovery program.
RESUMEN
The synthesis and structure-activity relationships (SAR) of Chk1 inhibitors based on a 5,10-dihydro-dibenzo[b,e][1,4]diazepin-11-one core are described. Specifically, an exploration of the 7 and 8 positions on this previously disclosed core afforded compounds with improved enzymatic and cellular potency.
Asunto(s)
Antineoplásicos/farmacología , Benzodiazepinonas/farmacología , Diseño de Fármacos , Inhibidores Enzimáticos/farmacología , Proteínas Quinasas/metabolismo , Antineoplásicos/síntesis química , Benzodiazepinonas/síntesis química , Línea Celular Tumoral/efectos de los fármacos , Quinasa 1 Reguladora del Ciclo Celular (Checkpoint 1) , Inhibidores Enzimáticos/síntesis química , Células HeLa , Humanos , Modelos Químicos , Unión Proteica , Relación Estructura-ActividadRESUMEN
A novel series of 5,10-dihydro-dibenzo[b,e][1,4]diazepin-11-ones have been synthesized as potent and selective checkpoint kinase 1 (Chk1) inhibitors via structure-based design. Aided by protein X-ray crystallography, medicinal chemistry efforts led to the identification of compound 46d, with potent enzymatic activity against Chk1 kinase. While maintaining a low cytotoxicity of its own, compound 46d exhibited a strong ability to abrogate G2 arrest and increased the cytotoxicity of camptothecin by 19-fold against SW620 cells. Pharmacokinetic studies revealed that it had a moderate bioavailabilty of 20% in mice. Two important binding interactions between compound 46b and Chk1 kinase, revealed by X-ray cocrystal structure, were hydrogen bonds between the hinge region and the amide bond of the core structure and a hydrogen bond between the methoxy group and Lys38 of the protein.
Asunto(s)
Antineoplásicos/síntesis química , Azepinas/síntesis química , Benzodiazepinonas/síntesis química , Inhibidores de Proteínas Quinasas/síntesis química , Proteínas Quinasas/metabolismo , Animales , Antineoplásicos/química , Antineoplásicos/farmacología , Azepinas/química , Azepinas/farmacología , Benzodiazepinonas/química , Benzodiazepinonas/farmacología , Disponibilidad Biológica , Camptotecina/farmacología , Línea Celular Tumoral , Quinasa 1 Reguladora del Ciclo Celular (Checkpoint 1) , Cristalografía por Rayos X , Doxorrubicina/farmacología , Diseño de Fármacos , Sinergismo Farmacológico , Humanos , Ratones , Modelos Moleculares , Unión Proteica , Inhibidores de Proteínas Quinasas/química , Inhibidores de Proteínas Quinasas/farmacología , Proteínas Quinasas/química , Relación Estructura-ActividadRESUMEN
Mammalian cells initiate cell cycle arrest at different phases of the cell cycle in response to various forms of genotoxic stress to allow time for DNA repair, and thus preserving their genomic integrity. The protein kinases checkpoint kinase 1 (Chk1), checkpoint kinase 2 (Chk2), and mitogen-activated protein kinase-activated protein kinase 2 (MK2) have all been shown to be involved in cell cycle checkpoint control. Recently, cell cycle checkpoint abrogation has been proposed as one way to sensitize cancer cells to DNA-damaging agents due to the expected induction of mitotic catastrophe. Due to their overlapping substrate spectra and redundant functions, it is still not clear which kinase is mainly responsible for the cell cycle arrests conferred by clinically relevant chemotherapeutics. Thus, the issue remains about which kinase is the most therapeutically relevant target and, more importantly, whether multiple kinases might need to be targeted to achieve the best efficacy in light of recent studies showing superior efficacy for pan-receptor tyrosine kinase inhibitors. To clarify this issue, we investigated the roles of the three kinases in response to different genotoxic stresses through small interfering RNA-mediated specific target knockdowns. Our result showed that only the down-regulation of Chk1, but not of Chk2 or MK2, abrogated camptothecin- or 5-fluorouracil-induced S-phase arrest or doxorubicin-induced G(2)-phase arrest. This was followed by mitotic catastrophe and apoptosis. Moreover, double inhibition of Chk1 and Chk2 failed to achieve better efficacy than Chk1 inhibition alone; surprisingly, inhibition of MK2, in addition to Chk1 suppression, partially reversed the checkpoint abrogation and negated mitotic catastrophe. We further showed that this is due to the fact that in MK2-deficient cells, Cdc25A protein, which is critically required for the mitotic progression following checkpoint abrogation, becomes greatly depleted. In summary, our findings show that Chk1 is the only relevant checkpoint kinase as a cancer drug target and inhibition of other checkpoint kinases in addition to Chk1 would be nonproductive.
Asunto(s)
Ciclo Celular/fisiología , Daño del ADN/fisiología , Proteínas Quinasas/metabolismo , Proteínas Serina-Treonina Quinasas/metabolismo , Antineoplásicos/farmacología , Camptotecina/farmacología , Ciclo Celular/efectos de los fármacos , Quinasa 1 Reguladora del Ciclo Celular (Checkpoint 1) , Quinasa de Punto de Control 2 , Doxorrubicina/farmacología , Femenino , Fluorouracilo/farmacología , Células HeLa , Histonas/efectos de los fármacos , Histonas/genética , Humanos , Péptidos y Proteínas de Señalización Intracelular , Neoplasias/tratamiento farmacológico , Proteínas Quinasas/efectos de los fármacos , Proteínas Quinasas/genética , Proteínas Serina-Treonina Quinasas/efectos de los fármacos , Proteínas Serina-Treonina Quinasas/genética , ARN Interferente Pequeño , Células Tumorales Cultivadas , Fosfatasas cdc25/efectos de los fármacos , Fosfatasas cdc25/metabolismoRESUMEN
Chk1 is the major mediator in the activation of cell-cycle checkpoints in response to a variety of genotoxic stresses. We have previously shown that inhibition of Chk1 sensitizes tumor cells to topoisomerase inhibitors such as camptothecin and doxorubicin through abrogation of cell-cycle arrest (S or G2/M checkpoints). However, it was not clear whether inhibition of Chk1 could potentiate antimetabolites, a mainstay of cancer therapy, which confer genotoxic stress through a different mechanism than topoisomerase inhibitors. 5-Fluorouracil (5-FU) is the most widely used antimetabolite in the treatment of colorectal, breast and other major types of cancers. Here we demonstrate that 5-FU activates Chk1 and induces an early S-phase arrest. Chk1 downregulation abrogates this arrest and dramatically sensitizes tumor cells to the cytotoxic effects of 5-FU. 5-FU confers S-phase arrest through Chk1-mediated Cdc25A proteolysis leading to inhibition of Cdk2. Chk1 elimination stabilizes the Cdc25A protein and results in the abrogation of the S checkpoint and resumption of DNA synthesis, which leads to excessive accumulation of double-stranded DNA breaks. As a result, downregulation of Chk1 potentiates 5-FU efficacy through induction of premature chromosomal condensation followed by apoptosis. Interestingly, the profiles of various cell-cycle markers indicate that cells progress to early M phase to induce apoptosis after checkpoint abrogation. Yet, cells fail to increase their DNA content to 4N as revealed by FACS analysis, probably due to the dramatic induction of double-stranded DNA breaks and chromosomal fragmentation. This is significantly different from the cell-cycle profiles observed in the potentiation of topoisomerase inhibitors by Chk1 siRNA, which showed mitotic progression with 4N DNA content leading to mitotic catastrophe after abrogation of the S or G2 checkpoint. Thus, our results illustrate a novel mode of checkpoint abrogation and cell death conferred by Chk1 inhibition. Additionally, we show that Chk1 deficiency potentiates 5-FU efficacy through the preferential induction of the caspase-8 pathway and subsequent caspase-3 activation. In conclusion, we have clearly demonstrated that inhibition of Chk1 not only potentiates the toxicity of conventional DNA-damaging agents such as ionizing radiation and topoisomerase inhibitors, but also enhances the toxicity of antimetabolites in cancer cell lines. This discovery reveals novel scope of checkpoint abrogation and will significantly broaden the potential application of Chk1 inhibitors in cancer therapy if they do not potentiate the toxicity of 5-FU in normal cells.
Asunto(s)
Antimetabolitos Antineoplásicos/toxicidad , Daño del ADN , Regulación hacia Abajo , Fluorouracilo/toxicidad , Proteínas Quinasas/fisiología , Apoptosis/fisiología , Caspasa 3 , Caspasa 8 , Caspasas/fisiología , Ciclo Celular/efectos de los fármacos , Ciclo Celular/fisiología , Núcleo Celular/química , Núcleo Celular/efectos de los fármacos , Quinasa 1 Reguladora del Ciclo Celular (Checkpoint 1) , ADN/análisis , ADN/efectos de los fármacos , Replicación del ADN/fisiología , Resistencia a Antineoplásicos , Células HeLa , Humanos , Fosforilación , Poli(ADP-Ribosa) Polimerasas/fisiología , Proteínas Quinasas/genética , ARN Interferente Pequeño/genéticaRESUMEN
The BCL-2/BCL-XL/BCL-W inhibitor ABT-263 (navitoclax) has shown promising clinical activity in lymphoid malignancies such as chronic lymphocytic leukemia. However, its efficacy in these settings is limited by thrombocytopenia caused by BCL-XL inhibition. This prompted the generation of the BCL-2-selective inhibitor venetoclax (ABT-199/GDC-0199), which demonstrates robust activity in these cancers but spares platelets. Navitoclax has also been shown to enhance the efficacy of docetaxel in preclinical models of solid tumors, but clinical use of this combination has been limited by neutropenia. We used venetoclax and the BCL-XL-selective inhibitors A-1155463 and A-1331852 to assess the relative contributions of inhibiting BCL-2 or BCL-XL to the efficacy and toxicity of the navitoclax-docetaxel combination. Selective BCL-2 inhibition suppressed granulopoiesis in vitro and in vivo, potentially accounting for the exacerbated neutropenia observed when navitoclax was combined with docetaxel clinically. By contrast, selectively inhibiting BCL-XL did not suppress granulopoiesis but was highly efficacious in combination with docetaxel when tested against a range of solid tumors. Therefore, BCL-XL-selective inhibitors have the potential to enhance the efficacy of docetaxel in solid tumors and avoid the exacerbation of neutropenia observed with navitoclax. These studies demonstrate the translational utility of this toolkit of selective BCL-2 family inhibitors and highlight their potential as improved cancer therapeutics.
Asunto(s)
Regulación Neoplásica de la Expresión Génica , Neoplasias/tratamiento farmacológico , Proteínas Proto-Oncogénicas c-bcl-2/antagonistas & inhibidores , Administración Oral , Compuestos de Anilina/uso terapéutico , Animales , Antineoplásicos/uso terapéutico , Benzotiazoles/química , Compuestos Bicíclicos Heterocíclicos con Puentes/uso terapéutico , Línea Celular Tumoral , Supervivencia Celular , Docetaxel , Perfilación de la Expresión Génica , Granulocitos/metabolismo , Humanos , Isoquinolinas/química , Cinética , Ratones , Trasplante de Neoplasias , Neoplasias/metabolismo , Neutropenia/inducido químicamente , Neutrófilos/efectos de los fármacos , Proteínas Proto-Oncogénicas c-bcl-2/metabolismo , Sulfonamidas/uso terapéutico , Taxoides/efectos adversos , Trombocitopenia/inducido químicamente , Proteína bcl-X/antagonistas & inhibidores , Proteína bcl-X/metabolismoRESUMEN
Myeloid cell leukemia 1 (MCL-1) is a BCL-2 family protein that has been implicated in the progression and survival of multiple tumor types. Herein we report a series of MCL-1 inhibitors that emanated from a high throughput screening (HTS) hit and progressed via iterative cycles of structure-guided design. Advanced compounds from this series exhibited subnanomolar affinity for MCL-1 and excellent selectivity over other BCL-2 family proteins as well as multiple kinases and GPCRs. In a MCL-1 dependent human tumor cell line, administration of compound 30b rapidly induced caspase activation with associated loss in cell viability. The small molecules described herein thus comprise effective tools for studying MCL-1 biology.
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Antineoplásicos/química , Antineoplásicos/farmacología , Diseño de Fármacos , Mieloma Múltiple/tratamiento farmacológico , Proteína 1 de la Secuencia de Leucemia de Células Mieloides/química , Neoplasias Pancreáticas/tratamiento farmacológico , Apoptosis/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Cristalografía por Rayos X , Bases de Datos Factuales , Ensayos Analíticos de Alto Rendimiento , Humanos , Simulación del Acoplamiento Molecular , Estructura Molecular , Mieloma Múltiple/metabolismo , Mieloma Múltiple/patología , Proteína 1 de la Secuencia de Leucemia de Células Mieloides/metabolismo , Neoplasias Pancreáticas/metabolismo , Neoplasias Pancreáticas/patología , Unión Proteica , Relación Estructura-Actividad , Células Tumorales CultivadasRESUMEN
Maintenance of nuclear architecture is crucial for gene regulation, cell proliferation and tissue development. However, during every open mitosis and meiosis, chromosomes are exposed to cytoskeletal forces until they are fully reassembled into mature nuclei. Here we discuss our recent study of nuclear assembly in Xenopus egg extracts, where we showed that the DNA binding protein Developmental pluripotency associated 2 (Dppa2) directly inhibits microtubule polymerization during nuclear formation, and that this is essential for normal nuclear shape and replication. We explore mechanisms by which microtubule dynamics could regulate nuclear formation and morphology, and discuss the importance of both spatial and temporal regulation of microtubules in this process. Moreover, expression of Dppa2 is limited to the early embryo and pluripotent tissues, and we highlight the specific demands of mitosis in these often rapidly dividing cells, in which telophase nuclear assembly must be expedited and may facilitate developmental changes in nuclear architecture.
Asunto(s)
Núcleo Celular/ultraestructura , Microtúbulos/metabolismo , Proteínas Nucleares/metabolismo , Proteínas de Xenopus/metabolismo , Animales , Cromosomas/genética , Replicación del ADN , Femenino , Meiosis , Mitosis , Proteínas Nucleares/genética , Xenopus/genética , Proteínas de Xenopus/genéticaRESUMEN
Immune escape is a fundamental trait of cancer. Dendritic cells (DC) that interact with T cells represent a crucial site for the development of tolerance to tumor antigens, but there remains incomplete knowledge about how DC-tolerizing signals evolve during tumorigenesis. In this study, we show that DCs isolated from patients with metastatic or locally advanced breast cancer express high levels of the adiponectin receptors AdipoR1 and AdipoR2, which are sufficient to blunt antitumor immunity. Mechanistic investigations of ligand-receptor interactions on DCs revealed novel signaling pathways for each receptor. AdipoR1 stimulated IL10 production by activating the AMPK and MAPKp38 pathways, whereas AdipoR2 modified inflammatory processes by activating the COX-2 and PPARγ pathways. Stimulation of these pathways was sufficient to block activation of NF-κB in DC, thereby attenuating their ability to stimulate antigen-specific T-cell responses. Together, our findings reveal novel insights into how DC-tolerizing signals evolve in cancer to promote immune escape. Furthermore, by defining a critical role for adiponectin signaling in this process, our work suggests new and broadly applicable strategies for immunometabolic therapy in patients with cancer.
Asunto(s)
Neoplasias de la Mama/inmunología , Células Dendríticas/metabolismo , Receptores de Adiponectina/metabolismo , Escape del Tumor , Adiponectina/fisiología , Animales , Neoplasias de la Mama/patología , Línea Celular Tumoral , Anergia Clonal , Ciclooxigenasa 2/metabolismo , Citotoxicidad Inmunológica , Progresión de la Enfermedad , Activación Enzimática , Femenino , Humanos , Interleucina-10/metabolismo , Sistema de Señalización de MAP Quinasas , Ratones Endogámicos C57BL , FN-kappa B/metabolismo , Trasplante de Neoplasias , PPAR gamma/metabolismo , Linfocitos T Citotóxicos/inmunologíaRESUMEN
A-1155463, a highly potent and selective BCL-XL inhibitor, was discovered through nuclear magnetic resonance (NMR) fragment screening and structure-based design. This compound is substantially more potent against BCL-XL-dependent cell lines relative to our recently reported inhibitor, WEHI-539, while possessing none of its inherent pharmaceutical liabilities. A-1155463 caused a mechanism-based and reversible thrombocytopenia in mice and inhibited H146 small cell lung cancer xenograft tumor growth in vivo following multiple doses. A-1155463 thus represents an excellent tool molecule for studying BCL-XL biology as well as a productive lead structure for further optimization.
RESUMEN
Nuclear shape and size vary between species, during development, and in many tissue pathologies, but the causes and effects of these differences remain poorly understood. During fertilization, sperm nuclei undergo a dramatic conversion from a heavily compacted form into decondensed, spherical pronuclei, accompanied by rapid nucleation of microtubules from centrosomes. Here we report that the assembly of the spherical nucleus depends on a critical balance of microtubule dynamics, which is regulated by the chromatin-binding protein Developmental pluripotency-associated 2 (Dppa2). Whereas microtubules normally promote sperm pronuclear expansion, in Dppa2-depleted Xenopus egg extracts excess microtubules cause pronuclear assembly defects, leading to abnormal morphology and disorganized DNA replication. Dppa2 inhibits microtubule polymerization in vitro, and Dppa2 activity is needed at a precise time and location during nascent pronuclear formation. This demonstrates a strict spatiotemporal requirement for local suppression of microtubules during nuclear formation, fulfilled by chromatin-bound microtubule regulators.
Asunto(s)
Núcleo Celular/metabolismo , Cromatina/metabolismo , Microtúbulos/metabolismo , Proteínas Nucleares/metabolismo , Proteínas de Xenopus/metabolismo , Animales , Núcleo Celular/ultraestructura , Replicación del ADN , Microtúbulos/ultraestructura , Proteínas Nucleares/genética , Polimerizacion , Xenopus , Proteínas de Xenopus/genéticaRESUMEN
Proteins in the B cell CLL/lymphoma 2 (BCL-2) family are key regulators of the apoptotic process. This family comprises proapoptotic and prosurvival proteins, and shifting the balance toward the latter is an established mechanism whereby cancer cells evade apoptosis. The therapeutic potential of directly inhibiting prosurvival proteins was unveiled with the development of navitoclax, a selective inhibitor of both BCL-2 and BCL-2-like 1 (BCL-X(L)), which has shown clinical efficacy in some BCL-2-dependent hematological cancers. However, concomitant on-target thrombocytopenia caused by BCL-X(L) inhibition limits the efficacy achievable with this agent. Here we report the re-engineering of navitoclax to create a highly potent, orally bioavailable and BCL-2-selective inhibitor, ABT-199. This compound inhibits the growth of BCL-2-dependent tumors in vivo and spares human platelets. A single dose of ABT-199 in three patients with refractory chronic lymphocytic leukemia resulted in tumor lysis within 24 h. These data indicate that selective pharmacological inhibition of BCL-2 shows promise for the treatment of BCL-2-dependent hematological cancers.
Asunto(s)
Antineoplásicos/farmacología , Plaquetas/efectos de los fármacos , Compuestos Bicíclicos Heterocíclicos con Puentes/farmacología , Neoplasias Hematológicas/tratamiento farmacológico , Proteínas Proto-Oncogénicas c-bcl-2/antagonistas & inhibidores , Sulfonamidas/farmacología , Compuestos de Anilina/farmacología , Animales , Antineoplásicos/uso terapéutico , Apoptosis/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Perros , Femenino , Células HeLa , Humanos , Ratones , Ratones SCID , Proteínas Proto-Oncogénicas c-bcl-2/química , Carga Tumoral , Ensayos Antitumor por Modelo de Xenoinjerto , Proteína bcl-X/antagonistas & inhibidoresRESUMEN
A hallmark of mitosis is the appearance of high levels of histone phosphorylation, yet the roles of these modifications remain largely unknown. Here, we demonstrate that histone H3 phosphorylated at threonine 3 is directly recognized by an evolutionarily conserved binding pocket in the BIR domain of Survivin, which is a member of the chromosomal passenger complex (CPC). This binding mediates recruitment of the CPC to chromosomes and the resulting activation of its kinase subunit Aurora B. Consistently, modulation of the kinase activity of Haspin, which phosphorylates H3T3, leads to defects in the Aurora B-dependent processes of spindle assembly and inhibition of nuclear reformation. These findings establish a direct cellular role for mitotic histone H3T3 phosphorylation, which is read and translated by the CPC to ensure accurate cell division.