RESUMEN
Cyclin dependent kinase-like 5 (CDKL5) Deficiency Disorder (CDD) is a rare neurodevelopmental disorder caused by a mutation in the X-linked CDKL5 gene. CDKL5 is a serine/threonine kinase that is critical for axon outgrowth, dendritic morphogenesis, as well as synapse formation, maturation, and maintenance. This disorder is characterized by early-onset epilepsy, hypotonia, and failure to reach cognitive and motor developmental milestones. Because the disease is monogenic, delivery of the CDKL5 gene to the brain of patients should provide clinical benefit. To this end, we designed a gene therapy vector, adeno-associated virus (AAV)9.Syn.hCDKL5, in which human CDKL5 gene expression is driven by the synapsin promoter. In biodistribution studies conducted in mice, intracerebroventricular (ICV) injection resulted in broader, more optimal biodistribution than did intracisterna magna (ICM) delivery. AAV9.Syn.hCDKL5 treatment increased phosphorylation of EB2, a bona fide CDKL5 substrate, demonstrating biological activity in vivo. Our data provides proof-of-concept that ICV delivery of AAV9.Syn.hCDKL5 to neonatal male Cdkl5 knockout mice reduces pathology and reduces aberrant behavior. Functional improvements were seen at doses of 3e11 to 5e11 vector genomes (vg)/g brain, which resulted in transfection of ≥50% of the neurons. Functional improvements were not seen at lower doses suggesting a requirement for broad distribution for efficacy.
RESUMEN
Dihydroorotate dehydrogenase (DHODH) is the enzyme that catalyzes a rate-determining step during the de novo synthesis of uridine, an important source of cellular pyrimidine nucleotides. Ability to modulate the activity of this enzyme may be used to control diseases associated with rapid, out-of-control cell growth in oncology, immunology, and virology. Emvododstat (PTC299) is a tetrahydro-ß-carboline DHODH inhibitor discovered through the GEMS technology (Gene Expression Modulation by Small-Molecules). Described in this paper is the lead optimization campaign that culminated in the discovery of this highly potent DHODH inhibitor.
Asunto(s)
Oxidorreductasas actuantes sobre Donantes de Grupo CH-CH , Dihidroorotato Deshidrogenasa , Inhibidores Enzimáticos/farmacología , CarbamatosRESUMEN
Blocking the pyrimidine nucleotide de novo synthesis pathway by inhibiting dihydroorotate dehydrogenase (DHODH) results in the cell cycle arrest and/or differentiation of rapidly proliferating cells including activated lymphocytes, cancer cells, or virally infected cells. Emvododstat (PTC299) is an orally bioavailable small molecule that inhibits DHODH. We evaluated the potential for emvododstat to inhibit the progression of acute myeloid leukemia (AML) using several in vitro and in vivo models of the disease. Broad potent activity was demonstrated against multiple AML cell lines, AML blasts cultured ex vivo from patient blood samples, and AML tumor models including patient-derived xenograft models. Emvododstat induced differentiation, cytotoxicity, or both in primary AML patient blasts cultured ex vivo with 8 of 10 samples showing sensitivity. AML cells with diverse driver mutations were sensitive, suggesting the potential of emvododstat for broad therapeutic application. AML cell lines that are not sensitive to emvododstat are likely to be more reliant on the salvage pathway than on de novo synthesis of pyrimidine nucleotides. Pharmacokinetic experiments in rhesus monkeys demonstrated that emvododstat levels rose rapidly after oral administration, peaking about 2 hours post-dosing. This was associated with an increase in the levels of dihydroorotate (DHO), the substrate for DHODH, within 2 hours of dosing indicating that DHODH inhibition is rapid. DHO levels declined as drug levels declined, consistent with the reversibility of DHODH inhibition by emvododstat. These preclinical findings provide a rationale for clinical evaluation of emvododstat in an ongoing Phase 1 study of patients with relapsed/refractory acute leukemias.
RESUMEN
Growing evidence has demonstrated that clonogenic cancer stem (initiating) cells are responsible for tumor regrowth and disease relapse. Bmi-1 plays a critical role in the self-renewal of adult stem cells. The Bmi-1 protein is elevated in many types of cancers, and experimental reduction of Bmi-1 protein levels by small interfering RNA (siRNA) causes apoptosis and/or senescence in tumor cells in vitro and increases susceptibility to cytotoxic agents. The Bmi-1 protein has no known enzymatic activity, but serves as the key regulatory component of the PRC1 complex (polycomb repressive complex-1). This complex influences chromatin structure and regulates transcriptional activity of a number of important loci including the Ink4a locus which encodes the tumor suppressor proteins p16(Ink4a) and p14(Arf) . In this prospective study, we will discuss the implication of BMI1 in cancers, the biology of BMI1, and the regulatory control of BMI1 expression. The target validation and the future prospects of targeting BMI1 in cancer therapy are also discussed.
Asunto(s)
Proteínas Nucleares/metabolismo , Proteínas Proto-Oncogénicas/metabolismo , Proteínas Represoras/metabolismo , Animales , Humanos , Células Madre Neoplásicas/metabolismo , Células Madre Neoplásicas/patología , Proteínas Nucleares/genética , Complejo Represivo Polycomb 1 , Proteínas del Grupo Polycomb , Proteínas Proto-Oncogénicas/genética , Proteínas Represoras/genéticaRESUMEN
PTC596 is an investigational small-molecule tubulin-binding agent. Unlike other tubulin-binding agents, PTC596 is orally bioavailable and is not a P-glycoprotein substrate. So as to characterize PTC596 to position the molecule for optimal clinical development, the interactions of PTC596 with tubulin using crystallography, its spectrum of preclinical in vitro anticancer activity, and its pharmacokinetic-pharmacodynamic relationship were investigated for efficacy in multiple preclinical mouse models of leiomyosarcomas and glioblastoma. Using X-ray crystallography, it was determined that PTC596 binds to the colchicine site of tubulin with unique key interactions. PTC596 exhibited broad-spectrum anticancer activity. PTC596 showed efficacy as monotherapy and additive or synergistic efficacy in combinations in mouse models of leiomyosarcomas and glioblastoma. PTC596 demonstrated efficacy in an orthotopic model of glioblastoma under conditions where temozolomide was inactive. In a first-in-human phase I clinical trial in patients with cancer, PTC596 monotherapy drug exposures were compared with those predicted to be efficacious based on mouse models. PTC596 is currently being tested in combination with dacarbazine in a clinical trial in adults with leiomyosarcoma and in combination with radiation in a clinical trial in children with diffuse intrinsic pontine glioma.
Asunto(s)
Bencimidazoles/farmacología , Glioblastoma/tratamiento farmacológico , Leiomiosarcoma/tratamiento farmacológico , Pirazinas/farmacología , Moduladores de Tubulina/farmacología , Adulto , Anciano , Anciano de 80 o más Años , Animales , Apoptosis , Bencimidazoles/farmacocinética , Proliferación Celular , Femenino , Glioblastoma/patología , Humanos , Leiomiosarcoma/patología , Masculino , Dosis Máxima Tolerada , Ratones , Ratones Desnudos , Persona de Mediana Edad , Pronóstico , Pirazinas/farmacocinética , Distribución Tisular , Moduladores de Tubulina/farmacocinética , Células Tumorales Cultivadas , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
The coronavirus disease 2019 (COVID-19) pandemic has created an urgent need for therapeutics that inhibit the SARS-COV-2 virus and suppress the fulminant inflammation characteristic of advanced illness. Here, we describe the anti-COVID-19 potential of PTC299, an orally bioavailable compound that is a potent inhibitor of dihydroorotate dehydrogenase (DHODH), the rate-limiting enzyme of the de novo pyrimidine nucleotide biosynthesis pathway. In tissue culture, PTC299 manifests robust, dose-dependent, and DHODH-dependent inhibition of SARS-COV-2 replication (EC50 range, 2.0-31.6 nM) with a selectivity index >3,800. PTC299 also blocked replication of other RNA viruses, including Ebola virus. Consistent with known DHODH requirements for immunomodulatory cytokine production, PTC299 inhibited the production of interleukin (IL)-6, IL-17A (also called IL-17), IL-17 F, and vascular endothelial growth factor (VEGF) in tissue culture models. The combination of anti-SARS-CoV-2 activity, cytokine inhibitory activity, and previously established favorable pharmacokinetic and human safety profiles render PTC299 a promising therapeutic for COVID-19.
Asunto(s)
Antivirales/farmacología , Carbamatos/farmacología , Carbazoles/farmacología , Citocinas/antagonistas & inhibidores , Oxidorreductasas actuantes sobre Donantes de Grupo CH-CH/antagonistas & inhibidores , SARS-CoV-2/efectos de los fármacos , Replicación Viral/efectos de los fármacos , Animales , Chlorocebus aethiops , Síndrome de Liberación de Citoquinas/tratamiento farmacológico , Citocinas/inmunología , Dihidroorotato Deshidrogenasa , Células HeLa , Humanos , Inflamación/tratamiento farmacológico , Inflamación/virología , Células Vero , Tratamiento Farmacológico de COVID-19RESUMEN
The coronavirus disease 2019 (COVID-19) pandemic has created an urgent need for therapeutics that inhibit the SARS-CoV-2 virus and suppress the fulminant inflammation characteristic of advanced illness. Here, we describe the anti-COVID-19 potential of PTC299, an orally available compound that is a potent inhibitor of dihydroorotate dehydrogenase (DHODH), the rate-limiting enzyme of the de novo pyrimidine biosynthesis pathway. In tissue culture, PTC299 manifests robust, dose-dependent, and DHODH-dependent inhibition of SARS CoV-2 replication (EC 50 range, 2.0 to 31.6 nM) with a selectivity index >3,800. PTC299 also blocked replication of other RNA viruses, including Ebola virus. Consistent with known DHODH requirements for immunomodulatory cytokine production, PTC299 inhibited the production of interleukin (IL)-6, IL-17A (also called IL-17), IL-17F, and vascular endothelial growth factor (VEGF) in tissue culture models. The combination of anti-SARS-CoV-2 activity, cytokine inhibitory activity, and previously established favorable pharmacokinetic and human safety profiles render PTC299 a promising therapeutic for COVID-19.
RESUMEN
PURPOSE: Pancreatic ductal adenocarcinoma (PDA) is a deadly cancer that is broadly chemoresistant, due in part to biophysical properties of tumor stroma, which serves as a barrier to drug delivery for most classical chemotherapeutic drugs. The goal of this work is to evaluate the preclinical efficacy and mechanisms of PTC596, a novel agent with potent anticancer properties in vitro and desirable pharmacologic properties in vivo.Experimental Design: We assessed the pharmacology, mechanism, and preclinical efficacy of PTC596 in combination with standards of care, using multiple preclinical models of PDA. RESULTS: We found that PTC596 has pharmacologic properties that overcome the barrier to drug delivery in PDA, including a long circulating half-life, lack of P-glycoprotein substrate activity, and high systemic tolerability. We also found that PTC596 combined synergistically with standard clinical regimens to improve efficacy in multiple model systems, including the chemoresistant genetically engineered "KPC" model of PDA. Through mechanistic studies, we learned that PTC596 functions as a direct microtubule polymerization inhibitor, yet a prior clinical trial found that it lacks peripheral neurotoxicity, in contrast to other such agents. Strikingly, we found that PTC596 synergized with the standard clinical backbone regimen gemcitabine/nab-paclitaxel, yielding potent, durable regressions in a PDX model. Moreover, similar efficacy was achieved in combination with nab-paclitaxel alone, highlighting a specific synergistic interaction between two different microtubule-targeted agents in the setting of pancreatic ductal adenocarcinoma. CONCLUSIONS: These data demonstrate clear rationale for the development of PTC596 in combination with standard-of-care chemotherapy for PDA.
Asunto(s)
Antineoplásicos/farmacología , Carcinoma Ductal Pancreático/metabolismo , Microtúbulos/metabolismo , Neoplasias Pancreáticas/metabolismo , Multimerización de Proteína/efectos de los fármacos , Moduladores de Tubulina/farmacología , Albúminas/farmacología , Animales , Antineoplásicos/administración & dosificación , Apoptosis/efectos de los fármacos , Carcinoma Ductal Pancreático/diagnóstico , Carcinoma Ductal Pancreático/tratamiento farmacológico , Carcinoma Ductal Pancreático/mortalidad , Puntos de Control del Ciclo Celular/efectos de los fármacos , Línea Celular Tumoral , Desoxicitidina/análogos & derivados , Desoxicitidina/farmacología , Modelos Animales de Enfermedad , Relación Dosis-Respuesta a Droga , Sinergismo Farmacológico , Humanos , Inmunohistoquímica , Ratones , Microtúbulos/química , Paclitaxel/farmacología , Neoplasias Pancreáticas/diagnóstico , Neoplasias Pancreáticas/tratamiento farmacológico , Neoplasias Pancreáticas/mortalidad , Moduladores de Tubulina/administración & dosificación , Ensayos Antitumor por Modelo de Xenoinjerto , GemcitabinaRESUMEN
PTC299 was identified as an inhibitor of VEGFA mRNA translation in a phenotypic screen and evaluated in the clinic for treatment of solid tumors. To guide precision cancer treatment, we performed extensive biological characterization of the activity of PTC299 and demonstrated that inhibition of VEGF production and cell proliferation by PTC299 is linked to a decrease in uridine nucleotides by targeting dihydroorotate dehydrogenase (DHODH), a rate-limiting enzyme for de novo pyrimidine nucleotide synthesis. Unlike previously reported DHODH inhibitors that were identified using in vitro enzyme assays, PTC299 is a more potent inhibitor of DHODH in isolated mitochondria suggesting that mitochondrial membrane lipid engagement in the DHODH conformation in situ is required for its optimal activity. PTC299 has broad and potent activity against hematologic cancer cells in preclinical models, reflecting a reduced pyrimidine nucleotide salvage pathway in leukemia cells. Archived serum samples from patients treated with PTC299 demonstrated increased levels of dihydroorotate, the substrate of DHODH, indicating target engagement in patients. PTC299 has advantages over previously reported DHODH inhibitors, including greater potency, good oral bioavailability, and lack of off-target kinase inhibition and myelosuppression, and thus may be useful for the targeted treatment of hematologic malignancies.
Asunto(s)
Neoplasias Hematológicas/tratamiento farmacológico , Imidazoles/administración & dosificación , Oxidorreductasas actuantes sobre Donantes de Grupo CH-CH/antagonistas & inhibidores , Tiazoles/administración & dosificación , Factor A de Crecimiento Endotelial Vascular/genética , Animales , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Dihidroorotato Deshidrogenasa , Neoplasias Hematológicas/sangre , Neoplasias Hematológicas/enzimología , Humanos , Imidazoles/farmacología , Células K562 , Ratones , Oxidorreductasas actuantes sobre Donantes de Grupo CH-CH/sangre , Tiazoles/farmacología , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Medulloblastoma (MB) is the most frequent malignant pediatric brain tumor, representing 20% of newly diagnosed childhood central nervous system malignancies. Although advances in multimodal therapy yielded a 5-year survivorship of 80%, MB still accounts for the leading cause of childhood cancer mortality. In this work, we describe the epigenetic regulator BMI1 as a novel therapeutic target for the treatment of recurrent human Group 3 MB, a childhood brain tumor for which there is virtually no treatment option beyond palliation. Current clinical trials for recurrent MB patients based on genomic profiles of primary, treatment-naive tumors will provide limited clinical benefit since recurrent metastatic MBs are highly genetically divergent from their primary tumor. Using a small molecule inhibitor against BMI1, PTC-028, we were able to demonstrate complete ablation of self-renewal of MB stem cells in vitro. When administered to mice xenografted with patient tumors, we observed significant reduction in tumor burden in both local and metastatic compartments and subsequent increased survival, without neurotoxicity. Strikingly, serial in vivo re-transplantation assays demonstrated a marked reduction in tumor initiation ability of recurrent MB cells upon re-transplantation of PTC-028-treated cells into secondary recipient mouse brains. As Group 3 MB is often metastatic and uniformly fatal at recurrence, with no current or planned trials of targeted therapy, an efficacious targeted agent would be rapidly transitioned to clinical trials.
Asunto(s)
Neoplasias Cerebelosas/tratamiento farmacológico , Meduloblastoma/tratamiento farmacológico , Células Madre Neoplásicas/efectos de los fármacos , Complejo Represivo Polycomb 1/antagonistas & inhibidores , Bibliotecas de Moléculas Pequeñas/administración & dosificación , Animales , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Neoplasias Cerebelosas/genética , Neoplasias Cerebelosas/metabolismo , Niño , Epigénesis Genética , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Humanos , Meduloblastoma/genética , Meduloblastoma/metabolismo , Ratones , Células Madre Neoplásicas/citología , Células Madre Neoplásicas/metabolismo , Complejo Represivo Polycomb 1/genética , Bibliotecas de Moléculas Pequeñas/farmacología , Resultado del Tratamiento , Regulación hacia Arriba/efectos de los fármacos , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Despite the development of the novel Bruton tyrosine kinase inhibitor ibrutinib, mantle cell lymphoma (MCL) remains an incurable B-cell non-Hodgkin lymphoma. BMI-1 is required for the self-renewal and maintenance of MCL-initiating stem cells. Upregulation of BMI-1 has been reported in MCL patients, especially in those with refractory/relapsed disease. We studied the effects of a novel small-molecule selective inhibitor of BMI1 expression, PTC596, in MCL cells. Eight MCL cell lines and patient-derived samples were exposed to PTC596. PTC596 induced mitochondrial apoptosis, as evidenced by loss of mitochondrial membrane potential, caspase-3 cleavage, BAX activation, and phosphatidylserine externalization. There was a positive correlation between baseline BMI-1 protein levels and PTC596-induced apoptosis. p53 status did not affect sensitivity to PTC596. PTC596 effectively decreased BMI-1-expressing and tumor-initiating side population MCL cells (IC50: 138 nM) compared with ibrutinib, which modestly decreased side population cells. Interestingly, PTC596, reported to target cancer stem cells, decreased MCL-1 expression levels and antagonized ibrutinib-induced increase in MCL-1 expression, leading to synergistic apoptosis induction in MCL cells. There are currently no drugs that specifically target cancer stem cell fractions, and a reduction in BMI-1 protein by PTC596 may offer a novel therapeutic strategy for MCL.
RESUMEN
With rising incidence rates, endometrial cancer is one of the most common gynecologic malignancies in the United States. Although surgery provides significant survival benefit to early-stage patients, those with advanced or recurrent metastatic disease have a dismal prognosis. Limited treatment options include chemotherapy and radiotherapy. Hence, there is a compelling need for developing molecularly targeted therapy. Here, we show that the polycomb ring finger protein BMI1, also known as a stem cell factor, is significantly overexpressed in endometrial cancer cell lines, endometrial cancer patient tissues as well as in nonendometrioid histologies and associated with poor overall survival. PTC-028, a second-generation inhibitor of BMI1 function, decreases invasion of endometrial cancer cells and potentiates caspase-dependent apoptosis, while normal cells with minimal expression of BMI1 remain unaffected. In an aggressive uterine carcinosarcoma xenograft model, single-agent PTC-028 significantly delayed tumor growth and increased tumor doubling time compared with the standard carboplatin/paclitaxel therapy. Therefore, anti-BMI1 strategies may represent a promising targeted approach in patients with advanced or recurrent endometrial cancer, a population where treatment options are limited. Mol Cancer Ther; 17(10); 2136-43. ©2018 AACR.
Asunto(s)
Antineoplásicos/farmacología , Neoplasias Endometriales/metabolismo , Complejo Represivo Polycomb 1/antagonistas & inhibidores , Adulto , Anciano , Anciano de 80 o más Años , Animales , Apoptosis/efectos de los fármacos , Caspasas/metabolismo , Línea Celular Tumoral , Movimiento Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Modelos Animales de Enfermedad , Neoplasias Endometriales/tratamiento farmacológico , Neoplasias Endometriales/mortalidad , Neoplasias Endometriales/patología , Femenino , Humanos , Inmunohistoquímica , Persona de Mediana Edad , Clasificación del Tumor , Estadificación de Neoplasias , Complejo Represivo Polycomb 1/genética , Complejo Represivo Polycomb 1/metabolismo , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
BMI-1, also known as a stem cell factor, is frequently upregulated in several malignancies. Elevated expression of BMI-1 correlates with poor prognosis and is therefore considered a viable therapeutic target in a number of malignancies including ovarian cancer. Realizing the immense pathologic significance of BMI-1, small-molecule inhibitors against BMI-1 are recently being developed. In this study, we functionally characterize PTC-028, an orally bioavailable compound that decreases BMI-1 levels by posttranslational modification. We report that PTC-028 treatment selectively inhibits cancer cells in clonal growth and viability assays, whereas normal cells remain unaffected. Mechanistically, hyperphosphorylation-mediated depletion of cellular BMI-1 by PTC-028 coupled with a concurrent temporal decrease in ATP and a compromised mitochondrial redox balance potentiates caspase-dependent apoptosis. In vivo, orally administered PTC-028, as a single agent, exhibits significant antitumor activity comparable with the standard cisplatin/paclitaxel therapy in an orthotopic mouse model of ovarian cancer. Thus, PTC-028 has the potential to be used as an effective therapeutic agent in patients with epithelial ovarian cancer, where treatment options are limited. Mol Cancer Ther; 17(1); 39-49. ©2017 AACR.
Asunto(s)
Bencimidazoles/farmacología , Carcinoma Epitelial de Ovario/tratamiento farmacológico , Complejo Represivo Polycomb 1/antagonistas & inhibidores , Proteínas Proto-Oncogénicas/antagonistas & inhibidores , Pirazinas/farmacología , Animales , Antineoplásicos/farmacología , Carcinoma Epitelial de Ovario/metabolismo , Carcinoma Epitelial de Ovario/patología , Línea Celular Tumoral , Femenino , Humanos , Ratones , Ratones Desnudos , Complejo Represivo Polycomb 1/metabolismo , Proteínas Proto-Oncogénicas/metabolismo , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
PTC299 is a novel small molecule that specifically blocks the production of protein from selected mRNAs that under certain conditions use noncanonical ribosomal translational pathways. Hypoxia, oncogenic transformation, and viral infections limit normal translation and turn on these noncanonical translation pathways that are sensitive to PTC299. Vascular endothelial cell growth factor (VEGF) is an example of a transcript that is posttranscriptionally regulated. Single doses of PTC299 (0.03 to 3 mg/kg) were administered orally to healthy volunteers in a phase 1 single ascending-dose study. In a subsequent multiple ascending-dose study in healthy volunteers, multiple-dose regimens (0.3 to 1.2 mg/kg twice a day or 1.6 mg/kg 3 times a day for 7 days) were evaluated. PTC299 was well tolerated in these studies. As expected in healthy volunteers, mean plasma VEGF levels did not change. Increases in Cmax and AUC of PTC299 were dose-proportional. The target trough plasma concentration associated with preclinical efficacy was achieved within 7 days at doses of 0.6 mg/kg twice daily and above. These data demonstrate that PTC299 is orally bioavailable and well tolerated and support clinical evaluation of PTC299 in cancer, certain viral infections, or other diseases in which deregulation of translational control is a causal factor.
Asunto(s)
Antineoplásicos/administración & dosificación , Imidazoles/administración & dosificación , Tiazoles/administración & dosificación , Factor A de Crecimiento Endotelial Vascular/sangre , Administración Oral , Adolescente , Adulto , Antineoplásicos/efectos adversos , Antineoplásicos/farmacocinética , Área Bajo la Curva , Disponibilidad Biológica , Relación Dosis-Respuesta a Droga , Método Doble Ciego , Femenino , Humanos , Imidazoles/efectos adversos , Imidazoles/farmacocinética , Masculino , Persona de Mediana Edad , Tiazoles/efectos adversos , Tiazoles/farmacocinética , Adulto JovenRESUMEN
PURPOSE: Current prostate cancer management calls for identifying novel and more effective therapies. Self-renewing tumor-initiating cells (TICs) hold intrinsic therapy resistance and account for tumor relapse and progression. As BMI-1 regulates stem cell self-renewal, impairing BMI-1 function for TIC-tailored therapies appears to be a promising approach. EXPERIMENTAL DESIGN: We have previously developed a combined immunophenotypic and time-of-adherence assay to identify CD49bhiCD29hiCD44hi cells as human prostate TICs. We utilized this assay with patient-derived prostate cancer cells and xenograft models to characterize the effects of pharmacologic inhibitors of BMI-1. RESULTS: We demonstrate that in cell lines and patient-derived TICs, BMI-1 expression is upregulated and associated with stem cell-like traits. From a screened library, we identified a number of post-transcriptional small molecules that target BMI-1 in prostate TICs. Pharmacologic inhibition of BMI-1 in patient-derived cells significantly decreased colony formation in vitro and attenuated tumor initiation in vivo, thereby functionally diminishing the frequency of TICs, particularly in cells resistant to proliferation- and androgen receptor-directed therapies, without toxic effects on normal tissues. CONCLUSIONS: Our data offer a paradigm for targeting TICs and support the development of BMI-1-targeting therapy for a more effective prostate cancer treatment. Clin Cancer Res; 22(24); 6176-91. ©2016 AACR.
Asunto(s)
Antineoplásicos/farmacología , Autorrenovación de las Células/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Células Madre Neoplásicas/efectos de los fármacos , Complejo Represivo Polycomb 1/metabolismo , Neoplasias de la Próstata/metabolismo , Animales , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Transformación Celular Neoplásica/efectos de los fármacos , Transformación Celular Neoplásica/metabolismo , Humanos , Masculino , Ratones , Ratones Endogámicos NOD , Ratones SCID , Células Madre Neoplásicas/metabolismo , Neoplasias de la Próstata/tratamiento farmacológico , Receptores Androgénicos/metabolismo , Ensayos Antitumor por Modelo de Xenoinjerto/métodosRESUMEN
Current anti-VEGF (Vascular Endothelial Growth Factor A) therapies to treat various cancers indiscriminately block VEGF function in the patient resulting in the global loss of VEGF signaling which has been linked to dose-limiting toxicities as well as treatment failures due to acquired resistance. Accumulating evidence suggests that this resistance is at least partially due to increased production of compensatory tumor angiogenic factors/cytokines. VEGF protein production is differentially controlled depending on whether cells are in the normal "homeostatic" state or in a stressed state, such as hypoxia, by post-transcriptional regulation imparted by elements in the 5' and 3' untranslated regions (UTR) of the VEGF mRNA. Using the Gene Expression Modulation by Small molecules (GEMS™) phenotypic assay system, we performed a high throughput screen to identify low molecular weight compounds that target the VEGF mRNA UTR-mediated regulation of stress-induced VEGF production in tumor cells. We identified a number of compounds that potently and selectively reduce endogenous VEGF production under hypoxia in HeLa cells. Medicinal chemistry efforts improved the potency and pharmaceutical properties of one series of compounds resulting in the discovery of PTC-510 which inhibits hypoxia-induced VEGF expression in HeLa cells at low nanomolar concentration. In mouse xenograft studies, oral administration of PTC-510 results in marked reduction of intratumor VEGF production and single agent control of tumor growth without any evident toxicity. Here, we show that selective suppression of stress-induced VEGF production within tumor cells effectively controls tumor growth. Therefore, this approach may minimize the liabilities of current global anti-VEGF therapies.
Asunto(s)
Inhibidores de la Angiogénesis/administración & dosificación , Antineoplásicos/administración & dosificación , Ensayos Analíticos de Alto Rendimiento/métodos , Neoplasias/tratamiento farmacológico , Regiones no Traducidas/efectos de los fármacos , Factor A de Crecimiento Endotelial Vascular/genética , Administración Oral , Inhibidores de la Angiogénesis/farmacología , Animales , Antineoplásicos/farmacología , Hipoxia de la Célula , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Células HEK293 , Células HeLa , Células Hep G2 , Humanos , Ratones , Neoplasias/genética , Factor A de Crecimiento Endotelial Vascular/antagonistas & inhibidores , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Lung cancer is the most common cause of cancer deaths. The expression of the transcription factor C/EBPα (CCAAT/enhancer binding protein α) is frequently lost in non-small cell lung cancer, but the mechanisms by which C/EBPα suppresses tumor formation are not fully understood. In addition, no pharmacological therapy is available to specifically target C/EBPα expression. We discovered a subset of pulmonary adenocarcinoma patients in whom negative/low C/EBPα expression and positive expression of the oncogenic protein BMI1 (B lymphoma Mo-MLV insertion region 1 homolog) have prognostic value. We also generated a lung-specific mouse model of C/EBPα deletion that develops lung adenocarcinomas, which are prevented by Bmi1 haploinsufficiency. BMI1 activity is required for both tumor initiation and maintenance in the C/EBPα-null background, and pharmacological inhibition of BMI1 exhibits antitumor effects in both murine and human adenocarcinoma lines. Overall, we show that C/EBPα is a tumor suppressor in lung cancer and that BMI1 is required for the oncogenic process downstream of C/EBPα loss. Therefore, anti-BMI1 pharmacological inhibition may offer a therapeutic benefit for lung cancer patients with low expression of C/EBPα and high BMI1.
Asunto(s)
Adenocarcinoma/patología , Adenocarcinoma/terapia , Proteína alfa Potenciadora de Unión a CCAAT/metabolismo , Neoplasias Pulmonares/patología , Neoplasias Pulmonares/terapia , Complejo Represivo Polycomb 1/metabolismo , Proteínas Proto-Oncogénicas/metabolismo , Adenocarcinoma/genética , Adenocarcinoma/metabolismo , Adenocarcinoma del Pulmón , Animales , Proteína alfa Potenciadora de Unión a CCAAT/genética , Regulación Neoplásica de la Expresión Génica/genética , Humanos , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/metabolismo , Ratones , Ratones Noqueados , Mutación/genética , Complejo Represivo Polycomb 1/genética , Proteínas Proto-Oncogénicas/genéticaRESUMEN
Tumor recurrence following treatment remains a major clinical challenge. Evidence from xenograft models and human trials indicates selective enrichment of cancer-initiating cells (CICs) in tumors that survive therapy. Together with recent reports showing that CIC gene signatures influence patient survival, these studies predict that targeting self-renewal, the key 'stemness' property unique to CICs, may represent a new paradigm in cancer therapy. Here we demonstrate that tumor formation and, more specifically, human colorectal CIC function are dependent on the canonical self-renewal regulator BMI-1. Downregulation of BMI-1 inhibits the ability of colorectal CICs to self-renew, resulting in the abrogation of their tumorigenic potential. Treatment of primary colorectal cancer xenografts with a small-molecule BMI-1 inhibitor resulted in colorectal CIC loss with long-term and irreversible impairment of tumor growth. Targeting the BMI-1-related self-renewal machinery provides the basis for a new therapeutic approach in the treatment of colorectal cancer.