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ABSTRACT: Hemophagocytic lymphohistiocytosis (HLH) comprises a severe hyperinflammatory phenotype driven by the overproduction of cytokines, many of which signal via the JAK/STAT pathway. Indeed, the JAK1/2 inhibitor ruxolitinib has demonstrated efficacy in preclinical studies and early-phase clinical trials in HLH. Nevertheless, concerns remain for ruxolitinib-induced cytopenias, which are postulated to result from the blockade of JAK2-dependent hematopoietic growth factors. To explore the therapeutic effects of selective JAK inhibition in mouse models of HLH, we carried out studies incorporating the JAK1 inhibitor itacitinib, JAK2 inhibitor fedratinib, and JAK1/2 inhibitor ruxolitinib. All 3 drugs were well-tolerated and at the doses tested, they suppressed interferon-gamma (IFN-γ)-induced STAT1 phosphorylation in vitro and in vivo. Itacitinib, but not fedratinib, significantly improved survival and clinical scores in CpG-induced secondary HLH. Conversely, in primary HLH, in which perforin-deficient (Prf1-/-) mice are infected with lymphocytic choriomeningitis virus (LCMV), itacitinib, and fedratinib performed suboptimally. Ruxolitinib demonstrated excellent clinical efficacy in both HLH models. RNA-sequencing of splenocytes from LCMV-infected Prf1-/- mice revealed that itacitinib targeted inflammatory and metabolic pathway genes in CD8 T cells, whereas fedratinib targeted genes regulating cell proliferation and metabolism. In monocytes, neither drug conferred major transcriptional impacts. Consistent with its superior clinical effects, ruxolitinib exerted the greatest transcriptional changes in CD8 T cells and monocytes, targeting more genes across several biologic pathways, most notably JAK-dependent proinflammatory signaling. We conclude that JAK1 inhibition is sufficient to curtail CpG-induced disease, but combined inhibition of JAK1 and JAK2 is needed to best control LCMV-induced immunopathology.
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Modelos Animais de Doenças , Linfo-Histiocitose Hemofagocítica , Nitrilas , Pirazóis , Pirimidinas , Animais , Pirimidinas/farmacologia , Linfo-Histiocitose Hemofagocítica/tratamento farmacológico , Linfo-Histiocitose Hemofagocítica/induzido quimicamente , Linfo-Histiocitose Hemofagocítica/patologia , Pirazóis/farmacologia , Pirazóis/uso terapêutico , Camundongos , Janus Quinase 1/antagonistas & inibidores , Janus Quinase 1/metabolismo , Janus Quinase 1/genética , Pirróis/farmacologia , Pirróis/uso terapêutico , Janus Quinase 2/antagonistas & inibidores , Janus Quinase 2/genética , Janus Quinase 2/metabolismo , Camundongos Endogâmicos C57BL , Sulfonamidas/farmacologia , Sulfonamidas/uso terapêutico , Fator de Transcrição STAT1/metabolismo , Fator de Transcrição STAT1/genética , Inibidores de Janus Quinases/farmacologia , Inibidores de Janus Quinases/uso terapêutico , Piperidinas/farmacologia , Humanos , Benzenossulfonamidas , Hidrocarbonetos Aromáticos com Pontes , PirrolidinasRESUMO
BACKGROUND: Pediatric-type diffuse high-grade glioma (pHGG) is the most frequent malignant brain tumor in children and can be subclassified into multiple entities. Fusion genes activating the MET receptor tyrosine kinase often occur in infant-type hemispheric glioma (IHG) but also in other pHGG and are associated with devastating morbidity and mortality. METHODS: To identify new treatment options, we established and characterized two novel orthotopic mouse models harboring distinct MET fusions. These included an immunocompetent, murine allograft model and patient-derived orthotopic xenografts (PDOX) from a MET-fusion IHG patient who failed conventional therapy and targeted therapy with cabozantinib. With these models, we analyzed the efficacy and pharmacokinetic properties of three MET inhibitors, capmatinib, crizotinib and cabozantinib, alone or combined with radiotherapy. RESULTS: Capmatinib showed superior brain pharmacokinetic properties and greater in vitro and in vivo efficacy than cabozantinib or crizotinib in both models. The PDOX models recapitulated the poor efficacy of cabozantinib experienced by the patient. In contrast, capmatinib extended survival and induced long-term progression-free survival when combined with radiotherapy in two complementary mouse models. Capmatinib treatment increased radiation-induced DNA double-strand breaks and delayed their repair. CONCLUSIONS: We comprehensively investigated the combination of MET inhibition and radiotherapy as a novel treatment option for MET-driven pHGG. Our seminal preclinical data package includes pharmacokinetic characterization, recapitulation of clinical outcomes, coinciding results from multiple complementing in vivo studies, and insights into molecular mechanism underlying increased efficacy. Taken together, we demonstrate the groundbreaking efficacy of capmatinib and radiation as a highly promising concept for future clinical trials.
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Neoplasias Encefálicas , Glioma , Proteínas Proto-Oncogênicas c-met , Ensaios Antitumorais Modelo de Xenoenxerto , Animais , Humanos , Glioma/patologia , Glioma/tratamento farmacológico , Glioma/genética , Glioma/terapia , Proteínas Proto-Oncogênicas c-met/antagonistas & inibidores , Proteínas Proto-Oncogênicas c-met/genética , Proteínas Proto-Oncogênicas c-met/metabolismo , Camundongos , Neoplasias Encefálicas/patologia , Neoplasias Encefálicas/tratamento farmacológico , Neoplasias Encefálicas/genética , Neoplasias Encefálicas/radioterapia , Benzamidas/farmacologia , Benzamidas/uso terapêutico , Linhagem Celular Tumoral , Proteínas de Fusão Oncogênica/genética , Proteínas de Fusão Oncogênica/metabolismo , Feminino , Inibidores de Proteínas Quinases/farmacologia , Inibidores de Proteínas Quinases/uso terapêutico , Piridinas/farmacologia , Piridinas/uso terapêutico , Crizotinibe/farmacologia , Crizotinibe/uso terapêutico , Modelos Animais de Doenças , Criança , Gradação de Tumores , Anilidas/farmacologia , Imidazóis , TriazinasRESUMO
Acute erythroid leukemia (AEL) is characterized by a distinct morphology, mutational spectrum, lack of preclinical models, and poor prognosis. Here, using multiplexed genome editing of mouse hematopoietic stem and progenitor cells and transplant assays, we developed preclinical models of AEL and non-erythroid acute leukemia and describe the central role of mutational cooperativity in determining leukemia lineage. Different combination of mutations in Trp53, Bcor, Dnmt3a, Rb1, and Nfix resulted in the development of leukemia with an erythroid phenotype, accompanied by the acquisition of alterations in signaling and transcription factor genes that recapitulate human AEL by cross-species genomic analysis. Clonal expansion during tumor evolution was driven by mutational cooccurrence, with clones harboring a higher number of founder and secondary lesions (eg, mutations in signaling genes) showing greater evolutionary fitness. Mouse and human AEL exhibited deregulation of genes regulating erythroid development, notably Gata1, Klf1, and Nfe2, driven by the interaction of mutations of the epigenetic modifiers Dnmt3a and Tet2 that perturbed methylation and thus expression of lineage-specific transcription factors. The established mouse leukemias were used as a platform for drug screening. Drug sensitivity was associated with the leukemia genotype, with the poly (ADP-ribose) polymerase inhibitor talazoparib and the demethylating agent decitabine efficacious in Trp53/Bcor-mutant AEL, CDK7/9 inhibitors in Trp53/Bcor/Dnmt3a-mutant AEL, and gemcitabine and bromodomain inhibitors in NUP98-KDM5A leukemia. In conclusion, combinatorial genome editing has shown the interplay of founding and secondary genetic alterations in phenotype and clonal evolution, epigenetic regulation of lineage-specific transcription factors, and therapeutic tractability in erythroid leukemogenesis.
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Edição de Genes , Leucemia Eritroblástica Aguda/genética , Animais , Sistemas CRISPR-Cas , Evolução Clonal , Epigênese Genética , Hematopoese , Humanos , Camundongos , Mutação , TranscriptomaRESUMO
Paediatric solid tumours arise from endodermal, ectodermal, or mesodermal lineages. Although the overall survival of children with solid tumours is 75%, that of children with recurrent disease is below 30%. To capture the complexity and diversity of paediatric solid tumours and establish new models of recurrent disease, here we develop a protocol to produce orthotopic patient-derived xenografts at diagnosis, recurrence, and autopsy. Tumour specimens were received from 168 patients, and 67 orthotopic patient-derived xenografts were established for 12 types of cancer. The origins of the patient-derived xenograft tumours were reflected in their gene-expression profiles and epigenomes. Genomic profiling of the tumours, including detailed clonal analysis, was performed to determine whether the clonal population in the xenograft recapitulated the patient's tumour. We identified several drug vulnerabilities and showed that the combination of a WEE1 inhibitor (AZD1775), irinotecan, and vincristine can lead to complete response in multiple rhabdomyosarcoma orthotopic patient-derived xenografts tumours in vivo.
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Neoplasias/tratamento farmacológico , Ensaios Antitumorais Modelo de Xenoenxerto/métodos , Animais , Bortezomib/farmacologia , Bortezomib/uso terapêutico , Camptotecina/análogos & derivados , Camptotecina/farmacologia , Camptotecina/uso terapêutico , Proteínas de Ciclo Celular/antagonistas & inibidores , Criança , Células Clonais , Quimioterapia Combinada , Epigênese Genética , Feminino , Xenoenxertos/efeitos dos fármacos , Xenoenxertos/metabolismo , Xenoenxertos/patologia , Xenoenxertos/transplante , Ensaios de Triagem em Larga Escala/métodos , Humanos , Ácidos Hidroxâmicos/farmacologia , Ácidos Hidroxâmicos/uso terapêutico , Indóis/farmacologia , Indóis/uso terapêutico , Irinotecano , Camundongos , Neoplasias/genética , Proteínas Nucleares/antagonistas & inibidores , Panobinostat , Proteínas Tirosina Quinases/antagonistas & inibidores , Pirazóis/farmacologia , Pirazóis/uso terapêutico , Pirimidinas/farmacologia , Pirimidinas/uso terapêutico , Pirimidinonas , Rabdomiossarcoma/tratamento farmacológico , Rabdomiossarcoma/genética , Vincristina/farmacologia , Vincristina/uso terapêuticoRESUMO
Clinical efficacy of the influenza antiviral baloxavir marboxil (baloxavir) is compromised by treatment-emergent variants harboring a polymerase acidic protein I38T (isoleucine-38-threonine) substitution. However, the fitness of I38T-containing influenza B viruses (IBVs) remains inadequately defined. After the pharmacokinetics of the compound were confirmed in ferrets, animals were injected subcutaneously with 8 mg/kg of baloxavir acid (BXA) at 24 h postinoculation with recombinant BXA-sensitive (BXA-Sen, I38) or BXA-resistant (BXA-Res, I38T) B/Brisbane/60/2008 (Victoria lineage) virus. BXA treatment of donor ferrets reduced virus replication and delayed transmission of the BXA-Sen but not the BXA-Res IBV. The I38 genotype remained dominant in the BXA-Sen-infected animals, even with BXA treatment. In competitive-mixture experiments, no transmission to aerosol contacts was seen from BXA-treated donors coinfected with the BXA-Sen and BXA-Res B/Brisbane/60/2008 viruses. However, in parallel mixed infections with the B/Phuket/3073/2013 (Yamagata lineage) virus background, BXA treatment failed to block airborne transmission of the BXA-Res virus, and the I38T genotype generally predominated. Therefore, the relative fitness of BXA-Res IBVs is complex and dependent on the virus backbone and within-host virus competition. BXA treatment of single-virus-infected ferrets hampers aerosol transmission of the BXA-Sen virus and does not readily generate BXA-Res variants, whereas mixed infections may result in propagation of BXA-Res IBVs of the Yamagata lineage. Our findings confirm the antiviral potency of baloxavir against IBVs, while supporting optimization of the dosing regimen to maximize clinical benefit.
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Influenza Humana , Preparações Farmacêuticas , Animais , Antivirais/farmacologia , Antivirais/uso terapêutico , Dibenzotiepinas , Farmacorresistência Viral/genética , Furões , Humanos , Vírus da Influenza B/genética , Influenza Humana/tratamento farmacológico , Morfolinas , Piridonas/uso terapêutico , Tempo para o Tratamento , Triazinas/uso terapêuticoRESUMO
BACKGROUND: The ongoing global malaria eradication campaign requires development of potent, safe, and cost-effective drugs lacking cross-resistance with existing chemotherapies. One critical step in drug development is selecting a suitable clinical candidate from late leads. The process used to select the clinical candidate SJ733 from two potent dihydroisoquinolone (DHIQ) late leads, SJ733 and SJ311, based on their physicochemical, pharmacokinetic (PK), and toxicity profiles is described. METHODS: The compounds were tested to define their physicochemical properties including kinetic and thermodynamic solubility, partition coefficient, permeability, ionization constant, and binding to plasma proteins. Metabolic stability was assessed in both microsomes and hepatocytes derived from mice, rats, dogs, and humans. Cytochrome P450 inhibition was assessed using recombinant human cytochrome enzymes. The pharmacokinetic profiles of single intravenous or oral doses were investigated in mice, rats, and dogs. RESULTS: Although both compounds displayed similar physicochemical properties, SJ733 was more permeable but metabolically less stable than SJ311 in vitro. Single dose PK studies of SJ733 in mice, rats, and dogs demonstrated appreciable oral bioavailability (60-100%), whereas SJ311 had lower oral bioavailability (mice 23%, rats 40%) and higher renal clearance (10-30 fold higher than SJ733 in rats and dogs), suggesting less favorable exposure in humans. SJ311 also displayed a narrower range of dose-proportional exposure, with plasma exposure flattening at doses above 200 mg/kg. CONCLUSION: SJ733 was chosen as the candidate based on a more favorable dose proportionality of exposure and stronger expectation of the ability to justify a strong therapeutic index to regulators.
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Antimaláricos/farmacologia , Isoquinolinas/farmacologia , Animais , Antimaláricos/farmacocinética , Antimaláricos/toxicidade , Disponibilidade Biológica , Cães , Hepatócitos/efeitos dos fármacos , Compostos Heterocíclicos de 4 ou mais Anéis/farmacocinética , Compostos Heterocíclicos de 4 ou mais Anéis/farmacologia , Compostos Heterocíclicos de 4 ou mais Anéis/toxicidade , Humanos , Isoquinolinas/farmacocinética , Isoquinolinas/toxicidade , Camundongos , Microssomos Hepáticos/efeitos dos fármacos , RatosRESUMO
3-Hydroxy-3-methylglutaryl coenzyme A reductase inhibitors were potent hits against a mouse ependymoma cell line, but their effectiveness against central nervous system tumors will depend on their ability to cross the blood-brain barrier and attain a sufficient exposure at the tumor. Among 3-hydroxy-3-methylglutaryl coenzyme A inhibitors that had activity in vitro, we prioritized simvastatin (SV) as the lead compound for preclinical pharmacokinetic studies based on its potential for central nervous system penetration as determined from in silico models. Furthermore, we performed systemic plasma disposition and cerebral microdialysis studies of SV (100 mg/kg, p.o.) in a murine model of ependymoma to characterize plasma and tumor extracellular fluid (tECF) pharmacokinetic properties. The murine dosage of SV (100 mg/kg, p.o.) was equivalent to the maximum tolerated dose in patients (7.5 mg/kg, p.o.) based on equivalent plasma exposure of simvastatin acid (SVA) between the two species. SV is rapidly metabolized in murine plasma with 15 times lower exposure compared with human plasma. SVA exposure in tECF was <33.8 ± 11.9 µg/l per hour, whereas the tumor to plasma partition coefficient of SVA was <0.084 ± 0.008. Compared with in vitro washout IC50 values, we did not achieve sufficient exposure of SVA in tECF to suggest tumor growth inhibition; therefore, SV was not carried forward in subsequent preclinical efficacy studies.
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Neoplasias do Sistema Nervoso Central/metabolismo , Citotoxinas/administração & dosagem , Citotoxinas/metabolismo , Microdiálise/métodos , Sinvastatina/análogos & derivados , Animais , Barreira Hematoencefálica/efeitos dos fármacos , Barreira Hematoencefálica/metabolismo , Linhagem Celular Tumoral , Neoplasias do Sistema Nervoso Central/tratamento farmacológico , Córtex Cerebral/efeitos dos fármacos , Córtex Cerebral/metabolismo , Avaliação Pré-Clínica de Medicamentos/métodos , Camundongos , Camundongos Nus , Sinvastatina/administração & dosagem , Sinvastatina/metabolismoRESUMO
Chemotherapies active in preclinical studies frequently fail in the clinic due to lack of efficacy, which limits progress for rare cancers since only small numbers of patients are available for clinical trials. Thus, a preclinical drug development pipeline was developed to prioritize potentially active regimens for pediatric brain tumors spanning from in vitro drug screening, through intracranial and intra-tumoral pharmacokinetics to in vivo efficacy studies. Here, as an example of the pipeline, data are presented for the combination of 5-fluoro-2'-deoxycytidine and tetrahydrouridine in three pediatric brain tumor models. The in vitro activity of nine novel therapies was tested against tumor spheres derived from faithful mouse models of Group 3 medulloblastoma, ependymoma, and choroid plexus carcinoma. Agents with the greatest in vitro potency were then subjected to a comprehensive series of in vivo pharmacokinetic (PK) and pharmacodynamic (PD) studies culminating in preclinical efficacy trials in mice harboring brain tumors. The nucleoside analog 5-fluoro-2'-deoxycytidine (FdCyd) markedly reduced the proliferation in vitro of all three brain tumor cell types at nanomolar concentrations. Detailed intracranial PK studies confirmed that systemically administered FdCyd exceeded concentrations in brain tumors necessary to inhibit tumor cell proliferation, but no tumor displayed a significant in vivo therapeutic response. Despite promising in vitro activity and in vivo PK properties, FdCyd is unlikely to be an effective treatment of pediatric brain tumors, and therefore was deprioritized for the clinic. Our comprehensive and integrated preclinical drug development pipeline should reduce the attrition of drugs in clinical trials.
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Antineoplásicos/administração & dosagem , Neoplasias Encefálicas/tratamento farmacológico , Encéfalo/efeitos dos fármacos , Desoxicitidina/análogos & derivados , Modelos Animais de Doenças , Avaliação Pré-Clínica de Medicamentos/métodos , Tetra-Hidrouridina/administração & dosagem , Animais , Antineoplásicos/sangue , Antineoplásicos/farmacocinética , Antineoplásicos/uso terapêutico , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Desoxicitidina/administração & dosagem , Desoxicitidina/sangue , Desoxicitidina/farmacocinética , Desoxicitidina/uso terapêutico , Relação Dose-Resposta a Droga , Epigênese Genética/efeitos dos fármacos , Camundongos , Camundongos Nus , Tetra-Hidrouridina/sangue , Tetra-Hidrouridina/farmacocinética , Tetra-Hidrouridina/uso terapêuticoRESUMO
PURPOSE: Retinoblastoma is a childhood cancer of the retina. Clinical trials have shown that local delivery of broad spectrum chemotherapeutic agents is efficacious. Recent studies characterizing the genomic and epigenomic landscape of retinoblastoma identified spleen tyrosine kinase (SYK) as a promising candidate for targeted therapy. The purpose of this study was to conduct preclinical testing of the SYK antagonist R406 to evaluate it as a candidate for retinoblastoma treatment. METHODS: The efficacy of the SYK antagonist R406 delivered locally in a human orthotopic xenograft mouse model of retinoblastoma was tested. Intraocular exposure of R406 was determined for various routes and formulations. RESULTS: There was no evidence of efficacy for subconjunctival. R406. Maximal vitreal concentration was 10-fold lower than the minimal concentration required to kill retinoblastoma cells in vitro. Dosage of R406 subconjunctivally from emulsion or suspension formulations, direct intravitreal injection of the soluble prodrug of R406 (R788), and repeated topical administration of R406 all increased vitreal exposure, but failed to reach the exposure required for retinoblastoma cell death in culture. CONCLUSION: Taken together, these data suggest that R406 is not a viable clinical candidate for the treatment of retinoblastoma. This study highlights the importance of pharmacokinetic testing of molecular targeted retinoblastoma therapeutics.
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Peptídeos e Proteínas de Sinalização Intracelular/antagonistas & inibidores , Oxazinas/farmacologia , Oxazinas/farmacocinética , Inibidores de Proteínas Quinases/farmacologia , Inibidores de Proteínas Quinases/farmacocinética , Proteínas Tirosina Quinases/antagonistas & inibidores , Piridinas/farmacologia , Piridinas/farmacocinética , Neoplasias da Retina/tratamento farmacológico , Retinoblastoma/tratamento farmacológico , Animais , Antineoplásicos/farmacocinética , Antineoplásicos/farmacologia , Morte Celular/efeitos dos fármacos , Modelos Animais de Doenças , Olho/efeitos dos fármacos , Feminino , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Quinase SykRESUMO
The cellular plasticity of neuroblastoma is defined by a mixture of two major cell states, adrenergic (ADRN) and mesenchymal (MES), which may contribute to therapy resistance. However, how neuroblastoma cells switch cellular states during therapy remains largely unknown and how to eradicate neuroblastoma regardless of their cell states is a clinical challenge. To better understand the lineage switch of neuroblastoma in chemoresistance, we comprehensively defined the transcriptomic and epigenetic map of ADRN and MES types of neuroblastomas using human and murine models treated with indisulam, a selective RBM39 degrader. We showed that cancer cells not only undergo a bidirectional switch between ADRN and MES states, but also acquire additional cellular states, reminiscent of the developmental pliancy of neural crest cells. The lineage alterations are coupled with epigenetic reprogramming and dependency switch of lineage-specific transcription factors, epigenetic modifiers and targetable kinases. Through targeting RNA splicing, indisulam induces an inflammatory tumor microenvironment and enhances anticancer activity of natural killer cells. The combination of indisulam with anti-GD2 immunotherapy results in a durable, complete response in high-risk transgenic neuroblastoma models, providing an innovative, rational therapeutic approach to eradicate tumor cells regardless of their potential to switch cell states.
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Retinoic acid (RA) is a standard-of-care neuroblastoma drug thought to be effective by inducing differentiation. Curiously, RA has little effect on primary human tumors during upfront treatment but can eliminate neuroblastoma cells from the bone marrow during post-chemo consolidation therapy-a discrepancy that has never been explained. To investigate this, we treated a large cohort of neuroblastoma cell lines with RA and observed that the most RA-sensitive cells predominantly undergo apoptosis or senescence, rather than differentiation. We conducted genome-wide CRISPR knockout screens under RA treatment, which identified BMP signaling as controlling the apoptosis/senescence vs differentiation cell fate decision and determining RA's overall potency. We then discovered that BMP signaling activity is markedly higher in neuroblastoma patient samples at bone marrow metastatic sites, providing a plausible explanation for RA's ability to clear neuroblastoma cells specifically from the bone marrow, seemingly mimicking interactions between BMP and RA during normal development.
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Chemical discovery efforts commonly target individual protein domains. Many proteins, including the EP300/CBP histone acetyltransferases (HATs), contain several targetable domains. EP300/CBP are critical gene-regulatory targets in cancer, with existing high potency inhibitors of either the catalytic HAT domain or protein-binding bromodomain (BRD). A domain-specific inhibitory approach to multidomain-containing proteins may identify exceptional-responding tumor types, thereby expanding a therapeutic index. Here, we discover that targeting EP300/CBP using the domain-specific inhibitors, A485 (HAT) or CCS1477 (BRD) have different effects in select tumor types. Group 3 medulloblastoma (G3MB) cells are especially sensitive to BRD, compared with HAT inhibition. Structurally, these effects are mediated by the difluorophenyl group in the catalytic core of CCS1477. Mechanistically, bromodomain inhibition causes rapid disruption of genetic dependency networks that are required for G3MB growth. These studies provide a domain-specific structural foundation for drug discovery efforts targeting EP300/CBP and identify a selective role for the EP300/CBP bromodomain in maintaining genetic dependency networks in G3MB.
Assuntos
Proteína p300 Associada a E1A , Redes Reguladoras de Genes , Meduloblastoma , Humanos , Meduloblastoma/genética , Meduloblastoma/tratamento farmacológico , Meduloblastoma/metabolismo , Meduloblastoma/patologia , Proteína p300 Associada a E1A/metabolismo , Proteína p300 Associada a E1A/genética , Proteína p300 Associada a E1A/antagonistas & inibidores , Linhagem Celular Tumoral , Redes Reguladoras de Genes/efeitos dos fármacos , Animais , Domínios Proteicos , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Camundongos , Neoplasias Cerebelares/genética , Neoplasias Cerebelares/tratamento farmacológico , Neoplasias Cerebelares/metabolismo , Neoplasias Cerebelares/patologia , Antineoplásicos/farmacologiaRESUMO
Acute lymphoblastic leukemia expressing the gamma delta T cell receptor (yo T-ALL) is a poorly understood disease. We studied 200 children with yo T-ALL from 13 clinical study groups to understand the clinical and genetic features of this disease. We found age and genetic drivers were significantly associated with outcome. yo T-ALL diagnosed in children under three years of age was extremely high-risk and enriched for genetic alterations that result in both LMO2 activation and STAG2 inactivation. Mechanistically, using patient samples and isogenic cell lines, we show that inactivation of STAG2 profoundly perturbs chromatin organization by altering enhancer-promoter looping, resulting in deregulation of gene expression associated with T-cell differentiation. High throughput drug screening identified a vulnerability in DNA repair pathways arising from STAG2 inactivation, which can be targeted by Poly(ADP-ribose) polymerase (PARP) inhibition. These data provide a diagnostic framework for classification and risk stratification of pediatric yo T-ALL.
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Despite improvement in the treatment of medulloblastoma over the last years, numerous patients with MYC- and MYCN-driven tumors still fail current therapies. Medulloblastomas have an intact retinoblastoma protein RB, suggesting that CDK4/6 inhibition might represent a therapeutic strategy for which drug combination remains understudied. We conducted high-throughput drug combination screens in a Group3 (G3) medulloblastoma line using the CDK4/6 inhibitor (CDK4/6i) ribociclib at IC20, referred to as an anchor, and 87 oncology drugs approved by FDA or in clinical trials. Bromodomain and extra terminal (BET) and PI3K/mTOR inhibitors potentiated ribociclib inhibition of proliferation in an established cell line and freshly dissociated tumor cells from intracranial xenografts of G3 and Sonic hedgehog (SHH) medulloblastomas in vitro. A reverse combination screen using the BET inhibitor JQ1 as anchor, revealed CDK4/6i as the most potentiating drugs. In vivo, ribociclib showed single-agent activity in medulloblastoma models whereas JQ1 failed to show efficacy due to high clearance and insufficient free brain concentration. Despite in vitro synergy, combination of ribociclib with the PI3K/mTOR inhibitor paxalisib did not significantly improve the survival of G3 and SHH medulloblastoma-bearing mice compared with ribociclib alone. Molecular analysis of ribociclib and paxalisib-treated tumors revealed that E2F targets and PI3K/AKT/MTORC1 signaling genes were depleted, as expected. Importantly, in one untreated G3MB model HD-MB03, the PI3K/AKT/MTORC1 gene set was enriched in vitro compared with in vivo suggesting that the pathway displayed increased activity in vitro. Our data illustrate the difficulty in translating in vitro findings in vivo. See related article in Mol Cancer Ther (2022) 21(8):1306-1317.
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Neoplasias Cerebelares , Meduloblastoma , Animais , Humanos , Camundongos , Neoplasias Cerebelares/tratamento farmacológico , Gencitabina , Proteínas Hedgehog , Alvo Mecanístico do Complexo 1 de Rapamicina , Meduloblastoma/genética , Inibidores de MTOR , Fosfatidilinositol 3-Quinases/uso terapêutico , Inibidores de Proteínas Quinases/farmacologia , Inibidores de Proteínas Quinases/uso terapêutico , Proteínas Proto-Oncogênicas c-akt , Serina-Treonina Quinases TOR/uso terapêuticoRESUMO
Obesity has been epidemiologically and empirically linked with more severe diseases upon influenza infection. To ameliorate severe disease, treatment with antivirals, such as the neuraminidase inhibitor oseltamivir, is suggested to begin within days of infection especially in high-risk hosts. However, this treatment can be poorly effective and may generate resistance variants within the treated host. Here, we hypothesized that obesity would reduce oseltamivir treatment effectiveness in the genetically obese mouse model. We demonstrated that oseltamivir treatment does not improve viral clearance in obese mice. While no traditional variants associated with oseltamivir resistance emerged, we did note that drug treatment failed to quench the viral population and did lead to phenotypic drug resistance in vitro. Together, these studies suggest that the unique pathogenesis and immune responses in obese mice could have implications for pharmaceutical interventions and the within-host dynamics of the influenza virus population. IMPORTANCE Influenza virus infections, while typically resolving within days to weeks, can turn critical, especially in high-risk populations. Prompt antiviral administration is crucial to mitigating these severe sequalae, yet concerns remain if antiviral treatment is effective in hosts with obesity. Here, we show that oseltamivir does not improve viral clearance in genetically obese or type I interferon receptor-deficient mice. This suggests a blunted immune response may impair oseltamivir efficacy and render a host more susceptible to severe disease. This study furthers our understanding of oseltamivir treatment dynamics both systemically and in the lungs of obese mice, as well as the consequences of oseltamivir treatment for the within-host emergence of drug-resistant variants.
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Vírus da Influenza A Subtipo H1N1 , Influenza Humana , Camundongos , Animais , Humanos , Oseltamivir/uso terapêutico , Camundongos Obesos , Influenza Humana/tratamento farmacológico , Antivirais/uso terapêutico , Antivirais/farmacologia , Neuraminidase , Farmacorresistência ViralRESUMO
BACKGROUND: Pediatric high-grade glioma (pHGG) is largely incurable and accounts for most brain tumor-related deaths in children. Radiation is a standard therapy, yet the benefit from this treatment modality is transient, and most children succumb to disease within 2 years. Recent large-scale genomic studies suggest that pHGG has alterations in DNA damage response (DDR) pathways that induce resistance to DNA damaging agents. The aim of this study was to evaluate the therapeutic potential and molecular consequences of combining radiation with selective DDR inhibition in pHGG. METHODS: We conducted an unbiased screen in pHGG cells that combined radiation with clinical candidates targeting the DDR and identified the ATM inhibitor AZD1390. Subsequently, we profiled AZD1390 + radiation in an extensive panel of early passage pHGG cell lines, mechanistically characterized response to the combination in vitro in sensitive and resistant cells and evaluated the combination in vivo using TP53 wild-type and TP53 mutant orthotopic xenografts. RESULTS: AZD1390 significantly potentiated radiation across molecular subgroups of pHGG by increasing mutagenic nonhomologous end joining and augmenting genomic instability. In contrast to previous reports, ATM inhibition significantly improved the efficacy of radiation in both TP53 wild-type and TP53 mutant isogenic cell lines and distinct orthotopic xenograft models. Furthermore, we identified a novel mechanism of resistance to AZD1390 + radiation that was marked by an attenuated ATM pathway response which dampened sensitivity to ATM inhibition and induced synthetic lethality with ATR inhibition. CONCLUSIONS: Our study supports the clinical evaluation of AZD1390 in combination with radiation in pediatric patients with HGG.
Assuntos
Neoplasias Encefálicas , Glioma , Humanos , Criança , Glioma/tratamento farmacológico , Glioma/genética , Glioma/radioterapia , Neoplasias Encefálicas/tratamento farmacológico , Neoplasias Encefálicas/genética , Neoplasias Encefálicas/radioterapia , Inibidores de Proteínas Quinases/farmacologia , Inibidores de Proteínas Quinases/uso terapêutico , Dano ao DNA , Proteínas Mutadas de Ataxia Telangiectasia/genética , Proteínas Mutadas de Ataxia Telangiectasia/metabolismoRESUMO
PURPOSE: Gamma delta T-cell receptor-positive acute lymphoblastic leukemia (γδ T-ALL) is a high-risk but poorly characterized disease. METHODS: We studied clinical features of 200 pediatric γδ T-ALL, and compared the prognosis of 93 cases to 1,067 protocol-matched non-γδ T-ALL. Genomic features were defined by transcriptome and genome sequencing. Experimental modeling was used to examine the mechanistic impacts of genomic alterations. Therapeutic vulnerabilities were identified by high throughput drug screening of cell lines and xenografts. RESULTS: γδ T-ALL in children under three was extremely high-risk with 5-year event-free survival (33% v. 70% [age 3-<10] and 73% [age ≥10], P =9.5 x 10 -5 ) and 5-year overall survival (49% v. 78% [age 3-<10] and 81% [age ≥10], P =0.002), differences not observed in non-γδ T-ALL. γδ T-ALL in this age group was enriched for genomic alterations activating LMO2 activation and inactivating STAG2 inactivation ( STAG2/LMO2 ). Mechanistically, we show that inactivation of STAG2 profoundly perturbs chromatin organization by altering enhancer-promoter looping resulting in deregulation of gene expression associated with T-cell differentiation. Drug screening showed resistance to prednisolone, consistent with clinical slow treatment response, but identified a vulnerability in DNA repair pathways arising from STAG2 inactivation, which was efficaciously targeted by Poly(ADP-ribose) polymerase (PARP) inhibition, with synergism with HDAC inhibitors. Ex-vivo drug screening on PDX cells validated the efficacy of PARP inhibitors as well as other potential targets including nelarabine. CONCLUSION: γδ T-ALL in children under the age of three is extremely high-risk and enriched for STAG2/LMO2 ALL. STAG2 loss perturbs chromatin conformation and differentiation, and STAG2/LMO2 ALL is sensitive to PARP inhibition. These data provide a diagnostic and therapeutic framework for pediatric γδ T-ALL. SUPPORT: The authors are supported by the American and Lebanese Syrian Associated Charities of St Jude Children's Research Hospital, NCI grants R35 CA197695, P50 CA021765 (C.G.M.), the Henry Schueler 41&9 Foundation (C.G.M.), and a St. Baldrick's Foundation Robert J. Arceci Innovation Award (C.G.M.), Gabriella Miller Kids First X01HD100702 (D.T.T and C.G.M.) and R03CA256550 (D.T.T. and C.G.M.), F32 5F32CA254140 (L.M.), and a Garwood Postdoctoral Fellowship of the Hematological Malignancies Program of the St Jude Children's Research Hospital Comprehensive Cancer Center (S.K.). This project was supported by the National Cancer Institute of the National Institutes of Health under the following award numbers: U10CA180820, UG1CA189859, U24CA114766, U10CA180899, U10CA180866 and U24CA196173. DISCLAIMER: The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health. The funding agencies were not directly involved in the design of the study, gathering, analysis and interpretation of the data, writing of the manuscript, or decision to submit the manuscript for publication.
RESUMO
BACKGROUND: SJ733, a newly developed inhibitor of P. falciparum ATP4, has a favorable safety profile and rapid antiparasitic effect but insufficient duration to deliver a single-dose cure of malaria. We investigated the safety, tolerability, and pharmacokinetics of a multidose SJ733 regimen and a single-dose pharmacoboost approach using cobicistat to inhibit CYP3A4, thereby increasing exposure. METHODS: Two multidose unboosted cohorts (nâ¯=â¯9) (SJ733, 300â¯mg and 600â¯mg daily for 3 days) followed by three single-dose boosted cohorts combining SJ733 (nâ¯=â¯18) (75-, 300-, or 600-mg single dose) with cobicistat (150-mg single dose) as a pharmacokinetic booster were evaluated in healthy volunteers (ClinicalTrials.gov: NCT02661373). FINDINGS: All participants tolerated SJ733 well, with no serious adverse events (AEs), dose-limiting toxicity, or clinically significant electrocardiogram or laboratory test findings. All reported AEs were Grade 1, clinically insignificant, and considered unlikely or unrelated to SJ733. Compared to unboosted cohorts, the SJ733/cobicistat-boosted cohorts showed a median increase in area under the curve and maximum concentration of 3·9â¯×â¯and 2·6 ×, respectively, and a median decrease in the ratio of the major CYP3A-produced metabolite SJ506 to parent drug of 4·6â¯×â¯. Incorporating these data in a model of parasite dynamics indicated that a 3-day regimen of SJ733/cobicistat (600â¯mg/150â¯mg daily) relative to a single 600-mg dose ± cobicistat would increase parasite clearance from 106 to 1012 parasites/µL. INTERPRETATION: The multidose and pharmacoboosted approaches to delivering SJ733 were well-tolerated and significantly increased drug exposure and prediction of cure. This study supports the further development of SJ733 and demonstrates an innovative pharmacoboost approach for an antimalarial. FUNDING: Global Health Innovative Technology Fund, Medicines for Malaria Venture, National Institutes of Health, and American Lebanese Syrian Associated Charities.
Assuntos
Antimaláricos , Antagonistas do Ácido Fólico , Malária Falciparum , Malária , Antimaláricos/efeitos adversos , Cobicistat/uso terapêutico , Compostos Heterocíclicos de 4 ou mais Anéis , Humanos , Isoquinolinas , Malária/tratamento farmacológico , Malária Falciparum/tratamento farmacológico , Malária Falciparum/parasitologia , Plasmodium falciparumRESUMO
Vitamin D3 (VD3) induces intestinal CYP3A that metabolizes orally administered anti-leukemic chemotherapeutic substrates dexamethasone (DEX) and dasatinib potentially causing a vitamin-drug interaction. To determine the impact of VD3 status on systemic exposure and efficacy of these chemotherapeutic agents, we used VD3 sufficient and deficient mice and performed pharmacokinetic and anti-leukemic efficacy studies. Female C57BL/6J and hCYP3A4 transgenic VD3 deficient mice had significantly lower duodenal (but not hepatic) mouse Cyp3a11 and hCYP3A4 expression compared to VD3 sufficient mice, while duodenal expression of Mdr1a, Bcrp and Mrp4 were significantly higher in deficient mice. When the effect of VD3 status on DEX systemic exposure was compared following a discontinuous oral DEX regimen, similar to that used to treat pediatric acute lymphoblastic leukemia patients, male VD3 deficient mice had significantly higher mean plasma DEX levels (31.7 nM) compared to sufficient mice (12.43 nM) at days 3.5 but not at any later timepoints. Following a single oral gavage of DEX, there was a statistically, but not practically, significant decrease in DEX systemic exposure in VD3 deficient vs. sufficient mice. While VD3 status had no effect on oral dasatinib's area under the plasma drug concentration-time curve, VD3 deficient male mice had significantly higher dasatinib plasma levels at t = 0.25 hr. Dexamethasone was unable to reverse the poorer survival of VD3 sufficient vs. deficient mice to BCR-ABL leukemia. In conclusion, although VD3 levels significantly altered intestinal mouse Cyp3a in female mice, DEX plasma exposure was only transiently different for orally administered DEX and dasatinib in male mice. Likewise, the small effect size of VD3 deficiency on single oral dose DEX clearance suggests that the clinical significance of VD3 levels on DEX systemic exposure are likely to be limited.
Assuntos
Dasatinibe , Vitamina D , Animais , Feminino , Masculino , CamundongosRESUMO
Vincristine (VCR) is one of the most widely prescribed medications for treating solid tumors and acute lymphoblastic leukemia (ALL) in children and adults. However, its major dose-limiting toxicity is peripheral neuropathy that can disrupt curative therapy. Peripheral neuropathy can also persist into adulthood, compromising quality of life of childhood cancer survivors. Reducing VCR-induced neurotoxicity without compromising its anticancer effects would be ideal. Here, we show that low expression of NHP2L1 is associated with increased sensitivity of primary leukemia cells to VCR, and that concomitant administration of VCR with inhibitors of NHP2L1 increases VCR cytotoxicity in leukemia cells, prolongs survival of ALL xenograft mice, but decreases VCR effects on human-induced pluripotent stem cell-derived neurons and mitigates neurotoxicity in mice. These findings offer a strategy for increasing VCR's antileukemic effects while reducing peripheral neuropathy in patients treated with this widely prescribed medication.