RESUMO
As lead optimization efforts have successfully reduced metabolic liabilities due to cytochrome P450 (CYP)-mediated metabolism, there has been an increase in the frequency of involvement of non-CYP enzymes in the metabolism of investigational compounds. Although there have been numerous notable advancements in the characterization of non-CYP enzymes with respect to their localization, reaction mechanisms, species differences and identification of typical substrates, accurate prediction of non-CYP-mediated clearance, with a particular emphasis with the difficulties in accounting for any extrahepatic contributions, remains a challenge. The current manuscript comprehensively summarizes the recent advancements in the prediction of drug metabolism and the in vitro to in vitro extrapolation of clearance for substrates of non-CYP drug metabolizing enzymes.
Assuntos
Sistema Enzimático do Citocromo P-450 , Microssomos Hepáticos , Sistema Enzimático do Citocromo P-450/metabolismo , Humanos , Inativação Metabólica , Taxa de Depuração Metabólica , Microssomos Hepáticos/metabolismoRESUMO
Oncogenic activation of BRAF fuels cancer growth by constitutively promoting RAS-independent mitogen-activated protein kinase (MAPK) pathway signalling. Accordingly, RAF inhibitors have brought substantially improved personalized treatment of metastatic melanoma. However, these targeted agents have also revealed an unexpected consequence: stimulated growth of certain cancers. Structurally diverse ATP-competitive RAF inhibitors can either inhibit or paradoxically activate the MAPK pathway, depending whether activation is by BRAF mutation or by an upstream event, such as RAS mutation or receptor tyrosine kinase activation. Here we have identified next-generation RAF inhibitors (dubbed 'paradox breakers') that suppress mutant BRAF cells without activating the MAPK pathway in cells bearing upstream activation. In cells that express the same HRAS mutation prevalent in squamous tumours from patients treated with RAF inhibitors, the first-generation RAF inhibitor vemurafenib stimulated in vitro and in vivo growth and induced expression of MAPK pathway response genes; by contrast the paradox breakers PLX7904 and PLX8394 had no effect. Paradox breakers also overcame several known mechanisms of resistance to first-generation RAF inhibitors. Dissociating MAPK pathway inhibition from paradoxical activation might yield both improved safety and more durable efficacy than first-generation RAF inhibitors, a concept currently undergoing human clinical evaluation with PLX8394.
Assuntos
Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Proteínas Quinases Ativadas por Mitógeno/metabolismo , Inibidores de Proteínas Quinases/farmacologia , Proteínas Proto-Oncogênicas B-raf/antagonistas & inibidores , Animais , Linhagem Celular Tumoral , Ativação Enzimática/efeitos dos fármacos , Feminino , Genes ras/genética , Compostos Heterocíclicos com 2 Anéis/efeitos adversos , Compostos Heterocíclicos com 2 Anéis/farmacologia , Humanos , Indóis/efeitos adversos , Indóis/farmacologia , Sistema de Sinalização das MAP Quinases/genética , Camundongos , Modelos Biológicos , Mutação/genética , Inibidores de Proteínas Quinases/efeitos adversos , Proteínas Proto-Oncogênicas B-raf/genética , Sulfonamidas/efeitos adversos , Sulfonamidas/farmacologia , VemurafenibRESUMO
Inflammation and cancer, two therapeutic areas historically addressed by separate drug discovery efforts, are now coupled in treatment approaches by a growing understanding of the dynamic molecular dialogues between immune and cancer cells. Agents that target specific compartments of the immune system, therefore, not only bring new disease modifying modalities to inflammatory diseases, but also offer a new avenue to cancer therapy by disrupting immune components of the microenvironment that foster tumor growth, progression, immune evasion, and treatment resistance. McDonough feline sarcoma viral (v-fms) oncogene homolog (FMS) and v-kit Hardy-Zuckerman 4 feline sarcoma viral oncogene homolog (KIT) are two hematopoietic cell surface receptors that regulate the development and function of macrophages and mast cells, respectively. We disclose a highly specific dual FMS and KIT kinase inhibitor developed from a multifaceted chemical scaffold. As expected, this inhibitor blocks the activation of macrophages, osteoclasts, and mast cells controlled by these two receptors. More importantly, the dual FMS and KIT inhibition profile has translated into a combination of benefits in preclinical disease models of inflammation and cancer.
Assuntos
Aminopiridinas/farmacologia , Inflamação/tratamento farmacológico , Modelos Moleculares , Metástase Neoplásica/tratamento farmacológico , Proteína Oncogênica gp140(v-fms)/antagonistas & inibidores , Inibidores de Proteínas Quinases/farmacologia , Proteínas Proto-Oncogênicas c-kit/antagonistas & inibidores , Pirróis/farmacologia , Aminopiridinas/síntese química , Aminopiridinas/química , Animais , Sobrevivência Celular/efeitos dos fármacos , Cromatografia de Afinidade , Cristalização , Escherichia coli , Células Endoteliais da Veia Umbilical Humana , Humanos , Indóis , Macrófagos/efeitos dos fármacos , Mastócitos/efeitos dos fármacos , Estrutura Molecular , Mutação de Sentido Incorreto/genética , Proteína Oncogênica gp140(v-fms)/química , Proteína Oncogênica gp140(v-fms)/genética , Osteoclastos/efeitos dos fármacos , Conformação Proteica , Inibidores de Proteínas Quinases/síntese química , Inibidores de Proteínas Quinases/química , Proteínas Proto-Oncogênicas c-kit/química , Proteínas Proto-Oncogênicas c-kit/genética , Pirróis/síntese química , Pirróis/química , Células Sf9 , SpodopteraRESUMO
PURPOSE: The Bromodomain and Extra-Terminal (BET) domain proteins facilitate the development of many human cancers via epigenetic regulation. BET inhibitors may be effective in reversing platinum resistance in ovarian cancer (OC) and may generate synthetic lethality with ARID1A loss. PLX2853 is an orally active, small-molecule inhibitor of BET bromodomain-mediated interactions that exhibits low nanomolar potency in blocking all four BET family members. METHODS: We conducted a multicenter and open-label study with two parallel arms: a phase IIa study of PLX2853 monotherapy in patients with advanced gynecologic malignancies with an ARID1A mutation and a phase Ib/IIa combination study of PLX2853 plus carboplatin in women with platinum-resistant OC. The primary objectives were safety and tolerability for phase Ib and efficacy for both phase IIa portions. Thirty-four of 37 enrolled patients completed at least one post-baseline response assessment. RESULTS: Of the 14 evaluable patients on the monotherapy arm, 1 (7.1%) achieved a best overall response of partial response (PR), 5 (35.7%) had stable disease (SD), and 8 (57.1%) had progressive disease (PD). Of the 20 evaluable patients on the combination arm, 1 (5.0%) had PR, 9 (45.0%) had SD, and 10 (50%) had PD. CONCLUSION: This study confirmed the safety profile of PLX2853 and demonstrated the feasibility of combination with carboplatin. Although these results did not meet the prespecified response criteria, evidence of clinical activity highlights the rationale for further exploration of BET inhibitors in patients with ARID1A-mutated gynecologic malignancies, possibly in combination with agents targeting potential feedback mechanisms such as the PI3K pathway.
Assuntos
Antineoplásicos , Neoplasias dos Genitais Femininos , Neoplasias Ovarianas , Feminino , Humanos , Carboplatina/uso terapêutico , Neoplasias dos Genitais Femininos/genética , Carcinoma Epitelial do Ovário/tratamento farmacológico , Carcinoma Epitelial do Ovário/genética , Epigênese Genética , Fosfatidilinositol 3-Quinases/genética , Antineoplásicos/uso terapêutico , Neoplasias Ovarianas/tratamento farmacológico , Neoplasias Ovarianas/genética , Mutação , Proteínas de Ligação a DNA/genética , Fatores de Transcrição/genéticaRESUMO
IMPORTANCE: Many cancer subtypes, including KIT-mutant gastrointestinal stromal tumors (GISTs), are driven by activating mutations in tyrosine kinases and may initially respond to kinase inhibitors but frequently relapse owing to outgrowth of heterogeneous subclones with resistance mutations. KIT inhibitors commonly used to treat GIST (eg, imatinib and sunitinib) are inactive-state (type II) inhibitors. OBJECTIVE: To assess whether combining a type II KIT inhibitor with a conformation-complementary, active-state (type I) KIT inhibitor is associated with broad mutation coverage and global disease control. DESIGN, SETTING, AND PARTICIPANTS: A highly selective type I inhibitor of KIT, PLX9486, was tested in a 2-part phase 1b/2a trial. Part 1 (dose escalation) evaluated PLX9486 monotherapy in patients with solid tumors. Part 2e (extension) evaluated PLX9486-sunitinib combination in patients with GIST. Patients were enrolled from March 2015 through February 2019; data analysis was performed from May 2020 through July 2020. INTERVENTIONS: Participants received 250, 350, 500, and 1000 mg of PLX9486 alone (part 1) or 500 and 1000 mg of PLX9486 together with 25 or 37.5 mg of sunitinib (part 2e) continuously in 28-day dosing cycles until disease progression, treatment discontinuation, or withdrawal. MAIN OUTCOMES AND MEASURES: Pharmacokinetics, safety, and tumor responses were assessed. Clinical efficacy end points (progression-free survival and clinical benefit rate) were supplemented with longitudinal monitoring of KIT mutations in circulating tumor DNA. RESULTS: A total of 39 PLX9486-naive patients (median age, 57 years [range, 39-79 years]; 22 men [56.4%]; 35 [89.7%] with refractory GIST) were enrolled in the dose escalation and extension parts. The recommended phase 2 dose of PLX9486 was 1000 mg daily. At this dose, PLX9486 could be safely combined with 25 or 37.5 mg daily of sunitinib continuously. Patients with GIST who received PLX9486 at a dose of 500 mg or less, at the recommended phase 2 dose, and with sunitinib had median (95% CI) progression-free survivals of 1.74 (1.54-1.84), 5.75 (0.99-11.0), and 12.1 (1.34-NA) months and clinical benefit rates (95% CI) of 14% (0%-58%), 50% (21%-79%), and 80% (52%-96%), respectively. CONCLUSIONS AND RELEVANCE: In this phase 1b/2a nonrandomized clinical trial, type I and type II KIT inhibitors PLX9486 and sunitinib were safely coadministered at the recommended dose of both single agents in patients with refractory GIST. Results suggest that cotargeting 2 complementary conformational states of the same kinase was associated with clinical benefit with an acceptable safety profile. TRIAL REGISTRATION: ClinicalTrials.gov Identifier: NCT02401815.
Assuntos
Tumores do Estroma Gastrointestinal , Mesilato de Imatinib , Inibidores de Proteínas Quinases , Sunitinibe , Tumores do Estroma Gastrointestinal/tratamento farmacológico , Tumores do Estroma Gastrointestinal/genética , Tumores do Estroma Gastrointestinal/patologia , Humanos , Mesilato de Imatinib/efeitos adversos , Masculino , Pessoa de Meia-Idade , Mutação , Recidiva Local de Neoplasia/tratamento farmacológico , Inibidores de Proteínas Quinases/efeitos adversos , Sunitinibe/efeitos adversosRESUMO
The purpose of the present study was to investigate the in vitro and in vivo activity of PLX9486, a tyrosine kinase inhibitor (TKI) targeting both primary KIT exon 9 and 11 and secondary exon 17 and 18 mutations in gastrointestinal stromal tumors (GISTs). Imatinib, a potent inhibitor of mutated KIT, has revolutionized the clinical management of advanced, metastatic GIST. However, secondary resistance develops mainly through acquired mutations in KIT exons 13/14 or exons 17/18. Second-line sunitinib potently inhibits KIT exon 13/14 mutants but is ineffective against exon 17 mutations. In our study, PLX9486 demonstrated in vitro nanomolar potency in inhibiting the growth and KIT phosphorylation of engineered BaF3 cells transformed with KIT exon 17 mutations (p.D816V) and with the double KIT exon 11/17 mutations (p.V560G/D816V). The in vivo efficacy of PLX9486 was evaluated using two imatinib-resistant GIST patient-derived xenograft (PDX) models. In UZLX-GIST9 (KIT: p.P577del;W557LfsX5;D820G), PLX9486 100 mg/kg/day resulted in significant inhibition of proliferation. Pharmacodynamic analysis showed a pronounced reduction in mitogen-activated protein kinase (MAPK) activation and other downstream effects of the KIT signaling pathway but no significant effect on KIT Y703 and Y719 phosphorylation. Similarly, in MRL-GIST1 (KIT: p.W557_K558del;Y823D) PLX9486 treatment led to significant tumor regression and strong inhibition of MAPK activation. Interestingly, the inhibitory effect on MAPK activation was evident even after a single dose of PLX9486. In conclusion, PLX9486 showed anti-tumor efficacy in patient-derived imatinib-resistant GIST xenograft models, mainly through inhibition of KIT signaling. These preclinical efficacy data encourage further testing of PLX9486 in the clinical setting.
Assuntos
Antineoplásicos/administração & dosagem , Inibidores Enzimáticos/administração & dosagem , Tumores do Estroma Gastrointestinal/tratamento farmacológico , Tumores do Estroma Gastrointestinal/patologia , Proteínas Mutantes/genética , Proteínas Tirosina Quinases/antagonistas & inibidores , Proteínas Proto-Oncogênicas c-kit/genética , Animais , Modelos Animais de Doenças , Avaliação Pré-Clínica de Medicamentos , Xenoenxertos , Humanos , Camundongos , Proteínas Mutantes/metabolismo , Transplante de Neoplasias , Proteínas Proto-Oncogênicas c-kit/metabolismo , Resultado do TratamentoRESUMO
Resistance to current therapies still impacts a significant number of melanoma patients and can be regulated by epigenetic alterations. Analysis of global cytosine methylation in a cohort of primary melanomas revealed a pattern of early demethylation associated with overexpression of oncogenic transcripts. Loss of methylation and associated overexpression of the CSF 1 receptor (CSF1R) was seen in a majority of tumors and was driven by an alternative, endogenous viral promoter in a subset of samples. CSF1R was particularly elevated in melanomas with BRAF and other MAPK activating mutations. Furthermore, rebound ERK activation after BRAF inhibition was associated with RUNX1-mediated further upregulation of CSF-1R and its ligand IL-34. Importantly, increased CSF-1R and IL-34 overexpression were detected in an independent cohort of resistant melanomas. Inhibition of CSF-1R kinase or decreased CSF-1R expression by RNAi reduced 3-D growth and invasiveness of melanoma cells. Coinhibition of CSF-1R and BRAF resulted in synergistic efficacy in vivo. To our knowledge, our data unveil a previously unknown role for the autocrine-regulated CSF-1R in BRAF V600E resistance and provide a preclinical rationale for targeting this pathway in melanoma.
Assuntos
Subunidade alfa 2 de Fator de Ligação ao Core/metabolismo , Resistencia a Medicamentos Antineoplásicos , Regulação Neoplásica da Expressão Gênica , Interleucinas/metabolismo , Melanoma/metabolismo , Proteínas Proto-Oncogênicas B-raf/metabolismo , Receptores de Fator Estimulador das Colônias de Granulócitos e Macrófagos/metabolismo , Animais , Antineoplásicos/farmacologia , Linhagem Celular Tumoral , Metilação de DNA , Sinergismo Farmacológico , Feminino , Humanos , Sistema de Sinalização das MAP Quinases , Melanoma/genética , Melanoma/patologia , Camundongos , Camundongos Endogâmicos BALB C , Quinases de Proteína Quinase Ativadas por Mitógeno/efeitos dos fármacos , Mutação , Proteínas Proto-Oncogênicas B-raf/antagonistas & inibidores , Receptores de Fator Estimulador das Colônias de Granulócitos e Macrófagos/antagonistas & inibidores , Receptores de Fator Estimulador das Colônias de Granulócitos e Macrófagos/genética , Células THP-1 , Transplante Heterólogo , Células U937RESUMO
Bromodomain and extra-terminal (BET) family proteins are key regulators of gene expression in cancer. Herein, we utilize BRD4 profiling to identify critical pathways involved in pathogenesis of chronic lymphocytic leukemia (CLL). BRD4 is overexpressed in CLL and is enriched proximal to genes upregulated or de novo expressed in CLL with known functions in disease pathogenesis and progression. These genes, including key members of the B-cell receptor (BCR) signaling pathway, provide a rationale for this therapeutic approach to identify new targets in alternative types of cancer. Additionally, we describe PLX51107, a structurally distinct BET inhibitor with novel in vitro and in vivo pharmacologic properties that emulates or exceeds the efficacy of BCR signaling agents in preclinical models of CLL. Herein, the discovery of the involvement of BRD4 in the core CLL transcriptional program provides a compelling rationale for clinical investigation of PLX51107 as epigenetic therapy in CLL and application of BRD4 profiling in other cancers.Significance: To date, functional studies of BRD4 in CLL are lacking. Through integrated genomic, functional, and pharmacologic analyses, we uncover the existence of BRD4-regulated core CLL transcriptional programs and present preclinical proof-of-concept studies validating BET inhibition as an epigenetic approach to target BCR signaling in CLL. Cancer Discov; 8(4); 458-77. ©2018 AACR.This article is highlighted in the In This Issue feature, p. 371.
Assuntos
Regulação Leucêmica da Expressão Gênica , Isoxazóis/uso terapêutico , Leucemia Linfocítica Crônica de Células B/tratamento farmacológico , Proteínas Nucleares/genética , Piridinas/uso terapêutico , Pirróis/uso terapêutico , Transdução de Sinais , Fatores de Transcrição/genética , Animais , Antineoplásicos/farmacologia , Antineoplásicos/uso terapêutico , Proteínas de Ciclo Celular , Linhagem Celular Tumoral , Proliferação de Células , Perfilação da Expressão Gênica , Humanos , Isoxazóis/farmacologia , Leucemia Linfocítica Crônica de Células B/genética , Leucemia Linfocítica Crônica de Células B/metabolismo , Leucemia Linfocítica Crônica de Células B/fisiopatologia , Camundongos , Camundongos SCID , Proteínas Nucleares/metabolismo , Piridinas/farmacologia , Pirróis/farmacologia , Fatores de Transcrição/metabolismo , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
UNLABELLED: Tyrosine kinase domain mutations are a common cause of acquired clinical resistance to tyrosine kinase inhibitors (TKI) used to treat cancer, including the FLT3 inhibitor quizartinib. Mutation of kinase "gatekeeper" residues, which control access to an allosteric pocket adjacent to the ATP-binding site, has been frequently implicated in TKI resistance. The molecular underpinnings of gatekeeper mutation-mediated resistance are incompletely understood. We report the first cocrystal structure of FLT3 with the TKI quizartinib, which demonstrates that quizartinib binding relies on essential edge-to-face aromatic interactions with the gatekeeper F691 residue, and F830 within the highly conserved Asp-Phe-Gly motif in the activation loop. This reliance makes quizartinib critically vulnerable to gatekeeper and activation loop substitutions while minimizing the impact of mutations elsewhere. Moreover, we identify PLX3397, a novel FLT3 inhibitor that retains activity against the F691L mutant due to a binding mode that depends less vitally on specific interactions with the gatekeeper position. SIGNIFICANCE: We report the first cocrystal structure of FLT3 with a kinase inhibitor, elucidating the structural mechanism of resistance due to the gatekeeper F691L mutation. PLX3397 is a novel FLT3 inhibitor with in vitro activity against this mutation but is vulnerable to kinase domain mutations in the FLT3 activation loop.
Assuntos
Aminopiridinas/farmacologia , Benzotiazóis/farmacologia , Resistencia a Medicamentos Antineoplásicos/genética , Mutação , Compostos de Fenilureia/farmacologia , Pirróis/farmacologia , Tirosina Quinase 3 Semelhante a fms/genética , Aminopiridinas/química , Animais , Antineoplásicos/química , Antineoplásicos/farmacologia , Benzotiazóis/química , Linhagem Celular Tumoral , Ativação Enzimática/efeitos dos fármacos , Xenoenxertos , Humanos , Leucemia Mieloide Aguda/tratamento farmacológico , Leucemia Mieloide Aguda/genética , Leucemia Mieloide Aguda/patologia , Camundongos , Modelos Moleculares , Conformação Molecular , Compostos de Fenilureia/química , Ligação Proteica , Domínios e Motivos de Interação entre Proteínas/genética , Inibidores de Proteínas Quinases/química , Inibidores de Proteínas Quinases/farmacologia , Pirróis/química , Recidiva , Relação Estrutura-Atividade , Tirosina Quinase 3 Semelhante a fms/químicaRESUMO
The colony-stimulating factor 1 receptor (CSF1R) is a key regulator of myeloid lineage cells. Genetic loss of the CSF1R blocks the normal population of resident microglia in the brain that originates from the yolk sac during early development. However, the role of CSF1R signaling in microglial homeostasis in the adult brain is largely unknown. To this end, we tested the effects of selective CSF1R inhibitors on microglia in adult mice. Surprisingly, extensive treatment results in elimination of â¼99% of all microglia brain-wide, showing that microglia in the adult brain are physiologically dependent upon CSF1R signaling. Mice depleted of microglia show no behavioral or cognitive abnormalities, revealing that microglia are not necessary for these tasks. Finally, we discovered that the microglia-depleted brain completely repopulates with new microglia within 1 week of inhibitor cessation. Microglial repopulation throughout the CNS occurs through proliferation of nestin-positive cells that then differentiate into microglia.