RESUMO
Nonclinical safety and pharmacokinetic data for monomethyl auristatin E (MMAE) and 14 vedotin antibody-drug conjugates (ADC) were evaluated to determine patterns of toxicity, consistency of pharmacokinetic results, and species differences between rats and monkeys. Most nonclinical toxicities were antigen-independent, common across ADCs, and included hematologic, lymphoid, and reproductive toxicity related to MMAE pharmacology. Hematologic toxicity was the dose-limiting toxicity (DLT) or predominant toxicity for the majority of vedotin ADCs in both species. Tissue expression of the targeted antigen of an ADC rarely correlated with DLT; only two ADCs had antigen-dependent skin DLTs. For two additional ADCs, antigen-dependent delivery of MMAE in the bone marrow may have exacerbated the antigen-independent hematologic DLT. The highest tolerated doses and pharmacokinetics were similar within a given species, with rats tolerating higher doses than monkeys. Studies longer than 1 month in duration detected the same or fewer toxicities than 1-month studies and had no additional findings that affected the human risk assessment. These data support opportunities to streamline ADC toxicity assessments without compromising human starting dose selection or target organ identification.
Assuntos
Imunoconjugados , Animais , Imunoconjugados/farmacocinética , Imunoconjugados/efeitos adversos , Ratos , Humanos , Brentuximab Vedotin/farmacocinética , Masculino , Feminino , Oligopeptídeos/farmacocinética , Macaca fascicularisRESUMO
PURPOSE: Tucatinib, a small molecule for the treatment of metastatic HER2-positive breast cancer, was extensively metabolized in humans to multiple oxidative metabolites. To fully understand the elimination and biotransformation pathways of tucatinib, we investigated the in vitro and in vivo metabolism of tucatinib, and also conducted a Phase I trial using [14C]tucatinib. METHODS: To identify the responsible enzymes for tucatinib clearance, we investigated the in vitro metabolism of tucatinib including enzyme phenotyping, which facilitated the discovery of several metabolites in human and monkey plasma and excreta, in particular M1 (ONT-993, an aliphatic hydroxylated metabolite). Stereoselective formation of M1 was further investigated in vitro, in vivo, and in silico. RESULTS: In humans, approximately 86% of the total radiolabeled dose was recovered in feces and 4% in urine; in plasma, approximately 76% of radioactivity circulated as parent drug, with 19% attributed to multiple metabolites. The primary isoforms responsible for the elimination of tucatinib were CYP2C8 and CYP3A4/5. CYP2C8 was shown to possess sole catalytic activity for the formation of M1, whereas CYP3A4/5 and aldehyde oxidase catalyzed the formation of the remaining metabolites. Subsequent investigation revealed that M1 was formed in a stereoselective manner. Examination of the enantiomeric ratio of M1 stereoisomers observed in humans relative to cynomolgus monkeys revealed comparable results, suggesting that the enantiomers that comprise M1 were not considered to be unique or disproportionately high in human. CONCLUSION: CYP2C8 and CYP3A4/5 are the primary drug-metabolizing enzymes involved in the in vitro metabolism of tucatinib, which provided the basis to describe human disposition of tucatinib and formation of the observed metabolites.
Assuntos
Antineoplásicos , Citocromo P-450 CYP3A , Antineoplásicos/metabolismo , Citocromo P-450 CYP2C8/metabolismo , Citocromo P-450 CYP3A/metabolismo , Humanos , Microssomos Hepáticos/metabolismo , Oxazóis , Inibidores de Proteínas Quinases/metabolismo , Piridinas , Quinazolinas , EstereoisomerismoRESUMO
OBJECTIVE: The goal of this work was to endothelialize silicone aneurysm tubes for use as in vitro models for evaluating endothelial cell interactions with neurovascular devices. The first objective was to establish consistent and confluent endothelial cell linings and to evaluate the silicone vessels over time. The second objective was to use these silicone vessels for flow diverter implantation and assessment. METHODS: Silicone aneurysm tubes were coated with fibronectin and placed into individual bioreactor systems. Human umbilical vein endothelial cells were deposited within tubes to create silicone vessels, then cultivated on a peristaltic pump and harvested at 2, 5, 7, or 10 days to evaluate the endothelial cell lining. A subset of silicone aneurysm vessels was used for flow diverter implantation, and evaluated for cell coverage over device struts at 3 or 7 days after deployment. RESULTS: Silicone vessels maintained confluent, PECAM-1 (platelet endothelial cell adhesion molecule 1) positive endothelial cell linings over time. These vessels facilitated and withstood flow diverter implantation, with robust cell linings disclosed after device deployment. Additionally, the endothelial cells responded to implanted devices through coverage of the flow diverter struts with increased cell coverage over the aneurysm seen at 7 days after deployment as compared with 3 days. CONCLUSIONS: Silicone aneurysm models can be endothelialized and successfully maintained in vitro over time. Furthermore, these silicone vessels can be used for flow diverter implantation and assessment.
Assuntos
Órgãos Artificiais , Vasos Sanguíneos , Endotélio Vascular , Modelos Anatômicos , Elastômeros de Silicone/farmacologia , Materiais Biocompatíveis/farmacologia , Endotélio Vascular/citologia , Endotélio Vascular/fisiologia , Desenho de Equipamento , Humanos , Aneurisma Intracraniano/cirurgia , Propriedades de Superfície , Procedimentos Cirúrgicos Vasculares/métodosRESUMO
BACKGROUND: Notwithstanding the widespread implementation of flow diverters (FDs) in the treatment of intracranial aneurysms, the exact mechanism of action of these devices remains elusive. We aimed to advance the understanding of cellular responses to FD implantation using a 3D tissue-engineered in vitro aneurysm model. METHODS: Aneurysm-like blood vessel mimics (aBVMs) were constructed by electrospinning polycaprolactone nanofibers onto desired aneurysm-like geometries. aBVMs were seeded with human aortic smooth muscle cells (SMCs) followed by human aortic endothelial cells (ECs). FDs were then deployed in the parent vessel of aBVMs covering the aneurysm neck and were cultivated for 7, 14, or 28 days (n=3 for each time point). The EC and SMC coverage in the neck was measured semi-quantitatively. RESULTS: At day 7, the device segment in contact with the parent vessel was partially endothelialized. Also, the majority of device struts, but not pores, at the parent vessel and neck interface were partially covered with ECs and SMCs, while device struts in the middle of the neck lacked cell coverage. At 14 days, histology verified a neointimal-like lining had formed, partially covering both the struts and pores in the center of the neck. At 28 days, the majority of the neck was covered with a translucent neointimal-like layer. A higher degree of cellular coverage was seen on the struts and pores at the neck at 28 days compared with both 7 and 14 days. CONCLUSION: aBVMs can be a valuable alternative tool for evaluating the healing mechanisms of endovascular aneurysm devices.
Assuntos
Órgãos Artificiais , Vasos Sanguíneos , Aneurisma Intracraniano/cirurgia , Miócitos de Músculo Liso , Engenharia Tecidual/métodos , Materiais Biocompatíveis/farmacologia , Procedimentos Endovasculares/instrumentação , Desenho de Equipamento , Humanos , Modelos Anatômicos , Miócitos de Músculo Liso/citologia , Miócitos de Músculo Liso/fisiologia , Poliésteres/farmacologia , Stents , Propriedades de SuperfícieRESUMO
Acalabrutinib is a targeted, covalent inhibitor of Bruton tyrosine kinase (BTK) with a unique 2-butynamide warhead that has relatively lower reactivity than other marketed acrylamide covalent inhibitors. A human [14C] microtracer bioavailability study in healthy subjects revealed moderate intravenous clearance (39.4 l/h) and an absolute bioavailability of 25.3% ± 14.3% (n = 8). Absorption and elimination of acalabrutinib after a 100 mg [14C] microtracer acalabrutinib oral dose was rapid, with the maximum concentration reached in <1 hour and elimination half-life values of <2 hours. Low concentrations of radioactivity persisted longer in the blood cell fraction and a peripheral blood mononuclear cell subfraction (enriched in target BTK) relative to plasma. [14C]Acalabrutinib was metabolized to more than three dozen metabolites detectable by liquid chromatography-tandem mass spectrometry, with primary metabolism by CYP3A-mediated oxidation of the pyrrolidine ring, thiol conjugation of the butynamide warhead, and amide hydrolysis. A major active, circulating, pyrrolidine ring-opened metabolite, ACP-5862 (4-[8-amino-3-[4-(but-2-ynoylamino)butanoyl]imidazo[1,5-a]pyrazin-1-yl]-N-(2-pyridyl)benzamide), was produced by CYP3A oxidation.Novel enol thioethers from the 2-butynamide warhead arose from glutathione and/or cysteine Michael additions and were subject to hydrolysis to a ß-ketoamide. Total radioactivity recovery was 95.7% ± 4.6% (n = 6), with 12.0% of dose in urine and 83.5% in feces. Excretion and metabolism characteristics were generally similar in rats and dogs. Acalabrutinib's highly selective, covalent mechanism of action, coupled with rapid absorption and elimination, enables high and sustained BTK target occupancy after twice-daily administration.
Assuntos
Tirosina Quinase da Agamaglobulinemia/antagonistas & inibidores , Antineoplásicos/farmacologia , Benzamidas/farmacologia , Citocromo P-450 CYP3A/metabolismo , Inibidores de Proteínas Quinases/farmacologia , Pirazinas/farmacologia , Administração Oral , Adulto , Animais , Antineoplásicos/análise , Antineoplásicos/metabolismo , Benzamidas/análise , Benzamidas/metabolismo , Disponibilidade Biológica , Cães , Fezes/química , Feminino , Meia-Vida , Voluntários Saudáveis , Humanos , Hidrólise , Absorção Intestinal , Linfoma de Célula do Manto/tratamento farmacológico , Masculino , Pessoa de Meia-Idade , Oxirredução , Inibidores de Proteínas Quinases/análise , Inibidores de Proteínas Quinases/metabolismo , Pirazinas/análise , Pirazinas/metabolismo , Ratos , Ratos Sprague-Dawley , Urina/química , Adulto JovemRESUMO
Optical coherence tomography (OCT) is an imaging modality that enables assessment of tissue structural characteristics. Studies have indicated that OCT is a useful method to assess both blood vessel morphology and the response of a vessel to a deployed stent. We evaluated the ability of OCT to visualize the cellular lining of a tissue-engineered blood vessel mimic (BVM) and the response of this lining to a bare metal stent. We develop a side-firing endoscope that obtains intraluminal, longitudinal scans within the sterile bioreactor environment, enabling time-serial assessment. Seventeen BVMs are imaged with the endoscopic OCT system. The BVMs are then evaluated via fluorescence microscopy and/or standard histologic techniques. We determine that (1) the OCT endoscope can be repeatedly inserted without visible damage to the BVM cellular lining, (2) OCT provides a precise measure of cellular lining thickness with good correlation to measurements obtained from histological sections, and (3) OCT is capable of monitoring the accumulation of cellular material in response to a metallic stent. Our studies indicate that OCT is a useful technique for monitoring the BVM cellular lining, and that OCT may facilitate the use of BVMs for early stage device assessment.