Gene-edited liver organoids for predictive hepatotoxicity
用于预测肝毒性的基因编辑肝脏类器官
基本信息
- 批准号:10758179
- 负责人:
- 金额:$ 28.46万
- 依托单位:
- 依托单位国家:美国
- 项目类别:
- 财政年份:2023
- 资助国家:美国
- 起止时间:2023-08-01 至 2024-07-31
- 项目状态:已结题
- 来源:
- 关键词:3-DimensionalABCB1 geneAddressAffinityAllelesAnimal ModelArchitectureAssessment toolBiological AssayBiomimeticsCRISPR/Cas technologyCYP2B6 geneCYP2C19 geneCYP2D6 geneCYP3A4 geneCaucasiansCause of DeathCell LineCellsCessation of lifeChemicalsClassificationClustered Regularly Interspaced Short Palindromic RepeatsDiffusionDisease modelDoxycyclineDrug toxicityEngineeringEnzymesEpigenetic ProcessEthnic PopulationFailureGenesGenetic EngineeringGenetic PolymorphismGenetic VariationGoalsGuide RNAHepG2HepaticHepatocyteHepatotoxicityHospitalizationHumanHuman EngineeringHydrogelsImageIn VitroIndividualIntestinesLaboratoriesLentivirusLinkLiverMarketingMeasuresMediator of activation proteinMetabolic BiotransformationMetabolismMicrofluidicsModelingMolecularMorbidity - disease rateNatureNutrientOrganoidsOxygenPathogenicityPatientsPerfusionPharmaceutical PreparationsProteinsProtocols documentationReproducibilitySmall Business Innovation Research GrantStainsSystemTechnologyTestingTimeTissue ModelTissuesToxic effectToxicity TestsUnited States Food and Drug AdministrationVariantWorkbioprintingcost estimatedrug candidatedrug developmentdrug efficacydrug induced liver injurydrug metabolismdrug withdrawalfeasibility testingfetalhepatoma cellhigh throughput screeningimprovedin vivoindividual patientindividual variationinduced pluripotent stem cellinsightinterestmanufacturing scale-upmatrigelmortalityoverexpressionpre-clinicalresponsesafety assessment
项目摘要
Project Summary/Abstract
Unexpected adverse drug responses (ADRs) including drug-induced liver injury (DILI) are the 4th
leading cause of death in the U.S. In addition, DILI in individuals is one of the major reasons for drug
withdrawal from the market and is difficult to predict using conventional in vitro hepatotoxicity tests and
preclinical animal models. Due to the critical link between DILI and drug failure, there is an urgent need
for improved human hepatotoxicity testing in the early stage of drug development by investigating the
major pathogenic mechanisms of DILI, such as genetic variations in drug metabolizing enzymes (DMEs)
and drug transporters. To address this need, we propose to develop genetically engineered human liver
organoids (HLOs) on a pillar/perfusion plate using gene-edited, induced pluripotent stem cell (iPSC) lines
carrying CRISPR/Cas9 synergistic activation mediator (CRISPR-SAM), inducible Csy4, and multiplexed
guide RNA (gRNA), and recapitulate poor and ultrafast drug metabolizers in different ethnic groups. Using
normal and engineered HLOs on the pillar/perfusion plate, together with high-throughput, high-content,
HLO imaging assays, we propose to decipher the cellular and molecular mechanisms underlying the
toxicity of drug candidates and chemicals and assess DILI potential.
Our core hypotheses are: (i) overexpression of DMEs and drug transporters can recapitulate ultrafast
drug metabolizers in different ethnic groups that may be critical in addressing ADRs; (ii) metabolism-
induced hepatotoxicity can be established using normal and engineered HLOs with model compounds;
and (iii) high-throughput, high-content analysis of HLOs on the pillar/perfusion plate can be used to
identify DILI, which in turn can improve predictability of compound hepatotoxicity in vivo.
The specific aims of the proposed work are to: (1) create genetically engineered HLOs containing
doxycycline-inducible, CRISPR-SAM for overexpression of multiple hepatic genes to model ultrafast
metabolizers; (2) validate normal and engineered HLOs with model compounds, which undergo
metabolism and lead to toxic cellular responses in the human liver.
Although several human hepatic cell/tissue models including primary hepatocytes (ATCC),
engineered hepatoma cell lines (HepG2-CYP cell panel from Hera BioLabs), liver spheroids (3D InSight
liver microtissues from InSphero), bioprinted liver tissues (ExVive human liver tissues from Organovo),
and microfluidic liver chips (Liver-Chip from Emulate) are commercially available, these in vitro liver
models have been used for assessing general hepatotoxicity of compounds for normal drug metabolizers
and cannot easily simulate poor and ultrafast drug metabolizers in their assays who suffer the most from
DILI. Thus, there is great potential to apply engineered HLOs on the pillar/perfusion plate as a safety
assessment tool. Genetically engineered human organoids can be used to express any endogenous
proteins of interest in the cells for disease modeling by using a combination of guide RNAs. There is a
great potential for genetically engineered human organoids to be used to incorporate genetic diversity
into toxicity testing and for disease modeling.
项目总结/文摘
项目成果
期刊论文数量(0)
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科研奖励数量(0)
会议论文数量(0)
专利数量(0)
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Pranav Joshi其他文献
Pranav Joshi的其他文献
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{{ truncateString('Pranav Joshi', 18)}}的其他基金
Pillar and perfusion well plate platforms for reproducible organoid culture from iPSC
用于从 iPSC 进行可重复类器官培养的支柱和灌注孔板平台
- 批准号:
10080406 - 财政年份:2020
- 资助金额:
$ 28.46万 - 项目类别:
Pillar and perfusion well plate platforms for reproducible organoid culture from iPSC
用于从 iPSC 进行可重复类器官培养的支柱和灌注孔板平台
- 批准号:
10210319 - 财政年份:2020
- 资助金额:
$ 28.46万 - 项目类别:
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