Mechanisms of cardiomyocyte dysfunction due to the E258K-MYBPC3 mutation modeled in patient-derived cardiomyocytes
在患者来源的心肌细胞中建模 E258K-MYBPC3 突变引起的心肌细胞功能障碍的机制
基本信息
- 批准号:10794930
- 负责人:
- 金额:$ 4.57万
- 依托单位:
- 依托单位国家:美国
- 项目类别:
- 财政年份:2022
- 资助国家:美国
- 起止时间:2022-09-30 至 2025-09-29
- 项目状态:未结题
- 来源:
- 关键词:3-DimensionalAccelerationActinsAddressAffectAge of OnsetBackBindingBiochemistryBiological AssayCRISPR/Cas technologyCardiac MyocytesCardiac MyosinsCell LineCellsClinicalCustomDataDevelopmentDiseaseDisease ProgressionEvaluationExtracellular MatrixFibrinFounder EffectFunctional disorderGenderGenerationsGeneticGenus HippocampusHeart DiseasesHeart TransplantationHeart failureHypertrophic CardiomyopathyInduced MutationInheritedItalyKineticsLeadLeftLengthLinkMass Spectrum AnalysisMetabolismMissense MutationModalityModelingMutationMyocardiumMyofibrilsMyosin ATPaseObstructionOperative Surgical ProceduresPathogenicityPathologicPatientsPatternPhenotypePhosphocreatinePrevalenceProductionProteinsProteomicsQuality of lifeRelaxationRespirationSamplingSarcomeresStructureSurfaceSymptomsTestingTherapeuticThick FilamentThrombinTissue constructsTissuesUniversity HospitalsVariantVentricularcardiac muscle diseasecardiac tissue engineeringcitrate carriercohortcostgenetic variantheart functionindividualized medicineinduced pluripotent stem cellinsightliquid chromatography mass spectrometrymetabolic profilemetabolomicsmutantmutation correctionmyosin-binding protein Cpre-clinicalprotein expressionprotein protein interactionrecruitscreeningstem cell differentiationstoichiometrysudden cardiac deathsymptom managementtargeted treatmentyoung adult
项目摘要
Hypertrophic cardiomyopathy (HCM) is the most common inherited heart disease, characterized by progressive
thickening of the left ventricular walls and potential for sudden cardiac death. Twenty-five percent of HCM
mutations occur in the sarcomere protein cardiac myosin binding protein-C (cMyBP-C). Currently, there is no
cure for HCM, only management of symptoms and disease progression, left ventricular obstruction surgery, or
heart transplantation. As such, there is great need to better understand the pathological mechanisms that underly
specific HCM mutations in order to better inform development of targeted therapeutics. For this project, I will
study a highly penetrant mutation in cMyBP-C, c.772G>A (p.E258K), that has an identified founder effect in the
north-east Tuscany region of Italy. To better explore the direct impacts of the mutation, I have generated patient
induced pluripotent stem cells (iPSCs) from six HCM patients carrying the E258K mutation and a representative
isogenic cell line using CRISPR/Cas9 by correcting the mutation. Initial studies have been performed on
myectomy samples from three of the above six HCM patients with the E258K mutation, however, such patient
tissue is limited and provides results from late stage of disease. Utilizing our patient iPSC lines, I can more
thoroughly probe mechanisms underlying HCM from an almost unlimited supply of tissue specific cells. I propose
to study multiple patient-derived iPSC lines all harboring the same E258K mutation, allowing me to probe
the mechanism of the E258K mutation as well as investigate how other factors such as gender and age
of onset may affect said mechanisms. Within the E258K patient cohort at the Careggi University Hospital,
myectomy samples demonstrate consistently lower expression of full-length cMyBP-C protein, suggesting a
potential haploinsufficiency disease mechanism. At the level of the sarcomere, myectomy samples indicate
accelerated cross-bridge cycling, accompanied by a greater energetic cost of tension generation. Taken
together, I hypothesize that the E258K mutation 1) destabilizes cMyBP-C’s ability to recruit and regulate
myosin, leading to reduced expression and/or incorporation of cMyBP-C into the sarcomere
(haploinsufficiency) and 2) shifts the sarcomere to a state of excessive ATP utilization during contraction
(energetic inefficiency). To test this hypothesis, I will use our patient iPSCs differentiated to cardiomyocytes,
and their isogenic control lines, cultured on linear, aligned substrate surfaces to enhance maturation of
cardiomyocyte structure and function. My hypothesis will be tested with multiple modalities: myofibril cross-bridge
kinetics, evaluation of myosin confirmations by stopped flow (disordered relaxed state vs. super-relaxed state),
cMyBP-C expression and stoichiometry in the sarcomere using mass spectrometry (MS) based proteomics,
cellular metabolism via Seahorse assay, substrate utilization via MS based metabolomics and energetic cost of
tension generation using engineered heart tissue (EHT) constructs. If successful, this study will help uncover the
mechanism of this highly penetrant HCM mutation and inform preclinical screening of potential therapeutics.
肥厚性心肌病(HCM)是最常见的遗传性心脏病,以进行性为特征
项目成果
期刊论文数量(0)
专著数量(0)
科研奖励数量(0)
会议论文数量(0)
专利数量(0)
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Sonette Steczina其他文献
Sonette Steczina的其他文献
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{{ truncateString('Sonette Steczina', 18)}}的其他基金
Mechanisms of cardiomyocyte dysfunction due to the E258K-MYBPC3 mutation modeled in patient-derived cardiomyocytes
在患者来源的心肌细胞中建模 E258K-MYBPC3 突变引起的心肌细胞功能障碍的机制
- 批准号:
10462968 - 财政年份:2022
- 资助金额:
$ 4.57万 - 项目类别:
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