Clinic for Cardiology and Pneumology, University Medical Center, 37075 Göttingen, Germany.
German Centre for Cardiovascular Research, 10785 Berlin, partner site Göttingen, Germany.
Sci Transl Med. 2021 Nov 3;13(618):eabd3079. doi: 10.1126/scitranslmed.abd3079.
Heterozygous truncating variants in (TTNtv), the gene coding for titin, cause dilated cardiomyopathy (DCM), but the underlying pathomechanisms are unclear and disease management remains uncertain. Truncated titin proteins have not yet been considered as a contributor to disease development. Here, we studied myocardial tissues from nonfailing donor hearts and 113 patients with end-stage DCM for titin expression and identified a TTNtv in 22 patients with DCM (19.5%). We directly demonstrate titin haploinsufficiency in TTNtv-DCM hearts and the absence of compensatory changes in the alternative titin isoform Cronos. Twenty-one TTNtv-DCM hearts in our cohort showed stable expression of truncated titin proteins. Expression was variable, up to half of the total titin protein pool, and negatively correlated with patient age at heart transplantation. Truncated titin proteins were not detected in sarcomeres but were present in intracellular aggregates, with deregulated ubiquitin-dependent protein quality control. We produced human induced pluripotent stem cell–derived cardiomyocytes (hiPSC-CMs), comparing wild-type controls to cells with a patient-derived, prototypical A-band-TTNtv or a CRISPR-Cas9–generated M-band-TTNtv. TTNtv-hiPSC-CMs showed reduced wild-type titin expression and contained truncated titin proteins whose proportion increased upon inhibition of proteasomal activity. In engineered heart muscle generated from hiPSC-CMs, depressed contractility caused by TTNtv could be reversed by correction of the mutation using CRISPR-Cas9, eliminating truncated titin proteins and raising wild-type titin content. Functional improvement also occurred when wild-type titin protein content was increased by proteasome inhibition. Our findings reveal the major pathomechanisms of TTNtv-DCM and can be exploited for new therapies to treat TTNtv-related cardiomyopathies.
肌联蛋白（TTN）基因的杂合截断变异（TTNtv）可导致扩张型心肌病（DCM），但其潜在的发病机制尚不清楚，疾病管理仍不确定。截断的肌联蛋白蛋白尚未被认为是导致疾病发展的因素之一。在这里，我们研究了非衰竭供体心脏和 113 例终末期 DCM 患者的心肌组织中的肌联蛋白表达，并在 22 例 DCM 患者（19.5%）中发现了 TTNtv。我们直接证明了 TTNtv-DCM 心脏中的肌联蛋白单倍不足，以及替代肌联蛋白同工型 Cronos 中不存在代偿性变化。我们队列中的 21 例 TTNtv-DCM 心脏显示截断的肌联蛋白蛋白稳定表达。表达是可变的，可达总肌联蛋白蛋白库的一半，与心脏移植时患者的年龄呈负相关。在肌节中未检测到截断的肌联蛋白蛋白，但存在于细胞内聚集体中，伴有失调的泛素依赖性蛋白质量控制。我们生成了人诱导多能干细胞衍生的心肌细胞（hiPSC-CMs），将野生型对照与具有患者衍生的典型 A 带-TTNtv 或 CRISPR-Cas9 生成的 M 带-TTNtv 的细胞进行比较。TTNtv-hiPSC-CMs 显示野生型肌联蛋白表达减少，并且含有截断的肌联蛋白蛋白，其比例在蛋白酶体活性抑制时增加。在由 hiPSC-CMs 生成的工程化心肌中，通过 CRISPR-Cas9 纠正突变可以逆转 TTNtv 引起的收缩力下降，消除截断的肌联蛋白蛋白并提高野生型肌联蛋白含量。通过蛋白酶体抑制增加野生型肌联蛋白蛋白含量也可实现功能改善。我们的研究结果揭示了 TTNtv-DCM 的主要发病机制，并可用于治疗 TTNtv 相关心肌病的新疗法。
