Heterozygous truncating variants in TTN (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.

中文翻译：

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由于 TTN 突变，截短的肌动蛋白和肌动蛋白单倍体不足是人类心肌病功能恢复的目标

TTN中的杂合截断变体(TTNtv) 是 Titin 的编码基因，可导致扩张型心肌病 (DCM)，但其潜在的病理机制尚不清楚，疾病管理仍不确定。截短的肌联蛋白尚未被认为是疾病发展的一个因素。在这里，我们研究了来自非衰竭供体心脏和 113 名终末期 DCM 患者的心肌组织的肌动蛋白表达，并在 22 名 DCM 患者（19.5%）中发现了 TTNtv。我们直接证明了 TTNtv-DCM 心脏中的肌动蛋白单倍体不足以及替代肌动蛋白亚型 Cronos 中没有补偿性变化。我们队列中的 21 颗 TTNtv-DCM 心脏显示出稳定的截短肌动蛋白表达。表达是可变的，高达总肌联蛋白库的一半，并且与心脏移植的患者年龄呈负相关。在肌节中未检测到截短的肌动蛋白，但存在于细胞内聚集体中，具有失调的泛素依赖性蛋白质质量控​​制。我们生产了人类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原型 A-band-TTNtv 或 CRISPR-Cas9 生成的 M-band-TTNtv 的细胞进行了比较。TTNtv-hiPSC-CMs 显示野生型肌联蛋白表达减少，并含有截短的肌联蛋白，其比例在抑制蛋白酶体活性后增加。在由 hiPSC-CM 生成的工程心肌中，由 TTNtv 引起的收缩性降低可以通过使用 CRISPR-Cas9 校正突变、消除截短的肌动蛋白和提高野生型肌动蛋白含量来逆转。当通过蛋白酶体抑制增加野生型肌联蛋白含量时，也发生了功能改善。我们的研究结果揭示了 TTNtv-DCM 的主要病理机制，可用于治疗 TTNtv 相关心肌病的新疗法。