b1AR-mediated EGFR transactivation in the heart

 描述(申请人提供)：心力衰竭(HF)是一种进行性疾病，在美国影响着数以百万计的人，这些患者的护理每年花费数十亿美元，而患者的症状和死亡率随着神经激素受体拮抗剂(如β阻滞剂)的使用而减少，它们对心脏重塑和患者寿命的长期影响是有限的。G蛋白偶联受体信号通路已经被证明是可调节的，其中保护性通路(通常是β依赖的)可以通过有偏见的配体来选择，而有害的通路(通常是G蛋白依赖性的)同时被抑制。然而，尤其是β肾上腺素能受体(β受体)的偏向配体的开发缺乏令人失望的进展，已被证明通过β依赖的表皮生长因子受体(EGFR)的反式激活来传递心脏保护结果。在过去的几年里，我们已经广泛地描述了在β1AR刺激后，心肌细胞和整个心脏中因急性和慢性EGFR反式激活而发生的亚细胞信号、转录和心脏重塑反应。我们的研究强调了β-1AR介导的表皮生长因子受体反式激活的潜在治疗益处，并为其作为心力衰竭治疗策略的追求提供了理论基础，但缺乏特定的工具阻碍了确定性研究的实施。特别是，真正的β1AR特异性βARR/EGFR偏向配体 具有增强的疗效，并且不影响G蛋白依赖的信号转导，则不存在。因此，迄今为止的表皮生长因子受体反式激活研究依赖于在存在β受体刺激的情况下抑制表皮生长因子受体，以间接测量其对心肌细胞/心脏信号、转录和生存结果的影响。此外，EGFR基因敲除动物是不可行的，因此在没有使用引入选择性和毒性问题的药物抑制剂的情况下，无法评估成年心力衰竭模型的EGFR功能。考虑到这些问题， 我们已经开始测试β蛋白，这是GPCRs细胞内环的小肽片段，可以变构地诱导有偏见的信号，特别是GPCRs 1AR，目的是在没有G蛋白激活的情况下驱动EGR的反式激活。这些新的分子工具将首次在不依赖药物抑制剂的情况下，对β1AR介导的心肌细胞EGFR反式激活进行阳性表征。此外，我们还建立了一种心肌细胞特异性可诱导的表皮生长因子受体缺失的小鼠模型，该模型将明确确定表皮生长因子受体信号在心脏中的影响，以响应βAR刺激，以及在更广泛形式的心衰发生的背景下。因此，这些研究将首次直接评估心肌细胞表达的表皮生长因子受体在心力衰竭过程中的重要性，并产生新的、潜在的治疗性药物，用于特异性地激活β1AR抑制的表皮生长因子受体反式激活，以促进生存信号并减少心力衰竭期间有害的结构和功能后果。

 DESCRIPTION (provided by applicant): Heart failure (HF) is a progressive disease affecting millions of people in the U.S. with the care of these patients costing billions of dollars annually While patient symptoms and mortality have been reduced with the use of neurohormone receptor antagonists, such as β-blockers, their long-term impact on cardiac remodeling and patient lifespan is limited. G protein-coupled receptor (GPCR) signaling pathways have been shown for some time to be tunable, wherein protective pathways (often β-arrestin-dependent) may be selected for through biased ligands, while deleterious pathways (usually G protein-dependent) are simultaneously inhibited. However, there has been a disappointing lack of progress in the development of biased ligands for β- adrenergic receptors (βARs) in particular, which have been shown to relay cardioprotective outcomes via β- arrestin-dependent transactivation of epidermal growth factor receptor (EGFR). Over the last few years we have extensively characterized the subcellular signaling, transcriptional and cardiac remodeling responses that occur in response to both acute and chronic EGFR transactivation in cardiomyocytes and in whole heart following β1AR stimulation. Our studies have highlighted the potential therapeutic benefit of β1AR-mediated EGFR transactivation and provided rationale for its pursuit as a HF treatment strategy, however lack of specific tools has prevented the execution of definitive studies. In particular, bona fide β1AR-specific βarr/EGFR- biased ligands with enhanced efficacy, and that do not impact G protein-dependent signaling, do not exist. Thus, EGFR transactivation studies to date have relied upon EGFR inhibition in the presence of βAR stimulation to indirectly measure its impact on cardiomyocyte/cardiac signaling, transcription and survival outcomes. Additionally, EGFR knockout animals are not viable, thereby preventing assessment of EGFR function in adult models of HF without the use of pharmacologic inhibitors that introduce selectivity and toxicity issues. With these issues in mind, we have begun to test pepducins, small peptide fragments of the intracellular loops of GPCRs that allosterically induce biased signaling, specifically for the β1AR with the intent to drive EGR transactivation in the absence of G protein activation. These new molecular tools will allow positive characterization of β1AR-mediated EGFR transactivation in cardiomyocytes for the first time, without the reliance upon pharmacological inhibitors. Additionally, we have generated a cardiomyocyte-specific inducible EGFR deletion mouse model that will definitively establish the impact of EGFR signaling in the heart in response to βAR stimulation, as well as in the context of more broad forms of HF development. These studies will therefore provide the first direct assessment of the importance of cardiomyocyte-expressed EGFR during HF, as well as generate novel and potentially therapeutic agents with which to specifically engage β1AR-depedent EGFR transactivation to promote survival signaling and reduce detrimental structural and functional outcomes during HF.