b1AR-mediated EGFR transactivation in the heart

 描述（由申请人提供）：心力衰竭（HF）是一种影响美国数百万人的进行性疾病，这些患者的护理每年花费数十亿美元。虽然使用神经激素受体拮抗剂（如β-阻滞剂）已减少患者症状和死亡率，但其对心脏重塑和患者寿命的长期影响有限。一段时间以来，G蛋白偶联受体（GPCR）信号传导途径已被证明是可调节的，其中保护性途径（通常是β-抑制蛋白依赖性的）可以通过偏向性配体来选择，而有害途径（通常是G蛋白依赖性的）同时被抑制。然而，特别是在β-肾上腺素能受体（β AR）的偏向性配体的开发方面缺乏令人失望的进展，其已被证明通过表皮生长因子受体（EGFR）的β-抑制蛋白依赖性反式激活来传递心脏保护结果。在过去的几年中，我们已经广泛表征了β1AR刺激后心肌细胞和整个心脏中急性和慢性EGFR反式激活引起的亚细胞信号传导、转录和心脏重塑反应。我们的研究强调了β 1AR介导的EGFR反式激活的潜在治疗获益，并为其作为HF治疗策略的追求提供了理论依据，但缺乏特定工具阻碍了确定性研究的执行。特别是，真正的β 1 AR特异性β 1 AR/EGFR偏向配体 具有增强的功效并且不影响G蛋白依赖性信号传导的药物不存在。因此，迄今为止，EGFR反式激活研究依赖于在βAR刺激存在下的EGFR抑制，以间接测量其对心肌细胞/心脏信号传导、转录和生存结局的影响。此外，EGFR敲除动物不能存活，因此无法在不使用引入选择性和毒性问题的药理学抑制剂的情况下评估成人HF模型中的EGFR功能。考虑到这些问题， 我们已经开始测试pepducins，GPCR细胞内环的小肽片段，其变构诱导偏向信号传导，特别是β 1 AR，目的是在G蛋白活化不存在的情况下驱动EGR反式活化。这些新的分子工具将首次允许心肌细胞中β 1AR介导的EGFR反式激活的阳性表征，而不依赖于药理学抑制剂。此外，我们还建立了一种心肌细胞特异性诱导型EGFR缺失小鼠模型，该模型将明确确定心脏中EGFR信号传导对βAR刺激的反应以及更广泛形式的HF发展的影响。因此，这些研究将首次直接评估心力衰竭期间心肌细胞表达的EGFR的重要性，并产生新型和潜在的治疗药物，用于特异性参与β 1AR依赖性EGFR反式激活，以促进存活信号传导并减少心力衰竭期间有害的结构和功能结局。

 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.