Targeting the M4 muscarinic acetylcholine receptor in neurodegenerative disease

本项目将结合化学遗传学、转基因小鼠模型、神经退行性疾病模型和独特的化学探针来确定选择性靶向M4毒蕈碱乙酰胆碱受体（M4 mAChRs）在神经退行性疾病中的影响。在与礼来公司的合作中，我们开发了一种小鼠神经退行性变（小鼠朊病毒疾病）模型，在该模型中，我们可以直接探测在疾病背景下靶向毒蕈碱受体的影响。这些朊病毒感染的小鼠表现出错误折叠的朊病毒蛋白（PrPsc）的积累、星形胶质细胞增生和突触功能障碍，并经历进行性神经元丢失，最终达到终末期临床疾病。此外，朊病毒感染小鼠表现出明显的胆碱能功能障碍，并伴有相关的记忆缺陷。这种记忆缺陷可以通过xanomeline治疗完全恢复，xanomeline是一种经临床验证的M1/M4毒蕈碱激动剂，也可以通过高选择性M1 mAChR阳性变构调节剂（PAMs）治疗。因此，我们知道M1 mAChR可以对朊病毒神经退行性变产生影响-我们不知道的是M4 mAChR在神经退行性变中的重要性。该博士课程将使用礼来公司提供的独特化学工具，以及化学遗传动物模型，探索M4 mAChR在神经变性中的作用。具体来说，我们将：采用一种新的化学遗传小鼠模型，其中编码M4 mAChR的野生型基因位点被表达该受体的突变形式的编码序列所取代，该受体不能被内源性配体乙酰胆碱激活，而只能被合成配体氯氮平- n -氧化物（CNO）激活。这种突变受体被称为设计药物独家激活的设计受体（DREADD），它允许我们特异性地探测M4 machr的生理功能，因为我们可以监测c4突变受体受到CNO刺激后的行为反应。利用这种小鼠模型，我们可以诱导朊病毒神经退行性变，并研究选择性的M4-mAChR刺激与正畸受体激动剂对神经退行性设置认知能力下降的影响。上述化学遗传方法的另一种替代方法是利用我们无与伦比的途径，通过礼来公司获得一系列新的M4 mAChR配体（包括orthosteric和allosteric），来测试以这种受体为靶点的药理学可能挽救朊病毒感染小鼠学习和记忆受损的可能性。除了研究对学习和记忆缺陷的影响外，我们还将测试新型M4 mAChR配体对神经退行性疾病进展的影响，特别是探索新型毒蕈碱配体对朊病毒患病小鼠存活的影响。在疾病的其他特征中，我们将探讨M4 mAChR激活对突触功能障碍、神经元丢失和错误折叠蛋白聚集的影响

Studentship strategic priority area:Basic and Clinical ResearchKeyword:Alzheimer's disease, neurodegeneration, muscarinic, GPCRThis studentship will combine chemical genetics, transgenic mouse models, models of neurodegenerative disease and unique chemical probes to define the impact of selectively targeting M4 muscarinic acetylcholine receptors (M4 mAChRs) in neurodegenerative disease. In collaboration with Eli Lilly, we have developed a mouse model of neurodegeneration (murine prion disease) in which we can directly probe the impact of targeting muscarinic receptors in a disease context. These prion-infected mice show accumulation of misfolded prion protein (PrPsc), astrogliosis and synaptic dysfunction and undergo progressive neuronal loss which ultimately reaches end-stage clinical disease. Furthermore, prion-diseased mice display significant cholinergic dysfunction with associated memory deficits. This memory-deficit can completely restored by treatment with xanomeline, an M1/M4 clinically validated muscarinic agonist, and also by highly selective M1 mAChR positive allosteric modulators (PAMs). We know therefore that M1 mAChRs can have an impact on prion neurodegeneration - what we do not know is how important the M4 mAChR is in neurodegeneration. This PhD programme will use unique chemical tools made available through Eli Lilly, together with chemical genetic animal models, to probe the role of the M4 mAChR in neurodegeneration. Specifically we will:. Employ a novel chemical genetic mouse model in which the wild type gene locus encoding the M4 mAChR has been replaced by a coding sequence that expresses a mutant form of this receptor that is unable to be activated by the endogenous ligand, acetylcholine, but only by the synthetic ligand, clozapine-N-oxide (CNO). This mutant receptor, called Designer Receptor Exclusively Activated by Designer Drug (DREADD), allows us to specifically probe the physiological function of M4 mAChRs since we can monitor the behavioural response following stimulation of the M4-mutant receptors with CNO. Using this mouse model we can induce prion-neurodegeneration and investigate the effects of selective M4-mAChR stimulation with an orthosteric agonist on cognitive decline in a neurodegenerative setting.. An alternative to the chemical genetic approach above is to employ our unparalleled access to an array of novel M4 mAChR ligands (including orthosteric and allosteric), through Eli Lilly, to test the possibility that pharmacological targeting this receptor may rescue impairments in learning and memory in prion-infected mice.. In addition to investigating effects on learning and memory deficits, we will test of the impact of novel M4 mAChR ligands on the progression of neurodegenerative disease, particularly to explore the impact of novel muscarinic ligands on the survival of prion-diseased mice. Amongst other hallmarks of disease, we will probe the effects of M4 mAChR activation on synaptic dysfunction, neuronal loss and aggregation of misfolded proteins