Molecular characterisation of MC2R/MRAP antagonists for the treatment of ACTH excess in pituitary and neuroendocrine tumours.

用于治疗垂体和神经内分泌肿瘤中 ACTH 过量的 MC2R/MRAP 拮抗剂的分子特征。

• 批准号: 2442100
• 金额: --
• 依托单位: Queen Mary University of London
• 依托单位国家: 英国
• 项目类别: Studentship
• 财政年份: 2020
• 资助国家: 英国
• 起止时间: 2020 至 无数据
• 项目状态: 已结题
• 来源: https://gtr.ukri.org/projects?ref=studentship-2442100
• 关键词: Molecular; characterisation; MC2R; MRAP; antagonists

Pituitary and neuroendocrine tumours (NETs) are known to secrete adrenocorticotropic hormone (ACTH), which acts on the adrenal gland to stimulate glucocorticoid (GC) production. GC excess worsens prognosis (1). Identifying and removal of the source of this excessive plasma ACTH is preferred but not always possible (1,2). Hence, the ability to effectively block the effects ACTH has important clinical utility. ACTH acts on its specific receptor, the melanocortin-2-receptor (MC2R) complexed to the MC2R accessory protein (MRAP). The project is to develop and characterise small molecule compounds that can specifically inhibit the MC2R/MRAP complex for the treatment of pituitary tumours and NETs. There is growing awareness of a need to develop ACTH antagonists to treat ACTH excess (3). The MC2R/MRAP complex is unique responding specifically to ACTH (4,5). We previously identified small molecules that specifically inhibited ACTH stimulated cAMP generation in HEK293-cells co-expressing MC2R/MRAP (3). The overarching aim is to characterize MC2R/MRAP antagonists at a molecular and physiological level. , starting with our own lead compounds (788/0371/9018/6741) with additional compounds as they are identified by our industrial partner. Aim 1. Characterization of MC2R/MRAP antagonists. The student will characterise the antagonist effect on MC2R/MRAP biologyinteraction, binding, signalling and receptor internalisation in real-time.. Techniques include: NanoBRET binding assays, cAMP accumulation assays using luciferase biosensors, p-Glo, or Ca2+ mobilization using the Gcamp6 fluorescent biosensor, beta-arrestin recruitment and G-protein recruitment using nanoBit complementation. With our industrial partners,At OMass they will experience drug modelling and discovery. Aim 2. Effects of MC2R/MRAP antagonists on ACTH stimulated steroid production. Currently, there are no ACTH responsive, glucocorticoid producing adrenal cell-lines available representative of typical adrenocortical cells. Therefore, the student will use murine and human primary adrenal cell cultures to study GC and steroid production with/without MC2R/MRAP antagonists. , by ELISA and mass-spectrometry. Aim 3: Determine effectiveness of MC2R/MRAP antagonists in animal models of ACTH excess. The student will measure GC/steroid production after administering MC2R/MRAP antagonists IP/IV/PO in our murine model of ACTH excess where ACTH is delivered via an osmotic pump. The student will be trained in state-of-the-art pharmacology/receptor signalling techniques and acquire skills working with primary cell cultures and the latest murine models. The supervising team are experts in their field and have a team of technicians, post-docs and research co-ordinators to support the student. Together with an industrial placement at one of the UKs most exciting new biotech companies, they will gain technical and transferable skills sought after in both academia and industry.

已知垂体瘤和神经内分泌肿瘤（NET）分泌促肾上腺皮质激素（ACTH），其作用于肾上腺以刺激糖皮质激素（GC）的产生。GC过量影响预后（1）。最好能识别并清除过量血浆ACTH的来源，但并非总是可行（1，2）。因此，有效阻断ACTH作用的能力具有重要的临床实用性。ACTH作用于其特异性受体，即与MC 2 R辅助蛋白（MRAP）复合的黑皮质素-2受体（MC 2 R）。该项目旨在开发和表征可以特异性抑制MC 2 R/MRAP复合物的小分子化合物，用于治疗垂体瘤和NET。人们越来越意识到需要开发促肾上腺皮质激素拮抗剂来治疗促肾上腺皮质激素过多（3）。MC 2 R/MRAP复合物是独特的，对ACTH有特异性反应（4，5）。我们先前鉴定了在共表达MC 2 R/MRAP的HEK 293细胞中特异性抑制ACTH刺激的cAMP生成的小分子（3）。总体目标是在分子和生理水平上表征MC 2 R/MRAP拮抗剂。，从我们自己的先导化合物（788/0371/9018/6741）开始，以及我们的工业合作伙伴确定的其他化合物。目标1。MC 2 R/MRAP拮抗剂的表征。学生将演示拮抗剂对MC 2 R/MRAP生物学相互作用，结合，信号传导和受体内化的实时影响。技术包括：NanoBRET结合测定、使用荧光素酶生物传感器的cAMP积累测定、使用Gcamp 6荧光生物传感器的p-Glo或Ca 2+动员、使用nanoBit互补的β-抑制蛋白募集和G-蛋白募集。与我们的工业合作伙伴一起，在OMass，他们将体验药物建模和发现。目标2. MC 2 R/MRAP拮抗剂对ACTH刺激的类固醇产生的影响。目前，还没有ACTH反应性的、产生糖皮质激素的肾上腺细胞系可用于代表典型的肾上腺皮质细胞。因此，学生将使用鼠和人的原代肾上腺细胞培养物来研究GC和类固醇的产生，有/没有MC 2 R/MRAP拮抗剂。用ELISA和质谱法检测。目的3：确定MC 2 R/MRAP拮抗剂在ACTH过量动物模型中的有效性。学生将在我们的小鼠ACTH过量模型中测量MC 2 R/MRAP拮抗剂IP/IV/PO给药后GC/类固醇的产生，其中ACTH通过渗透泵输送。学生将接受最先进的药理学/受体信号传导技术的培训，并获得与原代细胞培养和最新鼠模型合作的技能。监督团队是各自领域的专家，并有一个技术人员，博士后和研究协调员团队来支持学生。再加上在英国最令人兴奋的新生物技术公司之一的工业实习，他们将获得学术界和工业界所追求的技术和可转移技能。