Design of highly immunogenic immunogens for development of monoclonal antibodies to selfpeptides: a combined simulation and experimental study

单克隆抗体（mAb）疗法是一种免疫疗法，它使用高度特异性的抗体与特定的细胞或蛋白质结合，以刺激免疫系统破坏这些细胞。因此，它可以用于靶向癌症。该过程涉及将来自肿瘤的小组分（肽）结合到纳米颗粒上。用这些肽包被的纳米颗粒免疫导致刺激脾细胞产生一系列抗体，这些抗体具有破坏生长中的肿瘤的潜力。使用杂交瘤技术，可分离和克隆产生仅针对一个靶标的高度特异性抗体（mAb）的单独分离和克隆的脾细胞，并在工业规模上生长以用于mAb治疗剂的开发。然而，需要进行大量的研究，不仅要找到正确的肽，而且要确保它们以正确的方向结合到纳米颗粒上，以便免疫系统随后将“自身肽”识别为靶标。（促性腺激素释放激素，GnRH），我们以前已经研究过使用模拟和实验，在那里，我们研究了如何最好地将GnRH结合到二氧化硅纳米颗粒（SiNPs）上。我们模拟的吸附GnRH（本地）和一个修改的类似物，是半胱氨酸标记（cys-GnRH）的二氧化硅，并发现，本地肽吸附通过其精氨酸残基离开其N（氨基）和C（羧基）末端松散结合，并可用于进一步的相互作用。相反，吸附的cys-GnRH仅将其N-末端暴露于溶液中。我们还模拟了cys-GnRH与BSA的共价结合（在碳二亚胺化学反应后进行交联），发现这导致更好的GnRH表位呈递到溶液中。还用吸附到SiNP上的肽和BSA-肽-SiNP系统进行了一系列免疫学研究。我们发现GnRH-SiNP由于其两个末端残基的可用性而诱导药物作用，而cys-GnRH-SiNP则没有。我们还发现BSA缀合物系统有效地诱导抗体产生。这一范围内的免疫和激素反应解释了模拟结果和介绍的肽的解决方案。因此，这项工作不仅为在分子水平上深入了解治疗作用铺平了道路，更重要的是，为在分子模拟指导下合理设计药物输送和疫苗系统铺平了道路;我们将在本项目中利用这种新策略。为了证明该技术的实用性，我们将在项目中使用癌症应用。建模将用于选择最佳候选物，并进行修改以结合到SiNP上，并设计最佳呈现的条件。设计的材料将通过实验进行部署，随后将评估抗体和细胞因子的肿瘤细胞杀伤潜力。最好的候选人将被用于单克隆抗体治疗。该项目将汇集来自斯特拉斯克莱德大学和TAC的多学科监督团队。学生的监督将来自化学和过程工程系，药学和生物医学科学的斯特拉斯克莱德研究所，和ARCHIE-WEST超级计算机中心。我们还将与格拉斯哥喀里多尼亚大学和格拉斯哥大学密切合作，他们将提供患者的临床样本，并提供患者信息。工业主管为Eric瓦格纳（TAC）。里程碑1：肽修饰和计算建模（第1-12个月; Mulheran博士，Kubiak-Ossowska实验室）里程碑2：免疫原制备和免疫增强评价（第13-18个月; Ferro labs博士，TAC）里程碑3：癌症研究（第19-36个月; Mulheran、Ferro博士与威廉姆斯和苏特实验室合作）里程碑4：传播和撰写（第43-48个月）;包括知识产权保护后的出版物。

Monoclonal antibody (mAb) therapy is an immunotherapy that uses highly specific antibodies to bind to particular cells or proteins, in order to stimulate the immune system to destroy those cells. It can therefore be used to target cancers. The process involves binding small components (peptides) derived from tumours to nanoparticles. Immunisation with these peptide-coated nanoparticles results in stimulation of spleen cells to produce a range of antibodies that have the potential to destroy growing tumours. Using hybridoma technology, individually isolated and cloned spleen cells that produce highly specific antibodies to only one target (mAbs) can be isolated and grown on an industrial scale for the development of mAb therapeutics. However, a lot of research is required to not only find the right peptides, but to then ensure they bind to the nanoparticle with the correct orientation so that the immune system subsequently identifies the "self-peptides" as targets.We will initially use a model self-peptide (gonadotrophin releasing hormone, GnRH), that we have previously studied using simulation and experiment, where we investigated how best to conjugate GnRH to silica nanoparticles (SiNPs). We simulated the adsorption of GnRH(native) and a modified analogue that was cysteine-tagged (cys-GnRH) to silica, and found that the native peptide adsorbed via its arginine residue to leave its N (amino) and C (carboxyl) termini loosely bound and available for further interactions. In contrast, the adsorbed cys-GnRH exposed only its N-terminus to solution. We also simulated cys-GnRH covalently conjugated to BSA (following a carbodiimide chemical reaction to enable crosslinking), finding that this led to better GnRH epitope presentation to solution. A series of immunological studies was also conducted with the peptides adsorbed to SiNPs and with the BSA-peptide-SiNP systems. We found that the GnRH-SiNP induced a drug effect due to its availability of both terminal residues, whereas the cys-GnRH-SiNP did not. We also found that the BSA conjugate systems effectively induced antibody production. This range of immunological and hormonal response is explained by the simulation results and the presentation of the peptides to solution. Hence, this work paves the way for not only a molecular scale insight into therapeutic action, but more significantly, for a rational design of drug delivery and vaccine systems guided by molecular simulation; it is this new strategy that we will exploit in this project.To demonstrate the utility of the technology, we will use a cancer application in the project. Modelling will be used to select the best candidates with modification to bind onto SiNPs and design conditions for optimum presentation. The designed materials will be deployed experimentally, and the subsequent antibodies and cytokines will be evaluated for tumour cell killing potential. The best candidate(s) will be taken forward into a mAb therapeutic(s).The project will bring together a multi-disciplinary supervisory team from the University of Strathclyde and TAC. Supervision of the student will come from the Department of Chemical and Process Engineering, the Strathclyde Institute of Pharmacy and Biomedical Sciences, and the ARCHIE-WeSt supercomputer centre. We will also work in close collaboration with Glasgow Caledonian University and University of Glasgow who will supply clinical samples from patients and also provide access to patient information. The industrial supervisor will be Eric Wagner (TAC).Milestone 1: Peptide Modification and Computational Modelling (Months 1-12; Drs Mulheran, Kubiak-Ossowska labs)Milestone 2: Immunogen Preparation and Immune Enhancement Evaluation (Months 13-18; Dr Ferro labs, TAC)Milestone 3: Cancer studies (Months 19-36; Drs Mulheran, Ferro in collaboration with Williams and Souter labs)Milestone 4: Dissemination and Write-up (Months 43-48); including publications once IP has been protected.