Investigation of Mergo? mechanism of action by quantitative serum protein profiling.

Sixfold Bioscience是一家总部位于伦敦的生物技术中小企业，正在开发Mergo？，设计用于将寡核苷酸治疗剂携带至肝外组织的基于RNA的递送系统。梅尔戈号技术允许开发一个平台，可以迅速设计，以探索和筛选一个大的和多样化的化学空间。事实上，梅尔戈斯可以很容易地进行化学修饰，以改变其物理化学性质。Sixfold已经证明，化学成分改变的Mergos可以在体内分布到肝外组织（小鼠模型）。更有趣的是，某些梅尔戈？候选物显示出细胞向性，从而开辟了细胞特异性药物递送的途径。然而，Mergo？具体地分配到某些细胞还没有解决。目前主要的假说改变分布是基于蛋白质指纹。我们假设，修改的化学身份的合并？导致生物特性（蛋白结合剂）改变，导致分布、细胞摄取和治疗效果的差异。事实上，在注射到生物液体（例如血清或脑脊液）中时，蛋白质显示与Mergo？结合，形成蛋白质冠。而且，梅尔戈？具有不同的体内分布曲线，定性地显示出不同的蛋白结合（PB）曲线。例如，整个亲和力血清的变化，并形成了更高层次的结构（蛋白质Mergo？复合物）有不同的形状和大小。而下拉实验分析的SDS-PAGE显示，蛋白结合剂之间的不同梅尔戈？候选人，他们的身份尚未得到澄清。解决PB指纹将有助于Sixfold，以及更全球化的RNAi药物递送领域，将化学身份和生物学结果联系起来。更具体地说，细胞外蛋白结合物可用于了解体内分布，而细胞内PB可提供毒性和治疗活性信息。此外，了解蛋白质结合物在不同物种之间是否保守将有助于了解在小动物（小鼠）中观察到的结果是否可以转化为更大的物种（NHP，人类），加速临床前阶段和平台开发。最后，一个强大的，潜在的高通量的方法来确定蛋白质结合剂的发展可能有助于合理设计Mergo？候选人，并集成在六重AI/ML化学设计循环中。目前，选择的修改添加到合并？候选人是基于小分子的ADMET属性。我们相信PB可以用来更好地预测体内行为。总之，蛋白结合剂鉴定可用于了解作用机制（MOA），毒性，跨物种差异，并更好地告知下一代Mergo？候选人

Sixfold Bioscience, a London-based biotech SME, are developing Mergo?, RNA-based delivery systems designed to carry oligonucleotides therapeutics to extrahepatic tissues. The Mergo? technology allows the development of a platform that can rapidly be engineered to explore and screen a large and diverse chemical space. Indeed, Mergos can be easily chemically modified to change their physicochemical properties. Sixfold has shown that Mergos with altered chemical composition can distribute to extrahepatic tissues _in vivo_ (mice models). More interestingly, certain Mergo? candidates display cell tropism, opening the way to cell-specific drug delivery. Yet, the mechanism by which Mergo? distribute to certain cells specifically has not been addressed. The current leading hypothesis for altered distribution is based on protein fingerprinting.We postulate that modifying the chemical identities of Mergo? results in altered biological identity (protein binders), leading to differences in distribution, cellular uptake and therapeutic effect. Indeed, upon injection to biological fluids (_e.g_ serum or cerebral spinal fluid), proteins are shown to bind Mergo?, forming a protein corona. Moreover, Mergo? with different _in vivo_ distribution profiles have been qualitatively shown to display different protein binding (PB) profiles. For example, whole affinity to serum changed, and the higher-order structures formed (protein-Mergo? complexes) have different shapes and sizes.While pulldown experiments analyzed by SDS-PAGE have revealed that protein binders differ between Mergo? candidates, their identities have not been yet elucidated. Resolving PB fingerprints would help Sixfold, and more globally the RNAi drug delivery field, to link chemical identity and biological outcomes. More specifically, the extracellular protein binders can be used to understand _in vivo_ distribution, while intracellular PB might inform on toxicity and therapeutic activity. Additionally, understanding whether protein binders are conserved from one species to another would help understanding whether results observed in small animals (mouse) can be translated to bigger species (NHP, human), accelerating preclinical phase and platform development. Finally, the development of a robust, potentially high-throughput method to identify protein binders could help to rationally design Mergo? candidates, and be integrated within the Sixfold AI/ML chemical design loop. Currently, the choice of the modifications added to Mergo? candidates is based on ADMET properties of small molecules. We believe that PB can be used to better predict _in vivo_ behaviour. In summary, protein binders identification could be used to understand mechanism of action (MOA), toxicity, cross-species differences and inform better the design of next-generation Mergo? candidates.