Defining physiological and pathophysiological roles of the Free Fatty Acid Receptor2 by analysis of novel transgenic mouse models

开发新药是一项长期的目标和挑战。最初的努力通常是基于对啮齿动物中激活或阻断特定蛋白质或信号转导途径的影响的观察和分析，这些蛋白质或信号转导途径被认为有助于或影响人类疾病的发展。做到这一点的方法包括消除小鼠中特定基因的表达或用合成药物样分子治疗动物，希望产生的反应将告知或模仿可能发生在人类中的反应。虽然许多重要的生理过程在小鼠和人类之间以相同的方式被控制和调节，但也有许多差异反映了9000万年的进化将物种分开。这方面的一个共同点是，来自不同物种的等价蛋白质之间通常存在序列差异，并且在许多情况下，已知这显著影响合成药物产生效果的能力。这是受体FFA 2的情况，其被存在于肠道中的细菌产生的短链脂肪酸激活。通过发现阻断人类形式的配体而鉴定的这种受体的拮抗剂根本不适用于啮齿动物直系同源物，因此，很难在小鼠中进行初步研究，以预测它们对人类的影响。为了克服这一点，我们已经产生了小鼠，其中我们用人类形式替换了小鼠形式的FFA 2，并且已经表明，在这些动物的组织中，鉴定的拮抗剂现在能够阻断FFA 2介导的作用。我们也在这方面更进了一步。由于在小鼠和人类中存在至少两种被短链脂肪酸激活的受体，我们进一步调整了人类FFA 2，使得其不被短链脂肪酸激活，而是被一组不激活FFA 2正常形式的合成分子激活。我们还产生了小鼠，其中表达了这种修饰的人类受体，而不是小鼠形式。有了这些基础知识，我们现在计划全面研究FFA 2在小鼠中控制的作用。我们将特别关注肠道中的作用，我们的初步研究表明，FFA 2是一种名为GLP-1的激素释放的关键调节剂，当餐后血糖水平升高时，GLP-1的释放在促进胰腺产生胰岛素方面起着核心作用。关于激活或阻断FFA 2是否可能是预防或治疗基于下肠道炎症的疾病（如溃疡性结肠炎）的有效方法，其他研究小组也有相互矛盾的报告。我们现在将能够在这种疾病和相关疾病的小鼠模型中解决这些问题，这是迄今为止不可能的。另一个存在争议和矛盾观点的关键领域是FFA 2的直接激活或阻断是否能够增强或限制人类，特别是II型糖尿病患者的胰岛素释放。有理由相信小鼠的控制机制与人类不同，现在，使用上述动物模型以及对小鼠和人类受体蛋白形式的详细分子研究，我们计划评估这个问题。这可能会影响制药行业的思维，即考虑作为治疗性治疗的最适当策略，以及在野生型小鼠中进行的研究是否实际上可能为人类治疗指出了错误的方向。

Development of new medicines is a long term ambition and challenge. Initial efforts are often based on observations and analysis of effects in rodents of either activating or blocking a specific protein or a signal transduction pathway that is believed to contribute to or influence the development of a disease in humans. Means to do this include elimination of expression of specific genes in mouse or treatment of animals with synthetic drug-like molecules with the hope that responses produced will inform or mimic those that might occur in humans. Although it is true that many important physiological process are controlled and regulated in equivalent ways between mice and humans there are also many differences that reflect the 90 million years of evolution separating the species. One common aspect of this is that there are often differences in sequence between the equivalent proteins from different species and in a number of cases this is known to markedly effect the ability of a synthetic drug to produce effects. This is the situation for the receptor FFA2 that is activated by short chain fatty acids produced by the bacteria that are present in the intestine. Antagonists of this receptor that were identified by finding ligands that block the human form simply don't work at rodent orthologues and, because of this, it has been difficult to perform preliminary studies in mice that might predict their effects in humans. To overcome this we have generated mice in which we replaced the mouse version of FFA2 with the human form, and have shown that in tissues from these animals the identified antagonists are now able to block FFA2-mediated effects. We have also taken this one step further. As there are at least two receptors in both mouse and human that are activated by short chain fatty acids we further adapted human FFA2 such that it NOT activated by short chain fatty acids but is activated instead by a set of synthetic molecules that don't activate the normal forms of FFA2. We have also generated mice in which this modified human receptor is expressed instead of the mouse form. With this underpinning knowledge we now plan to fully investigate the roles that are controlled by FFA2 in mouse. We will pay particular attention to actions in both the intestine where our preliminary studies have shown that FFA2 is a key regulator of release of a hormone called GLP-1 that after its release plays a central role in promoting insulin production from the pancreas when blood glucose levels rise after a meal. There have also been conflicting reports from other groups as to whether activating or blocking FFA2 might be an effective approach to either prevent the development of, or to treat, diseases such as ulcerative colitis that are based on inflammation of the lower intestine. We will now be able to address these questions in mouse models of this and related diseases, something that has hitherto been impossible. Another key area where there has been debate and contradictory views is whether direct activation or blockade of FFA2 would be able to enhance or limit release of insulin in humans, and specifically in type II diabetics. There are reasons to believe the control mechanisms in mouse are different from in humans and now, using both the animal models described above and detailed molecular studies on both mouse and human forms of the receptor protein we plan to assess this question. This is likely to influence thinking with the pharmaceutical industry as to the most appropriate strategy to consider as a therapeutic treatment and whether studies that have been performed in wild type mice might actually be pointing in the wrong direction for therapy in humans.