Biological and Medicinal Chemistry Approaches to Human Beta Cell Regeneration

1型和2型糖尿病(T1D和T2D)都是由于正常功能的β细胞数量不足造成的。抑制激酶DYRK1a的小分子药物，如骆驼蓬碱等，可以重复地诱导成年人类β细胞复制，但复制速度很低(~2%/天)。最近，我们已经证明，在转化生长因子-β超家族抑制剂(转化生长因子βI‘s)或GLP1受体激动剂(如GLP1或exendin-4)中加入任何小分子DYRK1a抑制剂都能显著增强这种复制诱导，平均速度为5-8%/天。这不仅被Ki67和BrdU等复制标记在体外和体内证明了这一点，而且还通过增加β细胞的实际数量来证明这一点。不幸的是，这些药物可能不只影响β细胞，还可能产生“偏离靶点”的效果。因此，糖尿病研究人员有有效的再生药物可以输送到人类的贝塔细胞，但可能需要一个“靶向分子”将它们带到贝塔细胞。因此，在过去的几年里，糖尿病研究人员已经确定了两类“原型β细胞靶向分子”：一种是针对β细胞表面分子的单抗，称为“ENTPD3”；另一类是GLP1受体类分子。虽然这些分子对于β细胞靶向来说可能是完美的，也可能不是完美的，但在该领域试图识别更完美的β细胞靶向分子时，它们无疑是可以与之合作的“原型”靶向分子。我们还合成了许多DYRK1A抑制剂和TGFbeta抑制剂以及可切割的化学连接物，使它们能够与任何潜在的β细胞靶向分子结合。因此，本申请的目的是：1.合成带有化学接头的转化生长因子-β抑制剂，以补充我们新型的DYRK1A抑制剂接头化合物，用于与原型靶向递送载体的结合。2.将三尖杉酯-Linker和转化生长因子-β-抑制剂-Linker化合物偶联到两个靶向分子：GLP1受体激动剂和ENTPD3单抗。3.人胰岛移植模型中三尖杉酯碱-Linker和转化生长因子-β抑制剂-Linker结合物的长期有效性、特异性和安全性的确定这些目标既是可实现的，也是对NIDDK目标的直接回应。如果GLP1受体激动剂和/或ENTPD3单抗被证明不适合于β细胞靶向，这里开发的方法和分子可以很容易地扩展和适应任何未来更具特异性的人类β细胞靶向分子。

Both Type 1 and Type 2 diabetes (T1D and T2D) result from inadequate numbers of normally functioning beta cells. Small molecule drugs that inhibit the kinase, DYRK1A, such as harmine and others, are reproducibly able to induce adult human beta cells to replicate, but at low rates (~2%/day). More recently, we have shown that adding any small molecule DYRK1A inhibitor to either a TGF-beta superfamily inhibitor (TGFβI's) or to a GLP1 receptor agonist such as GLP1 or exendin-4 markedly enhances this replicative induction to rates averaging 5-8%/day. This has been documented not only by “markers” of replication such as Ki67 and BrdU, both in vitro and in vivo, but also by enhancing actual numbers of beta cells. Unfortunately, these drugs may not exclusively affect the beta cell, but instead may have “off target” effects as well. Thus, diabetes researchers have effective regenerative drugs to deliver to the human beta cell, but may require a “targeting molecule” to bring them to the beta cell. Accordingly, over the past few years, diabetes researchers have identified two classes of “prototype beta cell targeting molecules”: a monoclonal antibody raised against the beta cell surface molecule called “ENTPD3”; and, the GLP1-receptor class of molecules. While these molecules may or may not be perfect for beta cell targeting, they are unquestionably “prototype” targeting molecules with which to work while the field attempts to identify more perfect beta cell targeting molecules. We have also synthesized numerous DYRK1A inhibitors and TGFbeta inhibitors and cleavable chemical linkers that enable their conjugation to any potential beta cell targeting molecule. Thus, the Aims of this application are: 1. Synthesis of TGF-beta Inhibitors With Chemical Linkers, to Complement our Novel DYRK1A inhibitor Linker Compounds, For Conjugation To Prototype Targeted Delivery Vehicles. 2. Conjugation of Harmine-Linker and TGF-beta-Inhibitor-Linker Compounds to Two Prototype Targeting Molecules: GLP1 Receptor Agonists and ENTPD3 Monoclonal Antibodies. 3. Definition of Long Term Efficacy, Specificity and Safety of the Harmine-Linker and TGF-beta Inhibitor- Linker Conjugates in vivo in Human Islet Engraftment Models. These goals are both achievable and directly responsive to the goals of aims of the NIDDK. If GLP1 receptor agonists and/or ENTPD3 MAbs prove suboptimal for beta cell targeting, the approaches and molecules developed here can readily be extended and adapted to any future more specific human beta cell targeting molecule.