Identification of Relaxin Receptor Antagonists

描述(申请人提供)：松弛素是一种异二聚体多肽激素，由怀孕和非怀孕女性的黄体、蜕膜和胎盘产生，在怀孕期间达到最高的血浆水平。除了在抑制子宫收缩活动中的作用外，松弛素还被证明可以诱导胶原重塑，从而导致产道组织(宫颈和阴道)的软化，乳腺的生长和分化，以及血管的扩张。因此，松弛素在妊娠期生理适应的整体调节中起着重要作用。根据一级和二级结构特征，松弛素与胰岛素和胰岛素样生长因子(IGFS)被归类为结构同源物。与胰岛素类似，松弛素的前体形式前松弛素具有类似于胰岛素和胰岛素样生长因子前体的结构域排列；成熟的松弛素似乎被转换酶处理，从前松弛素生成双链异二聚体。在人类中，除了两个几乎相同的松弛蛋白基因外，还有另外五个松弛蛋白家族基因，包括编码INSL3/RLF、INSIA/Epil、INSL5/Rif2、INSL6/Rif1和relasin3的基因。在胎儿期被动免疫抗松弛蛋白抗体会降低啮齿类动物的宫颈生长和延伸性，并扰乱其生育。此外，许多研究表明，血清松弛素是人类怀孕多个阶段早产风险的独立预测因子。这些早期的研究提出了这样的观点，即通过受体拮抗剂或免疫降低松弛素活性，与针对其他信号通路的传统方法相结合，对于早产和分娩的治疗可能是重要的。我们最近的研究证实，松弛素能激活两个孤立的G蛋白偶联受体LGR7和LGR8(Hsu等人，2002年)。科学295：671-4)以及靶细胞中cAMP依赖的途径，而密切相关的松弛蛋白3和INSL3分别是LGR7和LGR8的选择性激动剂。为了利用松弛蛋白信号转导方面的这些新发现，以利于生殖健康研究，我们建议筛选保持受体结合活性而不具有受体激活活性的突变型重组松弛蛋白拮抗剂。一旦获得了仅具有受体结合活性的突变多肽，将在共处理试验中测试这些突变多肽的拮抗作用，以确定这些突变多肽是否能够拮抗松弛素诱导的LGR7和LGR8表达细胞中cAMP的产生，并延缓体内妊娠大鼠的分娩。这些拮抗剂可用于阻断早产时的松弛作用，并为进一步研究LGR7和LGR8在生殖道生理中的作用提供工具。

DESCRIPTION (provided by applicant): Relaxin is a heterodimeric peptide hormone produced by the corpus luteum, decidua, and placenta in pregnant and nonpregnant females attaining the highest plasma levels during pregnancy. Other than its role in the inhibition of uterine contractile activity, relaxin has been shown to effect the induction of collagen remodeling and the consequent softening of the tissues of the birth canal (cervix and vagina), growth and differentiation of the mammary gland, and dilation of the blood vessels. Thus, relaxin plays a significant role in the overall regulation of physiological adaptation during pregnancy. Based on: primary and secondary structural characteristics, relaxin was grouped with insulin and insulin-like growth factors (IGFs) as a structural homolog. Analogous to insulin, prorelaxin, the precursor form of relaxin, has a domain arrangement similar to insulin and IGF precursors; mature relaxin appears to be processed by convertases to generate a two-chain heterodimer from the prorelaxin. In addition to two almost identical relaxin genes in human, there are five additional relaxin family genes including those encoding INSL3/RLF, INSIA/EPIL, INSL5/RIF2, INSL6/RIF1, and relaxin3. Passive immunization with anti-relaxin antibodies during the anteparmm period reduces cervical growth and extensibility as well as disrupts birth in rodents. In addition, numerous studies have shown that serum relaxin is an independent predictor of the risk of preterm delivery at multiple stages of pregnancy in humans. These earlier studies have propelled the proposition that a reduction of relaxin activity through receptor antagonists or immunization could be important for the treatment of preterm labor and birth in conjunction with traditional methods targeting other signaling pathways. Our recent studies have established that relaxin activates two orphan G protein-coupled receptors, LGR7 and LGR8 (Hsu et al., 2002. Science 295:671-4) as well as the cAMP-dependent pathway in target cells, whereas the closely related relaxin3 and INSL3 are selective agonists for LGR7 and LGR8, respectively. To take advantage of these new findings in relaxin signaling for the benefit of reproductive health research, we propose to screen for mutant recombinant relaxin antagonists that retain receptor-binding activity while being devoid of receptor-activation activity. Once mutant peptides with only the receptor binding activity are obtained, the antagonistic effect of these peptides will be tested in co-treatment assays to determine whether the mutant peptides are capable of antagonizing the relaxin-induced cAMP production in LGR7- and LGR8- expressing cells as well as delaying parturition in pregnant rats in vivo. These antagonists could be used to block relaxin action during preterm labor as well as provide a tool for further research on the role of LGR7 and LGR8 in reproductive tract physiology.