We have used combinatorial chemistry with amino acid mixtures (X) at positions 6 to 23 in vasoactive intestinal peptide (VIP) to optimize binding affinity and selectivity to the rat VPAC(1) receptor. The most efficient amino acid replacement was a substitution of alanine at position 18 to diphenylalanine (Dip), increasing the displacement efficiency of (125)I-VIP by 370-fold. The [Dip(18)]VIP(6-23) was subsequently used to find a second replacement, employing the same approach. Tyrosine at position 9 was selected and the resulting [Tyr(9),Dip(18)]VIP(6-23) analog has a K(i) value of 90 nM. This analog was unable to stimulate cAMP production at 10(-6) M but was able to inhibit VIP-induced cAMP stimulation (K(b) = 79 nM). The K(i) values of [Tyr(9),Dip(18)]VIP(6-23) using the rat VPAC(2) and PAC(1) receptors were 3,000 nM and >10,000 nM, respectively. Thus, [Tyr(9),Dip(18)]VIP(6-23) is a selective VPAC(1) receptor antagonist. The C-terminally extended form, [Tyr(9),Dip(18)]VIP(6-28), displays improved antagonistic properties having a K(i) and K(b) values of 18 nM and 16 nM, respectively. On the contrary, the fully extended form, [Tyr(9),Dip(18)]VIP(1-28), was a potent agonist with improved binding affinity (K(i) = 0.11 nM) and ability to stimulate cAMP (EC(50) = 0.23 nM) compared with VIP (K(i) = 1.7 nM, EC(50) = 1.12 nM). Furthermore, the specificity of this agonist to the VPAC(1) receptor was high, the K(i) values for the VPAC(2) and PAC(1) receptors were 53 nM and 3,100 nM, respectively. Seven other analogs with the [Tyr(9),Dip(18)] replacement combined with previously published VIP modifications have been synthesized and described in this work.

译文

我们已使用血管活性肠肽 (VIP) 中6至23位氨基酸混合物 (X) 的组合化学来优化对大鼠VPAC(1) 受体的结合亲和力和选择性。最有效的氨基酸置换是将第18位的丙氨酸替换为二苯丙氨酸 (Dip),将 (125)I-VIP的置换效率提高了370倍。随后使用 [Dip(18)]VIP(6-23) 来寻找第二种替代品,采用相同的方法。选择位置9的酪氨酸,所得 [Tyr(9),Dip(18)]VIP(6-23) 类似物的K(i) 值为90 nM。该类似物无法刺激10(-6) M的cAMP产生,但能够抑制VIP诱导的cAMP刺激 (K(b) = 79 nM)。使用大鼠VPAC(2) 和PAC(1) 受体的 [Tyr(9),Dip(18)]VIP(6-23) 的K(i) 值分别为3,000 nM和> 10,000 nM。因此,[Tyr(9),Dip(18)]VIP(6-23) 是选择性VPAC(1) 受体拮抗剂。C端扩展形式 [Tyr(9),Dip(18)]VIP(6-28) 显示出改善的拮抗特性,其K(i) 和K(b) 值分别为18 nM和16 nM。相反,完全扩展的形式,[Tyr(9),Dip(18)]VIP(1-28),与VIP (K(i) = 0.11纳米,EC(50) = 1.12纳米) 相比,是一种有效的激动剂,具有改善的结合亲和力 (K(i) = 纳米) 和刺激cAMP的能力 (EC(50) = 0.23纳米)。此外,该激动剂对VPAC(1) 受体的特异性高,VPAC(2) 和PAC(1) 受体的K(i) 值分别为53 nm和3,100 nM。在这项工作中,已经合成并描述了与先前发布的VIP修饰相结合的 [Tyr(9),Dip(18)] 替换的其他七个类似物。

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