Design and Synthesis of a Short-Chain Bitistatin Analogue for Imaging Thrombi and Emboli
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- 作者：Kwamena E. Baidoo ; Linda C. Knight ; Kuo-Shyan Lin ; Jerome L. Gabriel ; and Jan E. Romano
- 刊名：Bioconjugate Chemistry
- 出版年：2004
- 出版时间：September 2004
- 年：2004
- 卷：15
- 期：5
- 页码：1068 - 1075
- 全文大小：185K
- 年卷期：v.15,no.5(September 2004)
- ISSN：1520-4812
文摘
Previously, we showed that labeled bitistatin analogues possessed excellent characteristics for imagingboth deep-vein thrombosis and pulmonary embolism. We hypothesized that the N-terminal aminoacid sequence of bitistatin, which is different from other disintegrins, likely interacts with the bindingsite of platelets to confer desirable properties to bitistatin for imaging. In this study, we present thedesign, synthesis, and initial biological testing of a short-chain analogue of the native 83-amino-acidbitistatin sequence. Our initial molecular modeling of the binding loop of bitistatin showed that theminimal sequence that represented the binding region was a cyclic 10 amino acid sequence cyclo[Cys-Arg-Ile-Ala-Arg-Gly-Asp-Trp-Asn-Cys(S)]. Systematic modeling of a truncated N-terminalsequence of bitistatin fused with the optimized binding region having a thioether sequence througha Gaba spacer ultimately yielded the 24-amino acid peptide, cyclo-[CH2CO-Arg-Ile-Ala-Arg-Gly-Asp-Trp-Asn-Cys(S-)]-Gaba-Gly-Asn-Glu-Ile-Leu-Glu-Gln-Gly-Glu-Asp-Ser-Asp-Ser-Lys-OH, 1. The peptide was then coupled to the hydrazino-nicotinic acid bifunctional chelating agentand the purified adduct labeled with 99mTc using tricine as a coligand. Binding of the unlabeled andlabeled peptide to stimulated human platelets was assayed in vitro. The 99mTc labeling yield was >90%. The in vitro binding assays showed that the IC50 for inhibition of platelet aggregation was 3694nM, while the Kd of the 99mTc labeled peptide was 185 nM, indicating moderate affinity for the receptor.The 99mTc-labeled peptide was able to identify sites of experimental thrombi and emboli in a caninemodel. The results suggest initial success in attempting to mimic the behavior of bitistatin for imagingthrombi and emboli.