Optical antisense tumor targeting in vivo with an improved fluorescent DNA duplex probe.
Optical antisense tumor targeting in vivo with an improved fluorescent DNA duplex probe.
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DOI：
10.1021/bc9000933
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发表时间：
2009-06
影响因子：
4.7
通讯作者：
Hnatowich DJ
中科院分区：
文献类型：
--
作者：
Liang M;Liu X;Cheng D;Nakamura K;Wang Y;Dou S;Liu G;Rusckowski M;Hnatowich DJ
用于mRNA的反义靶向的荧光缀合的DNA寡核苷酸具有改善肿瘤/正常组织比率的潜力，超过通过核反义成像可实现的比率。通过将Cy5.5发射体缀合至25聚体硫代磷酸酯（PS）反义主要DNA的3 '等同端并与较短的18聚体磷酸二酯（PO）互补次要DNA（cDNA）杂交，在其5'端具有黑洞抑制剂BHQ 3（即，PS DNA 25-Cy5.5/PO cDNA 18-BHQ 3），我们先前在小鼠中实现了反义光学成像作为该概念的证明。在优化过程中，我们现在已经评估了一小系列具有可变长度次要链的双链体的稳定性。从这些结果中，选择具有10聚体次要链的新研究抗mdr 1反义双链体（即，PS DNA 25-Cy5.5/PO cDNA 10-BHQ 3）。新的研究双链体在37 °C的血清环境中显示出稳定性，并且与KB-31（pgp±）相比，在KB-G2（pgp++）细胞中提供了显著增强的荧光，作为其mdr 1 mRNA靶标的反义解离的证据。将该双链体也给予KB-G2荷瘤小鼠，与先前使用的双链体相比，来自肿瘤大腿的荧光更明显，肿瘤与背景荧光的比率得到改善。总之，通过设计用于优化光学反义肿瘤靶向的双链体的方法，在细胞和肿瘤小鼠中的荧光信号都得到了改善。
Fluorescent conjugated DNA oligonucleotides for antisense targeting of mRNA has the potential of improving tumor/normal tissue ratios over that achievable by nuclear antisense imaging. By conjugating the Cy5.5 emitter to the 3′ equivalent end of a 25 mer phosphorothioate (PS) antisense major DNA and hybridizing with a shorter 18 mer phosphodiester (PO) complementary minor DNA (cDNA) with the Black Hole inhibitor BHQ3 on its 5′ end (i.e., PS DNA25-Cy5.5/PO cDNA18-BHQ3), we previously achieved antisense optical imaging in mice as a proof of this concept. In a process of optimization, we have now evaluated the stability of a small series of duplexes with variable-length minor strands. From these results, a new study anti-mdr1 antisense duplex was selected with a 10 mer minor strand (i.e., PS DNA25-Cy5.5/PO cDNA10-BHQ3). The new study duplex shows stability in serum environments at 37 °C and provides a dramatically enhanced fluorescence in KB-G2 (pgp++) cells when compared with KB-31 (pgp±) as evidence of antisense dissociation at its mdr1 mRNA target. The duplex was also administered to KB-G2 tumor bearing mice, and when compared to the duplex used previously, the fluorescence from the tumor thigh was more obvious and the tumor-to-background fluorescence ratio was improved. In conclusion, by a process designed to optimize the duplex for optical antisense tumor targeting, the fluorescence signal was improved both in cells and in tumored mice.