CFTR：cystic fibrosis transmembrane conductance regulator protein

mR-509-3p：表达CFTR的mRNA

miR-509-3p：一种可以调节mR-509-3p的miRNA

PNA：肽核酸

囊胞性纤维症是一种常见的基因突变导致的疾病，每3000个高加索人中就有一个患有此病。这个疾病使由于基因突变导致cystic fibrosis transmembrane conductance regulator (CFTR) protein 的表达发生变化，比如蛋白结构发生改变或者表达量减少，致使细胞某些功能功能丧失或者减弱。有不止一种突变类型的突变，CFTR蛋白还保留部分功能，因此，通过提高CFTR蛋白的表达量可以治疗Cystic Fibrosis。 研究表明，细胞中存在miRNA，会调节CFTR的mRNA（miR-509-3p ）的翻译，导致CFTR蛋白表达量减少。因此，可以通过减少miR-509-3p 对mR-509-3p 的影响来提高CFTR的表达量。之前的研究是直接设计一条跟miR-509-3p互补的PNA，通过PNA可以竞争性的与miR-509-3p的结合，从而抑制miR-509-3p对mR-509-3p的抑制作用。但研究表明，虽然针对miRNA的 PNA可以减少miRNA对相应mRNA的影响，但是同时也会影响很多其它的mRNA。这是因为一条miRNA可以调节几百甚至几千种mRNA 。

鉴于此，我们设计了另外一种方法来减少miR-509-3p对mR-509-3p的调节作用。 设计与mR-509-3p 的3′UTR 互补的PNA。 PNA与3′UTR 结合后，位点被保护，miR-509-3p就不会对mR-509-3p 有调节作用，从而miR-509-3p对mR-509-3p的抑制作用就减弱，CFTR蛋白的表达量就会得到明显提高，患者的症状得到缓解。

实验结果： 设计的13bp 的PNA 能显著的提高CFTR蛋白的表达量（增加70%）。PNA能与相应的mRNA位点牢固结合，形成heteroduplexes ，从而保护了mRNA，不会被miRNA识别并结合。

Molecules 2017, 22(7), 1144; doi:10.3390/molecules22071144

原文要点：

In this paper, we have proposed for the first time the use of PNAs as miRNA target protectors to increase the expression of CFTR in CF. The negatively charged PNAs 1 and 2, conveniently modified at their C-ends and fully complementary to the 3’UTR region of the CFTR mRNA recognized by the seed region of miR-509-3p, were synthesized and characterized. To demonstrate the sequence dependent activity of 1 and 2, two other PNAs , containing scrambled sequences of 1 and 2, respectively, were designed and synthesized. Spectroscopic data confirmed the ability of 1 and 2 to bind their complementary ODN target by forming stable PNA/ODN heteroduplexes. The structural features of 1/RNA and 2/RNA heteroduplexes were also determined through molecular dynamics simulations. The results indicated that the presence of the negatively charged tetra peptide at the C-end of 1 and 2 slightly affected the structural features of the resulting PNA/RNA heteroduplexes (with respect to the experimentally determined PNA/RNA heteroduplexes). Biological studies show that the PNA molecules are suitable to counteract the action of miR-509-3p (in this case) as miRNA target protectors. These data confirm, once again, that a mRNA-targeted approach of a gene to increase the expression of the protein could be a good therapeutic strategy (that, of course, could be also used in other monogenic diseases). Moreover, the PNAs, showing a high affinity towards nucleic acid targets and forming stable heteroduplex complexes, represent excellent candidates to be used for this approach. Furthermore, this type of approach has the advantage of being relatively mutation-independent. For this reason, it would be sufficient to increase the expression of a protein, even if mutated, to exceed the threshold of minimal activity needed to obtain an optimal clinical phenotype.