Identification of microRNA that correctly inhibits KRAS-related downstream signal transduction in KRAS mutant colorectal cancer

抗EGFR单抗对KRAS野生型转移性结直肠癌有效。然而，KRAS突变型mCRC对抗EGFR抗体表现出耐药性。为了鉴定抑制KRAS突变的crc的miRNAs，我们首先将KRASG12V基因导入HEK293和MRC5中。在miRNA芯片分析后，我们使用SRE和AP1荧光素酶报告分析来寻找哪个候选miRNAs会抑制两个主要的KRAS相关信号转导通路。我们发现了2个miRNAs。MIR-4689通过直接与KRAS的3UTR和Akt1的编码序列结合，不仅调控RAF/MEK/ERK通路，还调控PI3K/Akt通路。Akt1的下调导致促凋亡因子的顺序诱导和抗凋亡分子的抑制。与miR-NC相比，全身应用含有超碳酸盐磷灰石纳米颗粒的miR-4689显著抑制了预先建立的DLD1移植瘤的生长，并证明了KRAS和Akt1的下调。

The antiEGFR monoclonal antibodies are effective in patients with metastatic CRC with KRAS wild type. However, KRAS mutant mCRC shows resistance to the antiEGFR antibodies. To identify miRNAs that inhibit KRAS mutant CRCs, we first introduced KRASG12V cDNA into HEK293 and MRC5. After miRNA microarray analyses we searched which candidate miRNAs would inhibit two major KRAS-associated signal transduction pathways using SRE and AP1 luciferase reporter assays. And we found 2 miRNAs. miR-4689 regulated not only RAF/MEK/ERK pathway but also PI3K/Akt pathway by directly binding to both 3UTR of KRAS and the coding sequence of Akt1. Down-regulation of Akt1 led to sequential induction of pro-apoptotic factors and inhibition of anti-apoptotic molecules. Systemic administration of miR-4689 with super carbonate apatite nanoparticles in nude mice significantly inhibited the growth of pre-established DLD1 xenografts compared to miR-NC with a proof of down-regulation of KRAS and Akt1.