Several studies have been performed with the aim of identifying drugs able in inhibiting Epithelial-Mesenchymal Transition (EMT), chiefly by blocking PI3K/Akt pathway. In previous work, we have demonstrated how pharmacological doses of myo-Inositol (myo-Ins) can block EMT in breast cancer cells by downregulating PI3K/Akt. Moreover, this effect is associated to cytoskeletal remodelling, finally triggering tumor reversion.
We have investigated the mechanism of action of myo-Ins in high (MDA-MB-231) and low invasive (MCF-7) human breast cancer cells, evidencing a key role of IP6K1 in myo-Ins effects. IP6K1, acts in inositol metabolism, inhibiting ISYNA1, responsible for myo-Ins biosynthesis, probably by inducing de novo DNA methylation via increase in DNMT3B.
In MDA cells, myo-Ins downregulates IP6K1 at 30' and 24h, whilst DNMT3B is reduced only at 30'. Furthermore, IP6K1 inhibition correlates with a decrease in cancer cells motility. Unexpectedly, in MCF-7 myo-Ins addiction does not induce any significant changes in the protein levels of the above-mentioned parameters (IP6K1, DNMT3B, ISYNA1).
By in silico analysis, we observed miR-125a-5p up-regulation by myo-Ins in MDA cells. We hypothesized thus the divergent response in between the two cancer cell lines can be ascribed to a different modulation of miR-125a-5p. In fact, we recorded a significant miR-125a-5p downregulation in breast cancer tissues and an its relevant increase after myo-Ins treatment in MDA cells. Moreover, a strong interaction was predicted between miR-125a-5p and IP6K1 in 3'-UTR site. As expected, miR-125a-5p levels were found unchanged upon myo-Ins treatment in MCF-7 cells.
These findings suggest that myo-Ins causes early gene expression changes, by miRNAs and methylation remodelling.
The aim of this project is to elucidate the role of miR-125a-5p/IP6K1 axis, by using RNAi and overexpression tools. Accordingly, this approach could allow us to identify key factors as well as new therapeutic targets.
The genetic/molecular heterogeneity of cancers may help in explaining the different, individual responses to standard systemic therapy. Given the unsatisfactorily clinical outcome up to date, there is an increasing pressure to characterize molecular profiles of individual patients and to use this information to develop personalized therapy programs. RNA interference (RNAi), due to its specificity, adaptability and breadth of targeting capability, has great potential to serve in assessing the personalized array of therapeutic targets. Due to their robustness and specificity, siRNA and shRNA have been extensively used to silence cancer-related targets (Mixson AJ et al, 2019). A large number of preclinical studies have presented favourable outcomes by silencing genes critical for tumor cell growth, metastasis, angiogenesis and chemoresistance. RNAi technology has been used to inhibit tumor metastasis (Wang Z et al, 2011). For example, in advanced prostate cancer, tumor cells frequently metastasize to bones and regional lymph nodes. A systemic delivery system has been developed to introduce siRNA to the bone-metastatic cancer sites using atelocollagen. siRNAs targeting either EZH2 orp 110-alpha were formulated with atelocollagen and delivered to bone-metastatic lesions in a xenograft model by tail vein injection (Minakuchi Y et al, 2004).
In literature, studies about miR-125a-5p / IP6K1 axis are lacking. Yet, it has been proposed that monitoring of miR-125a-5p expression in breast cancer patients could represent a reliable marker. In fact, downregulation of miR-125a-5p in breast tumor tissue is correlated with a poor prognosis. In this case, IP6K1 inhibitors could be evaluated as a useful therapeutic device (Terao Y et al, inventors; WO/2018/182051, 2018). Given that myo-Ins has been proven to efficiently and safely down-regulate IP6K1, the clinical utility of myo-Ins as natural IP6K1-silencing factor, deserves to be investigated in depth.