Internal Oligoguanidinium Transporter: Mercury Free Scalable Synthesis, Improvement of Cellular Localization, Endosomal Escape, Mitochondrial Localization and Conjugation with Antisense Morpholino for NANOG Inhibition to Induce Chemosensitisation of Taxol

A non toxic delivery vehicle is essential for the therapeutic applications of antisense phosphorodiamidate morpholino oligonucleotides (PMOs). Though guanidinium rich or arginine rich cellular transporter conjugated Vivo-PMO or PPMO has been developed for in vivo application, however, either their toxicity or stability has become an issue. Previously we reported non-peptidic internal guanidinium transporter (IGT) mediated delivery of PMO for gene silencing and got encouraging results. In this paper we report the synthesis of IGT using Hg-free method for scale up and N-terminal modification of IGT with a suitable hydrophobic or lipophilic group to improve the cell permeability, endosomal escape, mitochondrial localization and to reduce toxicity in MTT assay. For the delivery of PMO, IGT-PMO conjugate was synthesized to target NANOG in cells, a transcription factor required for cancer stem cell proliferation and embryonic development and is involved in many cancers. Our data shows IGT-PMO-facilitated NANOG inhibition and thereby the prevention of EpCAM-N-Cadherin-Vimentin axis mediated epithelial to mesenchymal transition (EMT) in MCF-7 cells. Moreover, unlike taxol, NANOG inhibition influences the expression of stemness factor c-Myc, Hh-Gli signaling proteins, other cancer related factors and their respective phenotypes in cancer cells. To the best of our knowledge, this is the first report to illustrate that the IGT-PMO-mediated NANOG inhibition increases the therapeutic potential of taxol and induces G0-G1 arrest in cancer cells to prevent the cancer progression. However, it warrants further investigations in other cancer cells and preclinical platforms.