Lipid nanovectors to use non-coding RNA oligonucleotides in glioblastoma in combination with standard therapy
Prof. Giuseppe De Rosa and Dr. Virginia Campani (postdoc fellow sponsored by the PRC) - Department of Pharmacy, University Federico II of Naples
Prof. Michele Caraglia, Dr. Silvia Zappavigna and Dr. Marianna Abate - Department of Precision Medicine, University of Campania “Luigi Vanvitelli”, Naples, Italy
Dr. Carlo Leonetti and Dr. Manuela Porru - Department of Research, Diagnosis and Innovate Technologies, Regina Elena National Cancer
Nanocarriers based on cationic lipids have been largely investigated to deliver DNA or RNA oligonucleotides in different forms of cancer. Self-assembling nanoparticles (SANPs) based on cationic lipids have been recently developed for the delivery of bisphosphonates in different form of tumors,1, 2) among them glioblastoma.3) Here, we propose to test SANPs to deliver microRNA (miRNA) in a novel therapeutic approach against glioblastoma. In the first stage, the project will be focused on the optimization of SANPs for miRNA delivery, namely by testing different cationic, PEGylated, and neutral lipids, as well as different lipid ratios. The most promising SANP formulation encapsulating the miRNA (miR-603 against the O6-methylguanine methyl transferase) will be tested on glioblastoma cells, alone or in association to the gold standard temozolamide to sensitize glioblastoma cells to chemotherapeutic drugs. Afterwards, biodistribution safety studies on plain formulations and miRNA-encapsulating SANPs selected in the previous phase of the study will be carried out.
This research activity should provide the proof-of-principle for using SANPs as delivery system for miRNA in the treatment of brain tumors, thus supporting the further studies.
Benefit for the community
The technology, that are self-assembling nanoparticles (SANPs), have already been successfully developed for the use of bisphosphonates in the treatment of brain tumor. The product patented by the proposer, with the support of a pharmaceutical company, obtained the Orphan Drug Designation on 29/08/2016-EU/3/16/1735, for the treatment of high-grade glioma, thus supporting the further development clinical development of the product. Here, we propose to use the same technology for RNA oligonucleotides, to deliver miRNAs that have been proposed as key regulator of several processes related to the tumor progression. The project aims to assess the correlation between the lipid composition of the nanoparticles and the miRNA intracellular uptake as well as the biodistribution of miRNA-containing SANPs. These findings certainly should provide new insights for the use of this technology (including for the delivery of bisphosphonates), but also give a novel opportunity to design RNA-based therapy.
On the other hand, although the project has not the ambition to provide conclusive results for novel therapies, it should provide the proof-of-principle for an innovative therapeutic strategy in the treatment of glioblastoma, by increasing the sensitivity of glioblastoma cells to chemotherapeutics already in the clinical practice. It is worthy of note that SANPs will be designed to rapidly move “from the bench to the bed side”, by avoiding the scale-up issues generally encountered for the industrialization of nanoparticle-based formulations.
During our research, we selected the lipid compositions, namely the SANP based on DOTAP (a cationic lipid) together with Cer-PEG (a PEGylated sphingosine-based lipid) or DSPE-PEG, respectively, that showed optimal physical-chemical characteristics in terms of size and polydispersity index, high miRNA encapsulation efficiency, good stability following incubation in serum, and no hemolytic activity. Moreover, these formulations allowed to achieve enhanced miRNA uptake, especially in an U87MG (glioblastoma) cell line, compared to the control and to the other tested formulations. The formulation containing the Cer-PEG showed the highest miRNA intracellular uptake. Finally, in an orthotopic model of glioblastoma, the formulation containing Cer-PEG showed the highest miRNA delivery into the brain, as well as in the other investigated organs such as lungs, kidneys, heart, and liver. Further studies could clarify the mechanism by which formulations containing Cer-PEG could provide highest transfection into the cells as well as higher miRNA delivery in some organs, such as the brain.
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