Cover Feature: Direct Conjugation of Gallium-(III)-Corroles to Short Interfering RNA(siRNA) Providing Real-Time siRNA Imaging and Gene Silencing (ChemPlusChem 6/2024).

Invited for this month's cover is the group of Jean-Paul Desaulniers at Ontario Tech University. The cover picture shows the successful conjugation of a GaIII-corrole to an siRNA to enable live cell imaging. Read the full text of the article at 10.1002/cplu.202400084.

Direct Conjugation of Gallium-(III)-Corroles to Short Interfering RNA(siRNA) Providing Real-Time siRNA Imaging and Gene Silencing.

Discovering new modifications for oligonucleotide therapeutics is essential for expanding its application to new targets and diseases. In this project, we focus on conjugating metaled ligands to short interfering RNAs (siRNAs) to investigate robust and simple conjugation methods for adding new properties such as real-time imaging to the siRNA. Here we report the chemical synthesis of novel Ga-(III)-corroles for their direct conjugation to siRNAs. Ga-(III)-corrole-siRNAs showed promising results when evaluated for gene silencing and live cell imaging. The knockdown activity of the firefly luciferase reporter gene was measured to evaluate gene silencing activity. Gene silencing studies from two 5'-Ga-(III)-labeled-siRNAs exhibited dose-dependent knockdown with IC50s of 812.7 and 451.4 pM, which is comparable to wild-type (IC50=439.7 pM) in the absence of red light, and IC50s of 562.9 and 354.5 pM, which is also comparable to wild-type (IC50=337.4 pM), in the presence of red light. In addition, imaging studies with Ga-(III)-corrole-modified siRNAs showed intense fluorescence in HeLa cells, highlighting that the Ga-(III)-corrole modification is an effective fluorophore for siRNA tracing and imaging. Moreover, the photodynamic activity of free base corrole vs the Ga-(III)-corrole was evaluated. Results show an increase of light cytotoxicity of the corrole ligand upon the addition of Ga-(III); however, no phototoxicity was observed when Ga-(III) ligands were linked to siRNA. In conclusion, Ga-(III)-corrole-siRNAs show promising results for applications in simultaneous real-time imaging and gene silencing.

Effective carrier-free gene-silencing activity of sphingosine-modified siRNAs.

RNA interference (RNAi) is a natural cellular process that silences the expression of target genes in a sequence-specific way by mediating targeted mRNA degradation. One of the main challenges in RNAi research is developing an effective career-free delivery system and targeting cells in the central nervous system (CNS). Recently, lipid-conjugated systems involving fatty acids have shown promising potential as safe and effective delivery systems of oligonucleotides to CNS cells due to their hydrophobic tails and interactions with the cell's hydrophobic membrane. Therefore, in this study, we are interested in creating career-free siRNA therapeutics for potential applications in drug delivery to the CNS. Here we explore different synthetic pathways of conjugating sphingolipids containing long-carbon chains to siRNA and assess their effectiveness as career-free delivery systems. In this project, a library of sphingosine-modified siRNAs was created, and their gene-silencing effect was evaluated in both the presence and absence of a transfection carrier. siRNAs modified with one or two sphingosine moieties resulted in dose-dependent gene knockdown while demonstrating promising results for their use as carrier-free agents. The IC50 values of single-modified siRNAs ranged from 49.9 nM to 670.7 nM, whereas double-modified siRNAs had IC50 values in the range of 49.9 nM to 66.4 nM. In conclusion, sphingosine-modified siRNAs show promising results in advancing carrier-free siRNA therapeutics.

SiRNAs with Neutral Phosphate Triester Hydrocarbon Tails Exhibit Carrier-Free Gene-Silencing Activity.

Short interfering RNAs (siRNAs) show promise as gene-silencing therapeutics, but their cellular uptake remains a challenge. We have recently shown the synthesis of siRNAs bearing a single neutral phenylethyl phosphotriester linkage within the sense strand. Here, we report the synthesis of siRNAs bearing three different hydrophobic phosphate triester linkages at key positions within the sense strand and assess their gene silencing in the absence of a transfection carrier. The best siRNAs bearing hydrophobic phosphate triester tails were not aromatic and exhibited effective gene silencing (IC50 ≈ 56-141 nM), whereas the aromatic derivative with three hydrophobic tails did not exhibit carrier-free gene silencing.

A dual therapeutic system based on corrole-siRNA conjugates.

Corrole molecules are a new generation of photosensitizers (PS) due to their ease of tunability for different medical applications. Their ability to initiate cellular death using a wide range of non-toxic wavelengths allows for the creation of non-invasive treatments. This work focuses on creating potent and non-invasive treatments by advancing siRNA therapeutics by directly conjugating siRNAs with the photosensitizer, corrole. Combining gene silencing with photodynamic therapy (PDT) creates a non-invasive dual therapy system. Basic synthetic designs were explored to create novel corrole-phosphoramidites and from these, a small library of corrole-functionalized short interfering RNAs (corrole-siRNAs) were synthesized. Corrole-siRNA conjugates showed promising results when evaluated for gene silencing and PDT therapy in vitro. Gene silencing effects were evaluated in cells by measuring the knockdown activity of the firefly luciferase reporter gene. Gene silencing studies from four siRNAs showed promising dose dependent knockdown with IC50s of 387.8, 77.8, 60.0, and 49.4 pM in the absence of red light, and 101.0, 57.2, 55.3, and 23.8 pM in the presence of red light. Furthermore, PDT showed approximately a 50% decrease in cell viability for red-light irradiated cells treated with corrole-siRNAs, demonstrating the effective role of corrole to act as a photosensitizer while still maintaining robust siRNA activity. In conclusion, corrole-siRNAs show a promising path for developing novel siRNA combination therapy.