Effect of Confinement in Nanopores on RNA Interactions with Functionalized Mesoporous Silica Nanoparticles

Abstract

Amine-functionalized mesoporous silica nanoparticles (MSNPAs) are ideal carriers for oligonucleotides for gene delivery and RNA interference. This investigation examines the thermodynamic driving force of interactions of double-stranded (ds) RNA with MSNPAs as a function of RNA length (84 and 282 base pair) and particle pore diameter (nonporous, 2.7, 4.3, and 8.1 nm) using isothermal titration calorimetry, extending knowledge of solution-based nucleic acid−polycation interactions to RNA confined in nanopores. Adsorption of RNA follows a two-step process: endothermic interactions driven by entropic contribution from counterion (and water) release and an exothermic regime dominated by short-range interactions within the pores. Evidence of hindered pore loading of the longer RNA and pore sizedependent confinement of RNA in the MSPAs is provided from the relative contributions of the endothermic and exothermic regimes. Reduction of endothermic and exothermic enthalpies in both regimes in the presence of salt for both lengths of RNA indicates the significant contribution of short-range electrostatic interactions, whereas ΔH and ΔG values are consistent with conformation changes and desolvation of nucleic acids upon binding with polycations. Knowledge of the interactions between RNA and functionalized porous nanoparticles will aid in porous nanocarrier design suitable for functional RNA delivery.

Mesoporous Silica Particles as Drug Delivery Systems—The State of the Art in Loading Methods and the Recent Progress in Analytical Techniques for Monitoring These Processes

“…ITC measures directly the heat released or absorbed during a binding, simultaneously providing information about a range of binding parameters such as entropy, enthalpy, the equilibrium constants, etc. It is relevant in some studies, involving nucleic acid incorporation into MSNs, to determine the nature of DNA or RNA interaction with the silica [ 256 , 257 ].…”

Relating Mobility of dsRNA in Nanoporous Silica Particles to Loading and Release Behavior

“…For the porous particles, these concentrations are very close to the end of the first binding region (endothermic) of dsRNA to silica nanoparticles as measured by ITC (0.12, 0.2, and 0.26, mol bp/mol amine for 2.7, 4.3, and 8.1 nm-diameter pores, respectively), which is attributed to saturation of the phosphate on RNA molecules by surface amines. 43 Here, concentration-dependent measurements indicate that at this ratio, the particles are saturated with dsRNA. A halo of fluorescence at the surface of the spherical particles indicates that the dsRNA is adsorbed to the surface but excluded from the pores for the 282 bp dsRNA incubated with microspheres with 3.9 nm pores.…”

“…The difference in pore accessibility by 84 and 282 bp RNA is consistent with thermodynamic interactions with pSNPs with a pore size of 2.7 and 4.3 nm, where higher enthalpy of binding was observed for 84 bp RNA compared to 282 bp RNA. 43 However, the difference is less pronounced for 4.3 nm particles (ΔH = 7.13 and 5.92 kJ/mol, for 84 and 282 bp RNA, respectively) as opposed to that for 2.7 nm particles (ΔH = 9.98 and 1.56 kJ/ mol for 84 and 282 bp RNA, respectively), suggesting the relative ease of access for longer RNA as the particle pore diameter increases. Steric hindrance due to the ability of larger RNA molecules to bend is probably responsible for not being able to penetrate 3.9 nm pores.…”

“…38 The understanding of adsorption and diffusion of stiff polyelectrolytes such as RNA confined in pores is incomplete, as indicated by a number of recent theoretical and computational studies. 39−42 In our previous work quantifying the thermodynamic interaction between dsRNA (84 and 282 bp) and aminefunctionalized mesoporous silica nanoparticles (nonporous, 2.7, 4.3, and 8.1 nm pore size; <200 nm-diameter particles) using isothermal titration calorimetry (ITC), 43 two different binding regimes were observed: first, an endothermic interaction from the release of surface counter-ions and water followed by an exothermic region dominated by shortrange electrostatic interactions within the pores. The end of the first binding phase indicates phosphate saturation between the dsRNA and the surface amine group.…”

Nanotechnology‐Assisted RNA Delivery: From Nucleic Acid Therapeutics to COVID‐19 Vaccines

“…It was shown that the efficient design of dsRNA nanocarriers offers the advantage of creating small pores to protect the RNAs from degradation, while a sufficient space is required for dsRNA threading into pores. [ 126 ] A layer of lipid conjugated with iRGD peptide was used to stabilize silica nanoparticles by copper‐free click‐chemistry in a tumor‐penetrating siRNA and miRNA codelivery investigation. The carrier efficiency in cytosolic RNA delivery was further enhanced by loading a near‐infrared‐responsive photosensitizer into nanoparticles for the local generation of reactive oxygen species (ROS).…”