Interaction Kinetics of Individual mRNA-Containing Lipid Nanoparticles with an Endosomal Membrane Mimic: Dependence on pH, Protein Corona Formation, and Lipoprotein Depletion.

Lipid nanoparticles (LNPs) have emerged as potent carriers for mRNA delivery, but several challenges remain before this approach can offer broad clinical translation of mRNA therapeutics. To improve their efficacy, a better understanding is required regarding how LNPs are trapped and processed at the anionic endosomal membrane prior to mRNA release. We used surface-sensitive fluorescence microscopy with single LNP resolution to investigate the pH dependency of the binding kinetics of ionizable lipid-containing LNPs to a supported endosomal model membrane. A sharp increase of LNP binding was observed when the pH was lowered from 6 to 5, accompanied by stepwise large-scale LNP disintegration. For LNPs preincubated in serum, protein corona formation shifted the onset of LNP binding and subsequent disintegration to lower pH, an effect that was less pronounced for lipoprotein-depleted serum. The LNP binding to the endosomal membrane mimic was observed to eventually become severely limited by suppression of the driving force for the formation of multivalent bonds during LNP attachment or, more specifically, by charge neutralization of anionic lipids in the model membrane due to their association with cationic lipids from earlier attached LNPs upon their disintegration. Cell uptake experiments demonstrated marginal differences in LNP uptake in untreated and lipoprotein-depleted serum, whereas lipoprotein-depleted serum increased mRNA-controlled protein (eGFP) production substantially. This complies with model membrane data and suggests that protein corona formation on the surface of the LNPs influences the nature of the interaction between LNPs and endosomal membranes.

The memristive effect as a novelty in drug monitoring.

Nanoscale devices exhibiting memristive properties show great potential in a plethora of applications. In this work, memristive nanowires are presented for the first time as ideal candidates for absolutely novel, ultrasensitive, highly specific and selective drug-biosensors, also paving the way for real-time monitoring applications, in coupling with the restoration properties of DNA-aptamers. The hysteretic properties exhibited by the hereby-presented special nanodevices, modified via surface treatments, are leveraged along the complete cycle consisting of DNA-aptamer immobilization, target binding, and DNA-aptamer regeneration for successful and effective detection of Tenofovir, an antiviral drug for HIV treatment, in buffer as well as in non-diluted human serum. This results in ultrasensitive, label-free monitoring of the therapeutic compound with a limit of detection of 3.09 pM in buffer and 1.38 nM in full serum. These LODs demonstrate 10 times higher sensitivity for the in-buffer drug detection, and twice better performance for drug sensing in full human serum, ever obtained. The selectivity of the memristive biosensor for Tenofovir detection was verified through both positive and negative controls in full human serum. In addition, the DNA-aptamer regeneration character is portrayed for the first time through a memristive effect, and scanning electron microscopy throws more light on the binding mechanism efficiency through the variation of the nanodevice surface properties at the nanoscale.The results presented in this work demonstrate that the coupling of the memristive effect and aptamer regeneration provides the best ever realized nano-biosensor for drug detection also in full human serum.

Enzymatic and Nonenzymatic Electrochemical Interaction of Abiraterone (Antiprostate Cancer Drug) with Multiwalled Carbon Nanotube Bioelectrodes.

Unexplored electrochemical behavior of abiraterone, a recent and widely used prostate cancer drug, in interaction with cytochrome P450 3A4 (CYP3A4) enzyme and multiwalled carbon nanotubes (MWCNTs) is investigated in this work. The results reported in this work are significant for personalized medicine and point-of-care chemical treatment, especially to improve the life expectancy and quality of life of patients with prostate-cancer. To this purpose, enzymatic and nonenzymatic electrochemical biosensors were developed and characterized with different concentrations of abiraterone. Nonenzymatic biosensors were functionalized with MWCNTs as a catalyst for signal enhancement, while enzymatic biosensors have been obtained with CYP3A4 protein immobilized on MWCNTs as recognition biomolecule. Enzymatic electrochemical experiments demonstrated an inhibition effect on the CYP3A4, clearly observed as a diminished electrocatalytic activity of the enzyme. Electrochemical responses of nonenzymatic biosensors clearly demonstrated the direct electroactivity of abiraterone when reacting with MWCNT as well as an electrode-fouling effect.