The recent success of mRNA vaccines in SARS-CoV-2 clinical trials is in part due to the development of lipid nanoparticle delivery systems that not only efficiently express the mRNA-encoded immunogen after intramuscular injection, but also play roles as adjuvants and in vaccine reactogenicity. We present an overview of mRNA delivery systems and then focus on the lipid nanoparticles used in the current SARS-CoV-2 vaccine clinical trials. The review concludes with an analysis of the determinants of the performance of lipid nanoparticles in mRNA vaccines.

The 1,4-diene motif, also known as a skipped diene, is widespread across various classes of natural products including alkaloids, fatty acids, terpenoids, and polyketides as part of either the finalized structure or a biosynthetic intermediate.

Protein–protein interactions lie at the heart of many biological processes and therefore represent promising drug targets. Despite this opportunity, identification of protein–protein interfaces remains challenging. We have previously developed a method that relies on coating protein surfaces with small-molecule dyes to discriminate between solvent-accessible protein surfaces and hidden interface regions. Dye-bound, solvent-accessible protein regions resist trypsin digestion, whereas hidden interface regions are revealed by denaturation and sequenced by MS. The small-molecule dyes bind promiscuously and with high affinity, but their binding mechanism is unknown. Here, we report on the optimization of a novel dye probe used in protein painting, Fast Blue B + naphthionic acid, and show that its affinity for proteins strongly depends on hydrophobic moieties that we call here “hydrophobic clamps.” We demonstrate the utility of this probe by sequencing the protein–protein interaction regions between the Hippo pathway protein Yes-associated protein 2 (YAP2) and tight junction protein 1 (TJP1 or ZO-1), uncovering interactions via the known binding domain as well as ZO-1's MAGUK domain and YAP's N-terminal proline-rich domain. Additionally, we demonstrate how residues predicted by protein painting are present exclusively in the complex interface and how these residues may guide the development of peptide inhibitors using a case study of programmed cell death protein 1 (PD-1) and programmed cell death 1 ligand 1 (PD-L1). Inhibitors designed around the PD-1/PD-L1 interface regions identified via protein painting effectively disrupted complex formation, with the most potent inhibitor having an IC₅₀ of 5 μm.

The aminopeptidase activity (AP) of the leukotriene A₄ hydrolase (LTA₄H) enzyme has emerged as a therapeutic target to modulate host immunity. Initial reports focused on the benefits of augmenting the LTA₄H AP activity and clearing its putative pro-inflammatory substrate Pro-Gly-Pro (PGP). However, recent reports have introduced substantial complexity disconnecting the LTA₄H modulator 4-methoxydiphenylmethane (4MDM) from PGP as follows: (1) 4MDM inhibits PGP hydrolysis and subsequently inhibition of LTA₄H AP activity, and (2) 4MDM activates the same enzyme target in the presence of alternative substrates. Differential modulation of LTA₄H by 4MDM was probed in a murine model of acute lung inflammation, which showed that 4MDM modulates the host neutrophilic response independent of clearing PGP. X-ray crystallography showed that 4MDM and PGP bind at the zinc binding pocket and no allosteric binding was observed. We then determined that 4MDM modulation is not dependent on the allosteric binding of the ligand, but on the N-terminal side chain of the peptide. In conclusion, our study revealed that a peptidase therapeutic target can interact with its substrate and ligand in complex biochemical mechanisms. This raises an important consideration when ligands are designed to explain some of the unpredictable outcomes observed in therapeutic discovery targeting LTA₄H.