In 2019, hydrodealkenylation was reported as a new synthetic method to replace an alkenyl C(sp2)–C(sp3) bond by a C(sp3)–H bond. Despite the versatility of this method, the ozonolysis step of hydrodealkenylation poses some challenges in functionalizing allylic alcohols, basic amines, and electron-rich aromatics as these three classes of alkene substrates suffer from Grob fragmentation, direct oxidation, and oxidative degradation, respectively. In this enclosed study, the Criegee ozonolysis step was modified to rescue these three classes of previously-incompatible alkenes. With acid additives, acetyl protecting group, and the control of O3 equivalents, hydrodealkenylation was successfully applied to these three classes of substrates.

With the advent of ribonucleic acid (RNA) gene therapies and vaccines, many different nanoparticles are being tested for their ability to effectively package and protect the RNA for cell-specific delivery. Among these nanoparticles are virus-like particles (VLPs) which exploit viruses’ natural life cycles in order to package exogenous RNA molecules. In particular, we are interested in Brome Mosaic Virus (BMV), a spherical virus capable of in vitro self-assembly from purified viral components, namely its capsid protein and an RNA molecule of approximately 3,200 nucleotides (nt) long. We posit that BMV VLP formation is optimized around the capsid protein’s isoelectric point (pI) of 6.5, at which point there is minimal capsid coat protein (CP) self-repulsion and thus maximization of the pro-associative hydrophobic interactions. In order to test this, we generated a “phase diagram” of the CP’s various aggregates across pH and ionic strength conditions in order to investigate which pH and salt conditions allow efficient formation of spherical particles and larger aggregates, including rods and multi shelled spheres. The main aim of this project is to use this phase diagram to improve our VLP formation protocol, which has been shown to produce broken particles which are ineffectual for RNA protection. We posit that increasing the pH used for VLP assembly to match the protein’s isoelectric point of 6.5 will allow particles to form more efficiently; we utilize negative stain electron microscopy (NSEM) to confirm this hypothesis.

This abstract has been withheld from publication.

Amide couplings are the most common reactions in the synthesis of pharmaceutical compounds. However, their often-limited reactivity and wide range of conditions leads to poor atom economy, meaning wasted chemicals. Efforts to predict the yield of amide bond formation as a function of substrate structure and reaction conditions have failed to result in a predictive model. It is hypothesized that one of the major reasons for the difficulty of predicting the yield of amide couplings is the presence of “reactivity cliffs”—pairs of substrates that are structurally similar, but undergo amide coupling in very different yields. To go about elucidating these reactivity cliffs from the CAS database of 8 million reaction, Tanimoto and Cosine similarity models were used. These models calculate molecular similarity based off molecular fingerprints and were used to find the “reactivity cliffs”, along with filtering for a large yield difference and commercial availability. Mechanistic experiments, like doping in an incompatible functional group, can be used to investigate what long-range interactions might be causing the decreased reactivity. By quantifying these affects, models can be better fit for subtle differences in structure and be able to make more substantiated predictions to aid in smoother lead optimization campaigns. The solubility of substrates was one factor discovered to be an important in creating a model sensitive of “reactivity cliffs”.

Exon skipping therapy is a promising treatment strategy for patients with Duchenne Muscular Dystrophy (DMD), a debilitating and fatal genetic disorder marked by muscle degeneration. Previous research has demonstrated that antisense oligonucleotides (ASOs), which are short, synthetic strands of DNA or RNA, can effectively skip specific exons adjacent to exon deletions in DMD to produce a functional dystrophin protein. In the present study, we synthesized a novel three-dimensional (3D) bridge ASO design aimed at enhancing exon 44 skipping, applicable to a subset of 6% of all DMD patients. To assess the efficacy of these ASOs, we differentiated inducible directly reprogrammable myotubes (iDRMs) isolated from two distinct DMD patients and transfected them with varying concentrations (nM) of the ASOs. Following RNA extraction from the cells, we measured exon skipping efficiency using RT-PCR and other biochemical assays. We quantified the gels with PCR products to determine exon skipping percentages, indicated by the ratio of skipped mRNA transcripts over total mRNA transcripts. Our results demonstrated that the 3D-bridge ASO design exhibited a significantly higher exon skipping rate compared to previously reported ASOs. We further screened different ASO backbone chemistries to identify the optimal formulation for 3D-bridge ASOs. These findings highlight the promising potential of this novel 3D-bridge ASO design, positioning it as an effective breakthrough therapy for DMD patients with exon 44 deletions.