Research suggests that recruitment, the number of fish entering a population in a given year, is influenced by the environment. However, environmental drivers are not currently used to refine the recruitment estimates of most statistical models used in fisheries management (hereafter, population assessment models). This is increasingly relevant as fish populations experience long-term productivity shifts due to climate change. One major goal of the National Oceanic and Atmospheric Administration (NOAA) in recent years has been to incorporate environmental drivers into population assessment models. This is achieved by using time series data of environmental drivers to inform model estimates. One ongoing challenge is determining which environmental drivers have potential to improve model estimates. In this project, we aimed to determine how correlated an environmental driver time series must be to historical recruitment deviations to improve key model estimates – recruitment deviations and population depletion – in recent years for a range of species. We used R to simulate mock environmental driver time series with varying correlation levels to recruitment deviations by randomly sampling data from a normal distribution. We then compared errors in estimates between population assessment models fit with and without the simulated environmental data. Our results suggest that the more correlated an environmental driver is to historical recruitment deviations, the more accurate estimates of both recruitment deviations and population depletion become. However, our results also reveal that the correlation level necessary for environmentally-driven models to consistently produce more accurate estimates than the original model varies across species. These correlation thresholds are also generally higher than observed correlations between recruitment deviations and environmental drivers in actual fish populations. We suggest fisheries scientists run similar simulation experiments to determine which correlation levels have the potential to improve population assessment models for their target species.

Peptide cancer vaccines have had limited clinical success despite their safety, characterization, and production advantages. We hypothesize that the poor immunogenicity of peptides can be surmounted by delivery vehicles that overcome the systemic and cellular drug delivery barriers faced by peptides. We introduce Man-VIPER, a self-assembling, pH-sensitive, mannosylated polymeric peptide delivery platform that targets dendritic cells in the lymph nodes and facilitates endosomal release of antigens through a conjugated membranolytic peptide melittin. We evaluated polymers with both releasable (Man-VIPER-R) or non-releasable (Man-VIPER-NR) D-melittin. Both Man-VIPER polymers exhibited superior endosomolysis and antigen cross-presentation compared to non-membranolytic D-melittin-free analogues (Man-AP) in vitro. In vivo, Man-VIPER polymers demonstrated an adjuvanting effect, induced the proliferation of antigen-specific cytotoxic T cells and helper T cells compared to free peptides and Man-AP. Remarkably, antigen delivery with Man-VIPER-NR generated significantly more antigen-specific cytotoxic T cells than Man-VIPER-R in vivo. As our candidate for a therapeutic vaccine, Man-VIPER-NR exerted superior efficacy in a B16F10-OVA tumor model. These results highlight Man-VIPER-NR as a safe and powerful peptide cancer vaccine platform for cancer immunotherapy.

Chronic kidney disease (CKD) is an incurable, progressive condition that affects up to 700 million people globally. Progression of CKD currently leads to a deteriorating quality of life on dialysis, often resulting in terminal end-stage renal disease (ESRD) and increased risk of cardiovascular disease. A hallmark feature of CKD progression is epithelial-to-mesenchymal transition (EMT), a process by which kidney cells obtain malignant properties like increased mobility and resistance to apoptosis. Despite the pressing burden of CKD, current therapeutics like angiotensin-converting enzyme (ACE) inhibitors and angiotensin-receptor blockers are unable to halt the fibrotic progression of CKD. In recent years, epigallocatechin-3-gallate (EGCG) has been shown to be a promising candidate to inhibit EMT in kidney tubular epithelial cells (TECs). In this project, we conjugated EGCG to a low molecular weight polymer previously engineered at the Pun Lab for enhanced localization to the kidney TECs. In a TGF-β1 induced fibrosis model in human kidney cells, we showed that this polymer-EGCG conjugate (poly-EGCG) could diminish the RNA and protein expression of mesenchymal markers compared to untreated controls. We also confirmed that poly-EGCG was well-tolerated across a broad range of concentrations through in vitro cell viability assays. Finally, immunohistochemistry staining of mouse kidney samples injured with anti-glomerular antibodies displayed partial fibrotic recovery when treated with unconjugated EGCG. Future in vivo studies will aim to optimize the efficacy of poly-EGCG treatments compared to unconjugated EGCG treatments by assessing histology and urine samples for markers of kidney dysfunction. Through improved delivery of EGCG to the kidney TECs, this novel polymer-EGCG conjugate has the potential to halt the progression of EMT for future patients with CKD.

The Stimulator of Interferon Genes (STING) pathway is a promising target for cancer immunotherapies. However, STING agonists have poor cell membrane permeability and also off-site toxicity that limit their therapeutic application. Our motivation is thus to develop a novel method for the delivery of non-cell membrane-permeable STING agonists. We propose incorporating the STING agonist in a self-assembling polymer, which then transports the drug into the cytosol, thereby avoiding membrane permeability issues. The first step is the incorporation of the STING agonist into Man-VIPER, a cytosolic drug delivery system developed by the Pun Lab. Man-VIPER mediates cytosolic delivery via endosomal escape, releasing the STING agonist cargo into the cytosol. Currently, I am working on synthesizing and purifying the small molecule STING agonist called SR-012. I tested many possible reaction pathways to make SR-012, which is unavailable commercially and has only been previously synthesized once. I developed a novel alternative reaction pathway and purification that works consistently. I am also currently synthesizing the polymer building blocks for the Man-VIPER copolymer that will deliver SR-012. After the optimized drug-polymer complex is completed, I will work with collaborators to quantify the max tolerated dose. We hope to demonstrate that specificity is improved through lower toxicity, allowing for a higher maximum tolerated dose than my control, the membrane permeable STING agonist SR-717. Next, I will assay the immunotherapeutic effectiveness of Man-VIPER delivered SR-012 using in-vivo tumor models.

An effective peptide-based cancer vaccine requires efficient intracellular delivery of antigen peptides to activate tumor-killing immune responses. However, the localization of peptide antigens during endosomal release to optimize the immune response remains under-investigated. The Pun Lab has developed the self-assembling Virus-Inspired Polymers for Endosomal Release (VIPER) that can induce endosomal escape of peptide antigens. The current VIPER formulation employs reducible disulfide bonds for antigen conjugation. We hypothesize that the controlled antigen peptides release mediated by endosomal proteases can result in more effective antigen presentation that leads to more potent tumor-killing cell responses. In this project, I replace VIPER’s antigen conjugation strategy with the pentafluorobenzyl (PFB) moiety (VIPER-NR) and introduce an enzyme-labile linker in the antigen sequence to determine the optimal peptide release kinetics and localization for optimal tumor-killing cell responses. I synthesized a library of enzyme-labile linkers for the ovalbumin antigen peptide of VIPER-NR. I utilized a dendritic cell cross-presentation assay to screen for linkers associated with that efficient antigen presentation in vitro. Subsequently, I conducted enzyme-mediated drug release assay and red blood cell lysis assay in vitro to evaluate the cleaved antigen profile and endosomal escape induction capacity of VIPER-NR with cleavable linkers compared to VIPER-NR. We expect early release of antigens during endocytosis with these modified antigen sequences in VIPER-NR in vitro and significantly elevated cytotoxic T-cell response against the model antigen in vivo. This project could improve the effectiveness of the VIPER system as a peptide vaccine delivery platform. It would also provide further understanding of optimal antigen release profiles for a better tumor-killing cell response in cancer vaccine applications.