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Abstract Infections by Clostridioides difficile , a bacterium that targets the large intestine (colon), impact a large number of people worldwide. Bacterial colonization is mediated by two exotoxins: toxins A and B. Short peptides that can be delivered to the gut and inhibit the biocatalytic activity of these toxins represent a promising therapeutic strategy to prevent and treat C. diff . infection. We describe an approach that combines a Pep tide B inding D esign (PepBD) algorithm, molecular-level simulations, a rapid screening assay to evaluate peptide:toxin binding, a primary human cell-based assay, and surface plasmon resonance (SPR) measurements to develop peptide inhibitors that block Toxin A in colon epithelial cells. One peptide, SA1, is found to block TcdA toxicity in primary-derived human colon (large intestinal) epithelial cells. SA1 binds TcdA with a K D of 56.1 ± 29.8 nM as measured by surface plasmon resonance (SPR). 

Most affinity-based biosensors are designed to be single-use devices, based on the measurement of irreversible binding events, which makes longitudinal monitoring resource-intensive, and typically prohibits the measurement of analyte fluctuations over time using the same device. Selective reversal of biorecognition events, i.e., regeneration, may enable repeated and longitudinal use of affinity-based biosensors; however, typical regeneration methods utilize additional chemical reagents, requiring longer processing times and increasing the likelihood of operator error. The development of a “solid-state” regeneration method provides significant value for extending the utility of affinity-based biosensors, such as electrochemical immunosensors and aptasensors. Herein, we report the characterization of a method for electronically controlling pH without additional reagents. Palladium was used to induce pH swings in aqueous electrolytes and buffers by application of an electric potential. The developed system was able to affect acidic and basic pH changes of ± 4. The efficacy of this method was further demonstrated by reversing common affinity-binding complexes and compared to conventional glycine-based regeneration. 

Gene therapies have shown great promise for the potential treatment of a broad range of diseases. Adeno-associated viruses (AAVs) are popular gene vectors because of their ability to target specific tissues, and they have demonstrated high transduction efficiencies in multiple neurological targets. While these therapeutics hold great promise, their biomanufacturing has limited potential cost-reduction and more widespread adoption. Herein, we report the preliminary development of an immunosensor for measuring the titer of adeno-associated virus 2 (AAV2), which may be deployed for rapid quantification of product yield during AAV biomanufacturing. We functionalized an interdigitated electrode array with anti-AAV2 antibodies, and electrochemical impedance spectroscopy was employed to investigate the response to AAV2 titer. A Faradaic sensing principle was utilized, in which the charge transfer resistance (Rct) of an electrochemical reporter was monitored after capture of AAV2 on the surface of the sensor. A linear response was measured over titers 1012 - 1013 capsids/mL. 

Viral vectors are poised to acquire a prominent position in modern medicine and biotechnology owing to their role as delivery agents for gene therapies, oncolytic agents, vaccine platforms, and a gateway to engineer cell therapies as well as plants and animals for sustainable agriculture. The success of viral vectors will critically depend on the availability of flexible and affordable biomanufacturing strategies that can meet the growing demand by clinics and biotech companies worldwide. In this context, a key role will be played by downstream process technology: while initially adapted from protein purification media, the purification toolbox for viral vectors is currently undergoing a rapid expansion to fit the unique biomolecular characteristics of these products. Innovation efforts are articulated on two fronts, namely (i) the discovery of affinity ligands that target adeno-associated virus, lentivirus, adenovirus, etc.; (ii) the development of adsorbents with innovative morphologies, such as membranes and 3D printed monoliths, that fit the size of viral vectors. Complementing these efforts are the design of novel process layouts that capitalize on novel ligands and adsorbents to ensure high yield and purity of the product while safeguarding its therapeutic efficacy and safety; and a growing panel of analytical methods that monitor the complex array of critical quality attributes of viral vectors and correlate them to the purification strategies. To help explore this complex and evolving environment, this study presents a comprehensive overview of the downstream bioprocess toolbox for viral vectors established in the last decade, and discusses present efforts and future directions contributing to the success of this promising class of biological medicines. 

Abstract Recyclable and biodegradable microelectronics, i.e., “green” electronics, are emerging as a viable solution to the global challenge of electronic waste. Specifically, flexible circuit boards represent a prime target for materials development and increasing the utility of green electronics in biomedical applications. Circuit board substrates and packaging are good dielectrics, mechanically and thermally robust, and are compatible with microfabrication processes. Poly(octamethylene maleate (anhydride) citrate) (POMaC) – a citric acid-based elastomer with tunable degradation and mechanical properties – presents a promising alternative for circuit board substrates and packaging. Here, we report the characterization of Elastomeric Circuit Boards (ECBs). Synthesis and processing conditions were optimized to achieve desired degradation and mechanical properties for production of stretchable circuits. ECB traces were characterized and exhibited sheet resistance of 0.599 Ω cm⁻², crosstalk distance of <0.6 mm, and exhibited stable 0% strain resistances after 1000 strain cycles to 20%. Fabrication of single layer and encapsulated ECBs was demonstrated. 

The recent uptick in the approval of ex vivo cell therapies highlights the relevance of lentivirus (LV) as an enabling viral vector of modern medicine. As labile biologics, however, LVs pose critical challenges to industrial biomanufacturing. In particular, LV purification—currently reliant on filtration and anion-exchange or size-exclusion chromatography—suffers from long process times and low yield of transducing particles, which translate into high waiting time and cost to patients. Seeking to improve LV downstream processing, this study introduces peptides targeting the enveloped protein Vesicular stomatitis virus G (VSV-G) to serve as affinity ligands for the chromatographic purification of LV particles. An ensemble of candidate ligands was initially discovered by implementing a dual-fluorescence screening technology and a targeted in silico approach designed to identify sequences with high selectivity and tunable affinity. The selected peptides were conjugated on Poros resin and their LV binding-and-release performance was optimized by adjusting the flow rate, composition, and pH of the chromatographic buffers. Ligands GKEAAFAA and SRAFVGDADRD were selected for their high product yield (50%–60% of viral genomes; 40%–50% of HT1080 cell-transducing particles) upon elution in PIPES buffer with 0.65 M NaCl at pH 7.4. The peptide-based adsorbents also presented remarkable values of binding capacity (up to 3·109 TU per mL of resin, or 5·1011 vp per mL of resin, at the residence time of 1 min) and clearance of host cell proteins (up to a 220-fold reduction of HEK293 HCPs). Additionally, GKEAAFAA demonstrated high resistance to caustic cleaning-in-place (0.5 M NaOH, 30 min) with no observable loss in product yield and quality. 

Abstract Poly(ethylene terephthalate) (PET) is a highly recyclable plastic that has been extensively used and manufactured. Like other plastics, PET resists natural degradation, thus accumulating in the environment. Several recycling strategies have been applied to PET, but these tend to result in downcycled products that eventually end up in landfills. This accumulation of landfilled PET waste contributes to the formation of microplastics, which pose a serious threat to marine life and ecosystems, and potentially to human health. To address this issue, our project leveraged synthetic biology to develop a whole‐cell biocatalyst capable of depolymerizing PET in seawater environments by using the fast‐growing, nonpathogenic, moderate halophileVibrio natriegens. By leveraging a two‐enzyme system—comprising a chimera ofIsPETase andIsMHETase fromIdeonella sakaiensis—displayed onV. natriegens, we constructed whole‐cell catalysts that depolymerize PET and convert it into its monomers in salt‐containing media and at a temperature of 30°C.