Abstract Background Bronchiolitis obliterans syndrome (BOS) is a late complication after allogeneic hematopoietic cell transplantation (HCT). While retrospective evidence shows symptomatic respiratory viral infections (RVI) may contribute to the development and progression of BOS, a similar role for asymptomatic RVI is unknown. This study aims to assess the incidence of asymptomatic and symptomatic RVI in a high-risk population. Methods Allogeneic HCT recipients with new-onset chronic graft-versus-host disease (cohort 1, at risk for BOS) and new onset BOS (cohort 2) were enrolled in an ongoing multi-center prospective longitudinal study (NCT05250037) to assess the role of RVI in development of BOS. Participants conducted weekly handheld home spirometry, weekly symptom surveys, and submitted self-administered nasal swabs for respiratory viral PCR testing every two weeks; additional swabs were prompted if symptom survey score reached ≥2. An asymptomatic episode was defined as one or more consecutive positive viral swab with a symptom score of 0-1 and no positive symptoms in the 2 weeks before or after. A symptomatic RVI episode was defined as the first positive PCR to the final positive PCR with any associated symptom score ≥2, allowing no more than ≥4 weeks or 2 negative samples between any 2 consecutive positive PCR samples. Results From March 2022 to July 2023, 73 participants were enrolled (cohort 1: n=50; cohort 2: n=23), with 47 completing one year of follow-up. Compliance was 68% for surveys and 62% for nasal swabs. Twenty percent of 744 swabs tested positive for ≥1 virus. Of 43 participants who tested positive for RVI, 21% (9/43) had 1 episode, 21% (9/43) had 2 episodes, and 52% (25/43) had ≥3 episodes. Ninety-five percent of RVI episodes (104/109) were symptomatic, most commonly rhinovirus (22%), followed by SARS-CoV-2 (21%), seasonal coronavirus (18%), parainfluenza (14%), RSV (6%), adenovirus (5%), and influenza (4%). Only 5% (5/109) of RVI episodes were asymptomatic with 2 episodes of SARS-CoV-2, 2 episodes of seasonal coronavirus, and 1 episode of parainfluenza. The median duration of a symptomatic episode was 1 week (range 1-18 weeks). During the follow-up period, 3 participants in cohort 1 satisfied diagnostic criteria for BOS, of which 2 had antecedent RVI. Conclusion The first year of this observational study revealed frequent symptomatic RVIs and a low rate of asymptomatic RVIs in patients at high risk for BOS. Ongoing follow-up and analyses will evaluate the impact of common RVI on BOS development and progression after HCT.

Electrochemical biosensors have shown great potential for simple, fast, and cost-effective point-of-care diagnostic tools. However, direct analysis of complex biological fluids such as plasma has been limited by the loss of sensitivity caused by biofouling. By increasing the surface area, the nanostructured electrode can improve detection sensitivity. However, like a double-edged sword, a large surface area increases the nonspecific adsorption of contaminating proteins. The use of nanoporous structures may prevent fouling proteins. However, there is no straightforward approach for creating nanostructured and nanoporous surfaces compatible with microfabricated thin-film electrodes. Herein, the preferential etching of chloride and surfactant-assisted anisotropic gold reduction to create homogeneous, nanostructured, and nanoporous gold electrodes is demonstrated, yielding a 190 ± 20 times larger surface area within a minute without using templates. This process, "surfactant-based electrochemical etch-deposit interplay for nanostructure/nanopore growth" (SEEDING), on electrodes enhances the sensitivity and antibiofouling capabilities of amperometric biosensors, enabling direct analysis of tumor-derived extracellular vesicles (tEVs) in complex biofluids with a limit of detection of 300 tEVs µL^-1 from undiluted plasma and good discrimination between patients with prostate cancer from healthy ones with an area under the curve of 0.91 in urine and 0.90 in plasma samples.

Abstract Background Bronchiolitis obliterans syndrome (BOS) is a late complication after allogeneic hematopoietic cell transplantation (HCT). While retrospective evidence shows symptomatic respiratory viral infections (RVI) may contribute to the development and progression of BOS, a similar role for asymptomatic RVI is unknown. This study aims to assess the incidence of asymptomatic and symptomatic RVI in a high-risk population. Methods Allogeneic HCT recipients with new-onset chronic graft-versus-host disease (cohort 1, at risk for BOS) and new onset BOS (cohort 2) were enrolled in an ongoing multi-center prospective longitudinal study (NCT05250037) to assess the role of RVI in development of BOS. Participants conducted weekly handheld home spirometry, weekly symptom surveys, and submitted self-administered nasal swabs for respiratory viral PCR testing every two weeks; additional swabs were prompted if symptom survey score reached ≥2. An asymptomatic episode was defined as one or more consecutive positive viral swab with a symptom score of 0-1 and no positive symptoms in the 2 weeks before or after. A symptomatic RVI episode was defined as the first positive PCR to the final positive PCR with any associated symptom score ≥2, allowing no more than ≥4 weeks or 2 negative samples between any 2 consecutive positive PCR samples. Results From March 2022 to July 2023, 73 participants were enrolled (cohort 1: n=50; cohort 2: n=23), with 47 completing one year of follow-up. Compliance was 68% for surveys and 62% for nasal swabs. Twenty percent of 744 swabs tested positive for ≥1 virus. Of 43 participants who tested positive for RVI, 21% (9/43) had 1 episode, 21% (9/43) had 2 episodes, and 52% (25/43) had ≥3 episodes. Ninety-five percent of RVI episodes (104/109) were symptomatic, most commonly rhinovirus (22%), followed by SARS-CoV-2 (21%), seasonal coronavirus (18%), parainfluenza (14%), RSV (6%), adenovirus (5%), and influenza (4%). Only 5% (5/109) of RVI episodes were asymptomatic with 2 episodes of SARS-CoV-2, 2 episodes of seasonal coronavirus, and 1 episode of parainfluenza. The median duration of a symptomatic episode was 1 week (range 1-18 weeks). During the follow-up period, 3 participants in cohort 1 satisfied diagnostic criteria for BOS, of which 2 had antecedent RVI. Conclusion The first year of this observational study revealed frequent symptomatic RVIs and a low rate of asymptomatic RVIs in patients at high risk for BOS. Ongoing follow-up and analyses will evaluate the impact of common RVI on BOS development and progression after HCT.

Electrochemical biosensors have shown great potential for simple, fast, and cost-effective point-of-care diagnostic tools. However, direct analysis of complex biological fluids such as plasma has been limited by the loss of sensitivity caused by biofouling. By increasing the surface area, the nanostructured electrode can improve detection sensitivity. However, like a double-edged sword, a large surface area increases the nonspecific adsorption of contaminating proteins. The use of nanoporous structures may prevent fouling proteins. However, there is no straightforward approach for creating nanostructured and nanoporous surfaces compatible with microfabricated thin-film electrodes. Herein, the preferential etching of chloride and surfactant-assisted anisotropic gold reduction to create homogeneous, nanostructured, and nanoporous gold electrodes is demonstrated, yielding a 190 ± 20 times larger surface area within a minute without using templates. This process, "surfactant-based electrochemical etch-deposit interplay for nanostructure/nanopore growth" (SEEDING), on electrodes enhances the sensitivity and antibiofouling capabilities of amperometric biosensors, enabling direct analysis of tumor-derived extracellular vesicles (tEVs) in complex biofluids with a limit of detection of 300 tEVs µL^-1 from undiluted plasma and good discrimination between patients with prostate cancer from healthy ones with an area under the curve of 0.91 in urine and 0.90 in plasma samples.

Electrochemical Pollination In article number 2200981, Yoon-Kyoung Cho and co-workers discuss electrochemical roughening, a method based on the etching and redeposition of material from a flat surface, to create homogeneous, nanostructured, and nanoporous gold electrodes. The process is not efficient, as most of the etched material typically diffuses away. However, the addition of surfactant generates micelles that can electrostatically capture the freshly etched material and, in a process that evokes pollination from bumblebees, increase redeposition efficiency.

Raman spectroscopy offers non-destructive and highly sensitive molecular insights into bacterial species, making it a valuable tool for detection, identification, and antibiotic susceptibility testing. However, achieving clinically relevant accuracy, quantitative data, and reproducibility remains challenging due to the dominance of bulk signals and the uncontrollable heterogeneity of analytes. In this study, we introduce an innovative diagnostic tool: a plasmonic fidget spinner (P-FS) incorporating a nitrocellulose membrane integrated with a metallic feature, referred to as a nanoplasmonic-enhanced matrix, designed for simultaneous bacterial filtration and detection. We developed a method to fabricate a plasmonic array patterned nitrocellulose membrane using photolithography, which is then integrated with a customized fidget spinner. Testing the P-FS device with various bacterial species (E. coli 25922, S. aureus 25923, E. coli MG1655, Lactobacillus brevis, and S. mutans 3065) demonstrated successful identification based on their unique Raman fingerprints. The bacterial interface with regions within the plasmonic array, where the electromagnetic field is most intensely concentrated-called nanoplasmonic hotspots-on the P-FS significantly enhances sensitivity, enabling more precise detection. SERS intensity mappings from the Raman spectrometer are transformed into digital signals using a threshold-based approach to identify and quantify bacterial distribution. Given the P-FS's ability to enhance vibrational signatures and its scalable fabrication under routine conditions, we anticipate that nanoplasmonic-enhanced Raman spectroscopy-utilizing nanostructures made from metals (specifically gold and silver) deposited onto a nitrocellulose membrane to amplify Raman scattering signals-will become the preferred technology for reliable and ultrasensitive detection of various analytes, including those crucial to human health, with strong potential for transitioning from laboratory research to clinical applications.

Tumor-derived extracellular vesicle (tEV)-associated RNAs hold promise as diagnostic biomarkers, but their clinical use is hindered by the rarity of tEVs among nontumor EVs. Here, we present EV-CLIP, a highly sensitive droplet-based digital method for profiling EV RNA. EV-CLIP utilizes the fusion of EVs with charged liposomes (CLIPs) in a microfluidic chip. Optimized CLIP surface charge enables exceptional sensitivity and selectivity for EV-derived miRNAs and mRNAs. This approach streamlines detection with minimal plasma volume (20 μL) and eliminates the need for prior EV isolation or RNA preparation, preventing loss of EVs or RNA. In testing with 83 patient samples, EV-CLIP detected EGFR L858R and T790M mutations with high AUC values of 1.0000 and 0.9784, respectively. Its success in serial monitoring during chemotherapy highlights its potential for precise quantification of rare EV subpopulations, facilitating the exploration of single EV RNA content and enhancing understanding of diverse EV populations in various disease states.

Extracellular vesicles (EVs) are emerging as crucial biomarkers in cancer diagnostics and therapeutics with their heterogeneity presenting both challenges and opportunities in prostate cancer research. However, existing methods for isolating and characterizing EV subtypes have been limited by inefficient separation and inadequate proteomic analysis. Here we show an optimized centrifugal microfluidic device, Exodisc, that efficiently isolates large quantities of EV subtypes from particle-enriched medium, enabling comprehensive proteomic analysis of small (EV-S, 20-200 nm) and large (EV-L, >200 nm) EVs. Using this device, we successfully separated EV-S and EV-L from prostate cancer cell lines LNCaP and PC3. Mass spectrometry-based proteomics revealed that EV proteins reflect parental cell characteristics more than EV size, with EV-L demonstrating increased expression of PSMA-correlated proteins. Our optimized protocol addresses challenges in EV isolation and characterization, providing a more effective method for studying cellular and molecular mechanisms of specific EV subtypes. This study extends the potential use of EVs as a liquid biopsy for cancer theranostics, paving the way for more precise isolation of EV subtypes and potentially leading to improved biomarker discovery and the development of personalized treatments.

Open Microfluidic Platform The cover features an innovative open microfluidic platform that enables direct culture and analysis of tumor spheroids interacting with blood vessels, developed by Yoon‐Kyoung Cho and co‐workers. The image showcases the high‐throughput chip where cancer spheroids (detailed in circular inset) can be grown and retrieved for genetic analysis while studying their interactions with the underlying vasculature, advancing our understanding of tumor drug resistance mechanisms. More details can be found in article number 2410659. [Image: see text]

The advent of Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)-based genome editing technologies has opened up groundbreaking possibilities for treating a wide spectrum of genetic disorders and diseases. However, the success of these technologies relies heavily on the development of efficient and safe delivery systems. Among the most promising approaches are natural and synthetic nanocarrier-mediated delivery systems, including viral vectors, extracellular vesicles (EVs), engineered cellular membrane particles, liposomes, and various nanoparticles. These carriers enhance the efficacy of the CRISPR system by providing a unique combination of efficiency, specificity, and reduced immunogenicity. Synthetic carriers such as liposomes and nanoparticles facilitate CRISPR delivery with high reproducibility and customizable functions. Viral vectors, renowned for their high transduction efficiency and broad tropism, serve as powerful vehicles for delivering CRISPR components to various cell types. EVs, as natural carriers of RNA and proteins, offer a stealth mechanism to evade immune detection, allowing for the targeted delivery of genome editors with minimal off-target effects. Engineered cellular membrane particles further improve delivery by simulating the cellular environment, enhancing uptake, and minimizing immune response. This review explores the innovative integration of CRISPR genome editors with various nanocarrier systems, focusing on recent advancements, applications, and future directions in therapeutic genome editing.

Although antibody-drug conjugates offer advanced targeted anticancer therapy that overcomes the limitations of conventional chemotherapy and therapeutic antibodies, they are restricted in their capacity to carry multiple hydrophobic payloads. Protein nanocages have emerged as versatile therapeutic platforms for targeted drug delivery, offering advantages like precise molecular assembly, biocompatibility, and multivalent targeting. This study presents the development of engineered E2 nanocages functionalized with anti-HER2 single-chain variable fragments (scFv) using the SpyTag/SpyCatcher ligation system to achieve controlled scFv display valency. The results demonstrate that increasing anti-HER2 scFv valency enhances HER2 binding affinity via avidity effects, with the highest valency nanocages showing the highest binding avidity. Furthermore, cysteine residues were introduced into the E2 nanocages to enable conjugation with monomethyl auristatin E (MMAE) through maleimide chemistry, achieving efficient drug loading. The resulting MMAE-conjugated nanocages displayed potent, subnanomolar cytotoxicity in HER2-positive SKBR3 and BT-474 cell lines while sparing HER2-negative MDA-MB-231 cells at concentrations up to 1 nM. These results underscore the critical role of scFv valency in enhancing HER2 targeting and highlight the potential of E2 protein nanocages as specific, potent platforms for targeted cancer therapy. In this study, we developed an enhanced targeted drug delivery system using E2 nanocages and scFv with SpyCatcher/SpyTag ligation to regulate binding avidity and encapsulate hydrophobic drugs. The modular design and pH-sensitive dissociation of these nanocages establish a foundation for next-generation precision medicine strategies.