RESUMO
In recent years, steady progress has been made in synthesizing and characterizing engineered nanoparticles, resulting in several approved drugs and multiple promising candidates in clinical trials. Regulatory agencies such as the Food and Drug Administration and the European Medicines Agency released important guidance documents facilitating nanoparticle-based drug product development, particularly in the context of liposomes and lipid-based carriers. Even with the progress achieved, it is clear that many barriers must still be overcome to accelerate translation into the clinic. At the recent conference workshop "Mechanisms and Barriers in Nanomedicine" in May 2023 in Colorado, U.S.A., leading experts discussed the formulation, physiological, immunological, regulatory, clinical, and educational barriers. This position paper invites open, unrestricted, nonproprietary discussion among senior faculty, young investigators, and students to trigger ideas and concepts to move the field forward.
Assuntos
Nanomedicina , Humanos , Portadores de Fármacos/química , Lipossomos/química , Nanopartículas/química , Estados UnidosRESUMO
Detecting nucleic acids at ultralow concentrations is critical for research and clinical applications. Particle-based assays are commonly used to detect nucleic acids. However, DNA hybridization on particle surfaces is inefficient due to the instability of tethered sequences, which negatively influences the assay's detection sensitivity. Here, we report a method to stabilize sequences on particle surfaces using a double-stranded linker at the 5' end of the tethered sequence. We termed this method Rigid Double Stranded Genomic Linkers for Improved DNA Analysis (RIGID-DNA). Our method led to a 3- and 100-fold improvement of the assays' clinical and analytical sensitivity, respectively. Our approach can enhance the hybridization efficiency of particle-based assays without altering existing assay workflows. This approach can be adapted to other platforms and surfaces to enhance the detection sensitivity.
Assuntos
DNA , Limite de Detecção , Hibridização de Ácido Nucleico , DNA/química , Humanos , Conformação de Ácido NucleicoRESUMO
Histological hematoxylin and eosin-stained (H&E) tissue sections are used as the gold standard for pathologic detection of cancer, tumor margin detection, and disease diagnosis. Producing H&E sections, however, is invasive and time-consuming. While deep learning has shown promise in virtual staining of unstained tissue slides, true virtual biopsy requires staining of images taken from intact tissue. In this work, we developed a micron-accuracy coregistration method [micro-registered optical coherence tomography (OCT)] that can take a two-dimensional (2D) H&E slide and find the exact corresponding section in a 3D OCT image taken from the original fresh tissue. We trained a conditional generative adversarial network using the paired dataset and showed high-fidelity conversion of noninvasive OCT images to virtually stained H&E slices in both 2D and 3D. Applying these trained neural networks to in vivo OCT images should enable physicians to readily incorporate OCT imaging into their clinical practice, reducing the number of unnecessary biopsy procedures.
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Redes Neurais de Computação , Tomografia de Coerência Óptica , Humanos , Tomografia de Coerência Óptica/métodos , Biópsia , Imageamento TridimensionalRESUMO
Gas vesicles (GVs) are genetically encoded, air-filled protein nanostructures of broad interest for biomedical research and clinical applications, acting as imaging and therapeutic agents for ultrasound, magnetic resonance, and optical techniques. However, the biomedical applications of GVs as systemically injectable nanomaterials have been hindered by a lack of understanding of GVs' interactions with blood components, which can significantly impact in vivo behavior. Here, we investigate the dynamics of GVs in the bloodstream using a combination of ultrasound and optical imaging, surface functionalization, flow cytometry, and mass spectrometry. We find that erythrocytes and serum proteins bind to GVs and shape their acoustic response, circulation time, and immunogenicity. We show that by modifying the GV surface we can alter these interactions and thereby modify GVs' in vivo performance. These results provide critical insights for the development of GVs as agents for nanomedicine.
Assuntos
Nanoestruturas , Proteínas , Ultrassonografia/métodos , Proteínas/química , Meios de Contraste , Nanoestruturas/química , Imageamento por Ressonância Magnética/métodosRESUMO
The effective treatment of patients with cancer hinges on the delivery of therapeutics to a tumor site. Nanoparticles provide an essential transport system. We present 5 principles to consider when designing nanoparticles for cancer targeting: (a) Nanoparticles acquire biological identity in vivo, (b) organs compete for nanoparticles in circulation, (c) nanoparticles must enter solid tumors to target tumor components, (d) nanoparticles must navigate the tumor microenvironment for cellular or organelle targeting, and (e) size, shape, surface chemistry, and other physicochemical properties of nanoparticles influence their transport process to the target. This review article describes these principles and their application for engineering nanoparticle delivery systems to carry therapeutics to tumors or other disease targets.
RESUMO
Gas vesicles (GVs) are genetically encoded, air-filled protein nanostructures of broad interest for biomedical research and clinical applications, acting as imaging and therapeutic agents for ultrasound, magnetic resonance, and optical techniques. However, the biomedical applications of GVs as a systemically injectable nanomaterial have been hindered by a lack of understanding of GVs' interactions with blood components, which can significantly impact in vivo performance. Here, we investigate the dynamics of GVs in the bloodstream using a combination of ultrasound and optical imaging, surface functionalization, flow cytometry, and mass spectrometry. We find that erythrocytes and serum proteins bind to GVs and shape their acoustic response, circulation time, and immunogenicity. We show that by modifying the GV surface, we can alter these interactions and thereby modify GVs' in vivo performance. These results provide critical insights for the development of GVs as agents for nanomedicine.
RESUMO
Nanoparticles enter tumours through endothelial cells, gaps or other mechanisms, but how they exit is unclear. The current paradigm states that collapsed tumour lymphatic vessels impair the exit of nanoparticles and lead to enhanced retention. Here we show that nanoparticles exit the tumour through the lymphatic vessels within or surrounding the tumour. The dominant lymphatic exit mechanism depends on the nanoparticle size. Nanoparticles that exit the tumour through the lymphatics are returned to the blood system, allowing them to recirculate and interact with the tumour in another pass. Our results enable us to define a mechanism of nanoparticle delivery to solid tumours alternative to the enhanced permeability and retention effect. We call this mechanism the active transport and retention principle. This delivery principle provides a new framework to engineer nanomedicines for cancer treatment and detection.
Assuntos
Vasos Linfáticos , Nanopartículas , Neoplasias , Humanos , Células Endoteliais , Neoplasias/tratamento farmacológico , Sistemas de Liberação de MedicamentosRESUMO
Nanobio interaction studies have generated a significant amount of data. An important next step is to organize the data and design computational techniques to analyze the nanobio interactions. Here we developed a computational technique to correlate the nanoparticle spatial distribution within heterogeneous solid tumors. This approach led to greater than 88% predictive accuracy of nanoparticle location within a tumor tissue. This proof-of-concept study shows that tumor heterogeneity might be defined computationally by the patterns of biological structures within the tissue, enabling the identification of tumor patterns for nanoparticle accumulation.
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Nanopartículas , Neoplasias , Humanos , Nanopartículas/químicaRESUMO
Highly functionalized skeletons of macrolide natural products gain access to rare spatial arrangements of atoms, where changes in stereochemistry can have a profound impact on the structure and function. Spliceosome modulators present a unique consensus motif, with the majority targeting a key interface within the SF3B spliceosome complex. Our recent preparative-scale synthetic campaign of 17S-FD-895 provided unique access to stereochemical analogues of this complex macrolide. Here, we report on the preparation and systematic activity evaluation of multiple FD-895 analogues. These studies examine the effects of modifications at specific stereocenters within the molecule and highlight future directions for medicinal chemical optimization of spliceosome modulators.
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Macrolídeos , Macrolídeos/farmacologiaRESUMO
Designing diagnostic assays to genotype rapidly mutating viruses remains a challenge despite the overall improvements in nucleic acid detection technologies. RT-PCR and next-generation sequencing are unsuitable for genotyping during outbreaks or in point-of-care detection due to their infrastructure requirements and longer turnaround times. We developed a quantum dot barcode multiplexing system to genotype mutated viruses. We designed multiple quantum dot barcodes to target conserved, wildtype, and mutated regions of SARS-CoV-2. We calculated ratios of the signal output from different barcodes that enabled SARS-CoV-2 detection and identified SARS-CoV-2 variant strains from a sample. We detected different sequence types, including conserved genes, nucleotide deletions, and single nucleotide substitutions. Our system detected SARS-CoV-2 patient specimens with 98% sensitivity and 94% specificity across 91 patient samples. Further, we leveraged our barcoding and ratio system to track the emergence of the N501Y SARS-CoV-2 mutation from December 2020 to May 2021 and demonstrated that the more transmissible N501Y mutation started to dominate infections by April 2021. Our barcoding and signal ratio approach can genotype viruses and track the emergence of viral mutations in a single diagnostic test. This technology can be extended to tracking other viruses. Combined with smartphone detection technologies, this assay can be adapted for point-of-care tracking of viral mutations in real time.
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COVID-19 , Ácidos Nucleicos , Humanos , SARS-CoV-2/genética , COVID-19/diagnóstico , Genótipo , Nucleotídeos , MutaçãoRESUMO
Pediatric acute myeloid leukemia (pAML) is typified by high relapse rates and a relative paucity of somatic DNA mutations. Although seminal studies show that splicing factor mutations and mis-splicing fuel therapy-resistant leukemia stem cell (LSC) generation in adults, splicing deregulation has not been extensively studied in pAML. Herein, we describe single-cell proteogenomics analyses, transcriptome-wide analyses of FACS-purified hematopoietic stem and progenitor cells followed by differential splicing analyses, dual-fluorescence lentiviral splicing reporter assays, and the potential of a selective splicing modulator, Rebecsinib, in pAML. Using these methods, we discover transcriptomic splicing deregulation typified by differential exon usage. In addition, we discover downregulation of splicing regulator RBFOX2 and CD47 splice isoform upregulation. Importantly, splicing deregulation in pAML induces a therapeutic vulnerability to Rebecsinib in survival, self-renewal, and lentiviral splicing reporter assays. Taken together, the detection and targeting of splicing deregulation represent a potentially clinically tractable strategy for pAML therapy.
Assuntos
Leucemia Mieloide Aguda , Células-Tronco , Adulto , Criança , Humanos , Splicing de RNA/genética , Leucemia Mieloide Aguda/tratamento farmacológico , Leucemia Mieloide Aguda/genética , Isoformas de Proteínas/genética , Mutação , Fatores de Processamento de RNA/genética , Proteínas Repressoras/genéticaRESUMO
Nanoparticles travel through blood vessels to reach disease sites, but the local environment they encounter may affect their surface chemistry and cellular interactions. Here, we found that as nanoparticles transit through injured blood vessels they may interact with a highly localized concentration of platelet factor 4 proteins released from activated platelets. The platelet factor 4 binds to the nanoparticle surface and interacts with heparan sulfate proteoglycans on endothelial cells, and induces uptake. Understanding nanoparticle interactions with blood proteins and endothelial cells during circulation is critical to optimizing their design for diseased tissue targeting and delivery.
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Nanopartículas , Coroa de Proteína , Células Endoteliais/metabolismo , Fator Plaquetário 4/metabolismo , Coroa de Proteína/metabolismo , Plaquetas/metabolismoRESUMO
Nanoparticles can reduce cytotoxicity, increase circulation time and increase accumulation in tumours compared to free drug. However, the value of using nanoparticles for carrying small molecules to treat tumours at the cellular level has been poorly established. Here we conducted a cytodistribution analysis on Doxorubicin-treated and Doxil-treated tumours to delineate the differences between the small molecule therapeutic Doxorubicin and its packaged liposomal formulation Doxil. We found that Doxil kills more cancer cells, macrophages and neutrophils in the 4T1 breast cancer tumour model, but there is delayed killing compared to its small molecule counterpart Doxorubicin. The cellular interaction with Doxil has slower uptake kinetics and the particles must be degraded to release the drug and kill the cells. We also found that macrophages and neutrophils in Doxil-treated tumours repopulated faster than cancer cells during the relapse phase. While researchers conventionally use tumour volume and animal survival to determine a therapeutic effect, our results show diverse cell killing and a greater amount of cell death in vivo after Doxil liposomes are administered. We conclude that the fate and behaviour of the nanocarrier influences its effectiveness as a cancer therapy. Further investigations on the interactions between different nanoparticle designs and the tumour microenvironment components will lead to more precise engineering of nanocarriers to selectively kill tumour cells and prolong the therapeutic effect.
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Nanopartículas , Neoplasias , Animais , Lipossomos/uso terapêutico , Microambiente Tumoral , Doxorrubicina/uso terapêutico , Neoplasias/tratamento farmacológico , PolietilenoglicóisRESUMO
Diagnostic assays are commonly performed in multiple steps, where reagents are added at specific times and concentrations into a reaction chamber. The reagents require storage, preparation, and addition in the correct sequence and amount. These steps rely on trained technicians and instrumentation to perform each task. The reliance on such resources hinders the use of these diagnostic assays by lay users. We developed a tablet that can sequentially introduce prequantified lyophilized diagnostic reagents at specific time points for a multistep assay. We designed the tablet to have multiple layers using cellulose-grade polymers, such as microcrystalline cellulose and hydroxypropyl cellulose. Our formulation allows each layer to dissolve at a controlled rate to introduce reagents into the solution sequentially. The release rate is controlled by modulating the compression force or chemical formulation of the layer. Controlling the reagent release time is important because different assays have specific times when reagents need to be added. As proof of concept, we demonstrated two different assays with our tablet system. Our tablet detected nucleic acid target (tpp47 gene from Treponema pallidum) and nitrite ions in an aqueous sample without user intervention. Our multilayer tablets can simplify multistep assay processes.