Drug Screening Applications Research Articles

Biofilm Formation Capability of Clinical Helicobacter pylori Isolates on MKN-45 Cells

Background: In vitro biofilm formation of H. pylori is demonstrated; however, its potential role in the persistent infection of the human stomach has not yet been addressed. Objectives: The aim of this study was to assess the biofilm formation of clinical H. pylori isolates on an epithelial cell line, a line that produces mucin. Methods: H. pylori isolates consisting of an efficient (19B) and a weak (4B) biofilm formation ability, were selected from screening of the clinical isolates. Their adhesion index was determined after 2h incubation with the semi-confluent monolayers of MKN-45 cells. Their biofilm formation was evaluated after 24 and 72 h incubation with MKN-45 cells using a modified adherence assay developed in this work. Production of biofilm was quantitatively assessed by CFU enumeration and qualitatively by the immunofluorescence, and scanning-electron-microscopic (SEM) methods. Due to the importance of mucin in the binding of H. pylori and biofilm formation, the binding strength of the mucin binding protein, MUC5AC, and MUC1 with docking was investigated using cluspro webserver. Results: Using MKN-45 epithelial cell line as a model, significant differences were observed between the adhesion index of 19B and 4B isolates. After 24h, both isolates were able to form biofilms with significantly higher numbers of CFU for the 19B isolate. These results were confirmed by immunofluorescence and SEM such that after 24h, a cluster of coccoid bacteria on the MKN-45 cells in the form of microcolonies was observed. The docking results showed that MUC5AC demonstrated the most favorable interaction with H. pylori urease and BabA with docking energy scores of -931.1 and -906.3 kcal.mol-1, respectively. Conclusions: By developing an appropriate in situ biofilm assay, we investigated biofilm formation by clinical H. pylori isolates on the MKN-45 epithelial cell line. The establishment of such an in-situ model for studying the biofilm formation ability of clinical isolates can also be used to study cell-bacteria interactions in the context of a complex biofilm and also as a model for drug screening applications.

3D bioprinting: novel approaches for engineering complex human tissue equivalents and drug testing.

Conventional approaches in drug development involve testing on 2D-cultured mammalian cells, followed by experiments in rodents. Although this is the common strategy, it has significant drawbacks: in 2D cell culture with human cells, the cultivation at normoxic conditions on a plastic or glass surface is an artificial situation that significantly changes energy metabolism, shape and intracellular signaling, which in turn directly affects drug response. On the other hand, rodents as the most frequently used animal models have evolutionarily separated from primates about 100 million years ago, with significant differences in physiology, which frequently leads to results not reproducible in humans. As an alternative, spheroid technology and micro-organoids have evolved in the last decade to provide 3D context for cells similar to native tissue. However, organoids used for drug testing are usually just in the 50–100 micrometers range and thereby too small to mimic micro-environmental tissue conditions such as limited nutrient and oxygen availability. An attractive alternative offers 3D bioprinting as this allows fabrication of human tissue equivalents from scratch with hollow structures for perfusion and strict spatiotemporal control over the deposition of cells and extracellular matrix proteins. Thereby, tissue surrogates with defined geometry are fabricated that offer unique opportunities in exploring cellular cross-talk, mechanobiology and morphogenesis. These tissue-equivalents are also very attractive tools in drug testing, as bioprinting enables standardized production, parallelization, and application-tailored design of human tissue, of human disease models and patient-specific tissue avatars. This review, therefore, summarizes recent advances in 3D bioprinting technology and its application for drug screening.

Induction and expansion of human PRRX1+ limb-bud-like mesenchymal cells from pluripotent stem cells.

Current protocols for the differentiation of human pluripotent stem cells (hPSCs) into chondrocytes do not allow for the expansion of intermediate progenitors so as to prospectively assess their chondrogenic potential. Here we report a protocol that leverages PRRX1-tdTomato reporter hPSCs for the selective induction of expandable and ontogenetically defined PRRX1+ limb-bud-like mesenchymal cells under defined xeno-free conditions, and the prospective assessment of the cells' chondrogenic potential via the cell-surface markers CD90, CD140B and CD82. The cells, which proliferated stably and exhibited the potential to undergo chondrogenic differentiation, formed hyaline cartilaginous-like tissue commensurate to their PRRX1-expression levels. Moreover, we show that limb-bud-like mesenchymal cells derived from patient-derived induced hPSCs can be used to identify therapeutic candidates for type II collagenopathy and we developed a method to generate uniformly sized hyaline cartilaginous-like particles by plating the cells on culture dishes coated with spots of a zwitterionic polymer. PRRX1+ limb-bud-like mesenchymal cells could facilitate the mass production of chondrocytes and cartilaginous tissues for applications in drug screening and tissue engineering.

Numerical Modeling of Flow through the Vertical Rectangular Microchannel for Drug Screening Applications

A numerical study of flow in the rectangular vertical microchannel for drug screening applications is described. The flow characteristics through the microchannel are presented by incompressible Naiver-Stroke equations. Flow rate is evaluated from numerical modeling to simulate the drug flow through the microchannel. Here in this study, computational fluid dynamics based finite element method commercial software COMSOL is used for simulation of drug flow through the artery for cancer drug screening applications. The simulation work also presents the diffusion velocity at the intersection area between the drug and cancer cells..

Generation of Hepatobiliary Cell Lineages from Human Induced Pluripotent Stem Cells: Applications in Disease Modeling and Drug Screening.

The possibility to reproduce key tissue functions in vitro from induced pluripotent stem cells (iPSCs) is offering an incredible opportunity to gain better insight into biological mechanisms underlying development and disease, and a tool for the rapid screening of drug candidates. This review attempts to summarize recent strategies for specification of iPSCs towards hepatobiliary lineages —hepatocytes and cholangiocytes—and their use as platforms for disease modeling and drug testing. The application of different tissue-engineering methods to promote accurate and reliable readouts is discussed. Space is given to open questions, including to what extent these novel systems can be informative. Potential pathways for improvement are finally suggested.

To Better Generate Organoids, What Can We Learn From Teratomas?

The genomic profile of animal models is not completely matched with the genomic profile of humans, and 2D cultures do not represent the cellular heterogeneity and tissue architecture found in tissues of their origin. Derived from 3D culture systems, organoids establish a crucial bridge between 2D cell cultures and in vivo animal models. Organoids have wide and promising applications in developmental research, disease modeling, drug screening, precision therapy, and regenerative medicine. However, current organoids represent only single or partial components of a tissue, which lack blood vessels, native microenvironment, communication with near tissues, and a continuous dorsal-ventral axis within 3D culture systems. Although efforts have been made to solve these problems, unfortunately, there is no ideal method. Teratoma, which has been frequently studied in pathological conditions, was recently discovered as a new in vivo model for developmental studies. In contrast to organoids, teratomas have vascularized 3D structures and regions of complex tissue-like organization. Studies have demonstrated that teratomas can be used to mimic multilineage human development, enrich specific somatic progenitor/stem cells, and even generate brain organoids. These results provide unique opportunities to promote our understanding of the vascularization and maturation of organoids. In this review, we first summarize the basic characteristics, applications, and limitations of both organoids and teratomas and further discuss the possibility that in vivo teratoma systems can be used to promote the vascularization and maturation of organoids within an in vitro 3D culture system.

Liver-Heart on chip models for drug safety.

Current pre-clinical models to evaluate drug safety during the drug development process (DDP) mainly rely on traditional two-dimensional cell cultures, considered too simplistic and often ineffective, or animal experimentations, which are costly, time-consuming, and not truly representative of human responses. Their clinical translation thus remains limited, eventually causing attrition and leading to high rates of failure during clinical trials. These drawbacks can be overcome by the recently developed Organs-on-Chip (OoC) technology. OoC are sophisticated in vitro systems capable of recapitulating pivotal architecture and functionalities of human organs. OoC are receiving increasing attention from the stakeholders of the DDP, particularly concerning drug screening and safety applications. When a drug is administered in the human body, it is metabolized by the liver and the resulting compound may cause unpredicted toxicity on off-target organs such as the heart. In this sense, several liver and heart models have been widely adopted to assess the toxicity of new or recalled drugs. Recent advances in OoC technology are making available platforms encompassing multiple organs fluidically connected to efficiently assess and predict the systemic effects of compounds. Such Multi-Organs-on-Chip (MOoC) platforms represent a disruptive solution to study drug-related effects, which results particularly useful to predict liver metabolism on off-target organs to ultimately improve drug safety testing in the pre-clinical phases of the DDP. In this review, we focus on recently developed liver and heart on chip systems for drug toxicity testing. In addition, MOoC platforms encompassing connected liver and heart tissues have been further reviewed and discussed.

A versatile microfluidic tool for the 3D culture of HepaRG cells seeded at various stages of differentiation

The development of livers-on-a-chip aims to provide pharmaceutical companies with reliable systems to perform drug screening and toxicological studies. To that end, microfluidic systems are engineered to mimic the functions and architecture of this organ. In this context we have designed a device that reproduces series of liver microarchitectures, each permitting the 3D culture of hepatocytes by confining them to a chamber that is separated from the medium conveying channel by very thin slits. We modified the structure to ensure its compatibility with the culture of hepatocytes from different sources. Our device was adapted to the migratory and adhesion properties of the human HepaRG cell line at various stages of differentiation. Using this device, it was possible to keep the cells alive for more than 14 days, during which they achieved a 3D organisation and acquired or maintained their differentiation into hepatocytes. Albumin secretion as well as functional bile canaliculi were confirmed on the liver-on-a-chip. Finally, an acetaminophen toxicological assay was performed. With its multiple micro-chambers for hepatocyte culture, this microfluidic device architecture offers a promising opportunity to provide new tools for drug screening applications.

Abstract 1648: Checkpoint molecule profiling in tumor cell lines and immune cell lines for application in immuno-oncology drug screening

Abstract The development of cancer immunotherapies has emerged as an exciting new method in treating cancer. Inhibition of immune checkpoint is one of the most promising approaches to unleash the potential of the anti-tumor immune response. Immune checkpoint blockades are monoclonal antibodies that bind to checkpoint molecules on the surface of immune and tumor cells. Tumor cells utilize immune checkpoints to suppress immune response and increase survival. There are several established and novel checkpoint molecules being studied to create cancer immune therapeutics. PD-1 (Programmable Death-1) is one of the most studied of these checkpoint molecules and is the basis for several FDA approved therapeutics on the market. Although immune checkpoint blockades have exhibited anti-tumor effects in multiple cancer types, there are still challenges to overcome such as resistance and low response rate. In addition, clinical studies have shown that the efficacy of these treatments vary depending on cancer type. There is a need for comprehensive data on the expression levels of checkpoint molecules based on cancer type, which can be utilized to guide specific treatment plans and combinations. ATCC provides the scientific community a vast portfolio of human tumor and immune cell lines. In this study, we profiled a cross-section of those cell lines with a number of established and novel checkpoint molecules. FACS analysis was used to evaluate expression level of the checkpoint markers including, but not limited to, PD-1, CTLA4, LAG-3, TIGIT, and VISTA. HLA typing was also performed on those cell lines. With this profiling data, we further tested the responses of the cell lines to several on-the-market immunotherapy biologics, such as Pembrolizumab (anti-PD1), Nivolumab (anti-PD1), and Ipilimumab (anti-CTLA4). In our physiological relevant checkpoint assay system, an activated human CD4+ T-cell line and primary CD8+ T-cells were co-cultured with target cell lines with the addition of a serial dilution of checkpoint blockade treatment. We were able to demonstrate T-cell proliferation and increased cytokine expression based on immunotherapy antibody treatment. In summary, this study presents a comprehensive checkpoint molecule profiling data set of T cells lines and tumor cell lines derived from various cancer types that can be used to develop custom assays, as required. The combination of established cell lines and primary T cells offers a unique and effective approach in checkpoint assay development, which provides a valuable tool for studying checkpoint molecule interactions and screening biologics as cancer immunotherapy treatments. Citation Format: Brian Della Fera, Haiyun Liu, Fang Tian. Checkpoint molecule profiling in tumor cell lines and immune cell lines for application in immuno-oncology drug screening [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 1648.

Applications of separation technology in novel coronavirus research, epidemic prevention and detection

新型冠状病毒肺炎(COVID-19)疫情的爆发给世界公共卫生安全带来前所未有的挑战。随着新型冠状病毒(SARS-CoV-2)相关研究的不断深入,众多分析检测技术相继被应用,推动了病毒检测、疫苗和创新疗法的研发,从而使疫情早日得到控制。分离技术作为生命科学、医学、药学领域的关键技术,操作简单,分离效率高,选择性强,在新型冠状病毒的分离、检测、诊疗及防疫中起到不可替代的作用。该文以SARS-CoV-2或COVID-19为关键词在ISI Web of Science中进行主题检索,归纳了2020年度新型冠状病毒相关的研究论文,简要介绍主要的研究方向,并对国际顶级学术期刊Nature, Science, Cell的论文发表情况进行了统计。通过检索影响因子较高的期刊,综述了新型冠状病毒研究中主要应用的分离技术,并从亲和色谱和尺寸排阻色谱、液相色谱、磁珠分离、离心、微纳分离以及电泳6个方面进行说明。综述统计了亲和色谱和尺寸排阻色谱纯化的病毒相关蛋白,并介绍了其在新型冠状病毒传播、感染机制以及药物筛选中的应用;介绍了液相色谱对病毒候选药物评估以及复杂基质中单一成分的鉴定;介绍了磁珠分离在细胞分离、核酸提取和免疫学检测中的应用;介绍了离心对病毒颗粒、细胞以及血清的分离;介绍了微纳分离结合其他技术以实现病毒蛋白的高灵敏检测;简要介绍了电泳在聚合酶链式反应(PCR)产物分析中的应用。该文综述了2020年度新型冠状病毒研究和防疫检测中分离技术的应用情况,分析了分离技术在新型冠状病毒检测中发挥的作用,旨在为从事分离研究的科研工作者提供一些参考。

Human iPSCs and Genome Editing Technologies for Precision Cardiovascular Tissue Engineering.

Induced pluripotent stem cells (iPSCs) originate from the reprogramming of adult somatic cells using four Yamanaka transcription factors. Since their discovery, the stem cell (SC) field achieved significant milestones and opened several gateways in the area of disease modeling, drug discovery, and regenerative medicine. In parallel, the emergence of clustered regularly interspaced short palindromic repeats (CRISPR)-associated protein 9 (CRISPR-Cas9) revolutionized the field of genome engineering, allowing the generation of genetically modified cell lines and achieving a precise genome recombination or random insertions/deletions, usefully translated for wider applications. Cardiovascular diseases represent a constantly increasing societal concern, with limited understanding of the underlying cellular and molecular mechanisms. The ability of iPSCs to differentiate into multiple cell types combined with CRISPR-Cas9 technology could enable the systematic investigation of pathophysiological mechanisms or drug screening for potential therapeutics. Furthermore, these technologies can provide a cellular platform for cardiovascular tissue engineering (TE) approaches by modulating the expression or inhibition of targeted proteins, thereby creating the possibility to engineer new cell lines and/or fine-tune biomimetic scaffolds. This review will focus on the application of iPSCs, CRISPR-Cas9, and a combination thereof to the field of cardiovascular TE. In particular, the clinical translatability of such technologies will be discussed ranging from disease modeling to drug screening and TE applications.

Ligand binding constants for human serum albumin evaluated by ratiometric analysis of DSC thermograms

This paper describes an expanded application of our recently reported method (Eskew et al., Analytical Biochemistry 621,1 2021) utilizing thermogram signals for thermal denaturation measured by differential scanning calorimetry. Characteristic signals were used to quantitatively evaluate ligand binding constants for human serum albumin. In our approach the ensemble of temperature dependent calorimetric responses for various protein-ligand mixtures and native HSA were compared, in a ratiometric manner, to extract binding constants and stoichiometries. Protein/ligand mixtures were prepared at various ligand concentrations and subjected to thermal denaturation analysis by calorimetry. Measurements provided the melting temperature, Tm, and free-energy ΔGcal(37°C) for melting ligand-bound Albumin as a function of ligand concentration. Concentration dependent behaviors of these parameters derived from protein/ligand mixtures were used to construct dose-response curves. Fitting of dose-response curves yielded quantitative evaluation of the ligand binding constant and semi-quantitative estimates of the binding stoichiometry. Many of the ligands had known binding affinity for Albumin with binding constants reported in the literature. Evaluated binding parameters for the ligands impressively agreed with reported literature values determined using other standard experimental methods. Results are reported for 29 drug ligands binding to Albumin. These validate our calorimetry-based process for applications in pre-clinical drug screening.

Microfluidic 3D Printing of a Photo-Cross-Linkable Bioink Using Insights from Computational Modeling.

Three-dimensional (3D) bioprinting of photo-cross-linkable hydrogel precursors has attracted great interest in various tissue engineering and drug screening applications, as the biochemical and biophysical properties of the resultant hydrogel structures can be tuned spatiotemporally to provide cells with physiologically relevant microenvironments. In particular, these bioinks benefit from great biofunctional versatility that can be designed to direct cells toward a desired behavior. Despite significant progress in the field, the 3D printing of cell-laden photo-cross-linkable bioinks with low polymer concentrations has remained a challenge, as rapidly stabilizing these bioinks and transforming them to hydrogel filaments is hindered by their low viscosity. Additionally, reaching an optimized print condition has often been challenging due to the large number of print parameters involved in 3D bioprinting setups. Therefore, computational modeling has occasionally been employed to understand the impact of various print parameters and reduce the time and resources required to determine these effects in experimental settings. Here, we report a novel 3D bioprinting strategy for fabricating hydrogel fibrous structures of gelatin methacryloyl (GelMA) with superior control over polymer concentration, particularly in a relatively low range from ∼1% (w/v) to 6% (w/v), using a microfluidic printhead. The printhead features a coaxial core-sheath flow, coupled with a photo-cross-linking system, allowing for the in situ cross-linking of GelMA and the generation of hydrogel filaments. A computational model was developed to determine the optimal ranges of process parameters and inform about the diffusive and fluid dynamic behavior of the coaxial flow. The cytocompatibility of the biofabrication system was determined via bioprinting cell-laden bioinks containing U87-MG cells. Notably, the established pipeline from computational modeling to bioprinting has great potential to be applied to a wide range of photo-cross-linkable bioinks to generate living tissues with various material and cellular characteristics.

Cost-Effective Real-Time Metabolic Profiling of Cancer Cell Lines for Plate-Based Assays

A fundamental phenotype of cancer cells is their metabolic profile, which is routinely described in terms of glycolytic and respiratory rates. Various devices and protocols have been designed to quantify glycolysis and respiration from the rates of acid production and oxygen utilization, respectively, but many of these approaches have limitations, including concerns about their cost-ineffectiveness, inadequate normalization procedures, or short probing time-frames. As a result, many methods for measuring metabolism are incompatible with cell culture conditions, particularly in the context of high-throughput applications. Here, we present a simple plate-based approach for real-time measurements of acid production and oxygen depletion under typical culture conditions that enable metabolic monitoring for extended periods of time. Using this approach, it is possible to calculate metabolic fluxes and, uniquely, describe the system at steady-state. By controlling the conditions with respect to pH buffering, O2 diffusion, medium volume, and cell numbers, our workflow can accurately describe the metabolic phenotype of cells in terms of molar fluxes. This direct measure of glycolysis and respiration is conducive for between-runs and even between-laboratory comparisons. To illustrate the utility of this approach, we characterize the phenotype of pancreatic ductal adenocarcinoma cell lines and measure their response to a switch of metabolic substrate and the presence of metabolic inhibitors. In summary, the method can deliver a robust appraisal of metabolism in cell lines, with applications in drug screening and in quantitative studies of metabolic regulation.

Experimental liver models: From cell culture techniques to microfluidic organs-on-chip.

The liver is one of the most studied organs of the human body owing to its central role in xenobiotic and drug metabolism. In recent decades, extensive research has aimed at developing in vitro liver models able to mimic liver functions to study pathophysiological clues in high-throughput and reproducible environments. Two-dimensional (2D) models have been widely used in screening potential toxic compounds but have failed to accurately reproduce the three-dimensionality (3D) of the liver milieu. To overcome these limitations, improved 3D culture techniques have been developed to recapitulate the hepatic native microenvironment. These models focus on reproducing the liver architecture, representing both parenchymal and nonparenchymal cells, as well as cell interactions. More recently, Liver-on-Chip (LoC) models have been developed with the aim of providing physiological fluid flow and thus achieving essential hepatic functions. Given their unprecedented ability to recapitulate critical features of the liver cellular environments, LoC have been extensively adopted in pathophysiological modelling and currently represent a promising tool for tissue engineering and drug screening applications. In this review, we discuss the evolution of experimental liver models, from the ancient 2D hepatocyte models, widely used for liver toxicity screening, to 3D and LoC culture strategies adopted for mirroring a more physiological microenvironment for the study of liver diseases.

Capture and Identification of Dual Specificity Mitogen-Activated Protein Kinase Kinase 7 as a Direct Proteomic Target of Berberine to Affect the c-JunN-Terminal Kinase Pathway

Capture and Identification of Dual Specificity Mitogen-Activated Protein Kinase Kinase 7 as a Direct Proteomic Target of Berberine to Affect the c-JunN-Terminal Kinase Pathway

Electrical stimulation of neonatal rat cardiomyocytes using conductive polydopamine-reduced graphene oxide-hybrid hydrogels for constructing cardiac microtissues

The development of diversified biomaterials in tissue engineering has been promoted by growing research into carbon-based nanomaterials. Usually, ideal scaffold materials should possess properties similar to the extracellular matrix of natural myocardial tissue. In this study, dopamine-reduced graphene oxide (GO), was prepared and doped into gelatin methacrylate (GelMA) hydrogels, resulting in novel conductive and mechanical properties for controlling cell growth. Cardiomyocytes (CMs) cultured on PDA-rGO-incorporated hydrogels (GelMA-PDA-rGO) had greater cytocompatibility than those cultured on GelMA hydrogels, as evidenced by higher cell survival rates and up-regulation of cardiac-relevant proteins. Finally, electrical stimulation was applied to facilitate the maturation of CMs which was seeded on different hydrogels. The findings revealed that electrical stimulation of conductive hybrid hydrogel scaffolds improved the orientational order parameter of sarcomeres (OOP). In addition, propagation of intercellular pacing signals, which improves the expression of gap junction proteins was noticed, likewise calcium handling capacity was present in conductive hybrid hydrogels compared to those in pure GelMA group. This study has shown that the combination of GelMA-PDA-rGO based conductive hydrogels and electrical stimulation possessed synergistic effects for engineering a more functional and mature myocardium layer as well as further application in drug screening and disease modeling in vitro.

An Efficient Method for Directed Hepatocyte-Like Cell Induction from Human Embryonic Stem Cells.

The potential functions of hepatocyte-like cells (HLCs) derived from human embryonic stem cells (hESCs) hold great promise for disease modeling and drug screening applications. Provided here is an efficient and reproducible method for differentiation of hESCs into functional HLCs. The establishment of an endoderm lineage is a key step in the differentiation to HLCs. By our method, we regulate the key signaling pathways by continuously supplementing Activin A and CHIR99021 during hESC differentiation into definitive endoderm (DE), followed by generation of hepatic progenitor cells, and finally HLCs with typical hepatocyte morphology in a stagewise method with completely defined reagents. The hESC-derived HLCs produced by this method express stage-specific markers (including albumin, HNF4α nuclear receptor, and sodium taurocholate cotransporting polypeptide (NTCP)), and show special characteristics related to mature and functional hepatocytes (including indocyanine green staining, glycogen storage, hematoxylin-eosin staining and CYP3 activity), and can provide a platform for the development of HLC-based applications in the study of liver diseases.

Impacts of OrX and cAMP-insensitive Orco to the insect olfactory heteromer activity.

Insect odorant receptors (ORs) have been suggested to function as ligand-gated cation channels, with OrX/Orco heteromers combining ionotropic and metabotropic activity. The latter is mediated by different G proteins and results in Orco self-activation by cyclic nucleotide binding. In this contribution, we co-express the odor-specific subunits DmOr49b and DmOr59b with either wild-type Orco or an Orco-PKC mutant lacking cAMP activation heterologously in mammalian cells. We show that the characteristics of heteromers strongly depend on both the OrX type and the coreceptor variant. Thus, methyl acetate-sensitive Or59b/Orco demonstrated 25-fold faster response kinetics over o-cresol-specific Or49b/Orco, while the latter required a 10-100 times lower ligand concentration to evoke a similar electrical response. Compared to wild-type Orco, Orco-PKC decreased odorant sensitivity in both heteromers, and blocked an outward current rectification intrinsic to the Or49b/Orco pair. Our observations thus provide an insight into insect OrX/Orco functioning, highlighting their natural and artificial tuning features and laying the groundwork for their application in chemogenetics, drug screening, and repellent design.

Establishment of Colorectal Cancer Organoids in Microfluidic-Based System.

Colorectal cancer is the second leading cause of cancer death worldwide. Significant advances in the molecular mechanisms underlying colorectal cancer have been made; however, the clinical approval of new drugs faces many challenges. Drug discovery is a lengthy process causing a rapid increase in global health care costs. Patient-derived tumour organoids are considered preclinical models with the potential for preclinical drug screening, prediction of patient outcomes, and guiding optimized therapy strategies at an individual level. Combining microfluidic technology with 3D tumour organoid models to recapitulate tumour organization and in vivo functions led to the development of an appropriate preclinical tumour model, organoid-on-a-chip, paving the way for personalized cancer medicine. Herein, a low-cost microfluidic device suitable for culturing and expanding organoids, OrganoidChip, was developed. Patient-derived colorectal cancer organoids were cultured within OrganoidChip, and their viability and proliferative activity increased significantly. No significant differences were verified in the organoids’ response to 5-fluorouracil (5-FU) treatment on-chip and on-plate. However, the culture within the OrganoidChip led to a significant increase in colorectal cancer organoid-forming efficiency and overall size compared with conventional culture on a 24-well plate. Interestingly, early-stage and late-stage organoids were predominantly observed on-plate and within the OrganoidChip, respectively. The OrganoidChip thus has the potential to generate in vivo-like organotypic structures for disease modelling and drug screening applications.