Multiorgan Microphysiological Systems for Drug Development: Strategies, Advances, and Challenges.

Drug screening is traditionally based on the pharmacodynamic models from 2D cell culture or animal experiments. However, these models suffer from poor drug efficacy prediction due to their difference from human in vivo cell microenvironments. In recent years, advances in biotechnology and tissue engineering have enabled rapid growth of in vitro organoid culturing. These organoids cultured in matrigel can mimic human in vivo cell microenvironment and physiology more accurately than traditional 2D cell culture and animal models. Among them, tumor organoids, especially patient-derived tumour organoids, can be applied as effective cancer models for drug screening and personalized medicine. By integrating organoid culturing and microfluidics, organoid-on-a-chip platform has been developed to simulate the structure and function of human tissues or organs. In 2010, Ingber et al. reported a biomimetic microfluidic platform called lung-on-a-chip. Microscale engineering technologies were firstly utilized to create biomimetic microchips. Henceforth, increasing kinds of organ-on-a-chips were reported all over the world to mimic different organs, such as liver, kidney and brain. 2D planar cells were cultured in PDMS chambers in majority organ-on-a-chips. More and more studies focused on 3D cells or organoids instead recently due to their better biomimetic capability for drug screening and disease models. In 2018, Qin et al. reported a liver organoid from human iPSCs in a 3D perfusable chip system, which is the first study that uses human organoid in a microfluidic chip and executed drug testing successfully. The established liver organoid-on-a-chip system may provide a promising platform for engineering stem cell-based organoids with applications in regenerative medicine, disease modeling and drug testing. Conventionally, drug efficacy evaluation in organoid-on-a-chip was performed by cell staining. However, this approach only presents static results of live/dead cells, but cannot monitor the cells dynamically for long-term recording. Sensor integration with organoid-on-a-chips paves a new way for dynamic recording. Currently, Wang et al. developed a 3D electric cell/matrigel-substrate impedance sensing (3D-ECMIS) platform for real-time and non-invasive monitoring of 3D cell viability and drug susceptibility. This platform integrated impedance sensors to monitor the impedance changes during 3D cell culturing. Additionally, Wang et al. reported the 3D cardiomyocytes detection by combining microelectrode arrays (MEAs). Heart cells of neonatal rat were cultured on a tissue engineering scaffold, which was fabricated by 3D printing and electro-spinning. The experimental results demonstrated that extracellular field potential (spike amplitude and firing rate) of 3D cardiomyocytes can be recorded in real time. Moreover, non-invasive measurement of cellular metabolism is important to study the metabolism mechanism and drug efficacy. Thus, light-addressable potentiometric sensor (LAPS) was integrated with organoid-on-a-chip for chemical imaging of ions (H+, Na+, K+, Ca2+) induced by cellular metabolism, which enables accurate detection of different chemical parameters in organoid-on-a-chip. Multiple organoid chip linked by microfluidics, called human-on-a-chip is a newly emerging and frontier technology to mimic the interaction of multiple human organs in recent years. It can recapitulate the physiologically relevant structures and functions of the organs, as well as the interaction between multiple organs as in vivo , thereby offering alternative models for predicting human responses to various drugs and environmental stimulus. Organoid-on-a-chip demonstrates outstanding potential in drug screening, disease modeling and personalized precision medicine.

The human kidney proximal tubule is responsible for glucose reabsorption and serves as a primary target for exogenous toxins. While conventional in vitro cell-based models offer cost-effective alternatives to animal testing, they often fail to replicate the structural and functional complexity of the native proximal tubule. Here, we developed a paper-based human kidney proximal tubule-on-a-chip that mimicked key physiological functions, bridging between traditional cell cultures and animal models. Utilizing porous paper, the chip recreated an in vivo-like three-dimensional microenvironment that supported proximal tubule-specific functions and reproduced essential physiological processes including dynamic glycogen metabolism, glucose reabsorption, and drug transport. The model enabled precise pharmacodynamics evaluation of sodium-glucose co-transporter 2 (SGLT2) inhibitors, yielding median effect concentrations of 0.954ng/mL for dapagliflozin and 2.685ng/mL for canagliflozin. The platform maintained consistently high glucose reabsorption inhibition rates (94.59%-95.03%) under different conditions following SGLT2 inhibitors treatment. Furthermore, the methotrexate (MTX)-induced nephrotoxicity evaluation was performed by MTT assay, LDH assay, and glucose reabsorption measurements. The chip accurately reproduced MTX transport dynamics, demonstrating its potential for pharmacokinetic studies. Thus, the paper-based model serves as a reliable platform for pharmacokinetic and nephrotoxicity assessments, offering a valuable tool to replace animal testing and support Reduce, Refine, and Replace experimentation.

Bone organoid construction and evolution.

This POSTnote summarises the emerging technology of human stem cell-based embryo models, discussions around their regulation and their wider ethical societal implications. It introduces the scientific background and the potential applications of the models. It also outlines the challenges and opportunities in introducing their regulation and discusses stakeholder initiatives to address regulatory gaps.

Simple SummaryMalignant peripheral nerve sheath tumors (MPNSTs) are highly aggressive tumors with a complex genetic landscape. Patients with neurofibromatosis type 1 syndrome (NF1) are at a high risk of MPNSTs, which usually develop from pre-existing benign Schwann cell tumors called plexiform neurofibromas. In this study, we aimed to find genes that, when altered, resulted in MPNST development. Our results suggest that the functional genetic landscape of human MPNST is complex and implicates the hippo/Yes Activated Protein (YAP) pathway in the transformation of neurofibromas. Malignant peripheral nerve sheath tumors (MPNSTs) are highly aggressive, genomically complex, have soft tissue sarcomas, and are derived from the Schwann cell lineage. Patients with neurofibromatosis type 1 syndrome (NF1), an autosomal dominant tumor predisposition syndrome, are at a high risk for MPNSTs, which usually develop from pre-existing benign Schwann cell tumors called plexiform neurofibromas. NF1 is characterized by loss-of-function mutations in the NF1 gene, which encode neurofibromin, a Ras GTPase activating protein (GAP) and negative regulator of RasGTP-dependent signaling. In addition to bi-allelic loss of NF1, other known tumor suppressor genes include TP53, CDKN2A, SUZ12, and EED, all of which are often inactivated in the process of MPNST growth. A sleeping beauty (SB) transposon-based genetic screen for high-grade Schwann cell tumors in mice, and comparative genomics, implicated Wnt/β-catenin, PI3K-AKT-mTOR, and other pathways in MPNST development and progression. We endeavored to more systematically test genes and pathways implicated by our SB screen in mice, i.e., in a human immortalized Schwann cell-based model and a human MPNST cell line, using CRISPR/Cas9 technology. We individually induced loss-of-function mutations in 103 tumor suppressor genes (TSG) and oncogene candidates. We assessed anchorage-independent growth, transwell migration, and for a subset of genes, tumor formation in vivo. When tested in a loss-of-function fashion, about 60% of all TSG candidates resulted in the transformation of immortalized human Schwann cells, whereas 30% of oncogene candidates resulted in growth arrest in a MPNST cell line. Individual loss-of-function mutations in the TAOK1, GDI2, NF1, and APC genes resulted in transformation of immortalized human Schwann cells and tumor formation in a xenograft model. Moreover, the loss of all four of these genes resulted in activation of Hippo/Yes Activated Protein (YAP) signaling. By combining SB transposon mutagenesis and CRISPR/Cas9 screening, we established a useful pipeline for the validation of MPNST pathways and genes. Our results suggest that the functional genetic landscape of human MPNST is complex and implicate the Hippo/YAP pathway in the transformation of neurofibromas. It is thus imperative to functionally validate individual cancer genes and pathways using human cell-based models, to determinate their role in different stages of MPNST development, growth, and/or metastasis.

Modelling the impact of ISO 14001 on environmental performance: A comparative approach

Lung cancer is a major malignancy that poses a significant threat to human health, with its complex pathogenesis and molecular characteristics presenting substantial challenges for treatment. Traditional two-dimensional cell cultures and animal models are limited in their ability to accurately replicate the characteristics of different lung cancer patients, thereby hindering research on disease mechanisms and treatment strategies. The development of organoid technology has enabled the growth of patient-derived tumor cells in three-dimensional cultures, which can stably preserve the tumor's tissue morphology, genomic features, and drug response. There have been significant advancements in the field of patient-derived lung cancer organoids (PDLCOs), challenges remain in the reproducibility and standardization of PDLCOs models due to variations in specimen sources, subsequent processing techniques, culture medium formulations, and Matrigel batches. This review summarizes the cultivation and validation processes of PDLCOs and explores their clinical applications in personalized treatment, drug screening after resistance, PDLCOs biobanks construction, and drug development. Additionally, the integration of PDLCOs with cutting-edge technologies in various fields, such as tumor assembloid techniques, artificial intelligence, organoid-on-a-chip, 3D bioprinting, gene editing, and single-cell RNA sequencing, has greatly expanded their clinical potential. This review, incorporating the latest research developments in PDLCOs, provides an overview of their cultivation, clinical applications, and interdisciplinary integration, while also addressing the prospects and challenges of PDLCOs in precision medicine for lung cancer.

Orchestrating the Parkinson’s disease microenvironment in 3D for pathogenesis study and therapeutic development

Cardiovascular disease is a major cause of mortality worldwide, and current preclinical models including traditional animal models and 2D cell culture models have limitations in replicating human native heart physiology and response to drugs. Heart-on-a-chip (HoC) technology offers a promising solution by combining the advantages of cardiac tissue engineering and microfluidics to create in vitro 3D cardiac models, which can mimic key aspects of human microphysiological systems and provide controllable microenvironments. Herein, recent advances in HoC technologies are introduced, including engineered cardiac microtissue construction in vitro, microfluidic chip fabrication, microenvironmental stimulation, and real-time feedback systems. The development of cardiac tissue engineering methods is focused for 3D microtissue preparation, advanced strategies for HoC fabrication, and current applications of these platforms. Major challenges in HoC fabrication are discussed and the perspective on the potential for these platforms is provided to advance research and clinical applications.

Animal models have provided invaluable information in biological evaluation and drug screening. It comes to predicting the human body’s response to candidate drugs; traditional laboratory animal models are woefully inadequate. However, increased concern for the welfare of the animals used, and a growing awareness of the concept of animal rights, has brought a greater focus on the related ethical issues.In this review, we seek to summarize the important anatomical aspects of Zebra fish as it is used as an alternative animal model for drug screening.

The use of stem cells to model the early human embryo promises to transform our understanding of developmental biology and human reproduction. In this review, we present our current knowledge of the first 2weeks of human embryo development. We first focus on the distinct cell lineages of the embryo and the derivation of stem cell lines. We then discuss the intercellular crosstalk that guides early embryo development and how this crosstalk is recapitulated invitro to generate stem cell-based embryo models. We highlight advances in this fast-developing field, discuss current limitations, and provide a vision for the future.

Because of recent failures of novel drugs in phase II and III trials innovative in vitro models are needed that may predict drug efficacy in patients. Although, numerous modifications of traditional cell culture and animal models were undertaken significant improvements in cell culture- based drug development have not been achieved. Three-dimensional (3D) cell culture may represents an modern alternative to bridge the gap between 2D cell cultures and animal models.

The incidence of respiratory diseases such as chronic obstructive pulmonary disease and pulmonary cancer is growing significantly around the world, making pulmonary disease one of the leading causes of mortality. However, the development of effective therapeutics for pulmonary diseases has been hindered by the lack of human‐mimetic physiological models that reliably emulate patient responses. Recent advances in technology and cell culture have led to the development of organoids and organ‐on‐a‐chip models that allow us to recapitulate the structure, cellular organization, and organ‐level responses of the target tissue in vitro. Here, we review the advances and milestones of lung organoid and lung‐on‐a‐chip models in the past decade and highlight their applications in mimicking pulmonary system development, physiology, disease, and regeneration. In addition, we discuss the ongoing challenges and the future prospects of integrating lung organoids and lung‐on‐a‐chip models to overcome current limitations and to enhance their physiological relevance. These human‐centric models are likely to provide important insights into pulmonary physiology and pathophysiology for drug discovery that complement and potentially replace traditional animal models.

Obesity, a pandemic of modern world, is a chronic disease that seriously affects human health. The causes of obesity are complicated, including environmental factors and genetic factors. Due to the fact that traditional animal models cannot fully mimic obesity in human, new animal models are urgently needed for basic drug research on obesity. Previous studies have demonstrated that the genomic diversity of the wild mice chromosome 1 substitution lines was significantly different from that of laboratory mice, suggesting that it might be accompanied by phenotypic diversity. We firstly screened the obesity-related phenotype of chromosome 1 substitution lines and found that the male chow-diet-fed B6-Chr1YP1 mice showed severe obesity-related phenotypes in body weight, lipid metabolism and liver lesions. Compared with C57BL/6J mice, B6-Chr1YP1 showed much more severe obesity-related phenotypes when fed with high-fat-diet. Furthermore, we evaluated efficacy effects of Tirzepatide and MGL-3196 in B6-Chr1YP1 mice, and obesity-related phenotypes were significantly rescued. Taken together, male B6-Chr1YP1 mice could serve as a novel, polygenic interaction-based obesity and hyperlipidemia model. This study could provide a novel animal model for accurate clinical diagnosis and precise medicine of obesity and hyperlipidemia. Disclosure H.Qi: None. X.Yang: None. J.Zheng: None. Y.Hou: None. Z.Chen: None. Z.Li: None. C.Ju: None. J.Zhao: None. X.Gao: None.

Breast cancer is the most common cancer worldwide, with an estimated 2.3 million new cases diagnosed every year. Effective measures for cancer prevention and cancer therapy require a detailed understanding of the individual key disease mechanisms involved and their interactions at the molecular, cellular, tissue, organ, and organism level. In that regard, the rapid progress of biomedical and toxicological research in recent years now allows the pursuit of new approaches based on non-animal methods that provide greater mechanistic insight than traditional animal models and therefore facilitate the development of Adverse Outcome Pathways (AOPs) for human diseases. We performed a systematic review of the current state of published knowledge with regard to breast cancer to identify relevant key mechanisms for inclusion into breast cancer AOPs, i.e. decreased cell stiffness and decreased cell adhesion, and to concurrently map non-animal methods addressing these key events. We conclude that the broader sharing of expertise and methods between biomedical research and toxicology enabled by the AOP knowledge management framework can help to coordinate global research efforts and accelerate the transition to advanced non-animal methods, which, when combined into powerful method batteries, closely mimic human physiology and disease states without the need for animal testing.