Monitoring Of Drug Response Research Articles

Human Data on Pharmacokinetic Interactions of Cannabinoids: A Narrative Review.

Concomitant use of cannabinoids with other drugs may result in pharmacokinetic drug-drug interactions, mainly due to the mechanism involving Phase I and Phase II enzymes and/or efflux transporters. Cannabinoids are not only substrates but also inhibitors or inducers of some of these enzymes and/or transporters. This narrative review aims to provide the available information reported in the literature regarding human data on the pharmacokinetic interactions of cannabinoids with other medications. A search on Pubmed/Medline, Google Scholar, and Cochrane Library was performed. Some studies were identified with Google search. Additional articles of interest were obtained through cross-referencing of published literature. All original research papers discussing interactions between cannabinoids, used for medical or recreational/adult-use purposes, and other medications in humans were included. Thirty-two studies with medicinal or recreational/adult-use cannabis were identified (seventeen case reports/series, thirteen clinical trials, and two retrospective analyses). In three of these studies, a bidirectional pharmacokinetic drug-drug interaction was reported. In the rest of the studies, cannabinoids were the perpetrators, as in most of them, concentrations of cannabinoids were not measured. In light of the widespread use of prescribed and non-prescribed cannabinoids with other medications, pharmacokinetic interactions are likely to occur. Physicians should be aware of these potential interactions and closely monitor drug levels and/or responses. The existing literature regarding pharmacokinetic interactions is limited, and for some drugs, studies have relatively small cohorts or are only case reports. Therefore, there is a need for high-quality pharmacological studies on cannabinoid-drug interactions.

A Review of Potential Roles for Micro RNAs as Drugs Sensitivity Biomarkers in Breast Cancer

MiR-202, part of the let-7 family, plays a role in cell division and tumor development, especially in breast cancer (BC) among females. We have conducted a detailed review on new biomarkers to enhance BC screening and enable early diagnosis, particularly in younger women. Early detection and monitoring drug responses are crucial for optimal BC therapy, prompting the need for new biomarkers in diagnosis and prognosis. This review discusses recent findings on microRNAs in detecting, analyzing, and treating breast cancer (miRNA-BC). Although breast cancer patients often start treatment with radiotherapy, its effectiveness may decline with radiation resistance. This review study compares docetaxel-resistant breast cancer (DRBC) and docetaxel-sensitive cell lines, exploring the role of non-coding RNAs, especially circular RNAs (circRNAs), in DRBC development and identifying predictive biomarkers for taxane-containing therapies. MicroRNA sequencing and bioinformatics predict differences in microRNA expression, genes, and pathways between breast cancer cell types. Current evidence suggests that miR-202 dysregulation is linked to signaling pathways, influencing its anti- or pro-tumorigenic effects, showcasing its potential as a diagnostic biomarker.

Urine-derived bladder cancer organoids (urinoids) as a tool for cancer longitudinal response monitoring and therapy adaptation.

Bladder cancer is one of the most common cancer types worldwide. Generally, research relies on invasive sampling strategies. Here, we generate bladder cancer organoids directly from urine (urinoids). In this project, we establish 12 urinoid lines from 22 patients with non-muscle and muscle-invasive bladder tumours, with an efficiency of 55%. The histopathological features of the urinoids accurately resemble those of the original bladder tumours. Genetically, there is a high concordance of single nucleotide polymorphisms (92.56%) and insertions & deletions (91.54%) between urinoids and original tumours from patient 4. Furthermore, these urinoids show sensitivity to bladder cancer drugs, similar to their tissue-derived organoid counterparts. Genetic analysis of longitudinally generated tumoroids and urinoids from one patient receiving systemic immunotherapy, identify alterations that may guide the choice for second-line therapy. Successful treatment adaptation was subsequently demonstrated in the urinoid setting. Therefore, urinoids can advance precision medicine in bladder cancer as a non-invasive platform for tumour pathogenesis, longitudinal drug-response monitoring, and therapy adaptation.

Detection of continuous hierarchical heterogeneity by single-cell surface antigen analysis in the prognosis evaluation of acute myeloid leukaemia

BackgroundAcute myeloid leukaemia (AML) is characterised by the malignant accumulation of myeloid progenitors with a high recurrence rate after chemotherapy. Blasts (leukaemia cells) exhibit a complete myeloid differentiation hierarchy hiding a wide range of temporal information from initial to mature clones, including genesis, phenotypic transformation, and cell fate decisions, which might contribute to relapse in AML patients.MethodsBased on the landscape of AML surface antigens generated by mass cytometry (CyTOF), we combined manifold analysis and principal curve-based trajectory inference algorithm to align myelocytes on a single-linear evolution axis by considering their phenotype continuum that correlated with differentiation order. Backtracking the trajectory from mature clusters located automatically at the terminal, we recurred the molecular dynamics during AML progression and confirmed the evolution stage of single cells. We also designed a ‘dispersive antigens in neighbouring clusters exhibition (DANCE)’ feature selection method to simplify and unify trajectories, which enabled the exploration and comparison of relapse-related traits among 43 paediatric AML bone marrow specimens.ResultsThe feasibility of the proposed trajectory analysis method was verified with public datasets. After aligning single cells on the pseudotime axis, primitive clones were recognized precisely from AML blasts, and the expression of the inner molecules before and after drug stimulation was accurately plotted on the trajectory. Applying DANCE to 43 clinical samples with different responses for chemotherapy, we selected 12 antigens as a general panel for myeloblast differentiation performance, and obtain trajectories to those patients. For the trajectories with unified molecular dynamics, CD11c overexpression in the primitive stage indicated a good chemotherapy outcome. Moreover, a later initial peak of stemness heterogeneity tended to be associated with a higher risk of relapse compared with complete remission.ConclusionsIn this study, pseudotime was generated as a new single-cell feature. Minute differences in temporal traits among samples could be exhibited on a trajectory, thus providing a new strategy for predicting AML relapse and monitoring drug responses over time scale.

Recent Analytical Advances for Decoding Metabolic Reprogramming in Lung Cancer.

Lung cancer is the leading cause of cancer-related death worldwide. Metabolic reprogramming is a fundamental trait associated with lung cancer development that fuels tumor proliferation and survival. Monitoring such metabolic pathways and their intermediate metabolites can provide new avenues concerning treatment strategies, and the identification of prognostic biomarkers that could be utilized to monitor drug responses in clinical practice. In this review, recent trends in the analytical techniques used for metabolome mapping of lung cancer are capitalized. These techniques include nuclear magnetic resonance (NMR), gas chromatography-mass spectrometry (GC-MS), liquid chromatography-mass spectrometry (LC-MS), and imaging mass spectrometry (MSI). The advantages and limitations of the application of each technique for monitoring the metabolite class or type are also highlighted. Moreover, their potential applications in the analysis of many biological samples will be evaluated.

Exploring Metabolomic Patterns in Type 2 Diabetes Mellitus and Response to Glucose-Lowering Medications-Review.

The spectrum of information related to precision medicine in diabetes generally includes clinical data, genetics, and omics-based biomarkers that can guide personalized decisions on diabetes care. Given the remarkable progress in patient risk characterization, there is particular interest in using molecular biomarkers to guide diabetes management. Metabolomics is an emerging molecular approach that helps better understand the etiology and promises the identification of novel biomarkers for complex diseases. Both targeted or untargeted metabolites extracted from cells, biofluids, or tissues can be investigated by established high-throughput platforms, like nuclear magnetic resonance (NMR) and mass spectrometry (MS) techniques. Metabolomics is proposed as a valuable tool in precision diabetes medicine to discover biomarkers for diagnosis, prognosis, and management of the progress of diabetes through personalized phenotyping and individualized drug-response monitoring. This review offers an overview of metabolomics knowledge as potential biomarkers in type 2 diabetes mellitus (T2D) diagnosis and the response to glucose-lowering medications.

Patient-Derived 3D Models for Preclinical Drug Response Evaluation in Neuroblastoma

Abstract Background Neuroblastoma is the most common solid childhood tumor outside the central nervous system. Metastatic and relapsed disease in neuroblastoma patients still has a poor prognosis. New treatment methods to improve therapy and outcomes for patients with advanced neuroblastoma are urgently needed. However, clinical trials currently only allow for testing few substances in even fewer patients. This increases the need to improve and advance preclinical tumor models to preselect favorable candidates for novel therapeutics. Aims This study investigates the advantages and limitations of using bioreactor-based perfused 3D models in for preclinical drug testing in solid tumors exemplified in neuroblastoma. Methods We established perfused bioreactor-based 3D models using neuroblastoma cell lines and tissue slice-culture and evaluated cell viability. Several inhibitors against hypoxia-related factors, associated with tumor progression, were tested including Aqb011 against aquaporin 1 (AQP1) and SLC-0111 against carbonic-anhydrase IX (CAIX). Isothermal microcalorimetry was used for measuring real time thermogenesis in 3D cell and tissue constructs. Results 3D neuroblastoma cell constructs presented with similar morphological features compared to neuroblastoma cells in intact tumor tissue featuring small, round and blue cells. Perfused neuroblastoma slice-culture maintained its primary structure in the bioreactor for up to 10 days. Both 3D neuroblastoma cell constructs and neuroblastoma tissue responded to inhibition with significant decrease of thermogenesis measured by microcalorimetry, if the correlating hypoxic factor (AQP1 or CAXI) was expressed. The AQP1 inhibitor Aqb011 and the CAIX inhibitor SLC-0111 decrease heat production of 3D neuroblastoma cultures. Conclusions We newly establish two preclinical models for neuroblastoma cells and tissue in a 3D structure. Drug response monitoring can be applied in these models using isothermal microcalorimetry. Monitoring times can be increased by use of the perfused bioreactor. Our models are an important first step to preclinically evaluate new drugs using patient-derived tumor tissue exemplified for neuroblastoma.

A microphysiological model of bone development and regeneration

Endochondral ossification (EO) is an essential biological process than underpins how human bones develop, grow, and heal in the event of a fracture. So much is unknown about this process, thus clinical manifestations of dysregulated EO cannot be adequately treated. This can be partially attributed to the absence of predictive in vitro models of musculoskeletal tissue development and healing, which are integral to the development and preclinical evaluation of novel therapeutics. Microphysiological systems, or organ-on-chip devices, are advanced in vitro models designed for improved biological relevance compared to traditional in vitro culture models. Here we develop a microphysiological model of vascular invasion into developing/regenerating bone, thereby mimicking the process of EO. This is achieved by integrating endothelial cells and organoids mimicking different stages of endochondral bone development within a microfluidic chip. This microphysiological model is able to recreate key events in EO, such as the changing angiogenic profile of a maturing cartilage analogue, and vascular induced expression of the pluripotent transcription factors SOX2 and OCT4 in the cartilage analogue. This system represents an advanced in vitro platform to further EO research, and may also serve as a modular unit to monitor drug responses on such processes as part of a multi-organ system.

Muscle-on-a-chip devices: a new era for in vitro modelling of muscular dystrophies.

Muscular dystrophies are a heterogeneous group of highly debilitating diseases that result in muscle atrophy and weakness. The lack of suitable cellular and animal models that reproduce specific aspects of their pathophysiology is one of the reasons why there are no curative treatments for these disorders. This highlights a considerable gap between current laboratory models and clinical practice. We strongly believe that organs-on-chip could help to fill this gap. Organs-on-chip, and in particular muscles-on-chip, are microfluidic devices that integrate functional skeletal muscle tissues. Biosensors in these systems allow monitoring of muscle homeostasis or drug responses in situ. This Perspective outlines the potential of organs-on-chip as advanced models for muscular dystrophies, as well as the current challenges and future opportunities for this technology.

Applications of single-cell RNA sequencing in drug discovery and development.

Single-cell technologies, particularly single-cell RNA sequencing (scRNA-seq) methods, together with associated computational tools and the growing availability of public data resources, are transforming drug discovery and development. New opportunities are emerging in target identification owing to improved disease understanding through cell subtyping, and highly multiplexed functional genomics screens incorporating scRNA-seq are enhancing target credentialling and prioritization. ScRNA-seq is also aiding the selection of relevant preclinical disease models and providing new insights into drug mechanisms of action. In clinical development, scRNA-seq can inform decision-making via improved biomarker identification for patient stratification and more precise monitoring of drug response and disease progression. Here, we illustrate how scRNA-seq methods are being applied in key steps in drug discovery and development, and discuss ongoing challenges for their implementation in the pharmaceutical industry.

P384 A higher red blood cell methotrexate polyglutamate 3 concentration is associated with methotrexate drug-survival in patients with Crohn’s Disease: first results of a prospective cohort

Abstract Background Prediction and monitoring of drug response of methotrexate (MTX) in patients with Crohn’s disease (CD) is an unmet need. MTX-polyglutamates (MTX-PGs) in red blood cells (RBC) are potential markers for response in other immune-mediated inflammatory diseases. Our objectives were investigating the relation between MTX-PGs and efficacy and defining predictors of response in CD patients treated with MTX. Methods In a multicenter prospective cohort study, CD patients starting subcutaneous (s.c.) MTX without biologics were included and followed for 12 months or until MTX s.c. was discontinued or a concomitant step-up therapy was started. At baseline clinical and biochemical predictors were recorded. At 2, 3, 6 and 12 months after start of therapy or when dropping out, blood samples were collected and individual MTX-PGs (MTX-PG1 - MTX-PG5) were analyzed in RBC using a validated UHPLC-MS/MS method. The outcome was either MTX s.c. discontinuation or initiation concomitant step-up therapy, due to disease activity or toxicity. Predictors were analysed in an univariate Cox regression model and MTX-PGs in an extended Cox model, corrected for prednisone (at start) and budesonide. Results Eighty CD patients were included (age mean±SD 55±13y, 35% male). The median Harvey Bradshaw Index (HBI) was 4 (IQR 2-7). After 12 months 21 patients were still on MTX s.c. monotherapy, 21 patients stopped because of disease activity, 29 because of toxicity, 4 because of a combination of both and 5 patients were censored (4 ended study participation, 1 MTX was stopped on patient’s own initiative) [Figure 1]. A higher HBI at baseline was associated with an increased rate of MTX s.c. monotherapy discontinuation (HR 1.08, 95% CI 1.02-1.16). Predictors of discontinuation because of disease activity (cause specific hazards) were male sex (3.83, 1.62-9.05), baseline eGFR (1.06, 1.02-1.09), baseline HBI (1.12, 1.02-1.23) and baseline plasma folate (0.94, 0.88-0.99). Sex and plasma folate were not correlated with HBI. No cause specific hazards for stopping MTX because of toxicity were identified. MTX-PG3 was the most abundant MTX-PG subspecie with a median concentration of 51 nmol/L RBC (IQR 37-62) at month 3 and was associated with better MTX survival (HR 0.98, 95% CI 0.971-0.999). For every ten points increase in the MTX-PG3 concentration, the rate of MTX s.c. monotherapy discontinuation decreases with 14%. Conclusion Higher eGFR, higher HBI, lower plasma folate at baseline and male sex are predictors for MTX failure in the first year in CD patients. The last two parameters are likely predictors of MTX response specifically rather than prognostic disease factors. The measurement of MTX-PG3 in packed RBC holds potential as a tool for therapeutic drug monitoring in CD.

When cancer drug resistance meets metabolomics (bulk, single-cell and/or spatial): Progress, potential, and perspective.

Resistance to drug treatment is a critical barrier in cancer therapy. There is an unmet need to explore cancer hallmarks that can be targeted to overcome this resistance for therapeutic gain. Over time, metabolic reprogramming has been recognised as one hallmark that can be used to prevent therapeutic resistance. With the advent of metabolomics, targeting metabolic alterations in cancer cells and host patients represents an emerging therapeutic strategy for overcoming cancer drug resistance. Driven by technological and methodological advances in mass spectrometry imaging, spatial metabolomics involves the profiling of all the metabolites (metabolomics) so that the spatial information is captured bona fide within the sample. Spatial metabolomics offers an opportunity to demonstrate the drug-resistant tumor profile with metabolic heterogeneity, and also poses a data-mining challenge to reveal meaningful insights from high-dimensional spatial information. In this review, we discuss the latest progress, with the focus on currently available bulk, single-cell and spatial metabolomics technologies and their successful applications in pre-clinical and translational studies on cancer drug resistance. We provide a summary of metabolic mechanisms underlying cancer drug resistance from different aspects; these include the Warburg effect, altered amino acid/lipid/drug metabolism, generation of drug-resistant cancer stem cells, and immunosuppressive metabolism. Furthermore, we propose solutions describing how to overcome cancer drug resistance; these include early detection during cancer initiation, monitoring of clinical drug response, novel anticancer drug and target metabolism, immunotherapy, and the emergence of spatial metabolomics. We conclude by describing the perspectives on how spatial omics approaches (integrating spatial metabolomics) could be further developed to improve the management of drug resistance in cancer patients.

Brca1 and Tp53 co-deficiency causes a PARP-inhibitor sensitive erythroproliferative neoplasm.

Mutations in the BRCA1 tumor suppressor gene, such as 5382insC (BRCA15382insC), give carriers an increased risk for breast, ovarian, prostate and pancreatic cancers. We have previously reported that, in mice, Brca1 deficiency in the hematopoietic system leads to pancytopenia and, as a result, early lethality. Here we explore the cellular consequences of Brca1 null and BRCA1 5382insC alleles in combination with Tp53 deficiency in the murine hematopoietic system. We find that Brca1 and Tp53 co-deficiency leads to a highly penetrant erythroproliferative disorder that is characterized by hepatosplenomegaly and expanded megakaryocyte erythroid progenitor (MEP) and immature erythroid blast populations. The expanded erythroid progenitor populations in both bone marrow and spleen have the capacity to transmit the disease into secondary mouse recipients, suggesting Brca1 and Tp53 co-deficiency provides a new murine model of hematopoietic neoplasia. This Brca1/Tp53 model replicates Poly (ADP-ribose) polymerase (PARP) inhibitor olaparib sensitivity seen in existing Brca1/Tp53 breast cancer models and has the benefits of monitoring disease progression and drug responses via peripheral blood analyses without sacrificing experimental animals. In addition, this erythroid neoplasia develops much faster than murine breast cancer, allowing for increased efficiency of future preclinical studies.

Identifying the prevalence of clinically actionable drug-gene interactions in a health system biorepository to guide pharmacogenetics implementation services.

Understanding patterns of drug-gene interactions (DGIs) is important for advancing the clinical implementation of pharmacogenetics (PGx) into routine practice. Prior studies have estimated the prevalence of DGIs, but few have confirmed DGIs in patients with known genotypes and prescriptions, nor have they evaluated clinician characteristics associated with DGI-prescribing. This retrospective chart review assessed prevalence of DGI, defined as a medication prescription in a patient with a PGx phenotype that has a clinical practice guideline recommendation to adjust therapy or monitor drug response, for patients enrolled in a research genetic biorepository linked to electronic health records (EHRs). The prevalence of prescriptions for medications with pharmacogenetic (PGx) guidelines, proportion of prescriptions with DGI, location of DGI prescription, and clinical service of the prescriber were evaluated descriptively. Seventy-five percent (57,058/75,337) of patients had a prescription for a medication with a PGx guideline. Up to 60% (n=26,067/43,647) of patients had at least one DGI when considering recommendations to adjust or monitor therapy based on genotype. The majority (61%) of DGIs occurred in outpatient prescriptions. Proton pump inhibitors were the most common DGI medication for 11 of 12 clinical services. Almost 25% of patients (n=10,706/43,647) had more than one unique DGI, and, among this group of patients, 61% had a DGI with more than one gene. These findings can inform future clinical implementation by identifying key stakeholders for initial DGI prescriptions, helping to inform workflows. The high prevalence of multigene interactions identified also support the use of panel PGx testing as an implementation strategy.

Blood-Lymphatic Integrated System with Heterogeneous Melanoma Spheroids via In-Bath Three-Dimensional Bioprinting for Modelling of Combinational Targeted Therapy.

Although metastatic melanoma can be managed with chemotherapy, its heterogeneity and resistance to therapy remain poorly understood. In addition to the spread of melanoma in the bloodstream, melanoma‐stroma interaction and the lymphatic system play active roles in said heterogeneity and resistance, leading to its progression and metastasis. Reproducing the complexities of the melanoma microenvironment in vitro will help understanding its progression and enhance the translatability of potential cancer therapeutics. A blood‐lymphatic integrated system with heterogeneous melanoma spheroids (BLISH) using the in‐bath bioprinting process is developed. The process uniformly prints size‐controllable metastatic melanoma spheroids along with biomimetic blood and lymphatic vessels (LVs). The system reproduces hallmark events of metastatic melanoma, such as tumor stroma interaction, melanoma invasion, and intravasation. The application of the system to investigate the anticancer effect of combinational targeted therapy suggests that it can be used to study the pathophysiology of melanoma and improve the accuracy of drug response monitoring in skin cancer.

A data-independent acquisition (DIA)-based quantification workflow for proteome analysis of 5000 cells

Data independent acquisition (DIA) has emerged as a powerful proteomic technique with exceptional reproducibility and throughput, and has been widely applied to clinical sample analysis. DIA approaches normally rely on project-specific spectral libraries generated by data dependent acquisition (DDA), requiring extensive off-line fractionation and large amounts of input material. In this study, we aimed to explore the utility of DIA for the analysis of samples with limited quantities. We employed three software tools (DIA-NN, Spectronaut, and EncyclopeDIA) for data analysis and generated three types of libraries, including an experiment-specific library built by DDA analysis of off-line fractions, a FASTA sequence database, and a library generated by gas-phase fractionation (GPF), resulting in eight analysis pipelines. Then we systematically compared the performance of the eight strategies by analyzing the DIA data from HEK293T cell tryptic peptides with sample loads of 500 ng, 100 ng, 20 ng, and 4 ng. The results showed that DIA-NN with GPF-based libraries outperformed the others in protein identification and retention time calibration. Next, we further evaluated the optimized workflow by analyzing the proteome alteration in 5000 peripheral blood mononuclear cells (PBMCs) induced by lipopolysaccharide (LPS) stimulation. As a result, 3179 protein groups were quantified, and functional analysis revealed activation of multiple signaling pathways, e. g., endocytosis, NF-kappa B signaling, and T cell receptor signaling. The results showed the practicability of using DIA for scarce samples, and the established workflow of PBMC analysis could be easily adapted for biomarker discovery, immune status evaluation, and drug response monitoring, especially for diseases involved with dysfunction of the immune system.

Real-time monitoring of immediate drug response and adaptation upon repeated treatment in a microfluidic chip system

Microfluidic tissue culture and organ-on-a-chip models provide efficient tools for drug testing in vivo and are considered to become the basis of in vitro test systems to analyze drug response, drug interactions and toxicity to complement and reduce animal testing. A major limitation is the efficient recording of drug action. Here we present an efficient experimental setup that allows long-term cultivation of cells in a microfluidic system in combination with continuous recording of luciferase reporter gene expression. The system combines a sensitive cooled luminescence camera system in combination with a custom build miniaturized incubation chamber. The setup allows to monitor time-dependent activation, but also the end of drug response. Repeated activation and recovery as well as varying durations of drug treatment periods can be monitored, and different modes of drug activity can be visualized.

Perfusion-Based Bioreactor Culture and Isothermal Microcalorimetry for Preclinical Drug Testing with the Carbonic Anhydrase Inhibitor SLC-0111 in Patient-Derived Neuroblastoma.

Neuroblastoma is a rare disease. Rare are also the possibilities to test new therapeutic options for neuroblastoma in clinical trials. Despite the constant need to improve therapy and outcomes for patients with advanced neuroblastoma, clinical trials currently only allow for testing few substances in even fewer patients. This increases the need to improve and advance preclinical models for neuroblastoma to preselect favorable candidates for novel therapeutics. Here we propose the use of a new patient-derived 3D slice-culture perfusion-based 3D model in combination with rapid treatment evaluation using isothermal microcalorimetry exemplified with treatment with the novel carbonic anhydrase IX and XII (CAIX/CAXII) inhibitor SLC-0111. Patient samples showed a CAIX expression of 18% and a CAXII expression of 30%. Corresponding with their respective CAIX expression patterns, the viability of SH-EP cells was significantly reduced upon treatment with SLC-0111, while LAN1 cells were not affected. The inhibitory effect on SH-SY5Y cells was dependent on the induction of CAIX expression under hypoxia. These findings corresponded to thermogenesis of the cells. Patient-derived organotypic slice cultures were treated with SLC-0111, which was highly effective despite heterogeneity of CAIX/CAXII expression. Thermogenesis, in congruence with the findings of the histological observations, was significantly reduced in SLC-0111-treated samples. In order to extend the evaluation time, we established a perfusion-based approach for neuroblastoma tissue in a 3D perfusion-based bioreactor system. Using this system, excellent tissue quality with intact tumor cells and stromal structure in neuroblastoma tumors can be maintained for 7 days. The system was successfully used for consecutive drug response monitoring with isothermal microcalorimetry. The described approach for drug testing, relying on an advanced 3D culture system combined with a rapid and highly sensitive metabolic assessment, can facilitate development of personalized treatment strategies for neuroblastoma.

Integrated microfluidic single-cell immunoblotting chip enables high-throughput isolation, enrichment and direct protein analysis of circulating tumor cells

Effective capture and analysis of a single circulating tumor cell (CTC) is instrumental for early diagnosis and personalized therapy of tumors. However, due to their extremely low abundance and susceptibility to interference from other cells, high-throughput isolation, enrichment, and single-cell-level functional protein analysis of CTCs within one integrated system remains a major challenge. Herein, we present an integrated multifunctional microfluidic system for highly efficient and label-free CTC isolation, CTC enrichment, and single-cell immunoblotting (ieSCI). The ieSCI-chip is a multilayer microfluidic system that combines an inertia force-based cell sorter with a membrane filter for label-free CTC separation and enrichment and a thin layer of a photoactive polyacrylamide gel with microwell arrays at the bottom of the chamber for single-cell immunoblotting. The ieSCI-chip successfully identified a subgroup of apoptosis-negative (Bax-negative) cells, which traditional bulk analysis did not detect, from cisplatin-treated cells. Furthermore, we demonstrated the clinical application of the ieSCI-chip with blood samples from breast cancer patients for personalized CTC epithelial-to-mesenchymal transition (EMT) analysis. The expression level of a tumor cell marker (EpCAM) can be directly determined in isolated CTCs at the single-cell level, and the therapeutic response to anticancer drugs can be simultaneously monitored. Therefore, the ieSCI-chip provides a promising clinical translational tool for clinical drug response monitoring and personalized regimen development.

Current Perspectives on the Unique Roles of Exosomes in Drug Resistance of Hepatocellular Carcinoma.

As a common malignant tumor worldwide, the prognosis of hepatocellular carcinoma (HCC) remains unsatisfactory, even though treatment methods have improved. Despite the developments in traditional chemotherapy and emerging targeted immunotherapy, the problem of recurrence and metastasis of HCC and adverse effects on survival and prognosis are still serious. Drug resistance is a daunting challenge that impedes HCC treatment. Exosomes, a class of extracellular vesicles ranging in size from 30 to 100 nm, have been the focus of recent studies. Exosomes can activate various signaling pathways and regulate the tumor microenvironment with their cargo, which includes functional lipids, proteins, and nucleic acids. Thus, they change the phenotype of recipient cells via exosome-mediated communication. Exosomes secreted by tumors or stromal cells can also transfer drug-resistant traits to other tumor cells. However, their effects on drug resistance in HCC are not completely understood. In this review, we summarize and discuss the underlying relationship between exosomes and drug resistance in HCC. In addition, we also show that exosomes may act as candidate biomarkers for predicting and monitoring drug responses and as potential targets or vectors to reverse the drug resistance of HCC.