Next-Generation Preclinical Functional Testing Models in Cancer Precision Medicine: CTC-Derived Organoids.

Basic and clinical cancer research requires tumor models that consistently recapitulate the characteristics of prima tumors. As ex vivo 3D cultures of patient tumor cells, patient-derived tumor organoids possess the biological properties of primary tumors and are therefore excellent preclinical models for cancer research. Patient-derived organoids can be established using primary tumor tissues, peripheral blood, pleural fluid, ascites, and other samples containing tumor cells. Circulating tumor cells acquired by non-invasive sampling feature dynamic circulation and high heterogeneity. Circulating tumor cell-derived organoids are prospective tools for the dynamic monitoring of tumor mutation evolution profiles because they reflect the heterogeneity of the original tumors to a certain extent. This review discusses the advantages and applications of patient-derived organoids. Meanwhile, this work highlights the biological functions of circulating tumor cells, the latest advancement in research of circulating tumor cell-derived organoids, and potential application and challenges of this technology.

Manganese vacancies and tunnel pillars synergistically improve the electrochemical performance of MnO2 in aqueous Zn ion batteries.

High-oxidation niobium was used for the first time in manganese dioxide cation doping to reduce the diffusion resistance of zinc ions, in order to improve its kinetic and electrochemical properties. The results show that using a simple hydrothermal process, all niobium ions were doped into the manganese dioxide lattice. As niobium(v) was incorporated into the [2 × 2] tunnel of α-MnO2, it induced manganese vacancies, which reduced the diffusion resistance of Zn2+ in manganese dioxide, improving the migration kinetics. It acted as a tunnel pillar, avoiding the collapse of the tunnel structure during the repeated insertion/extraction of the Zn2+ process, and prevented a rapid degradation of the cycling performance. In particular, the sample with the Nb/Mn molar ratio of 0.003 exhibited the best kinetic reversibility and rate performance. After 400 cycles at 1C, the capacity retention of Nb-doped MnO2 significantly increased to 89%, which was only 55% for the undoped sample. Meanwhile, at a power density of 400 W kg-1, it presented the highest energy density of 765 W h kg-1 due to the existing doping of metal ions.

Development and Validation of the Oxidative Stress Related lncRNAs for Prognosis in Esophageal Squamous Cell Carcinoma.

Esophageal squamous cell cancer (ESCC) is an aggressive disease associated with a poor prognosis. Long non-coding RNAs (lncRNAs) and oxidative stress play crucial roles in tumor progression. We aimed to identify an oxidative stress-related lncRNA signature that could predict the prognosis in ESCC. In the GSE53625 dataset, we identified 332 differentially expressed lncRNAs (DElncRNAs) between ESCC and control samples, out of which 174 were oxidative stress-related DElncRNAs. Subsequently, seven oxidative stress-related DElncRNAs (CCR5AS, LINC01749, PCDH9-AS1, TMEM220-AS1, KCNMA1-AS1, SNHG1, LINC01672) were selected based on univariate and LASSO Cox to build a prognostic risk model, and their expression was detected by RT-qPCR. The model exhibited an excellent ability for the prediction of overall survival (OS) and other clinicopathological traits using Kaplan-Meier (K-M) survival curves, receiver operating characteristic (ROC) curves, and the Wilcoxon test. Additionally, analysis of infiltrated immune cells and immune checkpoints indicated differences in immune status between the two risk groups. Finally, the in vitro experiments showed that PCDH9-AS1 overexpression inhibited proliferation ability and promoted apoptosis and oxidative stress levels in ESCC cells. In conclusion, our study demonstrated that a novel oxidative stress-related DElncRNA prognostic model performed favorably in predicting ESCC patient prognosis and benefits personalized clinical applications.

The possible molecular mechanism underlying the involvement of the variable shear factor QKI in the epithelial-mesenchymal transformation of oesophageal cancer.

OBJECTIVE: Based on the GEO, TCGA and GTEx databases, we reveal the possible molecular mechanism of the variable shear factor QKI in epithelial mesenchymal transformation (EMT) of oesophageal cancer. METHODS: Based on the TCGA and GTEx databases, the differential expression of the variable shear factor QKI in oesophageal cancer samples was analysed, and functional enrichment analysis of QKI was performed based on the TCGA-ESCA dataset. The percent-spliced in (PSI) data of oesophageal cancer samples were downloaded from the TCGASpliceSeq database, and the genes and variable splicing types that were significantly related to the expression of the variable splicing factor QKI were screened out. We further identified the significantly upregulated circRNAs and their corresponding coding genes in oesophageal cancer, screened the EMT-related genes that were significantly positively correlated with QKI expression, predicted the circRNA-miRNA binding relationship through the circBank database, predicted the miRNA-mRNA binding relationship through the TargetScan database, and finally obtained the circRNA-miRNA-mRNA network through which QKI promoted the EMT process. RESULTS: Compared with normal control tissue, QKI expression was significantly upregulated in tumour tissue samples of oesophageal cancer patients. High expression of QKI may promote the EMT process in oesophageal cancer. QKI promotes hsa_circ_0006646 and hsa_circ_0061395 generation by regulating the variable shear of BACH1 and PTK2. In oesophageal cancer, QKI may promote the production of the above two circRNAs by regulating variable splicing, and these circRNAs further competitively bind miRNAs to relieve the targeted inhibition of IL-11, MFAP2, MMP10, and MMP1 and finally promote the EMT process. CONCLUSION: Variable shear factor QKI promotes hsa_circ_0006646 and hsa_circ_0061395 generation, and downstream related miRNAs can relieve the targeted inhibition of EMT-related genes (IL11, MFAP2, MMP10, MMP1) and promote the occurrence and development of oesophageal cancer, providing a new theoretical basis for screening prognostic markers of oesophageal cancer patients.

In Situ Microreaction Platform Based on Acoustic Droplet Manipulation for Ultra-High-Precision Multiplex Bioassay.

Liquid droplets rectors have been used in clinical diagnosis, high throughput screening and bioassay. However, it is challenging for droplet reactors to be used in practical applications due to the difficulty of uniformly mixing ultrasmall volumes of samples and the lack of rapid and high-precision detection protocols. Here, we have developed an acoustic droplet system for rapid and efficient biological detection and chemical screening. By employing acoustic wave devices, rapid and nondestructive uniform mixing of ∼nL-µL droplets can be achieved. By the acoustophoretic force, the perturbation of the droplets can quickly concentrate the sample and increase the detection limit by five times. Through the color reaction and the coordinated detection of photodiodes, we have developed a biomarker detection protocol with short reaction time and high accuracy. As a proof-of-concept application, we demonstrated that this system can detect ultrasmall or low-abundance samples faster and more accurately, highlighting its wide application in analytical chemistry, basic research, and clinical medicine.

Engineered exosomes as an in situ DC-primed vaccine to boost antitumor immunity in breast cancer.

BACKGROUND: Dendritic cells (DCs) are central for the initiation and regulation of innate and adaptive immunity in the tumor microenvironment. As such, many kinds of DC-targeted vaccines have been developed to improve cancer immunotherapy in numerous clinical trials. Targeted delivery of antigens and adjuvants to DCs in vivo represents an important approach for the development of DC vaccines. However, nonspecific activation of systemic DCs and the preparation of optimal immunodominant tumor antigens still represent major challenges. METHODS: We loaded the immunogenic cell death (ICD) inducers human neutrophil elastase (ELANE) and Hiltonol (TLR3 agonist) into α-lactalbumin (α-LA)-engineered breast cancer-derived exosomes to form an in situ DC vaccine (HELA-Exos). HELA-Exos were identified by transmission electron microscopy, nanoscale flow cytometry, and Western blot analysis. The targeting, killing, and immune activation effects of HELA-Exos were evaluated in vitro. The tumor suppressor and immune-activating effects of HELA-Exos were explored in immunocompetent mice and patient-derived organoids. RESULTS: HELA-Exos possessed a profound ability to specifically induce ICD in breast cancer cells. Adequate exposure to tumor antigens and Hiltonol following HELA-Exo-induced ICD of cancer cells activated type one conventional DCs (cDC1s) in situ and cross-primed tumor-reactive CD8+ T cell responses, leading to potent tumor inhibition in a poorly immunogenic triple negative breast cancer (TNBC) mouse xenograft model and patient-derived tumor organoids. CONCLUSIONS: HELA-Exos exhibit potent antitumor activity in both a mouse model and human breast cancer organoids by promoting the activation of cDC1s in situ and thus improving the subsequent tumor-reactive CD8+ T cell responses. The strategy proposed here is promising for generating an in situ DC-primed vaccine and can be extended to various types of cancers. Scheme 1. Schematic illustration of HELA-Exos as an in situ DC-primed vaccine for breast cancer. (A) Allogenic breast cancer-derived exosomes isolated from MDA-MB-231 cells were genetically engineered to overexpress α-LA and simultaneously loaded with the ICD inducers ELANE and Hiltonol (TLR3 agonist) to generate HELA-Exos. (B) Mechanism by which HELA-Exos activate DCs in situ in a mouse xenograft model ofTNBC. HELA-Exos specifically homed to the TME and induced ICD in cancer cells, which resulted in the increased release of tumor antigens, Hiltonol, and DAMPs, as well as the uptake of dying tumor cells by cDC1s. The activated cDC1s then cross-primed tumor-reactive CD8+ T cell responses. (C) HELA-Exos activated DCs in situ in the breast cancer patient PBMC-autologous tumor organoid coculture system. ABBREVIATIONS: DCs: dendritic cells; α-LA: α-lactalbumin; HELA-Exos: Hiltonol-ELANE-α-LA-engineered exosomes; ICD: immunogenic cell death; ELANE: human neutrophil elastase; TLR3: Toll-like receptor 3; TNBC: triple-negative breast cancer; TME: tumor microenvironment; DAMPs: damage-associated molecular patterns; cDC1s: type 1 conventional dendritic cells; PBMCs: peripheral blood mononuclear cells.

Acoustic Droplet Printing Tumor Organoids for Modeling Bladder Tumor Immune Microenvironment within a Week.

Current organoid models are limited by the incapability of rapidly fabricating organoids that can mimic the immune microenvironment for a short term. Here, an acoustic droplet-based platform is presented to facilitate the rapid formation of tumor organoids, which retains the original tumor immune microenvironment and establishes a personalized bladder cancer tumor immunotherapy model. In combination with a hydrophobic substrate, the acoustic droplet printer can yield a large number of homogeneous and highly viable bladder tumor organoids in vitro within a week. The generated organoids consist of all components of bladder tumor, including diverse immune elements and tumor cells. By coculturing tumor organoids with autologous immune cells for 2 days, tumor reactive T cells are induced in vitro. Furthermore, it is also demonstrated that these tumor-reactive T cells can also enhance the killing efficiency of matched organoids. Because of the easy operation, repeatability, and stability, the proposed acoustic droplet platform will provide a reliable approach for personalized tumor immunotherapy.

Circular RNAs Are Promising Biomarkers in Liquid Biopsy for the Diagnosis of Non-small Cell Lung Cancer.

The high incidence and mortality of lung cancer make early detection of lung cancer particularly important. At present, the diagnosis of lung cancer mainly depends on diagnostic imaging and tissue biopsy. However, current diagnostics are not satisfactory owing to the low specificity and inability of multiple sampling. Accumulating evidence indicates that circular RNAs (circRNAs) play a critical role in cancer progression and are promising cancer biomarkers. In particular, circRNAs are considered novel specific diagnostic markers for non-small cell lung cancer (NSCLC). Liquid biopsy is an important method in the early diagnosis of cancer due to its high sensitivity and specificity, as well as the possibility of performing multiple sampling. circRNAs are stably present in exosomes and sometimes become part of circulating nucleic acids, making them ideal for liquid biopsy. In this review, we summarize the advances in the research on circRNAs in NSCLC, and also highlight their potential applications for NSCLC detection.

Acoustic Droplet Vitrification Method for High-Efficiency Preservation of Rare Cells.

Cryopreservation is a key step for current translational medicine including reproductive medicine, regenerative medicine, and cell therapy. However, it is challenging to preserve rare cells for practical applications due to the difficulty in handling low numbers of cells as well as the lack of highly efficient and biocompatible preservation protocols. Here, we developed an acoustic droplet vitrification method for high-efficiency handling and preservation of rare cells. By employing an acoustic droplet ejection device, we can encapsulate rare cells into water-in-air droplets with a volume from ∼pL to ∼nL and deposit these cell-containing droplets into a droplet array onto a substrate. By incorporating a cooling system into the droplet array substrate, we can vitrify hundreds to thousands of rare cells at an ultrafast speed (about ∼2 s) based on the high surface to volume ratio of the droplets. By optimizing this method with three different cell lines (a human lung cancer cell line, A549 cells, a human liver cell line, L02 cells, and a mouse embryonic fibroblast cell line, 3T3-L1 cells), we developed an effective protocol with excellent cell viability (e.g., >85% for days, >70% for months), proliferation, and adhesion. As a proof-of-concept application, we demonstrated that our method can rapidly handle and efficiently preserve rare cells, highlighting its broad applications in species diversity, basic research, and clinical medicine.

Aptamer-Exosomes for Tumor Theranostics.

As carriers of biomolecules (proteins, nucleic acids, and lipids) from parent cells, exosomes play a significant role in physiology and pathology. In any diseased state, the morphology of the released exosomes remained similar. The contents of exosomes change depending on the disease or its stage; thus, exosomes are generally considered as a "source of biomarkers". Therefore, they are considered promising biomarkers for the diagnosis and prognosis of tumors. As natural delivery vehicles, exosomes can protect their cargo from immune clearance and deliver them to other cells through membrane fusion. After being genetically edited at the cell or exosome level, exosomes can be used for treatment with aptamers. Aptamers are short stretches of oligonucleotide sequences or short polypeptides that have been selected in vitro or in vivo, and have a wide range of targets and show excellent binding affinity and specificity. Aptamers have been widely used as molecular probes, and the combination of aptamers with exosomes has become a new direction for exosome-related research and therapeutic development. Here, we summarized various applications of exosomes and aptamers in cancer research, and further analyzed their combination as an "aptamer-exosome". Finally, we propose future directions for the aptamer-exosome in the precise diagnosis or personalized treatment of cancer.

COVID-19: advance in laboratory diagnostic strategy and technology.

SARS-CoV-2 is one of the beta-coronaviruses with the spike protein. It invades host cells by binding to angiotensin converting enzyme 2 (ACE2). This newly discovered virus can result in excessive inflammation and immune pathological damage, as shown by a decreased number of peripheral lymphocytes, increased levels of cytokines, and damages of lung, heart, liver, kidney, and other organs. Effective therapeutic modalities such as new antiviral drugs and vaccines against this emerging virus need to be thoroughly studied and developed. However, so far the only recognized but mild progress in this area is the screening of old drugs for new uses. Therefore, rapid and accurate laboratory SARS-CoV-2 testing approaches are the important basis of identification and blockage of COVID-19 transmission. For COVID-19 patients with different clinical classifications (mild, common, severe, and critically severe), dynamic monitoring of functional indicators of susceptible and vital organs is an important strategy for evaluating therapeutic efficacy and prognosis. In this review, we summarized SARS-CoV-2 laboratory diagnostic schemes, pathophysiological indices of tissues and organs of COVID-19 patients, and laboratory diagnostic strategies for distinct disease stages. Further, we discussed the importance of hierarchical management and dynamic observation in SARS-CoV-2 laboratory diagnostics. We then summed up the advance in SARS-CoV-2 testing technology and described the prospect of intelligent medicine in the prevention of infectious disease outbreaks.

SARS-CoV-2 vaccine research and development: Conventional vaccines and biomimetic nanotechnology strategies.

The development of a massively producible vaccine against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), a novel coronavirus, is essential for stopping the current coronavirus disease (COVID-19) pandemic. A vaccine must stimulate effective antibody and T cell responses in vivo to induce long-term protection. Scientific researchers have been developing vaccine candidates for the severe acute respiratory syndrome (SARS) and Middle East respiratory syndrome (MERS) since the outbreaks of these diseases. The prevalence of new biotechnologies such as genetic engineering has shed light on the generation of vaccines against novel viruses. In this review, we present the status of the development of coronavirus vaccines, focusing particularly on the biomimetic nanoparticle technology platform, which is likely to have a major role in future developments of personalized medicine.

Circulating tumor cells in hepatocellular carcinoma: single-cell based analysis, preclinical models, and clinical applications.

Circulating tumor cells (CTCs) are shed into the bloodstream from primary tumors and metastatic lesions and provide significant information about tumor progression and metastasis. CTCs contribute to tumor metastasis through the epithelial-to-mesenchymal transition (EMT). CTC clusters and stem-like phenotypes lead to a more aggressive and metastatic potential. CTCs retain the heterogeneity and imitate the nature of corresponding primary tumors. Therefore, it is important to use single-cell based analysis to obtain information on tumor heterogeneity and biology. CTCs are also good candidates for building preclinical models (especially 3D organoid cultures) for drug screening, disease modeling, genome editing, tumor immunity research, and organ-like biobank establishment. In this article, we summarize the current CTC capture technology, dissect the phenotypes associated with CTC metastasis, and review the progress in single-cell based analysis and preclinical modeling of the pattern and kinetics of CTCs. In particular, we discuss the use of CTCs to assess the progression of hepatocellular carcinoma (HCC).

Co-administration of sulforaphane and doxorubicin attenuates breast cancer growth by preventing the accumulation of myeloid-derived suppressor cells.

Sulforaphane (SFN) is a compound derived from cruciferous plants shown to be effective in cancer prevention and suppression. Myeloid-derived suppressor cells (MDSCs) are known to inhibit anti-tumor immunity; however, whether SFN regulates the anti-tumor activity of MDSCs in breast cancer is still unknown. In the current study, we found that SFN blocked prostaglandin E2 (PGE2) synthesis in parental and doxorubicin (DOX)-resistant breast cancer 4T1 cell lines by activating NF-E2-related factor 2 (Nrf2). Nrf2-mediated reduction of PGE2 was dependent on the enhanced expression of heme oxygenase 1 (HO-1) and glutamate-cysteine ligase (GCLC), and decreased COX-2 expression in breast cancer cells. Moreover, our study further revealed that reduced PGE2 secretion from SFN-treated 4T1 cells triggered MDSCs to switch to an immunogenic phenotype, enhancing the anti-tumor activities of CD8+ T cells. Co-administration of SFN and DOX was more efficacious for the treatment of breast cancer in a mouse model than either agent alone, as evidenced by the significant decrease in tumor volume, MDSC expansion, and increase in cytotoxic CD8+ T cells. Taken together, our data indicate that SFN reverses the immunosuppressive microenvironment and is a potent adjuvant chemotherapeutic candidate in breast cancer.

Nanoscale tungsten nitride/nitrogen-doped carbon as an efficient non-noble metal catalyst for hydrogen and oxygen recombination at room temperature in nickel-iron batteries.

A nanoscale tungsten nitride/nitrogen-doped carbon (WN/NC) catalyst was synthesized through a facile route, and it exhibited efficient catalytic performance for hydrogen and oxygen recombination at room temperature with an average catalytic velocity of 140 µmol h-1 gcat -1 and long catalytic life of 954 660 s without decay in the catalytic performance. With the WN/NC catalyst, a nickel-iron battery could be sealed and maintenance-free, and it also exhibited low cost; thus, the nickel-iron battery can be used for large-scale energy storage systems in rural/remote areas.

Heterologous expression and functional characterization of a novel chitinase from the chitinolytic bacterium Chitiniphilus shinanonensis.

Chitiniphilus shinanonensis strain SAY3(T) is a chitinolytic bacterium isolated from moat water of Ueda Castle in Nagano Prefecture, Japan. Fifteen genes encoding putative chitinolytic enzymes (chiA-chiO) have been isolated from this bacterium. Five of these constitute a single operon (chiCDEFG). The open reading frames of chiC, chiD, chiE, and chiG show sequence similarity to family 18 chitinases, while chiF encodes a polypeptide with two chitin-binding domains but no catalytic domain. Each of the five genes was successfully expressed in Escherichia coli, and the resulting recombinant proteins were characterized. Four of the recombinant proteins (ChiC, ChiD, ChiE, and ChiG) exhibited endo-type chitinase activity toward chitinous substrates, while ChiF showed no chitinolytic activity. In contrast to most endo-type chitinases, which mainly produce a dimer of N-acetyl-D-glucosamine (GlcNAc) as final product, ChiG completely split the GlcNAc dimer into GlcNAc monomers, indicating that it is a novel chitinase.

Isolation of genes coding for chitin-degrading enzymes in the novel chitinolytic bacterium, Chitiniphilus shinanonensis, and characterization of a gene coding for a family 19 chitinase.

Chitiniphilus shinanonensis type strain SAY3(T) is a strongly chitinolytic bacterium, originally isolated from the moat water in Ueda, Japan. To elucidate the chitinolytic activity of this strain, 15 genes (chiA-chiO) coding for putative chitin-degrading enzymes were isolated from a genomic library. Sequence analysis revealed the genes comprised 12 family 18 chitinases, a family 19 chitinase, a family 20 ß-N-acetylglucosaminidase, and a polypeptide with a chitin-binding domain but devoid of a catalytic domain. Two operons were detected among the sequences: chiCDEFG and chiLM. The gene coding for the polypeptide (chiN) showed sequence similarity to family 19 chitinases and was successfully expressed in Escherichia coli. ChiN demonstrated a multi-domain structure, composed of the N-terminal, two chitin-binding domains connected by a Pro- and Thr-rich linker, and a family 19 catalytic domain located at the C-terminus. The recombinant protein rChiN catalyzed an endo-type cleavage of N-acetyl-d-glucosamine oligomers, and also degraded insoluble chitin and soluble chitosan (degree of deacetylation of 80%). rChiN exhibited an inhibitory effect on hyphal growth of the fungus Trichoderma reesei. The chitin-binding domains of ChiN likely play an important role in the degradation of insoluble chitin, and are responsible for a growth inhibitory effect on fungi.