Tumor-derived extracellular vesicles as a biomarker for breast cancer diagnosis and metastasis monitoring.

It is imperative to explore biomarkers that are both precise and readily accessible in the comprehensive management of breast cancer. A multicenter cohort, including 512 breast cancer patients and 198 nonneoplastic individuals, was recruited to detect the level of tumor-derived extracellular vesicles using our method based on dual DNA tetrahedral nanostructures. The level of tumor-derived extracellular vesicles was significantly higher in newly diagnosed breast cancer patients than in nonneoplastic individuals at a cutoff value of 3.58 U/µL. For postoperative metastasis monitoring, the level of tumor-derived extracellular vesicles was significantly higher in breast cancer patients with metastasis than in those without metastasis at a cutoff value of 3.91 U/µL. Its efficacy of diagnosis and metastasis monitoring was superior to traditional tumor markers. Elevated level of tumor-derived extracellular vesicles served as a predictive biomarker for diagnosis and metastasis monitoring in breast cancer patients.

Cross-talk of Three Molecular Subtypes of Telomere Maintenance Defines Clinical Characteristics and Tumor Microenvironment in Gastric Cancer.

Background: Telomere maintenance takes part in the regulation of gastric cancer (GC) pathogenesis and is essential for patients' clinical features. Though the correlation between a single telomere maintenance-related gene and GC has previously been published, comprehensive exploration and systematic analysis remain to be studied. Our study is aimed at determining telomere maintenance-related molecular subtypes and examining their role in GC. Methods: By analyzing the transcriptome data, we identified three telomere maintenance-associated clusters (TMCs) with heterogeneity in clinical features and tumor microenvironment (TME). Then, we screened five prognostic telomere maintenance-related genes and established corresponding TM scores. Additionally, the expression level and biological function of tubulin beta 6 class V (TUBB6) were validated in GC tissues and cells. Results: TMC1 was correlated with EMT and TGF-beta pathway and predicted low tumor mutation burden (TMB) as well as bad prognostic outcomes. TMC3 was associated with cell cycle and DNA repair. In terms of TMB and overall survival, TMC3 exhibited opposite results against TMC1. Significant heterogeneity was observed between TMCs. TUBB6 was upregulated and could promote GC proliferation, migration, and invasion. Conclusion: Altogether, combining bioinformatics and functional experiments, we identified three molecular subtypes based on telomere maintenance-associated genes in GC, which could bring new ideas and novel biomarkers to the clinic.

Roles of estrogen receptors during sexual reversal in Pelodiscus sinensis.

BACKGROUND: The Chinese soft-shelled turtle, Pelodiscus sinensis, exhibits distinct sexual dimorphism, with the males growing faster and larger than the females. During breeding, all-male offspring can be obtained using 17ß-estradiol (E2). However, the molecular mechanisms underlying E2-induced sexual reversal have not yet been elucidated. Previous studies have investigated the molecular sequence and expression characteristics of estrogen receptors (ERs). METHODS AND RESULTS: In this study, primary liver cells and embryos of P. sinensis were treated with ER agonists or inhibitors. Cell incubation experiments revealed that nuclear ERs (nERs) were the main pathway for the transmission of estrogen signals. Our results showed that ERα agonist (ERα-ag) upregulated the expression of Rspo1, whereas ERα inhibitor (ERα-Inh) downregulated its expression. The expression of Dmrt1 was enhanced after ERα-Inh + G-ag treatment, indicating that the regulation of male genes may not act through a single estrogen receptor, but a combination of ERs. In embryos, only the ERα-ag remarkably promoted the expression levels of Rspo1, Wnt4, and ß-catenin, whereas the ERα-Inh had a suppressive effect. Additionally, Dmrt1, Amh, and Sox9 expression levels were downregulated after ERß inhibitor (ERß-Inh) treatment. GPER agonist (G-ag) has a significant promotion effect on Rspo1, Wnt4, and ß-catenin, while the inhibitor G-Inh does not affect male-related genes. CONCLUSIONS: Overall, these results suggest that ERs play different roles during sexual reversal in P. sinensis and ERα may be the main carrier of estrogen-induced sexual reversal in P. sinensis. Further studies need to be performed to analyze the mechanism of ER action.

Hypoxia stimulates CTC-platelet cluster formation to promote breast cancer metastasis.

Circulating tumor cell clusters/micro-emboli (CTM) possess greater metastatic capacity and survival advantage compared to individual circulating tumor cell (CTC). However, the formation of CTM subtypes and their role in tumor metastasis remain unclear. In this study, we used a microfluidic Cluster-Chip with easy operation and high efficiency to isolate CTM from peripheral blood, which confirmed their correlation with clinicopathological features and identified the critical role of CTC-platelet clusters in breast cancer metastasis. The correlation between platelets and CTM function was further confirmed in a mouse model and RNA sequencing of CTM identified high-expressed genes related to hypoxia stimulation and platelet activation which possibly suggested the correlation of hypoxia and CTC-platelet cluster formation. In conclusion, we successfully developed the Cluster-Chip platform to realize the clinical capture of CTMs and analyze the biological properties of CTC-platelet clusters, which could benefit the design of potential treatment regimens to prevent CTM-mediated metastasis and tumor malignant progression.

Construction of a Low-Resistivity Carbon Layer To Improve Performance in Dimethyl Sulfoxide-Based Kesterite Solar Cells.

Morphology of the absorber plays a decisive role in photoelectric conversion efficiency (PCE) of kersterite solar cells. Cu2ZnSn(S,Se)4 (CZTSSe) grain prepared from dimethyl sulfoxide (DMSO)-based solution easily grows into large grains, which can lead to the formation of some holes at the back of the absorber. These holes cause the recombination of photocarriers and greatly weaken the performance of CZTSSe devices. Here, trace amounts of thioglycolic acid (TGA) are introduced to the DMSO-based solution, and a combination of TGA and metal is formed in the absorber, leading to the formation of fine grains in the CZTSSe absorber. Next, post-annealing (PA) in a N2 atmosphere is performed to promote Na diffusion, helping the transition from a fine-grain layer to a low-resistivity carbon layer at the interface between CZTSSe and Mo and avoiding the drawbacks of the DMSO-based system. Finally, the champion PCE of the CZTSSe device can be improved to 10.05% from 8.06%. The conclusions demonstrate that the construction of a carbon layer can boost the performance of CZTSSe devices.

Engineering the Tat-secretion pathway of Bacillus licheniformis for the secretion of cytoplasmic enzyme arginase.

The industrial bacterium Bacillus licheniformis has long been used as a microbial factory for the production of enzymes due to its ability to secrete copious amounts of native extracellular proteins and its generally regarded as safe (GRAS) status. However, most attempts to use B. licheniformis to produce heterologous and cytoplasmic enzymes primarily via the general secretory (Sec) pathway have had limited success. The twin-arginine transport (Tat) pathway offers a promising alternative for the extracellular export of Sec-incompatible proteins because it transports full, correctly folded proteins. However, compared to the Sec pathway, the yields of the Tat pathway have historically been too low for commercial use. To improve the export efficiency of the Tat pathway, we identified the optimal Tat-dependent signal peptides and increased the abundance of the Tat translocases, the signal peptidase (SPase), and the intracellular chaperones. These strategic modifications significantly improved the Tat-dependent secretion of the cytoplasmic enzyme arginase into the culture medium using B. licheniformis. The extracellular enzymatic activity of arginase showed a 5.2-fold increase after these modifications. Moreover, compared to the start strain B. licheniformis 0F3, the production of extracellular GFP was improved by 3.8 times using the strategic modified strain B. licheniformis 0F13, and the extracellular enzymatic activity of SOX had a 1.3-fold increase using the strain B. licheniformis 0F14. This Tat-based production chassis has the potential for enhanced production of Sec-incompatible enzymes, therefore expanding the capability of B. licheniformis as an efficient cellular factory for the production of high-value proteins. KEY POINTS: • Systematic genetic modification of Tat-pathway in B. licheniformis. • Significant enhancement of the secretion capacity of Tat pathway for delivery the cytoplasmic enzyme arginase. • A new platform for efficient extracellular production of Sec-incompatible enzymes.

Novel Covalent Probe Selectively Targeting Glutathione Peroxidase 4 In Vivo: Potential Applications in Pancreatic Cancer Therapy.

Glutathione peroxidase 4 (GPX4) emerges as a promising target for the treatment of therapy-resistant cancer through ferroptosis. Thus, there is a broad interest in the development of GPX4 inhibitors. However, a majority of reported GPX4 inhibitors utilize chloroacetamide as a reactive electrophilic warhead, and the selectivity and pharmacokinetic properties still need to be improved. Herein, we developed a compound library based on a novel electrophilic warhead, the sulfonyl ynamide, and executed phenotypic screening against pancreatic cancer cell lines. Notably, one compound A16 exhibiting potent cell toxicity was identified. Further chemical proteomics investigations have demonstrated that A16 specifically targets GPX4 under both in situ and in vivo conditions, inducing ferroptosis. Importantly, A16 exhibited superior selectivity and potency compared to reported GPX4 inhibitors, ML210 and ML162. This provides the structural diversity of tool probes for unraveling the fundamental biology of GPX4 and exploring the therapeutic potential of pancreatic cancer via ferroptosis induction.

Global Reactivity Profiling of the Catalytic Lysine in Human Kinome for Covalent Inhibitor Development.

Advances in targeted covalent inhibitors (TCIs) have been made by using lysine-reactive chemistries. Few aminophiles possessing balanced reactivity/stability for the development of cell-active TCIs are however available. We report herein lysine-reactive activity-based probes (ABPs; 2-14) based on the chemistry of aryl fluorosulfates (ArOSO2 F) capable of global reactivity profiling of the catalytic lysine in human kinome from mammalian cells. We concurrently developed reversible covalent ABPs (15/16) by installing salicylaldehydes (SA) onto a promiscuous kinase-binding scaffold. The stability and amine reactivity of these probes exhibited a broad range of tunability. X-ray crystallography and mass spectrometry (MS) confirmed the successful covalent engagement between ArOSO2 F on 9 and the catalytic lysine of SRC kinase. Chemoproteomic studies enabled the profiling of >300 endogenous kinases, thus providing a global landscape of ligandable catalytic lysines of the kinome. By further introducing these aminophiles into VX-680 (a noncovalent inhibitor of AURKA kinase), we generated novel lysine-reactive TCIs that exhibited excellent in vitro potency and reasonable cellular activities with prolonged residence time. Our work serves as a general guide for the development of lysine-reactive ArOSO2 F-based TCIs.

Estrogen receptor beta suppresses the androgen receptor oncogenic effects in triple-negative breast cancer.

BACKGROUND: Triple-negative breast cancer (TNBC) is an aggressive type of breast cancer associated with poor prognosis and limited treatment options. The androgen receptor (AR) has emerged as a potential therapeutic target for luminal androgen receptor (LAR) TNBC. However, multiple studies have claimed that anti-androgen therapy for AR-positive TNBC only has limited clinical benefits. This study aimed to investigate the role of AR in TNBC and its detailed mechanism. METHODS: Immunohistochemistry and TNBC tissue sections were applied to investigate AR and nectin cell adhesion molecule 4 (NECTIN4) expression in TNBC tissues. Then, in vitro and in vivo assays were used to explore the function of AR and estrogen receptor beta (ERß) in TNBC. Chromatin immunoprecipitation sequencing (ChIP-seq), co-immunoprecipitation (co-IP), molecular docking method, and luciferase reporter assay were performed to identify key molecules that affect the function of AR. RESULTS: Based on the TNBC tissue array analysis, we revealed that ERß and AR were positive in 21.92% (32/146) and 24.66% (36/146) of 146 TNBC samples, respectively, and about 13.70% (20/146) of TNBC patients were ERß positive and AR positive. We further demonstrated the pro-tumoral effects of AR on TNBC cells, however, the oncogenic biology was significantly suppressed when ERß transfection in LAR TNBC cell lines but not in AR-negative TNBC. Mechanistically, we identified that NECTIN4 promoter -42 bp to -28 bp was an AR response element, and that ERß interacted with AR thus impeding the AR-mediated NECTIN4 transcription which promoted epithelial-mesenchymal transition in tumor progression. CONCLUSIONS: This study suggests that ERß functions as a suppressor mediating the effect of AR in TNBC prognosis and cell proliferation. Therefore, our current research facilitates a better understanding of the role and mechanisms of AR in TNBC carcinogenesis.

Integrating High-Sensitivity Photodetector and High-Energy Aqueous Battery in All-in-One Triple-Twisted Fiber.

Fiber-shaped photodetectors (FPDs) have attracted special attention to wearable health monitoring due to their 3D absorption capabilities. However, the practical application of traditional FPDs is severely limited by the irreversible degradation of performance caused by vulnerable interface compatibility on complex deformation and a single function. Here, an integrated photoelectrochemical FPD/battery device (FPDB) is designed, consisting of a common electrode, photoanode, anode, and sol-gel electrolyte as an isolation layer, which not only effectively avoids the short circuit problem of FPD but also endows high-efficiency energy storage capacity. As expected, the resulting all-in-one triple-twisted fiber-shaped FPDB simultaneously achieves high responsiveness of 151.45 mA W-1 and excellent volume capacity of 18.75 mAh cm-3. Such a stable architectural design and multifunctional integration of functional fibers accelerate the development of next-generation wearable fabrics.

Multitarget inhibitors/probes that target LRRK2 and AURORA A kinases noncovalently and covalently.

Herein, we report a salicylaldehyde-based, reversible covalent inhibitor (A2) that possesses moderate cellular activity against AURKA with a prolonged residence time and shows significant non-covalent inhibition towards LRRK2. Our results indicated that this multitarget kinase inhibitor may be used as the starting point for future development of more potent, selective and dual-targeting covalent kinase inhibitors against AURKA and LRRK2 for mitophagy.

Midbrain dopamine oxidation links ubiquitination of glutathione peroxidase 4 to ferroptosis of dopaminergic neurons.

Parkinson's disease (PD) is a neurodegenerative disorder characterized by the gradual loss of midbrain dopaminergic neurons in association with aggregation of α-synuclein. Oxidative damage has been widely implicated in this disease, though the mechanisms involved remain elusive. Here, we demonstrated that preferential accumulation of peroxidized phospholipids and loss of the antioxidant enzyme glutathione peroxidase 4 (GPX4) were responsible for vulnerability of midbrain dopaminergic neurons and progressive motor dysfunctions in a mouse model of PD. We also established a mechanism wherein iron-induced dopamine oxidation modified GPX4, thereby rendering it amenable to degradation via the ubiquitin-proteasome pathway. In conclusion, this study unraveled what we believe to be a novel pathway for dopaminergic neuron degeneration during PD pathogenesis, driven by dopamine-induced loss of antioxidant GPX4 activity.

Ynamide Coupling Reagent for the Chemical Cross-Linking of Proteins in Live Cells.

Chemical cross-linking of proteins coupled with mass spectrometry analysis (CXMS) is a powerful method for the study of protein structure and protein-protein interactions (PPIs). However, the chemical probes used in the CXMS are limited to bidentate reactive warheads, and the available zero-length cross-linkers are restricted to 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride/N-hydroxysuccinimide (EDC/NHS) and 4-(4,6-dimethoxy-1,3,5-triazin-2-yl)-4-methylmorpholinium chloride (DMTMM). To alleviate this issue, an efficient coupling reagent, sulfonyl ynamide, was developed as a new zero-length cross-linker that can connect high-abundance carboxyl residues (D/E) with lysine (K) to form amide bonds in the absence of any catalyst. Significant improvement in the cross-linking efficiency and specificity in comparison with traditional EDC/NHS was achieved with model proteins, which includes inter- and intramolecular conjugations. The cross-linked structures were validated by X-ray crystallography. Importantly, this coupling reagent can be successfully used to capture interacting proteins in the whole proteome and can be a useful reagent for probing potential protein-protein interactions in situ.

c-Myc-Targeting PROTAC Based on a TNA-DNA Bivalent Binder for Combination Therapy of Triple-Negative Breast Cancer.

Triple-negative breast cancer (TNBC) is highly aggressive with a poor clinical prognosis and no targeted therapy. The c-Myc protein is a master transcription factor and a potential therapeutic target for TNBC. In this study, we develop a PROTAC (PROteolysis TArgeting Chimera) based on TNA (threose nucleic acid) and DNA that effectively targets and degrades c-Myc. The TNA aptamer is selected in vitro to bind the c-Myc/Max heterodimer and appended to the E-box DNA sequence to create a high-affinity, biologically stable bivalent binder. The TNA-E box-pomalidomide (TEP) conjugate specifically degrades endogenous c-Myc/Max, inhibits TNBC cell proliferation, and sensitizes TNBC cells to the cyclin-dependent kinase inhibitor palbociclib in vitro. In a mouse TNBC model, combination therapy with TEP and palbociclib potently suppresses tumor growth. This study offers a promising nucleic acid-based PROTAC modality for both chemical biology studies and therapeutic interventions of TNBC.

Development and validation of a risk prediction model for diabetic retinopathy in type 2 diabetic patients.

To establish a risk prediction model and make individualized assessment for the susceptible diabetic retinopathy (DR) population in type 2 diabetic mellitus (T2DM) patients. According to the retrieval strategy, inclusion and exclusion criteria, the relevant meta-analyses on DR risk factors were searched and evaluated. The pooled odds ratio (OR) or relative risk (RR) of each risk factor was obtained and calculated for ß coefficients using logistic regression (LR) model. Besides, an electronic patient-reported outcome questionnaire was developed and 60 cases of DR and non-DR T2DM patients were investigated to validate the developed model. Receiver operating characteristic curve (ROC) was drawn to verify the prediction accuracy of the model. After retrieving, eight meta-analyses with a total of 15,654 cases and 12 risk factors associated with the onset of DR in T2DM, including weight loss surgery, myopia, lipid-lowing drugs, intensive glucose control, course of T2DM, glycated hemoglobin (HbA1c), fasting plasma glucose, hypertension, gender, insulin treatment, residence, and smoking were included for LR modeling. These factors, followed by the respective ß coefficient was bariatric surgery (- 0.942), myopia (- 0.357), lipid-lowering drug follow-up < 3y (- 0.994), lipid-lowering drug follow-up > 3y (- 0.223), course of T2DM (0.174), HbA1c (0.372), fasting plasma glucose (0.223), insulin therapy (0.688), rural residence (0.199), smoking (- 0.083), hypertension (0.405), male (0.548), intensive glycemic control (- 0.400) with constant term α (- 0.949) in the constructed model. The area under receiver operating characteristic curve (AUC) of the model in the external validation was 0.912. An application was presented as an example of use. In conclusion, the risk prediction model of DR is developed, which makes individualized assessment for the susceptible DR population feasible and needs to be further verified with large sample size application.

Circulating Tumor DNA as a Novel Biomarker Optimizing Treatment for Triple Negative Breast Cancer.

Triple-negative breast cancer is a sub-type of clinically and molecularly heterogeneous malignant disease with a worse prognosis and earlier recurrence than HER2-amplified or hormone-receptor positive breast cancer. Because of the lack of personalized therapy, genetic information is essential to early diagnosing, identifying the high risk of recurrence, guiding therapeutic management, and monitoring treatment efficiency. Circulating tumor DNA (ctDNA) is a novel noninvasive, timely, and tumor specified biomarker that reliably reflects the comprehensive tumor genetic profiles. Thus, it holds significant expectations in personalized therapy, including accurate diagnosis, treatment monitoring, and early detection of recurrence of TNBC. In this review, we summarize the results from recent and ongoing ctDNA-based biomarker-driven clinical trials, with respect to ctDNA analysis' predictive role, in adjuvant, neo-adjuvant, and metastatic settings. Collectively, we anticipate that ctDNA will ultimately be integrated into the management of TNBC to foster precise treatment.

2-Ethynylbenzaldehyde-Based, Lysine-Targeting Irreversible Covalent Inhibitors for Protein Kinases and Nonkinases.

Lysine-targeting irreversible covalent inhibitors have attracted growing interests in recent years, especially in the fields of kinase research. Despite encouraging progress, few chemistries are available to develop inhibitors that are exclusively lysine-targeting, selective, and cell-active. We report herein a 2-ethynylbenzaldehyde (EBA)-based, lysine-targeting strategy to generate potent and selective small-molecule inhibitors of ABL kinase by selectively targeting the conserved catalytic lysine in the enzyme. We showed the resulting compounds were cell-active, capable of covalently engaging endogenous ABL kinase in K562 cells with long-residence time and few off-targets. We further validated the generality of this strategy by developing EBA-based irreversible inhibitors against EGFR (a kinase) and Mcl-1 (a nonkinase) that covalently reacted with the catalytic and noncatalytic lysine within each target.

Novel n-i CeO2/a-Al2O3 Heterostructure Electrolyte Derived from the Insulator a-Al2O3 for Fuel Cells.

Heterostructure technologies have been regarded as promising methods in the development of electrolytes with high ionic conductivity for low-temperature solid oxide fuel cells (LT-SOFCs). Here, a novel semiconductor/insulator (n-i) heterostructure strategy has been proposed to develop composite electrolytes for LT-SOFCs based on CeO2 and the insulator amorphous alumina (a-Al2O3). The constructed CeO2/a-Al2O3 electrolyte exhibits an ionic conductivity of up to 0.127 S cm-1, and its fuel cell achieves a maximum power density (MPD) of 1017 mW cm-2 with an open-circuit voltage (OCV) of 1.14 V at 550 °C without the short-circuiting problem, suggesting that the introduction of a-Al2O3 can effectively suppress the electron conduction of CeO2. It is found that the potential energy barrier at the heterointerfaces caused by the ultrawide band gap of the insulator a-Al2O3 plays an important role in restraining electron conduction. Simultaneously, the thermoelectric effect of the insulator induces more oxygen vacancies because of interface charge compensation, which further promotes ionic transport and results in high ionic conductivity and fuel cell performance. This study presents a practical n-i heterostructure electrolyte design, and further research confirmed the advanced functionality of the CeO2/a-Al2O3 electrolyte. Our study may open frontiers in the field of developing high-efficiency electrolytes of LT-SOFCs using insulating materials such as amorphous alumina.

Inhibition of MYC suppresses programmed cell death ligand-1 expression and enhances immunotherapy in triple-negative breast cancer.

BACKGROUND: Cancer immunotherapy has emerged as a promising strategy against triple-negative breast cancer (TNBC). One of the immunosuppressive pathways involves programmed cell death-1 (PD-1) and programmed cell death ligand-1 (PD-L1), but many patients derived little benefit from PD-1/PD-L1 checkpoint blockades treatment. Prior research has shown that MYC, a master transcription amplifier highly expressed in TNBC cells, can regulate the tumor immune microenvironment and constrain the efficacy of immunotherapy. This study aims to investigate the regulatory relationship between MYC and PD-L1, and whether a cyclin-dependent kinase (CDK) inhibitor that inhibits MYC expression in combination with anti-PD-L1 antibodies can enhance the response to immunotherapy. METHODS: Public databases and TNBC tissue microarrays were used to study the correlation between MYC and PD-L1. The expression of MYC and PD-L1 in TNBCs was examined by quantitative real-time polymerase chain reaction and Western blotting. A patient-derived tumor xenograft (PDTX) model was used to evaluate the influence of a CDK7 inhibitor THZ1 on PD-L1 expression. Cell proliferation and migration were detected by 5-ethynyl-2'-deoxyuridine (EdU) cell proliferation and cell migration assays. Tumor xenograft models were established for in vivo verification. RESULTS: A high MYC expression level was associated with a poor prognosis and could alter the proportion of tumor-infiltrating immune cells (TIICs). The positive correlation between MYC and PD-L1 was confirmed by immunostaining samples from 165 TNBC patients. Suppression of MYC in TNBC caused a reduction in the levels of both PD-L1 messenger RNA and protein. In addition, antitumor immune response was enhanced in the TNBC cancer xenograft mouse model with suppression of MYC by CDK7 inhibitor THZ1. CONCLUSIONS: The combined therapy of CDK7 inhibitor THZ1 and anti-PD-L1 antibody appeared to have a synergistic effect, which might offer new insight for enhancing immunotherapy in TNBC.