Clinical efficacy of tumor organoid-guided cancer therapy for locally advanced unresectable or metastatic breast cancer.

Patient-derived organoids (PDOs) may facilitate treatment selection. This retrospective cohort study evaluated the feasibility and clinical benefit of using PDOs to guide personalized treatment in metastatic breast cancer (MBC). Patients diagnosed with MBC were recruited between January 2019 and August 2022. PDOs were established and the efficacy of customized drug panels was determined by measuring cell mortality after drug exposure. Patients receiving organoid-guided treatment (OGT) were matched 1:2 by nearest neighbor propensity scores with patients receiving treatment of physician's choice (TPC). The primary outcome was progression-free survival. Secondary outcomes included objective response rate and disease control rate. Targeted gene sequencing and pathway enrichment analysis were performed. Forty-six PDOs (46 of 51, 90.2%) were generated from 45 MBC patients. PDO drug screening showed an accuracy of 78.4% (95% CI 64.9%-91.9%) in predicting clinical responses. Thirty-six OGT patients were matched to 69 TPC patients. OGT was associated with prolonged median progression-free survival (11.0 months vs. 5.0 months; hazard ratio 0.53 [95% CI 0.33-0.85]; p = .01) and improved disease control (88.9% vs. 63.8%; odd ratio 4.26 [1.44-18.62]) compared with TPC. The objective response rate of both groups was similar. Pathway enrichment analysis in hormone receptor-positive, human epidermal growth factor receptor 2-negative patients demonstrated differentially modulated pathways implicated in DNA repair and transcriptional regulation in those with reduced response to capecitabine/gemcitabine, and pathways associated with cell cycle regulation in those with reduced response to palbociclib. Our study shows that PDO-based functional precision medicine is a feasible and effective strategy for MBC treatment optimization and customization.

Site-Directed Electrochemical Grafting for Amplified Detection of Antibody Pharmaceuticals.

Antibody pharmaceuticals have become the most popular immunotherapeutic drugs and are often administered with low serum drug dosages. Hence, the development of a highly sensitive method for the quantitative assay of antibody levels is of great importance to individualized therapy. On the basis of the dual signal amplification by the glycan-initiated site-directed electrochemical grafting of polymer chains (glyGPC), we report herein a novel strategy for the amplified electrochemical detection of antibody pharmaceuticals. The target of interest was affinity captured by a DNA aptamer ligand, and then the glycans of antibody pharmaceuticals were decorated with the alkyl halide initiators (AHIs) via boronate cross-linking, followed by the electrochemical grafting of the ferrocenyl polymer chains from the glycans of antibody pharmaceuticals through the electrochemically controlled atom transfer radical polymerization (eATRP). As the glycans can be decorated with multiple AHIs and the grafted polymer chains are composed of tens to hundreds of electroactive tags, the glyGPC-based strategy permits the dually amplified electrochemical detection of antibody pharmaceuticals. In the presence of trastuzumab (Herceptin) as the target, the glyGPC-based strategy achieved a detection limit of 71.5 pg/mL. Moreover, the developed method is highly selective, and the results of the quantitative assay of trastuzumab levels in human serum are satisfactory. Owing to its uncomplicated operation and cost-effectiveness, the glyGPC-based strategy shows great promise in the amplified electrochemical detection of antibody pharmaceuticals.

Genomics of predictive radiation mutagenesis in oilseed rape: modifying seed oil composition.

Rapeseed is a crop of global importance but there is a need to broaden the genetic diversity available to address breeding objectives. Radiation mutagenesis, supported by genomics, has the potential to supersede genome editing for both gene knockout and copy number increase, but detailed knowledge of the molecular outcomes of radiation treatment is lacking. To address this, we produced a genome re-sequenced panel of 1133 M2 generation rapeseed plants and analysed large-scale deletions, single nucleotide variants and small insertion-deletion variants affecting gene open reading frames. We show that high radiation doses (2000 Gy) are tolerated, gamma radiation and fast neutron radiation have similar impacts and that segments deleted from the genomes of some plants are inherited as additional copies by their siblings, enabling gene dosage decrease. Of relevance for species with larger genomes, we showed that these large-scale impacts can also be detected using transcriptome re-sequencing. To test the utility of the approach for predictive alteration of oil fatty acid composition, we produced lines with both decreased and increased copy numbers of Bna.FAE1 and confirmed the anticipated impacts on erucic acid content. We detected and tested a 21-base deletion expected to abolish function of Bna.FAD2.A5, for which we confirmed the predicted reduction in seed oil polyunsaturated fatty acid content. Our improved understanding of the molecular effects of radiation mutagenesis will underpin genomics-led approaches to more efficient introduction of novel genetic variation into the breeding of this crop and provides an exemplar for the predictive improvement of other crops.

CCT2 prevented ß-catenin proteasomal degradation to sustain cancer stem cell traits and promote tumor progression in epithelial ovarian cancer.

BACKGROUND: Epithelial ovarian cancer (EOC) is featured by rapid progression and dismal outcomes clinically. Chaperonin Containing TCP1 Subunit 2 (CCT2) was identified as a crucial regulator for tumor progression, however, its exact role in EOC remained largely unknown. METHODS: CCT2 expression and prognostic value in EOC samples were assessed according to TCGA dataset. Proliferation and mobility potentials were assessed by CCK8, colony-formation, wound healing, and Transwell assays. Cancer stem cell (CSC) traits were evaluated by RT-PCR, WB assays, sphere-forming assay and chemoresistance analysis. Bioinformatic analysis, co-IP assays and ubiquitin assays were performed to explore the mechanisms of CCT2 on EOC cells. RESULTS: CCT2 highly expressed in EOC tissues and predicted poor prognosis of EOC patients by TCGA analysis. Silencing CCT2 significantly restrained cell proliferation, migration, and invasion. Moreover, CCT2 could effectively trigger epithelial-mesenchymal transition to confer extensive invasion potentials to EOC cells, Importantly, CCT2 positively correlated with CSC markers in EOC, and CCT2 knockdown impaired CSC traits and sensitize EOC cells to conventional chemotherapy regimens. Contrarily, overexpressing CCT2 achieved opposite results. Mechanistically, CCT2 exerted its pro-oncogene function by triggering Wnt/ß-catenin signaling. Specifically, CCT2 could recruit HSP105-PP2A complex, a well-established dephosphorylation complex, to ß-catenin via direct physical interaction to prevent phosphorylation-induced proteasomal degradation of ß-catenin, resulting in intracellular accumulation of active ß-catenin and increased signaling activity. CONCLUSIONS: CCT2 was a novel promotor for EOC progression and a crucial sustainer for CSC traits mainly by preventing ß-catenin degradation. Targeting CCT2 may represent a promising therapeutic strategy for EOC.

The relation between effortful control and executive function training in preschoolers.

In recent years, the question of whether executive function (EF) is malleable has been widely documented. Despite using the same training tasks, transfer effects remain uncertain. Researchers suggested that the inconsistency might be attributed to individual differences in temperamental traits. In the current study, we investigated how effortful control, a temperamental trait, would affect EF training outcomes in children. Based on parent rating, 79 6-year-old preschoolers were identified as having higher or lower effort control and were assigned to three conditions: working memory (WM) training, inhibitory control (IC) training, and a business-as-usual control group. Children completed assessments at baseline, 1 week after intervention (posttest), and 3 months after intervention (follow-up). As compared with the control group, the WM and IC training groups showed improvement in both trained tasks and nontrained measures. At baseline, children with higher effortful control scores showed greater WM capacity and better IC. Furthermore, effortful control was positively correlated with training gain in both training groups, with children with higher effortful control benefitting more through training. In the WM training group, effortful control was positively correlated with near transfer on WM outcomes both immediately and longitudinally. At posttest, the WM and IC training groups showed a positive correlation between effortful control and fluid intelligence performance. Our results underscore the importance of individual differences in training benefits, in particular the role of effortful control, and further illustrate the potential avenues for designing more effective individualized cognitive training programs to foster learning and optimize children's development.

Screening of active components in Astragalus mongholicus Bunge and Panax notoginseng formula for anti-fibrosis in CKD: nobiletin inhibits Lgals1/PI3K/AKT signaling to improve renal fibrosis.

The Astragalus mongholicus Bunge and Panax notoginseng formula (A&P) has been clinically shown to effectively slow down the progression of chronic kidney disease (CKD) and has demonstrated significant anti-fibrosis effects in experimental CKD model. However, the specific active ingredients and underlying mechanism are still unclear. The active ingredients of A&P were analyzed by Ultra-high performance liquid chromatography-tandem mass spectrometry (UPLC-HR-MS). A mouse model of CKD was constructed by 5/6 nephrectomy. Renal function was assessed by creatinine and urea nitrogen. Real-time PCR and Western Blot were performed to detect the mRNA and protein changes in kidney and cells. An in vitro fibrotic cell model was constructed by TGF-ß induction in TCMK-1 cells. The results showed that thirteen active ingredients of A&P were identified by UPLC-HR-MS, nine of which were identified by analysis with standards, among which the relative percentage of NOB was high. We found that NOB treatment significantly improved renal function, pathological damage and reduced the expression level of fibrotic factors in CKD mice. The results also demonstrated that Lgals1 was overexpressed in the interstitial kidney of CKD mice, and NOB treatment significantly reduced its expression level, while inhibiting PI3K and AKT phosphorylation. Interestingly, overexpression of Lgals1 significantly increased fibrosis in TCMK1 cells and upregulated the activity of PI3K and AKT, which were strongly inhibited by NOB treatment. NOB is one of the main active components of A&P. The molecular mechanism by which NOB ameliorates renal fibrosis in CKD may be through the inhibition of Lgals1/PI3K/AKT signaling pathway.

The Relationship between Socioeconomic Status and Academic Achievement is Mediated and Moderated by Executive Functions.

Executive function is an underlying mechanism linking family socioeconomic status (SES) and academic achievement. Previous studies mainly investigated either the mediating or moderating role of executive function within this relationship, which either overlook the individual differences that are independent of the environment or neglect the influence of the environment on shaping personal factors. To avoid a piecemeal approach to theory, the current study aimed to test the mediating and moderating roles of executive function in a single analytic model. Two hundred and thirty-six Chinese fifth graders (Mage = 10.70 years, SD = 0.49, range = 10.23-11.75 years, and 40.30% girls) were recruited. Their executive function performance was measured using eight different tasks, and their Chinese literacy skills and mathematics achievement were assessed by routine school evaluations. One year after the initial assessment, children's academic achievements were evaluated again. Results demonstrated that parental SES positively predicted children's academic achievement when controlling for prior academic achievement, and children's executive function mediated this relationship. Also, executive function moderated the association between SES and academic achievement in that, the negative predictive effect of low SES on academic achievement was only significant for children with lower levels of executive function, which is not shown in children with higher levels of executive function. By demonstrating the dual roles of executive function in the SES-achievement link, this work provides evidence for supporting the optimal development of children from diverse socioeconomic backgrounds and emphasizes the significance of developing individualized intervention strategies on executive function to mitigate the negative effect of low SES on children's academic achievement.

Target-Synergized Biologically Mediated RAFT Polymerization for Electrochemical Aptasensing of Femtomolar Thrombin.

The assay of thrombin levels is integral to the assessment of coagulation function and clinical screening of coagulation disorder-related diseases. In this work, we illustrate the ingenious use of the target-synergized biologically mediated reversible addition-fragmentation chain transfer (RAFT) polymerization (tsBMRP) as a novel amplification strategy for the electrochemical aptamer-based biosensing of thrombin at the femtomolar levels. Briefly, the tsBMRP-based strategy relies on the boronate affinity-mediated decoration of the glycan chain(s) of the target itself with RAFT agents and the subsequent recruitment of signal labels via BMRP, mediated by the direct reduction of RAFT agents by NADH into initiating/propagating radicals. Obviously, the tsBMRP-based strategy is biologically friendly, low-cost, and simple in operation. As thrombin is a glycoconjugate, its electrochemical aptasensing involves the use of the thrombin-binding aptamer (TBA) as the recognition receptor, the site-specific decoration of RAFT agents to the glycan chain of thrombin via boronate affinity, and further the recruitment of ferrocene signal labels via the BMRP of ferrocenylmethyl methacrylate (FcMMA). As boronate affinity results in the decoration of each glycan chain with tens of RAFT agents while BMRP recruits hundreds of signal labels to each RAFT agent-decorated site, the tsBMRP-based strategy allows us to detect thrombin at a concentration of 35.3 fM. This electrochemical aptasensor is highly selective, and its applicability to thrombin detection in serum samples has been further demonstrated. The merits of high sensitivity and selectivity, low cost, good anti-interference capability, and simple operation make the tsBMRP-based electrochemical thrombin aptasensor great promise in biomedical and clinical applications.

Amplification-Free Ratiometric Electrochemical Aptasensor for Point-of-Care Detection of Therapeutic Monoclonal Antibodies.

The rapid quantification of therapeutic monoclonal antibodies (mAbs) is of great significance to their pharmacokinetics/pharmacodynamics (PK/PD) research and the personalized medication for disease treatment. Taking advantage of the direct decoration of tens of redox tags to the target of interest, we illustrate herein an amplification-free ratiometric electrochemical aptasensor for the point-of-care (POC) detection of trace amounts of therapeutic mAbs. The POC detection of therapeutic mAbs involved the use of the methylene blue (MB)-conjugated aptamer as the affinity element and the decoration of therapeutic mAbs with ferrocene (Fc) tags via the boronate crosslinking, in which the MB-derived peak current was used as the reference signal, and the peak current of the Fc tag was used as the output signal. As each therapeutic mAb carries tens of diol sites for the site-specific decoration of the Fc output tags, the boronate crosslinking enabled the amplification-free detection, which is cost-effective and quite simple in operation. In the presence of bevacizumab (BevMab) as the target, the resulting ratiometric signal (i.e., the IFc/IMB value) exhibited a good linear response over the range of 0.025-2.5 µg/mL, and the limit of detection (LOD) of the electrochemical aptasensor was 6.5 ng/mL. Results indicated that the aptamer-based affinity recognition endowed the detection of therapeutic mAbs with high selectivity, while the ratiometric readout exhibited satisfactory reproducibility and robustness. Moreover, the ratiometric electrochemical aptasensor is applicable to the detection of therapeutic mAbs in serum samples. Taking together, the amplification-free ratiometric electrochemical aptasensor holds great promise in the POC detection of therapeutic mAbs.

Dually Amplified Electrochemical Aptasensor for Endotoxin Detection via Target-Assisted Electrochemically Mediated ATRP.

As the entering of bacterial endotoxin into blood can cause various life-threatening pathological conditions, the screening and detection of low-abundance endotoxin are of great importance to human health. Taking advantage of signal amplification by target-assisted electrochemically mediated atom transfer radical polymerization (teATRP), we illustrate herein a simple and cost-effective electrochemical aptasensor capable of detecting endotoxin with high sensitivity and selectivity. Specifically, the aptamer receptor was employed for the selective capture of endotoxin, of which the glycan chain was then decorated with ATRP initiators via covalent coupling between the diol sites and phenylboronic acid (PBA) group, followed by the recruitment of ferrocene signal reporters via the grafting of polymer chains through potentiostatic eATRP under ambient temperature. As the glycan chain of endotoxin can be decorated with hundreds of ATRP initiators while the further grafting of polymer chains through eATRP can recruit hundreds to thousands of signal reporters to each initiator-decorated site, the teATRP-based strategy allows for the dual amplification of the detection signal. This dually amplified electrochemical aptasensor has the ability to sensitively and selectively detect endotoxin at a concentration as low as 1.2 fg/mL, and its practical applicability has been further demonstrated using human serum samples. Owing to the simplicity, high efficiency, biocompatibility, and inexpensiveness of the teATRP-based amplification strategy, this electrochemical aptasensor holds great application potential in the sensitive and selective detection of low-abundance endotoxin and many other glycan chain-containing bio-targets.

Boosting the Methanol Oxidation Reaction Activity of Pt-Ru Clusters via Resonance Energy Transfer.

The sluggish kinetics of the methanol oxidation reaction (MOR) with PtRu electrocatalyst severely hinder the commercialization of direct methanol fuel cells (DMFCs). The electronic structure of Pt is of significant importance for its catalytic activity. Herein, it is reported that low-cost fluorescent carbon dots (CDs) can regulate the behavior of the D-band center of Pt in PtRu clusters through resonance energy transfer (RET), resulting in a significant increase in the catalytic activity of the catalyst participating in methanol electrooxidation. For the first time, the bifunction of RET is used to provide unique strategy for fabrication of PtRu electrocatalysts, not only tunes the electronic structure of metals, but also provides an important role in anchoring metal clusters. Density functional theory calculations further prove that charge transfer between CDs and Pt promotes the dehydrogenation of methanol on PtRu catalysts and reduces the free energy barrier of the reaction associated with the oxidation of CO* to CO2 . This helps to improve the catalytic activity of the systems participating in MOR. The performance of the best sample is 2.76 times higher than that of commercial PtRu/C (213.0 vs 76.99  mW cm - 2 mg Pt - 1 ${\rm{mW\ cm}}^{ - 2}{\rm{\ mg}}_{{\rm{Pt}}}^{ - 1}$ ). The fabricated system can be potentially used for the efficient fabrication of DMFCs.

Sarcopenia and mild kidney dysfunction and risk of all-cause and cause-specific mortality in older adults.

BACKGROUND: Sarcopenia has been identified as a risk factor for increased mortality in individuals with CKD. However, when considering individuals with mild kidney dysfunction prior to CKD, the impact of sarcopenia on adverse outcomes, particularly mortality, remains uncertain. METHODS: This study included 323 801 participants from the UK Biobank. Mild kidney dysfunction was defined as eGFR between 60 and 89.9 mL/min/1.73 m2, and sarcopenia was defined according to the criteria of the 2019 European Working Group of Sarcopenia in Older People. Cox proportional hazard models with inverse probability weighting and competing risk models were used for analysis. RESULTS: During a median follow-up of 11.8 years, 20 146 participants died from all causes. Compared with participants with normal kidney function and without sarcopenia, those with mild kidney dysfunction or sarcopenia had significantly increased risks of all-cause mortality [hazard ratio (HR): 1.16, 95% confidence interval (CI): 1.12 to 1.19; HR: 1.29, 95% CI: 1.20 to 1.37]; those with both mild kidney dysfunction and sarcopenia had an even higher risk of all-cause mortality (HR: 1.61, 95% CI: 1.52 to 1.71), with a significant overall additive interaction (relative risk due to interaction 0.17, 95% CI: 0.05 to 0.29). Further subgroup analyses revealed that the associations of probable sarcopenia with all-cause and cause-specific mortality (non-accidental cause, non-communicable diseases, and cancer) were stronger among participants with mild kidney dysfunction than those with normal kidney function. CONCLUSIONS: The study indicates that sarcopenia and mild kidney dysfunction synergistically increase the risk of all-cause and cause-specific mortality. Early recognition and improvement of mild kidney function or sarcopenia in older people may reduce mortality risk but would require more prospective confirmation.

Fused diethynylbenzenes and phenanthrenes via arynes with alkynylsilanes.

A method for the hydroalkynylation and catalytic cyclization reactions of hexadehydro-Diels-Alder-derived benzynes is described. Diethynylbenzene derivatives are generated in a one-step reaction via trimethylsilyl-alkyne groups with benzyne formed by heating the appropriate tetrayne substrate. Trimethyl(phenylethynyl)silane loses TMS and binds to the electron-deficient site on HDDA-derived benzynes, and then phenanthrene was synthesized under mild reaction conditions by transition-metal-free, base promoted intramolecular cyclization.

The Antimicrobial Peptide Esculentin-1a(1-21)NH2 Stimulates Wound Healing by Promoting Angiogenesis through the PI3K/AKT Pathway.

Delayed wound healing is a persistent medical problem mainly caused by decreased angiogenesis. Esculentin-1a(1-21)NH2 [Esc-1a(1-21)NH2], has broad-spectrum antibacterial properties which comes from frog skins. It has shown promise as a treatment for wound healing. However, its effects on angiogenesis as well as the mechanism by which esc-1a(1-21)NH2 enhanced wound healing remained unclear. In this study, we analyzed the structural properties and biocompatibility of esc-1a(1-21)NH2 and evaluated its effect on wound closure using a full-thickness excision model in mice. Our results showed that esc-1a(1-21)NH2 significantly accelerated wound healing by increasing collagen deposition and angiogenesis, characterized by elevated expression levels of platelet, endothelial cell adhesion molecule-1 (CD31) and proliferating cell nuclear antigen (PCNA). Furthermore, the angiogenic activity of esc-1a(1-21)NH2 was confirmed in vitro by various assays. Esc-1a(1-21)NH2 significantly promoted cell migration and cell proliferation in human umbilical vein vascular endothelial cells (HUVECs) via activation of the phosphatidylinositol 3'-kinase (PI3K)/protein kinase B (AKT) pathway, and upregulated the expression of CD31 at both mRNA and protein levels. The effect of esc-1a(1-21)NH2 on angiogenesis was diminished by LY294002, a PI3K pathway inhibitor. Taken together, this study demonstrates that esc-1a(1-21)NH2 accelerates wound closure in mice by promoting angiogenesis via the PI3K/AKT signaling pathway, suggesting its effective application in the treatment of wound healing.

Nur77 Serves as a Potential Prognostic Biomarker That Correlates with Immune Infiltration and May Act as a Good Target for Prostate adenocarcinoma.

Prostate adenocarcinoma (PRAD) is the most frequent malignancy, and is the second leading cause of death due to cancer in men. Thus, new prognostic biomarkers and drug targets for PRAD are urgently needed. As we know, nuclear receptor Nur77 is important in cancer development and changes in the tumor microenvironment; whereas, the function of Nur77 in PRAD remains to be elucidated. The TCGA database was used to explore the Nur77 expression and its role in the prognosis of PRAD. It was shown that Nur77 was down regulated in PRAD, and low Nur77 expression was correlated with advanced clinical pathologic characteristics (high grade, histological type, age) and poor prognosis. Furthermore, key genes screening was examined by univariate Cox analysis and Kaplan-Meier survival. Additionally, Nur77 was closely related to immune infiltration and some anti-tumor immune functions. The differentially expressed genes (DEGs) were presented by protein-protein interaction (PPI) network analysis. Therefore, the expression level of Nur77 might help predict the survival of PRAD cases, which presents a new insight and a new target for the treatment of PRAD. In vitro experiments verified that natural product malayoside targeting Nur77 exhibited significant therapeutic effects on PRAD and largely induced cell apoptosis by up-regulating the expression of Nur77 and its mitochondrial localization. Taken together, Nur77 is a prognostic biomarker for patients with PRAD, which may refresh the profound understanding of PRAD individualized treatment.

Electrochemically Controlled Atom Transfer Radical Polymerization for Electrochemical Aptasensing of Tumor Biomarkers.

Tumor biomarkers are of great value in the liquid biopsy of malignant tumors. In this work, a simple and cost-friendly electrochemical aptasensor was presented for the highly sensitive and selective detection of glycoprotein tumor biomarkers. The DNA aptamer-modified electrode was used as the sensing interface to specifically capture the target glycoprotein tumor biomarkers, to which the alkyl halide initiators for atom transfer radical polymerization (ATRP) were then attached via the esterification crosslinking between the boronic acid group and the cis-dihydroxyl sites of the conjugated oligosaccharide chains on glycoprotein tumor biomarkers followed by the growth of long-chain polymers through electrochemically controlled ATRP (eATRP) to efficiently recruit the ferrocene detection tags. As there are tens to hundreds of cis-dihydroxyl sites on a glycoprotein tumor biomarker for attaching ATRP initiators while each long-chain polymer can recruit hundreds to thousands of ferrocene detection tags, a significantly high current signal can be generated even in the presence of ultralow-abundance targets. Hence, the eATRP-based electrochemical aptasensor is capable of sensitively and selectively detecting glycoprotein tumor biomarkers. Using alpha-fetoprotein as the model target, the limit of detection was demonstrated to be 0.32 pg/mL. Moreover, the aptasensor has been successfully applied to detect glycoprotein tumor biomarkers in human serum samples. In view of its high sensitivity and selectivity, simple operation, and cost-friendliness, the eATRP-based electrochemical aptasensor shows great promise in the glycoprotein-based liquid biopsy of malignant tumors, even at the early stage of development.

Boronate-Affinity Cross-Linking-Based Ratiometric Electrochemical Detection of Glycoconjugates.

Despite the widespread application of the boronate-affinity cross-linking (BAC) in the separation, enrichment, and sensing of glycoconjugates, it remains a huge challenge to integrate the BAC into the selective electrochemical detection of glycoconjugates due to the poor selectivity of the BAC. Herein, we demonstrate a BAC-based ratiometric electrochemical method for the simple, low-cost, and highly sensitive and selective detection of glycoconjugates. Briefly, the methylene blue (MB)-tagged nucleic acid aptamer is exploited as the recognition element to selectively capture target glycoconjugate, to which a large number of ferrocene (Fc) tags are subsequently labeled via the BAC between the phenylboronic acid (PBA) group and the cis-diol site of the oligosaccharide chains on the captured targets. Using the MB tag as the internal reference and the Fc tag as the reporter of the target capture, the dual-signal output enables the ratiometric detection. Due to the presence of a high density of the cis-diol sites on a glycoconjugate, sufficiently high sensitivity can be obtained even without using any amplification strategies. Using glycoprotein mucin 1 (MUC1) as the model target, the signal ratio (IFc/IMB) exhibits good linearity over the range from 0.05 to 50 U/mL, with a detection limit of 0.021 U/mL. In addition to the high sensitivity and selectivity, the results of the analysis of MUC1 in serum samples are acceptable. By virtue of its simplicity, cost-effectiveness, and high robustness and reproducibility, this BAC-based ratiometric electrochemical method holds great promise in the highly sensitive and selective detection of glycoconjugates.

Biologically Mediated RAFT Polymerization for Electrochemical Sensing of Kinase Activity.

The assay of kinase activity with ultrahigh sensitivity is important to medical diagnostics and drug discovery. Herein, we report the biologically mediated RAFT polymerization (BMRP) and its potential use as an efficient amplification strategy in the ultrasensitive electrochemical sensing of kinase activity. In BMRP, the reversible addition-fragmentation chain-transfer (RAFT) process is initiated and sustained by the reduced form of coenzyme I (i.e., NADH), which can efficiently mediate the direct fragmentation of thiocarbonylthio (TCT) compounds (or the TCT-capped dormant chains) to produce an initiating/propagating radical under mild conditions. Due to the absence of exogenous radicals, the notorious radical termination in RAFT equilibrium can be greatly suppressed. For the sensing of kinase activity, the recognition peptides, without carboxyl groups, are immobilized via the Au-S self-assembly. After phosphorylation, TCT compounds (as RAFT agents) are tethered to the enzymatically generated phosphate groups via the carboxylate-Zr(IV)-phosphate (CZP) linkage. Subsequently, the BMRP of ferrocenylmethyl methacrylate (FcMMA) results in the labeling of each phosphate group with hundreds to thousands of Fc tags, thereby greatly amplifying the sensing signal. Obviously, the BMRP-based strategy is biologically friendly, highly efficient, uncomplicated, and quite low-cost. The detection limit of 1.85 mU/mL has been achieved toward the selective sensing of the cAMP-dependent protein kinase (PKA). Moreover, the proposed kinase sensor is applicable to inhibitor screening and kinase activity sensing in serum samples. By virtue of its low cost, high sensitivity and selectivity, and uncomplicated operation, the proposed kinase sensor holds great potential in medical diagnostics and drug discovery.

Boronate Affinity-Amplified Electrochemical Aptasensing of Lipopolysaccharide.

As lipopolysaccharide (LPS) is closely associated with sepsis and other life-threatening conditions, the point-of-care (POC) detection of LPS is of significant importance to human health. In this work, we illustrate an electrochemical aptasensor for the POC detection of low-abundance LPS by utilizing boronate affinity (BA) as a simple, efficient, and cost-effective amplification strategy. Briefly, the BA-amplified electrochemical aptasensing of LPS involves the tethering of the aptamer receptors and the BA-mediated direct decoration of LPS with redox signal tags. As the polysaccharide chain of LPS contains hundreds of cis-diol sites, the covalent crosslinking between the phenylboronic acid group and cis-diol sites can be harnessed for the site-specific decoration of each LPS with hundreds of redox signal tags, thereby enabling amplified detection. As it involves only a single-step operation (∼15 min), the BA-mediated signal amplification holds the significant advantages of unrivaled simplicity, rapidness, and cost-effectiveness over the conventional nanomaterial- and enzyme-based strategies. The BA-amplified electrochemical aptasensor has been successfully applied to specifically detect LPS within 45 min, with a detection limit of 0.34 pg/mL. Moreover, the clinical utility has been validated based on LPS detection in complex serum samples. As a proof of concept, a portable device has been developed to showcase the potential applicability of the BA-amplified electrochemical LPS aptasensor in the POC testing. In view of its simplicity, rapidness, and cost-effectiveness, the BA-amplified electrochemical LPS aptasensor holds broad application prospects in the POC testing.

Electrochemical Detection of Femtomolar DNA via Boronate Affinity-Mediated Decoration of Polysaccharides with Electroactive Tags.

In view of their high efficiency and cost-effectiveness, polymers are of great promise as carriers for signal tags in amplified detection. Herein, we present a polysaccharide-amplified method for the electrochemical detection of a BRCA1 breast cancer gene-derived DNA target at the femtomolar levels. Briefly, peptide nucleic acid (PNA) with a complementary sequence was tethered as the capture probe for the DNA target, to which carboxyl group-containing polysaccharides were then attached via facile phosphate-Zr(IV)-carboxylate crosslinking, followed by the decoration of polysaccharide chains with electroactive ferrocene (Fc) signal tags via affinity coupling between a cis-diol site and phenylboronic acid (PBA) group. As the polysaccharide chain contains hundreds of cis-diol sites, boronate affinity can enable the site-specific decoration of each polysaccharide chain with hundreds of Fc signal tags, efficiently transducing each target capture event into the decoration of many Fc signal tags. As polysaccharides are cheap, renewable, ubiquitous, and biodegradable natural biopolymers, the use of polysaccharides for signal amplification offers the benefits of high efficiency, cost-effectiveness, excellent biocompatibility, and environmental friendliness. The linear range of the polysaccharide-amplified method for DNA detection was demonstrated to be from 10 fM to 10 nM (R2 = 0.996), with the detection limit as low as 2.9 fM. The results show that this method can also discriminate single base mismatch with satisfactory selectivity and can be applied to DNA detection in serum samples. In view of these merits, the polysaccharide-amplified PNA-based electrochemical method holds great promise in DNA detection with satisfactory sensitivity and selectivity.