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Abstract Aberrant signaling pathway activity is a hallmark of tumorigenesis and progression, which has guided targeted inhibitor design for over 30 years. Yet, adaptive resistance mechanisms, induced by rapid, context-specific signaling network rewiring, continue to challenge therapeutic efficacy. Leveraging progress in proteomic technologies and network-based methodologies, we introduce Virtual Enrichment-based Signaling Protein-activity Analysis (VESPA)—an algorithm designed to elucidate mechanisms of cell response and adaptation to drug perturbations—and use it to analyze 7-point phosphoproteomic time series from colorectal cancer cells treated with clinically-relevant inhibitors and control media. Interrogating tumor-specific enzyme/substrate interactions accurately infers kinase and phosphatase activity, based on their substrate phosphorylation state, effectively accounting for signal crosstalk and sparse phosphoproteome coverage. The analysis elucidates time-dependent signaling pathway response to each drug perturbation and, more importantly, cell adaptive response and rewiring, experimentally confirmed by CRISPR knock-out assays, suggesting broad applicability to cancer and other diseases.

Abstract Induced oncoproteins degradation provides an attractive anti-cancer modality. Activation of anaphase-promoting complex (APC/CCDH1) prevents cell-cycle entry by targeting crucial mitotic proteins for degradation. Phosphorylation of its co-activator CDH1 modulates the E3 ligase activity, but little is known about its regulation after phosphorylation and how to effectively harness APC/CCDH1 activity to treat cancer. Peptidyl-prolyl cis-trans isomerase NIMA-interacting 1 (PIN1)-catalyzed phosphorylation-dependent cis-trans prolyl isomerization drives tumor malignancy. However, the mechanisms controlling its protein turnover remain elusive. Through proteomic screens and structural characterizations, we identify a reciprocal antagonism of PIN1-APC/CCDH1 mediated by domain-oriented phosphorylation-dependent dual interactions as a fundamental mechanism governing mitotic protein stability and cell-cycle entry. Remarkably, combined PIN1 and cyclin-dependent protein kinases (CDKs) inhibition creates a positive feedback loop of PIN1 inhibition and APC/CCDH1 activation to irreversibly degrade PIN1 and other crucial mitotic proteins, which force permanent cell-cycle exit and trigger anti-tumor immunity, translating into synergistic efficacy against triple-negative breast cancer.

Abstract Background Stem cell transplantation shows great potential to improve the long-term survival of cirrhosis patients. However, therapeutic effects may not be homogeneous across the whole study population. This study constructed an easy-to-use nomogram to improve prognostic prediction and aid in treatment decision making for cirrhotic patients. Methods From August 2005 to April 2019, 315 patients with decompensated cirrhosis receiving autologous peripheral blood stem cell (PBSC) transplantation were enrolled in this study. They were randomly classified into training (2/3) and validation (1/3) groups. A predictive model was developed using Cox proportional hazard models and subsequently validated. The predictive performance of the model was evaluated and also compared with other prognostic models. Results Age, creatinine, neutrophil-to-lymphocyte ratio, and Child–Turcotte–Pugh class were included in the nomogram as prognostic variables. The nomogram showed high discrimination power concerning the area under receiver operating characteristic curves (3/5-year AUC: 0.742/0.698) and good consistency suggested by calibration plots. Patients could be accurately stratified into poor- and good-outcome groups regarding liver-transplantation free survival after receiving PBSC therapy (P

For patients with chronic spinal cord injury, the conventional treatment is rehabilitation and treatment of spinal cord injury complications such as urinary tract infection, pressure sores, osteoporosis, and deep vein thrombosis. Surgery is rarely performed on spinal cord injury in the chronic phase, and few treatments have been proven effective in chronic spinal cord injury patients. Development of effective therapies for chronic spinal cord injury patients is needed. We conducted a randomized controlled clinical trial in patients with chronic complete thoracic spinal cord injury to compare intensive rehabilitation (weight-bearing walking training) alone with surgical intervention plus intensive rehabilitation. This clinical trial was registered at ClinicalTrials.gov (NCT02663310). The goal of surgical intervention was spinal cord detethering, restoration of cerebrospinal fluid flow, and elimination of residual spinal cord compression. We found that surgical intervention plus weight-bearing walking training was associated with a higher incidence of American Spinal Injury Association Impairment Scale improvement, reduced spasticity, and more rapid bowel and bladder functional recovery than weight-bearing walking training alone. Overall, the surgical procedures and intensive rehabilitation were safe. American Spinal Injury Association Impairment Scale improvement was more common in T7–T11 injuries than in T2–T6 injuries. Surgery combined with rehabilitation appears to have a role in treatment of chronic spinal cord injury patients.

Abstract CD8+ T cells play an important role in anti-tumor immunity. Better understanding of their regulation could advance cancer immunotherapies. Here we identify, via stepwise CRISPR-based screening, that CUL5 is a negative regulator of the core signaling pathways of CD8+ T cells. Knocking out CUL5 in mouse CD8+ T cells significantly improves their tumor growth inhibiting ability, with significant proteomic alterations that broadly enhance TCR and cytokine signaling and their effector functions. Chemical inhibition of neddylation required by CUL5 activation, also enhances CD8 effector activities with CUL5 validated as a major target. Mechanistically, CUL5, which is upregulated by TCR stimulation, interacts with the SOCS-box-containing protein PCMTD2 and inhibits TCR and IL2 signaling. Additionally, CTLA4 is markedly upregulated by CUL5 knockout, and its inactivation further enhances the anti-tumor effect of CUL5 KO. These results together reveal a negative regulatory mechanism for CD8+ T cells and have strong translational implications in cancer immunotherapy.

Summary: Neutrophils are important innate immune cells with plasticity, heterogenicity, and functional ambivalency. While bone marrow is often regarded as the primary source of neutrophil production, the roles of extramedullary production in regulating neutrophil plasticity and heterogenicity in autoimmune diseases remain poorly understood. Here, we report that the lack of wingless-type MMTV integration site family member 5 (WNT5) unleashes anti-inflammatory protection against colitis in mice, accompanied by reduced colonic CD8+ T cell activation and enhanced splenic extramedullary myelopoiesis. In addition, colitis upregulates WNT5 expression in splenic stromal cells. The ablation of WNT5 leads to increased splenic production of hematopoietic niche factors, as well as elevated numbers of splenic neutrophils with heightened CD8+ T cell suppressive capability, in part due to elevated CD101 expression and attenuated pro-inflammatory activities. Thus, our study reveals a mechanism by which neutrophil plasticity and heterogenicity are regulated in colitis through WNT5 and highlights the role of splenic neutrophil production in shaping inflammatory outcomes.

Concrete slump, a crucial indicator of fluidity, directly affects the pumpability and construction efficiency of concrete. Conventionally, concrete slumps are assessed through multiple test iterations conducted by skilled professionals to obtain accurate results. In this study, a predictive model was established to predict concrete slumps directly based on concrete mix proportions, thereby mitigating labor and time costs. An open-source dataset was used in this study. The selected water-cement ratio ranged from 0.29 to 0.66. Cement, slag, fly ash, water, superplasticizer, coarse aggregate, and fine aggregate were employed as input parameters. Furthermore, the performances of a variety of algorithms in predicting concrete slumps were analyzed and compared. The algorithms included SVM, KNN, Extra-Trees, Gradient Boosting, Decision Tree, Elastic Net, Lasso, Ridge, Random Forest, Bagging, AdaBoost, and XGBoost. Through a comprehensive comparison of the prediction performance of these models, AdaBoost, Bagging, Extra-Trees, and Random Forest were adopted as base models. Moreover, the SVM algorithm optimized using Differential Evolution was employed as a secondary model to construct an enhanced integrated prediction model. The final prediction model boasts an MSE of 2.099984, MAE of 1.225597, RMSE of 1.449132, and R2 of 0.970418. Compared to conventional prediction models, the proposed model has the potential to substantially improve prediction performance.

BackgroundSarcopenia adversely affects the treatment outcomes in Cirrhosis and NAFLD. However, such research is limited in primary biliary cholangitis (PBC) patients. This study was performed to examine the prevalence of sarcopenia and its impact on PBC patients’ prognoses.MethodsThis study enrolled confirmed PBC patients who had an abdominal CT scan. Sarcopenia was determined by the L3-skeletal muscle index with a Chinese population-based cut-off value. Laboratory test values and liver stiffness measurements values were obtained from the electronic medical records.ResultsIn total, 174 PBC patients with a median age of 54 (IQR, 48, 62) years old, were enrolled. 45 (25.9%) patients among them were diagnosed with sarcopenia. Univariate and multivariate logistic regression results illustrated that male gender (OR = 9.152, 95%CI = 3.131–26.751, p

This paper studies the existence of solutions for fractional dynamical systems with two damping terms in Banach space. First, we generalize the well-known Gronwall inequality. Next, according to fixed-point theorems and inequalities, the existence results for the considered system are obtained. At last, an example is used to support the main results.

Abstract The serine/threonine kinase AKT is a central node in cell signaling. While aberrant AKT activation underlies the development of a variety of human diseases, how different patterns of AKT-dependent phosphorylation dictate downstream signaling and phenotypic outcomes remains largely enigmatic. Herein, we perform a systems-level analysis that integrates methodological advances in optogenetics, mass spectrometry-based phosphoproteomics, and bioinformatics to elucidate how different intensity, duration, and pattern of Akt1 stimulation lead to distinct temporal phosphorylation profiles in vascular endothelial cells. Through the analysis of ~35,000 phosphorylation sites across multiple conditions precisely controlled by light stimulation, we identify a series of signaling circuits activated downstream of Akt1 and interrogate how Akt1 signaling integrates with growth factor signaling in endothelial cells. Furthermore, our results categorize kinase substrates that are preferably activated by oscillating, transient, and sustained Akt1 signals. We validate a list of phosphorylation sites that covaried with Akt1 phosphorylation across experimental conditions as potential Akt1 substrates. Our resulting dataset provides a rich resource for future studies on AKT signaling and dynamics.

Calponin 2 (CNN2) is a prominent actin stabilizer. It regulates fatty acid oxidation (FAO) by interacting with estrogen receptor 2 (ESR2) to determine kidney fibrosis. However, whether CNN2 is actively involved in acute kidney injury (AKI) remains unclear. Here, we report that CNN2 was induced in human and animal kidneys after AKI. Knockdown of CNN2 preserved kidney function, mitigated tubular cell death and inflammation, and promoted cell proliferation. Distinct from kidney fibrosis, proteomics showed that the key elements in the FAO pathway had few changes during AKI, but we identified that 3-hydroxymethylglutaryl-CoA synthase 2 (Hmgcs2), a rate-limiting enzyme of endogenous ketogenesis that promotes cell self-renewal, was markedly increased in CNN2-knockdown kidneys. The production of ketone body β-hydroxybutyrate and ATP was increased in CNN2-knockdown mice. Mechanistically, CNN2 interacted with ESR2 to negatively regulate the activities of mitochondrial sirtuin 5. Activated sirtuin 5 subsequently desuccinylated Hmgcs2 to produce energy for mitigating AKI. Understanding CNN2-mediated discrete fine-tuning of protein posttranslational modification is critical to optimize organ performance after AKI.

Summary: Here, we present a standardized, “off-the-shelf” proteomics pipeline working in a single 96-well plate to achieve deep coverage of cellular proteomes with high throughput and scalability. This integrated pipeline streamlines a fully automated sample preparation platform, a data-independent acquisition (DIA) coupled with high-field asymmetric waveform ion mobility spectrometer (FAIMS) interface, and an optimized library-free DIA database search strategy. Our systematic evaluation of FAIMS-DIA showing single compensation voltage (CV) at −35 V not only yields the deepest proteome coverage but also best correlates with DIA without FAIMS. Our in-depth comparison of direct-DIA database search engines shows that Spectronaut outperforms others, providing the highest quantifiable proteins. Next, we apply three common DIA strategies in characterizing human induced pluripotent stem cell (iPSC)-derived neurons and show single-shot mass spectrometry (MS) using single-CV (−35 V)-FAIMS-DIA results in >9,000 quantifiable proteins with

Rarely, penetrating injuries to the spinal cord result from wooden objects, creating unique challenges to mitigate neurological injury and high rates of infection and foreign body reactions. We report a man who sustained a penetrating cervical spinal cord injury from a sharpened stick. While initially tetraparetic, he rapidly recovered function. The risks of neurological deterioration during surgical removal made the patient reluctant to consent to surgery despite the impalement of the spinal cord. A repeat MRI on day 3 showed an extension of edema indicating progressive inflammation. On the 7th day after injury, fever and paresthesias occurred with a large increase in serum inflammatory indicators, and the patient agreed to undergo surgical removal of the wooden object. We discuss the management nuances related to wood, the longitudinal evolution of MRI findings, infection risk, surgical risk mitigation and technique, an inflammatory marker profile, long-term recovery, and the surprisingly minimal neurological deficits associated with low-velocity midline spinal cord injuries. The patient had an excellent clinical outcome. The main lessons are that a wooden penetrating central nervous system injury has a high risk for infection, and that surgical removal from the spinal cord should be performed soon after injury and under direct visualization.

In this paper, we mainly study the following boundary value problems of fractional discontinuous differential equations with impulses: $ \hskip 3mm \left\{ \begin{array}{lll} _{t}^{C} \mathcal {D}^{\mathfrak{R}}_{0^{+}}\Lambda(t) = \mathcal {E}(t)\digamma(t, \Lambda(t)), \ a.e.\ t\in Q, \\ \triangle \Lambda|_{t = t_{{\kappa}}} = \Phi_{{\kappa}}(\Lambda(t_{{\kappa}})), \ {\kappa} = 1, \ 2, \ \cdots, \ m, \\ \triangle \Lambda'|_{t = t_{{\kappa}}} = 0, \ {\kappa} = 1, \ 2, \ \cdots, \ m, \\ {\vartheta} \Lambda(0)-{\chi} \Lambda(1) = \int_{0}^{1}\varrho_{1}({\upsilon})\Lambda({\upsilon})d{\upsilon}, \\ {\zeta} \Lambda'(0)-\delta \Lambda'(1) = \int_{0}^{1}\varrho_{2}({\upsilon})\Lambda({\upsilon})d{\upsilon}, \end{array}\right. $ where $ {\vartheta} > {\chi} > 0, \ {\zeta} > \delta > 0 $, $ \Phi_{{\kappa}}\in C(\mbox{ $\mathbb{R}$ }^{+}, \mbox{ $\mathbb{R}$ }^{+}) $, $ \mathcal {E}, \ \varrho_{1}, \ \varrho_{2} \geq 0 $ a.e. on $ Q = [0, 1] $, $ \mathcal {E}, \ \varrho_{1}, \ \varrho_{2} \in L^{1}(0, 1) $ and $ \digamma:[0, 1]\times \mbox{ $\mathbb{R}$ }^{+}\rightarrow \mbox{ $\mathbb{R}$ }^{+} $, $ \mbox{ $\mathbb{R}$ }^{+} = [0, +\infty) $. By using Krasnosel skii's fixed point theorem for discontinuous operators on cones, some sufficient conditions for the existence of single or multiple positive solutions for the above discontinuous differential system are established. An example is given to confirm the main results in the end.

This paper investigates a new class of non-autonomous second-order measure evolution systems involving state-dependent delay and non-instantaneous impulses. We introduce a stronger concept of exact controllability called total controllability. The existence of mild solutions and controllability for the considered system are obtained by applying strongly continuous cosine family and the Mönch fixed point theorem. Finally, an example is used to verify the practical application of the conclusion.

Abstract Fenofibrate (FF) has shown potential benefits in patients with primary biliary cholangitis (PBC) who have an incomplete response to ursodeoxycholic acid (UDCA). However, the efficacy and safety of FF in patients with cirrhosis remain unclear. To evaluate the efficacy and safety of additional FF therapy in patients with PBC‐related cirrhosis with an incomplete response to UDCA, we conducted a retrospective analysis comparing the clinical results of additional FF therapy and continued UDCA monotherapy. A total of 59 patients were included; 27 cases underwent UDCA monotherapy and 32 cases underwent UDCA combined with FF therapy. A significant difference in alkaline phosphatase (ALP) normalization was achieved in the FF group compared to the UDCA group (37% vs. 11%, respectively; p = 0.020). Additional FF therapy was an independent risk factor for ALP normalization (hazard ratio, 7.679; 95% confidence interval, 2.059–28.633; p = 0.003). Hepatic deterioration was experienced by 40% versus 48% (p = 0.562) while 11% vs. 37% (p = 0.111) experienced liver‐related mortality or liver transplantation in the FF and UDCA groups, respectively. Compared to UDCA monotherapy, additional FF therapy was associated with lower United Kingdom (UK)‐PBC risk score and surrogate serum indices of liver fibrosis. After 12 months of add‐on FF therapy, median ALP level and UK‐PBC risk score decreased 35% and 52% from baseline (p = 0.001 and 0.210, respectively). Serum aminotransferase, triglyceride, and cholesterol decreased progressively, while total bilirubin, serum creatinine, blood urea, estimated glomerular filtration rate, aspartate aminotransferase‐to‐platelet ratio index, and fibrosis‐4 index remained stable in FF‐treated cirrhotic cases during follow‐up. No significant adverse effects associated with additional FF therapy were observed in our cohort. Conclusion: Additional FF therapy was associated with higher ALP normalization rates and lower UK‐PBC risk scores in patients with cirrhotic PBC with an incomplete response to UDCA. In addition, FF therapy seemed safe and well tolerated with a low frequency of adverse effects in patients with cirrhosis.

This paper aims to introduce robot technology to carry out the safety inspection of transmission towers in long-distance power transmission, so as to improve the safety and efficiency of inspection. However, aiming at the problem that the existing climbing robots are mainly used for large load applications, which leads to the large size and lack of flexibility of the robot, we propose an innovative solution. Firstly, a lightweight quadruped climbing robot is designed to improve portability and operational flexibility. Then, a one-dimensional force sensor is added at the end of each leg of the robot, and a special swing phase trajectory is designed. The robot can judge whether the electromagnetic adsorption is effective and avoid potential safety hazards. Finally, based on the principle of virtual model control (VMC), a foot-end force balancing algorithm is proposed to achieve uniform distribution and continuous change in force, and improve safety and load capacity. The experiments show that the scheme has a stable climbing ability in the environments of angle steel, vertical ferromagnetic plane and transmission tower.

Abstract Background Despite emerging evidence on the therapeutic potential of mesenchymal stem cells (MSCs) for liver fibrosis, the underlying mechanisms remain unclear. At present, MSC-derived exosomes (MSC-EXOs) are widely accepted as crucial messengers for intercellular communication. This study aimed to explore the therapeutic effects of MSC-EXOs on liver fibrosis and identify the mechanisms underlying the action of MSC-EXOs. Methods Carbon tetrachloride was used to induce a liver fibrosis model, which was intravenously administered with MSCs or MSC-EXOs to assess treatment efficacy. The resulting histopathology, fibrosis degree, inflammation and macrophage polarization were analyzed. RAW264.7 and BMDM cells were used to explore the regulatory effects of MSC-EXOs on macrophage polarization. Then, the critical miRNA mediating the therapeutic effects of MSC-EXOs was screened via RNA sequencing and validated experimentally. Furthermore, the target mRNA and downstream signaling pathways were elucidated by luciferase reporter assay, bioinformatics analysis and western blot. Results MSCs alleviated liver fibrosis largely depended on their secreted exosomes, which were visualized to circulate into liver after transplantation. In addition, MSC-EXOs were found to modulate macrophage phenotype to regulate inflammatory microenvironment in liver and repair the injury. Mechanically, RNA-sequencing illustrates that miR-148a, enriched in the MSC-EXOs, targets Kruppel-like factor 6 (KLF6) to suppress pro-inflammatory macrophages and promote anti-inflammatory macrophages by inhibiting the STAT3 pathway. Administration of miR-148a-enriched MSC-EXOs or miR-148a agomir shows potent ameliorative effects on liver fibrosis. Conclusions These findings suggest that MSC-EXOs protect against liver fibrosis via delivering miR-148a that regulates intrahepatic macrophage functions through KLF6/STAT3 signaling and provide a potential therapeutic target for liver fibrosis.

Abstract Human peripheral blood mononuclear cells (PBMCs) originate from hematopoietic stem cells in the bone marrow, which mainly includes lymphocytes (T cells, B cells, and natural killer cells) and monocytes. Cryopreserved PBMCs providing biobank resources are crucial for clinical application or scientific research. Here, we used flow cytometry to explore the influence of long-term cryopreservation on the quality of PBMCs with the aim of providing important evidence for the effective utilization of biobank resources. The PBMCs were isolated from the peripheral blood, which was collected from volunteers in the hospital. After long-term cryopreservation in liquid nitrogen, we analyzed the changes in cell numbers, viability, and multiple subtypes of PBMCs and studied the apoptosis, proliferation, activation, function, and status of T cells in comparison with freshly isolated PBMCs by flow cytometry, and then further tracked the effects of long-term cryopreservation on the same sample. Although the different cell types in the PBMCs dynamically changed compared with those in the freshly isolated samples, PBMC recovery and viability remained stable after long-term cryopreservation, and the number of most innate immune cells (e.g., monocytes and B cells) was significantly reduced compared to that of the freshly isolated PBMCs or long-term cryopreserved PBMCs; more importantly, the proportion of T cell subtypes, apoptosis, proliferation, and functional T cells, except for Tregs, were not affected by long-term cryopreservation. However, the proportions of activated T, naïve T, central memory T, effector T, and effector memory T cells dynamically changed after long-term cryopreservation. This article provides important evidence for the effective utilization of biobank resources. Long-term cryopreserved PBMCs can be partly used as biological resources for clinical research or basic studies, but the effect of cryopreservation on PBMCs should be considered when selecting cell samples, especially in research relating to activating or inhibiting function.

The diagnosis of primary biliary cholangitis (PBC) in patients with seropositive anti‐mitochondrial antibody (AMA) but normal alkaline phosphatase (ALP) depends on a liver biopsy. We aimed to reveal potential serum biomarkers that could suggest the necessity of a liver biopsy in such patients. Retrospective analysis was performed. Subjects who were treatment naive with seropositive AMA but normal ALP and who underwent at least one liver biopsy between 2008 and 2020 were included in this study. Histologic biopsies were evaluated by two experienced pathologists blinded to the serum tests. A total of 115 patients who were treatment naive were included in this study. Of these, 77 patients (67%) exhibited histologic PBC features and nonspecific histologic features were found in the remaining 38 (33%) patients. Multivariate analysis suggested that baseline serum immunoglobulin M (IgM) >0.773 × upper limit of normal (ULN) (P 42 years (P = 0.002) were associated with the diagnosis of PBC through liver biopsies. A significant decrease in the median levels of gamma‐glutamyl transpeptidase (GGT) and IgM was found in 54 patients with PBC who received ursodeoxycholic acid (UDCA). Conclusion: For patients who were treatment naive with seropositive AMA but normal ALP, baseline serum IgM >0.773 × ULN and age >42 years were the factors that strongly suggested a diagnosis of PBC. In these patients receiving UDCA, a dynamic monitoring of GGT and IgM might be helpful in evaluating therapeutic responses.

Abstract Background The biguanide drug metformin is a safe and widely prescribed drug for type 2 diabetes. Interestingly, hundreds of clinical trials have been set to evaluate the potential role of metformin in the prevention and treatment of cancer including colorectal cancer (CRC). However, the “metformin signaling” remains controversial. Aims and Methods To interrogate cell signaling induced by metformin in CRC and explore the druggability of the metformin‐rewired phosphorylation network, we performed integrative analysis of phosphoproteomics, bioinformatics, and cell proliferation assays on a panel of 12 molecularly heterogeneous CRC cell lines. Using the high‐resolute data‐independent analysis mass spectrometry (DIA‐MS), we monitored a total of 10,142 proteins and 56,080 phosphosites (P‐sites) in CRC cells upon a short‐ and a long‐term metformin treatment. Results and Conclusions We found that metformin tended to primarily remodel cell signaling in the long‐term and only minimally regulated the total proteome expression levels. Strikingly, the phosphorylation signaling response to metformin was highly heterogeneous in the CRC panel, based on a network analysis inferring kinase/phosphatase activities and cell signaling reconstruction. A “MetScore” was determined to assign the metformin relevance of each P‐site, revealing new and robust phosphorylation nodes and pathways in metformin signaling. Finally, we leveraged the metformin P‐site signature to identify pharmacodynamic interactions and confirmed a number of candidate metformin‐interacting drugs, including navitoclax, a BCL‐2/BCL‐xL inhibitor. Together, we provide a comprehensive phosphoproteomic resource to explore the metformin‐induced cell signaling for potential cancer therapeutics. This resource can be accessed at https://yslproteomics.shinyapps.io/Metformin/.

Immune checkpoint genes (ICGs) have recently been proven to perform instrumental functions in the maintenance of immune homeostasis and represent a promising therapeutic strategy; however, their expression patterns and prognostic values are not fully elucidated in hepatocellular carcinoma (HCC). In this investigation, we focused on establishing and validating a prognostic gene signature to facilitate decision-making in clinical practice. Clinical information, as well as transcriptome data, was obtained from the Cancer Genome Atlas (TCGA) and International Cancer Genome Consortium (ICGC) database. Univariate Cox regression and least absolute shrinkage and selection operator (LASSO) Cox method were employed to build a multi-gene signature in the TCGA database, while the ICGC database was used for validation. Subsequently, utilizing the six-gene signature, we were able to categorize patients into high- and low-risk groups. In two cohorts, survival analysis findings revealed a dismal outlook for the high-risk group. The receiver operating characteristic curves were utilized to estimate the gene signature's prediction ability. Moreover, correlation analysis showed high-risk group was linked to advanced pathological stage, infiltration of immune cells and therapeutic response. In summary, this unique gene profile might serve not only as a useful prognostic indicator but also as a marker of therapy responsiveness in HCC.

Abstract Background Hepatocellular carcinoma (HCC) is one of the most common malignancies worldwide. Costimulatory molecules have been proven to be the foundation of immunotherapy. However, the potential roles of costimulatory molecule genes (CMGs) in HCC remain unclear. Our study is aimed to develop a costimulatory molecule-related gene signature that could evaluate the prognosis of HCC patients. Methods Based on The Cancer Gene Atlas (TCGA) database, univariate Cox regression analysis was applied in CMGs to identify prognosis-related CMGs. Consensus clustering analysis was performed to stratify HCC patients into different subtypes and compared them in OS. Subsequently, the LASSO Cox regression analysis was performed to construct the CMGs-related prognostic signature and Kaplan–Meier survival curves as well as ROC curve were used to validate the predictive capability. Then we explored the correlations of the risk signature with tumor-infiltrating immune cells, tumor mutation burden (TMB) and response to immunotherapy. The expression levels of prognosis-related CMGs were validated based on qRT-PCR and Human Protein Atlas (HPA) databases. Results All HCC patients were classified into two clusters based on 11 CMGs with prognosis values and cluster 2 correlated with a poorer prognosis. Next, a prognostic signature of six CMGs was constructed, which was an independent risk factor for HCC patients. Patients with low-risk score were associated with better prognosis. The correlation analysis showed that the risk signature could predict the infiltration of immune cells and immune status of the immune microenvironment in HCC. The qRT-PCR and immunohistochemical results indicated six CMGs with differential expression in HCC tissues and normal tissues. Conclusion In conclusion, our CMGs-related risk signature could be used as a prediction tool in survival assessment and immunotherapy for HCC patients.

This paper is mainly concerned with the existence of mild solutions and exact controllability for a class of fractional semilinear system of order q∈1,2 with instantaneous and noninstantaneous impulses. First, combining the Kuratowski measure of noncompactness and the Mönch fixed point theorem, we investigated the existence result for the considered system. It is remarkable that our assumptions for impulses and the nonlinear term are weaker than the Lipschitz conditions. Next, on this basis, the exact controllability for the considered system is determined. In the end, an example is provided to support the main findings.

Abstract The aim of this paper is to establish some results about the existence of multiple solutions for the following singular semipositone boundary value problem of fourth-order differential systems with parameters: { u ( 4 ) ( t ) + β 1 u ″ ( t ) − α 1 u ( t ) = f 1 ( t , u ( t ) , v ( t ) ) , 0 < t < 1 ; v ( 4 ) ( t ) + β 2 v ″ ( t ) − α 2 v ( t ) = f 2 ( t , u ( t ) , v ( t ) ) , 0 < t < 1 ; u ( 0 ) = u ( 1 ) = u ″ ( 0 ) = u ″ ( 1 ) = 0 ; v ( 0 ) = v ( 1 ) = v ″ ( 0 ) = v ″ ( 1 ) = 0 , $$ \textstyle\begin{cases} u^{(4)}(t)+\beta _{1}u''(t)-\alpha _{1}u(t)=f_{1}(t,u(t),v(t)),\quad 0< t< 1; \\ v^{(4)}(t)+\beta _{2}v''(t)-\alpha _{2}v(t)=f_{2}(t,u(t),v(t)),\quad 0< t< 1; \\ u(0)=u(1)=u''(0)=u''(1)=0; \\ v(0)=v(1)=v''(0)=v''(1)=0, \end{cases} $$ where f 1 , f 2 ∈ C [ ( 0 , 1 ) × R 0 + × R , R ] $f_{1},f_{2}\in C[(0,1)\times \mathbb{R}^{+}_{0}\times \mathbb{R}, \mathbb{R}]$ , R 0 + = ( 0 , + ∞ ) $\mathbb{R}_{0}^{+}=(0,+\infty )$ . By constructing a special cone and applying fixed point index theory, some new existence results of multiple solutions for the considered system are obtained under some suitable assumptions. Finally, an example is worked out to illustrate the main results.

This paper is mainly concerned with the existence of multiple solutions for the following boundary value problems of fractional differential equations with generalized Caputo derivatives: $ \hskip 3mm \left\{ \begin{array}{lll} ^{C}_{0}D^{\alpha}_{g}x(t)+f(t, x) = 0, \ 0 0 $, $ f\in C([0, 1]\times \mathbb{R}^{+}, \mathbb{R}^{+}) $, $ g' > 0 $, $ h\in C([0, 1], \mathbb{R}^{+}) $, $ \mathbb{R}^{+} = [0, +\infty) $. Applying the fixed point theorem on cone, the existence of multiple solutions for considered system is obtained. The results generalize and improve existing conclusions. Meanwhile, the Ulam stability for considered system is also considered. Finally, three examples are worked out to illustrate the main results.

Metastatic breast cancer remains a major cause of cancer-related deaths in women, and there are few effective therapies against this advanced disease. Emerging evidence suggests that key steps of tumor progression and metastasis are controlled by reversible epigenetic mechanisms. Using an in vivo genetic screen, we identified WDR5 as an actionable epigenetic regulator that is required for metastatic progression in models of triple-negative breast cancer. We found that knockdown of WDR5 in breast cancer cells independently impaired their tumorigenic as well as metastatic capabilities. Mechanistically, WDR5 promotes cell growth by increasing ribosomal gene expression and translation efficiency in a KMT2-independent manner. Consistently, pharmacological inhibition or degradation of WDR5 impedes cellular translation rate and the clonogenic ability of breast cancer cells. Furthermore, a combination of WDR5 targeting with mTOR inhibitors leads to potent suppression of translation and proliferation of breast cancer cells. These results reveal novel therapeutic strategies to treat metastatic breast cancer.

Abstract Background and aim Acute severe autoimmune hepatitis (AS-AIH) is a rare cause of acute liver failure (ALF), which is often neglected and delayed in treatment. The purpose of this study was to analyze the clinical characteristics and therapeutic effects of AS-AIH. Methods Retrospective analysis was performed. AIH was diagnosed according to the International Autoimmune Hepatitis Group (IAIHG) criteria revised in 1999. AS-AIH was defined as an acute presentation (onset of symptoms to presentation of ≤ 26 weeks) and INR of ≥ 1.5, and no histologic evidence of cirrhosis. Results Twelve patients were diagnosed as AS-AIH. At baseline, median immunoglobulin G was 28.35 g/L (range, 11.4–49.2). Ten (83.3%) patients were antinuclear antibodies and/or anti-smooth muscle antibodies positive. The prominent histologic characteristics were lobular necrosis/inflammation (91.7%) and plasma cell infiltration (100%). All patients received corticosteroid therapy. Death occurred in 2 (16.7%) patients within 30 days resulted from ALF. The average interval between the onset of symptoms and initiation of corticosteroid therapy in deceased patients was 65 days, compared with 19 days for survivors. Conclusions AS-AIH is an uncommon disease with poor outcomes. Patients with acute severe hepatitis of unknown cause should be minded the possibility of AS-AIH and corticosteroids should be considered as soon as possible.

Although a large number of trials in the SCI field have been conducted, few proven gains have been realized for patients. In the present study, we determined the efficacy of a novel combination treatment involving surgical intervention and long-term weight-bearing walking training in spinal cord injury (SCI) subjects clinically diagnosed as complete or American Spinal Injury Association Impairment Scale (AIS) Class A (AIS-A). A total of 320 clinically complete SCI subjects (271 male and 49 female), aged 16–60 years, received early (≤ 7 days, n = 201) or delayed (8–30 days, n = 119) surgical interventions to reduce intraspinal or intramedullary pressure. Fifteen days post-surgery, all subjects received a weight-bearing walking training with the “Kunming Locomotion Training Program (KLTP)” for a duration of 6 months. The neurological deficit and recovery were assessed using the AIS scale and a 10-point Kunming Locomotor Scale (KLS). We found that surgical intervention significantly improved AIS scores measured at 15 days post-surgery as compared to the pre-surgery baseline scores. Significant improvement of AIS scores was detected at 3 and 6 months and the KLS further showed significant improvements between all pair-wise comparisons of time points of 15 days, 3 or 6 months indicating continued improvement in walking scores during the 6-month period. In conclusion, combining surgical intervention within 1 month post-injury and weight-bearing locomotor training promoted continued and statistically significant neurological recoveries in subjects with clinically complete SCI, which generally shows little clinical recovery within the first year after injury and most are permanently disabled. This study was approved by the Science and Research Committee of Kunming General Hospital of PLA and Kunming Tongren Hospital, China and registered at ClinicalTrials.gov (Identifier: NCT04034108) on July 26, 2019.

Clinical proteomics critically depends on the ability to acquire highly reproducible data over an extended period of time. Here, the authors assess reproducibility over four months across different mass spectrometers and develop a computational approach to mitigate variation among instruments over time.

This paper addresses the finite-time controllability and set controllability of impulsive probabilistic Boolean control networks (IPBCNs). Firstly, using the algebraic state space representation (ASSR) method, the transition probability matrix of IPBCNs is established. Secondly, a kind of finite step reachability matrix with probability one is constructed, based on which, several effective criteria are proposed for the finite-time controllability with probability one of IPBCNs. Thirdly, a necessary and sufficient condition is presented for the finite-time set controllability with probability one of IPBCNs by constructing the set controllability probability distribution vector. Finally, the obtained results are extended to switching topology case.

Fish nocardiosis is a chronic, systemic, granulomatous disease in aquaculture. Nocardia seriolae has been reported to be one of the main pathogenic bacteria of fish nocardiosis. There are few studies on the associated virulence factors and pathogenesis of N. seriolae. Alanine dehydrogenase (ALD), which may be a secreted protein, was discovered by analysis using bioinformatics methods throughout the whole genomic sequence of N. seriolae. Nevertheless, the roles of ALD and its homologs in the pathogenesis of N. seriolae are not demonstrated. In this study, the function of N. seriolae ALD (NsALD) was preliminarily investigated by gene cloning, host cell subcellular localization, secreted protein identification, and cell apoptosis detection. Identification of the extracellular products of N. seriolae via mass spectrometry (MS) analysis revealed that NsALD is a secreted protein. In addition, subcellular localization of NsALD-GFP recombinant protein in fathead minnow (FHM) cells showed that the strong green fluorescence co-localized with the mitochondria. Moreover, apoptosis assays demonstrated that the overexpression of NsALD induces apoptosis in FHM cells. This study may lay the foundation for further exploration of the function of NsALD and facilitate further understanding of the pathogenic mechanism and the associated virulence factors of N. seriolae.

Surface-enhanced Raman scattering (SERS) as a label-free, non-contact, highly sensitive, and powerful technique has been widely applied in determining bio- and chemical molecules with fingerprint recognitions. 2-dimensional (2D) materials with layered structures, tunable optical properties, good chemical/physical stabilities, and strong charge–transfer interaction with molecules have attracted researchers’ interests. Two-D materials with a large and flat surface area, as well as good biocompatibility have been considered promising candidates in SERS and widely applied in chemical and bio-applications. It is well known that the noble metallic nanostructures with localized surface plasmon effects dominate the SERS performance. The combination of noble metallic nanostructure with 2D materials is becoming a new and attractive research domain. Until now, the SERS substrates combined with 2D materials, such as 2D graphene/metallic NPs, 2D materials@metallic core-shell structures, and metallic structure/2D materials/metallic structure are intensely studied. In this review, we introduce different kinds of fabrication strategies of 2D and 3D SERS substrates combing with 2D materials as well as their applications. We hope this review will help readers to figure out new ideas in designing and fabricating SERS substrates with high SERS performance that could enlarge the applicable domains of SERS.

Hepatocellular carcinoma (HCC), a highly aggressive tumor, has high incidence and mortality rates. Recently, immunotherapies have been shown to be a promising treatment in HCC. The results of either the CheckMate-040 or IMbrave 150 trials demonstrate the importance of immunotherapy in the systemic treatment of liver cancer. Thus, in this study, we tried to establish a reliable prognostic model for liver cancer based on immune-related genes (IRGs) and to provide a new insight for immunotherapy of HCC. In this study, we used four datasets that incorporated 851 HCC samples, including 340 samples with complete clinical information from the cancer genome atlas (TCGA) database, to establish an effective model for predicting the prognosis of HCC patients based on the differential expression of IRGs and validated the prognostic model using the data from International Cancer Genome Consortium (ICGC). The top 6 characteristic IRGs identified by protein-protein interaction (PPI) network analysis, MMP9, FOS, CAT, ESR1, ANGPTL3, and KLKB1, were selected for further study. In addition, we assessed the correlations of the six characteristic IRGs with the tumor immune microenvironment, clinical stage, and sensitivity to anti-cancer drugs. We also explored whether the differential expression of the characteristic IRGs was specific to HCC or present in pan-cancer. The expression levels of the six characteristic IRGs were significantly different between most tumor tissues and adjacent normal tissues. In addition, these characteristic IRGs showed a strong association with immune cell infiltration in HCC patients. We found that MMP9 and ESR1 were independent prognostic factors for HCC, while CAT, ESR1, and KLKB1 were associated with the clinical stage. We collected HCC paraffin sections from 24 patients from Xijing hospital to identify the differential expression of the five genes (MMP9, ESR1, CAT, FOS, and KLKB1). Finally, the results of decision curve analysis (DCA) and nomogram revealed that our models provided a prognostic benefit for most HCC patients and the predicted overall survival (OS) was consistent with the actual OS. In conclusion, we systemically constructed a novel prognostic model that provides new insights into HCC.

With the development of robot technology and the extensive application of robots, the research on special robots for some complex working environments has gradually become a hot topic. As a special robot applied to transmission towers, the climbing robot can replace humans to work at high altitudes to complete bolt tightening, detection, and other tasks, which improves the efficiency of transmission tower maintenance and ensures personal safety. However, it is mostly the ability to autonomously locate in the complex environment of the transmission tower that limits the industrial applications of the transmission tower climbing robot. This paper proposes an intelligent positioning method that integrates the three-dimensional information model of transmission tower and visual sensor data, which can assist the robot in climbing and adjusting to the designated working area to guarantee the working accuracy of the climbing robots. The experimental results show that the positioning accuracy of the method is within 1 cm.

Summary: Mitochondria are highly dynamic organelles subjected to fission and fusion events. During mitosis, mitochondrial fission ensures equal distribution of mitochondria to daughter cells. If and how this process can actively drive mitotic progression remains largely unknown. Here, we discover a pathway linking mitochondrial fission to mitotic progression in mammalian cells. The mitochondrial fission factor (MFF), the main mitochondrial receptor for the Dynamin-related protein 1 (DRP1), is directly phosphorylated by Protein Kinase D (PKD) specifically during mitosis. PKD-dependent MFF phosphorylation is required and sufficient for mitochondrial fission in mitotic but not in interphasic cells. Phosphorylation of MFF is crucial for chromosome segregation and promotes cell survival by inhibiting adaptation of the mitotic checkpoint. Thus, PKD/MFF-dependent mitochondrial fission is critical for the maintenance of genome integrity during cell division.

Protein degradation in clinical samples is largely unexplored. Here, the authors analyze the transcriptome and proteome of clinical tissue samples and develop an algorithm to assess protein degradation, showing that protein degradation is negligible in most tissue samples and does not correlate with transcript degradation.

This paper is concerned with the existence of mild solutions and total controllability for a class of non-autonomous measure evolution systems with non-instantaneous impulses and state-dependent delay. By using the theory of evolution family and Krasnoselskii’s fixed point theorem, the existence of mild solutions and total controllability for the considered systems is obtained. Finally, we give two applications to support the validity of the study.

This paper deals with a class of boundary value problems of second-order differential equations with impulses and discontinuity. The existence of single or multiple positive solutions to discontinuous differential equations with impulse effects is established by using the nonlinear alternative of Krasnoselskii’s fixed point theorem for discontinuous operators on cones. Finally, an example is given to illustrate the main results.

Abstract This paper is concerned with the following nonlinear fractional Kirchhoff equation: (a+b∫R3|(−Δ)s2u|2dx)(−Δ)su+V(x)u=f(u),x∈R3, $$\biggl(a+b \int _{\mathbb{R}^{3}} \bigl\vert (-\Delta )^{\frac{s}{2}}u \bigr\vert ^{2}\,dx \biggr) (- \Delta )^{s}u+V(x)u=f(u), \quad x\in \mathbb{R}^{3}, $$ where s∈(34,1) $s\in (\frac{3}{4}, 1)$, a>0 $a>0$, b≥0 $b\geq 0$, (−Δ)s $(-\Delta )^{s}$ denotes the fractional Laplacian operator. Based on the methods of mountain pass theorem and invariant sets of descending flow, the existence of a positive solution, a negative solution, and multiple sign-changing solutions is obtained.

BackgroundEvidence from prevailing studies show that hepatocellular carcinoma (HCC) is among the top cancers with high mortality globally. Gene regulation at post-transcriptional level orchestrated by RNA-binding proteins (RBPs) is an important mechanism that modifies various biological behaviors of HCC. Currently, it is not fully understood how RBPs affects the prognosis of HCC. In this study, we aimed to construct and validate an RBP-related model to predict the prognosis of HCC patients.MethodsDifferently expressed RBPs were identified in HCC patients based on the GSE54236 dataset from the Gene Expression Omnibus (GEO) database. Integrative bioinformatics analyses were performed to select hub genes. Gene expression patterns were validated in The Cancer Genome Atlas (TCGA) database, after which univariate and multivariate Cox regression analyses, as well as Kaplan-Meier analysis were performed to develop a prognostic model. Then, the performance of the prognostic model was assessed using receiver operating characteristic (ROC) curves and clinicopathological correlation analysis. Moreover, data from the International Cancer Genome Consortium (ICGC) database were used for external validation. Finally, a nomogram combining clinicopathological parameters and prognostic model was established for the individual prediction of survival probability.ResultsThe prognostic risk model was finally constructed based on two RBPs (BOP1 and EZH2), facilitating risk-stratification of HCC patients. Survival was markedly higher in the low-risk group relative to the high-risk group. Moreover, higher risk score was associated with advanced pathological grade and late clinical stage. Besides, the risk score was found to be an independent prognosis factor based on multivariate analysis. Nomogram including the risk score and clinical stage proved to perform better in predicting patient prognosis.ConclusionsThe RBP-related prognostic model established in this study may function as a prognostic indicator for HCC, which could provide evidence for clinical decision making.

This paper is mainly concerned with a class of fractional p,q-difference equations under p,q-integral boundary conditions. Multiple positive solutions are established by using the topological degree theory and Krein–Rutman theorem. Finally, two examples are worked out to illustrate the main results.

In budding yeast, the mitotic exit network (MEN), a GTPase signaling cascade, integrates spatial and temporal cues to promote exit from mitosis. This signal integration requires transmission of a signal generated on the cytoplasmic face of spindle pole bodies (SPBs; yeast equivalent of centrosomes) to the nucleolus, where the MEN effector protein Cdc14 resides. Here, we show that the MEN activating signal at SPBs is relayed to Cdc14 in the nucleolus through the dynamic localization of its terminal kinase complex Dbf2-Mob1. Cdc15, the protein kinase that activates Dbf2-Mob1 at SPBs, also regulates its nuclear access. Once in the nucleus, priming phosphorylation of Cfi1/Net1, the nucleolar anchor of Cdc14, by the Polo-like kinase Cdc5 targets Dbf2-Mob1 to the nucleolus. Nucleolar Dbf2-Mob1 then phosphorylates Cfi1/Net1 and Cdc14, activating Cdc14. The kinase-primed transmission of the MEN signal from the cytoplasm to the nucleolus exemplifies how signaling cascades can bridge distant inputs and responses.

Abstract Profiling of biological relationships between different molecular layers dissects regulatory mechanisms that ultimately determine cellular function. To thoroughly assess the role of protein post‐translational turnover, we devised a strategy combining pulse stable isotope‐labeled amino acids in cells (pSILAC), data‐independent acquisition mass spectrometry (DIA‐MS), and a novel data analysis framework that resolves protein degradation rate on the level of mRNA alternative splicing isoforms and isoform groups. We demonstrated our approach by the genome‐wide correlation analysis between mRNA amounts and protein degradation across different strains of HeLa cells that harbor a high grade of gene dosage variation. The dataset revealed that specific biological processes, cellular organelles, spatial compartments of organelles, and individual protein isoforms of the same genes could have distinctive degradation rate. The protein degradation diversity thus dissects the corresponding buffering or concerting protein turnover control across cancer cell lines. The data further indicate that specific mRNA splicing events such as intron retention significantly impact the protein abundance levels. Our findings support the tight association between transcriptome variability and proteostasis and provide a methodological foundation for studying functional protein degradation.

Abstract This paper deals with the existence of infinitely many solutions for a class of impulsive fractional boundary value problems with p-Laplacian. Based on a variant fountain theorem, the existence of infinitely many nontrivial high or small energy solutions is obtained. In addition, two examples are worked out to illustrate the effectiveness of the main results.

This paper is mainly concerned with the exact controllability for a class of impulsive ψ-Caputo fractional evolution equations with nonlocal conditions. First, by generalized Laplace transforms, a mild solution for considered problems is introduced. Next, by the Mönch fixed point theorem, the exact controllability result for the considered systems is obtained under some suitable assumptions. Finally, an example is given to support the validity of the main results.

Trisomy 21 (T21) is a major cause of Down syndrome but little is known about its impact on the cellular proteome. Here, the authors define the proteome of T21 fibroblasts and its turnover and also map proteomic differences in monozygotic T21-discordant twins, revealing extensive, organelle-specific changes caused by T21.

SWATH-mass spectrometry consists of a data-independent acquisition and a targeted data analysis strategy that aims to maintain the favorable quantitative characteristics on the scale of thousands of proteins. Here, using data generated by eleven groups worldwide, the authors show that SWATH-MS is capable of generating highly reproducible data across different laboratories.

Summary: Alternative splicing is a critical determinant of genome complexity and, by implication, is assumed to engender proteomic diversity. This notion has not been experimentally tested in a targeted, quantitative manner. Here, we have developed an integrative approach to ask whether perturbations in mRNA splicing patterns alter the composition of the proteome. We integrate RNA sequencing (RNA-seq) (to comprehensively report intron retention, differential transcript usage, and gene expression) with a data-independent acquisition (DIA) method, SWATH-MS (sequential window acquisition of all theoretical spectra-mass spectrometry), to capture an unbiased, quantitative snapshot of the impact of constitutive and alternative splicing events on the proteome. Whereas intron retention is accompanied by decreased protein abundance, alterations in differential transcript usage and gene expression alter protein abundance proportionate to transcript levels. Our findings illustrate how RNA splicing links isoform expression in the human transcriptome with proteomic diversity and provides a foundation for studying perturbations associated with human diseases. : Liu et al. have developed an integrative approach to ask whether perturbations in mRNA splicing patterns alter the composition of the proteome. Their findings illustrate how RNA splicing links isoform expression in the human transcriptome with proteomic diversity and provides a foundation for studying perturbations associated with human diseases. Keywords: alternative splicing, proteomics, RNA

Summary: Accurate classification of breast tumors is vital for patient management decisions and enables more precise cancer treatment. Here, we present a quantitative proteotyping approach based on sequential windowed acquisition of all theoretical fragment ion spectra (SWATH) mass spectrometry and establish key proteins for breast tumor classification. The study is based on 96 tissue samples representing five conventional breast cancer subtypes. SWATH proteotype patterns largely recapitulate these subtypes; however, they also reveal varying heterogeneity within the conventional subtypes, with triple negative tumors being the most heterogeneous. Proteins that contribute most strongly to the proteotype-based classification include INPP4B, CDK1, and ERBB2 and are associated with estrogen receptor (ER) status, tumor grade status, and HER2 status. Although these three key proteins exhibit high levels of correlation with transcript levels (R > 0.67), general correlation did not exceed R = 0.29, indicating the value of protein-level measurements of disease-regulated genes. Overall, this study highlights how cancer tissue proteotyping can lead to more accurate patient stratification. : Bouchal et al. explore and confirm the suitability of SWATH-MS for proteotyping of human tumor samples at relatively high throughput. Results indicate that proteotype-based classification resolves more variability than is apparent from conventional subtyping and potentially improves current classification. Keywords: breast cancer, proteomics, tumor classification, tissue, SWATH-MS, data independent acquisition, transcriptomics