Protein Tyrosine Kinase Activity Research Articles

Magnetic photoactive Fe3O4@Sb2S3 nanospheres assisted homogeneous photoelectrochemical–electrochemical biosensing platform integrating with MOFs-derived bifunctional NPC@Thi nanopolyhedra-mediated dual “signal-on” strategy for protein tyrosine kinase activity

Abnormal protein tyrosine kinase (PTK) expression is associated with various diseases and thus it is of great significance in analysis of PTK activity via a reliable method. Herein, magnetic photoactive Fe3O4@Sb2S3 nanospheres and bifunctional thionine-embedded metal organic frameworks (MOFs)-derived nitrogen-doped porous carbon (NPC@Thi) nanopolyhedra were successfully synthesized for constructing a highly sensitive magnetic-assisted homogeneous photoelectrochemical (PEC)–electrochemical (EC) dual-mode biosensing platform for the detection of PTK (c-Src) activity. The Fe3O4@Sb2S3 nanospheres with good magnetic and photoactive properties assisted adequate biological recognition in a homogeneous system, simplified sample separation and facilitated sensor construction. After c-Src-mediated specific recognition, bifunctional NPC@Thi nanopolyhedra were connected to Fe3O4@Sb2S3 nanospheres. NPC@Thi not only enhanced the photocurrent of Fe3O4@Sb2S3 but also acted as the electrochemical probe. Thus, highly sensitive and reliable dual “signal-on” assay of c-Src activity with wide linear response ranges and low detection limits was achieved by the successful cooperation of PEC (10 fM to 10 nM, 7.2 fM) and EC (5 fM to 10 nM, 3.8 fM) biosensing strategies. The proposed homogeneous PEC–EC dual-mode biosensing platform is prospective to be extended to the analysis of other PTK activity and the discovery of PTK-targeted drug.

RESPONSE OF BLOOD PRESSURE TO RENAL DENERVATION IS NOT ASSOCIATED WITH GENETIC VARIANTS

Objective: Renal denervation (RDN) is an evidence-based therapeutic option in patients with hypertension. However, the blood pressure (BP) responses to RDN in individual patients is currently impossible to predict. We conducted an unbiased genomic screen to identify genetic variants that are associated with response to RDN. Design and method: We included 268 patients (34.7% female, mean age 62 ± 10 years, BMI 30.5 ± 5.1 kg/m2) with uncontrolled treatment resistant hypertension (baseline BP 166 ± 21 / 90 ± 15 mmHg) who underwent endovascular RDN using the Symplicity™ catheter (Medtronic Inc, USA). Response to 24 hour ambulatory BP was assessed after 6 months and divided into two groups, above and below the median response of 6.0 mmHg, taking pre-intervention 24-hour ambulatory BP and regression to the mean into account. Whole exome sequencing assessing 249,669 variants was conducted using Illumina NovaSeq technology read on a Nova Seq S4 Flow Cell device. Read quality was overall excellent with the eight poorest performing samples still covering 30 reads per base for 80% of the exome. Reads were analysed by a standardised variant calling pipeline, using principal component analysis followed by individual logistic regressions to model association of variants with phenotype where appropriate. Results: There has been no linkage found between individual gene variants and the response variable. These neutral findings were further confirmed after adjustment for sex and in a sensitivity analysis looking at tertiles of BP response. We also explored specific variants in AGT, ADD1, ADRB1, ADRB2 and SCNN1A that have been proposed as potential candidate genes for response of BP to RDN and again found no association (all p>0.13). Gene ontology analysis of variants across the two responder groups highlighted differences in biological processes such as cell adhesion and molecular function such as protein tyrosine kinase activity. Conclusions: Response to RDN is not robustly associated with the genetic make-up of patients with uncontrolled treatment resistant hypertension. Our data do not support the use of a genetic score to predict BP response. We found, however, different regulation of biological processes and cellular/molecular function that could provide insights into the mechanisms of RDN.

Bioinformatics analysis reveals the landscape of immune cell infiltration and novel immune-related biomarkers in moyamoya disease.

Objective: This study aimed to identify immune infiltration characteristics and new immunological diagnostic biomarkers in the cerebrovascular tissue of moyamoya disease (MMD) using bioinformatics analysis. Methods: GSE189993 and GSE141022 were downloaded from the GEO database. Differentially expressed gene and PPI analysis were performed. After performing WGCNA, the most significant module associated with MMD was obtained. Next, functional pathways according to GSEA, GO, and KEGG were enriched for the aforementioned core genes obtained from PPI and WGCNA. Additionally, immune infiltration, using the CIBERSORT deconvolution algorithm, immune-related biomarkers, and the relationship between these genes, was further explored. Finally, diagnostic accuracy was verified with ROC curves in the validation dataset GSE157628. Results: A total of 348 DEGs were screened, including 89 downregulated and 259 upregulated genes. The thistlel module was detected as the most significant module associated with MMD. Functional analysis of the core genes was chiefly involved in the immune response, immune system process, protein tyrosine kinase activity, secretory granule, and so on. Among 13 immune-related overlapping genes, 4 genes (BTK, FGR, PTPN11, and SYK) were identified as potential diagnostic biomarkers, where PTPN11 showed the highest specificity and sensitivity. Meanwhile, a higher proportion of eosinophils, not T cells or B cells, was demonstrated in the specific immune infiltration landscape of MMD. Conclusion: Immune activities and immune cells were actively involved in the progression of MMD. BTK, FGR, PTPN11, and SYK were identified as potential immune diagnostic biomarkers. These immune-related genes and cells may provide novel insights for immunotherapy in the future.

Genome-wide association study reveals candidate markers related to field fertility and semen quality traits in Holstein-Friesian bulls

In vitro assessment of bull semen quality is routinely used in bull semen processing centres in order to ensure that semen destined to be used in the field has passed minimum standards. Despite these stringent quality control checks, individual bulls that pass the quality control checks can still vary in field fertility by up to 25%. A genome-wide association study was undertaken to determine genetic markers associated with prefreeze and post-thaw bull sperm quality traits as well as field fertility. Genome-wide association analysis was performed using a single nucleotide polymorphism (SNP) regression mixed linear model in WOMBAT. Genes within a 250 Kb span of a suggestive (P ≤ 1 × 10−5) SNP were considered as candidate genes. One SNP was associated with adjusted pregnancy rate, and 21 SNPs were associated across the seven semen quality traits (P ≤ 1 × 10−5). Functional candidate genes include SIPA1L2 which was associated with adjusted pregnancy rate. This encodes a Rap GTPase-activating protein involved in Rap1 signalling pathway and was previously found to play a role in the process of sperm differentiation. Gene ontology (GO) analysis also identified significantly enriched biological processes involved protein tyrosine kinase activity including genes such as DYRK1A, TEC and TXK that were associated with sperm motility prior to freezing. Another candidate gene associated with post-thaw sperm motility was FHDC1 which coordinates actin filament and microtubule dynamics. The induced 11 GO terms in the ejaculates rejected after freezing trait were related to ATPase, phosphatase and hydrolase activity. These results reveal novel specific genomic regions and candidate genes associated with economically important phenotypes such as field fertility and semen quality traits.

Genes influencing deposition of melanin in breast muscle of the Xuefeng black bone chicken based on bioinformatic analysis.

The Xuefeng black bone chicken (XFBC) represents an important poultry genetic resource. However, the darkness in breast muscle is heterogeneous. The molecular genetic mechanisms underlying melanogenesis of breast muscle in XFBC remains unclear. This study used RNA-seq to compare the difference in transcriptome between hyperpigmentation and hypopigmentation of breast muscle. Six cDNA libraries were constructed for hyperpigmentation and hypopigmentation groups in XFBC. We identified 395 differently expressed genes (DEGs) between hyperpigmentation and hypopigmentation group (P<0.05, |log2FC|≥1). Gene ontology (GO) enrichment and the Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis indicated several differentially enriched biological functions and pathways involved in melanogenesis of the breast muscle. Gene set enrichment analysis (GSEA) GO analysis identified two significant gene sets, including the pathways of pigment metabolic process and transmembrane receptor protein tyrosine kinase activity. GSEA-KEGG analysis identified the process of tyrosine metabolism and several genes related with melanogenesis in breast muscle of the XFBC. The protein-protein interaction network was constructed and eight genes were clustered in the module. We identified nine hub genes, including TYR, TYRP1, DCT, GPR143, MLANA, SLC24A5, GPNMB, MLPH, and EDNRB2. Taken together, the DEGs and hub genes identified in the study provide a solid basis for the study of the genetic regulatory mechanisms involved the melanogenesis in the breast muscle of the XFBC.

Abstract LB060: Integrated analysis of transcriptomics and kinase activity data for better characterization of cancer models

Abstract Introduction: Quantitative measurements of transcripts and proteins are key to investigate the basal state of a biological system, while functional proteomics inform about the active state of regulatory networks. Here we describe how the integration of transcriptomics and kinase activity data lead to a better characterization of various cancer models. Methods: We performed RNA sequencing (RNAseq) and kinase activity profiling of 63 Patient Derived Xenograft (PDX) models from six tumor types (Breast, Ovarian, Colon, Melanoma, Lung and Acute Myeloid Leukemia, AML). RNAseq was performed on an Illumina NovaSeq platform. The data was DESeq2-normalized and log2-transformed. Protein Tyrosine Kinase and Serine-Threonine Kinase activities were profiled on PamChip® peptide microarray. To identify the role of kinase signaling related genes we defined a set of signaling-specific genes (n=2932), based on the elements from the reactome signal transduction pathway database (n=2560) and additional kinases (n=372) represented on Pamchip, that was used for further analysis. Integrated analysis of transcriptomics and kinase activity data was performed using Multi Omics Factor Analysis (MOFA). Results: Principal Component Analysis (PCA) of RNAseq data using all included genes or 2932 kinase signaling-specific genes showed clustering of the data according to cancer type, with ovarian cancer showing most heterogeneity, which indicates the importance of kinase signaling in these malignancies. Interestingly, with integrated RNAseq-Kinase activity data all except ovarian cancer show clustering of cancer types on the MOFA Factor 1 - Factor 3. Pathway analysis on the highest ranking 100 genes from principal component 1 of RNAseq data (capturing variation between AML and the other tumor types) resulted in 60 KEGG pathways. Importantly, highest ranking 50 genes and 47 peptides comprising MOFA Factor1 identified 115 significant KEGG pathways, and the statistical score of pathways identified by RNAseq alone was further improved. Finally, significant correlation between gene expression and kinase activity was found for selected PDX model per malignancy. Furthermore, ranking PDX models based on correlation score provided suitable tool to select PDX models for disease or pathway specific research question. Conclusion: Integrating transcriptomics with kinase activity data can be used to confirm transcriptomics findings on a functional level and provides deeper biological insights than transcriptomics alone. We show that integrative analysis leads to more significant and a higher number of enriched pathways. High correlation between two datasets allows for selecting animal models addressing specific research questions. Integrated analysis of transcriptomics and kinase activity data has great potential in improving diagnosis, prognosis and prediction of response to treatment. Citation Format: Dóra Schuller, Rik de Wijn, Dirk Pijnenburg, Tobias Deigner, Julia Schueler, Simar Pal Singh. Integrated analysis of transcriptomics and kinase activity data for better characterization of cancer models [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 2 (Clinical Trials and Late-Breaking Research); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(8_Suppl):Abstract nr LB060.

Abstract 2864: Selective suppression of melanoma lacking IFN-γ pathway by JAK inhibition depends on T cells and host TNF signaling

Abstract Therapeutic resistance to immune checkpoint blockers (ICBs) is a pressing issue, limiting ICB efficacy to a small proportion of cancer patients. Concerted efforts from our group and others have identified that loss of IFN-γ signaling genes in melanoma is a major mechanism of resistance to ICBs. However, strategies of overcoming this resistance have been largely elusive. Moreover, given the indispensable role of T cells in governing ICBs, little is known about how tumor-intrinsic loss of IFN-γ signaling impacts tumor-infiltrating T cells (TILs). In this study, we report that melanomas with defective IFN-γ signaling (IFNγR1KO) have reduced infiltration and function of TILs. Intriguingly, IFNγR1KO melanoma cells harbor a network of constitutively active protein tyrosine kinases (PTKs) centered on JAK1/2, essential downstream components of the IFN-γ signaling. Additional mechanistic studies reveal that JAK1/2 activation is mediated by augmentation of the mTOR pathway. Importantly, targeting active JAK1/2 with FDA-approved Ruxolitinib (Ruxo) selectively suppresses growth of IFNγR1KO but not scrambled control melanoma, in a T cell and host TNF-dependent manner. Together, our results highlight an important role of tumor-intrinsic IFN-γ signaling in shaping TILs and manifest a therapeutic target that can be leveraged to bypass ICB resistance of melanomas defective of IFN-γ signaling. Citation Format: Hongxing Shen, Fengyuan Huang, Xiangmin Zhang, Oluwagbemiga A. Ojo, Yuebin Li, Hoa Quang Trummell, Joshua C. Anderson, John Fiveash, Markus Bredel, Eddy Yang, Christopher Willey, Zechen Chong, James Bonner, Lewis Z. Shi. Selective suppression of melanoma lacking IFN-γ pathway by JAK inhibition depends on T cells and host TNF signaling [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2864.

Molecular mechanism of resveratrol combined with irinotecan in treatment of colorectal cancer

This study aimed to investigate the mechanism of resveratrol(RES) combined with irinotecan(IRI) in the treatment of colorectal cancer(CRC). The targets of RES, IRI, and CRC were obtained from databases, and the targets of RES combined with IRI in the treatment of CRC were acquired by Venn diagram. The protein functional cluster analysis, GO and KEGG enrichment analyses were performed. In addition, the protein-protein interaction(PPI) network was constructed. The core target genes were screened out and the target-signaling pathway network was set up. IGEMDOCK was used to dock the core target gene molecules. Besides, the relationship between the expression level of key target genes and the prognosis and immune infiltration of CRC was analyzed. Based on the in vitro cell experiment, the molecular mechanism of RES combined with IRI in the treatment of CRC was explored and analyzed. According to the results, 63 potential targets of RES combined with IRI were obtained for CRC treatment. Furthermore, cluster analysis revealed that protein functions included 23% transmembrane signal receptors, 22% protein modifying enzymes, and 14% metabolite converting enzymes. GO analysis indicated that BPs were mainly concentrated in protein autophosphorylation, CCs in receptor complex and plasma membrane, and MFs in transmembrane receptor protein tyrosine kinase activity. Moreover, KEGG signaling pathways were mainly enriched in central carbon metabolism in cancer. The key targets of RES combined with IRI in the treatment of CRC were PIK3CA, EGFR, and IGF1R, all of which were significantly positively correlated with the immune infiltration of CRC. As shown by the molecular docking results, PIK3CA had the most stable binding with RES and IRI. Compared with the results in the control group, the proliferation ability and EGFR protein expression of CRC cells in the RES-treated group, the IRI-treated group, and the RES+IRI treated group significantly decreased. Moreover, the cell proliferation ability and EGFR protein expression level of CRC cells in the RES+IRI treated group were remarkably lower than those in the IRI-treated group. In conclusion, PIK3CA, EGFR, and IGF1R are the key targets of RES combined with IRI in CRC treatment. In addition, RES can inhibit the proliferation of CRC cells and improve IRI chemoresistance by downregulating the EGFR signaling pathway.

Antidiabetic effect of Withania Somnifera Root in STZ induced diabetic rats

Objective: To study the antidiabetic effect of Withania Somnifera root powder (WSR) in Streptozotocin induced diabetic rats. Methods: Diabetes in rats were induced by injecting Streptozotocin intraperitoneally (45 mg/kg of body weight) to overnight fasting rats. Withania Somnifera root was given to group C (100 mg/ kg of body weight) and Pioglitazone to group D (20 mg/kg of body weight) orally for 4 weeks respectively. Blood glucose level, glycated Hb, insulin, protein tyrosine kinase activity and serum DPP-4 levels were measured. Results: Significant Improvement in body weight, glycemic profile, dyslipidemia and tyrosine kinase activity were observed in WSR treated rats at week 4 of the study compared to diabetic control rats. Blood glucose level, glycated Hb and Serum DPP4 levels, were also found to be decreased in WSR treated rats compared to diabetic control rats at the end of study. This is possibly due to augmented serum insulin levels and increased insulin sensitivity after treatment with WSR. Conclusion: The results from above study it was concluded that Withania Somnifera root powder has antidiabetic activity. Further study required to confirm the exact mechanism of action.

Drug Repurposing and Systems Biology approaches of Enzastaurin can target potential biomarkers and critical pathways in Colorectal Cancer

Colorectal cancer (CRC) is a severe health concern that results from a cocktail of genetic, epigenetic, and environmental abnormalities. Because it is the second most lethal malignancy in the world and the third-most common malignant tumor, but the treatment is unavailable. The goal of the current study was to use bioinformatics and systems biology techniques to determine the pharmacological mechanism underlying putative important genes and linked pathways in early-onset CRC.Computer-aided methods were used to uncover similar biological targets and signaling pathways associated with CRC, along with bioinformatics and network pharmacology techniques to assess the effects of enzastaurin on CRC.The KEGG and gene ontology (GO) pathway analysis revealed several significant pathways including in positive regulation of protein phosphorylation, negative regulation of the apoptotic process, nucleus, nucleoplasm, protein tyrosine kinase activity, PI3K-Akt signaling pathway, pathways in cancer, focal adhesion, HIF-1 signaling pathway, and Rap1 signaling pathway. Later, the hub protein module identified from the protein-protein interactions (PPIs) network, molecular docking and molecular dynamics simulation represented that enzastaurin showed strong binding interaction with two hub proteins including CASP3 (−8.6 kcal/mol), and MCL1 (−8.6 kcal/mol), which were strongly implicated in CRC management than other the five hub proteins. Moreover, the pharmacokinetic features of enzastaurin revealed that it is an effective therapeutic agent with minimal adverse effects.Enzastaurin may inhibit the potential biological targets that are thought to be responsible for the advancement of CRC and this study suggests a potential novel therapeutic target for CRC.

Comparative Expression Analysis of Breakpoint Cluster Region-Abelson Oncogene in Leukemia Patients.

Leukemia is a proliferative disorder of myeloid and lymphoid cells that may lead to death. Different types of leukemia have been reported, and several genetic and environmental factors are involved in their development. The Philadelphia chromosome causes the most common mutation known as breakpoint cluster region-Abelson oncogene (BCR-ABL1), which shows abnormal protein tyrosine kinase (PTK) activity. Basically, this activity is accountable for activating multiple pathways, including the inhibition of cell differentiation, controlled proliferation, and cell death. As a result of the absence of kinase activity, this mutation leads to the uncontrolled proliferation of leukocytes, causing chronic myeloid leukemia (CML), acute myeloid leukemia (AML), acute lymphoid leukemia (ALL), and chronic lymphocytic leukemia (CLL). This study aimed to evaluate the level of BCR-ABL1 expression in patients with these types of leukemias through qPCR. In brief, PBMCs were isolated from blood samples of patients, RNA was extracted from PBMCs, cDNA was synthesized, and the transcript levels of BCR-ABL1 in patients with each type of leukemia were determined by qPCR. The clinical, demographical, and experimental data were analyzed among CML, AML, and ALL patients. Results: The BCR-ABL1 expression levels are variable in all studied groups and are 90, 30-35, and 1-2.5% in CML, ALL, and AML, respectively. Demographic characteristics such as gender, BMI, age, family history, and clinical parameters along with CBC are also associated with the prevalence and diagnosis of leukemia. In a comparative expression analysis, the expression of BCR-ABL1 is onefold high in AML, but four- and sevenfold high in ALL and CML, respectively, as compared with normal levels. Conclusions: In this study, a significant difference was observed in the expression levels of BCR-ABL1 between CML (p = 0.0043) and ALL (p = 0.0006) and between CML and AML groups, and a high expression of BCR-ABL1 was noted in CML as compared with ALL and AML.

Application of hybrid biophysical-biochemical methods to unravel the molecular basis for auto-inhibition and activation of protein tyrosine phosphatase TCPTP/PTPN2.

Since the discovery of protein tyrosine phosphorylation as one of the critical post-translational modifications, it has been well known that the activity of protein tyrosine kinases (PTKs) is tightly regulated. On the other hand, protein tyrosine phosphatases (PTPs) are often regarded to act constitutively active, but recently we and others have shown that many PTPs are expressed in an inactive form due to allosteric inhibition by their unique structural features. Furthermore, their cellular activity is highly regulated in a spatiotemporal manner. In general, PTPs share a conserved catalytic domain comprising about 280 residues that is flanked by either an N-terminal or a C-terminal non-catalytic segment, which differs significantly in size and structure from each other and is known to regulate specific PTP's catalytic activity. The well-characterized non-catalytic segments can be globular or intrinsically disordered. In this work, we have focused on the T-Cell Protein Tyrosine Phosphatase (TCPTP/PTPN2) and demonstrated how the hybrid biophysical-biochemical methods can be applied to unravel the underlying mechanism through which TCPTP's catalytic activity is regulated by the non-catalytic C-terminal segment. Our analysis showed that TCPTP is auto-inhibited by its intrinsically disordered tail and trans-activated by Integrin alpha-1's cytosolic region.

Епідеміологія метаболічного синдрому та концепції механізмів його розвитку

The purpose of the study was to analyze literary sources with the study of modern views on the epidemiology of the metabolic syndrome and pro-inflammatory concepts of the mechanisms of the development of insulin resistance and type 2 diabetes mellitus, as one of its components. Materials and methods. Analytical and bibliosemantic methods were used in the research. During the scientific search, 53 sources of modern domestic and foreign literature were reviewed and analyzed. Results and discussion. Metabolic syndrome is the most common disorder of endocrine regulation, which is one of the threatening health care problems of the 21st century. In recent years, it remains almost the most important problem of endocrinology, cardiology, dietology, internal and family medicine. In recent decades, foreign researchers assign the most important role in the mechanism of metabolic syndrome development to adipocytes of adipose tissue. Most scientists consider insulin resistance to be the pathogenetic basis of metabolic syndrome. Insulin resistance manifests itself as the resistance of body cells to the action of insulin and occurs as a result of a violation of the insulin signal in these cells, the implementation of which is carried out by a membrane-mediated mechanism. Molecular manifestations of insulin resistance should be sought among the components of the insulin cascade, which includes the receptor, the substrate of the insulin receptor (IRS protein), the PI3-kinase cascade, and the GLUT-4 glucose transporter activation system. It should be noted that within the framework of the relationship between obesity, inflammation and insulin resistance, the following cytokines, TNF-α and IL-6, deserve more attention. TNF-α reduces the tyrosine protein kinase activity of the insulin receptor, the insulin-stimulated phosphorylation of its substrates (serine phosphorylation) and the expression of matrix ribonucleic acid GLUT-4 in muscle and adipose tissues, which is accompanied by impaired glucose transport. Also, TNF-α reduces the expression of the lipoprotein lipase gene, stimulates lipogenesis and synthesis of fatty acids, which additionally increases the degree of IP and obesity. IL-6 reduces the expression of the transporter GLUT-4 and the substrate of the insulin receptor, while also having an anti-inflammatory effect due to the reduction of TNF-α and interferon. Under the influence of this cytokine, the level of glycerol and free fatty acids in blood serum increases, which is accompanied by a lipotoxic effect on β-cells of the pancreas. Conclusion. Metabolic syndrome is a pathological process, the spread of which has become epidemic both in developed foreign countries and in Ukraine. One of the important components of the pathogenesis of the metabolic syndrome is insulin resistance, the mechanisms of which, in particular, are related to pro-inflammatory cytokine-mediated mechanisms affecting insulin signaling, glucose transport, involved in pathological changes in lipid synthesis and metabolism, as well as pro-oxidant and cytotoxic processes

Genetic Alterations of Melanoma Brain Metastases: A Systematic Review and Meta-Analysis.

Data on molecular alterations harbored by melanoma brain metastases (MBMs) are limited, and this has hampered the development of more effective therapeutic strategies. We conducted a systematic review and meta-analysis of all the studies reporting DNA sequencing data of MBMs, in order to identify recurrently mutated genes and molecular pathways significantly enriched for genetic alterations. We searched PubMed, Embase and Scopus for articles published from the inception of each database to June 30, 2021. We included in the analysis all the studies that reported individual patient data on DNA sequencing of MBMs, assessing single nucleotide variants (SNVs) and/or gene copy number variations (CNVs) in at least five tumor samples. Meta-analysis was performed for genes evaluated for SNVs and/or CNVs in at least two studies. Pooled proportions of samples with SNVs and/or CNVs was calculated by applying random-effect models based on the DerSimonian-Laird method. Gene-set enrichment analysis (GSEA) was performed to identify molecular pathways significantly enriched for mutated genes. Ten studies fulfilled the inclusion criteria and were included in the analysis, for a total of 531 samples of MBMs evaluated. Twenty-seven genes were found recurrently mutated with a meta-analytic rate of SNVs higher than 5%. GSEA conducted on the list of these 27 recurrently mutated genes revealed vascular endothelial growth factor-activated receptor activity and transmembrane receptor protein tyrosine kinase activity to be among the top 10 gene ontology (GO) molecular functions significantly enriched for mutated genes, while regulation of apoptosis and cell proliferation were among the top 10 significantly enriched GO biological processes. Notably, a high meta-analytic rate of SNVs was found in several actionable cancer-associated genes, such as all the vascular endothelial growth factor (VEGF) receptor isoforms (i.e., Flt1 and Flt2 genes, for both SNV rate: 0.22, 95% CI 0.04-0.49; KDR gene, SNV rate: 0.1, 95% CI 0.05-0.16). Finally, two tumor suppressor genes were characterized by a high meta-analytic rate of CNVs: CDKN2A/B (CNV rate: 0.59, 95% CI 0.23-0.90) and PTEN (CNV rate: 0.31, 95% CI 0.02-0.95). MBMs harbored actionable molecular alterations that could be exploited as therapeutic targets to improve the poor prognosis of patients.

PKMYT1 inhibits lung adenocarcinoma progression by abrogating AKT1 activity.

AKT hyperactivation drives malignant phenotypes in lung cancer via promoting tumor cell proliferation and survival. However, the relationship between dysregulation of cell cycle progression and AKT1 kinase activity is still not clear. Following the expression level of PKMYT1 in lung cancer, we performed cell proliferation, migration, invasion, and xenograft assays to determine the function of PKMYT1. We used RNA-seq to explore the anti-tumor mechanism of PKMYT1 and examined the effect of PKMYT1 on AKT1 activity. In this study, we report that PKMYT1 is downregulated in lung adenocarcinoma (LUAD) tissues and its low expression predicts a poor prognosis in LUAD patients. PKMYT1 exerts potent tumor-suppressive functions in LUAD cells by inhibiting AKT1 activation and thereby repressing cell cycle progression, which depends on its tyrosine and threonine protein kinase activity. Interestingly, PKMYT1 could directly bind AKT1 to abrogate AKT1 activation. Moreover, silencing AKT1 and inhibitors targeting the AKT pathway effectively reverse the promoting effects of PKMYT1 knockdown on proliferation, migration and invasion of LUAD cells. This work reveals the anti-tumor effect of PKMYT1 in LUAD and provides evidence to clarify the dual roles of PKMYT1 in tumor progression. Moreover, our findings broaden the current understandings on AKT1 activation and identify PKMYT1 as a potential negative regulator of AKT1 kinase activity, providing further insights into targeting the AKT pathway in LUAD.

Improving endothelial cell junction integrity by diphenylmethanone derivatives at oxidative stress: A dual-action directly targeting caveolar caveolin-1

Directly targeting caveolar caveolin-1 is a potential mechanism to regulate endothelial permeability, especially during oxidative stress, but little evidence on the topic limits therapeutics discoveries. In this study, we investigated the pharmacological effect of an antioxidant LM49 (5,2′-dibromo-2,4′,5′-trihydroxydiphenylmethanoe) and its five diphenylmethanone derivatives on endothelial permeability and establish two distinct mechanisms of action. Multiplex molecular assays with theoretical modeling indicate that diphenylmethanone molecules, including LM49, directly bind the caveolin-1 steric pocket of ASN53/ARG54, ILE49/ASP50, ILE18, LEU59, ASN60, GLU48 and ARG19 residues. They also indicated dynamic binding-affinity for diphenylmethanone derivatives. First, this molecular interaction at caveolin-1 pocket inhibits its phosphorylation at TYR14 residue in H2O2-injured endothelial cell. A positive correlation was established between diphenylmethanone derivative binding-affinity and caveolin-1 phosphorylation inhibition. Inhibition of caveolin-1 phosphorylation, however, was independent of the LM49-mediated variation of protein tyrosine kinase activity, suggesting a direct blockage of adenosine triphosphate substrate diffusion into cavelion-1 structure. Second, LM49 increases the expression of cellular adhesive and tight junction proteins, VE-cadherin and occludin, in H2O2-injured cell, in a dose dependent manner. A leakage assay of fluorescein isothiocyanate-labeled dextran 40 across cell monolayer suggested improvement in endothelial barrier integrity with diphenylmethanone treatments. Our results demonstrate a direct targeting effect of caveolin-1 on endothelial permeability, and should guide the diphenylmethanone therapy against oxidative stress-induced junction dysfunction, especially at caveolar membrane invagination.

A SERIES OF GENETIC EVENTS: FIP1L1/PDGFR REARRANGEMENT LEADING TO EOSINOPHILIC MYOCARDITIS

A SERIES OF GENETIC EVENTS: FIP1L1/PDGFR REARRANGEMENT LEADING TO EOSINOPHILIC MYOCARDITIS

Exploring the Molecular Mechanism of Zhi Bai Di Huang Wan in the Treatment of Systemic Lupus Erythematosus Based on Network Pharmacology and Molecular Docking Techniques

Objective: To investigate the molecular mechanism and simulated validation of Zhi Bai Di Huang Pill (ZBDHP) for the treatment of systemic lupus erythematosus (SLE) using network pharmacology and molecular docking techniques. Methods: The active ingredients of ZBDHP were obtained through the TCMSP database and the Canonical SMILES of the active ingredients were queried through Pubchem. The targets of the active ingredients were predicted in the SwissTarget database based on the SMILES. The SLE-related disease targets were obtained through the GeneCards, OMIM and DisGenets databases, and the intersection targets of ZBDHP and SLE were obtained using the Venny 2.1.0 online platform. Intersection targets build a visual protein interaction network (PPI) through the STRING database, and the core targets were identified by network topology analysis. GO analysis and KEGG pathway enrichment analysis of the intersecting targets were performed using the DAVID database. Finally, the molecular docking of the first four active ingredients and the first four core target genes were verified by Pubchem, the PDB database and CB-Dock online molecular docking technology. Results: ZBDHP screened 91 potential active ingredients and 816 potential targets. Among them, 141 genes were intersected by ZBDHP and SLE. The network topology analysis showed that the main active ingredients were Hydroxygenkwanin, Alisol B, asperglaucide, Cerevisterol, etc., and the key target genes were TNF, AKT1, EGFR, STAT3, etc. GO and KEGG enrichment analysis showed that common targets interfere with biological processes or molecular functions such as signal transduction protein phosphorylation, inflammatory response, transmembrane receptor protein tyrosine kinase activity, etc., through multiple signaling pathways, such as pathways in cancer, Kaposi sarcoma-associated herpesvirus infection, the PI3K-Akt signaling pathway, lipid and atherosclerosis, hepatitis B, etc. Molecular docking results showed that the active components of ZBDHP have good binding activity to the core targets of SLE. Conclusions: This study reveals that the ZBDHP treatment of SLE is a complex mechanistic process with multi-components, multi-targets and multi-pathways, and it may play a therapeutic role in SLE by inhibiting the production, proliferation and apoptosis of inflammatory factors. In conclusion, the present study provides a theoretical basis for further research on ZBDHP.

Ultraviolet Radiation Exposure and its Impacts on Cutaneous Phosphorylation Signaling in Carcinogenesis: Focusing on Protein Tyrosine Phosphatases†.

Sunlight exposure is a significant risk factor for UV-induced deteriorating transformations of epidermal homeostasis leading to skin carcinogenesis. The ability of UVB radiation to cause melanoma, as well as basal and squamous cell carcinomas, makes UVB the most harmful among the three known UV ranges. UVB-induced DNA mutations and dysregulation of signaling pathways contribute to skin cancer formation. Among various signaling pathways modulated by UVB, tyrosine phosphorylation signaling which is mediated by the action of protein tyrosine kinases (PTKs) on specific tyrosine residues is highly implicated in photocarcinogenesis. Following UVB irradiation, PTKs get activated and their downstream signaling pathways contribute to photocarcinogenesis by promoting the survival of damaged keratinocytes and increasing cell proliferation. While UVB activates oncogenic signaling pathways, it can also activate tumor suppressive signaling pathways as initial protective mechanisms to maintain epidermal homeostasis. Tyrosine dephosphorylation is one of the protective mechanisms and is mediated by the action of protein tyrosine phosphatases (PTPs). PTP can counteract UVB-mediated PTK activation and downregulate oncogenic signaling pathways. However, PTPs have not been studied extensively in photocarcinogenesis with previous studies regarding their inactivation induced by UVB. This current review will summarize the recent progress in the protective function of PTPs in epidermal photocarcinogenesis.

Selective suppression of melanoma lacking IFN-γ pathway by JAK inhibition depends on T cells and host TNF signaling

Therapeutic resistance to immune checkpoint blockers (ICBs) in melanoma patients is a pressing issue, of which tumor loss of IFN-γ signaling genes is a major underlying mechanism. However, strategies of overcoming this resistance mechanism have been largely elusive. Moreover, given the indispensable role of tumor-infiltrating T cells (TILs) in ICBs, little is known about how tumor-intrinsic loss of IFN-γ signaling (IFNγR1KO) impacts TILs. Here, we report that IFNγR1KO melanomas have reduced infiltration and function of TILs. IFNγR1KO melanomas harbor a network of constitutively active protein tyrosine kinases centered on activated JAK1/2. Mechanistically, JAK1/2 activation is mediated by augmented mTOR. Importantly, JAK1/2 inhibition with Ruxolitinib selectively suppresses the growth of IFNγR1KO but not scrambled control melanomas, depending on T cells and host TNF. Together, our results reveal an important role of tumor-intrinsic IFN-γ signaling in shaping TILs and manifest a targeted therapy to bypass ICB resistance of melanomas defective of IFN-γ signaling.