MAPK Family Research Articles

Design and synthesis of JNK1-targeted PROTACs and research on the activity.

Kinase dysregulation is greatly associated with cell growth, proliferation, differentiation and apoptosis, which indicates their great potential as therapeutic targets for treatment of numerous progressive disorders, including inflammatory, metabolic and autoimmune disorders, organ fibrosis and cancer. The c‑Jun N‑Terminal Kinase (JNK), as a member of MAPK family, is proved to be a potential target for the treatment of pulmonary fibrosis, which is the most common progressive and fatal fibrotic lung disease. As a new strategy, small-molecule-mediated targeted protein degradation pathway has the advantages of catalytic properties, overcoming drug resistance and expanding target space, which can circumvent the limitations associated with kinase inhibitors. Proteolysis targeting chimeras (PROTAC) contains a linker to concatenate a ligand of E3 ubiquitin ligase and a ligand for a protein of interest (POI). We developed a total of 20 JNK1-targeted PROTACs that induce proteasomal degradation of JNK1 components. The most active PROTAC molecule PA2 was then investigated by JNK1 enzyme assay and protein degradation assay, which suggested that PA2 had an anti-JNK1 ability and provided insights for the future use of JNK1-targeted PROTAC as treatment drugs for pulmonary fibrosis.

Ganoderic acid a alleviates Aβ25-35-induced HT22 cell apoptosis through the ERK/MAPK pathway: a system pharmacology and in vitro experimental validation.

Alzheimer's disease (AD) is a neurodegenerative disorder that occurs with aging. Ganoderma lucidum (Curtis.) P. Karst. (G. lucidum) is a traditional medicinal fungus believed to nourish the brain and anti-aging. Ganoderic acid A (GAA), a triterpenoid from G. lucidum, has demonstrated natural neuroprotective effects. This study aims to explore the therapeutic effect and molecular mechanism of GAA on AD. Systematic network pharmacology identified 95 targets, 8 biological functions, and multiple pathways. The results highlighted MAPK family members as core genes, with MAPK1 (ERK2) showing the highest binding affinity to GAA in molecular docking. In vitro experiments revealed that GAA dose-dependently increased the viability of Aβ25-35-injured HT22 cells and inhibited MAPK pathway-related protein expression. Similar to FR180204, 100 µM GAA significantly reversed ERK protein expression, oxidative stress markers, and mitochondrial damage in AD cell model. GAA also downregulated cleaved caspase-3 protein levels, apoptosis rates, Aβ and p-Tau expression by inhibiting the ERK signaling pathway. The therapeutic effect of GAA on AD was predicted and validated through network pharmacology and in vitro experiments. The ability of GAA to inhibit apoptosis via the ERK/MAPK signaling pathway positions it as a promising candidate for AD treatment.

An updated review of small-molecule HPK1 kinase inhibitors (2016-present).

Hematopoietic progenitor kinase 1 (HPK1) is a serine-threonine kinase specific to hematopoiesis and a member of the MAP4K family of Ste20-related protein kinases. Targeting HPK1 to ameliorate T cell exhaustion and enhance T cell functions is a promising strategy for clinical immunotherapies. Numerous studies have reported the progress in developing effective HPK1 inhibitors and elucidating their mechanisms of action. However, most inhibitors affect multiple signaling pathways, resulting in unintended side effects that limit their clinical development and application. Herein, we reviewed HPK1-related signaling pathways, clinical candidates and recent advances in small-molecule inhibitors targeting HPK1. Additionally, we present our perspectives on current challenges and potential future research field, hoping to provide inspiration for the development of novel HPK1 inhibitors.

Design, Synthesis, and Biological Evaluation of a Series of Spiro Analogues as Novel HPK1 Inhibitors.

Hematopoietic progenitor kinase 1 (HPK1) negatively affects T cell activation and proliferation and is a promising target for immunotherapy. Although HPK1 inhibitors have shown promising efficacy in preclinical models, none have been approved for clinical use. One significant challenge in developing an HPK1 inhibitor is the difficulty in designing a potent inhibitor with good kinase selectivity and pharmacokinetic properties. Here, we report a series of spiro HPK1 inhibitors with good potency and selectivity. Specifically, compound 16 exhibited potent HPK1 inhibition (IC50 = 2.67 nM), adequate selectivity toward the MAP4K family (>100-fold), and good selectivity against selected kinases (>300-fold). Compound 16 demonstrated moderate in vivo clearance and reasonable oral exposure in mice and rats. Notably, compound 16 possessed good antitumor efficacy in the CT26 murine colon cancer and a synergistic effect when combined with anti-PD-1. These exciting preclinical results support the continued development of this class of HPK1 inhibitors.

Map3k1 suppresses terminal differentiation of migratory eye progenitors in planarian regeneration.

Proper stem cell targeting and differentiation is necessary for regeneration to succeed. In organisms capable of whole body regeneration, considerable progress has been made identifying wound signals initiating this process, but the mechanisms that control the differentiation of progenitors into mature organs are not fully understood. Using the planarian as a model system, we identify a novel function for map3k1, a MAP3K family member possessing both kinase and ubiquitin ligase domains, to negatively regulate terminal differentiation of stem cells during eye regeneration. Inhibition of map3k1 caused the formation of multiple ectopic eyes within the head, but without controlling overall head, brain, or body patterning. By contrast, other known regulators of planarian eye patterning like WntA and notum also regulate head regionalization, suggesting map3k1 acts distinctly. Eye resection and regeneration experiments suggest that unlike Wnt signaling perturbation, map3k1 inhibition did not shift the target destination of eye formation in the animal. Instead, map3k1(RNAi) ectopic eyes emerge in the regions normally occupied by migratory eye progenitors, and the onset of ectopic eyes after map3k1 inhibition coincides with a reduction to eye progenitor numbers. Furthermore, RNAi dosing experiments indicate that progenitors closer to their normal target are relatively more sensitive to the effects of map3k1, implicating this factors in controlling the site of terminal differentiation. Eye phenotypes were also observed after inhibition of map2k4, map2k7, jnk, and p38, identifying a putative pathway through which map3k1 prevents differentiation. Together, these results suggest that map3k1 regulates a novel control point in the eye regeneration pathway which suppresses the terminal differentiation of progenitors during their migration to target destinations.

Roles of naïve CD4+ T cells and their differentiated subtypes in lung adenocarcinoma and underlying potential regulatory pathways

BackgroundNaïve CD4+ T cells and their differentiated counterparts play a significant regulatory role in the tumor immune microenvironment, yet their effects on lung adenocarcinoma (LUAD) are not fully understood.MethodsWe utilized Mendelian randomization to assess the causal association between naïve CD4+ T cells and LUAD. Employing a modified single-sample Gene Set Enrichment Analysis (ssGSEA) algorithm with The Cancer Genome Atlas (TCGA) database, we determined the infiltration levels of naïve CD4+ T cells and their differentiation subtypes and investigated their correlation with clinical characteristics. Potential regulatory pathways of T helper cells were identified through Mantel tests and Kyoto Encyclopedia of Genes and Genomes (KEGG) database enrichment analysis.ResultsMendelian randomization analysis revealed an inhibitory effect of naïve CD4+ T cells on LUAD (false discovery rate < 0.05), which was corroborated by observational experiments using TCGA database. Specifically, T helper cell type 2 demonstrated a promotive effect on LUAD in terms of overall, disease-free, and progression-free survival (p < 0.05). Moreover, regulatory T cells exhibited a protective effect on LUAD in terms of disease-specific survival (p < 0.01). Concurrently, we explored the overall impact of naïve CD4+ T cell differentiation subtypes on LUAD, revealing upregulation in pathways such as neutrophil degranulation, MAPK family signaling pathways, and platelet activation, signaling, and aggregation.ConclusionNaïve CD4+ T cells and their differentiated counterparts play essential regulatory roles in the tumor immune microenvironment, demonstrating bidirectionality in their effects.Thus, elucidating the mechanisms and developing novel cell differentiation-inducing agents will benefit anti-cancer therapy.

Synergistic drug interactions of the histone deacetylase inhibitor givinostat (ITF2357) in CRLF2-rearranged pediatric B-cell precursor acute lymphoblastic leukemia identified by high-throughput drug screening

Combining multiple drugs broadens the window of therapeutic opportunities and is crucial for diseases that are currently lacking fully curative treatments. A powerful emerging tool for selecting effective drugs and combinations is the high-throughput drug screening (HTP). The histone deacetylase inhibitor (HDACi) givinostat (ITF2357) has been shown to act effectively against CRLF2-rearranged pediatric B-cell precursor acute lymphoblastic leukemia (BCP-ALL), a subtype characterized by poor outcome and enriched in children with Down Syndrome, very fragile patients with a high susceptibility to treatment-related toxicity. The aim of this study is to investigate possible synergies with givinostat for these difficult-to-treat patients by performing HTP screening with a library of 174 drugs, either approved or in preclinical studies. By applying this approach to the CRLF2-r MHH-CALL-4 cell line, we identified 19 compounds with higher sensitivity in combination with givinostat compared to the single treatments. Next, the synergy between givinostat and the promising candidates was further validated in CRLF2r cell lines with a broad matrix of concentrations. The combinations with trametinib (MEKi) or venetoclax (BCL2i) were found to be the most effective and with the greatest synergy across three metrics (ZIP, HAS, Bliss). Their efficacy was confirmed in primary blasts treated ex vivo at concentration ranges with a safe profile on healthy cells. Finally, we described givinostat-induced modifications in gene expression of MAPK and BCL-2 family members, supporting the observed synergistic interactions. Overall, our study represents a model of drug repurposing strategy using HTP screening for identifying synergistic, efficient, and safe drug combinations.

The effect of the absence of nucleus on the amount of JNK enzyme from the MAPK family of red blood cells and its comparison with white blood cells

The effect of the absence of nucleus on the amount of JNK enzyme from the MAPK family of red blood cells and its comparison with white blood cells

A retrospective analysis of the clinicopathological features and prognostic value of MAPK12 protein expression in diffuse large B-cell lymphoma

PurposeMitogen-activated protein kinase 12 (MAPK12), also known as p38γ, is a member of the p38 MAPK family and plays a crucial role in tumor occurrence and invasion. However, there is still uncertainty regarding MAPK12 involvement in diffuse large B-cell lymphoma (DLBCL).MethodsOur study investigated the expression of MAPK12 mRNA in various types of cancer using bioinformatic analysis. Furthermore, we performed immunohistochemistry (IHC) to detect the expression of MAPK12 in patients with DLBCL and compared clinical indicators and survival rates.ResultsWe found that the high expression rate of MAPK12 was 43.1% in DLBCL patients. Several clinical indicators, including IPI scores, Hans classifications, LDH levels, and Ki-67 expression were closely associated with MAPK12 expression. Survival analysis revealed that higher expression of MAPK12 was significantly correlated with shorter progression-free survival (PFS) and overall survival (OS) in DLBCL patients. In addition, both univariate and multivariate analyses revealed IPI score, MAPK12 expression, and rituximab use as the independent OS risk factors (P < 0.05). To explore the functional role of MAPK12 in DLBCL, weighted gene co-expression network analysis (WGCNA) and gene ontology (GO) were used to confirm the involvement of MAPK12 in the regulation of type II interferon production, positive regulation of lymphocyte proliferation, and other related biological processes.ConclusionDLBCL patients have poor prognoses when MAPK12 levels are high, which is expected to be a therapeutic target and prognostic factor.

Comprehensive prognostic and immune analysis of sterol O-acyltransferase 1 in patients with hepatocellular carcinoma.

Sterol O-acyltransferase 1 (SOAT1) is an important target in the diagnosis and treatment of liver cancer. However, the prognostic value of SOAT1 in patients with hepatocellular carcinoma (HCC) is still not clear. To investigate the correlation of SOAT1 expression with HCC, using RNA-seq and gene expression data of The Cancer Genome Atlas (TCGA)-liver hepatocellular carcinoma (LIHC) and pan-cancer. The correlation between SOAT1 expression and HCC was analyzed. Cox hazard regression models were conducted to investigate the prognostic value of SOAT1 in HCC. Overall survival and disease-specific survival were explored based on TCGA-LIHC data. Biological processes and functional pathways mediated by SOAT1 were characterized by gene ontology (GO) analysis and the Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis of differentially expressed genes. In addition, the protein-protein interaction network and co-expression analyses of SOAT1 in HCC were performed to better understand the regulatory mechanisms of SOAT1 in this malignancy. SOAT1 and SOAT2 were highly expressed in unpaired samples, while only SOAT1 was highly expressed in paired samples. The area under the receiver operating characteristic curve of SOAT1 expression in tumor samples from LIHC patients compared with para-carcinoma tissues was 0.748, while the area under the curve of SOAT1 expression in tumor samples from LIHC patients compared with GTEx was 0.676. Patients with higher SOAT1 expression had lower survival rates. Results from GO/KEGG and gene set enrichment analyses suggested that the PI3K/AKT signaling pathway, the IL-18 signaling pathway, the calcium signaling pathway, secreted factors, the Wnt signaling pathway, the Jak/STAT signaling pathway, the MAPK family signaling pathway, and cell-cell communication were involved in such association. SOAT1 expression was positively associated with the abundance of macrophages, Th2 cells, T helper cells, CD56bright natural killer cells, and Th1 cells, and negatively linked to the abundance of Th17 cells, dendritic cells, and cytotoxic cells. Our findings demonstrate that SOAT1 may serve as a novel target for HCC treatment, which is helpful for the development of new strategies for immunotherapy and metabolic therapy.

Abstract 2652: NDI-101150 is a potent and highly selective HPK1 inhibitor that both synergizes with and differentiates from anti-PD1 immune checkpoint blockade

Abstract Introduction HPK1 is a member of the MAP4K family of protein serine/threonine kinases that negatively regulates activation signals in multiple immune cells and is an attractive therapeutic target for many cancers. Using structure-based drug design, we developed a highly selective HPK1 inhibitor, NDI-101150, with nanomolar potency and physiochemical properties suitable for once daily oral administration. Methods &amp; Results HPK1 inhibition with NDI-101150 has a differentiated pharmacology profile when compared directly to PD1 blockade. NDI-101150 was able to overcome the immunosuppressive effects of prostaglandin E2, adenosine, and TGF-β on human T cell activation. HPK1 inhibition also directly limited the suppressive capacity of T regulatory cells. In contrast, anti-PD1 treatment was not able to overcome the immunosuppressive activity associated with any of these mechanisms. In vivo, HPK1 inhibition significantly enhanced the production of antigen-specific antibodies in response to immunization with both T-dependent and T-independent antigens, while anti-PD1 treatment had minimal to no effects on antibody production. In the EMT-6 syngeneic model, 7/10 mice receiving NDI-101150 exhibited complete tumor regressions, compared to only 1/10 from an anti-PD1 cohort. Furthermore, NDI-101150 established a robust and durable immune memory response as 100% of treated mice showed complete rejection of tumor growth upon subsequent re-challenge. HPK1 inhibition with NDI-101150 also shows robust synergy with PD1 blockade in vitro and in vivo. Both NDI-101150 and anti-PD1 were able to reinvigorate exhausted human T cells, restoring cytokine production and proliferative effects in mixed lymphocyte reactions. Further, in combination HPK1 inhibition and PD1 blockade synergized to enhance the activity of exhausted human T cells to an extent exceeding that observed with naïve human T cells in similar experiments. In vivo, while NDI-101150 or anti-PD1 treatments alone induced tumor growth inhibition in the murine CT-26 syngeneic tumor model, NDI-101150 and anti-PD1 in combination mediated complete tumor regressions in several mice. Furthermore, NDI-101150 seemed to induce a durable immune memory response as evidenced by the combination-treated animals showing complete rejection of tumor growth upon subsequent re-challenge, without any further dosing of NDI-101150. Conclusions Pharmacological inhibition of HPK1 with NDI-101150 represents a powerful system-wide immunomodulatory approach with the potential to enhance anti-tumor immunity in patients failing to respond to currently approved immune checkpoint therapies or in combination with immune checkpoint blockade therapies. NDI-101150 is currently being tested in a Phase 1/2 trial (NCT05128487) in patients with advanced solid tumors. Citation Format: David Ciccone, Fu-Shan Kuo, Scott Daigle, Neelu Kaila, Gene Yau, Mark Carlson, Denise Levasseur, Bhaskar Srivastava, Frank G. Basile, Christine Loh. NDI-101150 is a potent and highly selective HPK1 inhibitor that both synergizes with and differentiates from anti-PD1 immune checkpoint blockade [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 2652.

Abstract 3967: ISM9182A, a novel HPK1 inhibitor, exhibits immune modulatory activity and robust monotherapy anti-tumor effects in preclinical studies

Abstract The hematopoietic progenitor kinase 1 (HPK1 or MAP4K1) is a member of the Ste20 serine/threonine kinase super family that negatively regulates immune cells, including T cells, B cells and dendritic cells (DCs). Thus HPK1 is an attractive immuno-oncology target in many tumor types. Here, we report ISM9182A as a novel, potent, and selective inhibitor of HPK1.ISM9182A demonstrated nanomolar enzymatic potency against HPK1 with &amp;gt;100-fold selectivity against other MAP4K family kinases. ISM9182A inhibited the phosphorylation of SLP-76 at Ser376 dose-dependently with an IC50 of 30nM in human peripheral blood mononuclear cells. Moreover, ISM9182A enhanced IL-2 and IFNγ secretion upon CD3/CD28 co-stimulation in a dose-dependent manner without affecting T cell proliferation. Treatment of human pan T cells with ISM9182A also significantly reversed PGE2-mediated immunosuppression of human pan T cell activation and further boosted IL-2 and IFNγ secretion compared to CD3/CD28 co-stimulation. In monocyte-derived DCs stimulated with TNF-α/IL-1β/PGE-2, ISM9182A significantly upregulated CD80, CD86, and CD83 expression, demonstrating its potency to promote DC maturation. ISM9182A showed no obvious toxicity towards human T cell, B cells, and monocytes without CD3/CD28 co-stimulation. In vivo PK data revealed favorable ADME properties and good oral bioavailability of ISM9182A across different species. In a CT-26 murine syngeneic mouse model, oral administration of ISM9182A exhibited potent antitumor effects with p-SLP76 inhibition in both murine spleen and tumor, without body weight loss. In summary, ISM9182A is a potent and selective HPK1 inhibitor and a promising immuno-oncology agent. Citation Format: Celia X.-J. Chen, Congying Pu, Jingjing Peng, Xiaoyu Ding, Hongfu Lu, Xiao Ding, Man Zhang, Supriya Bavadekar, Sujata Rao, Feng Ren, Alex Zhavoronkov. ISM9182A, a novel HPK1 inhibitor, exhibits immune modulatory activity and robust monotherapy anti-tumor effects in preclinical studies [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 3967.

COMP promotes pancreatic fibrosis by activating pancreatic stellate cells through CD36-ERK/AKT signaling pathways

BackgroundPancreatic fibrosis is one of the most important pathological features of chronic pancreatitis (CP) and pancreatic stellate cells (PSCs) are the key cells of fibrosis. As an extracellular matrix (ECM) glycoprotein, cartilage oligomeric matrix protein (COMP) is critical for collagen assembly and ECM stability and recent studies showed that COMP exert promoting fibrosis effect in the skin, lungs and liver. However, the role of COMP in activation of PSCs and pancreatic fibrosis remain unclear. We aimed to investigate the role and specific mechanisms of COMP in regulating the profibrotic phenotype of PSCs and pancreatic fibrosis. MethodsELISA method was used to determine serum COMP in patients with CP. Mice model of CP was established by repeated intraperitoneal injection of cerulein and pancreatic fibrosis was evaluated by Hematoxylin-Eosin staining (H&E) and Sirius red staining. Immunohistochemical staining was used to detect the expression changes of COMP and fibrosis marker such as α-SMA and Fibronectin in pancreatic tissue of mice. Cell Counting Kit-8, Wound Healing and Transwell assessed the proliferation and migration of human pancreatic stellate cells (HPSCs). Western blotting, qRT-PCR and immunofluorescence staining were performed to detect the expression of fibrosis marker, AKT and MAPK family proteins in HPSCs. RNA-seq omics analysis as well as small interfering RNA of COMP, recombinant human COMP (rCOMP), MEK inhibitors and PI3K inhibitors were used to study the effect and mechanism of COMP on activation of HPSCs. ResultsELISA showed that the expression of COMP significantly increased in the serum of CP patients. H&E and Sirius red staining analysis showed that there was a large amount of collagen deposition in the mice in the CP model group and high expression of COMP, α-SMA, Fibronectin and Vimentin were observed in fibrotic tissues. TGF-β1 stimulates the activation of HPSCs and increases the expression of COMP. Knockdown of COMP inhibited proliferation and migration of HPSCs. Further, RNA-seq omics analysis and validation experiments in vitro showed that rCOMP could significantly promote the proliferation and activation of HPSCs, which may be due to promoting the phosphorylation of ERK and AKT through membrane protein receptor CD36. rCOMP simultaneously increased the expression of α-SMA, Fibronectin and Collagen I in HPSCs. ConclusionIn conclusion, this study showed that COMP was up-regulated in CP fibrotic tissues and COMP induced the activation, proliferation and migration of PSCs through the CD36-ERK/AKT signaling pathway. COMP may be a potential therapeutic candidate for the treatment of CP. Interfering with the expression of COMP or the communication between COMP and CD36 on PSCs may be the next direction for therapeutic research.

Genome-wide identification of MAPK family in papaya (Carica papaya) and their involvement in fruit postharvest ripening

BackgroundPapaya (Carica papaya) is an economically important fruit cultivated in the tropical and subtropical regions of China. However, the rapid softening rate after postharvest leads to a short shelf-life and considerable economic losses. Accordingly, understanding the mechanisms underlying fruit postharvest softening will be a reasonable way to maintain fruit quality and extend its shelf-life.ResultsMitogen-activated protein kinases (MAPKs) are conserved and play essential roles in response to biotic and abiotic stresses. However, the MAPK family remain poorly studied in papaya. Here, a total of nine putative CpMAPK members were identified within papaya genome, and a comprehensive genome-wide characterization of the CpMAPKs was performed, including evolutionary relationships, conserved domains, gene structures, chromosomal locations, cis-regulatory elements and expression profiles in response to phytohormone and antioxidant organic compound treatments during fruit postharvest ripening. Our findings showed that nearly all CpMAPKs harbored the conserved P-loop, C-loop and activation loop domains. Phylogenetic analysis showed that CpMAPK members could be categorized into four groups (A-D), with the members within the same groups displaying high similarity in protein domains and intron–exon organizations. Moreover, a number of cis-acting elements related to hormone signaling, circadian rhythm, or low-temperature stresses were identified in the promoters of CpMAPKs. Notably, gene expression profiles demonstrated that CpMAPKs exhibited various responses to 2-chloroethylphosphonic acid (ethephon), 1-methylcyclopropene (1-MCP) and the combined ascorbic acid (AsA) and chitosan (CTS) treatments during papaya postharvest ripening. Among them, both CpMAPK9 and CpMAPK20 displayed significant induction in papaya flesh by ethephon treatment, and were pronounced inhibition after AsA and CTS treatments at 16 d compared to those of natural ripening control, suggesting that they potentially involve in fruit postharvest ripening through ethylene signaling pathway or modulating cell wall metabolism.ConclusionThis study will provide some valuable insights into future functional characterization of CpMAPKs, and hold great potential for further understanding the molecular mechanisms underlying papaya fruit postharvest ripening.

Proteomic and phosphoproteomic profiling of urinary small extracellular vesicles in hepatocellular carcinoma.

Hepatocellular carcinoma (HCC) is the most prevalent form of primary liver cancer and a major cause of cancer-related mortality worldwide. Small extracellular vesicles (sEVs) are heterogeneous populations of membrane-structured vesicles that can be found in many biological fluids and are currently considered as a potential source of disease-associated biomarkers for diagnosis. The purpose of this study was to define the proteomic and phosphoproteomic landscape of urinary sEVs in patients with HCC. Mass spectrometry-based methods were used to detect the global proteome and phosphoproteome profiles of sEVs isolated by differential ultracentrifugation. Label-free quantitation analysis showed that 348 differentially expressed proteins (DEPs) and 548 differentially expressed phosphoproteins (DEPPs) were identified in the HCC group. Among them, multiple phosphoproteins related to HCC, including HSP90AA1, IQGAP1, MTOR, and PRKCA, were shown to be upregulated in the HCC group. Pathway enrichment analysis indicated that the upregulated DEPPs participate in the regulation of autophagy, proteoglycans in cancer, and the MAPK/mTOR/Rap1 signaling pathway. Furthermore, kinase-substrate enrichment analysis revealed activation of MTOR, AKT1, MAP2Ks, and MAPKs family kinases in HCC-derived sEVs, indicating that dysregulation of the MAPK and mTOR signaling pathways may be the primary sEV-mediated molecular mechanisms involved in the development and progression of HCC. This study demonstrated that urinary sEVs are enriched in proteomic and phosphoproteomic signatures that could be further explored for their potential use in early HCC diagnostic and therapeutic applications.

Hematopoietic Progenitor Kinase 1 Inhibitors

Cancer, a general name for wide range of diseases that have the ability to make an impact on any parts of the human body and has been a fatal health issue that affects people around the world. Hematopoietic Progenitor Kinase 1 (HPK1), one member from the MAP4K family, is a kind of serine/threonine kinase. Destruction of HPK1 kinase raises the secretion of cytokine and T cell signaling, having positive impact on clearance of virus, and inhibition of tumor growth. Thus, HPK1 has been seen as a promising target that can be used in drug development for immunotherapy of tumors. HPK1 has a pivotal role of inhibition on the signaling pathway of the T-cell receptor, it also inhibits the proliferation and differentiation of B cells, DC cells, and NK cells. In this paper, we aimed to supply a detailed overview of research including the introduction of HPK1, research process on HPK1 inhibitors and different methods of developing new kinase inhibitors.

Mir-142-3P regulates MAPK protein family by inhibiting 14-3-3η to enhance bone marrow mesenchymal stem cells osteogenesis

Clinical studies have found 14-3-3η to be associated with osteoporosis through undefined mechanisms. We aimed to investigate the role of 14-3-3η in osteoporosis and its potential associations with miRNAs. The Gene Expression Omnibus(GEO) and Human Protein Atlas 1 databases were analyzed to examine both the mRNA and protein expression of 14-3-3η in OP. Gene enrichment analyses were performed to explore the underlying mechanism of 14-3-3η based on DAVID. miRWalk was used to predict the associated miRNAs. The statistics were analysed by R software and SPSS software. 14-3-3η was overexpressed and knock down expressed in BMSCs by lentiviral vector transfecting. And BMSCs were induced by hypoxia. qRT-PCR and Western-Blot verified the expression of mRNA and protein. Scratch assay detected the migration of osteocytes. Co-immunoprecipitation and luciferase assay studied the 14-3-3η targeted protein and miRNA. overexpression and knock down of miRNA to verify the relationship of 14-3-3η and target genes. The 14-3-3η mRNA expression level was low in patients with osteoporosis, as corroborated by immunohistochemical staining images. Functional analyses revealed enrichment of the MAPK-associated cascade. 14-3-3η was correlated with MAPK family proteins and five key miRNAs, including mir-142-3p. In addition, 14-3-3η knockdown in BMSCs increased the mRNA and protein expression levels of Hif-α, VEGF, BMP-2, OPN, OST, and Runx2, and enhanced the cells migration ability. Under hypoxic conditions, Hif-α and BMP-2 protein expression levels were upregulated, whereas those of 14-3-3η and MAPK3 were downregulated. Co-immunoprecipitation experiments showed decreased binding of 14-3-3η to MAPK3. 14-3-3η knockdown produced the same results as hypoxia induction. Adding caspase3 inhibitor and knocking down 14-3-3η again prevented MAPK3 cleavage by caspase3 and inhibited BMP-2 expression. Moreover, under hypoxic conditions, miR-142-3P expression was upregulated and luciferase assays revealed 14-3-3η as its target gene. miR-142-3P overexpression decreased mRNA and protein levels of 14-3-3η and MAPK3, while increasing BMP-2 expression. miR-142-3P knockdown reversed these results. BMSC osteogenesis was suppressed by 14-3-3η, whereas miRNA-142-3p promoted it through the inhibition of 14-3-3η.

Targeting RBM39-MEK5 Axis Synergizes with Bortezomib to Inhibit Multiple Myeloma Malignancy

Multiple myeloma (MM) is a type of haematological malignancy characterised by the accumulation and expansion of malignant plasma cells in the bone marrow (BM). Despite recent advances in chemotherapy, MM remains an incurable disease for most patients and therefore requires novel therapies. Abnormal alternative splicing is a hallmark of cancer and is associated with poor survival and an ultra-high-risk population in MM (Bauer et. al. Haematologica 2021). Targeting splicing is a promising therapeutic strategy in MM. Our previous study showed that RBM39 is highly expressed in primary MM cells and cell lines and correlates with poor overall survival (Tong et. al. Haematologica 2020). To investigate the underlying role of RBM39 in MM, we knocked down RBM39 in MM cell lines. We found that knockdown of RBM39 inhibited MM cell proliferation, induced apoptosis and cell cycle arrest at the G2/M phase, and inhibited subcutaneous tumour growth. Indisulam has been reported to induce RBM39 degradation via the proteasomal pathway by acting as a molecular glue to link RBM39 to the E3 ubiquitin ligase DCAF15 (Han et. al. Science 2017). In our study, Indisulam treatment in MM cells was shown to degrade RBM39 via DCAF15. Indisulam could induce apoptosis and G2/M phase arrest in MM cell lines and inhibite tumour growth in vivo. It also significantly decreased MM cell viability in a co-culture system with MM BM-derived mesenchymal stromal cells. Indisulam was also able to inhibit primary CD138+ cells isolated from newly diagnosed MM patients. To explore the mechanistic role of RBM39 in promoting MM malignancy, we performed RNA immunoprecipitation (RIP)-Seq and RNA sequencing in RBM39 knockdown MM cells to investigate the alternative splicing targets of RBM39. KEGG pathway analysis of differentially expressed and alternatively spliced genes revealed the enrichment of the MAPK pathway. We then identified MEK5, a non-redundant member of the MAPK family, as a potential target of RBM39, which is highly expressed in MM. Full-length MEK5 contributes to increased proliferation and survival of MM cells. We further demonstrated that MEK5 is bound and alternatively spliced by RBM39 and that splicing of full-length MEK5 is dependent on RBM39. Given the importance of the RBM39-MEK5 axis in MM, we also found that inhibition of RBM39 with Indisulam or MEK5 with the inhibitor BIX02189 increased the cytotoxicity of bortezomib in vivo and in vitro. In conclusion, we have demonstrated that the RBM39-MEK5 axis, in which RBM39 maintains splicing of full-length MEK5, is essential for MM cell survival and proliferation. Mechanistically, shRNA-mediated knockdown or Indisulam-mediated degradation of RBM39 caused extensive altered splicing, including mis-splicing of MEK5. Targeting RBM39 or MEK5 could not only inhibit MM malignancy, but also enhance the cytotoxicity of bortezomib for MM.

Ginsenoside Rg1 promotes β‑amyloid peptide degradation through inhibition of the ERK/PPARγ phosphorylation pathway in an Alzheimer's disease neuronal model.

β-Amyloid peptide (Aβ) deposition in the brain is an important pathological change in Alzheimer's disease (AD). Insulin-degrading enzyme (IDE), which is regulated transcriptionally by peroxisome proliferator-activated receptor γ (PPARγ), is able to proteolyze Aβ. One of the members of the MAPK family, ERK, is able to mediate the phosphorylation of PPARγ at Ser112, thereby inhibiting its transcriptional activity. Ginsenoside Rg1 is one of the active ingredients in the natural medicine ginseng and has inhibitory effects on Aβ production. The present study was designed to investigate whether ginsenoside Rg1 is able to affect the regulation of PPARγ based on the expression of its target gene, IDE, and whether it is able to promote Aβ degradation via inhibition of the ERK/PPARγ phosphorylation pathway. In the present study, primary cultured rat hippocampal neurons were treated with Aβ1-42, ginsenoside Rg1 and the ERK inhibitor PD98059, and subsequently TUNEL staining was used to detect the level of neuronal apoptosis. ELISA was subsequently employed to detect the intra- and extracellular Aβ1-42 levels, immunofluorescence staining and western blotting were used to detect the translocation of ERK from the cytoplasm to the nucleus, immunofluorescence double staining was used to detect the co-expression of ERK and PPARγ, and finally, western blotting was used to detect the phosphorylation of PPARγ at Ser112 and IDE expression. The results demonstrated that ginsenoside Rg1 or PD98059 were able to inhibit primary cultured hippocampal neuron apoptosis induced by Aβ1-42 treatment, reduce the levels of intra- and extraneuronal Aβ1-42 and inhibit the translocation of ERK from the cytoplasm to the nucleus. Furthermore, administration of ginsenoside Rg1 or PD98059 resulted in attenuated co-expression of ERK and PPARγ, inhibition of phosphorylation of PPARγ at Ser112 mediated by ERK and an increase in IDE expression. In addition, the effects when PD98059 to inhibit ERK followed by treatment with ginsenoside Rg1 were found to be more pronounced than those when using PD98059 alone. In conclusion, ginsenoside Rg1 was demonstrated to exert neuroprotective effects on AD via inhibition of the ERK/PPARγ phosphorylation pathway, which led to an increase in IDE expression, the promotion of Aβ degradation and the decrease of neuronal apoptosis. These results could provide a theoretical basis for the clinical application of ginsenoside Rg1 in AD.

Identification of mitogen-activated protein kinase 7 inhibitors from natural products: Combined virtual screening and dynamic simulation studies.

Mitogen-activated protein kinase 7 (MAPK7) is a serine/threonine protein kinase that belongs to the MAPK family and plays a vital role in various cellular processes such as cell proliferation, differentiation, gene transcription, apoptosis, metabolism, and cell survival. The elevated expression of MAPK7 has been associated with the onset and progression of multiple aggressive tumors in humans, underscoring the potential of targeting MAPK7 pathways in therapeutic research. This pursuit holds promise for the advancement of anticancer drug development by developing potential MAPK7 inhibitors. To look for potential MAPK7 inhibitors, we exploited structure-based virtual screening of natural products from the ZINC database. First, the Lipinski rule of five criteria was used to filter a large library of ~90,000 natural compounds, followed by ADMET and pan-assay interference compounds (PAINS) filters. Then, top hits were chosen based on their strong binding affinity as determined by molecular docking. Further, interaction analysis was performed to find effective and specific compounds that can precisely bind to the binding pocket of MAPK7. Consequently, two compounds, ZINC12296700 and ZINC02123081, exhibited significant binding affinity and demonstrated excellent drug-like properties. All-atom molecular dynamics simulations for 200 ns confirmed the stability of MAPK7-ZINC12296700 and MAPK7-ZINC02123081 docked complexes. According to the molecular mechanics Poisson-Boltzmann surface area investigation, the binding affinities of both complexes were considerable. Overall, the result suggests that ZINC12296700 and ZINC02123081 might be used as promising leads to develop novel MAPK7 inhibitors. Since these compounds would interfere with the kinase activity of MAPK7, therefore, may be implemented to control cell growth and proliferation in cancer after required validations.