Mitogen-Activated Protein Kinase 15 Is a New Predictive Biomarker and Potential Therapeutic Target for Ovarian Cancer.

Mitogen-activated protein kinase 15 (MAPK15) has been reported to be associated with several cancers. This study aimed to explore for the first time on the relationship between MAPK15 expression and cancer progression/drug responsiveness in ovarian carcinoma. To this end, MAPK15 expression level was examined by immunohistochemistry (IHC) staining of an ovarian tissue array (10 normal and 70 malignant samples). Drug sensitivity of ovarian cancer cell lines (including OVCAR3 and SKOV3) was measured by MTS assay. The modulation of MAPK15 expression in OVCAR3 and SKOV3 was verified by immunoblot and real-time PCR analyses. The prognostic value of MAPK15 in ovarian cancer patients was assessed using the Kaplan-Meier Plotter database and Gene Expression Omnibus (GEO) datasets. The IHC results showed that MAPK15 expression was negatively associated with tumor grade, TNM stage, tumor size, and regional lymph node metastasis of ovarian carcinoma. Importantly, overexpressing MAPK15 increased cisplatin toxicity in ovarian carcinoma cells and online database analysis indicated that patients with high MAPK15 expression had favorable prognosis with/without chemotherapy. Taken together, our results indicate that a decreased MAPK15 expression is associated with advanced-stage ovarian cancer and unfavorable survival outcomes. MAPK15 may be a new biomarker for ovarian cancer, and the encouraging therapeutic strategy would be found by combining the regulation of MAPK15 expression.

Regulation of human mitogen-activated protein kinase 15 (extracellular signal-regulated kinase 7/8) and its functions: A recent update.

Mitogen-activated protein kinase 15 (MAPK15), originally also known as extracellular signal-regulated kinase 7/8, is the most recently identified atypical MAPK and the least studied so far. Examinations of the role of MAPK15 in various cell lines and model systems indicate that MAPK15 participates in a variety of cellular activities such as promoting cell proliferation, cell transformation, and apoptosis; stimulating autophagy; regulating cell division, ciliogenesis, and protein secretion; and maintaining genome stability. As multiple roles of MAPK15 were observed among these studies, therefore, it remains unclear whether MAPK15 acts as a proto-oncogene or tumor suppressor. Here, the recent literature on human MAPK15 and the resulting functions will be discussed.

Atypical MAPKs in cancer.

Impaired kinase signalling leads to various diseases, including cancer. At the same time, kinases make up the majority of the druggable genome and targeting kinase activity has proven to be a successful first-line therapy for many cancers. Among the best-studied kinases are the mitogen-activated protein kinases (MAPKs), which regulate cell proliferation, differentiation, motility, and survival. However, the MAPK family also contains the atypical members ERK3 (MAPK6), ERK4 (MAPK4), ERK7/ERK8 (MAPK15), and NLK that are functionally and structurally different from their conventional family members and have long been neglected. Nevertheless, in recent years, important roles in carcinogenesis, actin cytoskeleton regulation and the immune system have been discovered, underlining the physiological importance of atypical MAPKs and the need to better understand their functions. This review highlights the distinctive features of the atypical MAPKs and summarizes the evidence on their regulation, physiological roles, and potential targeting strategies for cancer therapies.

The Atypical MAP Kinase MAPK15 Is Required for Lung Adenocarcinoma Metastasis via Its Interaction with NF-κB p50 Subunit and Transcriptional Regulation of Prostaglandin E2 Receptor EP3 Subtype.

Studying the relatively underexplored atypical MAP Kinase MAPK15 on cancer progression/patient outcomes and its potential transcriptional regulation of downstream genes would be highly valuable for the diagnosis, prognosis, and potential oncotherapy of malignant tumors such as lung adenocarcinoma (LUAD). Here, the expression of MAPK15 in LUAD was detected by immunohistochemistry and its correlation with clinical parameters such as lymph node metastasis and clinical stage was analyzed. The correlation between the prostaglandin E2 receptor EP3 subtype (EP3) and MAPK15 expression in LUAD tissues was examined, and the transcriptional regulation of EP3 and cell migration by MAPK15 in LUAD cell lines were studied using the luciferase reporter assay, immunoblot analysis, qRT-PCR, and transwell assay. We found that MAPK15 is highly expressed in LUAD with lymph node metastasis. In addition, EP3 is positively correlated with the expression of MAPK15 in LUAD tissues, and we confirmed that MAPK15 transcriptionally regulates the expression of EP3. Upon the knockdown of MAPK15, the expression of EP3 was down-regulated and the cell migration ability was decreased in vitro; similarly, the mesenteric metastasis ability of the MAPK15 knockdown cells was inhibited in in vivo animal experiments. Mechanistically, we demonstrate for the first time that MAPK15 interacts with NF-κB p50 and enters the nucleus, and NF-κB p50 binds to the EP3 promoter and transcriptionally regulates the expression of EP3. Taken together, we show that a novel atypical MAPK and NF-κB subunit interaction promotes LUAD cell migration through transcriptional regulation of EP3, and higher MAPK15 level is associated with lymph node metastasis in patients with LUAD.

MAPK15-ULK1 signaling regulates mitophagy of airway epithelial cell in chronic obstructive pulmonary disease.

Airway epithelial mitochondrial oxidative stress and damage is an important pathology in chronic obstructive pulmonary disease (COPD). Mitophagy involves MAPK15-ULK1 signaling, the role of which is unknown in COPD. This study investigated MAPK15-ULK1 signaling in airway epithelial cells of COPD patients and its activation by cigarette smoke extract (CSE) in isolated human airway epithelial cells. Significant increased phosphorylation of MAPK15 and ULK1 (Ser555) was detected in the airway epithelium of COPD patients. This pathology was maintained in isolated primary COPD-epithelial cells. Compared to control cells, the protein expression of Beclin1 and the ratio of LC3II to LC3I were both significantly increased in COPD-epithelial cells. In human airway epithelial cells, CSE significantly increased the phosphorylation of MAPK15, ULK1 (Ser555), the expression of Beclin1, and the LC3II/LC3I ratio in a concentration- and time-dependent manner. Transfection with MAPK15 siRNA significantly inhibited the CSE-induced ULK1 (Ser555) phosphorylation in airway epithelial cells. Silencing of MAPK15 or ULK1 significantly reduced CSE-induced mitophagy and mitochondrial oxidative stress, thereby improving cell viability. In summary, cigarette smoke activated MAPK15-ULK1 signaling, thereby promoting mitophagy and mitochondrial oxidative stress in airway epithelial cells. This signaling pathway is activated in COPD-epithelial cells and therefore might present a novel therapeutic target for COPD.

IGF1R and MAPK15 Emerge as Potential Targets of Pentabromobenzylisothioureas in Lung Neuroendocrine Neoplasms.

Pentabromobenzylisothioureas are antitumor agents with diverse properties, including the inhibition of MAPK15, IGF1R and PKD1 kinases. Their dysregulation has been implicated in the pathogenesis of several cancers, including bronchopulmonary neuroendocrine neoplasms (BP-NEN). The present study assesses the antitumor potential of ZKKs, a series of pentabromobenzylisothioureas, on the growth of the lung carcinoid H727 cell line. It also evaluates the expression of MAPK15, IGF1R and PKD1 kinases in different BP-NENs. The viability of the H727 cell line was assessed by colorimetric MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-tetrazolium bromide) and its proliferation by BrdU (5-bromo-2'-deoxyuridine) assay. Tissue kinase expression was measured using TaqMan-based RT-PCR and immunohistochemistry. ZKKs (10-4 to 10-5 M) strongly inhibited H727 cell viability and proliferation and their antineoplastic effects correlated with their concentrations (p < 0.001). IGF1R and MAPK15 were expressed at high levels in all subtypes of BP-NENs. In addition, the SCLC (small cell lung carcinoma) patients demonstrated higher mRNA levels of IGF1R (p = 0.010) and MAPK15 (p = 0.040) than the other BP-NEN groups. BP-NENs were characterized by low PKD1 expression, and lung neuroendocrine cancers demonstrated lower PKD1 mRNA levels than carcinoids (p = 0.003). ZKKs may suppress BP-NEN growth by inhibiting protein kinase activity. Our results suggest also a possible link between high IGF1R and MAPK15 expression and the aggressive phenotype of BP-NEN tumors.

Quantitative Proteomic Analysis Identifies MAPK15 as a Potential Regulator of Radioresistance in Nasopharyngeal Carcinoma Cells.

Since resistance to radiotherapy remains refractory for the clinical management of nasopharyngeal cancer (NPC), further understanding the mechanisms of radioresistance is necessary in order to develop more effective NPC treatment and improve prognosis. In this study, an integrated quantitative proteomic approach involving tandem mass tag labeling and liquid chromatograph-mass spectrometer was used to identify proteins potentially responsible for the radioresistance of NPC. The differential radiosensitivity in NPC model cells was examined through clonogenic survival assay, CCK-8 viability assay, and BrdU incorporation analysis. Apoptosis of NPC cells after exposure to irradiation was detected using caspase-3 colorimetric assay. Intracellular reactive oxygen species (ROS) was detected by a dichlorofluorescin diacetate fluorescent probe. In total, 5,946 protein groups were identified, among which 5,185 proteins were quantified. KEGG pathway analysis and protein-protein interaction enrichment analysis revealed robust activation of multiple biological processes/pathways in radioresistant CNE2-IR cells. Knockdown of MAPK15, one up-regulated protein kinase in CNE2-IR cells, significantly impaired clonogenic survival, decreased cell viability and increased cell apoptosis following exposure to irradiation, while over-expression of MAPK15 promoted cell survival, induced radioresistance and reduced apoptosis in NPC cell lines CNE1, CNE2, and HONE1. MAPK15 might regulate radioresistance through attenuating ROS accumulation and promoting DNA damage repair after exposure to irradiation in NPC cells. Quantitative proteomic analysis revealed enormous metabolic processes/signaling networks were potentially involved in the radioresistance of NPC cells. MAPK15 might be a novel potential regulator of radioresistance in NPC cells, and targeting MAPK15 might be useful in sensitizing NPC cells to radiotherapy.

Identification and functional analysis of a stress-responsive MAPK15 in Entamoeba invadens.

E. histolytica, a protozoan parasite is the causative agent of amoebiasis in human beings. It exists in two different forms - the motile trophozoite form which undergoes encystation under starvation conditions to form the non-motile, osmotically resistant cyst form. Cellular stresses stimulate several signaling cascades which assist the parasite in counter-attacking such conditions thereby, promoting cell survival. To study the stress-associated pathways activated during encystation, we have used Entamoeba invadens, a reptilian parasite as a model organism because of its ability to undergo encystation under in vitro conditions. In this study, we have identified a stress-responsive MAPK which gets upregulated under different stress conditions, including encystation. Sequence analysis and phylogenetic classification show that the MAPK belongs to the atypical MAPK15 family (henceforth, named EiMAPK15), which does not require an upstream MAPKK for its phosphorylation and activation. The in vitro kinase activity of recombinant EiMAPK15 exhibits its auto-phosphorylation ability. Immunolocalization studies reveal that the protein is mainly cytosolic under normal growing conditions but gets translocated into the nucleus under stress conditions. Knockdown of EiMAPK15 using double-stranded RNA was found to reduce the expression of other encystation-specific genes which in turn, resulted in the decline of the overall encystation efficiency of the cells. Overall, the present work has laid the platform for further characterization of this important MAPK gene in Entamoeba invadens.

Primary Cilium Formation and Ciliary Protein Trafficking Is Regulated by the Atypical MAP Kinase MAPK15 in Caenorhabditis elegans and Human Cells.

Motile and immotile (or primary) cilia are microtubule-based structures that mediate multiple cellular functions, including the transduction of environmental cues, developmental signaling, cellular motility, and modulation of fluid flow. Although their core architectures are similar, motile and primary cilia exhibit marked structural differences that underlie distinct functional properties. However, the extent to which ciliogenesis mechanisms are shared between these different cilia types is not fully described. Here, we report that the atypical MAP kinase MAPK15 (ERK7/8), implicated in the formation of vertebrate motile cilia, also regulates the formation of primary cilia in Caenorhabditis elegans sensory neurons and human cells. We find that MAPK15 localizes to a basal body subdomain with the ciliopathy protein BBS7 and to cell-cell junctions. MAPK15 also regulates the localization of ciliary proteins involved in cilium structure, transport, and signaling. Our results describe a primary cilia-related role for this poorly studied member of the MAPK family in vivo, and indicate a broad requirement for MAPK15 in the formation of multiple ciliary classes across species.

Extracellular signal-regulated kinase 8-mediated NF-κB activation increases sensitivity of human lung cancer cells to arsenic trioxide.

Extracellular signal-regulated kinase 8 (ERK8), also known as mitogen-activated protein kinase 15 (MAPK15), is the most recently identified protein kinase of the ERK family members and yet the least has been studied so far. Here, we report that ERK8 is highly expressed in several human lung cancer cell lines and is positively correlated with their sensitivities to the anti-cancer drug arsenic trioxide (As2O3). As2O3 at physiologically relevant concentrations (5-20 µM) potently stimulates the phosphorylation of ERK8 at Thr175 and Tyr177 within the TEY motif in the kinase domain, leading to its activation. Interestingly, activated ERK8 interacts and directly phosphorylates IkappaBalpha (IκBα) at Ser32 and Ser36, resulting in IκBα degradation. This in turn promotes nuclear factor-kappaB (NF-κB) p65 nuclear translocation and chromatin-binding, as well as the subsequent induction and activation of proteins involved in apoptosis. We also show that stable short-hairpin RNA-specific knockdown of endogenous ERK8 or inhibition of NF-κB activity by NF-κB inhibitor in high ERK8 expressing lung cancer H1299 cells blunted the As2O3-induced NF-κB activation and cytotoxicity towards these cells, indicating the critical role of ERK8 and NF-κB in mediating the As2O3 effects. Taken together, our findings suggest for the first time a regulatory paradigm of NF-κB activation by ERK8 upon As2O3 treatment in human lung cancer cells; and implicate a potential therapeutic advantage of As2O3 that might gain more selective killing of cancer cells with high ERK8 expression.

Overexpression of MAPK15 in gastric cancer is associated with copy number gain and contributes to the stability of c-Jun.

This study was aimed at understanding the functional and clinicopathological significance of MAPK15 alteration in gastric cancer. Genome-wide copy number alterations (CNAs) were first investigated in 40 gastric cancers using Agilent aCGH-244K or aCGH-400K, and copy number gains of MAPK15 found in aCGH were validated in another set of 48 gastric cancer tissues. The expression of MAPK15 was analyzed using immunohistochemistry in concurrent lesions of normal, adenoma, and carcinoma from additional 45 gastric cancer patients. The effects of MAPK15 on cell cycle, c-Jun phosphorylation, and mRNA stability were analyzed in gastric cancer cells. Copy number gains of MAPK15 were found in 15 (17%) of 88 tumor tissues. The mRNA levels of MAPK15 were relatively high in the gastric cancer tissues and gastric cancer cells with higher copy number gains than those without. Knockdown of MAPK15 using siRNA in gastric cancer cells significantly suppressed cell proliferation and resulted in cell cycle arrest at G1-S phase. Reduced c-Jun phosphorylation and c-Jun half-life were observed in MAPK15-knockdowned cells. In addition, transient transfection of MAPK15 into AGS gastric cancer cells with low copy number resulted in an increase of c-Jun phosphorylation and stability. The overexpression of MAPK15 occurred at a high frequency in carcinomas (37%) compared to concurrent normal tissues (2%) and adenomas (21%). In conclusion, the present study suggests that MAPK15 overexpression may contribute to the malignant transformation of gastric mucosa by prolonging the stability of c-Jun. And, patients with copy number gain of MAPK15 in normal or premalignant tissues of stomach may have a chance to progress to invasive cancer.