RBM25 depletion suppresses the growth of colon cancer cells through regulating alternative splicing of MNK2.

Increasing evidence suggests that deregulated RNA splicing factors play critical roles in tumorigenesis; however, their specific involvement in colon cancer remains largely unknown. Here we report that the splicing factor RBM25 is overexpressed in colon cancer, and this increased expression correlates with a poor prognosis of patients with colon cancer. Functionally, RBM25 ablation suppresses the growth of colon cancer cells both in vitro and in vivo. Mechanistically, our transcriptome-wide analysis of splicing events revealed that RBM25 regulates a large number of cancer-related alternative splicing events across the human genome in colon cancer. Particularly, RBM25 regulates the splicing of MNK2 by interacting with the poly G rich region in exon 14a, thereby inhibiting the selection of the proximal 3' splice site (ss), resulting in the production of the oncogenic short isoform, MNK2b. Knockdown of RBM25 leads to an increase in the MNK2a isoform and a decrease in the MNK2b isoform. Importantly, re-expression of MNK2b or blocking the 3' ss of the alternative exon 14a with ASO partially reverses the RBM25 knockdown mediated tumor suppression. Moreover, MNK2b levels were significantly increased in colon cancer tissues, which is positively correlated with the expression level of RBM25. Collectively, our findings uncover the critical role of RBM25 as a key splicing factor in colon cancer, suggesting its potential as a prognostic marker and therapeutic target.

Selective and effective suppression of pancreatic cancer through MNK inhibition.

OBJECTIVE: The study aimed to explore the role of the Wnt/ß-catenin signaling pathway in pancreatic cancer progression and chemoresistance, with a focus on identifying specific factors that distinguish between normal and tumor cells, thereby offering potential therapeutic targets. MATERIALS AND METHODS: We analyzed levels of total and phosphorylated eukaryotic translation initiation factor 4E (eIF4E) and ß-catenin in pancreatic cancer and normal pancreatic tissues. Functional assays were used to assess the impact of eIF4E phosphorylation on ß-catenin signaling, cell proliferation, and chemoresistance, with MNK kinase involvement determined through gene depletion studies. The MNK kinase inhibitor eFT508 was evaluated for its effects on eIF4E phosphorylation, ß-catenin activation, and cell viability in both in vitro and in vivo models of pancreatic cancer. RESULTS: Both total and phosphorylated eIF4E, along with ß-catenin, were significantly elevated in pancreatic cancer tissues compared to normal tissues. Phosphorylation of eIF4E at serine 209 was shown to activate ß-catenin signaling, enhance cell proliferation, and contribute to chemoresistance in pancreatic cancer. Importantly, these effects were dependent on MNK kinase activity. Depletion of eIF4E reduced cell viability in both pancreatic cancer and normal cells, while depletion of MNK selectively decreased viability in pancreatic cancer cells. Treatment with eFT508 effectively inhibited eIF4E phosphorylation, suppressed ß-catenin activation, and reduced pancreatic cancer cell growth and survival in vitro and in vivo, with minimal impact on normal cells.Conclusions: The MNK-eIF4E-ß-catenin axis plays a critical role in pancreatic cancer progression and chemoresistance, distinguishing pancreatic cancer cells from normal cells. Targeting MNK kinases with inhibitors like eFT508 presents a promising therapeutic strategy for pancreatic cancer, with potential for selective efficacy and reduced toxicity.

Interleukin-6 induces nascent protein synthesis in human dorsal root ganglion nociceptors primarily via MNK-eIF4E signaling.

Plasticity of dorsal root ganglion (DRG) nociceptors in the peripheral nervous system requires new protein synthesis. This plasticity is believed to be responsible for the physiological changes seen in DRG nociceptors in animal models of chronic pain. Experiments in human DRG (hDRG) neurons also support this hypothesis, but a direct observation of nascent protein synthesis in response to a pain promoting substance, like interleukin-6 (IL-6), has not been measured in these neurons. To fill this gap in knowledge, we used acutely prepared human DRG explants from organ donors. These explants provide a physiologically relevant microenvironment, closer to in vivo conditions, allowing for the examination of functional alterations in DRG neurons reflective of human neuropathophysiology. Using this newly developed assay, we demonstrate upregulation of the target of the MNK1/2 kinases, phosphorylated eIF4E (p-eIF4E), and nascently synthesized proteins in a substantial subset of hDRG neurons following exposure to IL-6. To pinpoint the specific molecular mechanisms driving this IL-6-driven increase in nascent proteins, we used the specific MNK1/2 inhibitor eFT508. Treatment with eFT508 resulted in the inhibition of IL-6-induced increases in p-eIF4E and nascent proteins. Additionally, using TRPV1 as a marker for nociceptors, we found that these effects occurred in a large number of human nociceptors. Our findings provide clear evidence that IL-6 drives nascent protein synthesis in human TRPV1+ nociceptors primarily via MNK1/2-eIF4E signaling. The work links animal findings to human nociception, creates a framework for additional hDRG signaling experiments, and substantiates the continued development of MNK inhibitors for pain.

Structural insights into trypanosomatid Mnk kinase orthologues (kMnks) suggest altered mechanism in the kinase domain.

Mitogen-activated protein kinase (MAPK) interacting protein kinases (Mnk1 and Mnk2) mediated phosphorylation of the eukaryotic initiation factor eIF4E is an important translation initiation control, in Mnk-mediated oncogenic activity and other disease conditions. Thus, Mnk kinases are an important target for therapy. Trypanosomatids are a class of kinetoplastids, some of which are protozoan parasites and cause diseases in humans. While protein translation initiation is well understood in eukaryotes and prokaryotes, there is a lack of sufficient structural information of this process in trypanosomatids. Here, we report that trypanosomatids have one orthologue of Mnk kinase with low overall sequence homology but high homology in the kinase domain and an additional C-terminal domain containing putative calmodulin binding site(s). We show that while many of the domains and motifs are conserved, homology modeling/structure prediction, docking analysis and molecular dynamics simulation studies suggest that trypanosomatid kMnk kinases, kinase domains are present in DFG-in conformation as opposed to the auto-inhibited DFD-out conformation of un-phosphorylated human Mnk1. Furthermore, we observed that several regulatory features are different in trypanosomatid kMnk kinases. Our study indicates that mechanism and regulation in the kinase domain of trypanosomatid kMnks are likely to be altered, and that they can be important drug targets.

The role of MNK1-mTORC1 pathway in modulating macrophage responses to Vibrio vulnificus infection.

Vibrio vulnificus (Vv) is known to cause life-threatening infections, particularly septicemia. These patients often exhibit elevated levels of pro-inflammatory cytokines. While it is established that mitogen-activated protein kinase (MAPK)-interacting kinase (MNK) contributes to the production of pro-inflammatory cytokines, the role of MNK in macrophages during Vv infection remains unclear. In this study, we investigate the impact of MNK on macrophages. We demonstrate that the inhibition of MNK in J774A.1 cells, when treated with lipopolysaccharide or Vv, resulted in decreased production of tumor necrosis factor alpha and interleukin-6, without affecting their transcription. Interestingly, treatment with MNK inhibitor CGP57380 led to enhanced phosphorylation of MNK1 but decreased phosphorylation of eIF4E. Moreover, MNK1 knockout cells exhibited an increased capacity for phagocytosis and clearance of Vv, with more acidic phagosomes than the parental cells. Notably, CGP57380 did not impact phagocytosis, bacterial clearance, or phagosome acidification in Vv-infected J774A.1 cells. Considering the reported association between MNK and mammalian target of rapamycin complex 1 (mTORC1) activation, we investigated the mTORC1 signaling in MNK1 knockout cells infected with Vv. Our results revealed that attenuation of the mTORC1 signaling in these cells and treatment with the mTORC1 inhibitor rapamycin significantly enhanced bacterial clearance in J774A.1 cells following Vv infection. In summary, our findings suggest that MNK promotes the Vv-induced cytokine production in J774A.1 cells without affecting their transcription levels. MNK1 appears to impair the phagocytosis, bacterial clearance, and phagosome acidification in Vv-infected J774A.1 cells through the MNK1-mTORC1 signaling pathway rather than the MNK1-eIF4E signaling pathway. Our findings highlight the importance of the MNK1-mTORC1 pathway in modulating macrophage responses to Vv infection. IMPORTANCE: Mitogen-activated protein kinase (MAPK)-interacting kinase (MNK) plays a role in promoting the production of tumor necrosis factor alpha and interleukin-6 in macrophages during Vibrio vulnificus (Vv) infection. Inhibition or knockout of MNK1 in J774A.1 cells resulted in reduced cytokine production without affecting their transcription levels. MNK1 also impairs phagocytosis, bacterial clearance, and phagosome acidification in Vv-infected cells through the MNK1-mammalian target of rapamycin complex 1 (mTORC1) signaling pathway. The findings highlight the importance of the MNK1-mTORC1 pathway in modulating macrophage responses to Vv infection.

Tomivosertib reduces ectopic activity in dorsal root ganglion neurons from patients with radiculopathy.

Spontaneous activity in dorsal root ganglion (DRG) neurons is a key driver of neuropathic pain in patients suffering from this largely untreated disease. While many intracellular signalling mechanisms have been examined in preclinical models that drive spontaneous activity, none have been tested directly on spontaneously active human nociceptors. Using cultured DRG neurons recovered during thoracic vertebrectomy surgeries, we showed that inhibition of mitogen-activated protein kinase interacting kinase (MNK) with tomivosertib (eFT508, 25 nM) reversibly suppresses spontaneous activity in human sensory neurons that are likely nociceptors based on size and action potential characteristics associated with painful dermatomes within minutes of treatment. Tomivosertib treatment also decreased action potential amplitude and produced alterations in the magnitude of after hyperpolarizing currents, suggesting modification of Na+ and K+ channel activity as a consequence of drug treatment. Parallel to the effects on electrophysiology, eFT508 treatment led to a profound loss of eIF4E serine 209 phosphorylation in primary sensory neurons, a specific substrate of MNK, within 2 min of drug treatment. Our results create a compelling case for the future testing of MNK inhibitors in clinical trials for neuropathic pain.

ephrin-B2 promotes nociceptive plasticity and hyperalgesic priming through EphB2-MNK-eIF4E signaling in both mice and humans.

Ephrin-B-EphB signaling can promote pain through ligand-receptor interactions between peripheral cells, like immune cells expressing ephrin-Bs, and EphB receptors expressed by DRG neurons. Previous studies have shown increased ephrin-B2 expression in peripheral tissues like synovium of rheumatoid and osteoarthritis patients, indicating the clinical significance of this signaling. The primary goal of this study was to understand how ephrin-B2 acts on mouse and human DRG neurons, which express EphB receptors, to promote pain and nociceptor plasticity. We hypothesized that ephrin-B2 would promote nociceptor plasticity and hyperalgesic priming through MNK-eIF4E signaling, a critical mechanism for nociceptive plasticity induced by growth factors, cytokines and nerve injury. Both male and female mice developed dose-dependent mechanical hypersensitivity in response to ephrin-B2, and both sexes showed hyperalgesic priming when challenged with PGE2 injection either to the paw or the cranial dura. Acute nociceptive behaviors and hyperalgesic priming were blocked in mice lacking MNK1 (Mknk1 knockout mice) and by eFT508, a specific MNK inhibitor. Sensory neuron-specific knockout of EphB2 using Pirt-Cre demonstrated that ephrin-B2 actions require this receptor. In Ca2+-imaging experiments on cultured DRG neurons, ephrin-B2 treatment enhanced Ca2+ transients in response to PGE2 and these effects were absent in DRG neurons from MNK1-/- and EphB2-PirtCre mice. In experiments on human DRG neurons, ephrin-B2 increased eIF4E phosphorylation and enhanced Ca2+ responses to PGE2 treatment, both blocked by eFT508. We conclude that ephrin-B2 acts directly on mouse and human sensory neurons to induce nociceptor plasticity via MNK-eIF4E signaling, offering new insight into how ephrin-B signaling promotes pain.

Computational insights into dynamics and conformational stability of N-acetylmannosamine kinase mutations.

The activity of UDP-GlcNAc 2-epimerase/ManNAc kinase (GNE) is essential for the biosynthesis of sialic acid, which is involved in cellular processes in health and diseases. GNE contains an N-terminal epimerase domain and a C-terminal kinase domain (N-acetylmannosamine kinase, MNK). Mutations of the GNE protein led to hypoactivity of the enzyme and cause sialurea or autosomal recessive inclusion body myopathy/Nonaka myopathy. Here, we used all-atom molecular dynamics (MD) simulations to comprehend the folding, dynamics and conformational stability of MNK variants, including the wild type (WT) and three mutants (H677R, V696M and H677R/V696M). The deleterious and destabilizing nature of MNK mutants were predicted using different prediction tools. Results predicted that mutations modulate the stability, flexibility and function of MNK. The effect of mutations on the conformational stability and dynamics of MNK was next studied through the free-energy landscape (FEL), hydrogen-bonds and secondary structure changes. The FEL results show that the mutations interfere with various conformational transitions in both WT and mutants, exposing the structural underpinnings of protein destabilization and unfolding brought on by mutation. We discover that, when compared to the other two mutations, V696M and H677R/V696M, H677R has the most harmful effects. These findings have a strong correlation with published experimental studies that demonstrate how these mutations disrupt MNK activity. Hence, this computational study describes the structural details to unravel the mutant effects at the atomistic resolution and has implications for understanding the GNE's physiological and pathological role.Communicated by Ramaswamy H. Sarma.

Design, synthesis and biological evaluation of MNK-PROTACs.

Mitogen-activated protein kinase (MAPK)-interacting kinases (MNKs) can regulate cellular mRNA translation by controlling the phosphorylation of the eukaryotic translation initiation factor 4E (eIF4E), which plays an important role in tumor initiation, development, and metastasis. Although small-molecule MNK inhibitors have made significant breakthroughs in the treatment of various malignancies, their clinical application can be limited by drug resistance, target selectivity and other factors. The strategy of MNK-PROTACs which selectively degrades MNK kinases provides a new approach for developing small-molecule drugs for related diseases. In this study, DS33059, a small-molecule compound modified based on the ongoing clinical trials drug ETC-206, was chosen as the target protein ligand. A series of novel MNK-PROTACs were designed, synthesized and evaluated biological activity. Several compounds showed good inhibitory activities against MNK1/2. Besides, compounds exhibited moderate to excellent anti-proliferative activity in A549 and TMD-8 cells in vitro. In particular, compound II-5 significantly inhibited A549 (IC50 = 1.79 µM) and TMD-8 (IC50 = 1.07 µM) cells. The protein degradation assay showed that compound II-5 had good capability to degrade MNK1. The MNK-PROTACs strategy represents a new direction in treating tumors and deserves further exploration.

High Expression of miR-6785-5p in the Serum Exosomes of Psoriasis Patients Alleviates Psoriasis-Like Skin Damage by Interfering with the MNK2/p-eIF4E Axis in Keratinocytes.

Psoriasis is a chronic inflammatory skin disease characterized by abnormal keratinocyte proliferation and inflammation. MiRNAs and serum exosomes participate in the pathogenesis of many diseases. The objective of this study is to explore the function of miR-6785-5p in psoriatic keratinocytes and its upstream and downstream mechanisms. For our study, we employed qRT-PCR and fluorescence in situ hybridization to evaluate miR-6785-5p in psoriatic keratinocytes and conducted a microRNA microarray for identifying differentially expressed miRNAs in patient serum exosomes. We then cocultured keratinocytes with these exosomes, using immunofluorescence staining and qRT-PCR to assess uptake and miR-6785-5p overexpression. We explored miR-6785-5p's role through transfection with specific mimics and inhibitors and confirmed MNK2 as its target using a luciferase assay. MNK2's function was further examined using siRNA technology. Lastly, we applied an imiquimod-induced psoriasis mouse model, also employing siRNA, to investigate MNK2's role in psoriasis. MiR-6785-5p demonstrates a notable overexpression in the keratinocytes of psoriasis patients as well as in their serum exosomes. These keratinocytes actively uptake the miR-6785-5p-enriched serum exosomes. Functionally, miR-6785-5p appears to alleviate psoriasis-like skin damage, observable both in vitro and in vivo, by downregulating MNK2 expression. Psoriasis keratinocytes uptake serum exosomes highly expressing miR-6785-5p. MiR-6785-5p inhibits the abnormal proliferation and inflammatory state of keratinocytes by reducing MNK2 expression and interfering with the MNK2/p-eIF4E axis.

MNK: A Novel Promising Target for Cancer Immunotherapy.

Cancer immunotherapy has demonstrated remarkable success in the treatment of multiple advanced malignancies, especially approaches to target the immune checkpoint. Nonetheless, the limited response rate remains a barrier to broader application. Identifying other ways to extend the beneficiaries to a large extent is needed. Emerging evidence has shown that mitogen-activated protein kinase-interacting kinases (MNKs) could be regarded as a novel, attractive target for cancer immunotherapy that is closely correlated with cancer biology and therapies. A comprehensive understanding of the role and mechanism of MNKs in cancer will shed light on the discovery of novel therapeutic strategies for cancer treatment. In this review, we outlined the structure of MNKs, their function and expression, and how MNKs affect tumor progression and elucidated the evidence supporting MNKs as a new promising treatment modality in human cancers.

MNK, mTOR or eIF4E-selecting the best anti-tumor target for blocking translation initiation.

Overexpression of eIF4E is common in patients with various solid tumors and hematologic cancers. As a potential anti-cancer target, eIF4E has attracted extensive attention from researchers. At the same time, mTOR kinases inhibitors and MNK kinases inhibitors, which are directly related to regulation of eIF4E, have been rapidly developed. To explore the optimal anti-cancer targets among MNK, mTOR, and eIF4E, this review provides a detailed classification and description of the anti-cancer activities of promising compounds. In addition, the structures and activities of some dual-target inhibitors are briefly described. By analyzing the different characteristics of the inhibitors, it can be concluded that MNK1/2 and eIF4E/eIF4G interaction inhibitors are superior to mTOR inhibitors. Simultaneous inhibition of MNK and eIF4E/eIF4G interaction may be the most promising anti-cancer method for targeting translation initiation.

VNLG-152R and its deuterated analogs potently inhibit/repress triple/quadruple negative breast cancer of diverse racial origins in vitro and in vivo by upregulating E3 Ligase Synoviolin 1 (SYVN1) and inducing proteasomal degradation of MNK1/2.

Triple-negative breast cancer (TNBC) and its recently identified subtype, quadruple negative breast cancer (QNBC), collectively account for approximately 13% of reported breast cancer cases in the United States. These aggressive forms of breast cancer are associated with poor prognoses, limited treatment options, and lower overall survival rates. In previous studies, our research demonstrated that VNLG-152R exhibits inhibitory effects on TNBC cells both in vitro and in vivo and the deuterated analogs were more potent inhibitors of TNBC cells in vitro. Building upon these findings, our current study delves into the molecular mechanisms underlying this inhibitory action. Through transcriptome and proteome analyses, we discovered that VNLG-152R upregulates the expression of E3 ligase Synoviolin 1 (SYVN1), also called 3-hydroxy-3-methylglutaryl reductase degradation (HRD1) in TNBC cells. Moreover, we provide genetic and pharmacological evidence to demonstrate that SYVN1 mediates the ubiquitination and subsequent proteasomal degradation of MNK1/2, the only known kinases responsible for phosphorylating eIF4E. Phosphorylation of eIF4E being a rate-limiting step in the formation of the eIF4F translation initiation complex, the degradation of MNK1/2 by VNLG-152R and its analogs impedes dysregulated translation in TNBC cells, resulting in the inhibition of tumor growth. Importantly, our findings were validated in vivo using TNBC xenograft models derived from MDA-MB-231, MDA-MB-468, and MDA-MB-453 cell lines, representing different racial origins and genetic backgrounds. These xenograft models, which encompass TNBCs with varying androgen receptor (AR) expression levels, were effectively inhibited by oral administration of VNLG-152R and its deuterated analogs in NRG mice. Importantly, in direct comparison, our compounds are more effective than enzalutamide and docetaxel in achieving tumor growth inhibition/repression in the AR+ MDA-MD-453 xenograft model in mice. Collectively, our study sheds light on the involvement of SYVN1 E3 ligase in the VNLG-152R-induced degradation of MNK1/2 and the therapeutic potential of VNLG-152R and its more potent deuterated analogs as promising agents for the treatment of TNBC across diverse patient populations.

Salinization Dramatically Enhance the Anti-Prostate Cancer Efficacies of AR/AR-V7 and Mnk1/2 Molecular Glue Degraders, Galeterone and VNPP433-3ß Which Outperform Docetaxel and Enzalutamide in CRPC CWR22Rv1 Xenograft Mouse Model.

Galeterone, 3ß-(hydroxy)-17-(1H-benzimidazole-1-yl)androsta-5,16-diene (Gal, 1) and VNPP433-3ß, 3ß-(1H-imidazole-1-yl-17-(1H-benzimidazole-1-yl)androsta-5,16-diene (2) are potent molecular glue degrader modulators of AR/AR-V7 and Mnk1/2-eIF4E signaling pathways, and are promising Phase 3 and Phase 1 drug candidates, respectively. Because appropriate salts can be utilized to create new chemical entities with enhanced aqueous solubility, in vivo pharmacokinetics, and enhanced in vitro and in vivo efficacies, the monohydrochloride salt of Gal (3) and the mono- and di-hydrochlorides salts of compound 2, compounds 4 and 5, respectively, were synthesized. The salts were characterized using 1H NMR, 13C NMR and HRMS analyses. Compound 3 displayed enhanced in vitro antiproliferative activity (7.4-fold) against three prostate cancer cell lines but surprisingly decreased plasma exposure in the pharmacokinetics study. The antiproliferative activities of the compound 2 salts (4 and 5) were equivalent to that of compound 2, but their oral pharmacokinetic profiles were significantly enhanced. Finally, and most importantly, oral administration of the parent compounds (1 and 2) and their corresponding salts (3, 4 and 5) caused dose-dependent potent inhibition/regression of aggressive and difficult-to-treat CWR22Rv1 tumor xenografts growth, with no apparent host toxicities and were highly more efficacious than the blockbuster FDA-approved prostate cancer drugs, Enzalutamide (Xtandi) and Docetaxel (Taxotere). Thus, the HCl salts of Gal (3) and VNPP433-3ß (4 and 5) are excellent orally bioavailable candidates for clinical development.

MNK inhibitor eFT508 (Tomivosertib) suppresses ectopic activity in human dorsal root ganglion neurons from dermatomes with radicular neuropathic pain.

Spontaneous activity in dorsal root ganglion (DRG) neurons is a key driver of neuropathic pain in preclinical models and in patients suffering from this largely untreated disease. While many intracellular signaling mechanisms have been examined in preclinical models that drive this spontaneous activity (SA), none of these have been tested directly on spontaneously active human nociceptors. Using cultured DRG neurons recovered during thoracic vertebrectomy surgeries, we show that inhibition of mitogen activated protein kinase interacting kinase (MNK) with eFT508 (25 nM) reverses SA in human sensory neurons associated with painful dermatomes. MNK inhibition in spontaneously active nociceptors decreased action potential amplitude and produced alterations in the magnitude of afterhyperpolarizing currents suggesting modification of Na+ and K+ channel activity downstream of MNK inhibition. The effects of MNK inhibition on SA took minutes to emerge and were reversible over time with eFT508 washout. MNK inhibition with eFT508 led to a profound loss of eIF4E Serine 209 phosphorylation, a specific target of the kinase, within 2 min of drug treatment, consistent with the rapid action of the drug on SA in electrophysiology experiments. Our results create a compelling case for the future testing of MNK inhibitors in clinical trials for neuropathic pain.

Pathophysiology of obesity and its associated diseases.

The occurrence of obesity has increased across the whole world. Many epidemiological studies have indicated that obesity strongly contributes to the development of cancer, cardiovascular diseases, type 2 diabetes, liver diseases and other disorders, accounting for a heavy burden on the public and on health-care systems every year. Excess energy uptake induces adipocyte hypertrophy, hyperplasia and formation of visceral fat in other non-adipose tissues to evoke cardiovascular disease, liver diseases. Adipose tissue can also secrete adipokines and inflammatory cytokines to affect the local microenvironment, induce insulin resistance, hyperglycemia, and activate associated inflammatory signaling pathways. This further exacerbates the development and progression of obesity-associated diseases. Although some progress in the treatment of obesity has been achieved in preclinical and clinical studies, the progression and pathogenesis of obesity-induced diseases are complex and unclear. We still need to understand their links to better guide the treatment of obesity and associated diseases. In this review, we review the links between obesity and other diseases, with a view to improve the future management and treatment of obesity and its co-morbidities.

Vinorelbine causes a neuropathic pain-like state in mice via STING and MNK1 signaling associated with type I interferon induction.

Type I interferons (IFNs) increase the excitability of dorsal root ganglion (DRG) neurons via activation of MNK-eIF4E translation signaling to promote pain sensitization in mice. Activation of STING signaling is a key component of type I IFN induction. Manipulation of STING signaling is an active area of investigation in cancer and other therapeutic areas. Vinorelbine is a chemotherapeutic that activates STING and has been shown to cause pain and neuropathy in oncology clinical trials in patients. There are conflicting reports on whether STING signaling promotes or inhibits pain in mice. We hypothesized that vinorelbine would cause a neuropathic pain-like state in mice via STING and signaling pathways in DRG neurons associated with type I IFN induction. Vinorelbine (10 mg/kg, i.v.) induced tactile allodynia and grimacing in WT male and female mice and increased p-IRF3 and type I IFN protein in peripheral nerves. In support of our hypothesis, vinorelbine-mediated pain was absent in male and female StingGt/Gt mice. Vinorelbine also failed to induce IRF3 and type I IFN signaling in these mice. Since type I IFNs engage translational control via MNK1-eIF4E in DRG nociceptors, we assessed vinorelbine-mediated p-eIF4E changes. Vinorelbine increased p-eIF4E in DRG in WT animals but not in StingGt/Gt or Mknk1-/- (MNK1 KO) mice. Consistent with these biochemical findings, vinorelbine had an attenuated pro-nociceptive effect in male and female MNK1 KO mice. Our findings support the conclusion that activation of STING signaling in the peripheral nervous system causes a neuropathic pain-like state that is mediated by type I IFN signaling to DRG nociceptors.

Dual targeting of melanoma translation by MNK/eIF4E and PI3K/mTOR inhibitors.

Melanoma is relatively resistant to chemotherapy, and no targeted therapies are fully effective. The most common mutations in melanoma result in hyperactivation of the mitogen-activated protein kinase (MAPK) and PI3K/AKT/ mTOR pathways responsible for initiating and controlling oncogenic protein translation. This makes both the signaling pathways potentially important therapeutic targets in melanoma. Our studies were carried out on human melanoma cell lines WM793 and 1205 LU with similar genomic alteration (BRAFV600E and PTEN loss). We used a highly specific PI3K/mTOR inhibitor, dactolisib (NVP-BEZ235), and Mnk inhibitor - CGP57380 alone and in combination. Here, we explore the mechanism of action of these drugs alone and in combination, as well as their effect on the viability and invasiveness of melanoma cells. Although when used independently, both drugs suppressed cell proliferation and migration, their combination has additional antitumor effects. We demonstrate that simultaneous inhibition of both pathways may prevent possible drug resistance.

Mitogen-activated protein kinase-interacting kinase 1 is involved in mTOR-mediated pancreatic exocrine function of bovine pancreatic acinar cells with leucine supply.

Leucine improves the exocrine capacity of the cow pancreas, but the mechanism was not revealed clearly. Mitogen-activated protein kinase-interacting kinase 1 (MNK1) is a pancreatic acinar cell-specific stress response kinase that regulates digestive enzyme abundance. We aimed to investigate the MNK1 gene and protein expression profiles among various organs or tissues of dairy cows and to demonstrate the mechanism by which leucine-stimulated MNK1 regulates pancreatic exocrine function. Firstly, the expression profiles of MNK1 protein and gene in the tissues and organs of dairy cows were measured using immunohistochemistry and RT-qPCR methods. Next, an in vitro model of cultured Holstein dairy calf pancreatic acinar cells was used to detect the role of MNK1 during pancreatic enzymes release which is stimulated by leucine. Cells were incubated in culture medium containing L-leucine (0.45 mM) for 180 min, and samples were collected hourly, with the control not containing L-leucine (0 mM). MNK1 was expressed at very high levels in the pancreatic tissue of dairy cows. Leucine supplementation increased the α-amylase level but not lipase level at three time-points (60, 120, and 180 min), and the interaction between treatments and times was significant only for α-amylase. Leucine treatment enhanced (P < 0.05) the phosphorylation of MNK1 and eIF4E. In addition, inhibition of MNK1 decreased leucine-mediated α-amylase and lipase release (P < 0.05) and the phosphorylation of Mnk1 and eIF4E but did not affect (P > 0.05) the phosphorylation of the mTOR signalling pathway factors 4EBP1 and S6K1. In summary, MNK1 is a key regulator of pancreatic exocrine function, which is regulated by leucine in the pancreas of dairy cows.