LKB1, Salt-Inducible Kinases, and MEF2C Are Linked Dependencies in Acute Myeloid Leukemia.

The lineage-specific transcription factor (TF) MEF2C is often deregulated in leukemia. However, strategies to target this TF have yet to be identified. Here, we used a domain-focused CRISPR screen to reveal an essential role for LKB1 and its Salt-Inducible Kinase effectors (SIK3, in a partially redundant manner with SIK2) to maintain MEF2C function in acute myeloid leukemia (AML). A key phosphorylation substrate of SIK3 in this context is HDAC4, a repressive cofactor of MEF2C. Consequently, targeting of LKB1 or SIK3 diminishes histone acetylation at MEF2C-bound enhancers and deprives leukemia cells of the output of this essential TF. We also found that MEF2C-dependent leukemias are sensitive to on-target chemical inhibition of SIK activity. This study reveals a chemical strategy to block MEF2C function in AML, highlighting how an oncogenic TF can be disabled by targeting of upstream kinases.

A novel SIK2 inhibitor SIC-19 exhibits synthetic lethality with PARP inhibitors in ovarian cancer.

PURPOSE: Ovarian cancer patients with HR proficiency (HRP) have had limited benefits from PARP inhibitor treatment, highlighting the need for improved therapeutic strategies. In this study, we developed a novel SIK2 inhibitor, SIC-19, and investigated its potential to enhance the sensitivity and expand the clinical utility of PARP inhibitors in ovarian cancer. METHODS: The SIK2 protein was modeled using a Molecular Operating Environment (MOE), and the most favorable model was selected based on a GBVI/WSA dG scoring function. The Chembridge Compound Library was screened, and the top 20 candidate compounds were tested for their interaction with SIK2 and downstream substrates, AKT-pS473 and MYLK-pS343. SIC-19 emerged as the most promising drug candidate and was further evaluated using multiple assays. RESULTS: SIC-19 exhibited selective and potent inhibition of SIK2, leading to its degradation through the ubiquitination pathway. The IC50 of SIC-19 correlated inversely with endogenous SIK2 expression in ovarian cancer cell lines. Treatment with SIC-19 significantly inhibited cancer cell growth and sensitized cells to PARP inhibitors in vitro, as well as in ovarian cancer organoids and xenograft models. Mechanistically, SIK2 knockdown and SIC-19 treatment reduced RAD50 phosphorylation at Ser635, prevented nuclear translocation of RAD50, disrupted nuclear filament assembly, and impaired DNA homologous recombination repair, ultimately inducing apoptosis. These findings highlight the crucial role of SIK2 in the DNA HR repair pathway and demonstrate the significant PARP inhibitor sensitization achieved by SIC-19 in ovarian cancer. CONCLUSIONS: SIC-19, a novel SIK2 inhibitor, effectively inhibits tumor cell growth in ovarian cancer by interfering with RAD50-mediated DNA HR repair. Furthermore, SIC-19 enhances the efficacy of PARP inhibitors, providing a promising therapeutic strategy to improve outcomes for ovarian cancer patients.

SIK2-positive tumor epithelial cells in breast cancer maybe potential anti-cancer messengers: A systematic analysis from a single-cell perspective.

Breast cancer is the most common malignancy in the world and one of the leading causes of cancer death, which is a heterogeneous disease involving genetic and environmental factors. Breast cancer stem cells (BCSCs) are the main players in the aggressiveness of different tumors, at the same time, these cells are the main challenge for cancer treatment. There are multiple treatment options for breast cancer (BC) patients and the lack of understanding of prognostic and predictive biomarkers for breast cancer is a potential research direction for us to develop better treatments in the future. In this paper, we conducted a correlation analysis between SIK2 and clinical traits by searching numerous BRCA datasets in the GEO database. The model was constructed and validated by incorporating tumor samples from the TCGA-BRCA cohort. Surprisingly, we found differential expression of SIK2 gene in individual tumor samples from the UCSC database. Subsequently, we found significantly high expression of SIK2 in epithelial cells by comparing the differential expression of SIK2 in different cell subpopulations and performed subsequent immune infiltration and pathway correlation analysis. Differential genes in SIK2+ epithelial cells, which may be potential therapeutic targets for breast cancer. In conclusion, our results suggest that SIK2 may be a potential prognostic and predictive biomarker that could serve as an oncogenic messenger for breast cancer. This discovery of SIK2 may provide more valuable references for potential therapeutic tools for breast cancer.

Salt-inducible kinase 2 regulates fibrosis during bleomycin-induced lung injury.

Idiopathic pulmonary fibrosis is a progressive and normally fatal disease with limited treatment options. The tyrosine kinase inhibitor nintedanib has recently been approved for the treatment of idiopathic pulmonary fibrosis, and its effectiveness has been linked to its ability to inhibit a number of receptor tyrosine kinases including the platelet-derived growth factor, vascular endothelial growth factor, and fibroblast growth factor receptors. We show here that nintedanib also inhibits salt-inducible kinase 2 (SIK2), with a similar IC50 to its reported tyrosine kinase targets. Nintedanib also inhibited the related kinases SIK1 and SIK3, although with 12-fold and 72-fold higher IC50s, respectively. To investigate if the inhibition of SIK2 may contribute to the effectiveness of nintedanib in treating lung fibrosis, mice with kinase-inactive knockin mutations were tested using a model of bleomycin-induced lung fibrosis. We found that loss of SIK2 activity protects against bleomycin-induced fibrosis, as judged by collagen deposition and histological scoring. Loss of both SIK1 and SIK2 activity had a similar effect to loss of SIK2 activity. Total SIK3 knockout mice have a developmental phenotype making them unsuitable for analysis in this model; however, we determined that conditional knockout of SIK3 in the immune system did not affect bleomycin-induced lung fibrosis. Together, these results suggest that SIK2 is a potential drug target for the treatment of lung fibrosis.

Inhibiting eukaryotic initiation factor 5A (eIF5A) hypusination attenuated activation of the SIK2 (salt-inducible kinase 2)-p4E-BP1 pathway involved in ovarian cancer cell proliferation and migration.

BACKGROUND: Eukaryotic initiation factor 5A hypusine (eIF5AHyp) stimulates the translation of proline repeat motifs. Salt inducible kinase 2 (SIK2) containing a proline repeat motif is overexpressed in ovarian cancers, in which it promotes cell proliferation, migration, and invasion. METHODS AND RESULTS: Western blotting and dual luciferase analyses showed that depletion of eIF5AHyp by GC7 or eIF5A-targeting siRNA downregulated SIK2 level and decreased luciferase activity in cells transfected with a luciferase-based reporter construct containing consecutive proline residues, whereas the activity of the mutant control reporter construct (replacing P825L, P828H, and P831Q) did not change. According to the MTT assay, GC7, which has a potential antiproliferative effect, reduced the viability of several ovarian cancer cell lines by 20-35% at high concentrations (ES2 > CAOV-3 > OVCAR-3 > TOV-112D) but not at low concentrations. In a pull-down assay, we identified eukaryotic translation initiation factor 4E-binding protein 1 (4E-BP1) and 4E-BP1 (p4E-BP1) phosphorylated at Ser 65 as downstream binding partners of SIK2, and we validated that the level of p4E-BP1(Ser 65) was downregulated by SIK2-targeting siRNA. Conversely, in ES2 cells overexpressing SIK2, the p4E-BP1(Ser 65) level was increased but decreased in the presence of GC7 or eIF5A-targeting siRNA. Finally, the migration, clonogenicity, and viability of ES2 ovarian cancer cells were reduced by GC7 treatment as well as by siRNA for eIF5A gene silencing and siRNA for SIK2 and 4E-BP1 gene silencing. Conversely, those activities were increased in cells overexpressing SIK2 or 4E-BP1 and decreased again in the presence of GC7. CONCLUSION: The depletion of eIF5AHyp by GC7 or eIF5A-targeting siRNA attenuated activation of the SIK2-p4EBP1 pathway. In that way, eIF5AHyp depletion reduces the migration, clonogenicity, and viability of ES2 ovarian cancer cells.

Designing of potent anti-diabetic molecules by targeting SIK2 using computational approaches.

Diabetes mellitus (DM) is one of the major health problems worldwide. WHO have estimated that 439 million people may have DM by the year 2030. Several classes of drugs such as sulfonylureas, meglitinides, thiazolidinediones etc. are available to manage this disease, however, there is no cure for this disease. Salt inducible kinase 2 (SIK2) is expressed several folds in adipose tissue than in normal tissues and thus SIK2 is one of the attractive targets for DM treatment. SIK2 inhibition improves glucose homeostasis. Several analogues have been reported and experimentally proven against SIK for DM treatment. But, identifying potential SIK2 inhibitors with improved efficacy and good pharmacokinetic profiles will be helpful for the effective treatment of DM. The objective of the present study is to identify selective SIK2 inhibitors with good pharmacokinetic profiles. Due to the unavailability of SIK2 structure, the modeled structure of SIK2 will be an important to understand the atomic level of SIK2 inhibitors in the binding site pocket. In this study, different molecular modeling studies such as Homology Modeling, Molecular Docking, Pharmacophore-based virtual screening, MD simulations, Density Functional Theory calculations and WaterMap analysis were performed to identify potential SIK2 inhibitors. Five molecules from different databases such as Binding_4067, TosLab_837067, NCI_349155, Life chemicals_ F2565-0113, Enamine_7623111186 molecules were identified as possible SIK2 inhibitors.

SIK2: A critical glucolipid metabolic reprogramming regulator and potential target in ovarian cancer.

AIM: To explore the role of salt-inducible kinase 2 (SIK2) on glucose and lipid metabolism in ovarian cancer (OC), so as to increase the understanding of potential inhibitors targeting SIK2 and lay a foundation for future precision medicine in OC patients. METHODS: We reviewed and summarized the regulation effect of SIK2 on glycolysis, gluconeogenesis, lipid synthesis, and fatty acids ß-oxidation (FAO) in OC, as well as the potential molecular mechanism and the prospects of potential inhibitors targeting SIK2 in future cancer treatments. RESULTS: Many pieces of evidence show that SIK2 is closed associated with glucose and lipid metabolism of OC. On the one hand, SIK2 enhances the Warburg effect by promoting glycolysis and inhibiting oxidative phosphorylation and gluconeogenesis, on the other hand, SIK2 regulates intracellular lipid metabolism through promoting lipid synthesis and FAO, all of which ultimately induces growth, proliferation, invasion, metastasis, and therapeutic resistance of OC. On this basis, SIK2 targeting may become a new solution for the treatment of a variety of cancer types including OC. The efficacy of some small molecule kinase inhibitors has also been demonstrated in tumor clinical trials. CONCLUSION: SIK2 displays significant effects in OC progression and treatment through regulating cellular metabolism including glucose and lipid metabolism. Therefore, future research needs to further explore the molecular mechanisms of SIK2 in other types of energy metabolism in OC, based on this to develop more unique and effective inhibitors.

Drug Repurposing: Escitalopram attenuates acute lung injury by inhibiting the SIK2/ HDAC4/ NF-κB signaling cascade.

Acute lung injury (ALI) is a significant cause of morbidity and mortality worldwide. To search for a new treatment for acute lung injury, we investigated the effect of escitalopram on lipopolysaccharide (LPS)-induced ALI. Our results showed that escitalopram inhibited salt-inducible kinase 2 (SIK2) activity (IC50 = 6.36 ± 0.93 µM) and triggered histone deacetylase 4 (HDAC4) dephosphorylation. Following its dephosphorylation, HDAC4 translocated into the nucleus, promoted deacetylation and cytoplasmic shuttling of p65, thus inhibited LPS-induced pro-inflammatory cytokine production. Moreover, escitalopram markedly ameliorated the inflammatory responses, reduced neutrophils infiltration and attenuated LPS-induced pulmonary injury in mice. Taken together, we identified a previously unexplored role for escitalopram in SIK2/HDAC4/NF-κB pathway, therefore escitalopram may be considered as a new treatment for ALI.

Salt-inducible kinase 2 (SIK2) inhibitor ARN-3236 attenuates bleomycin-induced pulmonary fibrosis in mice.

BACKGROUND: Pulmonary fibrosis is a fatal lung disease with complex pathogenesis and limited effective therapies. Salt-inducible kinase 2 (SIK2) is a kinase that phosphorylates CRTCs and regulates many physiological processes. However, the role of SIK2 on pulmonary fibrosis remains unclear, and whether SIK2 inhibitor can attenuate pulmonary fibrosis is unknown. METHOD: We subjected human fetal lung fibroblasts (HFLs) to transforming growth factor-ß1 (5 ng/mL) for 12 h, and examined the expression of SIK2, CRTCs and pCRTCs in fibroblasts by western-blot. To address the roles of SIK2 and CRTCs involved in the progression of pulmonary fibrosis, HFLs were treated with a small-molecule inhibitor ARN-3236 or by siRNA-mediated knockdown of SIK2 expression. Pulmonary fibrosis model was established with mice by exposing to bleomycin, and assessed by H&E and Masson's trichrome staining. COL1A and α-SMA distributions were detected in lung tissues by immunohistochemical staining. RESULTS: We discovered that SIK2 and phosphorylated-CRTC2 were expressed at a low basal level in normal lung tissues and quiescent fibroblasts, but increased in fibrotic lung tissues and activated fibroblasts. Inhibition of SIK2 by ARN-3236 prevented the fibroblasts differentiation and extracellular matrix expression in HFLs and attenuated bleomycin-induced pulmonary fibrosis in mice. Mechanistically, inactivation of SIK2 resulted in the dephosphorylation and nuclear translocation of CRTC2. Within the nucleus, CRTC2 binds to CREB, promoting CREB-dependent anti-fibrotic actions. CONCLUSION: In conclusion, our results elucidated a previously unexplored role of SIK2 in pulmonary fibrosis, and identified SIK2 as a new target for anti-fibrosis medicines.

MYBL2-induced PITPNA-AS1 upregulates SIK2 to exert oncogenic function in triple-negative breast cancer through miR-520d-5p and DDX54.

BACKGROUND: In recent years, long non-coding RNAs (lncRNAs) have attracted much attention because of its regulatory role in occurrence and progression of tumors, including triple-negative breast cancer (TNBC). LncRNA PITPNA antisense RNA 1 (PITPNA-AS1) has been explored in some cancers, whereas its function and molecular mechanism in TNBC remain unclear. METHODS: PITPNA-AS1 expression in TNBC tissues and cells was determined by RT-qPCR. TNBC cell viability, proliferation, migration, invasion were assessed with CCK-8, colony formation, wound healing, transwell assays. Cell apoptosis was evaluated by flow cytometry. Expression of EMT-related markers was detected by western blot analyses. The molecular mechanism of PITPNA-AS1 was explored by RNA pull down, luciferase reporter, RIP and ChIP assays. RESULTS: PITPNA-AS1 showed high expression levels in TNBC tissues and cells. PITPNA-AS1 knockdown suppressed TNBC cell viability, proliferation, migration, invasion in vitro and inhibited xenograft tumor growth in mice. Mechanistically, PITPNA-AS1 upregulated SIK2 expression by sponging miR-520d-5p and recruiting DDX54 protein. Results of rescue assays suggested that the inhibitive effects of silenced PITPNA-AS1 on TNBC cellular processes were partially rescued by overexpressing SIK2 or combination of miR-520d-5p inhibition and DDX54 overexpression. More importantly, we found that the upregulation of PITPNA-AS1 in TNBC cells was attributed to transcription factor MYBL2. CONCLUSION: PITPNA-AS1 activated by MYBL2 plays an oncogenic role in TNBC through upregulating SIK2.

Antagonistic modulation of SIK1 and SIK2 isoforms in high blood pressure and cardiac hypertrophy triggered by high-salt intake.

Salt-inducible kinases (SIKs) represent a subfamily of AMPK family kinases. SIK1 has been shown to act as a mediator during the cellular adaptation to variations in intracellular sodium in a variety of cell types. SIK2, as an isoform of the SIK family, modulates various biological functions and acts as a signal transmitter in various pathways. To evaluate the role of both SIK1 and SIK2 isoforms in blood pressure (BP), body fluid regulation and cardiac hypertrophy development, we made use of constitutive sik1-/- (SIK1-KO), sik2-/- (SIK2-KO), double sik1-/-sik2-/- (double SIK1*2-KO) knockout and wild-type (WT) mice challenged to a standard (0.3% NaCl) or chronic high-salt (HS, 8% NaCl) diet intake for 12 weeks.Mice, under a standard diet intake, had similar and normal BP. On a chronic HS intake, SIK1-KO and double SIK1*2-KO mice showed increased BP, but not WT and SIK2-KO mice. A chronic HS intake led to the development of cardiac left ventricle hypertrophy (LVH) in normotensive WT and hypertensive SIK1-KO mice, but not in SIK2-KO mice. Double SIK1*2-KO mice under standard diet intake show normal BP but an increased LV mass. Remarkably, in response to a dietary stress condition, there is an increase in BP but LVH remained unchanged in double SIK1*2-KO mice.In summary, SIK1 isoform is required for maintaining normal BP in response to HS intake. LVH triggered by HS intake requires SIK2 isoform and is independent of high BP.

Salt-inducible kinase 2 functions as a tumor suppressor in hepatocellular carcinoma.

Salt-inducible kinase 2 (SIK2) has been reported to be involved in cancer progression in a dichotomous manner. However, the role and mechanism of action of SIK2 in hepatocellular carcinoma (HCC) progression remain elusive. SIK2 expression in HCC tissues in The Cancer Genome Atlas (TCGA) database was analyzed using the AIPuFu platform. SIK2 expression in HCC cells was examined by quantitative real-time PCR and western blot analysis. The expression of N-cadherin, E-cadherin, ß-catenin, and c-Myc was detected by western blot analysis. SIK2 was downregulated in HCC tissues compared with normal patients, and low SIK2 expression was correlated with poor prognosis in HCC patients in TCGA database. SIK2 was lowly expressed in HCC cells than that in normal human liver epithelial cells. SIK2 overexpression inhibited cell proliferation and invasion and promoted apoptosis in HCC cells, while SIK2 silencing exerted the opposite effects. Additionally, SIK2 overexpression inactivated the Wnt/ß-catenin pathway in HCC cells, as evidenced by the reduced expression of ß-catenin and c-Myc. ß-catenin overexpression rescued the inhibitory effects of SIK2 on the malignant properties of HCC cells. Xenograft tumor experiment confirmed that SIK2 suppressed the growth of HCC cells in vivo. In conclusion, SIK2 exerted anti-tumor activity in HCC via inactivating the Wnt/ß-catenin signaling pathway.

Tumor-suppressor Fbxw7 targets SIK2 for degradation to interfere with TORC2-AKT signaling in pancreatic cancer.

The tumor suppressor F-box/WD repeat-containing protein 7 (Fbxw7) is a substrate-recognition subunit of a ubiquitin ligase complex. We have previously proposed that Fbxw7 inhibited pancreatic cancer cell proliferation and invasion by targeting ß-catenin. To identify other targets of Fbxw7 involved in pancreatic carcinogenesis, we screened the human protein database for Fbxw7 target candidates using the conserved Fbxw7-recognizing sequences. Twenty-three candidates are identified, including five known Fbxw7 targets and two cancer-related genes (salt inducible kinase 2 [SIK2] and ZMIZ1). We identified SIK2 as an Fbxw7 target for degradation by binding to the "TPPPS" motif of SIK2 in pancreatic cancer cells. We also demonstrated that SIK2 promoted proliferation and mitotic progression of pancreatic cancer cells. Moreover, endogenous Fbxw7 downregulates SIK2 protein level for controlling cell cycle progression, possibly by interfering the SIK2/TORC2/AKT signaling pathway to modulate p21 expression. Collectively, these data demonstrate that Fbxw7 targets the cell cycle controller, SIK2, for degradation, thereby leading to the disruption of downstream TORC2/AKT signaling to inhibit pancreatic cancer cell proliferation and cell cycle progression.

M2 Macrophagy-derived exosomal miRNA-5106 induces bone mesenchymal stem cells towards osteoblastic fate by targeting salt-inducible kinase 2 and 3.

BACKGROUND: Osteoblast differentiation is a vital process for fracture healing, and exosomes are nanosized membrane vesicles that can deliver therapeutic drugs easily and safely. Macrophages participate in the regulation of various biological processes in vivo, and macrophage-derived exosomes (MD-Exos) have recently been a topic of increasing research interest. However, few study has explored the link between MD-Exos and osteoblast differentiation. Herein, we sought to identify miRNAs differentially expressed between M1 and M2 macrophage-derived exosomes, and to evaluate their roles in the context of osteoblast differentiation. RESULTS: We found that microRNA-5106 (miR-5106) was significantly overexpressed in M2 macrophage-derived exosomes (M2D-Exos), while its expression was decreased in M1 macrophage-derived exosomes (M1D-Exos), and we found that this exosomal miRNA can induce bone mesenchymal stem cell (BMSC) osteogenic differentiation via directly targeting the Salt-inducible kinase 2 and 3 (SIK2 and SIK3) genes. In addition, the local injection of both a miR-5106 agonist or M2D-Exos to fracture sites was sufficient to accelerate healing in vivo. CONCLUSIONS: Our study demonstrates that miR-5106 is highly enriched in M2D-Exos, and that it can be transferred to BMSCs wherein it targets SIK2 and SIK3 genes to promote osteoblast differentiation.

Parathyroid hormone (PTH) promotes ADSC osteogenesis by regulating SIK2 and Wnt4.

Developing methods for regenerating bones and uncovering the molecular mechanism underlying bone formation have great significance to human health. In the last decade, people have been using adipose-derived stem cells (ADSCs), that are capable of multilineage differentiation, to reconstruct defected bones. Uncovering the molecular mechanisms of the osteoblast differentiation of ADSCs will provide more understanding of ADSCs in the application of bone regeneration and perhaps new methods for osteoporosis treatment. Here we studied how parathyroid hormone (PTH1-34) acts on osteoinduced ADSCs to orchestrate bone formation and how Wnt4 signaling is involved in PTH-promoted bone formation from ADSCs. We found that PTH1-34 can phosphorylate SIK2, upregulate RANKL and downregulate SOST, thereby upregulating Wnt4 to promote the osteogenesis process of ADSCs. Though the knockdown of Wnt4 with shRNA interference barely affects the expression of upstream proteins (i.e., RANKL, SOST), it affects the expression of other downstream osteogenic proteins (i.e., Runx2, Osterix, and Osteocalcin), and then inhibit the osteogenesis process of ADSCs. Overall, PTH can affect the osteogenesis process of ADSCs by regulating SIK2 and Wnt4. We anticipate that this work will provide researchers with new insights into the bone regeneration with ADSCs.

Upregulation of miR-874-3p and miR-874-5p inhibits epithelial ovarian cancer malignancy via SIK2.

Based on miR-874 expression levels in the GSE47841 microarray, we hypothesized that the mature products of miR-874, miR-874-3p, or miR-874-5p, would inhibit epithelial ovarian cancer (EOC) cell proliferation, metastasis, and chemoresistance. We first examined miR-874-3p and miR-874-5p expression levels in primary EOC tumor tissue samples and found that they were significantly decreased. 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) cell proliferation and transwell assays revealed that miR-874-3p and miR-874-5p significantly inhibit EOC cell proliferation, migration, and invasion. Then, using MTT and soft agar assays of paclitaxel-treated Caov3 and SKOV3 cells transfected with miR-874-3p and miR-874-5p, we found that miR-874-3p and miR-874-5p enhance EOC cell chemosensitivity. We then confirmed that serine/threonine-protein kinase 2 (SIK2) was a target gene of miR-874-3p and miR-874-5p. Overall, the results of this study indicate that SIK2 expression can serve as a prognostic biomarker for EOC and that miR-874-3p and miR-874-5p have the potential to enhance clinical treatment of EOC.

Salt-inducible kinase 2 and -3 are downregulated in adipose tissue from obese or insulin-resistant individuals: implications for insulin signalling and glucose uptake in human adipocytes.

AIMS/HYPOTHESIS: Salt-inducible kinases (SIKs) are related to the metabolic regulator AMP-activated protein kinase (AMPK). SIK2 is abundant in adipose tissue. The aims of this study were to investigate the expression of SIKs in relation to human obesity and insulin resistance, and to evaluate whether changes in the expression of SIKs might play a causal role in the development of disturbed glucose uptake in human adipocytes. METHODS: SIK mRNA and protein was determined in human adipose tissue or adipocytes, and correlated to clinical variables. SIK2 and SIK3 expression and phosphorylation were analysed in adipocytes treated with TNF-α. Glucose uptake, GLUT protein levels and localisation, phosphorylation of protein kinase B (PKB/Akt) and the SIK substrate histone deacetylase 4 (HDAC4) were analysed after the SIKs had been silenced using small interfering RNA (siRNA) or inhibited using a pan-SIK-inhibitor (HG-9-91-01). RESULTS: We demonstrate that SIK2 and SIK3 mRNA are downregulated in adipose tissue from obese individuals and that the expression is regulated by weight change. SIK2 is also negatively associated with in vivo insulin resistance (HOMA-IR), independently of BMI and age. Moreover, SIK2 protein levels and specific kinase activity display a negative correlation to BMI in human adipocytes. Furthermore, SIK2 and SIK3 are downregulated by TNF-α in adipocytes. Silencing or inhibiting SIK1-3 in adipocytes results in reduced phosphorylation of HDAC4 and PKB/Akt, less GLUT4 at the plasma membrane, and lower basal and insulin-stimulated glucose uptake in adipocytes. CONCLUSION/INTERPRETATION: This is the first study to describe the expression and function of SIKs in human adipocytes. Our data suggest that SIKs might be protective in the development of obesity-induced insulin resistance, with implications for future treatment strategies.

Bosutinib Attenuates Inflammation via Inhibiting Salt-Inducible Kinases in Experimental Model of Intracerebral Hemorrhage on Mice.

BACKGROUND AND PURPOSE: Intracerebral hemorrhage (ICH) is a subtype of stroke with highest mortality and morbidity. Pronounced inflammation plays a significant role in the development of the secondary brain injury after ICH. Recently, SIK-2 (salt-inducible kinase-2) was identified as an important component controlling inflammatory response. Here we sought to investigate the role of SIK-2 in post-ICH inflammation and potential protective effects of SIK-2 inhibition after ICH. METHODS: Two hundred and ninety-three male CD-1 mice were used. ICH was induced via injection of 30 µL of autologous blood. Recombinant SIK-2 was administrated 1 hour after ICH intracerebroventricularly. SIK-2 small interfering RNA was injected intracerebroventricularly 24 hours before ICH. Bosutinib, a clinically approved tyrosine kinase inhibitor with affinity to SIK-2, was given intranasally 1 hour or 6 hours after ICH. Effects of treatments were evaluated by neurological tests and brain water content calculation. Molecular pathways were investigated by Western blots and immunofluorescence studies. RESULTS: Endogenous SIK-2 was expressed in microglia and neurons. SIK-2 expression was reduced after ICH. Exogenous SIK-2 aggravated post-ICH inflammation, leading to brain edema and the neurobehavioral deficits. SIK-2 inhibition attenuated post-ICH inflammation, reducing brain edema and ameliorating neurological dysfunctions. Bosutinib inhibited SIK-2-attenuating ICH-induced brain damage. Protective effects of Bosutinib were mediated, at least partly, by CRTC3 (cyclic amp-response element binding protein-regulated transcription coactivator 3)/cyclic amp-response element binding protein/NF-κB (nuclear factor-κB) pathway. CONCLUSIONS: SIK-2 participates in inflammation induction after ICH. SIK-2 inhibition via Bosutinib or small interfering RNA decreased inflammation, attenuating brain injury. SIK-2 effects are, at least partly, mediated by CRTC3-cyclic amp-response element binding protein-NF-κB signaling pathway.

Overexpression of miR-203 sensitizes paclitaxel (Taxol)-resistant colorectal cancer cells through targeting the salt-inducible kinase 2 (SIK2).

MicroRNAs (miRNAs) are short non-coding RNAs that regulate gene expression through the endogenous RNA interference machinery. Treatments with combination of chemotherapy with surgery are essential for advanced-stage colorectal cancer. However, the development of chemoresistance is a major obstacle for clinical application of anticancer drugs. In this study, we report a miR-203-SIK2 axis that involves in the regulation of Taxol sensitivity in colon cancer cells. MiR-203 is downregulated in human colon tumor specimens and cell lines compared with their normal counterparts. We report miR-203 is correlated with Taxol sensitivity: overexpression of miR-203 sensitizes colon cancer cells and the Taxol-resistant cells display downregulated miR-203 compared with Taxol-sensitive cells. We identify SIK2 as a direct target of miR-203 in colorectal cancer cells. Overexpression of miR-203 complementary pairs to the 3' untranslated region (UTR) of SIK2, leading to the sensitization of Taxol resistant cells. In addition, miR-203 and the salt-inducible kinase 2 (SIK2) are reverse expressed in human colorectal tumors. Finally, we demonstrate recovery of SIK2 by overexpression of SIK2-desensitized Taxol-resistant cells, supporting the miR-203-mediated sensitization to Taxol, is through the inhibition of SIK2. In general, our study will provide mechanisms of the microRNA-based anti-tumor therapy to develop anti-chemoresistance drugs.

The association between Salt-inducible kinase 2 (SIK2) and gamma isoform of the regulatory subunit B55 of PP2A (B55gamma) contributes to the survival of glioma cells under glucose depletion through inhibiting the phosphorylation of S6K.

BACKGROUND: PPP2R2C encodes a gamma isoform of the regulatory subunit B55 subfamily consisting PP2A heterotrimeric with A and C subunits. Currently, the precise functions of B55gamma in cancer are still under investigating. In this project, we reported a novel function of B55gamma in the regulation of glucose metabolism in Glioma cells. METHODS: Western blot and immunoprecipitation were performed to determine protein expression and interaction. Cell viability was measured by Typan Blue staining and direct cell counting using hematocytometer. siRNA technology was used to down regulate protein expression. RESULTS: Glucose uptake and lactate product were suppressed by overexpression of B55gamma in Glioma cells. In addition, cancer cells with larger amount of B55gamma showed higher survival advantages in response to glucose starvation through the dephosphorylation of S6K. From proteomic analysis, we found B55gamma binds with and up regulates SIK2 through the stabilization of SIK2 protein which is required for the B55gamma-mediated suppression of S6K pathway. Knocking down of SIK2 in B55gamma over expressing cells recovered the phosphorylation of S6K. CONCLUSION: In summary, our project will provide novel insight into the design and development of therapeutic strategies to target the B55gamma-mediated glucose metabolism for the treatment of human brain tumor patients.