Renal cell carcinoma (RCC) is a malignant tumor originating from the epithelial cells of the renal tubules. The clear cell RCC subtype is closely linked to a poor prognosis due to its rapid progression. Circular RNA (circRNA) is a novel class of regulatory RNA molecules that play a role in the development of ccRCC, although their functions have not been fully elucidated. In this study, we identified a significant downregulation of circ-IP6K2 in ccRCC tissues based on data from the GSE100186 dataset. The decreased expression of circ-IP6K2 correlated with the progression of TNM stage and histological grade, and was also associated with decreased overall survival rates in ccRCC patients. Moreover, our findings revealed that circ-IP6K2 expression suppressed proliferation, migration, and invasion capabilities in vitro, and inhibited xenograft growth in vivo. Mechanistically, circ-IP6K2 acted as a sponge for miR-1292-5p in ccRCC cells, which in turn targeted the 3\'UTR of CAMK2N1, leading to a decrease in its expression. CAMK2N1 was identified as a tumor suppressor that negatively regulated the β-catenin/c-Myc oncogenic signaling pathway. Additionally, we confirmed a positive correlation between the expression of circ-IP6K2 and CAMK2N1 in ccRCC. Circ-IP6K2 functions to impede the progression of ccRCC by modulating the miR-1292-5p/CAMK2N1 axis. These findings shed new light on the molecular mechanisms driving ccRCC progression and suggest potential therapeutic targets for the treatment of ccRCC.

Methyltransferase like 3 (METTL3) has been reported to promote tumorigenesis of multiple myeloma (MM), however, the molecular mechanism still needs further research. The N6-methyladenosine (m6A) level in tissues or cells was measured by m6A kit and dot blot assay. The mRNA and protein expression were detected by quantitative real-time PCR (RT-qPCR) and Western blot, respectively. The cell counting kit-8 and colony formation assay were used to detect the cell proliferation. Coimmunoprecipitation (Co-IP) experiment verified the binding of two proteins. The luciferase reporter experiment demonstrated the targeted binding of miR-182-5p and CaMKII inhibitor 1 (CAMK2N1). More importantly, tumor growth was measured in xenograft mice. Our data showed that the expression of METTL3 was significantly increased in MM patients\' samples and MM cells. METTL3 overexpression promoted MM cells proliferation, and METTL3 knockdown inhibited MM cells proliferation. Mechanically, METTL3-dependent m6A participated in DiGeorge syndrome critical region 8 (DGCR8)-mediated maturation of pri-miR-182. Upregulation of miR-182-5p further enhanced the promoting proliferation effect of METTL3 overexpression on MM cells. Moreover, the luciferase reporter gene experiment proved that miR-182-5p targetedly regulated CAMK2N1 expression. Xenograft tumor in nude mice further verified that METTL3 promoted MM tumor growth through miR-182/CAMK2N1 signal axis. In summary, the METTL3/miR-182-5p/CAMK2N1 axis plays an important role in MM tumorigenesis, which may provide a new target for MM therapy.

Gastric cancer (GC) is still notorious for its poor prognosis and aggressive characteristics. Though great developments have been made in diagnosis and therapy for GC, the prognosis of patient is still perishing. In this study, differentially expressed genes (DEGs) in GC were first screened using three Gene Expression Omnibus (GEO) datasets (GSE13911, GSE29998, and GSE26899). Second, The Cancer Genome Atlas (TCGA) and Genotype-Tissue Expression (GTEx) data were used to validate expression of these DEGs and perform survival analysis. We selected seven candidate genes (CAMK2N1, OLFML2B, AKR7A3, CYP4X1, FMO5, MT1H, and MT1X) to carry out the next analysis. To construct the ceRNA network, we screened the most potential upstream ncRNAs of the candidate genes. A series of bioinformatics analyses, including expression analysis, correlation analysis, and survival analysis, revealed that the SNHG10-hsa-miR-378a-3p might be the most potential regulatory axis in GC. Then, the expression of CAMK2N1, miR-378a-3p, and SNHG10 was verified in GC cell lines (GES-1, MGC-803, BGC-823, HGC-27, MKN-45, and AGS) by qRT-PCR and Western blotting. We found that SNHG10 and CAMK2N1 were highly expressed in gastric cancer lines, and the miR-378a-3p was lowly expressed in BGC-823, HGC-27, and MKN-45. Furthermore, CAMK2N1 levels were significantly negatively associated with tumor immune cell infiltration, biomarkers of immune cells, and immune checkpoint expression. In summary, our results suggest that the ncRNA-mediated high expression of CAMK2N1 is associated with poor prognosis and tumor immune infiltration of GC.

The deregulation of calcium/calmodulin-dependent protein kinase II inhibitor 1 (CAMK2N1) is linked to the carcinogenesis reported in several malignancies. To date, studies describing the role of CAMK2N1 in colorectal carcinoma are scarce. The current project was carried out to study the relationship between CAMK2N1 and colorectal carcinoma progression. CAMK2N1 levels were lowered in colorectal carcinoma tissue, which also correlated to poor overall survival in patients. Colorectal carcinoma cell lines with overexpressed CAMK2N1 showed a reduction in transformative phenotypes, including proliferation suppression, the blocking of cell cycle progression, metastasis inhibition and chemoresistance reduction, whereas CAMK2N1-silenced cells showed the opposite effect. Mechanistic studies revealed a novel regulatory role of CAMK2N1 on Wnt/β-catenin transduction. Up-regulation of CAMK2N1 lowered the level of disheveled 2, phosphorylated GSK-3β, β-catenin, c-myc and cyclin D1. Re-expression of β-catenin decreased the CAMK2N1-mediated tumor-inhibiting effects. Moreover, blocking of Wnt/β-catenin diminished CAMK2N1-silencing-elicited cancer-enhancing effect. Critically, the tumorigenicity of CAMK2N1-overexpressed cells was markedly weakened in nude mice. To conclude, the study demonstrated a cancer-suppressive function of CAMK2N1 in colorectal carcinoma and illustrated that CAMK2N1 exerts the tumor-inhibiting effects via suppression of the Wnt/β-catenin pathway.

Prostate cancer (PCa) is the most frequently diagnosed malignancy in men, and its mechanism remains poorly understood. Therefore, it is urgent to discover potential novel diagnostic biomarkers and therapeutic targets that can potentially facilitate the development of efficient anticancer strategies.
A series of functional in vitro and in vivo experiments were conducted to evaluate the biological behaviors of PCa cells. RNA pulldown, Western blot, luciferase reporter, immunohistochemistry and chromatin immunoprecipitation assays were applied to dissect the detailed underlying mechanisms. High-throughput sequencing was performed to screen for differentially expressed circRNAs in PCa and adjacent normal tissues.
Upregulation of protein arginine methyltransferase 5 (PRMT5) is associated with poor progression-free survival and the activation of multiple signaling pathways in PCa. PRMT5 inhibits the transcription of CAMK2N1 by depositing the repressive histone marks H4R3me2s and H3R8me2s on the proximal promoter region of CAMK2N1, and results in malignant progression of PCa both in vitro and in vivo. Moreover, the expression of circSPON2, a candidate circRNA in PCa tissues identified by RNA-seq, was found to be associated with poor clinical outcomes in PCa patients. Further results showed that circSPON2 induced PCa cell proliferation and migration, and that the circSPON2-induced effects were counteracted by miR-331-3p. Particularly, circSPON2 acted as a competitive endogenous RNA (ceRNA) of miR-331-3p to attenuate the repressive effects of miR-331-3p on its downstream target PRMT5.
Our findings showed that the epigenetic regulator PRMT5 aggravates PCa progression by inhibiting the transcription of CAMK2N1 and is modulated by the circSPON2/miR-331-3p axis, which may serve as a potential therapeutic target for patients with aggressive PCa.

Inflammation response and subsequent ventricular remodeling are critically involved in the development of ventricular arrhythmia post myocardial infarction (MI). However, as the vital endogenous inhibitor of calcium/calmodulin-dependent protein kinase II (CaMKII), the effects of CaMKII inhibitor 1 (Camk2n1) on the process of arrhythmia substrate generation following MI remains unclear. In this study, we investigated the role of Camk2n1 in ventricular arrhythmia post-MI and the underlying mechanisms.
Camk2n1 was mainly expressed in cardiomyocytes and inhibited the phosphorylation of CaMKIIδ in the infarcted border zone. Compared to wild type (WT) littermates mice, Camk2n1 knockout mice (Camk2n1-/-) manifested exacerbated cardiac dysfunction, larger fibrosis area, higher incidence of premature ventricular contractions (PVCs) and higher vulnerability to ventricular tachycardia (VT) or ventricular fibrillation (VF) after MI. The results of RNA sequencing (RNA-seq) identified that excessive activation of NLRP3 inflammasome was responsible for aggravated inflammation response which led to adverse cardiac remodeling in Camk2n1-/- mice subjected to MI. More importantly, both in vivo and in vitro experiments verified that aggravated NLRP3 inflammasome activation occurred via CaMKIIδ-p38/JNK pathway in Camk2n1-/- mice.
Collectively, our results highlight the importance of Camk2n1 in alleviating ventricular remodeling and malignant ventricular arrhythmia post-MI by reducing cardiomyocytes inflammation activation via CaMKIIδ-p38/JNK-NLRP3 inflammasome pathway, targeting Camk2n1 might serve as a novel therapeutic strategy after MI.

CaMK2N1 and CaMK2N2 (also known as CaMKIINα and β) are endogenous inhibitors of calcium/calmodulin-dependent kinase II (CaMKII), an enzyme critical for memory and long-term potentiation (LTP), a form of synaptic plasticity thought to underlie learning. CaMK2N1/2 mRNAs are rapidly and differentially upregulated in the hippocampus and amygdala after acquisition or retrieval of fear memory. Moreover, CaMK2N2 protein levels increase after contextual fear conditioning. Therefore, it was proposed that CaMK2N1/2 genes (Camk2n1/2) could be immediate-early genes transcribed promptly (30-60 min) after training. As a first approach to explore a role in synaptic plasticity, we assessed a possible regulation of Camk2n1/2 during the expression phase of LTP in hippocampal CA3-CA1 connections in rat brain slices. Quantitative PCR revealed that Camk2n1, but not Camk2n2, is upregulated 60 min after LTP induction by Schaffer collaterals high-frequency stimulation. We observed a graded, significant positive correlation between the magnitude of LTP and Camk2n1 change in individual slices, suggesting a coordinated regulation of these properties. If mRNA increment actually resulted in the protein upregulation in plasticity-relevant subcellular locations, CaMK2N1 may be involved in CaMKII fine-tuning during LTP maintenance or in the regulation of subsequent plasticity events (metaplasticity).

Purpose: Prostate cancer (PCa) varies clinically from very indolent, not requiring therapeutic intervention, to highly aggressive, entailing radical treatment. Currently, stratification of PCa aggressiveness is mostly based on Gleason score, serum PSA and TNM stage, but outcome prediction in an individual basis is suboptimal. Thus, perfecting pre-therapeutic discrimination between indolent and aggressive PCa, avoiding overtreatment is a major challenge. Epithelial to mesenchymal transition (EMT) allows epithelial cells to acquire mesenchymal properties, constituting a critical step in tumor invasion and metastization. Thus, we hypothesized that EMT-related markers might allow for improved assessment of PCa aggressiveness. Methods and Results: Using RealTime ready Custom Panel 384 assay, 93 EMT-related genes were assessed in normal prostate tissues (NPT, n=5), stage pT2a+b-PCa (n=5) and stage pT3b-PCa (n=5), from which CAMK2N1, CD44, KRT14, TGFβ3 and WNT5A genes emerged as the most significantly altered. Expression levels were then evaluated in a larger series (16 NPT and 94 PCa) of frozen tissues using quantitative RT-PCR. Globally, CAMK2N1, CD44 and WNT5A displayed higher expression levels at higher stages and less differentiated PCa. CAMK2N1 and WNT5A immunoexpression analysis disclosed significantly lower expression in NPT and increasing proportion of high-expression cases from pT2a+b to pT3b and metastatic PCa. Furthermore, higher CAMK2N1 and WNT5A transcript levels associated with shorter disease-free and disease-specific survival. In multivariable analysis, a trend for WNT5A expression levels to independently predict DFS was disclosed (p=0.056). Conclusions: Globally, our findings suggest an association between PCa aggressiveness and increased expression of CAMK2N1 and WNT5A, reflecting the acquisition of effective EMT characteristics by PCa cells.

Synaptotagmin12 (SYT12) has been well characterized as the regulator of transmitter release in the nervous system, however the relevance and molecular mechanisms of SYT12 in oral squamous cell carcinoma (OSCC) are not understood. In the current study, we investigated the expression of SYT12 and its molecular biological functions in OSCC by quantitative reverse transcriptase polymerase chain reaction, immunoblot analysis, and immunohistochemistry. SYT12 were up-regulated significantly in OSCC-derived cell lines and primary OSCC tissue compared with the normal counterparts (P<0.05) and the SYT12 expression levels were correlated significantly with clinical indicators, such as the primary tumoral size, lymph node metastasis, and TNM stage (P<0.05). SYT12 knockdown OSCC cells showed depressed cellular proliferation, migration, and invasion with cell cycle arrest at G1 phase. Surprisingly, we found increased calcium/calmodulin-dependent protein kinase 2 (CAMK2) inhibitor 1 (CAMK2N1) and decreased CAMK2-phosphorylation in the knockdown cells. Furthermore, treatment with L-3, 4-dihydroxyphenylalanine (L-dopa), a drug approved for Parkinson\'s disease, led to down-regulation of SYT12 and similar phenotypes to SYT12 knockdown cells. Taken together, we concluded that SYT12 plays a significant role in OSCC progression via CAMK2N1 and CAMK2, and that L-dopa would be a new drug for OSCC treatment through the SYT12 expression.

OBJECTIVE: Emerging evidence suggests that the propagation of oral squamous cell carcinoma (OSCC) is influenced by the abnormal expression of microRNAs (miRNAs). This study aimed to characterize the involvement of miR-182-5p in OSCC by targeting the calcium/ calmodulin-dependent protein kinase II inhibitor CAMK2N1.
METHODS: miR-182-5p expression was quantified in OSCC tissues and cell lines with reverse transcription polymerase chain reaction (RT-PCR). Cell colony formation, Cell Counting Kit-8 (CCK-8), Ki-67, and nude mouse xenograft assays were used to characterize the role of miR-182-5p in the proliferation of OSCC. A miR-182-5p target gene was identified with western blotting, RT-PCR, and luciferase activity assays. OSCC patient survival based on CAMK2N1 expression was also analyzed.
RESULTS: miR-182-5p was up-regulated in in vitro cell lines and in vivo clinical OSCC samples. CCK-8, colony formation, and Ki-67 assays revealed that miR-182-5p promoted the growth and proliferation of OSCC cells. miR-182-5p directly targeted CAMK2N1, as evidenced by luciferase assays and target prediction algorithms. CAMK2N1 operated as a tumor suppressor gene in patients with OSCC. Down-regulating miR-182-5p expression in the CAL-27 cell line restored CAMK2N1-mediated OSCC cell proliferation. miR-182-5p expression inhibited the activation of AKT, ERK1/2, and NF-κB. Mice injected with CAL-27 cells transfected with miR-182-5p-inhibitor demonstrated a significant increase in tumor size and weight and increased CAMK2N1 mRNA and protein expression compared with the miR-negative control group.
CONCLUSIONS: The miR-182-5p-CAMK2N1 pathway can be potentially targeted to regulate the proliferation of OSCC cells.