Detection of porcine-derived ingredients from adulterated meat based on real-time loop-mediated isothermal amplification.

Increasingly globalized and complex food supply chains contribute to a growing problem of meat fraud. Meat adulteration with pork is especially exceptionable to the global population for health concern and religious faith reasons. To prevent unfair competition and protect consumer rights, an efficient and rapid assay to identify the species of meat products is crucial. In this study, a real-time loop-mediated isothermal amplification (real-time LAMP) assay was developed for the detection of a porcine gene in meat products. The designed primers were highly selective for the porcine gene. The amplification showed no cross-reactivity with 11 other meats. The established method required 20 min with an initial amplification curve of approximately 10 min and demonstrated a detection limit of 1.76 pg/µL porcine DNA, which is 1000 times more sensitive than PCR. This study is the first attempt at detecting porcine-derived ingredients using a real-time LAMP assay in commercial products. This method meets specificity, rapidness, robustness, and sensitivity criteria; its practical application will greatly aid in battling adulteration in the food industry.

Molecular and immunological characterizations of interleukin-11 in large yellow croaker (Larimichthys crocea).

Interleukin (IL)-11 is a multifunctional cytokine that exerts a series of important immunomodulatory effects and exists in many tissues and cells. A 1106-bp nucleotide sequence representing the complete cDNA of IL-11 was obtained from large yellow croaker (Larimichthys crocea), containing an open reading frame (ORF) of 603 bp encoding for 200 amino acids (aa). The predicted LcIL-11 protein included a 12aa signal peptide and a conserved IL-11 domain. The polypeptide sequence identities between LcIL-11 and its counterparts in mammals and other fish are from 84% to 92% with known fish IL-11a and 22%-27% with fish IL-11b. LcIL-11 mRNA existed in most tissues with the most predominant expression in the gill. After immune challenge, the expression levels of LcIL-11 were induced largely in vivo and in vitro, with the peak-value of 32 times as much as the control in the liver at 24 h after Vibrio parahaemolyticus injection (p < 0.05) and the greatest value of 13.9 times as much as the control in LCK cells at 12 h after poly I:C stimulation (p < 0.05). Furthermore, the overexpression vector pcDNA3.1-LcIL-11 was constructed and transfected to LCK cells. Our results showed that the transcriptional expression levels of tumor necrosis factor (TNF)-α and myxovirus resistant protein (Mx) significantly up-regulated in LCK cells after LcIL-11 overexpression (p < 0.05). However, no significant changes of IL-1ß, janus kinase (JAK)2 and signal transducers and activators of transcription (STAT)5 was detected. Our finding indicated that LcIL-11 might enhance TNF-α and antiviral protein Mx expression in large yellow croaker.

EYA1 promotes tumor angiogenesis by activating the PI3K pathway in colorectal cancer.

Blood vessels are one of the major routes for the dissemination of cancer cells. Malignant tumors release growth factors such as vascular endothelial growth factor(VEGF) to induce angiogenesis, thereby promoting metastasis. Here, we report that The Drosophila Eyes Absent Homologue 1 (EYA1), which is overexpressed in colorectal tumor cells, can promote colorectal tumor angiogenesis by coordinating with the hypoxia-inducible factor 1 (HIF-1α) to increase the expression of VEGF-A. Moreover, data indicated that the enhancement of HIF-1α expression by Eya1 depended on its ability to activate the phosphatidylinositol 3-kinase (PI3K) signaling pathways to increase the phosphorylation of AKT subunits. Overexpression of Eya1 increased tumor angiogenesis in vivo and in vitro. Our study suggested that Eya1 is essential in regulating cancer cell-mediated angiogenesis and contributes to tumor growth, and that Eya1 provides a potential and specific target for new anti-angiogenesis drug development.

There is No Association Between the Omega-3 Index and Depressive Symptoms in Patients With Heart Disease Who Are Low Fish Consumers.

BACKGROUND: Long chain Omega-3 polyunsaturated fatty acids (LCn3PUFAs) may improve cardiovascular health and depression. This study investigated the relationships between erythrocyte membrane LCn3PUFA status, depression and angina symptoms in patients with heart disease. METHODS: We recruited 91 patients (65 males and 26 females, mean age 59.2±10.3 years) with heart disease and depressive symptoms (Center for Epidemiological Studies Depression Scale, CES-D ≥ 16) and low fish/fish oil intakes. The Omega-3 Index (EPA+DHA) of erythrocyte membranes (as a percentage of total fatty acids) was assessed by gas chromatography. Depression status was measured by both self-report and clinician-report scales; CES-D and the Hamilton depression scale (HAM-D). Angina symptoms were measured using the Seattle Angina Questionnaire and the Canadian Cardiovascular Society Classification for Angina Pectoris. RESULTS: The mean Omega-3 Index was 4.8±1.0% (±SD). Depression scores measured by CES-D and HAM-D were 29.2±8.8 (moderate to severe) and 11.0±5.7 (mild) (arbitrary units) respectively reflecting a different perception of depressive symptoms between patients and clinicians. Angina status was inversely associated with depression scores (r>-0.26, P<0.03). There were no significant relationships between individual LCn3PUFA or the Omega-3 Index and either the depression scores or the angina symptoms. CONCLUSION: Worse angina status was associated with worse depression, but the Omega-3 Index was not associated with symptoms of depression or angina in patients with heart disease.

c-Myc enhances colon cancer cell-mediated angiogenesis through the regulation of HIF-1α.

Angiogenesis plays a pivotal role in tumor growth. The hypoxia-inducible factor 1, α subunit (HIF-1α)/vascular endothelial growth factor pathway is the most important pathway for regulating angiogenesis in the tumor microenvironment. c-Myc is an important oncogene that has many biological functions. In this study, we investigated the role of c-Myc in tumor angiogenesis. We found that the overexpression of c-Myc in colon cancer cells could promote the expression of HIF-1α and that of vascular endothelial growth factor. Moreover, we found that c-Myc regulated HIF-1α at the post-transcriptional level. The results revealed c-Myc-dependent regulation of HIF-1α instead of HIF-1α-dependent c-Myc regulation for the first time. They also showed that c-Myc was essential to regulate colon cancer cell-mediated angiogenesis and contributed to tumor growth. This research provides the theoretical basis for clinical trials of new therapeutic targets of c-Myc and HIF-1α in colon cancer cells.

Prognostic value of neutrophil-to-lymphocyte ratio for patients with triple-negative breast cancer: A meta-analysis.

We aimed to evaluate the prognostic role of the neutrophil-to-lymphocyte ratio (NLR) in patients with triple-negative breast cancer (TNBC). We searched the PubMed Database, Cochrane Library, Web of science, and Embase. Overall survival (OS), disease-free survival (DFS), and hazard ratio (HR) were the endpoints of the study. Eleven studies involving 2355 patients with TNBC were included in this meta-analysis. Among them, 10 studies involving 2069 patients with TNBC investigated the role of NLR in predicting OS; elevated NLR was associated with poor prognosis (hazard ratio [HR]: 2.48, 95% confidence interval [CI]: 2.03-3.03, P < 0.001). Ten studies involving 2054 patients with TNBC explored the role of NLR in predicting DFS; elevated NLR was associated with a significantly worse prognosis with a pooled HR of 2.20 (95% CI: 1.88-2.58, P < 001). This meta-analysis suggests that patients with TNBC who have a higher NLR have poorer prognoses. As a clinical parameter that we can easily obtain, NLR might be a potential predictor of patients' survival, and may be useful for physicians' treatment decisions.

Effect of KRAS mutations and p53 expression on the postoperative prognosis of patients with colorectal cancer.

BACKGROUND: In the occurrence and development of colorectal cancer, p53 is an important regulator downstream of the MAPK signaling pathway and plays an important role in inhibiting abnormal proliferation signals generated by KRAS mutations. The purpose of this study is to explore the correlation between KRAS mutations and p53 expression and evaluate their prognosis values in colorectal cancer. METHODS: PCR technology and immunohistochemical (IHC) staining were used to detect KRAS mutation status and p53 expression level in 266 specimens of colorectal adenocarcinoma. Based on p53 expression level, these were divided into high expression and normal groups. Patients with KRAS mutations were divided into mutant and wild-type groups. The two were combined with each other to analyze the relationship between patients' clinical data and their impact on the prognosis. RESULTS: KRAS mutations were found in 38.6% of the patients and 40.8% had a high p53 expression. There was no significant difference in the overall survival rate of patients, with or without KRAS gene mutations, and p53 expression level. In the group of patients with KRAS mutations, the survival time of patients with a high p53 expression was significantly lower than that of patients with a normal p53 expression (p = 0.020, log-rank test). Multivariate analysis showed that p53 high expression is an independent risk factor for the overall survival time of patients with KRAS mutations (HR = 2.330, 95% CI = 1.041-5.216, p < 0.05). CONCLUSION: Colorectal cancer patients with KRAS mutations with a high p53 expression have a poor prognosis.

Not All Kirsten Rat Sarcoma Viral Oncogene Homolog Mutations Predict Poor Survival in Patients With Unresectable Colorectal Liver Metastasis.

Objective: To assess the characteristics of Kirsten rat sarcoma viral oncogene homolog (KRAS) mutations and investigated whether all KRAS mutations predict poor prognosis in patients with unresectable colorectal liver metastasis (CRLM). Methods: Correlations between KRAS-mutation status and clinicopathological characteristics of 93 patients with unresectable CRLM at our institution between 2010 and 2018 were retrospectively analyzed. Kaplan-Meier and Cox proportional hazard models were used to evaluate the prognostic significance of KRAS mutations. Results:KRAS were primarily single-point mutations, identified in 41.9% of patients. There were no significant differences in clinicopathological characteristics between wild-type KRAS and mutant KRAS. Patients with mutant KRAS had significantly worse overall survival (OS) and progression-free survival (PFS) than those with wild-type KRAS. Moreover, patients with codon 12 mutations had worse OS and PFS than those with wild-type KRAS, whereas mutations in codon 13 were not associated with a worse prognosis. Among the 5 most common mutations in codons 12, G12V, and G12D were associated with worse OS, furthermore, G12C mutation seemed to associated with worse PFS than patients with wild-type KRAS. Conclusion:KRAS codon 12 mutations were predictive for a poor prognosis in patients with unresectable CRLM. G12D and G12V mutations were associated with worse OS, whereas G12C mutation seemed to be associated with decreased PFS.

Roles of HDAC2, eIF5, and eIF6 in Lung Cancer Tumorigenesis.

OBJECTIVE: The expression levels of histone deacetylase 2 (HDAC2), eukaryotic initiation factor 5 (eIF5), and eukaryotic initiation factor 6 (eIF6), and relationship between HDAC2 and eIF5 or eIF6 in lung cancer tissues were investigated, in order to charify the relationship between HDAC2 and the prognosis of lung cancer patients and its influence on the expression of eIF5 and eIF6. METHODS: The expression of HDAC2, eIF5, and eIF6 in lung cancer tissues was detected by quantitative reverse transcription polymerase chain reaction. The expression correlation between HDAC2 and eIF5 or eIF6 was tested using a t test. The correlation between HDAC2 and eIF5 or eIF6 was analyzed using the TCGA database. The identified cells were constructed with small interfering siRNA and HDAC2 overexpression plasmid. The proliferation and migration ability of the identified cells was investigated by CCK8 and Transwell assays, respectively. RESULTS: HDAC2, eIF5, and eIF6 were overexpressed in lung cancer tissues, and HDAC2 expression level was negatively correlated with the prognosis of lung cancer patients. HDAC2 expression level was positively correlated with eIF5 and eIF6 expression levels. HDAC2 could regulate the expression of eIF5 and eIF6. The regulation of proliferation and invasion of lung cancer cells by HDAC2 depended on eIF5 and eIF6. CONCLUSION: HDAC2, eIF5, and eIF6 were closely related with lung cancer tumorigenesis, which might be potential biological markers and therapeutic targets for lung cancer.

Lysine-specific histone demethylase 1B (LSD2/KDM1B) represses p53 expression to promote proliferation and inhibit apoptosis in colorectal cancer through LSD2-mediated H3K4me2 demethylation.

Epigenetic alterations have been reported to play critical roles in the development of colorectal cancer (CRC). However, the biological function of the lysine-specific histone demethylase 1B (LSD2/KDM1B) in CRC is not well understood. Therefore, we investigated the characteristics of LSD2 in CRC. We observed significant upregulation of LSD2 in CRC tissue compared to that in normal colorectal tissue. LSD2 promotes CRC cell proliferation and inhibits cell apoptosis through cell cycle regulation, promoting CRC progression both in vitro and in vivo. We found that LSD2 performs these functions by inhibiting the p53-p21-Rb pathway. Finally, we found that LSD2 directly binds to p53 and represses p53 expression via H3K4me2 demethylation at the p53 promoter. Our results revealed that LSD2 acts as an oncogene by binding and inhibiting p53 activity in CRC. Thus, LSD2 may be a new molecular target for CRC treatment.

Large tumor size is a poor prognostic factor of gastric cancer with signet ring cell: Results from the surveillance, epidemiology, and end results database.

There has been a steady increase in the incidence of signet ring cell (SRC) carcinoma, a distinct histological type with cells containing abundant intracytoplasmic mucin. We aimed to analyze the clinicopathological characteristics and prognostic value of patients with SRC gastric cancer (GC) who underwent gastrectomy.Clinical data of 10,312 GC patients who underwent D2 radical gastrectomy were obtained from the Surveillance, Epidemiology, and End Results database and were retrospectively analyzed. X-tile plots were constructed to illustrate the optimal cut-off points using the minimum P-value from the log-rank Chi-squared test. The Kaplan-Meier method was used for the analysis of the overall cumulative probability of survival. Their differences were evaluated using the log-rank test. The Cox multiple factors analysis was performed using the logistic regression method.In total, 946 (9.17%) SRC GC patients with pT1a-4bN0-3bM0 stage cancer were recruited. The optimal cut-off point for size was 49 mm. The 3-year overall survival (OS) rates of the SRC GC, large-size, and small-size groups were 35.89%, 30.63%, and 44.96%, respectively (P < .05). Cox multivariate analysis showed that tumor size (odds ratio [OR] = 2.032), T3 category (OR = 1.324), T4a category (OR = 1.945), and T4b category (OR = 2.163) were independent hazard prognostic factors.SRC GC has a distinct biological behavior, presents as a large-sized tumor (≥49 mm), and is associated with worse outcomes. SRC GC patients have 2.032 times risk of mortality. SRC patients with larger tumors are at higher risk for infiltrative growth, lymph node metastasis, and distant metastasis.

[Application of intracorporeal uncut Roux-en-Y anastomosis in digestive tract reconstruction after laparoscopic total gastrectomy].

OBJECTIVE: To explore the safety, feasibility and short-term efficacy of intracavitary uncut Roux-en-Y (URY) anastomosis in digestive tract reconstruction following laparoscopic total gastrectomy (LTG). METHODS: From November 2015 to January 2018, 67 gastric cancer patients underwent intracavitary URY following LTG to reconstruct the digestive tract at Oncological Surgery Department of Fujian Provincial Hospital. There were 41 males and 26 females with age of 50 to 81 (61.9±7.4) years and body mass index (BMI) of (23.4±3.2) kg/m². Among 67 patients, 19 were gastric cardia carcinomas, 33 were gastric body carcinomas, and 15 were gastric fundus carcinomas; tumor size was (3.4±2.3) cm; 22 were Borrmann type I, 15 were type II, 21 were type III, and 19 were type IV; 29 were highly or moderately differentiated adenocarcinoma, 23 were lowly differentiated adenocarcinoma, and 15 were signet-ring cell carcinoma. After conventional laparoscopic D2 radical gastrectomy, the duodenum was closed and dissociated at 2 cm below the pyloric ring using the Echelon-flex endoscopic articulated linear Endo-GIA stapler, and the esophagus was dissociated above the esophagogastric junction (EGJ).URY and digestive tract reconstruction were performed under the direct vision of laparoscope: (1) Side-to-side esophagojejunostomy: An incision of 0.5 cm was made in the left lower edge of the esophageal closed end; jejunum about 25 cm distal away from the Treitz ligament was elevated to the lower end of esophagus; another incision of 0.5 cm was made in the contralateral of mesenteric side; both arms of the linear Endo-GIA stapler were inserted into the windows opened through esophagus and jejunum respectively to complete side-to-side anastomosis. The common opening of esophagus and jejunum was closed to complete esophagojejunostomy, forming the chyme outflow tract. (2) Side-to-side Braun jejunojejunostomy: Incisions of 0.5 cm were made in the proximal jejunum about 10 cm away from the esophagojejunal anastomosis and 35-40 cm away from the contralateral of mesenteric side of distal jejunum respectively for proximal-distal side-to-side jejunojejunostomy. The common opening was closed to form the biliopancreatic duodenal juice outflow tract. (3) Closure of the input loop jejunum in the esophagojejunal anastomosis: The input loop jejunum 2-3 cm away from the esophagojejunal anastomosis was closed using the non-blade linear stapler (ATS45NK), and the biliopancreatic duodenal juice reflux was blocked. Clinical data of these patients were collected for retrospective case series study. Surgical and digestive tract functional recovery, perioperative complications, as well as postoperative nutritional status were observed. Moreover, related indexes, such as anastomosis function and tumor recurrence were evaluated through endoscopic and imaging examinations during postoperative follows-up. RESULTS: All the 67 patients completed the surgery successfully. The mean operative time was (259.4±38.5) minutes, digestive tract reconstruction time was (38.2±13.2) minutes, intraoperative blood loss was (73.4±38.4) ml, and number of harvested lymph node was 36.2±14.2. The mean distance from upper resection margin to upper tumor edge was (3.3±1.2) cm, distance from upper resection margin to dentate line was (1.2±0.7) cm, and 1 case had positive upper incisal margin, which became negative after the second resection. Moreover, the average length of the auxiliary incision was (3.2±0.4) cm. The mean postoperative intestinal exhaust time was (52.8±26.4) hours, time to liquid diet was (64.8±28.8) hours, and postoperative hospital stay was (8.4±2.5) days. The morbidity of postoperative complication was 10.4%(7/67). Among these 7 cases, 4 cases were grade IIIa of Clavien-Dindo classification, including 2 with esophagojejunal anastomosis leakage, 1 with duodenal stump leakage, and 1 with abdominal infection, and all these patients were recovered after conservative treatment. All the 67 patients were followed up. The mean nutrition index 12 months after surgery was 53.4±4.2, diameter of esophagojejunal anastomosis was (3.9±0.6) cm, the incidence of Roux-en-Y stasis syndrome was 3.0% (2/67), and the incidence of reflux esophagitis was 4.5% (3/67). No patient had recanalization of the closed input loop of esophagojejunal anastomosis, anastomotic stenosis, obstruction, or tumor recurrence at anastomosis. CONCLUSION: Intracavitary URY anastomosis following LTG for digestive tract reconstruction is safe and feasible, leading to fast postoperative recovery of digestive tract function and favorable short-term efficacy.

Not all mutations of KRAS predict poor prognosis in patients with colorectal cancer.

The mutation of Kirsten rat sarcoma viral oncogene homolog (KRAS) has been reported to be prognostically important in patients with colorectal cancer (CRC). In this study, we investigated whether all KRAS mutations predict poor prognosis in patients with CRC. Our analysis of characteristics of KRAS mutations revealed the mutation rate for codon 12 was 72.7%, of which G12D was the highest (47.5%) followed by G12V (30.6%), and the mutation rate for codon 13 was 22.0%, of which all were G13D. In support of the concept that prognostic value of the KRAS codon-12 mutations is different from the codon-13 mutations, results from our Cox proportional hazard model studies showed that codon-12 mutations correlated with worse overall survival (OS; HR = 2.846, 95% CI: 1.967-4.118, P < 0.001) and progression free survival (PFS; HR = 2.011, 95% CI: 1.450-2.789, P < 0.001). No prognostic significance was revealed for codon-13 mutations. On further analysis, we found that mortality risk was significantly increased with G12D and G12V (G12D: HR = 2.802, 95% CI: 1.793-4.381, P < 0.001; G12V: HR = 2.802, 95% CI: 1.793-4.381, P < 0.001), as was the risk of disease progression (G12D: HR = 2.079, 95% CI: 1.396-3.099, P < 0.001; G12V: HR = 2.408, 95% CI: 1.517-3.822, P < 0.001). To conclude, our results support the concept that codon-12 mutations were predictive for a poor prognosis in Chinese patients with CRC. Specifically, G12D and G12V were independent prognostic factors for worse OS and PFS.

Real-time loop-mediated isothermal amplification for rapid detection of Enterocytozoon hepatopenaei.

BACKGROUND: Enterocytozoon hepatopenaei (EHP) is a newly emerged microsporidian parasite that causes retarded shrimp growth in many countries. But there are no effective approaches to control this disease to date. The EHP could be an immune risk factor for increased dissemination of other diseases. Further, EHP infection involves the absence of obvious clinical signs and it is difficult to identify the pathogen through visual examination, increasing the risk of disease dissemination. It is urgent and necessary to develop a specific, rapid and sensitive EHP-infected shrimp diagnostic method to detect this parasite. In the present study, we developed and evaluated a rapid real-time loop-mediated isothermal amplification (real-time LAMP) for detection of EHP. METHODS: A rapid and efficient real-time LAMP method for the detection of EHP has been developed. Newly emerged EHP pathogens in China were collected and used as the sample, and three sets of specificity and sensitivity primers were designed. Three other aquatic pathogens were used as templates to test the specificity of the real-time LAMP assay. Also, we compared the real-time LAMP with the conventional LAMP by the serial dilutions of EHP DNA and their amplification curves. Application of real-time LAMP was carried out with clinical samples. RESULTS: Positive products were amplified only from EHP, but not from other tested species, EHP was detected from the clinical samples, suggesting a high specificity of this method. The final results of this assay were available within less than 45 min, and the initial amplification curve was observed at about 6 min. We found that the amplification with an exponential of sixfold dilutions of EHP DNA demonstrated a specific positive signal by the real-time LAMP, but not for the LAMP amplicons from the visual inspection. The real-time LAMP amplification curves demonstrated a higher slope than the conventional LAMP. DISCUSSION: In this study, pathogen virulence impacts have been increased in aquaculture and continuous observation was predominantly focused on EHP. The present study confirmed that the real-time LAMP assay is a promising and convenient method for the rapid identification of EHP in less time and cost. Its application greatly aids in the detection, surveillance, and prevention of EHP.

HINT2 downregulation promotes colorectal carcinoma migration and metastasis.

Histidine triad nucleotide-binding 2 (HINT2), a member of the histidine triad proteins family, sensitizes cells to apoptosis in hepatocellular carcinoma. Here, we showed that HINT2 expression is lower in primary colorectal cancer (CRC) and metastasis tissues than in normal colorectal tissues, and that HINT2 abundance is inversely correlated with CRC tumor stage. Treating CRC cells with 5-aza-2'-deoxycytidine, a demethylating agent, upregulated HINT2, suggesting HINT2 downregulation is caused by methylation of the gene promoter. HINT2 downregulation increased tumor migration and invasion in vitro, promoted CRC cell metastasis in vivo, and increased expression of epithelial-to-mesenchymal transition (EMT) markers. Furthermore, HINT2 downregulation depended on hypoxia inducible factor (HIF)-2α-mediated transcriptional activation of zinc finger E-box-binding homeobox 1 (ZEB1). These results suggest that HINT2 downregulation promotes HIF-2α expression, which induces EMT and enhances CRC cell migration and invasion. HINT2 may thus a useful clinical indicator of CRC progression and metastasis risk.

Isorhamnetin suppresses colon cancer cell growth through the PI3K­Akt­mTOR pathway.

Isorhamnetin, a flavonoid isolated from the fruits of herbal medicinal plants, such as Hippophae rhamnoides L., exerts anticancer effects similar to other flavonoids. However, the effect of isorhamnetin on colorectal cancer (CRC) and the underlying molecular mechanism are unclear. This study aimed to determine the effect of isorhamnetin on the proliferation of cells from the human CRC cell lines, HT­29, HCT116 and SW480. It was demonstrated that isorhamnetin suppressed the proliferation of cells from all three cell lines, induced cell cycle arrest at the G2/M phase and suppressed cell proliferation by inhibiting the PI3K­Akt­mTOR pathway. Isorhamnetin also reduced the phosphorylation levels of Akt (ser473), phosph­p70S6 kinase and phosph­4E­BP1 (t37/46) protein, and enhanced the expression of Cyclin B1 protein. Therefore, this compound was revealed to be a selective PI3K­Akt­mTOR pathway inhibitor, and may be a potent anticancer agent for the treatment of CRC, as it restrains the proliferation of CRC cells.

The tumor suppressive role of CAMK2N1 in castration-resistant prostate cancer.

Prostate cancer at advanced stages including metastatic and castration-resistant cancer remains incurable due to the lack of effective therapies. The CAMK2N1 gene, cloned and characterized as an inhibitor of CaMKII (calcium/calmodulin-dependent protein kinase II), has been shown to affect tumorigenesis and tumor growth. However, it is still unknown whether CAMK2N1 plays a role in prostate cancer development. We first examined the protein and mRNA levels of CAMK2N1 and observed a significant decrease in human prostate cancers comparing to normal prostate tissues. Re-expression of CAMK2N1 in prostate cancer cells reduced cellular proliferation, arrested cells in G0/G1 phases, and induced apoptotic cell death accompanied by down-regulation of IGF-1, ErbB2, and VEGF downstream kinases PI3K/AKT, as well as the MEK/ERK-mediated signaling pathways. Conversely, knockdown of CAMK2N1 had a significant opposite effects on these phenotypes. Our analyses suggest that CAMK2N1 plays a tumor suppressive role in prostate cancer cells. Reduced CAMK2N1 expression correlates to human prostate cancer progression and predicts poor clinical outcome, indicating that CAMK2N1 may serve as a biomarker. The inhibition of tumor growth by expressing CAMK2N1 established a role of CAMK2N1 as a therapeutic target.

CAMK2N1 inhibits prostate cancer progression through androgen receptor-dependent signaling.

Castration resistance is a major obstacle to hormonal therapy for prostate cancer patients. Although androgen independence of prostate cancer growth is a known contributing factor to endocrine resistance, the mechanism of androgen receptor deregulation in endocrine resistance is still poorly understood. Herein, the CAMK2N1 was shown to contribute to the human prostate cancer cell growth and survival through AR-dependent signaling. Reduced expression of CAMK2N1 was correlated to recurrence-free survival of prostate cancer patients with high levels of AR expression in their tumor. CAMK2N1 and AR signaling form an auto-regulatory negative feedback loop: CAMK2N1 expression was down-regulated by AR activation; while CAMK2N1 inhibited AR expression and transactivation through CAMKII and AKT pathways. Knockdown of CAMK2N1 in prostate cancer cells alleviated Casodex inhibition of cell growth, while re-expression of CAMK2N1 in castration-resistant cells sensitized the cells to Casodex treatment. Taken together, our findings suggest that CAMK2N1 plays a tumor suppressive role and serves as a crucial determinant of the resistance of prostate cancer to endocrine therapies.

EYA1 phosphatase function is essential to drive breast cancer cell proliferation through cyclin D1.

The Drosophila Eyes Absent Homologue 1 (EYA1) is a component of the retinal determination gene network and serves as an H2AX phosphatase. The cyclin D1 gene encodes the regulatory subunits of a holoenzyme that phosphorylates and inactivates the pRb protein. Herein, comparison with normal breast showed that EYA1 is overexpressed with cyclin D1 in luminal B breast cancer subtype. EYA1 enhanced breast tumor growth in mice in vivo, requiring the phosphatase domain. EYA1 enhanced cellular proliferation, inhibited apoptosis, and induced contact-independent growth and cyclin D1 abundance. The induction of cellular proliferation and cyclin D1 abundance, but not apoptosis, was dependent upon the EYA1 phosphatase domain. The EYA1-mediated transcriptional induction of cyclin D1 occurred via the AP-1-binding site at -953 and required the EYA1 phosphatase function. The AP-1 mutation did not affect SIX1-dependent activation of cyclin D1. EYA1 was recruited in the context of local chromatin to the cyclin D1 AP-1 site. The EYA1 phosphatase function determined the recruitment of CBP, RNA polymerase II, and acetylation of H3K9 at the cyclin D1 gene AP-1 site regulatory region in the context of local chromatin. The EYA1 phosphatase regulates cell-cycle control via transcriptional complex formation at the cyclin D1 promoter.

Dachshund binds p53 to block the growth of lung adenocarcinoma cells.

Hyperactive EGF receptor (EGFR) and mutant p53 are common genetic abnormalities driving the progression of non-small cell lung cancer (NSCLC), the leading cause of cancer deaths in the world. The Drosophila gene Dachshund (Dac) was originally cloned as an inhibitor of hyperactive EGFR alleles. Given the importance of EGFR signaling in lung cancer etiology, we examined the role of DACH1 expression in lung cancer development. DACH1 protein and mRNA expression was reduced in human NSCLC. Reexpression of DACH1 reduced NSCLC colony formation and tumor growth in vivo via p53. Endogenous DACH1 colocalized with p53 in a nuclear, extranucleolar location, and shared occupancy of -15% of p53-bound genes in ChIP sequencing. The C-terminus of DACH1 was necessary for direct p53 binding, contributing to the inhibition of colony formation and cell-cycle arrest. Expression of the stem cell factor SOX2 was repressed by DACH1, and SOX2 expression was inversely correlated with DACH1 in NSCLC. We conclude that DACH1 binds p53 to inhibit NSCLC cellular growth.