p21-activated protein kinase 1 induces the invasion of gastric cancer cells through c-Jun NH2-terminal kinase-mediated activation of matrix metalloproteinase-2.

Gastric cancer (GC) is one of the most common malignancies worldwide. The prognosis of GC is poor, mostly due to widespread metastasis. p21-activated kinase 1 (Pak1), the best characterized member of an evolutionarily conserved family of serine/threonine kinases, plays an important role in the regulation of cell morphogenesis, motility, mitosis and angiogenesis. By qRT-PCR and Gelatin zymograph assay, we demonstrated in the present study that stable overexpression of Pak1 induced matrix metalloproteinase (MMP)-2 mRNA expression and activity in the human MKN45 GC cell line. Conversely, knockdown of endogenous Pak1 expression by small interfering RNA (siRNA) decreased MMP-2 mRNA expression and activity in the MKN45 GC cells. Activation of c-Jun N-terminal kinase (JNK) was required for Pak1-induced upregulation of MMP-2 mRNA level and activity. Moreover, upregulation of MMP-2 by Pak1 via the JNK pathway notably promoted the invasion of MKN45 GC cells. Overexpression of MMP-2 mRNA was once again confirmed to be associated with GC metastasis. In conclusion, our results demonstrated for the first time that Pak1 stimulated MMP-2 mRNA expression and activity in MKN45 GC cells. The JNK signaling pathway was involved in Pak1 modulation of MMP-2, which was important for MKN45 GC cell invasiveness.

Prognostic role of phospho-PRAS40 (Thr246) expression in gastric cancer.

INTRODUCTIONS: Phospho-PRAS40(Thr246) (phosphorylated proline-rich Akt substrate of 40 kilodaltons at Thr246) is a biomarker for phosphatidylinositol 3-kinase (PI3K) pathway activation and AKT inhibitors sensitivity. MATERIAL AND METHODS: In this study, we immunohistochemically investigated the expression of phospho-PRAS40(Thr246) in 141 gastric cancer tumors, and evaluated its clinicopathological and prognostic significance. RESULTS: Sixty-four cases (45.4%) were defined as phospho-PRAS40(Thr246) positive. Phospho-PRAS40(Thr246) correlated positively with lymph node metastasis, lymphatic infiltration, vascular infiltration and shorter survival. Furthermore, phospho-PRAS40(Thr246) is an independent prognostic factor for gastric cancer. CONCLUSIONS: Our data suggest that phospho-PRAS40(Thr246) was frequently expressed in gastric cancers, and correlated with malignant progression and poor prognosis of patients. PI3K pathway-targeted therapies should be considered in the future treatment of gastric cancers.

Clinical significance of combined testing of YKL-40 with CEA in Chinese colorectal cancer patients.

BACKGROUND: To investigate the practical value of individual and combined testing of plasma levels of YKL-40, CEA, and CA199 for auxiliary diagnosis and detection of recurrence of colorectal cancer. METHODS: ELISA and ECLIA were used to evaluate levels of YKL-40, CEA, and CA199 in 120 colorectal cancer patients (56 initial-diagnosis, 42 post-operative, and 22 recurrent cases). Forty-three patients with benign colorectal disease and 36 healthy patients were enrolled as controls. The relationship between YKL-40 and clinical indicators of tumor pathology was analyzed. The positive rate and diagnostic efficacy of single and combined YKL-40, CEA, and CA199 testing were assessed in patients with colorectal cancer. RESULTS: Plasma YKL-40 in the cancer group was significantly higher than in the benign control and healthy control group, and the mean values were 145.4 ng/mL, 107.7 ng/mL, and 51.3 ng/mL (p < 0.05), respectively. With 72 ng/mL as the diagnostic threshold, the sensitivity and specificity of YKL-40 in colorectal cancer diagnosis were found to be 73.2% and 66.7%, respectively. Early-stage colorectal cancer patients showed a YKL-40 positive rate (73%) significantly higher than those of CEA and CA199 (50% and 32%, respectively; p < 0.05). When YKL-40 testing was combined with CEA or CA199, the positive rate increased to 82.1% and 80.3%, respectively. Through ROC curve analysis of the post-operative recurrent group against the non-recurrent group, the areas under the curve for YKL-40, CEA, and CA199 were found to be 0.907, 0.714, and 0.759, respectively. Based on the Dukes classification, the mean YKL-40 value for stages A/B, C, and D were 120.1 ng/mL, 131.7 ng/mL, and 226.8 ng/mL (p = 0.008), respectively. The plasma YKL-40 level gradually increased as the disease progressed. Lower degrees of tumor differentiation were correlated with higher YKL-40 levels. The mean YKL-40 values of high, medium, and low tumor differentiation groups were 96.8 ng/mL, 127.5 ng/mL, and 225.7 ng/mL (p = 0.004), respectively. CONCLUSIONS: The benefits of using YKL-40 testing are higher than CEA and CA199 for the monitoring of colorectal cancer recurrence. Combined testing of both YKL-40 and CEA was found to be optimal for auxiliary diagnosis of colorectal cancer. Plasma YKL-40 was found to be suitable for auxiliary diagnosis of colorectal cancer.

A study on the extraction process of active ingredients from Akebia Stem and an analysis of their anti-gastric cancer activity.

The study investigated the extraction process of active ingredients from akebia stem and an analysis of their anti-gastric cancer activity. Three different extraction methods were used to obtain extracts, namely the decoction method (group A), reflux extraction method (group B), and maceration method (group C), of which reflux extraction method and maceration method used ethanol as the extraction solvent, while decoction method used distilled water for extraction. The differences in anti-gastric cancer activity of the three extracts were compared. MTT assay was used to test and compare the inhibitory effects of extracts obtained in A, B, and C groups on gastric cancer cells. The results showed that the dry extract obtained by heat reflux extraction with "water-ethanol" ratio of 1:2, extractant volume of 70 ml, with ethanol as extraction solvent presented the best inhibitory activity on gastric cancer SGC-7901 cells in this study. Its inhibitory effect did not change over time, and was directly proportional to the concentration.

P21-activated protein kinase 1 is overexpressed in gastric cancer and induces cancer metastasis.

P21-activated protein kinase (Pak1), a main downstream effector of small Rho GTPases, plays an important role in the regulation of cell morphogenesis, motility, mitosis and angiogenesis. However, the role of Pak1 in gastric cancer metastasis remains unclear. Here, we showed that Pak1 is overexpressed in gastric cancer tissues from 74 patients by immunohistochemistry. Overexpression of Pak1 was associated with metastasis and prognosis of gastric cancer. In addition, overexpression of Pak1 increased gastric cancer cell motility and invasion, whereas downregulation of Pak1 expression reduced gastric cancer cell migration and invasion. In further study, data showed that activated Pak1 inhibited stress fiber and focal adhesion complex formation in gastric cancer cells and led to the formation of motile phenotypes. Importantly, activated Pak1 elicited phosphorylation of the ERK and JNK-dependent pathway in gastric cancer cell lines. In conclusion, our results suggest that Pak1 is overexpressed in gastric cancer and plays an important role in the metastasis of gastric cancer. The mechanism by which Pak1 induces cancer metastasis may involve activation of ERK and JNK.

Alterations of tumor-related genes do not exactly match the histopathological grade in gastric adenocarcinomas.

AIM: To investigate the diverse characteristics of different pathological gradings of gastric adenocarcinoma (GA) using tumor-related genes. METHODS: GA tissues in different pathological gradings and normal tissues were subjected to tissue arrays. Expressions of 15 major tumor-related genes were detected by RNA in situ hybridization along with 3' terminal digoxin-labeled anti-sense single stranded oligonucleotide and locked nucleic acid modifying probe within the tissue array. The data obtained were processed by support vector machines by four different feature selection methods to discover the respective critical gene/gene subsets contributing to the GA activities of different pathological gradings. RESULTS: In comparison of poorly differentiated GA with normal tissues, tumor-related gene TP53 plays a key role, although other six tumor-related genes could also achieve the Area Under Curve (AUC) of the receiver operating characteristic independently by more than 80%. Comparing the well differentiated GA with normal tissues, we found that 11 tumor-related genes could independently obtain the AUC by more than 80%, but only the gene subsets, TP53, RB and PTEN, play a key role. Only the gene subsets, Bcl10, UVRAG, APC, Beclin1, NM23, PTEN and RB could distinguish between the poorly differentiated and well differentiated GA. None of a single gene could obtain a valid distinction. CONCLUSION: Different from the traditional point of view, the well differentiated cancer tissues have more alterations of important tumor-related genes than the poorly differentiated cancer tissues.

[A preliminary study on construction of targeting double suicide gene therapy vector pcDNA3.1 (-) Cp-CD-TK].

OBJECTIVE: To construct the targeting double suicide gene therapy vector pcDNA3.1 (-) Cp-CD-TK driven by carcino-embryonic antigen promoter (CEA promoter, Cp), and to investigate whether the vector could control the expression of CD-TK gene specificity and impact on proliferation of CEA positive colon cancer cells. METHODS: Three kinds of target gene, Cp, CD, and TK were obtained by PCR, the products were double digested and inserted into pcDNA3.1 (-), then the targeting vector pcDNA3.1 (-) Cp-CD-TK was transfected into CEA positive human colon cancer SW480 cells and CEA negative Hela cells, the expression of CD-TK was examined by RT-PCR. The sensitivities of SW480 cells transfected with pcDNA3.1 (-) Cp-CD-TK to pro-drug 5-Fluorocytosine (5-Fc) and Ganciclovir (GCV) were detected by MTT analysis. RESULTS: Recombinants with Cp, CD and TK insert were obtained. Constructed targeting gene therapy vector pcDNA3.1 (-) Cp-CD-TK was confirmed by gel electrophoresis and sequencing. RT-PCR analysis further confirmed CD-TK gene was expressed in SW480 cells and was not expressed in CEA negative Hela cells. MTT assay demonstrated that SW480 cells transfected with targeting vector pcDNA3.1 (-) Cp-CD-TK were sensitive to pro-drug 5-Fc and GCV. CONCLUSION: Targeting double suicide gene therapy vector pcDNA3.1 (-) Cp-CD-TK was constructed correctly. The vector could make CD-TK gene express specifically in CEA positive cells for the purpose of targeting killing colon cancer.

[Anti-tumor and bystander effects of cationic liposome-mediated CD/5-FC suicide gene system combined with interferon-gamma in vivo].

OBJECTIVE: To study the antitumor and distant bystander effects of cationic liposome-mediated cytosine deaminase (CD)/5-fluorocytosine (5-FC) suicide gene system combined with interferon-gamma (IFN-gamma) in vivo. METHODS: Murine hepatoma 22 (H22) cells transfected by CD gene were inoculated subcutaneous in Kunming mice in the left axillary region, and the H22 cells without CD gene transfection were inoculated in the right axillary region. The mice were randomly divided into 4 groups and treated with normal saline , 5-FC, IFN-gamma, and 5-FC+ IFN-gamma on a daily basis. The tumor inhibition and distant bystander effects were observed in the mice. RESULTS: Exposure of CD gene-transfected tumor to 5-Fc resulted in obvious tumor growth inhibition with an inhibition rate of 78.38%, which was significantly increased to 93.21% (P<0.01) with 5-Fc +IFN-gamma treatment. A notable distant bystander effect in the CD/5-FC suicide gene system was observed in vivo, with a tumor inhibition rate of was 54.42%; when combined with IFN-gamma, the inhibition rate increased significantly to 87.57% (P<0.05). CONCLUSION: When combined with IFN-gamma, CD/5-FC suicide system has stronger anti-tumor and distant bystander effects. CD/5-FC suicide gene system combined with IFN-gamma may provide a potential therapy for malignant tumors.