OBJECTIVETo study the clinical phenotype and its prognostic value of PRAM1 in patients with acute myeloid leukemia(AML).

METHODSBased on the gene expression microarray platform of 486 AML cases, the PRAM1 expression phenotypes were summarized in all of AML subtypes. The PRAM1 expression features were explored in every differentiation stage of hematocytes through normal human stem cell chips and bone marrow gene expression microarray. The clinical drugs which could up-regulate PRAM1 expression in AML cell lines should be found out.

RESULTSThe PRAM1 expression was the richest in the inv(16) AML and the lowest in the t(15;17)M3, almost the same in the other subtypes of AML. By the classification of molecular abnormalities, PRAM1 expression was more in the panel of CN-AML with CEBPAdm than the other two panels. Interestingly, high/low expression of PRAM1 could be re-classified in the CN-AML, and the EFS is statistically significant. It was proven again that PRAM1 is more expressed in the mature granulocytes. Finally, it was confirmed that decitabine and the chidamide could up-regulate PRAM1 expression in AML cell lines, and chidamide effect is better.

CONCLUSIONPRAM1 expression is the lowest in t(15;17) M3 and the highest in inv(16) AML. The high expression of PRAM1 is a sign for favorable prognosis in the CN-AML. PRAM1 is more expressed in mature granulocytes, chidamide can up-regulate PRAM1 expression in AML cell lines.

Adaptor Proteins, Signal Transducing ; Bone Marrow ; Gene Expression ; Humans ; Leukemia, Myeloid, Acute ; Microarray Analysis ; Prognosis

BACKGROUND/AIMS: In order to identify microsatellite instability (MSI), the test based on the polymerase chain reaction (PCR) can be used. However, PCR is not routinely performed in all hospital laboratories. Recently, immunohistochemistry (IHC) for MLH1 and MSH2 proteins has been reported as a rapid and useful method for MSI. However, the efficacy of IHC in the detection of the MSI has not been well established. The aim of this study was to evaluate the usefulness of IHC in the detection of the MSI by comparing it with the test results using PCR in colorectal cancer (CRC). METHODS: Paraffin-embedded normal and tumor tissues from seventy-five patients who underwent surgical resection of CRC were used. Abnormal expression of MLH1 and MSH2 protein was determined by IHC using MLH1 and MSH2 antibodies. Normal and tumor DNAs were obtained from thirty CRC tissues that showed abnormal expression of MLH1 and MSH2 proteins by IHC. The MSI status was confirmed by PCR using five markers. RESULTS: Thirty tumors showed abnormal expression of MLH1 and MSH2 proteins by IHC, but only three tumors out of them were confirmed to have MSI by PCR. CONCLUSIONS: This result suggests that IHC with MLH1 and MSH2 antibodies does not seem to be a useful method to identify MSI in CRC, therefore PCR is required for detection of the MSI.
Adaptor Proteins, Signal Transducing ; Aged ; Carrier Proteins ; Colorectal Neoplasms/*genetics ; DNA-Binding Proteins/*analysis ; Female ; Humans ; Immunohistochemistry ; Male ; *Microsatellite Repeats ; Middle Aged ; MutS Homolog 2 Protein ; Neoplasm Proteins/*analysis ; Nuclear Proteins ; Polymerase Chain Reaction ; Proto-Oncogene Proteins/*analysis

BACKGROUNDLaryngeal carcinoma is a common malignant tumor of the upper respiratory tract, and in 95% of cases the tumor is laryngeal squamous cell carcinoma (LSCC). The abnormity of SH3-domain GRB2-like 2 (SH3GL2) gene was found in LSCC. In order to clarify the relationship between SH3GL2 gene and LSCC, we evaluated the expression of the SH3GL2 gene in LSCC.

METHODReal-time PCR, immunohistochemistry and Western blotting were used to detect the mRNA and protein expression and find the various rules of SH3GL2 gene in LSCC.

RESULTSThe result of real-time PCR showed that the expression level of SH3GL2 mRNA in LSCC tissue was apparently down-regulated; immunohistochemical analysis showed that SH3GL2 protein was mainly located in cytoplasm, the rate of positive cells and SH3GL2 protein expression level were fluctuated with the pathological classification of LSCC; the result of Western blotting showed that SH3GL2 protein was down-regulated significantly in LSCC samples, especially in metastatic lymph nodes.

CONCLUSIONSThese results suggest that SH3GL2 is a LSCC related gene and its expression level is fluctuated with the pathological classification which indicate that SH3GL2 participates in the development and progression of LSCC. And it may be considered as a novel tumor marker to find both a new anti-oncogene and relative factors of invasion and metastasis of laryngeal carcinoma.

Adaptor Proteins, Signal Transducing ; analysis ; genetics ; Blotting, Western ; Carcinoma, Squamous Cell ; chemistry ; genetics ; Humans ; Immunohistochemistry ; Laryngeal Neoplasms ; chemistry ; genetics ; Polymerase Chain Reaction ; src Homology Domains

OBJECTIVETo detect SH3BP2 gene mutation in a cherubism family.

METHODSPeripheral blood samples were obtained from the family of cherubism. Genomic DNA was extracted. Polymerase chain reaction and direct sequencing were performed to identify the mutation.

RESULTSA transition in exon 9 in SH3BP2 gene was detected in the family, which led to a missense mutation (Arg 415 Pro).

CONCLUSIONSMissense mutation in the SH3BP2 gene was responsible for the phenotypes of this Chinese cherubism family.

Adaptor Proteins, Signal Transducing ; genetics ; Adult ; Cherubism ; genetics ; Child ; DNA Mutational Analysis ; Exons ; genetics ; Female ; Humans ; Infant ; Male ; Middle Aged ; Mutation, Missense ; genetics ; Pedigree

Adaptor Proteins, Signal Transducing ; analysis ; Apoptosis ; drug effects ; Female ; Humans ; Inhibitor of Apoptosis Proteins ; analysis ; Male ; Neoplasm Proteins ; analysis ; PTEN Phosphohydrolase ; analysis ; Retinal Neoplasms ; drug therapy ; Retinoblastoma ; drug therapy ; Topoisomerase I Inhibitors ; pharmacology ; Topotecan ; pharmacology

BACKGROUNDMultidrug resistance to chemotherapeutic agents is an important clinical problem during the treatment of leukemia. The resistance process is multifactorial. To realize the total factors involved in multidrug resistance, we analyzed the differentially expressed proteins of K562 and K562/ADM cells and we investigated one of the up-regulated proteins (CRKL) using siRNA to determine its role in K562/ADM cells.

METHODSAltered protein expressions between K562/S (K562 ADM-sensitive cell line) and K562/ADM (K562 multidrug resistant cell line induced by adriamycin) were identified by 2D-DIGE coupled with mass spectrometry. Meanwhile, we confirmed the differential expression of CRKL and Stathmin in both K562 and K562/ADM cells by Western blot analysis. Furthermore, we used RNA interference to silence the CRKL gene expression.

RESULTSAmong the 9 differentially expressed proteins, 3 were up-regulated in K562/ADM cells, while 6 were down-regulated in the K562/ADM cells compared with its parent cell line. The expression of CRKL was up-regulated significantly in K562/ADM cells, and it can be decreased by recombinant lentivirus. Moreover, the multidrug resistance of K562/ADM cells was efficiently reversed by silence of CRKL gene expression.

CONCLUSIONSThe data provided the differentially expressed proteins in K562 and its resistant cell line and highlights the power of 2D-DIGE for the discovery of resistance markers in cancer. We found CRKL may be a new protein involved in the multidrug resistance of leukaemia cells.

Adaptor Proteins, Signal Transducing ; analysis ; antagonists & inhibitors ; genetics ; Amino Acid Sequence ; Doxorubicin ; pharmacology ; Drug Resistance, Multiple ; Humans ; K562 Cells ; chemistry ; drug effects ; Molecular Sequence Data ; Neoplasm Proteins ; analysis ; Nuclear Proteins ; analysis ; antagonists & inhibitors ; genetics ; Proteomics ; Stathmin ; analysis

BACKGROUNDCD44v6 plays an important role in invasion and metastasis of tumor, Livin has anti-apoptotic effects. The present study aimed to explore the expression and clinical significance of CD44v6 and Livin in gastric cancer tissue.

METHODSStreptavidin-peroxidase linked immunohistochemical method was used to determine the expression of CD44v6 and Livin in gastric cancer tissue and adjacent normal gastric tissues from 59 patients with histopathologically confirmed gastric cancer, and in gastric tissue specimens of 15 patients with gastric polyps, and 15 patients with chronic non-atrophic gastritis. The chi-square test was used for comparison of the relevant factors, Spearman's rank correlation test was applied for relationship among positive expression of the proteins.

RESULTSThe expresion of CD44v6 was positive in 64.4% of the gastric cancer patients; 5.1%, 0 and 13.3% in specimens of normal tissues adjacent to the cancer tissues, in gastric tissue specimens of patients with gastric polyps, and patients with chronic non-atrophic gastritis, respectively. The expression of Livin was positive in 52.5% of the gastric cancer tissues, 6.8%, 0 and 6.7% in the adjacent normal gastric tissue, specimens of patients with gastric polyps and chronic non-atrophic gastritis, respectively. The expression of CD44v6 was significantly correlated with the depth of invasion, the degree of differentiation, and lymphnode metastasis of gastric cancer (P < 0.05). The positive expression rate of Livin protein was also significantly correlated with degree of differentiation of gastric cancer cells and metastasis to lymphnodes (P < 0.05), but not correlated with the depth of invasion and pathological types (P > 0.05). The expression of CD44v6 and Livin in the gastric cancer tissue was positively correlated (r(s) = 0.286, P = 0.028).

CONCLUSIONSThe increased expression of CD44v6 and Livin in gastric cancer tissue may be closely related with development and progression of gastric cancer. CD44v6 and Livin may be new biological markers of gastric cancer.

Adaptor Proteins, Signal Transducing ; analysis ; physiology ; Aged ; Female ; Humans ; Hyaluronan Receptors ; analysis ; physiology ; Immunohistochemistry ; Inhibitor of Apoptosis Proteins ; analysis ; physiology ; Male ; Middle Aged ; Neoplasm Proteins ; analysis ; physiology ; Stomach Neoplasms ; chemistry ; metabolism ; pathology

OBJECTIVETo investigate Daxx expression and its clinical significance in children with acute leukemia.

METHODSThe expression of Daxx protein was detected by immunohistochemical assay in 50 children with newly diagnosed acute leukemia (34 cases of acute lymphocytic leukemia and 16 cases of acute non-lymphocytic leukemia). Twenty children with normal bone marrow were used as the control group.

RESULTSDaxx protein was expressed in 38.0% of 50 children with acute leukemia, which was significantly higher than that of the control group (5.0%) (P < 0.05). The children with acute non-lymphocytic leukemia had significantly higher Daxx expression levels (62.5%) than those with acute lymphocytic leukemia (26.5%; P < 0.05) as well as the control group (P < 0.05). There were no significant differences in the Daxx expression between acute lymphocytic leukemia children and the control group. Daxx protein was expressed in 55.6% of high risk group of acute lymphocytic leukemia but it was not expressed in standard risk group of acute lymphocytic leukemia (P < 0.05).

CONCLUSIONSDaxx expression is abnormal in children with acute leukemia and associated with some clinical features of acute leukemia, suggesting that it may play an important role in the genesis and development of acute leukemia.

Adaptor Proteins, Signal Transducing ; analysis ; Adolescent ; Child ; Child, Preschool ; Female ; Humans ; Immunohistochemistry ; Infant ; Leukemia, Myeloid, Acute ; drug therapy ; metabolism ; Male ; NF-kappa B ; metabolism ; Nuclear Proteins ; analysis ; Precursor Cell Lymphoblastic Leukemia-Lymphoma ; drug therapy ; metabolism

This study was aimed to investigate the aven mRNA expression level of leukocytes from peripheral blood(PB)of de novo patients with acute myeloid leukemia (AML) and analyze its clinical significance, so as to provide a experimental basis for evaluating prognosis. The aven mRNA expression levels in PB samples from 69 de novo AML patients were detected by using real-time quantitative PCR. The relation of aven mRNA level with clinical and hematological characteristics (age, sex, WBC, Hb, Plt, LDH, Blast% in PB and BM, FAB subtype) and treatment outcome (CR rate and relapse rate) were analyzed. 21 normal individuals served as controls. The results showed that the expression level of aven mRNA was between 11.72% and 178.93% (median 37.2%) in de novo AML and between 10.81% and 50.98% (median 28.81%) in normal individuals. Aven mRNA expression level was higher in the AML group than that in the controls (p = 0.006). As aven mRNA expression level was compared with other clinical and hematological parameters, there were significant correlations between aven mRNA expression level and age (r = 0.25, p = 0.039), and between hemoglobin level (r = 0.29, p = 0.019), FAB subtype(r = 0.253, p = 0.036). The median expression level (50.08%) of aven mRNA in older patients (> or = 44 years) was higher then that (32.41%) in younger patients (< 44 years) (p = 0.018). The complete remission (CR) rate after two cycles of chemotherapy in patients with lower aven mRNA level (25/30, 83.33%) was higher than that in patients with higher aven mRNA level(21/30, 70%), but the difference was not significant(p = 0.22). The difference of aven mRNA expression level between AML patients with relapse and AML patents without relapse was not significant (p = 0.076). It is concluded that the expression level of aven mRNA in de novo AML patients obviously increases, the overexpression of aven mRNA likely involves in genesis of AML. The definite relation of aven mRNA expression level with treatment outcome and relapse was not been found.
Adaptor Proteins, Signal Transducing ; genetics ; Adolescent ; Adult ; Aged ; Apoptosis Regulatory Proteins ; genetics ; Case-Control Studies ; Female ; Humans ; Leukemia, Myeloid, Acute ; genetics ; Male ; Membrane Proteins ; genetics ; Middle Aged ; Prognosis ; RNA, Messenger ; genetics ; Sequence Analysis ; Treatment Outcome ; Young Adult

PRAS40 (proline-rich Akt substrate 40 kD) associates with mammalian target of rapamycin complex 1(mTORC1), serine 183 site (Ser183) of PRAS40 can be phosphorylated by mTORC1. To prepare the phosphorylated PRAS40 (Ser183) antibody, We chosen 10-amino acid including Ser183 as antigen peptide through antigenicity and hydrophobicity analysis, hinged on keyhole limpet hemocyanin (KLH), and used the KLH-peptide to immunize rabbits. After antibody serum titer detection by enzyme linked immunosorbent assay (ELISA), the antibody was purified with rProtein A sepharose fast flow and dephosphorylated antigen membrane. The antibody titrate reached 1:10 000 after purification and its special property was enhanced with absorption treatment of dephosphorylated antigen membrane. In addition, we used rabbit anti-PRAS40 antibody and the phosphorylated PRAS40 (Ser183) antibody to detect PRAS40 expression in several cell lines, including the normal cells HL7702, HEK293, tumor cells HepG2, A549 and S180. There were no quite difference among these cells; otherwise, we observed the decreased phosphorylation level of Ser183 after amino acid withdrawal treatment. Therefore, the polyclonal phosphorylated PRAS40 (Ser183) antibody was specific to PRAS40 (Ser183) site and could be used for the function study of PRAS40.
Adaptor Proteins, Signal Transducing ; Animals ; Antibodies ; analysis ; Cell Line, Tumor ; Humans ; Male ; Mechanistic Target of Rapamycin Complex 1 ; Multiprotein Complexes ; Phosphoproteins ; chemistry ; immunology ; Proteins ; genetics ; metabolism ; Proto-Oncogene Proteins c-akt ; metabolism ; Rabbits ; Serine ; metabolism ; TOR Serine-Threonine Kinases