Breast Neoplasms ; genetics ; DNA Probes ; Female ; Genes, erbB-2 ; Humans

Animals ; Genes, BRCA2 ; Genes, erbB-1 ; Genes, erbB-2 ; Genes, p16 ; Genes, p53 ; Genes, ras ; Genetic Predisposition to Disease ; genetics ; Humans ; Pancreatic Neoplasms ; classification ; genetics ; pathology

Breast Neoplasms ; chemistry ; genetics ; Chromogenic Compounds ; Female ; Genes, erbB-2 ; Humans ; In Situ Hybridization ; methods ; Receptor, ErbB-2 ; analysis

To purify the extracellular domain of HER2 in vitro and improve its prokaryotic expression abundance, the cDNA fragment encoding extracellular domain of HER2 was obtained by PCR and cloned into the expression vector pGEX-6P-1. After transforming it into Escherichia coli BL21, we instituted an investigation of different inducing conditions to try out the optimal condition for expressing soluble fusion protein. As for insoluble inclusion bodies, they were dissolved in 8 M Urea and refolded in refolding buffer. The soluble protein and the refolded protein were purified with Glutathione Sepharose 4B, respectively. The results showed that both the soluble and insoluble protein existed in Escherichia coli, but the majority was insoluble. It is beneficial to the expression of soluble fusion protein by induction at lower temperature (30 degrees C) and higher optical density (A600= 1.8) with the use of certain additive in medium. By purification of the supernatant of the lysate and refolded protein, the yield of the fusion protein was about 1.23 mg per liter culture. As a result, we have obtained the maximum soluble extracellular domain of HER2 protein, and thus have laid a foundation for further work on functional study and antibody preparation for HER2.
Cloning, Molecular ; Escherichia coli ; genetics ; Genes, erbB-2 ; genetics ; Humans ; Prokaryotic Cells ; metabolism ; Protein Folding ; Receptor, ErbB-2 ; biosynthesis ; genetics ; Recombinant Fusion Proteins ; biosynthesis ; genetics ; isolation & purification

OBJECTIVETo construct a plasmid carrying small interfering RNA (siRNA) targeting human C-erbB2 gene (pGenesil- erbB2) and test its effect on Her-2 expression at the post-transcriptional level in human colon cancer cell lines HT-29 cells that highly express erbB2.

METHODSA HT-29 cell line that highly expressed CerbB-2 was selected using immunohistochemical method. The double-stranded siRNA targeting human CerbB-2 cDNA and the negative control fragment were cloned into pGenesil-1 vector, and after identification and sequence analysis, the constructed pGenesil-erbB2 plasmid was transfected into the selected HT-29 cell line.

RESULTSThe pGenesil-erbB2 plasmid was successfully constructed and stably transferred into HT-29 cells. The transfection resulted in significant inhibition of Her-2 protein expression in the HT-29 cells, as shown by Western blotting.

CONCLUSIONThe pGenesil-erbB plasmid we constructed can be stably transfected into HT-29 cells to inhibit the expression of Her-2 protein, and can be useful in further studies of increasing the radiosensitivity of HT-29 cell lines.

Base Sequence ; Genes, erbB-2 ; genetics ; HT29 Cells ; Humans ; Molecular Sequence Data ; Plasmids ; genetics ; RNA Interference ; RNA, Small Interfering ; Receptor, ErbB-2 ; biosynthesis ; genetics ; Transfection

Genetic damages are frequently found in both tumor and normal cells at carcinogen exposed areas in the patients with upper aerodigestive tract cancer. These phenomena are explained by the multistage process and/or field cancerization theories. The c-erbB-2 proto-oncogene has been amplified in many human tumors including breast, stomach, kidney and lung cancers. To study the possible evidence of multistage process and/or field cancerization in the development of gastric adenocarcinoma, the amplification statuses of c-erbB-2 proto-oncogene using the Southern hybridization technique were evaluated at the 45 gastric adenocarcinoma specimen sets consisting of tumor tissue, adjacent normal tissue (within 2 cm of the primary tumor), metastatic tissue and normal stomach tissue (at least 5 cm away from primary tumor). As a result, c-erbB-2 proto-oncogene at 2 specimen sets (4.4%) was amplified 2- to 4-fold to normal control status. In these 2 cases, c-erbB-2 proto-oncogene at histologically normal tissue adjacent to tumor tissue was amplified. And, the metastatic tissue of 1 case also exhibited c-erbB-2 proto-oncogene amplification of which the degree was less than that of tumor tissue. From these results, we were able to suspect that c-erbB-2 proto-oncogene amplification in the normal tissue adjacent to tumor tissue could be a biomarker of premalignant changes in a small proportion of gastric adenocarcinoma patients. And, this result might suggest the possible role of multistage process and/or field cancerization in the development of gastric adenocarcinoma.
Adenocarcinoma/secondary ; Adenocarcinoma/genetics* ; Blotting, Southern ; Cell Differentiation/genetics ; Gene Amplification ; Genes, erbB-2* ; Human ; Reference Values ; Stomach Neoplasms/genetics*

In order to explore a potential indicator of predicting the occurrence and development of gestational trophoblastic tumor, the expression of c-erbB2 oncogene in human normal placenta, hydatidiform mole and choriocarcinoma was investigated. The expression of c-erbB2 was detected immunohistochemically by monoclonal antibody against the gene on the formalin-fixed paraffin sections of 21 hydatidiform moles, 21 invasive moles, 20 choriocarcinomas and 30 normal placentas. Results showed that the expression level of c-erbB2 was significantly higher in gestational trophoblastic tumor than in hydatidiform mole and normal placenta of midterm and term pregnancy (P < 0.05), while there was no significant difference between patients with gestational trophoblastic tumor of stage III, IV and those of stage I, II. It was demonstrated that overexpression of c-erbB2 may closely associated with malignant transformation of hydatidiform mole, not only providing important insight into pathogenesis of gestational trophoblastic tumor, but also having an important significance for the early diagnosis and early treatment of gestational trophoblastic tumor.
Biomarkers, Tumor ; Choriocarcinoma ; metabolism ; Female ; Genes, erbB-2 ; Humans ; Hydatidiform Mole ; metabolism ; Hydatidiform Mole, Invasive ; metabolism ; Placenta ; metabolism ; Pregnancy ; Receptor, ErbB-2 ; biosynthesis ; genetics ; Uterine Neoplasms ; metabolism

OBJECTIVETo evaluate the clinical value of HER2 overexpression in breast cancer and its prognostic implication in patients with lymph node negative breast carcinoma.

METHODSThe following electronic database were extracted using appropriate inclusive and exclusive standards: Cochrane library, PUBMED, Embase (1984 - 2003), OVID, CMCC and CNKI. Excel and RevMan 4.2 were used for statistical analysis.

RESULTSFifty-six articles were extracted to calculate the positive rate of HER2 overexpression. The pooled positive rate was 23.14% [19.54%, 26.73%], with positive immunohistochemistry (IHC) rate of 23.13% [19.49%, 26.77%] and positive FISH rate of 20.90% [15.54%, 26.25%]. Seven articles were used to evaluate prognostic predication of HER2 expression. It was concluded that in patients with lymph node negative breast carcinoma, HER2 overexpression (both IHC and FISH) independently predicted a poor prognosis based on disease-free survival (DFS) and overall survival (OS) with a P < 0.05. For DFS, the pooled RR was 1.38 [1.07, 1.80] with 1.16 [1.02, 1.31] for IHC and 1.98 [1.56, 2.52] for FISH. For OS, the pooled RR was 1.58 [1.16, 2.14] with 1.37 [1.14 to 1.64] for IHC and 2.33 [1.45 to 3.75] for FISH. HER2 overexpression effectively predicted DFS/OS of patients without adjuvant therapy and OS of patients with the therapy, but not for DFS, with the pooled RR of 1.46 [1.02, 2.09] and 1.11 [0.95, 1.31] for DFS, respectively and the pooled RR of 1.93 [1.44 to 2.58] and 1.25 [1.01, 1.56] for OS, respectively.

CONCLUSIONSIn patients with lymph node negative breast carcinoma, the positive rate of HER2 overexpression is 23.14%. HER2 overexpression indicates a poor prognosis and adjuvant therapy after surgery should be recommended.

Breast Neoplasms ; genetics ; metabolism ; pathology ; therapy ; Chemotherapy, Adjuvant ; Disease-Free Survival ; Female ; Genes, erbB-2 ; Humans ; Lymph Nodes ; pathology ; Mastectomy ; Prognosis ; Receptor, ErbB-2 ; metabolism ; Survival Rate

Breast Neoplasms ; genetics ; Carcinoma, Ductal, Breast ; genetics ; Female ; Genes, erbB-2 ; Humans ; In Situ Hybridization ; methods ; In Situ Hybridization, Fluorescence ; Silver

Breast Neoplasms ; blood supply ; classification ; genetics ; pathology ; Female ; Genes, BRCA1 ; Genes, MDR ; Genes, erbB-2 ; Humans ; Lymphatic Metastasis ; Mutation ; Neovascularization, Pathologic ; Sentinel Lymph Node Biopsy