are no major side effects with Trastuzumab, it is ideal that all patients with gastriccarcinoma are tested for HER-2/neu and be given the benefit of it. The laboratories in the hospital should be well equipped with availability both IHC and FISH to test HER-2/neu gene. Patients may develop pimary and acquired resistance to Trastuzumab due to crosstalk between pathways. Research should also be done with combination of targeted therapies to overcome this resistance to improve the outcome of these patients. Beyond all, only a quarter of gastric cancer patients will have amplification of HER-2/neu gene leaving behind the rest 75% of patients without any benefit. Therefore it is time to concentrate on these patients and find out an effective chemotherapeutic or targeting agents which can really change the outcome of gastric cancer treatment. Also more trials are necessary in adjuvant setup to improve the disease free survival in patients who presented at an early stage though equal importance has to be given to prolong the progression free survival of patients who present in advanced disease. Each country should try to identify if any epidemiological factors are influencing the cause and take steps to get rid of those causative agent which will enable us to decrease the incidence of this disease.
In this study, c-Met protein expression was determined by IHC and scored by both membranous and cytoplasmic stain- ing. In our study, c-Met overexpression was 58.7% in 125 patients. The literature reports a difference in the frequency of c-Met overexpression in gastric cancer. This difference may be caused by the different methods used for c-Met expression. In our study, we used both membranous and cytoplasmic staining for c-Met expression, and accordingly, we have created a scoring system as given in Table 1. Some studies used only cytoplasmic staining, only membranous staining, Table 4 Relationship between clinicopathological factors and
survival (113) . The frequency of EGFR over expression and ⁄ or amplification in Gastriccarcinoma has been variously reported as ranging from 18 to 44%. (114-116). Recently, the new therapeutic agents targeting EGFR has attracted attention. They either bind to extracellular domain of the receptor (eg:cetuximab, matuzumab and panitumumab), where they compete with the natural ligand binding to the receptor and block activation of that receptor or they compete with adenosine triphosphate binding to the tyrosine kinase portion of endodomain of the receptor and they abrogate the receptor‘s catalytic activity (eg: gefitinib, erlotinib and lapatinib). Both groups of compounds are effective in blocking the downstream receptor-dependant signaling pathway. For patients with gastric cancer, a few clinical trials have been performed with these drugs but with ambiguous results. One clinical trial has found that a subpopulation of Gastriccarcinoma patients showed evidence of gefitinib sensitivity. Accurate evaluation of the EGFR protein and ⁄ or EGFR gene expression status in patients with Gastriccarcinoma is important for determining patient‘s eligibility for new targeted therapy.
Objective: HER-2/neustatus in gastric carcinomas (GCs) has not been studied before in the Egyp- tian population. Materials and Methods: Seventy-three GCs were evaluated for the expression of HER-2/neu and Ki-67 using immunohistochemistry (IHC). Fluorescence in situ Hybridization (FISH) was done for HER2/neu IHC score 2+ cases. Results: Out of the 73 gastric carcinomas, 23 (31.5%) were score 0, 17 (23.3%) were score 1+, 23 (31.5%) were score 2+, and 10 (13.7%) were score 3+. FISH analysis revealed that the HER-2/neu gene was amplified in 11 out of the 23 cases (47.8%) scored 2+ by IHC. Therefore, the overall HER-2/neu positivity rate was 28.8% and it was signifi- cantly associated with higher T-stage and lymphovascular invasion (LVI). The Ki-67 expression rate ranged between 10% and 100%, with 84.9% of the cases (n = 62) featuring high Ki-67 scores. High Ki-67 score was significantly associated with male sex, tumor grade, and number of positive nodes. HER-2/neu protein expression correlated significantly with Ki-67 score. Twenty tumors showed combined HER-2/neu positivity and high Ki-67 score and were significantly associated with higher T-stage and occurrence of LVI, implying a more aggressive behavior. Conclusions: The rate of HER-2/neu positive GCs in our series simulates universal rates, thereby mandating routine evaluation of HER-2/neustatus in all GCs submitted to our laboratory to benefit from trastuzumab therapy. Further studies on a larger number of GCs are required to prove that the concurrent HER-2/neu positivity and high Ki-67 score are markers of worse prognosis.
bersome and more economical; the signal intensity is permanent; and pathologists are familiar with IHC signals and are able to correlate findings with the underlying tumor histomorphology. CISH allows observers to select fields of invasive ductal carcinoma, avoiding foci of intraductal car- cinoma for which Her-2/neu has a different clinical signifi- cance. CISH is a specific, sensitive, and easily applicable method for the detection of Her-2/neu gene amplification, and it can be used together with IHC for the evaluation of patients with breast carcinoma . Ross and colleagues recently reviewed the current status of Her-2/neu testing, and they concluded that CISH is a more convenient gene- based technique that is 'waiting in the wings' . In this present study, the concordance between 3+ IHC and CISH-amplified cases was 100% (40 of 40), denoting all gene amplified cases to be overexpressing the Her-2/ neu protein. In contrast, the agreement on 2+ IHC and CISH-amplified cases was only 45%, which is lower than previously reported (93%) . The 13.75% (22 of 160) IHC-positive/CISH non-amplified tumors in this study, all of which had 2+ IHC scores, is higher than the 6% false pos- itive result by Zhao and colleagues , although compa- rable to the 17% result (IHC CB11 versus FISH) obtained by Tubbs and colleagues . Apparently, we find that the 2+ IHC score is equivocal when compared with CISH. This subset might benefit from further CISH reflex testing. All negative IHC cases were CISH non-amplified, showing complete agreement of all negative results. The 86.25% overall concordance between IHC (using CB11 antibody) and CISH in this study is lower than the previous reports (94–96%) [24,30]. We concur with Sapino and colleagues that gene amplification analysis can be avoided in all 0, 1+, and 3+ IHC cases because results are predictable from IHC and are completely concordant with CISH . The utility of CISH is therefore in further testing Her-2/neu alter- ation on equivocal cases in IHC, which remains the primary screening method. The combined IHC–CISH algorithm provides an economical and comprehensive Her-2/neu data to guide clinicians in mapping treatment options. The low-amplified CISH category (6–10 signals) was the most difficult to interpret, requiring an accurate enumera- tion of gene copy. Signal clustering, more probably a result of intrachromosomal amplification of homogeneously stain- ing regions, was immediately apparent in highly amplified cases and was easily evaluated [23,31].
Gastriccarcinoma is the second common cause of cancer related worldwide deaths. The HER-2/neu is the Human epidermal growth factor receptor-2 and it is a gene localized on chromosome 17q21 that encodes a growth factor receptor like molecule with tyrosine kinase activity and has a structure similar to that of epidermal growth factor receptor. Its expression has been detected in several human cancers and is believed to be associated with poor prognosis, aggressive biological behavior and metastatic potential. The present study is to evaluate the presence of HER-2/neu in gastric cancers of all histological types. IHC is an excellent technique to detect HER-2/neu expression in Gastriccarcinoma. Their prevalence in gastric cancer ranges from 9-38% in most studies. This study included 50 cases of gastriccarcinoma and these were subjected to immunohistochemical staining for HER-2/neu oncoprotein. Out of 50 cases , only 8 cases of Gastric adenocarcinoma (16%) were found to be positive for HER-2/oncoprotein. In this study Her2 neu expression is more in Oesophageal gastric junctional tumours & corpus location of stomach tumours (35.3%) compared to Pylorus-Antral tumours. And also we observed Her2 neu expression in patients with signet ring cell carcinoma (40%) compared to Non signet ring cell carcinoma.
Background: Gastriccarcinoma is a deadly disease with high mortality. A better understanding of the molecular basis of gastric cancer has contributed to the development of rationally designed molecular targeted therapies which will improve the survival rate. A genetic alteration that could help in targeted therapy and prognostication includes Human Epidermal Growth Factor Receptor 2 (HER2/neu) overexpression in gastriccarcinoma. The objective of the present study was to identify and evaluate the HER2/neu protein immunohistochemical expression in gastric cancer from biopsies and surgical resection specimens and to evaluate their correlation with histopathological features. Methods: Total/subtotal gastrectomy specimens and gastric biopsies from a tertiary care center in South India were included in the study and assessed by light microscopy and Immunohistochemistry (IHC).
HER-2/neu over-expression or mutation results in quantitative and qualitative alteration in the membrane proteins which is the basis of its detection. , Even though clinical correlation had not been established, many series had suggested that amplification or over-expression of the oncogene might be a marker of poor prognosis in cancers of the ovary, endometrium, breast etc. ,, There were reports in the literature that HER-2/neu over-expression correlates with reduced benefit of adjuvant therapy as in cases of carcinoma breast with tamoxifen therapy. Many studies have shown that regression of HER-2/neu suppresses the malignant phenotypes of cancer cells over-expressing this oncoprotein, which may serve as an excellent target for developing anti-cancer agents specific for HER-2/neu over- expression.  HER-2 expression can be estimated either by immunohistochemistry or by fluorescent in situ hybridization (FISH). FISH can be used for doubtful cases in IHC to confirm, HER2/neu expression. This combination is not necessary for low (0-1+) or high (3+) grades of immunohistochemical stain as the correlation with gene amplification status is high.
Her-2/neu-positive breast carcinoma was used as a positive control. For negative control, the primary antibody was omitted during the incubation step. The EHBC-TMA was incubated for 16 min with a Her-2/neu rabbit monoclonal antibody (clone 4B5, using the Benchmark XT; Ventana Medical Systems) and detected by ultraview universal DAB detection kit (using the Benchmark XT; Ventana Medical Systems, Inc.). Antigen retrieval was performed using the Ventana Cell Conditioner #1 for 32 min. The Her-2/neu protein expression was evaluated using the modified ToGA trial scoring criteria established for gastric and gastroesophageal carcinomas 12–14 as follows: 1) no
A role for CREB in the development of tumors has been suggested since it is often overexpressed and more active in many solid and hematopoietic tumors [23-26]. This could be influenced by the modulation of signalling cascades upstream of CREB as well as the induction of the CRE-dependent gene expression downstream of CREB, e.g. matrix metalloproteinases (MMPs), adhesion molecules and different survival factors [20, 27] leading to an increased tumor growth, prevention of cell death, enhanced metastasis formation and angiogenesis . Furthermore, overexpression of CREB in tumors often correlates with disease progression, poor patient survival and chemotherapy resistance [21, 24-26, 29-31]. On the other hand oxygenation in breast cancer is markedly reduced compared to normal tissue . In mammary carcinoma the CREB pathway is often upregulated [33, 34]. An important role of CREB in breast cancer has been further demonstrated by global profiling of signalling networks in breast cancer stem cells . Recently, a correlation between CREB expression/ activation, altered in vitro and in vivo tumor growth properties and HER-2/neu has been described in both in
Immunohistochemistry was performed on formalin-fixed paraffin blocks which cut at a thickness of 2 μm. These sections were deparaffinized in xylene followed by hy- dration in a graded series of alcohols. Sections were left under running water for 15 min. Endogenous peroxidase activity was blocked by incubation in 3% hydrogen per- oxide for 30 min at room temperature. After rinsing in TBS buffer (pH 7.4) for 30 min, the sections were incu- bated with primary antibody at 4 °C overnight. The fol- lowing primary antibodies were used: EGFR (SantaCruz Biotechnology, Santa Cruz, CA), GADD45A (Abcam, Cambridge, MA), and Her-2-neu (Biogenex). Slides were then washed again in TBS. Secondary antibody was added, and slides were washed again after 30 min. The labeled antigen–antibody complexes were visualized as brown pigments via a standard DAB (Zhongshan Jin- qiao Co., Beijing, China) protocol. DAB was added and kept for 10 min until brown color developed. Slides were washed in running water, and counterstaining was done with hematoxylin. Slides were then blotted, dried, and mounted.
Breast cancer is the most common cancer of women in developed countries and 12% breast cancers occur be- tween 20 - 34 years . In India, breast cancer is second to cancer of the cervix among women, but is consi- dered the leading cancer in certain metros such as Mumbai and Bangalore. It is estimated that approximately 80,000 cases occur annually. Breast cancer is strongly related to age, with only 5% of all breast cancers occur- ring in women under 40 years old . Several pathological features have prognostic significance in breast car- cinoma which includes histologic subtypes, grade, lymph node status, estrogen receptor (ER)/progesterone re- ceptor (PR) status, growth factor and its receptors, proliferation activity and DNA content with tumor suppressor genes and oncogenes . Prognosis of breast carcinoma is worse with higher grade, tumor subtypes like medul- lary and lobular carcinoma, lymph node metastasis, negativity for ER, PR and positivity for Her-e/neu, with excess of oncogenes and also presence of BRCA gene. Out of these, ER, PR and Her-2/neu immunostaining are a very useful tool for rapid diagnosis, treatment and prognosis of the patient. Determination of ER, PR status on biopsy specimens prior to therapeutic intervention is advocated as a standard practice . Due to this, breast cancer is better represented by its combined receptor expression than by a single receptor status alone. Recent management protocol is focusing on separating triple negative and triple positive tumors for treatment and prognosis. With this background, the present study is undertaken to analyze the pattern of expression of hor- mone receptor ER, PR and Her-2/neu in invasive breast carcinoma and to correlate them with various clinicopa- thological parameters, especially in this part of country where studies are limited.
The observation that morphologically similar neoplas- matic lesions of the breast can exhibit different biology has necessitated the identification of biological parame- ters that might improve risk assessment; the evaluation of HER-2/neu expression is a typical example . Indeed, several studies have demonstrated that HER-2/neu ampli- fication represents a prognostic and predictive marker; its expression is associated with early disease recurrence, rel- ative resistance to chemo- and/or hormonotherapy and short survival [2,10]. In addition it has been shown that genetic alterations of the HER-2/neu oncogene represent early events involved in breast carcinogenesis and tumour initiation, while their presence is observed in all stages of malignant development from in situ carcinomas to meta- static lesions . As a result, some researchers have maintained that HER-2/neu amplification and/or protein overexpression may also represent not only an important marker of prognosis but also a key indicator of the aetio- logical heterogeneity of breast carcinogenesis. [3,7-9]. On the other hand, the contribution of even well estab- lished breast cancer risk factors to the aetiology of carcino- genesis in the breast remains obscure, ill-defined and tenuous, mostly because of the existence of different path- ways for the initiation and the evolution of a breast tumour . In order to explain this incompatibility, sev- eral researchers have claimed that classification of tumours on the basis of HER-2/neu overexpression or amplification may define a subset of breast cancer in which the net effect of a risk factor could be rather more obvious and its impact on breast cancer development more clear [3,7,8].
The combinatorial treatment complicated our attempts to detect vaccine-induced Her2-specific anti- bodies in the vaccinated patients. However, a recently established l-subclass specific ELISA allowed evaluation of endogenous Her2-specific antibody responses without detection of or interference by the IgG1 antibody tras- tuzumab . Notably, the majority of evaluable patients demonstrated increased antibody binding activ- ity after completion of the vaccine trial and in most of the long term survivors these endogenous Her2-specific antibodies persisted or were increased in samples obtained several years after the last vaccine administra- tion. Due to co-administration of trastuzumab we were not able to evaluate the contribution of endogenous Her2-specific antibodies of the -subclass to the overall humoral immune response. Considering previous vac- cine trials  it is likely that IgG antibodies were also induced.
Expression of estrogen receptors and progesterone receptors within tumors correlates well with low histologic grade and responsiveness to hormone manipulation, especially in postmenopausal patients . No much correla- tion exists between the cytoarchitectural type of breast carcinoma and presence of hormone receptor protein, no statistically significant difference has been found between ductal type and lobular type tumors . Because the growth of breast cancer is often regulated by the female sex steroids, the determinations of the cellular concen- trations of ER and PR in the tumor are currently used to predict which patients are of good prognosis as they may also benefit from antihormonal therapy . The ER cannot be clearly classified as an oncoprotein or tumor suppressor protein, although it clearly mediates onset and progression of the disease. In striking contrast to its normal function in ER—positive cell lines, ER expressed in ER—negative cell lines functions to suppress cell growth, in spite of its normal action to regulate expression of certain hormonally responsive genes, thus the mul- tiple differences between ER positive and ER negative breast cancer appear to include incompatible growth reg- ulatory mechanisms . However, there is only 8% to 20% absolute difference in disease free survival between women with ER—positive node negative invasive breast carcinoma and those with ER—negative disease and some studies have shown that any survival advantage of ER—PR positivity is lost after 5 years of follow up . More than 60% of breast cancer patients are ER—positive tumors, and respond to endocrine therapy. In addi- tion, 5% to 10% of the patients designated ER—negative appear to initially respond to endocrine therapy . To improve the value of determination of the ER for tumor prognosis, the presence of the estrogen—regulated PR protein is now routinely determined. In many breast tumor cell lines and in normal, ER containing tissue, such as endometrium and brain, PR expression also has been shown to be positively regulated by estrogen .
The expression of HER-2/neu marker was examined by immunohistochemical staining by Envision method using Hercep Test Kit (Dako, Denmark) according to manufacture's instructions. Briefly, first, 3-4 microns sections of paraffin blocks were prepared and placed on sinalized slides. Then deparaffinisation and rehydration were performed and antigenic recovery was performed in a microwave oven in the presence of citrate buffer. After antigen retrieval, peroxidase was blocked to avoid endogenous peroxidase activity. Then staining with DAB chromogen followed by counterstaining with hematoxylin were performed (15). Also, a sample of breast carcinoma with over- expression of HER-2/neu was used (positive control) during coloration. Then, the slides were scored based on HercepTest Scoring in Dako instruction for gastric biopsies. Zero score was for the cases where the tumor cells or membrane were not stained. +1 score was for weak and diffuse membrane staining of tumor cell clusters (at least 5 cells). +2 score was for weak to moderate, lateral or basolateral complete membrane staining of tumor cell clusters; and+3 score was for strong, lateral or basolateral complete membrane staining of tumor cell clusters (Figures 1-4).
“. .. the gastric volume was smaller, its anterior surface is normal and on the right side there is a close adhesion with the inferior left liver and perforated in the center. The perforation was sealed by the liver adhesion. On opening stomach,over Greater curvature its capacity appeared filled with a Coffee coloured liquid. The interiorof the stomach was a cancerous ulcer whose center was near the lesser curve and the induration spread from the cardiac to pylorus, with a scirrhous thickening of the wall.”
In addition to new classes of cytotoxic agents, there is also interest in the use of oral fluorinated pyrimidines. These are FU prodrugs that offer the ease of oral administration and mimic the benefits of long-term infusional therapy. Different mechanisms of action offer another advantage. Three fluorinated pyrimidines have been studied in gastric cancer as single agents: UFT, S1, and capecitabine. In most studies, the overall response rate for all three drugs is approximately 20% to 30%, similar to the rate reported in the past for FU. Whether the oral fluoropyrimidines are equivalent to infusional FU in gastric cancer is not yet clear.
Sections were immersed in citrate and tris/ EDTA buffers and processed in a water bath by boiling for 30 min for E-cadherin and HER-2/ neu. The boiling time for hMLH1 and hMSH2 was 60 min with subsequent incubation in an autoclave (15 min at 120°C). After washing, sections were treated by the streptavidin-bio- tin complex method using a DAKO Universal LSAB kit (DAKO, Denmark) with diamino- benzidine tetrahydrochloride colorization. In the evaluation of hMLH1 and hMSH2, the dysplastic glands were regarded as negative only when stromal cells or lymphocytes in the adjacent area of dysplasia showed a positive nuclear reaction. For MSH2 staining, slides were only analyzed when the cytoplasmic re- action did not disturb evaluation of the nuclei. We used the chi-square test to analyze statis- tical differences between LG, HG and ID for biomarker gene expression and a p-value of <0.05 was considered significant.
An important member of the epidermal growth factor re- ceptor (EGFR) family, the proto-oncogene HER-2/neu en- codes a 185-kD transmembrane glycoprotein with tyrosine kinase activity . HER-2/neu over-expression typically oc- curs in the placenta, embryonic epithelial tissue, and several types of tumor cells. In contrast, HER-2/neu is absent or minimally expressed in normal tissues . The positive ex- pression rate of the HER-2/neu protein in endometrial car- cinoma is associated with clinical staging, a lower degree of tissue differentiation, and lymph node metastasis . We have applied RT-PCR and ELISA to detect the expression of HER-2/neu, COX-2, p450arom and PGE2 in normal endometrium, hyperplasia endometrium and endometrial carcinoma respectively. The results showed that the expres- sion of HER-2/neu was significantly correlated with patho- logic grading, FIGO staging, and lymph node metastasis. But it has no correlation with menopausal status . There are some studies also shows that the HER-2/neu gene con- tributes to the progression of carcinomas and tumor resist- ance to chemotherapy [9-11]. A better characterization of this proto-oncogene can lend insight to the pathogenesis and molecular mechanisms involved in the development of endometrial carcinoma.