Dr.N.Vani,postgraduate student in the department of pathology,Coimbatore Medical College is conducting a study on “HER2neu EXPRESSION IN COLORECTAL ADENOCARCINOMA AND ITS CORRELATION WITH CLINICOPATHOLOGIC VARIABLES”. Colectomy done for various colorectal lesions are received in pathology department and are processed and examined under a microscope to obtain diagnostic information or is tested for other studies. I have been informed to my satisfaction regarding the nature of procedure. The data used herein may be used for research and publication.
Half et al studied HER2 receptor expression in colorectal cancer cell lines. They analyzed the protein expression with respect to the mRNA levels, HER2 amplification, and the clinicopathological variables. There was strong membranous staining in 5% of the primary colorectal carcinomas. Cytoplasmic staining was found in 63.5% of the primary tumours. They found a significant correla- tion between the HER2 cytoplasmic staining and the tumour differentiation . Their results were very similar to our results, especially the correlation with grade, as our grade III tumours showed 100% positive Her-2/neu staining.
I, Dr.K.KOKILA, solemnly declare that the dissertation entitled
“IMMUNOHISTOCHEMICAL EXPRESSION OF HER2NEU, P53 & P63 IN UROTHELIAL BLADDER CARCINOMA AND ITS CORRELATION WITH CLINICO-PATHOLOGICAL VARIABLES” is the bonafide work done by me at the Institute Of Pathology, Madras Medical College under the expert guidance and supervision of Prof.Dr.Geetha Devadas, M.D., DCP, Professor of Pathology and Dr.R.NARMADHA, M.D., Assistant professor of Pathology, Institute Of Pathology, Madras Medical College. The dissertation is submitted to the Tamilnadu Dr.M.G.R Medical University towards partial fulfillment of requirement for the award of M.D., Degree (Branch III) in Pathology.
Thus, a close correlation of a risk factor with HER-2/neu overexpression could indicate either that a HER-2/neu alteration is the way that this risk factor evolves into the carcinogenesis or that there is a parallel interaction between them that leads to breast tumour initiation and development. Since the data in the literature supporting the above hypothesis are few and conflicting, we decided to investigate, in a group of patients from a geographical area with a low incidence of breast cancer, whether HER- 2/neu positive tumours are correlated with established or suspected risk factors for breast cancer and thus to identify distinct subgroups of high risk women.
Methods: A total of 24 locally advanced, non-metastatic oral cancer patients were included in the study. Tumor biopsies were taken prior to the start of neoadjuvant therapy for expression of EGFR, Her-2-Neu, and GADD45 by immunohistochemistry and for HPV by PCR. The response was evaluated using Response Evaluation Criteria in Solid Tumors (RECIST) criteria after three cycles of chemotherapy. Statistical analysis was performed using correlation and Kaplan – Meier analysis; the difference in survival was calculated with log rank test.
Correlation of ER, PR and Her-2/neu with grade of tumor has been shown in Table 3. Grade 2 tumors showed maximum ER positivity (40 cases or 40%) and minimum Her-2/neu positivity.
Table 1. Age distribution of patients of invasive ductal carcinoma-NOS of breast.
Age group Cases Percentage (%)
classification of Invasive ductal carcinoma -NOS type tumour is one of exclusion: “Invasive ductal carcinoma is most frequently encountered malignant tumour, not falling into any other categories of invasive mammary carcinoma”. 17
Microscopically WHO classification requires a non-specialized pattern in over 50% of the tumour area to be classified as NOS- type. Typically combinations of infiltrative margins, trabecular, diffuse sheet like, acinar and nested patterns are noted. Histological grading of the tumour depends on tubular formation, pleomorphism and mitotic count. They are graded into well differentiated, moderately differentiated and poorly differentiated grades. 14, 15
apart from a variety of human cancers such as breast cancer, colorectal cancer, gastric, lung and prostrate tumours.
Many strategies directed against EGFR and HER-2/neu have been developed such as monoclonal anti-HER-2/neu antibodies and small molecule kinase inhibitors. There is sufficient data from the clinical trials demonstrating positive effects of applying EGFR tyrosine kinase inhibitors to oesophageal SCC and ADC. These facts indicate that HER- 2/neu is a very useful molecular marker with a range of therapeutic implications in the overall survival rates of oesophageal squamous cell carcinomas and adenocarcinomas.
In our study co-expression of both markers varied with clinical parameters and histopathological parameters. Significant association with tumor histopathological parameters was observed. Co-expression of both markers was demonstrated in 25 % of low grade, 40 % of intermediate grade and 78.8% of high grade. In muscle invasive tumors 70.6% showed co- expression as compared to non muscle invasive tumors (30.4%). Co-expression of both markers correlated well with tumor grade and muscle invasion. Evaluation of both the markers expression was more accurate in predicting aggression and progression of the tumor. Thus, Ki67 and Her2neu co- expression was a superior to single marker expression in predicting tumor prognosis. We had 3 each cases of recurrence and death on follow up 1 year which had high co-expression of both the markers. Our study failed to identify a significant difference in survival in tumors with or without both markers (Ki67&Her2neu) over expression. This limitation may be due to our shorter study duration and small sample size. Larger cohort studies may be necessary to obtain a significant correlation in the above mentioned variables.
The incidence of Invasive ductal carcinoma NOS was higher in this study. Many of our patients presented in younger age with large sized tumors accounting for aggressive nature of breast cancer in our population. Her2 molecular types were more frequent and Basal type tumors were less frequent in the present study when compared to the western literature. Her2 overexpression was common with high grade, hormone receptor negative tumors. Higher Bmi 1 gene expression was associated with premenopausal, small size, ER negative, luminal A subtype tumors in concordance to the study conducted in Korean population. 67
showed that the cells developed resistant to paclitaxel and Baselga et al. . Showed drugs inhibition need more detailed research is needed.
The synergism, summation and antagonism among Herceptin and other anti-cancer compound is not de- pends on the expression of HER-2 or the change of phosphorylation . Herceptin single drug has no cyto- toxicity in cancer cell in vitro. But by blocking its down- stream signaling pathways, to inhibit the cell survival or drug resistance gene expression to achieve synergistic ef- fect in combination therapy. According to previous re- ports, pretreatment of doxorubicin can increase the expression of EGFR or TGF-α in cells, therefore the combined with anti -EGFR antibody treatment maybe achieve synergy effect in most of cell lines including U87MG cells even that has no or little HER2 expression.
over-expression but also is indicative of the degree of receptor activation.
Ali and coworkers  and Fehm and colleagues 
examined the question of whether serum HER-2/neu reflects tumor aggressiveness or is simply a surrogate marker of disease bulk. In the report by Ali and coworkers  pre- treatment samples from 566 MBC patients were tested for serum HER-2/neu levels as well as for levels of CA15-3, a surrogate marker of tumor burden. They concluded that serum HER-2/neu is a significant independent predictive and prognostic factor in hormone receptor positive MBC patients, even when adjusted for tumor burden using CA15-3. In addition, Fehm and colleagues , using a multivariate analysis, also concluded that when serum HER-2/neu results were adjusted for tumor load with CA15-3, serum HER-2/neu remained an independent marker of tumor aggressiveness and reflected the biologic behavior of the tumor. Several other studies have also concluded that the combination of serum HER-2/neu and CA15-3 identifies a subset of patients with a worse prognosis when compared with one marker alone. However, it has also been reported on several occasions that there is a weak correlation between serum HER-2/neu and CA 15-3, but each provides important information about the biology of the tumor.
Gastric carcinoma is one among the top five causes of cancer in both males and females. It is also listed among the top cancer related mortalities. Despite the advancements in the treatment of gastric cancer, none has shown to improve the survival significantly. There was no additional improvement in overall survival with addition of Epirubicin or Docetaxol to Cisplatin and 5FU combinations although the response rate was better. The role of targeted therapy has recently come into play after the establishment of the role of HER-2/neu over-expression in gastric malignancies. Trastuzumab ,an anti HER-2/neu antibody ,in combination with Cisplatin and 5FU in advanced gastric carcinoma patients has shown to improve overall survival in TOGA trial This study was undertaken to find out the prevalence and the clinicopathological correlation in HER-2/neu over –expression in gastric carcinoma patients.
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.
Amplification of the gene coding for HER-2/ neu has been described in breast, ovary, prostate, gastric, salivary gland, lung, colon and squamous cell carcinomas. Over all, 25% to 30% of primary breast cancers and about one third of primary ovarian cancers demonstrate amplification of the gene. These patients appear to have an increased rate of recurrence and decreased survival.[3,4].The amplification of the HER-2/neu gene is accompanied by increased expression of the protein gene product, which can be detected in frozen or paraffin section by immunohistochemical techniques (IHC) . There is near uniform correlation between gene amplification and IHC reactivity for HER-2/neu on frozen or paraffin sections, such that IHC has became the standard method for analysis for HER-2/ neu gene amplification . When over expressed, the protein accumulates at the cell membrane and is seen as a crisp membrane stain .
Previous studies on HER-2 expression in bladder carcinoma using Western blot analysis and IHC have found a correlation between increased HER-2 expression and both higher tumor stage and grade ( 20,21).but our study contradicted these studies, we found no significant correlation between HER-2 over-expression and histological subtype of bladder cancer (TCC, SCC) (p=0.55), and also no significant correlation between HER-2 over expression and tumors grade (P=0.06). Others (16, 19, 22) have reported a significant association between HER-2 over-expression and a higher tumor grade. Thus, shawky et al. ( 23) studied HER-2 over-expression in invasive bladder carcinoma among cohort patients. HER-2 over-expression was associated with high-grade bladder cancers. Similarly, in a recent study ( 24) on 59 patients, HER-2 over-expression was significantly correlated with the differentiation grade, our study contradicted for these studies;
Objective: T o study the clinical pathologic characteristics of β -catenin, K i 67 and H er- 2 /neu in gastric cancer and the correlation of β -catenin and K i 67 to the protein expression and gene conditions of Her-2/Neu. Methods: T he protein expression of β -catenin, K i 67 and H er- 2 / N eu was detected by immunohistochemistry in 101 cases of gastric cancer and the gene conditions of Her-2/Neu by fluorescence in situ hybridization ( FISH ) . Results: T he protein expression of β -catenin, K i 67 and H er- 2 / N eu had close relationship with the clinical pathologic characteristics of gastric cancer. T he β -catenin and K i 67 had obvious correlation to the differentiation, infiltration and lymphatic metastasis of the gastric cancer ( P< 0 . 05 ) . T he K i 67 had close relationship with the tumor-node-metastasis staging staging of gastric cancer ( P< 0 . 05 ) . H er- 2 / N eu had close relationship with the differentiation and tumor-node-metastasis staging of gastric cancer ( P< 0 . 05 ) but had no relationship with the infiltration and lymphatic metastasis of the gastric cancer ( P< 0 . 05 ) . T he protein expression of K i 67 had significantly positive correlation to the protein expression and gene amplification conditions of H er- 2 / N eu ( r= 0 . 567 , P< 0 . 05 for protein; r= 0 . 304 , P< 0 . 05 for gene ) . Conclusions: C ombined detection of β -catenin, K i 67 and H er- 2 / N eu can be used as a reliable method to help the observation of biological behavior, diagnosis and prognosis of gastric cancer, and K i 67 can be used to serve the preliminary screening of Her-2/Neu gene state.
Becker 18 and Kameda et al. 19 described as no Her-2/Neu gene amplification was found in diffuse type gastric cancer and they found 20% of amplification in intestinal type. Correlation is high between HER2 expression - intestinal type had been reported by many authors in late 20 the century . In Finnish study,associtation of her2/neu amplification with poor carcinoma-specific survival is strong particularly more in intestinal type subgroup of cancers , but the contradicting fact is it should have better prognosis than diffuse type of gastric carcinoma.  the same also reported in a Korean study.  Finally, in ToGA trial, HER2 positivity varied by histological subtype (intestinal 34%, diffuse 6%, mixed 20%). But in our study we found no HER2/NEU positivity in intestinal type of adenocarcinomas and we found only in diffuse type carcinomas (p<0.05). All the three had a poor prognosis and two did not respond for chemotherapy as well.
At least three mice from each group were tested including the control tumors. We clearly see more mutations occur- ring in Her-2/neu from tumors from treated versus untreated control mice (Table 1). Only two residues were mutated in tumors from control vaccinated mice and neither of these occurred in known CTL epitopes. The Flk-E1 vaccinated mice show more mutations in the extracellular fragment than the intracellular fragment, however, Flk-11 show mutations in both fragments. Also, the Flk-11 mutations seem to occur at a higher frequency in the region between residues 1026 and 1051 and several mutations overlap with mutations previ- ously identified from Her-2/neu directed vaccinations. 11 We do not see any correlation between frame shift, substitutions, deleted residues, and vaccination. It is possible that certain regions of the Her-2/neu molecule are susceptible to mutation or sensitive to selective pressure, such that certain mutations do not impact on signaling and tumor growth but do provide a selective advantage for survival.
The usage of EHBC-TMA in our study allowed the entire cohort, NNB and EHBC tissues, to be analyzed on one slide, and the benefits of using TMA are well documented. 11 A known 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 staining or no membranous staining of tumor cells was scored as “0”; 2) tumor cells with faint membrane staining irrespective of the percentage of tumor cells were scored as “1 + ”; 3) tumor cells with weak to moderate membrane staining irrespective of the percentage of tumor cells were scored as “2 + ”; and 4) tumor cells with strong complete, basolateral, or lateral membrane reactivity irrespective of the percentage of tumor cells were scored as “3 + ”. 14 We used the presence of complete, basolat- eral, or lateral membranous reactivity in ≥ 10% of cells as the cutoff for positivity, as required by the published guidelines. 14 The Her-2/neu IHC score was calculated by two independent pathologists (RS and DC) and a consensus score was reached for each specimen.