Mohamad Nidal Khabaz

Association of Epstein-Barr virus infection and breast carcinoma

Introduction A controversy regarding the association of Epstein-Barr virus (EBV) with breast carcinomas has recently been reported in the literature. The present study was carried out in an attempt to determine whether there is a relationship between latent infection with EBV and breast carcinomas in Jordanian females. Material and methods Extraction of DNA from the archive samples of breast carcinoma cases embedded in paraffin wax was performed and the extracted DNA was subjected to polymerase chain reaction amplification to detect the EBV genome using four sets of primers for EBER 2, BNLF-1, EBNA 2, and Gp220. Immunohistochemistry study was performed on sections of 4 µm which were cut from paraffin blocks of tumor and control groups. Monoclonal antibody against EBNA-1 was applied to all slides to identify the EBV-infected tumor cells. Detection was performed using the Dako envision dual link system. Results DNA was successfully extracted from 92 paraffin embedded samples of breast carcinoma patients, and from 49 normal samples. The extracted DNA was confirmed by using glyceraldehyde-3-phosphate-dehydrogenase (GAPDH) primers. Twenty-four out of 92 breast carcinoma specimens was found to be infected with EBV as compared to 3 out of 49 control group specimens, which represented a statistically significant difference (p-value using χ2 = 0.008). Immunohistochemically, 24 (26%) of the 92 studied samples were found to be positive, showing EBNA-1 granular nuclear staining in tumor epithelial cells. Conclusions These findings suggest an association between EBV infection and breast carcinoma development.

Immunohistochemistry Subtypes (ER/PR/HER) of Breast Cancer: Where Do We Stand in the West of Saudi Arabia?

In Saudi Arabia, cancer of breast is ranked the most frequent neoplasm and second source of cancer death in the female population. Breast cancer (BC) fast diagnosis, prognosis and medication management necessitate, these days, immunohistochemistry (IHC) assessment of hormone receptors and HER2 expression profile. The present report defines the IHC profile of ER, PR and HER2 in Saudi female breast neoplasms of ductal and lobular types and associations ER, PR and HER2 expression patterns with various clinicopathological factors (age, type of tumor, size, laterality, histological grade, and involvement of axillaries lymph nodes). Ninety nine cases of breast tumors were recruited from the pathology department archive of King Abdulaziz University Hospital, Kingdom of Saudi Arabia. ER, PR and HER2 expression was assessed using IHC staining. Ductal carcinomas with a variety of histological grades constituted 88 (88.8%) of total cases. Seventy four (77.8%), 59 (62.1%), and 35 (36.8%) of ductal carcinomas showed positive staining for ER, PR and HER2, in that order. Remaining breast cancer cases were four (4%) lobular carcinomas and two (2%) mixed form of ductal and lobular types, which were ER+, PR+, and HER2-. Breast cancer expression pattern of ER, PR and HER2 in Saudi female is different from that of Tunisian and Jordanian female populations and closer to the expression pattern of Egyptian, Lebanese, Iraqi and western country females. Furthermore, the present study found two IHC patterns of breast cancer ER+/PR-/HER2+ (5%) and ER+/PR-/HER2- (11.1%), which had not been reported in other Arabic studies. Thus the rates of IHC expression patterns in breast cancer show some variation among Arabic female populations.

The GSTP1 Ile105Val Polymorphism is not Associated with Susceptibility to Colorectal Cancer

The glutathione S transferase (GST) family is a major part of cellular defense mechanisms against endogenous and exogenous substances, many of which have carcinogenic potential. Alteration in the expression level or structure of the glutathione-S-transferase (GST) enzymes may lead to inadequate detoxification of potential carcinogens and consequently contribute to cancer development. A member of the glutathione-S-transferase (GST) family, GSTP1, is an attractive candidate for involvement in susceptibility to carcinogen-associated colorectal cancer. An A>G transition in exon 5 resulting in an Ile105Val amino acid substitution has been identified which alters catalytic efficiency. The present study investigated the possible impact of Ile105Val GSTP1 polymorphism on susceptibility to colorectal cancer. in Jordan We examined 90 tissue samples previously diagnosed with colorectal carcinoma, and 56 non-cancerous colon tissues. DNA was extracted from paraffin embedded tissues and the status of the GSTP1 polymorphism was determined using a polymerase chain reaction restriction fragment length polymorphism (RFLP) method. No statistically significant differences were found between colorectal cancer cases and controls for the GSTP1 Ile/Ile, Ile/Val and Val/Val genotypes. The glutathione S-transferase polymorphism was not associated with risk in colorectal cancer cases in Jordan stratified by age, sex, site, grade or tumor stage. In conclusion, the GSTP1 Ile105Val polymorphism is unlikely to affect the risk of colorectal cancer.

Infrequent Immunohistochemical Expression of Napsin A in Endometrial Carcinomas.

Introduction: Many studies described napsin A as a specific diagnostic marker that aids in differentiating lung adenocarcinomas from other respiratory tumors. This study describes the expression phenotype of napsin A in endometrial neoplasms, it investigates the relationship between this expression profile and the clinicopathologic parameters, and assess its utilization as an independent predictive marker. Methods: A total of 76 cases of previously diagnosed endometrial carcinoma (including 53 endometrioid adenocarcinomas, 6 endometrioid adenocarcinomas with squamous differentiation, 9 serous adenocarcinomas, 6 clear cell adenocarcinomas, and 2 malignant mixed mullerian tumors) and 30 tissue samples of noncancerous endometrium (including 16 proliferative endometriums, 10 secretory endometriums and 4 endometrial polyps) were retrieved from the archives of Pathology Department at King Abdulaziz University, Jeddah, Saudi Arabia. For napsin A detection, tissue microarrays and immunostaining were used. Results: A total number of 12 (15.78%) cases were positive for napsin A immunostaining. Brown granular cytoplasmic expression of napsin A was detected in 9.4% of endometrioid adenocarcinomas, 16.7% of endometrioid adenocarcinomas with squamous differentiation, 22.2% of papillary serous endometrial carcinomas, and 66.7% of clear cell carcinomas. Three (10%) control cases showed similar granular cytoplasmic expression. Positive napsin A immunostaining was more frequent in clear cell carcinoma, and there is a significant association between positive napsin A immunostaining and clear cell carcinoma (P-value=0.007). Significant associations have been found also between napsin A expression and older ages (above 60 y) and higher stage (IVB), the P-values of which were 0.035 and 0.043, respectively, but not with the tumor recurrence or survival rate. Conclusions: Although napsin A is infrequently expressed in endometrial carcinomas, positive results of napsin A immunostaining in endometrial neoplasms might support the diagnosis of clear cell carcinoma when the pathologic differential diagnosis includes other histologic subtypes.

Cyclin D1 is significantly associated with stage of tumor and predicts poor survival in endometrial carcinoma patients

Cyclin D1 overexpression has been described to have oncogenic role and association with diagnosis, prognosis and survival in various tumors. This study will describe the immunohistochemical phenotype of cyclin D1, and investigate the correlation between these patterns of expression and clinicopathological parameters of endometrial carcinomas, to conclude the clinical relevance of cyclin D1 expression in the evolution of endometrial neoplasms. This study employed 101 endometrial tissue samples which include 71 endometrial carcinomas and thirty normal and benign endometrium cases. All these tissue samples were used in the assembly of tissue microarrays which have been utilized afterward in immunohistochemistry staining to detect cyclin D1 expression. Forty (56.3%) cases of endometrial carcinomas showed brown nuclear expression of cyclin D1 including 36 (61%) cases of endometrioid carcinomas, and 3 (33.3%) cases of serous carcinomas. Twenty three (76.6%) cases of control group demonstrated nuclear expression. High score cyclin D1 immunohistochemical staining has been significantly linked with patient age (P=0.0001). Large proportion of high score cyclin D1 immunohistochemical staining was observed in females who are <40years of age while high proportions of negative staining were observed in older age groups. Histologic type of tissue was also significantly related to cyclin D1 immunohistochemical staining (P-value=0.0001), high staining is more common in normal proliferative and secretory endometrium while serous carcinoma is more prevalent with negative staining. Stage of tumor was significantly associated with cyclin D1 immunohistochemical staining (P-value=0.029), proportion of stage III and IV are higher in negative cyclin D1 immunostaining. Significantly higher proportion of high score cyclin D1 immunostaining is observed in controls while higher proportion of negative cyclin D1 immunostaining is observed among carcinoma cases (P-value=0.0001). No significant associations between cyclin D1 immunohistochemical staining and grade, recurrence and alive status were observed. Significant different survival distributions were observed (P-value=0.011) and poor survival behavior was correlated with negative cyclin D1 immunohistochemical staining. In conclusion, greater frequency of cyclin D1 expression was revealed in normal endometrial tissues in comparison with carcinomas. The distribution pattern of cyclin D1 immunoexpression suggests poor prognoses in endometrial carcinoma patients.

Clusterin immunoexpression is associated with early stage endometrial carcinomas

Clusterin has anti-apoptotic, regeneration and migration stimulating effects on tumor cells. This study investigates the relation between clusterin expression and the clinicopathological parameters in endometrial carcinomas. Seventy one cases of previously diagnosed endometrial carcinoma (including 59 endometrioid adenocarcinoma, 9 serous adenocarcinoma, 1 clear cell adenocarcinoma, and 2 malignant mixed Mullerian tumor) and 30 tissue samples of non-cancerous endometrium (including 16 proliferative endometrium, 10 secretory endometrium and 4 endometrial polyps) were employed for clusterin detection using tissue microarrays and immunostaining. A total number of 23 (32.4%) cases were positive for clusterin immunostaining. Brown granular cytoplasmic expression of clusterin was detected in 33.9% of endometrioid adenocarcinomas, 22.2% papillary serous endometrial carcinomas. Three (10%) control cases showed granular cytoplasmic expression. Positive clusterin immunostaining was found more frequent in well differentiated and stage I endometrial carcinomas, showing significant statistical association (p-value=0.036 and p-value=0.002 respectively). Significant difference in clusterin expression was observed between tumor cases and control group (P-Value=0.019), i.e., endometrial carcinoma cases are more than four times likely to show positive clusterin immunostaining (odds ratio 4.313 with 95% confidence interval 1.184-15.701). This study did not find relation between clusterin expression and disease recurrence, survival or any of the other clinicopathological parameters in endometrial tumors. The results of our study confirms the diagnostic values of clusterin in supporting the diagnosis of endometrioid carcinoma. When clusterin is expressed in endometrial tumors, it is associated with lower stage. The correlation of clusterin with tumor stage suggests involvement of this molecule in endometrial tumor progression.

Association between GSTP1 Genotypes and Hormone Receptor Phenotype in Invasive Ductal Carcinomas of Breast

Eighty six cases of invasive ductal breast carcinomas were utilized to investigate GSTP1 polymorphisms in certain immunohistochemistry (IHC) subtypes of breast cancer with respect to ER, PR and HER2 expression. The frequency of wild allele homozygote, heterozygote and variant allele homozygote genotypes were 46.5%, 52.3% and 1.16% respectively; Whereas 54.3% of the control subjects were GSTP1 wild type allele homozygous, 40.0% were heterozygous and 5.71% mutant allele homozygous. There was dramatic inverted relation between positive IHC ER staining and increasing grade of tumors in general (100%, 88.6%, 40.4%) and especially among tumors with heterozygote genotype of GSTP1 (70%, 35.4%, 22.7). There was increase in positive IHC HER2 staining consistent with higher grades in general (20%, 29.6%, 50.0%), especially among tumors with GSTP1 wild allele homozygote genotype (5.0%, 9.1%, 31.8%). A remarkable reverse relation was also observed between the fraction of IHC hormone receptor phenotype ER+/PR+/ HER2- and increased grade of tumors (60.0%, 45.5%, and 27.3%) especially among tumors with GSTP1 heterozygote genotype, and a similar link was noted regarding ER+/PR-/ HER2- and tumor grade. There was increase in frequency of ER-/PR-/ HER2- (0.0%, 6.8%, and 18.2%) and ER-/PR-/ HER2+ (0.0%, 4.54%, and 40.9%) consistent with the higher grades of tumors in general and especially GSTP1 heterozygote genotype tumors. As a conclusion, there is no correlation between GSTP1 polymorphism and increased risk of breast cancer i.e. the mutant allele is randomly distributed in cancer and control cases. However, there is a link between GSTP1 genotypes and hormone receptor expression status and certain phenotypes of breast cancer, which may have clinical importance.

Immunohistochemical expression of galectin-3 is significantly associated with grade, stage and differentiation of endometrial carcinomas

This study describes galectin-3 immunohistochemical phenotype and its association with clinicopathological factors in the carcinoma of endometrium. Seventy one cases of endometrial carcinoma and 30 cases of benign and normal endometrium were employed for the detection of galectin-3 protein using tissue microarrays and immunohistochemistry staining. Thirty nine (55%) cases, including 54.2% of endometrioid adenocarcinomas and 55.5% serous carcinomas, were positively stained for galectin-3. Brown granular expression of this glycoprotein was detected in transformed epithelial cells of 36 cases including 28 cases with membranous and cytoplasmic staining and 8 cases with only cytoplasmic staining; nuclear expression was present in stromal cells of the remaining 3 cases. Twenty-four (80%) control cases showed granular cytoplasmic and membranous expression, and six control cases were negative. Tumor grade, stage and differentiation were significantly associated with galectin-3 immunoreactivity (p-values are 0.043, 0.016, and 0.044 respectively), cases with membranous and cytoplasmic staining is significantly associated with grade I and stage II, while cases with loss of staining are more frequent in grade II, III and poorly differentiated tumors. No significant association of galectin-3 staining was observed with age, diagnosis, recurrence and alive status. The current study supports the tumor suppression role of galectin-3 in endometrial carcinoma. Greater galectin-3 immunostaining has been found in control endometrial tissues compared to endometrial tumors. Loss or decreased galectin-3 immunoexpression gives a sign for poor prognoses in endometrial carcinoma patients.

AMPK expression patterns are significantly associated with poor prognosis in breast cancer patients

Many investigators have examined the functions of AMP-activated protein kinase (AMPK) in cancer biology and its anti-neoplastic features in cancer models. The goal of this research is to assess the association of the immunohistochemical expression of AMPK in human mammary tumours with the clinical data of breast cancer patients. 449 cases of previously diagnosed breast cancer, and 27 tissue samples of fibroadenomas and normal breast were utilized for detection of AMPK expression using tissue microarrays and immunohistochemistry. Brownish nuclear and cytoplasmic staining were present in epithelial cells and stromal cells in 333 (74.16%) and 348 (77.5%) cancer cases respectively indicating AMPK expression. Twenty two (81.48%) control cases showed AMPK immunoexpression in both epithelial and stromal cells. Significant statistical association has been found between advanced stages of breast cancer and increased intensity of AMPK immunostaining only in epithelial cells (p-value=0.0001). Histotypes have been correlated with AMPK immunostaining in epithelial cells only (p-value=0.029). Low AMPK immunostaining scores were more dominant in DCIS, ductal and mixed types ductal and mucinous histotypes, while high intense staining was more common in the lobular type. Furthermore, breast tumour cases with lymph node metastases showed significant AMPK expression in both epithelial and stromal cells (p-value=0.0001 and p-value=0.026). Low scores of AMPK immunostaining were common in breast cancer cases with positive vascular invasion (p-value=0.007) and disease recurrence (p-value=0.008). No significant differences in survival behavior distributions were observed for the different categories of AMPK immunostaining in epithelial and stromal cells. In conclusion, our results showed decreased AMPK expression in breast cancer in comparison with the control group. AMPK expression was significantly correlated with some clinicopathological factors like advanced stage, lymph node involvement, vascular invasion and disease recurrence which give indications for poor clinical outcomes. Immunohistochemical staining of AMPK protein is a valuable method which could predict cases of breast cancer with poor prognosis.

Mesenchymal stem cell therapy of hepatocellular carcinoma in rats: Detection of cell homing and tumor mass by magnetic resonance imaging using iron oxide nanoparticles

BACKGROUND Bone marrow-derived mesenchymal stem cells (MSCs) are reported to improve hepatic fibrosis, and may impact the signaling mechanisms leading to the induction of hepatocellular carcinoma (HCC) in animal models of liver cirrhosis. OBJECTIVES The aim of this study was to clarify and explain the therapeutic role played by MSCs in hepatic cirrhosis and HCC by tracking them using nanoparticles. MATERIAL AND METHODS Liver cirrhosis and HCC were established in rats with the use of carbon tetrachloride and diethylnitrosamine injection. Magnetic resonance imaging (MRI) was used to track nanoparticlelabeled MSCs in the intact animal following injection and to monitor the changes in the hepatic parenchyma. RESULTS Labeling of MSCs with iron oxide nanoparticles did not adversely affect their viability and proliferation. MRI indicated a significant reduction in tumor mass in the labeled MSCs group compared to the control group. Histopathologic examination of the liver, following MSCs treatment, showed an apparently normal looking liver with no evidence of neoplastic cellular changes. The biochemical results support these findings. CONCLUSIONS This work documents that MSCs could be labeled with nanoparticles and traced in normal and cirrhotic liver and in liver with HCC in animals using MRI. MRI monitors the homing and localization of MSCs in the liver. MSCs infusion in animal models of cirrhosis and carcinoma may prove to be useful in limiting the cirrhotic process. Also, it may have a possible therapeutic potential on the carcinogenic process.