The Clinical Significance of Septin 9 and Colon Cancer Specific Antigen-2 (CCSA-2) in Colorectal Cancer.

INTRODUCTION: Colorectal cancer (CRC) is a major health problem Worldwide, Egypt shows a high rate of early CRC in the world as 35% of 1,600 Egyptian CRC patients were under 40 with threefold increased risk of death within 5 years. DNA methylation-based biomarkers as methylated Septin9 (mSEPT9) has a promising role for detecting CRC. As well as set of nuclear matrix proteins associated with changes in the nuclear structure/architecture. detection of these nuclear proteins resulted in identification of biomarkers that are specific for colon cancer. Particular interest has been placed on colon cancer specific antigen-2(CCSA-2). METHODS: A total of 30 newly diagnosed CRC patients, 7 colonic adenoma patients, and 15 age- and sex-matched control subjects were recruited in this study. Plasma mSEPT9was assayed by Epi procolon kit, CCSA-2 by ELISA and, Occult blood in stool by Guaiac-based fecal occult blood test. The level of Colon Cancer mSEPT9 and CCSA-2 were carried on CRC patients both preoperatively and three months postoperatively. RESULTS: mSEPT9 has 96.7% sensitivity and 95.5% specificity in differentiating colorectal cancer patients from non-malignant cases. Also, our study showed a highly statistically significant difference between the pre and three months postoperative expression of mSEPT9 in colorectal cancer as there was a dramatically decrease in the expression of mSEPT9 postoperatively (p value < 0.001). The CCSA-2 at the cutoff level of >1.43 would provide 93.3% sensitivity and 90.9% specificity in differentiation between malignant and non-malignant cases. Also, the study showed that there is a statistically significant difference between colorectal cancer patients preoperatively and postoperatively according to CCSA-2 with dramatic decrease in its level postoperatively (p value > 0.001). CONCLUSION: The plasma SEPT9 DNA methylation level and Serum CCSA-2 could be used as promising non-invasive methods for observing the CRC patients postsurgical response to predict the occurrence of complete remission or relapses.

Evaluation of Osteopontin as a Biomarker in Hepatocellular Carcinomas in Egyptian Patients with Chronic HCV Cirrhosis

Background: Hepatocellular carcinoma (HCC) is a high incidence disease in Egypt with a poor prognosis and survival. Biomarkers are important for diagnosis of HCC at an early stage. Osteopontin (OPN), a glycoprotein secreted by macrophages, osteoblasts, and T cells, is also highly expressed in a variety of tumors, such as examples in the breast, colon, and stomach. The present study aimed to correlate the serum level of OPN in HCV-positive hepatocellular carcinoma patients, with OPN expression in tumor and non-tumor liver tissues in order to identify its efficacy as a biomarker for diagnosis. Material and Methods: Out of total of 146 patients, 80 were selected for inclusion in the study. Blood samples as well as specimens of tumor and non-tumor liver tissue were collected. In addition, blood samples from 20 healthy volunteers were obtained as controls. Serum OPN and alpha-fetoprotein (AFP) were evaluated by ELISA for HCC and control groups. OPN and AFP gene expression were examined by real-time PCR, after homogenization and DNA extraction from serum samples and liver tissues. Results: It was found that serum OPN levels were significantly higher in the HCC group compared to normal group (P=0.009), with a strong positive correlation with AFP expression. However, there was no significant difference between OPN expression in tumor and non-tumor liver tissue. Conclusion: Serum OPN is highly suggested to be a professional candidate for HCC early diagnosis, with a diagnostic ability and accuracy equal or higher than for AFP.

Differential Expression of Glypican-3 and Insulin­Like Growth Factor-II mRNAs and Alpha-Fetoprotein and Ki-67 Markers in HCV Related Hepatocellular Carcinomas In Egyptian Patients

Background: Increasing evidence indicates that in hepatocellular carcinomas (HCCs) abnormal gene expression, for example of glypican-3 (GPC-3) and insulin-like growth factor-II (IGF-II), are associated with the occurrence and progression of HCC. The objective of this study was to evaluate the differential expression of GPC-3 and IGF-II mRNAs in HCC tissues with a background of chronic hepatitis C virus (HCV) genotype 4 cirrhosis, in relation to Ki-67 and alpha-feto protein (AFP) tissue markers. Methods: One hundred and five patients with HCCs who had undergone hepatectomy, were included, after obtaining informed consent. Total RNA was extracted from malignant and corresponding peri-malignant liver tissues, and GPC-3 and IGF-II mRNAs in addition to beta-actin mRNA as an internal control, were evaluated in all samples by reverse transcriptase-polymerase chain reactions (RT-PCR). Routine histopathological diagnosis as well as immunohistochemical (IHC) staining using monoclonal antibodies for Ki-67 and AFP were also performed. Result: Expression of GPC-3 mRNA was positive in all HCC malignant tissue, with overexpression in 86/105 (81.9%); in respect to the grade of the tumor (1-3 grades), while in peri-malignant tissue it was over expressed only in 20/105 (19%). The IGF-II mRNA was over expressed in only 10/105 (9.5%) malignant and peri-malignant samples. AFP was expressed in 33.3% of malignant samples but absent in peri-malignant tissues. Ki-67 expression was significantly increased in malignant compared to peri-malignant tissue. Conclusion: GPC-3 and IGF II mRNAs may be good molecular markers for HCC, especially with a background of cirrhosis due to chronic HCV infection. Significant correlations were noted with the pattern of AFP and Ki-67 expression.

E1/E2 of Hepatitis C Virus Genotype4 and Apoptosis.

Several studies have addressed the possible role of hepatitis C virus genotype4 (HCV GT4) in apoptosis. However, this still not fully understood. In the current study a reconstructed clone of E1/E2 polyprotein region of the HCV GT4 was transfected into the Huh7 cell line and a human apoptotic PCR array of 84 genes was used to investigate its possible significance for apoptosis. Out of the 84 genes, only 35 showed significant differential expression, 12 genes being upregulated and 23 downregulated. The highestup regulated genes were APAF1 (apoptotic peptidaseactivating factor 1), BID (BH3 interacting domain death agonist) and BCL 10 (Bcell CLL/ lymphoma protein 10) with fold regulation of 33.2, 30.1 and 18.9, respectively. The most downregulated were FAS (TNF receptor super family), TNFRSF10B (tumor necrosis factor receptor superfamily member 10b) and FADD (FASassociated death domain) with fold regulation of 30.2, 27.7 and 14.9, respectively. These results suggest that the E1/E2 proteins may be involved in HCVinduced pathogenesis by modulating apoptosis through the induction of the intrinsic apoptosis pathway and disruption of the BCL2 gene family.

Diagnostic and Prognostic Relevance of Bone Marrow Microenvironment Components in Non Hodgkin's Lymphoma Cases Before and After Therapy

Objective: To evaluate stromal cells of the bone marrow microenvironment (BMM) in bone marrow trephine biopsy (BMTB) specimens, with a focus on fibronectin, tumor necrosis factor- alpha (TNF-α) and L-selectin in Non- Hodgkin's lymphoma (NHL) patients, before and after therapy. Materials and Methods: A total of 80 de novo NHL patients, 64 with B-cell lymphomas 80% , (follicular cell lymphoma (FCL) in 32, chronic lymphocytic leukemia/ small lymphocytic lymphoma (CLL/SLL) in 12, and diffuse large cell lymphoma in 20) and 16 with T-cell lymphomas (20%) all diagnosed as T-Lymphoblastic lymphomas, were evaluated before and after therapy. For comparison, 25 age and sex matched BM donors, were included as a control group. BMTB material and BM aspirates were taken for morphological assessment of stromal cells, the plasma of these samples being examined for TNFα and L-selectin by ELISA, and fibronectin by radial immunodiffusion (RID). Results: BM stromal cells comprising reticular macrophages and fibroblasts were elevated in 53.3% of NHL cases at diagnosis, while BM fibronectin levels were decreased and BM TNFα and L-selectin were higher than in controls (p<0.05). In NHL cases, elevated values of BM TNFα and BM L-selectin were associated with signs of aggressive disease, including >1 extra nodal sites, detectable B symptoms, high grade, BM and CNS invasion, and a high International prognostic index (IPI) (p<0.05). Conclusion: BMM components, TNFα, L-selectin and fibronectin, in NHL can be useful in evaluating disease activity, extent and response to treatment and as prognostic markers according to the IPI.