Alterations of PTEN and SMAD4 methylation in diagnosis of breast cancer: implications of methyl II PCR assay.

BACKGROUND: Diagnosis of breast cancer is more complicated due to lack of minimal invasive biomarker with sufficient precision. DNA methylation is a promising marker for cancer diagnosis. In this study, authors evaluated methylation patterns for PTEN and SMAD4 in blood samples using EpiTect Methyl II QPCR assay quantitative PCR technology. RESULTS: Methylation status for PTEN and SMAD4 were statistically significant as breast cancer patients reported hypermethylation compared to benign and control groups (77.1 ± 17.9 vs. 24.9 ± 4.5 and 15.1 ± 1.4 and 70.1 ± 14.4 vs. 28.2 ± 0.61 and 29.5 ± 3.6, respectively). ROC curve analysis revealed that both PTEN (AUC = 0.992) and SMAD4 (AUC = 0.853) had good discriminative power for differentiating BC from all non-cancer individuals (benign and healthy combined) compared to routine tumor markers CEA (AUC = 0.538) and CA15.3 (AUC = 0.686). High PTEN methylation degree was associated with late stages (84.2 ± 17.4), positive lymph node (84.2 ± 18.5), positive ER (81.3 ± 19.7), positive PgR (79.5 ± 19.1), and positive HER2 (80.7 ± 19.0) vs. 67.4 ± 13.8, 70.6 ± 14.8, 72.8 ± 14.9, 72.5 ± 14.7, and 70.2 ± 13.5 in early stages, negative lymph node, negative ER, negative PgR, and negative HER2, respectively. Similar results were obtained regarding SMAD4 methylation. Sensitivity, specificity, positive and negative predictive values, and accuracy for methylated PTEN were 100%, 95%, 99.1%, 100%, and 95%, respectively when differentiated BC from all-non cancer controls. Interestingly, PTEN could distinguish early BC stages with good sensitivity 84.4%, 51.4%, 69.1%, 72%, and 70%, respectively. CONCLUSION: Methylation status of PTEN and SMAD4 is a promising blood marker for early detection of breast cancer. Future studies are needed for their role as prognostic markers.

Serum MiRNA-27a as potential diagnostic nucleic marker for breast cancer.

BACKGROUND: Accumulating evidence reveals that microRNA 27a (miR 27a) is implicated in the pathogenesis of cancer. However, its diagnostic role in breast cancer (BC) still needs investigation. MATERIALS AND METHODS: MiR 27a expression was assessed in serum samples from patients with primary BC (n = 100), benign breast lesions (n = 30) and control group served as healthy volunteers (n = 20) using quantitative real-time PCR. RESULTS: Both expression and mean rank of miR 27a and tumor markers among BC patients as compared to the other two groups. Clinicopathological characteristics showed significant relation with miRN 27a expression for clinical stage, histological grading, ER receptor and HER-2/neu. The diagnostic efficacy for miR 27a was superior to both tumor markers for early detection of BC especially high-risk BC groups. CONCLUSION: Detection of miR 27a expression may serve as a potential sensitive minimally invasive molecular marker for early detection of primary BC.

Influence of copper (I) nicotinate complex and autophagy modulation on doxorubicin-induced cytotoxicity in HCC1806 breast cancer cells.

PURPOSE: Doxorubicin is regarded as the most therapeutic active agent available for triple-negative breast cancer (TNBC) treatment. However, the development of drug resistance and toxicity limits its effectiveness. Thus, developing novel strategies for TNBC treatment remains a significant challenge and doxorubicin-based combinations either by metal complexes (Copper I nicotinate complex) or with autophagy modulators could provide novel strategies and alternative strategies contributed to cancer cell death pathways, autophagy and apoptosis. MATERIALS AND METHODS: The viability of HCC1806 TNBC cells and IC50 values of Doxorubicin (DOX), Torin-1 (TOR), Chloroquine (CQ) and Copper (I) nicotinate complex (CNC) were assessed by MTT assay. ELISA was used for detecting microtubule-associated protein 1 light chain 3 (LC3) level. Real time PCR was used to determine (NBR1) gene expression. Cell cycle analysis and quantitative detection of acid vesicular organelles (AVOs) was performed by flow cytometry. TOR and CQ were used as autophagy modulators for induction and suppression of autophagy, respectively. RESULTS: The half-maximal inhibition effect of TOR combination with DOX was revealed to the induction of autophagic cell death and apoptotic cell death. On the other hand, combination of CQ with DOX increased the growth inhibitory effect, induced accumulation of AVOs and suppressed apoptotic cell death. However, combination of CNC with DOX inhibited autophagy and induced cell cycle arrest. CONCLUSION: Doxorubicin drug based combinations either with TOR, CQ or CNC could positively affect DOX effectiveness and reduce DOX doses applied on HCC1806 cells through modulation of autophagy.

Genetic polymorphisms of TP53 Arg72Pro and Pro47Ser among Egyptian patients with colorectal carcinoma.

BACKGROUND: Previous reports demonstrated the role of TP53 gene polymorphisms with CRC risk among several ethnic populations. The purpose of this study is to assess the association of the TP53 Arg72Pro and Pro47Ser variants with CRC risk among Egyptian patients. SUBJECTS AND METHODS: This work was conducted on 120 unrelated CRC Egyptian patients who were compared to 140 healthy controls. DNA was genotyped for these variants using the PCR-RFLP technique. RESULTS: CRC patients observed a significant association of the rare genotype of TP53 Arg72Pro polymorphism compared with healthy controls. On the contrast, all genetic models showed no statistical association of TP53 Pro47Ser polymorphism among CRC patients compared with healthy controls. On the contrast, CRC patients of the TP53 gene polymorphisms indicated no significant difference regarding their clinical and laboratory markers. CONCLUSIONS: Our findings indicate a strong association with TP53 Arg72Pro variant within increased risk of CRC among Egyptian patients.

Role of some circulating MiRNAs on breast cancer diagnosis.

Aberrant expression of miRNAs has a link with tumorgenesis and their deregulation is reported in biological fluids of cancer patients. Authors aimed to investigate the diagnostic role of miRNA-17-5p, miR-155 and miRNA-222 in serum samples from breast cancer patients (n = 80), benign breast patients (n = 40) and healthy individuals (n = 30) using quantitative real-time PCR technique. Median levels of investigated markers revealed significant increase in primary breast cancer followed by benign and control groups. Investigated miRNAs reported significant relation with clinical stages and histological grading, while only miRNA-17-5p showed significant relation with hormone receptors. When considering investigated miRNAs as compared to tumor marker, their sensitivities were superior over tumor markers for early diagnosis of breast cancer, detection of early stages and low grades breast cancer patients. In conclusion, detection of the miRNA-17-5p, miR-155 and miRNA-222 expression levels in serum samples is significant promising molecular markers for early breast cancer diagnosis.

miRNA-155 and miRNA-181a as prognostic biomarkers for pediatric acute lymphoblastic leukemia.

Mortality rates of acute lymphoblastic leukemia (ALL) have improved over the past 20 years; however, a significant portion of deaths stems from the lack of prognostic biomarkers, which can direct therapy and overcome drug resistance. microRNA-155a (miRNA-155a) and miRNA-181a are two single-stranded miRNAs involved in the pathogenesis of many types of leukemia and lymphoma and is linked to drug resistance. We investigated their expression levels in 55 patients, 45 diagnosed with ALL and 10 as a control group. We found that miRNA-155a and miRNA-181a were significantly upregulated in the ALL group with both being linked to high levels of minimal residual disease and poor prognosis. miRNA-155a cutoff value was significant in discriminating between high- and low-risk ALL patients as well as between ALL patients and healthy controls, miRNA-181a cutoff value, however, was not significant. Both markers levels were significantly downregulated after therapy. We conclude that miR-155 is correlated with poor prognosis in ALL, whereas we couldn't link miRNA-181a to the prognosis in ALL. Moreover, the marked decrease in their expression after therapy could reflect their impact on disease outcome.

Prognostic Implication of MIF Gene Expression in Childhood Acute Lymphoblastic Leukemia.

BACKGROUND: Cytogenetic and molecular genetic markers have been used recently to stratify risk and predict prognosis of different types of cancers. Macrophage migration inhibitory factor (MIF) is a pro-inflammatory cytokine that plays an important role in the pathogenesis of many diseases, such as autoimmune diseases and cancers. We studied MIF gene expression behavior to investigate its prognostic impact in childish patients with acute lymphoblastic leukemia (ALL). METHODS: MIF expression was analyzed using quantitative real-time (QRT) PCR. Patients are classified into two groups, high and low MIF expressers, according to median MIF gene expression. RESULTS: We did not find any significant difference between both groups as regards the clinical and hematological picture. However, high MIF expressers had significantly lower incidence of CR (25.8% vs. 72.4%, p = 0.001), higher incidence of refractory (51.6% vs. 24.1, p = 0.029), relapse rates (19.4% vs. 10.3%, p = 0.32), and higher mortality rate (54.8% vs. 20.7, p = 0.007) than lower MIF expressers. In addition, high MIF expressers show significantly shorter DFS (12.48 vs. 25.11 months, cumulative survival 38.7% vs. 72.2%, p < 0.01) and inferior overall survival (20.61 vs. 29.74 months, cumulative survival 45.2% vs. 79.3%, p < 0.001) than low MIF expressers. Multivariate cox-regression analysis adjustment confirmed that high MIF expression was the only independent prognostic factor in ALL patients for OS and DFS in our study (p = 0.004, p = 0.01 respectively). CONCLUSIONS: We found that MIF expression is an important prognostic factor in ALL patients with normal karyotype, and its incorporation into novel risk-adapted therapeutic strategies will improve the current cure rates for this group of patients.

Angiotensin II type 1 receptor gene polymorphism and serum angiotensin-converting enzyme level in Egyptian children with systemic lupus erythematosus.

Angiotensin II, the major effective molecule of the renin-angiotensin system, plays a vital role in the development of systemic lupus erythematosus (SLE). To study angiotensin II type 1 receptor (AT1R) gene polymorphism at (A1166C) in Egyptian children with SLE and its correlation with serum ACE level and SLE manifestations. AT1R gene polymorphism (A1166C) was done in 123 children with SLE in comparison to 100 healthy controls using polymerase chain reaction-based restriction fragment length polymorphism method (PCR-RFLP) and the tetra-primer amplification refractory mutation system-polymerase chain reaction (T-ARMS-PCR) to confirm the results of the genotyping. Serum ACE level measurement was done using ELISA technique. The frequencies of C-containing genotypes (AC + CC) and C-allele of AT1R (A1166C) were significantly higher in SLE patients compared to controls (p < 0.0001, OR = 4.9, 95% CI = 2.7-8.8; p Ë‚ 0.0001, OR = 3.6, 95% CI = 2.2-5.9, respectively). Lupus nephritis (LN) patients had significantly higher frequency of (AC + CC) genotypes and C-allele compared with controls (p Ë‚ 0.0001, OR = 5.1, 95% CI = 2.7-9.7; p Ë‚ 0.0001, OR = 3.5, 95% CI = 2.1-6.02, respectively). Mean serum ACE levels were significantly higher in SLE patients compared to controls (p Ë‚ 0.0001). There were no associations between AT1R gene polymorphism and serum ACE level and the clinical manifestations of SLE. The AT1R gene polymorphism can be considered a risk factor for the development of SLE in Egyptian children.

MIF-173G/C (rs755622) polymorphism as a risk factor for acute lymphoblastic leukemia development in children.

BACKGROUND: Macrophage inhibitory factor (MIF) is a pro-inflammatory cytokine modulating monocyte motility and a pleiotropic regulator of different biological and cellular processes. The MIF-173G/C (rs755622) polymorphism is found in the promoter region and affects its activity. The present study investigated the MIF polymorphism as a risk factor for the development of acute lymphoblastic leukemia (ALL) in Egyptian children. METHODS: We analyzed the MIF-173G/C (rs755622) polymorphism in 180 ALL cases and 150 healthy control children by amplification of the gene using a polymerase chain reaction followed by restriction endonuclease digestion and running on an agarose gel for visualization of the product. RESULTS: We found a significant incidence of the homozygous polymorphic (CC) genotype and the combined polymorphic genotypes (GC + CC) in ALL patients compared to healthy controls (p = 0.001 and p = 0.007, respectively), whereas the wild-type genotype (GG) was more common in healthy controls (p = 0.006). Multivariate logistic regression analysis adjustment for MIF different genotypes and other potential risk factors such as age, sex and parental smoking indicated that the CC genotype is the only significant risk factor for the test (p = 0.02). We also noted that, by increasing the C-allele representation within the gene [GC, CC], there was an increase in total leukocytic count (p = 0.09 and p = 0.001, respectively) that may reflect the bad prognostic impact of the polymorphic allele, although further studies are needed. CONCLUSIONS: The results of the present study indicate that the MIF-173G/C (rs755622) polymorphism is a risk factor for childhood ALL development with respect to both homozygous and combined polymorphic genotypes. In addition, the increased leukocytic count in synchronization with the increased representation of the polymorphic C-allele may reflect its bad prognostic impact.

Neutralizing activity and safety of human monoclonal antibodies against hepatitis C virus.

AIM: Assessment of the neutralizing activity of human monoclonal antibodies against HCV and also study their safety in experimental small animals (Swiss mice). MATERIALS AND METHODS: Assessment of neutralizing activity of human monoclonal antibodies against HCV envelope regions (E1, E2) by two methods: by HCV cc infectious system 1) and by using positive HCV positive serum as source of HCV particles genotype 4a (neutralizing assay 2). Dot ELISA was used to study the activity of the generated antibodies. Safety and toxicity of the generated human antibodies were tested by assessing the changes in the biochemistry of liver function and kidney function tests, Complete blood counts (CBC) and studying the pathological changes with different concentrations of purified human antibodies were carried out.. RESULTS: Human Abs # 5 & 11 showed neutralizing activity by (neutralizing assay 2) but were not neutralizing by HCV cc assay. Human Abs # 12 & 15 showed neutralizing activity by the two methods i.e our generated human antibodies Abs# 5 &11 & 12 & 15 were neutralizing for HCV genotype 4a and Abs # 12 & 15 were neutralizing for HCV genotypes 4a and 2a. Liver and kidney functions and CBC results indicated that doses of 10 µg, 100 µg were safe. The histopathological results indicated that the dose of 10 µg of purified human monoclonal antibodies per mouse body weight was safe. CONCLUSION: The generated human monoclonal antibodies can be used to develop potent immunotherapy agents that can be administrated for the post-transplantation patients to prevent the recurrence of HCV infection. Also, the monoclonal antibodies can be used to develop a candidate vaccine against HCV.

Antitumor Activity of Copper (I)-Nicotinate Complex and Autophagy Modulation in HCC1806 Breast Cancer Cells.

BACKGROUND: Squamous cell cancer is a heterogeneous aggressive disease, therefore, its treatment is challenging. Increased attention has been paid to metal complexes as anticancer drugs. However, new insights towards autophagy have been recognized due to its role in tumor cell death or survival. OBJECTIVE: To clarify the antitumor activity of copper (I) nicotinate complex (CNC) as new therapeutic agent and understand the role of autophagy modulation as a prospective target for the advancement of efficient therapeutic agent for treatment. METHOD: Viability of MDA-MB-231 and HCC1806 cells and IC50 values of CNC for both cell lines were assessed by MTT assay. Also, the viability and IC50 values of Torin1 and Chloroquine (CQ) were assessed only in HCC1806 cells by MTT assay. The level of microtubule-associated protein 1 light chain 3 (LC3) was assessed by ELISA. Real time PCR was used to detect the changes in NBR1 gene expression. Cell cycle distribution and quantitative detection of acid vesicular organelles (AVOs) were determined by flow cytometry. Fluorescence microscope was used for qualitative detection of AVOs. Modulation of autophagy was carried out by Torin1 as inducer and CQ as inhibitor. RESULTS: CNC restrained the growth, in a dose-dependent manner, and induced cell death in human HCC1806 cell line. In addition, the CNC treated cells displayed inhibition of autophagy, as indicated by reduction of AVOs, decrease in LC3 protein level and up regulation of NBR1 gene expression. CONCLUSION: CNC, as an autophagy inhibitor and pro-apoptotic agent, could be a promising anti-cancer agent either alone or in combination with other therapeutic drugs.

Association of monoallelic MUTYH mutation among Egyptian patients with colorectal cancer.

Colorectal cancer (CRC) is a heterogeneous triat that involves both environmental and genetic factors. Genetic mutations of MUTYH (p.Y179C and p.G396D) have been reported to be associated with increased risk of CRC among several ethnic populations. The aim of this work is to assess the association of the monoallelic MUTYH mutations (p.Y179C and p.G396D) with increased risk of CRC among Egyptian patients. This study included 120 unrelated CRC Egyptian patients who were compared with 100 healthy controls from the same locality. For all individuals, DNA was genotyped for MUTYH p.Y179C and MUTYH p.G396D mutations using the T-ARMS-PCR technique. The frequencies of monoallelic MUTYH mutations showed a strong association with the increased risk of CRC among Egyptian patients compared with controls (12.5 vs. 4.0 %, OR = 3.49, 95 % CI = 1.12-10.90, P = 0.03). Moreover, the frequency of MUTYH p.Y179C mutation was noted to be significantly higher among CRC patients compared to controls rather than MUTYH p.G396D mutation. Interestingly, CRC patients with tumors in the right side colon showed an evidence for association with the MUTYH p.Y179C mutation compared with tumors in the left side colon (p = 0.01). MUTYH p.Y179C mutation was associated with an increased risk of CRC among Egyptian patients rather than MUTYH p.G396D mutation.

Association of APC I1307K and E1317Q polymorphisms with colorectal cancer among Egyptian subjects.

Colorectal cancer is a multifactorial disease that involves both environmental and genetic factors. The gene encoding adenomatous polyposis coli (APC) has been reported to be associated with colorectal cancer (CRC) risk in several ethnic populations. The aim of this work is to assess the association of the APC I1307K and E1317Q polymorphisms with CRC risk among Egyptian subjects. This study included 120 unrelated CRC Egyptian patients who were compared to 100 healthy controls from the same locality. For all subjects, DNA was genotyped for APC I1307K and E1317Q polymorphisms using the PCR-ARMS technique. The frequency of APC I1307K carrier (TA+AA genotypes) was noted to be significantly higher among cases with CRC compared to controls (18.3 vs. 9.0 %, OR 2.58, 95 % CI 1.09-6.09, p = 0.03). Also the frequency of the APC I1307K A allele was significantly higher among cases compared to controls (10.4 vs. 4.5 %, OR 2.47; 95 % CI 1.12-5.42, p = 0.03). On the contrast, the frequencies of APC E1317Q GC genotype and C allele showed no significant difference among CRC patients compared to controls (3.3 vs. 2.0 %, OR 1.69; 95 % CI 0.30-9.42, p = 0.69 and 2.1 vs. 1.0 %, OR 2.11; 95 % CI 0.40-10.97, p = 0.46, respectively). Cases of the APC I1307K and E1317Q carriers (TA+AA and GC) showed no significant difference compared to those with I1307K and E1317Q non-carriers (TT and GG) regarding their clinical and laboratory markers. APC I1307K variant was associated with an increased risk of CRC among Egyptian subjects.

A simple diagnostic index comprising epithelial membrane antigen and fibronectin for hepatocellular carcinoma.

UNLABELLED: Background and rationale for the study. Continuing search for suitable tumor-markers is of clinical value in managing patients with various malignancies. These markers may be presented as intracellular substances in tissues or may be released into the circulation and appear in serum. Therefore, this work is concerned with identification and quantitative determination of epithelial membrane antigen (EMA) and fibronectin and estimating their performances as surrogate markers for identifying hepatocellular carcinoma (HCC). RESULTS: A total of 627 individuals constituted this study [fibrosis (F1-F3) = 217; cirrhosis = 191; HCC = 219]. Western-blot was used for identifying EMA and fibronectin in sera. As a result, a single immunoreactive band was shown at 130-kDa and 90-kDa corresponding to EMA and fibronectin, respectively. They were quantified using ELISA providing values in HCC higher than fibrosis or cirrhosis with a significant difference (P < 0.0001). For identifying HCC, EMA showed 0.82 area under receiver-operating characteristic curve (AUC) with sensitivity = 70% and specificity = 78% while fibronectin yielded AUC = 0.70 with sensitivity = 67% and specificity = 82%. FEBA-Test comprising fibronectin and EMA together with total-bilirubin and AFP was constructed yielding AUC = 0.92 for identifying HCC from cirrhosis with sensitivity = 89% and specificity = 85%. FEBA-Test was then tested for differentiating HCC from fibrosis showing AUC = 0.97 with sensitivity = 90% and specificity = 89%. FEBA-Test enabled the correct identification of HCC patients with CLIP 0-1 and size ≤ 3 cm with AUC = 0.80 and AUC = 0.84, respectively, indicating its ability in identifying early HCC. CONCLUSIONS: A four-marker index may improve the early detection of HCC with a high degree of accuracy.

Fibro-check: a combination of direct and indirect markers for liver fibrosis staging in chronic hepatitis C patients.

UNLABELLED: BACKGROUND AND RATIONALE FOR THE STUDY: The assessment of liver fibrosis provides useful information not only for diagnosis but also for therapeutic decision. This study was concerned with determining the levels of collagen III and its degrading enzyme matrix metalloproteinase-1 (MMP-1) as direct and complementary markers for liver fibrosis staging. RESULTS: A total of 269 chronic hepatitis C patients constituted this study. Western blotting was used for identifying collagen III and MMP-1 in serum samples. As a result, collagen III and MMP-1 were identified, respectively, at 70 and 245 kDa using their respective mono-specific antibodies. These two markers were quantified in sera of patients using ELISA. Next, Fibro-check was constructed combining collagen III and MMP-1 together with other indirect markers which reflect alteration in hepatic functions that proved useful to stage liver fibrosis. Fibro-check produced area under the receiver-operating characteristic curve (AUC) 0.91 and 0.83 for significant (F2-F4) and cirrhosis (F4), respectively. Additionally, we estimated the performance of Fibro-check in comparison with aspartate to platelet ratio index (APRI) and fibrosis index. Fibro-check seems to be more efficient than both of them. Fibro-check was then applied to the validation study to test its accuracy and reproducibility showing AUCs 0.90 for F2-F4 and 0.86 for F4. CONCLUSIONS: Fibro-check combining 'direct' and 'indirect' markers using a mathematical formula may improve the staging of liver fibrosis with a high degree of accuracy and seems more efficient than APRI and Fibrosis index in this group of Egyptian patients.

Antiviral activity of virocidal peptide derived from NS5A against two different HCV genotypes: an in vitro study.

This study aimed at assessment of the antiviral activity of an amphipathic α-helical peptide derived from the hepatitis C virus NS5A known as C5A virocidal peptide against different HCV genotypes. Two sources of HCV virus for in vitro study: HCV genotype 4 sera samples and JFH-1 infectious culture system genotype 2a were used. Several virocidal peptide concentrations were tested to determine the concentration that inhibits HCV propagation in Huh 7.5 cells according to three different prortocols (pre-infection, coinfection, and post infection). The capacity of the virocidal peptide to block HCV in Huh7.5 cells infected with different 10 individual serum samples was evaluated. In the pre-infection protocol, virocidal concentration (20, 50, and 75 µM) showed no viral RNA. In the co-infection protocol, virocidal concentrations (10, 20, 50, 75 µM) showed no viral RNA while in post-infection protocol, 75 µM was the only concentration that blocked the HCV activity. Results of Huh7.5 cell line transfected with HCV cc J6/JFH and treated with virocidal peptide revealed that only the higher virocidal concentration (75 µM) showed no amplification. The percentage of virocidal blocking in the 10 HCV individual serum samples was 60%. In conclusion, the C5A virocidal peptide has potent antiviral activity against HCV.

Apoptosis and cell proliferation: correlation with BCL-2 and P53 oncoprotein expression in human hepatocellular carcinoma.

BACKGROUND/AIM: Occurrence and biological characteristics of tumors are related not only to over-proliferation of carcinoma cells but also to decrease of apoptosis. The present study was suggested to evaluate the correlation between P53 and Bcl-2 oncoprotein expression with apoptosis and cell proliferative activity in HCC patients. METHODOLOGY: P53 and Bcl-2 protein expression were estimated in the sera and in liver tissues of 45 HCC cases using ELISA and immunohistochemistry. Apoptosis was estimated as apoptotic index (AI) and cell proliferative activity was detected using AgNORs. RESULTS: Serum p53 antigen in HCC patients (0.46±0.331ng/ml) showed significant elevation than healthy individuals (0.24±0.11ng/ml, p<0.05). P53 protein was immunostained in 41% of HCC; 37.5% of these positive cases were in diffuse pattern representing the mutant p53. Serum Bcl-2 was elevated in HCC cases (50.28±25.83u/ ml) than healthy individuals (26.65±8.63u/ml, p<0.05). Bcl-2 was immunohistochemically localized in 35.9% of HCC and the positivity was inversely proportional with the histological grade (47.4%, 25%, 25% in grade I,II,III respectively). Bcl-2 showed a positive linear correlation with p53 in the sera of carcinoma patients (p<0.05). CONCLUSION: Bcl-2 may play a role in hepatocarcinogenesis as an inhibitor of apoptosis. However, a positive linear correlation was found between bcl-2 and p53 suggesting that bcl-2/p53 co-expression pattern may be of value in the development of more effective medical therapies in HCC.

Impact of PIVKA-II in diagnosis of hepatocellular carcinoma.

Liver cancer grows silently with mild or no symptoms until advanced. In the absence of an effective treatment for advanced stage of hepatic cancer hope lies in early detection, and screening for high-risk population. Among Egyptians viral hepatitis is the most common risk factor for hepatocellular carcinoma (HCC). The current work was designed to determine the level of prothrombin induced by vitamin K absence-II (PIVKA-II) in sera of patients suffering from HCC and hepatitis C virus (HCV) patients being the most common predisposing factor for HCC. Our ultimate goal is diagnosis of HCC at its early stage. The current study was carried out on 83 individuals within three groups; Normal control, HCV and HCC groups. Patients were subdivided into cirrhotic and non-cirrhotic. Complete clinicopathological examination was carried out for each individual to confirm diagnosis. Individuals' sera were subjected to quantitative determination of alpha-fetoprotein (AFP), PIVKA-II and other parameters. PIVKA-II proved to be superior to AFP for early detection of HCC patients being highly sensitive and specific. Furthermore it has the ability to discriminate between different histopathological grades of HCC and It has a powerful diagnostic validity to evaluate the thrombosis of portal vein and to differentiate between early and late stages of HCC. The direct relation between the level of PIVKA-II and the size of tumor makes it an attractive tool for early HCC diagnosis and surveillance. Using the best cut-off value of AFP (>28), showed a sensitivity of (44%) and specificity of (73.3%). While cut-off value of PIVKA-II (>53.7) showed 100% sensitivity and specificity.

Evaluation of cytokeratin-1 in the diagnosis of hepatocellular carcinoma.

BACKGROUND: This study was undertaken to investigate whether serum cytokeratin-1 (CK1) could complement alpha-fetoprotein (AFP) to improve the diagnosis of hepatocellular carcinoma (HCC). METHODS: CK1 was identified using western blot and ELISA in serum samples from 250 Egyptian patients including 150 with HCC, 100 with liver cirrhosis (LC) and 50 healthy controls. Multivariate discriminant analysis (MDA) and ROC curve analyses were used to create a predictive model including CK1 in addition to a panel of routine blood markers. RESULTS: CK1 was identified at 67 kDa and quantified in sera of HCC patients using western blot and ELISA. MDA selected a score for the prediction of HCC from LC patients based on levels of CK1, albumin and AFP. An area under the ROC curves (AUC) of the score was 0.87. The score showed a sensitivity of 87% vs 39% sensitivity of AFP at cutoff value of 200 IU/ml for prediction HCC. Absolute specificity (100%) was obtained to discriminate HCC from healthy individuals. CONCLUSIONS: This study suggests that the use of a combination of score including CK1, AFP and albumin in clinical practice provides a non invasive and simple test that could increase significantly the sensitivity of HCC diagnosis.

Identification of a specific marker for hepatitis C virus infection using capillary zone electrophoresis.

BACKGROUND: Hepatitis C virus (HCV) infection is now becoming a common health problem in both developed and developing countries. The limitation of the available diagnostic approaches enhances the efforts to have a rapid, sensitive, and specific diagnostic testing for HCV infection. Capillary zone electrophoresis (CZE) is a fully automated analytical technique whose popularity is quickly increasing in the clinical chemistry laboratory. CZE can analyze nanoliters or less of samples with detection sensitivity at the attomole level (10(-18) mol) or less. METHODS: CZE was optimized for the identification of a specific marker of HCV infection. The performance characteristics of the CZE for the detection of HCV RNA peak were evaluated in comparison with standard nested PCR. RESULTS: A characteristic peak at 2.72 min was identified only in the CZE electropherogram of urine samples from HCV-infected individuals. The nature of the characteristic peak was investigated and confirmed to be HCV RNA using PCR and other biochemical treatments including RNase, DNase, and trypsin enzymes. CZE showed high degrees of sensitivity (94%) and specificity (96%) in comparison with the nested PCR. CONCLUSION: CZE provides a rapid, inexpensive, sensitive, and specific analytical method for diagnosis and mass screening of a large number of HCV-infected individuals.