Ismail Sagap

Integrated analysis of copy number variation and genome-wide expression profiling in colorectal cancer tissues.

Integrative analyses of multiple genomic datasets for selected samples can provide better insight into the overall data and can enhance our knowledge of cancer. The objective of this study was to elucidate the association between copy number variation (CNV) and gene expression in colorectal cancer (CRC) samples and their corresponding non-cancerous tissues. Sixty-four paired CRC samples from the same patients were subjected to CNV profiling using the Illumina HumanOmni1-Quad assay, and validation was performed using multiplex ligation probe amplification method. Genome-wide expression profiling was performed on 15 paired samples from the same group of patients using the Affymetrix Human Gene 1.0 ST array. Significant genes obtained from both array results were then overlapped. To identify molecular pathways, the data were mapped to the KEGG database. Whole genome CNV analysis that compared primary tumor and non-cancerous epithelium revealed gains in 1638 genes and losses in 36 genes. Significant gains were mostly found in chromosome 20 at position 20q12 with a frequency of 45.31% in tumor samples. Examples of genes that were associated at this cytoband were PTPRT, EMILIN3 and CHD6. The highest number of losses was detected at chromosome 8, position 8p23.2 with 17.19% occurrence in all tumor samples. Among the genes found at this cytoband were CSMD1 and DLC1. Genome-wide expression profiling showed 709 genes to be up-regulated and 699 genes to be down-regulated in CRC compared to non-cancerous samples. Integration of these two datasets identified 56 overlapping genes, which were located in chromosomes 8, 20 and 22. MLPA confirmed that the CRC samples had the highest gains in chromosome 20 compared to the reference samples. Interpretation of the CNV data in the context of the transcriptome via integrative analyses may provide more in-depth knowledge of the genomic landscape of CRC.

Helicobacter pylori cagA gene variants in Malaysians of different ethnicity.

We have defined DNA repeat variability in the 3′-terminus of the cagA gene of Helicobacter pylori strains from Malaysian patients of different ethnicities. We identified different alleles based on the EPIYA repeats. cagA types A-B-D and A-B-B-D are more similar to the sequence of Japanese strains, whereas cagA types A-B-C, A-B-C-C, A-B and A-C displayed similarity to strain 26695 sequences. A significant association was found between cagA genotypes and patients’ ethnicity, with cagA type A-B-D being predominantly isolated from Chinese patients and cagA type A-B-C from Malays and Indians. Our data further corroborate the possibility that variant biological activity of CagA may affect the host specificity and/or pathogenicity of H. pylori.

Prospective Randomized Clinical Trial on Suction Elastic Band Ligator Versus Forceps Ligator in the Treatment of Haemorrhoids

OBJECTIVE This prospective randomized clinical trial was undertaken to compare the use of a single-operator vacuum suction ligator and the traditional forceps ligator in terms of pain perception following the procedure, intra-procedure bleeding and other complications. METHODS One hundred consecutive patients with second- and third-degree haemorrhoids presenting between July 2002 and September 2003 were randomized into suction and forceps groups for rubber band ligations. They were equally distributed in both groups, with a mean age of 48.7 years (range, 15-83 years). The immediate, 24-hour, 7-day and 14-day pain scores after the procedure were evaluated using a visual analogue scale. Intra-procedure bleeding and other complications at follow-up were evaluated. RESULTS Pain perception was worse in the forceps group immediately after ligation, with a mean score of 6.08 compared with 3.08 in the suction group (p < 0.001). Pain score remained high among the forceps patients at 24 hours post-banding, with a mean score of 4.00 compared with 1.92 in the suction group (p < 0.001). There was no significant difference in terms of immediate and 24-hour pain perceptions whether two or three haemorrhoids were banded per session (p = 0.904 and p = 0.058). The amount of analgesia consumed after banding correlated well with the severity of pain reported, being higher among the forceps group with a mean of 4.48 tablets (p = 0.003). Intra-procedure bleeding occurred in 25 patients in the forceps group compared with five in the suction group (p < 0.001). There were no severe complications such as perianal sepsis, urinary retention, sphincter dysfunction or bleeding during the trial. CONCLUSION Suction band ligation is superior to forceps ligation for the treatment of second- and third-degree haemorrhoids in terms of pain tolerance, amount of analgesia consumed and intra-procedure bleeding.

Identification of diagnostic markers in colorectal cancer via integrative epigenomics and genomics data

Apart from genetic mutations, epigenetic alteration is a common phenomenon that contributes to neoplastic transformation in colorectal cancer. Transcriptional silencing of tumor-suppressor genes without changes in the DNA sequence is explained by the existence of promoter hypermethylation. To test this hypothesis, we integrated the epigenome and transcriptome data from a similar set of colorectal tissue samples. Methylation profiling was performed using the Illumina InfiniumHumanMethylation27 BeadChip on 55 paired cancer and adjacent normal epithelial cells. Fifteen of the 55 paired tissues were used for gene expression profiling using the Affymetrix GeneChip Human Gene 1.0 ST array. Validation was carried out on 150 colorectal tissues using the methylation-specific multiplex ligation-dependent probe amplification (MS-MLPA) technique. PCA and supervised hierarchical clustering in the two microarray datasets showed good separation between cancer and normal samples. Significant genes from the two analyses were obtained based on a ≥2-fold change and a false discovery rate (FDR) P-value of <0.05. We identified 1,081 differentially hypermethylated CpG sites and 36 hypomethylated CpG sites. We also found 709 upregulated and 699 downregulated genes from the gene expression profiling. A comparison of the two datasets revealed 32 overlapping genes with 27 being hypermethylated with downregulated expression and 4 hypermethylated with upregulated expression. One gene was found to be hypomethylated and downregulated. The most enriched molecular pathway identified was cell adhesion molecules that involved 4 overlapped genes, JAM2, NCAM1, ITGA8 and CNTN1. In the present study, we successfully identified a group of genes that showed methylation and gene expression changes in well-defined colorectal cancer tissues with high purity. The integrated analysis gives additional insight regarding the regulation of colorectal cancer-associated genes and their underlying mechanisms that contribute to colorectal carcinogenesis.

Telepointer technology in telemedicine : a review

Telepointer is a powerful tool in the telemedicine system that enhances the effectiveness of long-distance communication. Telepointer has been tested in telemedicine, and has potential to a big influence in improving quality of health care, especially in the rural area. A telepointer system works by sending additional information in the form of gesture that can convey more accurate instruction or information. It leads to more effective communication, precise diagnosis, and better decision by means of discussion and consultation between the expert and the junior clinicians. However, there is no review paper yet on the state of the art of the telepointer in telemedicine. This paper is intended to give the readers an overview of recent advancement of telepointer technology as a support tool in telemedicine. There are four most popular modes of telepointer system, namely cursor, hand, laser and sketching pointer. The result shows that telepointer technology has a huge potential for wider acceptance in real life applications, there are needs for more improvement in the real time positioning accuracy. More results from actual test (real patient) need to be reported. We believe that by addressing these two issues, telepointer technology will be embraced widely by researchers and practitioners.

Cost analysis of colorectal cancer (CRC) management in UKM Medical Centre using clinical pathway

Background Cost analysis has become a huge concern in today’s healthcare due to the rising healthcare cost and the pressure for policy makers to make appropriate resource allocations. One of the approaches to cost calculation is by activity based costing using clinical pathway. Colorectal cancer (CRC) is fast becoming a threat to an industrialising country like Malaysia due to its rapidly rising incidence trend. Therefore, this study aims to develop a clinical pathway in managing CRC in Universiti Kebangsaan Malaysia Medical Centre (UKMMC) and to determine the cost of managing CRC according to disease stages.

Health-related quality of life among colorectal cancer patients in Malaysia: a study protocol

BackgroundColorectal cancer is a major public health problem in Malaysia. However, it is also one of the most treatable cancers, resulting in significant numbers of survivors. Therefore, the impact of surviving treatment for colorectal cancer on health related quality of life is important for the patients, clinicians and policy makers, and may differ in different cultures and populations. The aim of this study was to validate the Malaysian versions of the European Organization for Research and Treatment of Cancer quality of life instruments among colorectal cancers patients.Methods/designThis is a cross sectional multi centre study. Three hospitals were included, the University of Malaya Medical Centre, the Universiti Kebangsaan Malaysia Medical Centre and Hospital Tuanku Jaafar Seremban. Malaysian citizens and permanent residence were studied and demographic and clinical information obtained from hospital records. The European Organization for Research and Treatment of Cancer Quality of life Core 30, colorectal cancer CR29, and the colorectal cancer liver metastasis LMC 21 were used and an observer assessment of performance obtained with the Karnofsky Performance Scale. Questionnaires were translated into three most commonly spoken languages in Malaysia (Bahasa Malaysia, Chinese and Tamil), then administered, scored and analyzed following the developers’ guidelines. Ethical approval was obtained from the participating centres. Tests of reliability and validity were performed to examine the validity of these instruments.ConclusionThe result of pilot testing shows that the use of the Malaysian versions of EORTC QLQ C30, CR29 instruments is feasible in our sample of colorectal cancer patients. Instructions for completion as well as questions were well understood except the questions on the overall quality of life, overall health status and sexual activity. Thus we anticipate obtaining good psychometric properties for the instruments at the end of the study.

Identification of Predictive DNA Methylation Biomarkers for Chemotherapy Response in Colorectal Cancer

Resistance to 5-Fluorouracil (5-FU) is a major obstacle to the successful treatment of colorectal cancer (CRC) and posed an increased risk of recurrence. DNA methylation has been suggested as one of the underlying mechanisms for recurrent disease and its contribution to the development of drug resistance remains to be clarified. This study aimed to determine the methylation phenotype in CRC for identification of predictive markers for chemotherapy response. We performed DNA methylation profiling on 43 non-recurrent and five recurrent CRC patients using the Illumina Infinium HumanMethylation450 Beadchip assay. In addition, CRC cells with different genetic backgrounds, response to 5-FU and global methylation levels (HT29 and SW48) were treated with 5-FU and DNA methylation inhibitor 5-aza-2′-deoxycytidine (5-azadC). The singular and combined effects of these two drug classes on cell viability and global methylation profiles were investigated. Our genome-wide methylation study on the clinical specimens showed that recurrent CRCs exhibited higher methylation levels compared to non-recurrent CRCs. We identified 4787 significantly differentially methylated genes (P < 0.05); 3112 genes were hyper- while 1675 genes were hypomethylated in the recurrent group compared to the non-recurrent. Fifty eight and 47 of the significantly hypermethylated and hypomethylated genes have an absolute recurrent/non-recurrent methylation difference of ≥20%. Most of the hypermethylated genes were involved in the MAPK signaling pathway which is a key regulator for apoptosis while the hypomethylated genes were involved in the PI3K-AKT signaling pathway and proliferation process. We also demonstrate that 5-azadC treatment enhanced response to 5-FU which resulted in significant growth inhibition compared to 5-FU alone in hypermethylated cell lines SW48. In conclusion, we found the evidence of five potentially biologically important genes in recurrent CRCs that could possibly serve as a new potential therapeutic targets for patients with chemoresistance. We postulate that aberrant methylation of CCNEI, CCNDBP1, PON3, DDX43, and CHL1 in CRC might be associated with the recurrence of CRC and 5-azadC-mediated restoration of 5-FU sensitivity is mediated at least in part by MAPK signaling pathway.

Identification of Differentially Expressed Proteins in the Serum of Colorectal Cancer Patients Using 2D-DIGE Proteomics Analysis

Early detection of colorectal cancer (CRC) is vital for the improvement of disease prognosis. However to date there are no blood-based biomarkers sensitive and specific enough for early diagnosis. We analysed the differences in serum protein expression of early stage CRC (Dukes’ A and B) and late stage CRC (Dukes’ C and D) against normal controls using 2D Fluorescence Difference Gel Electrophoresis (2D-DIGE). Analysis of the 2D maps showed that 23 proteins were differentially expressed between groups (p ≤ 0.05) and these proteins were identified with LC-MS/MS. Eight proteins were up-regulated and 2 down-regulated in patients with early CRC, whereas 14 proteins were up-regulated and 4 down-regulated in those with late CRC compared to normal controls (p ≤ 0.05). Five proteins, namely apolipoprotein A1 (APOA1), apolipoprotein E (APOE), complement factor H (CFH), galectin-7 (GAL7) and synaptojanin-2 (SYNJ2) were validated using ELISA and only APOA1 and GAL-7 showed consistent findings. Further validation using immunohistochemistry showed negative immunoreactivity for GAL-7 in CRC tissues, suggesting that GAL-7 detected in the serum did not originate from the CRC tumour. APOA1 showed positive immunoreactivity but its expression did not correlate with Dukes’ staging (p = 0.314), tumour grading (p = 0.880) and lymph node involvement (p = 0.108). Differences in APOA1 isoforms and/or conformation between serum and tissue samples as well as tumour heterogeneity may explain for the discrepancies between DIGE and ELISA when compared to immunohistochemistry. Structural and functional studies of APOA1 in future would best describe the role of APOA1 in CRC.

Profiling of cytokines, chemokines and other soluble proteins as a potential biomarker in colorectal cancer and polyps

Graphical abstract Figure. No Caption available. HighlightsConsistent significant high levels of IL‐8, G‐CSF and Eotaxin concentration in CRC‐derived serum could be the biomarker for the CRC initiation and progression.MIP‐1&bgr; appears both in CRC‐ and P‐ derived serum.The higher sensitivity of the CBA analysis as compared to ELISA showed some promise of employing multiplexing in clinical application. Abstract Soluble proteins including cytokines, chemokines and growth factors are small proteins that mediate and regulate immunity. They involved in the pathogenesis of many diseases including cancers. The concentration of these proteins in biological fluids (serum or plasma) and tissues in diseases may suggest pathway activation that leads to inflammatory response or disease progression. Therefore, these soluble proteins may be useful as a tool for screening, diagnosis classification between stages of disease or surveillance for therapy. Enzyme‐linked immunosorbent assays (ELISA) and bioassay have been used as a gold standard in cytokine level measurements in clinical practice. However, these methods allow only single cytokine detection at a time and ineffective for screening purposes. Hence, the innovation of multiplexing technology allows measurement of many these soluble proteins simultaneously, thus allowing rapid, cost effective and better efficiency by using a minute amount of sample. In this study, we explored the profiles of key inflammatory cytokines, chemokines and other soluble proteins from the serum derived from colorectal carcinoma (CRC, n = 20), colorectal polyps (P, n = 20) and healthy volunteers (N, n = 20) using multiplexed bead‐based immunoassays. We aimed to evaluate if the levels of these soluble proteins can classify these groups of populations and explore the possible application of the soluble proteins as biomarkers in early stage screening and/or surveillance. We observed significant high IL‐4, MIP‐1&bgr;, FasL and TGF‐&bgr;1 levels but lower levels for RANTES in P‐derived serum as compared to N‐derived serum. Significant high IL‐8, VEGF, MIP‐1&bgr;, Eotaxin and G‐CSF observed in CRC‐derived serum when compared to N‐derived serum. Between CRC‐ and P‐derived serum, significantly higher levels of IL‐8, Eotaxin and G‐CSF but lower levels for TGF‐&bgr;1 were detected in CRC‐derived serum. These preliminary results were obtained from small sample size and could be further validated with larger sample size cohort to produce a panel of biomarkers for CRC and P patients. Our findings might be useful in developing a disease‐specific panel for biomarker screening assay. This could be used for early diagnosis and/or treatment surveillance.