The HLA DPB1*02:01:02 and DQB1*05:02:01 alleles as possible risk factors for colorectal carcinoma in the Malaysian population

Many studies have shown that the immune response highly depends on the inheritance of specific HLA genes in promoting the generation of T cells for the elimination of pathogens. Loss or alteration of HLA antigen expression in tumor cells has been observed in a variety of human malignancies leading to immune escape or immune resistance. We investigated whether the inheritance of certain alleles of HLA class II genes confers susceptibility or resistance towards the development of colorectal carcinoma (CRC). Molecular typing of HLA DRB1, DQB1 and DPB1 alleles in 42 patients diagnosed with CRC and 50 ethnically matched healthy controls using the PCR-sequence based typing (PCR-SBT) was conducted. The HLA DPB1*02:01:02 was significantly higher in CRC patients (38.1%, p=0.0189) compared to healthy controls (16%). Also, HLA DQB1*05:02:01 was present in 28.6% of CRC patients but only 10% of healthy controls (p=0.0278). The odds ratios for HLA DPB1*02:01:02 and HLA DQB1*05:02:01were 3.23 and 3.60, respectively. There were no significant association observed for the DRB1 allele with CRC. Our study suggests that the HLA DPB1*02:01:02 and HLA DQB1*05:02:01 alleles may confer a higher risk for CRC

Preliminary study shows novel variant detected in the screening of RET gene in Malaysian patients with Hirschsprung’s disease

Hirschsprung’s disease (HSCR) is a disorder associated with congenital absence of ganglion cells in the gastrointestinal tract. Molecular analyses have identified variants in various genes including RET, GDNF, EDN3 and EDNRB that are involved in the development, migration and survival of neural cells. Variants in the receptor tyrosine kinase (RET) are most common and have been identified in 10-20% of sporadic HSCR patients. The objective of this study was to screen for RET gene variants in Malaysian patients with HSCR. Thirty-two patients with HSCR and 30 normal controls were recruited for this study. Mutations were screened using the Polymerase Chain Reaction – Denaturing High Performance Liquid Chromatography (PCR-dHPLC) approach. Mutations identified were then confirmed using Sanger sequencing. We identified one novel rare variant in exon 4 (A268A c807 G>C) in one patient. We also identified the common coding sequence variantsA45A (c135G>A), A432A (c1296A>G), L769L (c2307 T>G) and the G691S in our cohort of patients. In conclusion, our Malaysian patients with HSCR diseases showed the presence of similar RET gene common variants which have been described in other populations. We have also identified a novel variant in exon 4 (A268A)

Identification of missense mutations in genes related to cancer pathways in glioma

Glioma is the most common primary brain tumour of the central nervous system. Many genetic alterations and mutations have been identified in glioma using various approaches. We performed DNA sequencing on the tumours of 16 patients with Grade I, II, III and IV glioma. The AmpliSeq Cancer Primers Pool was used to generate the amplicons. The targeted-ion sphere particles were prepared using the Ion One Touch and Ion Enrichment systems. DNA sequencing was performed on the Ion Torrent Personal Genome Machine (PGM) and the data were analysed using the Torrent Suite Software. In total, 14 mutations were identified in the following genes: KDR (Q472H), MLH1 (V384D), MET (N375S), PTPN11 (E69K), BRAF (V600E), TP53 (D149E, E154K, V157F), IDH1 (R132H), PIK3CA (H1047R), CSF1R (c1061_1061 ins A), KIT (M541L), PTEN (c1373_1373 del A) and PDGFRA (E556V). In addition, there were four novel mutations identified; TP53 (E154K, and D149E), CSF1R (c1061_1061 ins A) and PDGFRA (E556V). The pathogenicity prediction showed that only three mutations were pathogenic: PTPN11 (E69K), BRAF (V600E) and Tp53 (E154K). These mutations result in changes of the proteins’ structure and could affect their functions. Pathway analyses suggested that these genes are closely related to the pathogenesis of GBM through several pathways such as proliferation and invasion, metabolism and angiogenesis. In conclusion, PGM in combination with the AmpliSeq Cancer Panel could be utilised as a potential molecular diagnostic tool not only for glioma but also for other cancers

Integrated microRNA, gene expression and transcription factors signature in papillary thyroid cancer with lymph node metastasis

Background. Papillary thyroid carcinoma (PTC) is the commonest thyroid malignancy originating from the follicle cells in the thyroid. Despite a good overall prognosis, certain high-risk cases as in those with lymph node metastasis (LNM) have progressive disease and poorer prognosis. MicroRNAs are a class of non-protein-coding, 19–24 nucleotides single-stranded RNAs which regulate gene expression and these molecules have been shown to play a role in LNM. The integrated analysis of miRNAs and gene expression profiles together with transcription factors (TFs) has been shown to improve the identification of functional miRNA-target gene-TF relationships, providing a more complete view of molecular events underlying metastasis process. Objectives. We reanalyzed The Cancer Genome Atlas (TCGA) datasets on PTC to identify differentially expressed miRNAs/genes in PTC patients with LNM-positive (LNM-P) versus lymph node negative (LNN) PTC patients and to investigate the miRNA-gene-TF regulatory circuit that regulate LNM in PTC. Results. PTC patients with LNM (PTC LNM-P) have a significantly shorter disease-free survival rate compared to PTC patients without LNM (PTC LNN) (Log-rank Mantel Cox test, p = 0.0049). We identified 181 significantly differentially expressed miRNAs in PTC LNM-P versus PTC LNN; 110 were upregulated and 71 were downregulated. The five topmost deregulated miRNAs were hsa-miR-146b, hsa-miR-375, hsa-miR-31, hsa-miR-7-2 and hsa-miR-204. In addition, 395 miRNAs were differentially expressed between PTC LNM-P and normal thyroid while 400 miRNAs were differentially expressed between PTC LNN and normal thyroid. We found four significant enrichment pathways potentially involved in metastasis to the lymph nodes, namely oxidative phosphorylation (OxPhos), cell adhesion molecules (CAMs), leukocyte transendothelial migration and cytokine–cytokine receptor interaction. OxPhos was the most significantly perturbed pathway (p = 4.70E−06) involving downregulation of 90 OxPhos-related genes. Significant interaction of hsa-miR-301b with HLF, HIF and REL/NFkB transcription factors were identified exclusively in PTC LNM-P versus PTC LNN. Conclusion. We found evidence of five miRNAs differentially expressed in PTC LNM-P. Alteration in OxPhos pathway could be the central event in metastasis to the lymph node in PTC. We postulate that hsa-miR-301b might be involved in regulating LNM in PTC via interactions with HLF, HIF and REL/NFkB. To the best of our knowledge, the roles of these TFs have been studied in PTC but the precise role of this miRNA with these TFs in LNM in PTC has not been investigated