AIMS: The aim of this study was to analyze the mutations in genes involved in CRC including MLH1, MSH2, KRAS, and APC genes.
METHODS: A total of 76 patients were recruited. We used the polymerase chain reaction-denaturing high-performance liquid chromatography for the detection of mutations in the mismatch repair (MMR) and APC genes and the PCR single-strand conformation polymorphism for screening of the KRAS gene mutations.
RESULTS: We identified 17 types of missense mutations in 38 out of 76 patients in our patients. Nine mutations were identified in the APC gene, five mutations were detected in the KRAS gene, and two mutations were identified in the MSH2 gene. Only one mutation was identified in MLH1. Out of these 17 mutations, eight mutations (47 %) were predicted to be pathogenic. Seven patients were identified with multiple mutations (3: MSH2 and KRAS, 1: KRAS and APC, 1: MLH1 and APC, 2: APC and APC).
CONCLUSIONS: We have established the PCR-DHPLC and PCR-SSCP for screening of mutations in CRC patients. This study has given a snapshot of the spectrum of mutations in the four genes that were analyzed. Mutation screening in patients and their family members will help in the early detection of CRC and hence will reduce mortality due to CRC.
METHODS: This is a cross-sectional study that conducted genome-wide copy number analysis using CytoScan 750K array on salivary samples from Malay subjects with NSCL/P with or without hypodontia aged 7-13 years. To confirm the significant results, simple logistic regression was employed to conduct statistical data analysis using SPSS software.
RESULTS: The results indicated the most common recurrent copy neutral LOH (cnLOH) observed at 1p33-1p32.3, 1q32.2-1q42.13 and 6p12.1-6p11.1 loci in 8 (13%), 4 (7%), and 3 (5%) of the NSCL/P subjects, respectively. The cnLOHs at 1p33-1p32.3 (D1S197), 1q32.2-1q42.13 (D1S160), and 6p12.1-6p11.1 (D1S1661) were identified observed in NSCL/P and noncleft children using microsatellite analysis markers as a validation analysis. The regions affected by the cnLOHs at 1p33-1p32.3, 1q32.2-1q42.13, and 6p12.1-6p11.1 loci contained selected genes, namely FAF1, WNT3A and BMP5, respectively. There was a significant association between the D1S197 (1p33-32.3) markers containing the FAF1 gene among NSCL/P subjects with or without hypodontia compared with the noncleft subjects (p-value = 0.023).
CONCLUSION: The results supported the finding that the genetic aberration on 1p33-32.3 significantly contributed to the development of NSCL/P with or without hypodontia. These results have an exciting prospect in the promising field of individualized preventive oral health care.
METHODS: Demographic, clinical and genotype data were determined for 1122 women (267 cases and 855 controls) recruited from the University of Malaya Medical Centre in the Klang Valley, Kuala Lumpur. Relevant articles were identified from Pubmed, Embase, MEDLINE, and Web of Science. Extraction of data was carried out and summary estimates of the association between rs780094 and GDM were examined.
RESULTS: The frequency of risk allele C was significantly higher in the cases than controls (OR 1.34, 95% CI 1.09-1.66, P = 0.006). The C allele was also associated with increased level of random 2-hour fasting plasma glucose and pregravid body mass index. Meta-analysis further confirmed the association of the GCKR rs780094 with GDM (OR 1.32, 95% CI 1.14-1.52, P = 0.0001).
CONCLUSION: This study strongly suggests that GCKR rs780094-C is associated with increased risk of GDM.
METHODS: PNMA5 mutants were generated through deletion or site-directed mutagenesis and transiently expressed in human cancer cell lines to investigate their role in apoptosis, subcellular localization, and potential interaction with MOAP-1 through apoptosis assays, fluorescence microscopy, and co-immunoprecipitation studies, respectively.
RESULTS: Over-expressed human PNMA5 exhibited nuclear localization pattern in both MCF-7 and HeLa cells. Deletion mapping and mutagenesis studies showed that C-terminus of PNMA5 is responsible for nuclear localization, while the amino acid residues (391KRRR) within the C-terminus of PNMA5 are required for nuclear targeting. Deletion mapping and co-immunoprecipitation studies showed that PNMA5 interacts with MOAP-1 and N-terminal domain of PNMA5 is required for interaction with MOAP-1. Furthermore, co-expression of PNMA5 and MOAP-1 in MCF-7 cells significantly enhanced chemo-sensitivity of MCF-7 to Etoposide treatment, indicating that PNMA5 and MOAP-1 interact synergistically to promote apoptotic signaling in MCF-7 cells.
CONCLUSIONS: Our results show that PNMA5 promotes apoptosis signaling in HeLa and MCF-7 cells and interacts synergistically with MOAP-1 through its N-terminal domain to promote apoptosis and chemo-sensitivity in human cancer cells. The C-terminal domain of PNMA5 is required for nuclear localization; however, both N-and C-terminal domains of PNMA5 appear to be required for pro-apoptotic function.
METHODS: The tricistronic expression construct that encodes MOAP-1, Bax, and RASSF1A (MBR) or its mutant, MOAP-1∆BH3L, Bax and RASSF1A (MBRX) was expressed from an IRES (Internal Ribosome Entry Site)-based tricistronic expression vector in human breast cancer cells, including MCF-7, MCF-7-CR (cisplatin resistant) and triple negative breast cancer cells, BMET05, for functional characterization through in vitro and in vivo models.
RESULTS: Transient expression of MBR potently promoted dose-dependent apoptotic signaling and chemo-sensitization in the cancer cells, as evidenced by loss of cell viability, nuclei condensation and Annexin-V positive staining while stable expression of MBR in MCF-7 cells significantly reduced the number of MBR stable clone by 86% and the stable clone exhibited robust chemo-drug sensitivity. In contrast, MBRX stable clone exhibited chemo-drug resistance while transiently over-expressed MOAP-1ΔBH3L inhibited the apoptotic activity of MBR. Moreover, the spheroids derived from the MBR stable clone displayed enhanced chemo-sensitivity and apoptotic activity. In mouse xenograft model, the tumors derived from MBR stable clone showed relatively high level of tumor growth retardation associated with the increase in apoptotic activity, leading to the decreases in both tumor weight and volume.
CONCLUSIONS: Expression of MBR in cancer cells induces apoptotic cell death with enhanced chemo-sensitization requiring the BH3L domain of MOAP-1. In animal model, the expression of MBR significantly reduces the growth of tumors, suggesting that MBR is a potent apoptotic sensitizer with potential therapeutic benefits for cancer treatment.
METHODS: We performed trio-based whole-exome sequencing (WES) in 210 families and case-only WES in 489 individuals with epileptic encephalopathies. The functional effect of CYFIP2 variants on WAVE signaling was evaluated by computational structural analysis and in vitro transfection experiments.
RESULTS: We identified three de novo CYFIP2 variants at the Arg87 residue in 4 unrelated individuals with early-onset epileptic encephalopathy. Structural analysis indicated that the Arg87 residue is buried at an interface between CYFIP2 and WAVE1, and the Arg87 variant may disrupt hydrogen bonding, leading to structural instability and aberrant activation of the WAVE regulatory complex. All mutant CYFIP2 showed comparatively weaker interactions to the VCA domain than wild-type CYFIP2. Immunofluorescence revealed that ectopic speckled accumulation of actin and CYFIP2 was significantly increased in cells transfected with mutant CYFIP2.
INTERPRETATION: Our findings suggest that de novo Arg87 variants in CYFIP2 have gain-of-function effects on the WAVE signaling pathway and are associated with severe neurological disorders. Ann Neurol 2018;83:794-806.
AIM: To investigate the genotype frequencies of MLH1 promoter polymorphism -93G>A and to determine whether it could play any role in modulating familial and sporadic CRC susceptibility risk.
METHODS: A case-control study comprising of 104 histopathologically confirmed CRC patients as cases (52 sporadic CRC and 52 Lynch syndrome patients) and 104 normal healthy individuals as controls was undertaken. DNA was extracted from peripheral blood and the polymorphism was genotyped employing PCR-RFLP methods. The genotypes were categorized into homozygous wild type, heterozygous and homozygous variants. The risk association between these polymorphisms and CRC susceptibility risk was calculated using binary logistic regression analysis and deriving odds ratios (ORs).
RESULTS: When risk association was investigated for all CRC patients as a single group, the heterozygous (G/A) genotype showed a significantly higher risk for CRC susceptibility with an OR of 2.273, (95%CI: 1.133-4.558 and p-value=0.021). When analyzed specifically for the 2 types of CRC, the heterozygous (G/A) genotype showed significantly higher risk for sporadic CRC susceptibility with and OR of 3.714, (95%CI: 1.416-9.740 and p-value=0.008). Despite high OR value was observed for Lynch syndrome (OR: 1.600, 95%CI: 0.715-3.581), the risk was not statistically significant (P=0.253).
CONCLUSION: Our results suggest an influence of MLH1 promoter polymorphism -93G>A in modulating susceptibility risk in Malaysian CRC patients, especially those with sporadic disease.