Anatomical Pathology
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Browsing Anatomical Pathology by browse.metadata.advisor "Kotze, Maritha J."
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- ItemAnalysis of hereditary haemochromatosis and clinical correlations in the elderly(Stellenbosch : Stellenbosch University, 2000-12) Bouwens, C. S. H.; Kotze, Maritha J.; Maritz, F. J.; De Villiers, J. N. P.; Stellenbosch University. Faculty of Medicine & Health Sciences. Dept. of Pathology.ENGLISH ABSTRACT: Hereditary haemochromatosis (HH) is an autosomal recessive iron storage disease where the accumulation of iron in parenchymal organs may lead to diabetes, heart failure, liver cirrhosis, arthropathy, weakness and a variety of other ailments if preventive measures are not taken. HH is often not considered as a cause of these conditions, particularly not in the elderly where the background frequencies of type II diabetes, osteoarthritis and heart failure are generally high. Heterozygosity for C282Y, the HFE-mutation causing HH in approximately 80% of affected individuals worldwide, has been linked to a raised incidence of malignancies of the colon and rectum, stomach and the haematological system. One of the highest carrier-frequencies (116) in the world for this mutation has been reported in the South-African Afrikaner population, resulting in C282Y-homozygosity in approximately 1 in every 115 people in this group. A sample of 197 elderly Afrikaner volunteers was recruited for genotype/phenotype association studies. Their clinical presentation was denoted, biochemical iron-status determined and HFE genotyping performed. Either an increase or decrease in survival, or both, were proposed, depending on possible gender effects. HH has been positively associated with various cancer types, but may also protect against iron-deficiency anaemia which is by far the most frequent cause of anaemia in the older person. This study has led to the following findings: 1. The carrier frequency of mutation C282Y was found to be 1/8 in the elderly population (similar in males and females), which is slightly lower than the 1/6 reported in younger adults from the same population. Only one C282Y homozygote and two C282YIH63D compound heterozygotes were detected, all of them female. 2. The prevalence of diabetes, heart disease, arthropathy or a combination of these conditions did not differ significantly in C282Y heterozygotes and the mutationnegative group. 3. Among 24 C282Y heterozygotes only one individual with rectal carcmoma was detected compared with two cases with rectal- and seven with colonic malignancies in 153 mutation-negative individuals. The single female C282Y homozygote identified suffered from both rectal and colon carcinoma and died approximately 6 months ago as a consequence of her colon malignancy. 4. Serum ferritin appears to be a highly unreliable parameter of iron status, particularly in the elderly where a variety of factors that may influence the levels are often present in elderly individuals. This may be due to ageing alone or as a result of multiple comorbidities. 5. Serum ferritin levels were lower than expected in elderly subjects with mutation C282Y and compound heterozygotes with both C282Y and H63D, which may be related to a variable penetrance of the HFE gene mutations. It is possible that variation in other genes exist that confer protection against iron-loading by gene-gene interaction. The probability that environmental factors (e.g. a low iron diet) are more important in this respect cannot be excluded, although this is considered less likely in the light of the fact that the same trend was observed in all mutation-positive elderly individuals. It is therefore highly likely that C282Y -positive subjects with significant iron loading have died before reaching their seventies, particularly since none of the males included in this study were homozygous or compound heterozygous for the mutations analysed. In conclusion, possession of a mutant HFE gene does not appear to confer a survival advantage in old age, neither does it seem that mutation carriers with significant ironloading are overlooked by the medical fraternity. Further investigations are warranted to shed more light on the contributions of gene-gene and gene-environment interaction in the clinical manifestation of Hll, and how these processes can be manipulated to prevent the symptoms of this largely underdiagnosed disease.
- ItemAnalysis of the clinical utility of gene expression profiling in relation to conventional prognostic markers in South African patients with breast carcinoma(Stellenbosch : Stellenbosch University, 2014-12) Grant, Kathleen Ann; Kotze, Maritha J.; Wright, Colleen A.; Apffelstaedt, Justus P.; Stellenbosch University. Faculty of Medicine and Health Sciences. Dept. of Pathology. Anatomical Pathology.ENGLISH ABSTRACT: Breast cancer is a heterogeneous disease characterised by marked inter-individual variability in presentation, prognosis and clinical outcome. The recognition that morphological assessment has limited utility in stratifying patients into prognostic subgroups led to clinico-pathological classification of tumour biology, based on receptor expression using immunohistochemical (IHC) techniques. This standard is currently complemented by the development of gene expression profiling methodology that led to the identification of intrinsic molecular subtypes, reflecting tumour genetics as the true driver of biological activity in breast cancer. The study was based on the hypothesis that molecular classification of breast carcinomas integrated with established clinico-pathological risk factors will improve current diagnostic and risk management algorithms used in clinical decision-making. A pathology-supported genetic testing strategy was used to evaluate microarray-based gene profiling against diagnostic pathology techniques as the current standard. Clinico-pathological factors including age, number of positive axillary nodes, tumour size, grade, proliferation index and hormone receptor status was documented for 141 breast cancer patients (143 tumours) referred for microarray-based gene expression profiling between 2007 and 2014. Subsets of patients were selected from the database based on the inclusion criteria defined for three phases in which the study was performed, in order to determine 1) the percentage of patients stratified as having a low as opposed to high risk of distant recurrence using the 70-gene MammaPrint profile within the inclusion criteria, 2) correlation of HER2 status as determined by IHC and fluorescence in situ hybridisation (FISH) with microarray-based mRNA readout (TargetPrint), and 3) the relationship between hormone receptor determination as reported by standard IHC and molecular subtyping using the 80-gene BluePrint profile. Similar distribution patterns for MammaPrint low- and high-risk profiles were obtained irrespective of whether fresh tumour biopsies or formalin-fixed paraffin embedded (FFPE) tissue was used. During the first phase of the study, 60% of the 106 tumour specimens analysed with MammaPrint were classified as low-risk and 40% as high-risk using a newly-developed MammaPrint pre-screen algorithm (MPA) aimed at cost-saving. In the second phase of the study, performed in 102 breast tumours, discordant or equivocal HER2 results were found in four cases. Reflex testing confirmed the TargetPrint results in discordant cases, achieving 100% concordance regardless of whether fresh tumour or FFPE tissue was used for microarray analysis. For the third phase of the study 74 HER2-negative tumour samples were selected for comparative analysis. Statistically significant positive correlations were found between protein expression (IHC score) and mRNA (TargetPrint) levels for estrogen receptor (ER) (R=0.53, p<0.0001) as well as progesterone receptor (PR) (R=0.62, p<0.0001), while combined ER/PR tumour status was reported concordantly in 82.4% of these tumours. BluePrint was essential for interpretation of these results used in treatment decision-making. The MPA developed in South Africa in 2009 was validated in this study as an appropriate strategy to prevent chemotherapy overtreatment in patients with early-stage breast cancer. The use of microarray-based analysis proved to be a reliable ancillary method of assessing HER2 status in breast cancer patients. Risk reclassification based on the TargetPrint results helped to avoid unnecessary high treatment costs in false-positive cases, in addition to providing potentially life-saving treatment to those for whom it was indicated. While neither IHC nor TargetPrint estimation of intrinsic subtype correlated independently with the molecular subtype as indicated by BluePrint profiling, the ability to distinguish between basal-like and luminal tumours was enhanced when the combined protein and mRNA values was considered. Genomic profiling provided information over and above that obtained from routine clinico-pathological assessments. This finding supports the relevance of a pathology-supported genetic testing approach to breast cancer management, whereby advanced genomic testing is combined with existing clinico-pathological risk stratification methods for improved patient management.
- ItemComparison of Xpert® Breast cancer STRAT4 assay and immunohistochemistry for the evaluation of breast cancer biomarkers in South African patients(Stellenbosch : Stellenbosch University, 2020-12) Dube, Welile Vumile; De Jager, Louis; Kotze, Maritha J.; Stellenbosch University. Faculty of Medicine and Health Sciences. Dept. of Pathology: Anatomical Pathology.Background: Breast cancer is one of the most common cancers diagnosed in women and approximately 60% of breast cancer related deaths are reported in low-and middle-income countries. Breast cancer is a highly heterogeneousdisease, and molecular subtyping is paramount foreffective treatment of patients. Therefore, it is important to validate new molecular methods for assessing cancer biomarkers for cost-effective use in resource-poor settings.Aim:Aretrospective study was performed to determine the concordance between aQuantitativeReverse TranscriptionPolymerase Chain Reaction(RT-qPCR) CE-IVD assay (Xpert® Breast Cancer STRAT4*) and the current gold standard methods of immunohistochemistry (IHC) and fluorescence in situ hybridization (FISH) for determining estrogenreceptor (ER),progesterone receptor(PR), human epidermal growth factor receptor 2 (HER2)andproliferation index(KI-67)expression in breast carcinomas.Method: One hundred and one cases of breast carcinoma were retrieved from the archives of the Division of Anatomical Pathology, Tygerberg Academic Hospital. The original stained slides were reviewed and IHC expression of ER, PR, HER2 and KI-67 scored. Three-micron sections were cut from formalin-fixed paraffin embedded (FFPE) tissue blocks and processed according to the instructions of the manufacturer. The assay was run on the resultant lysates.Results:The overall percentageagreement between the Xpert® STRAT4 assay and IHC / FISH results were 85.15% for ESR, 89.90% for PGR,91.09% for ERBB2, 90.72% for MKI67 (when using a cut off of 10%) and 84.54% for MKI67 (when using a cut-off of 20%). The positive percentage agreement for ESR, PGR, ERBB2, MKI67 with 10% cut-off andMKI67 with 20% cut-offwere 82.76%, 94.64%, 68.97%, 91.30% and 96.05%, respectively, and the negative percentage agreement were 100%, 84.09%, 91.67%, 80.00% and 42.86%, respectively. Conclusion:The studyhas shownthattheXpert® BreastCancer STRAT4 assay shows good concordance with IHC and FISH in detecting breast cancer biomarkers, and may become a supplementary or alternativestandard of care aftervalidation studies are performed.
- ItemDevelopment and application of a pathology supported pharmacogenetic test for improved clinical management of South African patients with breast cancer and associated co-morbidities(Stellenbosch : Stellenbosch University, 2016-03) Van der Merwe, Nicole; Kotze, Maritha J.; Pienaar, Fredrieka; Janse van Rensburg, Susan; Bezuidenhout, Juanita; Stellenbosch University. Faculty of Medicine and Health Sciences. Dept. of Pathology. Anatomical Pathology.ENGLISH ABSTRACT: Three major challenges in the field of breast cancer have been identified as research priorities for this study. The first is the need to combine genetic testing of high-risk patients with familial breast cancer with pharmacogenetics to reduce recurrence risk in cancer survivors due to drug failure as a consequence of anti-cancer treatment that does not match the patient’s genotype. The second is the delineation of key pathways through which genes implicated in breast cancer and associated co-morbidities can serve as nutritional and drug targets across diagnostic boundaries. The third is the discovery of genetic alterations underlying familial breast cancer not attributed to mutations in the two major tumour suppressor genes, BRCA1 and BRCA2. The study population consisted of 164 breast cancer patients (60 Coloured/Mixed Ancestry and 104 Caucasian), of whom 88 patients were selected from a total of 813 individuals who provided informed consent for inclusion of their data in a genomics database resource generated at the interface between the laboratory and routine clinical practice. In addition, DNA samples of 101 cancer-free individuals above the age of 65 years were available for clinical validation of potentially causative variants in an extended female control group. In the first phase of this study, real-time polymerase chain reaction (PCR) TaqMan© technology was used to confirm the potential value of adding pharmacogenetic testing (CYP2D6 allele 4) to standard immunohistochemistry (IHC)-based breast tumour subtyping complemented by BRCA mutation screening and/or microarray gene profiling in eligible patients. In phase two of the study, common genetic risk factors for cardiovascular disease (CVD) were shown to be significantly associated with earlier age (10 years on average) of breast cancer onset/diagnosis (APOE E4 allele p=0.003; 95% CI: 4-15) and body mass index (BMI) (MTHFR 1298 A>C; p=0.01; 95% CI: 3-14) in patients stratified according to estrogen receptor (ER) status, after adjustment for potential confounders. Age at diagnosis/onset of breast cancer was significantly lower in patients with ER-negative versus ER-positive tumours, after adjustment for ethnicity (p=0.022), while BMI was significantly higher in patiens with ER-positive compared to ERnegative tumours after adjustment for age, ethnicity, and family history of cancer (p=0.035). These findings contributed to the development of an exome pre-screening algorithm (EPA) used in part 3 of this study to select three genetically uncharacterized breast cancer patients for whole exome sequencing (WES) performed in comparison with three ethnically concordant cancer-free controls. WES followed by variant calling using both the standard human genome reference sequence (hg19) and an ethnically concordant major allele reference genome (MARS) revealed a more than 20% discrepancy in the number of gene variants identified in the same samples. After exclusion of a large number of false-positives caused by minor alleles in hg19, two rare missense mutations (<1%) were identified in a family with ER-positive breast cancer: RAD50 R385C and MUC1 Q67E. Three different bioinformatics tools were used to predict functionality and both mutations were confirmed by Sanger sequencing and/or real-time PCR in the Pathology Research Facility (PRF) laboratory. Neither the RAD50 nor the MUC1 missense mutation were identified in the exomes of an unrelated breast cancer patient with triple-negative breast cancer or three population-matched control individuals. This study led to the development of a pathology-supported genetic testing framework for WES beyond the limitations of single-gene BRCA mutation screening in South African breast cancer patients. Our findings support previous WES results indicating that the majority of genetically uncharacterised familial breast cancer may be caused by a combination of low-moderate penetrance mutations exerting their effect in a high-risk environment reflected by high BMI. WES enables identification of genetic risk factors of relevance to both cancer development and tailored therapeutic intervention in a single genetic test.
- ItemDevelopment of a novel pre-screen algorithm for cardio-metabolic risk management using a genomics database resource(Stellenbosch : Stellenbosch University, 2016-03) Luckhoff, Hilmar Klaus; Kotze, Maritha J.; Janse van Rensburg, Susan; Stellenbosch University. Faculty of Medicine and Health Sciences. Dept. of Pathology. Anatomical Pathology.ENGLISH ABSTRACT: The timely assessment and treatment of dyslipidaemia is an important component of cardiovascular risk screening and intervention. The apolipoprotein E (APOE) ε-2/ε-3/ε-4 polymorphism associated with impaired lipid homeostasis provides a genetic link between cardiovascular disease (CVD) and late-onset Alzheimer’s disease (AD). Realization that the phenotypic expression of the risk associated APOEε-2 and ε-4 alleles may be dependent on non-genetic factors supports the inclusion of APOE genotyping in chronic disease screening programs. The lack of well-defined selection criteria for APOE genotyping, however, limits the use of this biomarker in clinical practice. The aim of the present study was to develop a pre-screen algorithm for identification of a target population most likely to benefit from APOE genotyping, performed in conjunction with a clinical and lifestyle assessment. Towards this goal, comprehensive patient data were evaluated from a total of 580 unrelated Caucasians enrolled in a chronic disease screening program over a five-year period (2010-2015), using an ethically approved study questionnaire. Biochemical tests performed according to standard laboratory protocols were extracted from the research database. All study participants were genotyped for the APOE ε- 2/ε-3/ε-4 polymorphism. APOE genotype distribution differed significantly (p<0.05) between study participants with and without a family history of AD. A positive association between dietary fat intake and lowdensity lipoprotein (LDL) cholesterol (p=0.001), as well as an inverse association with highdensity lipoprotein (HDL) cholesterol (p=0.002), were observed in patients with a family history of AD. Body mass index (BMI) was positively associated with LDL cholesterol and inversely associated with HDL cholesterol levels (p<0.001), irrespectively of an AD family history. Smoking was associated with higher triglycerides (p<0.001) and lower HDL cholesterol levels (p=0.004) in the total study group. Alcohol intake was positively associated with BMI (p=0.008) as well as triglyceride levels (p=0.021) in patients with a positive family history of AD. The clinical expression of a hypercholesterolaemic phenotype in APOE ε-4 allele carriers, as well as apparent mitigation by regular physical activity, were dependent on the interaction between a family history of AD and APOE genotype (p<0.001). APOE ε-2 carriers without an AD family history showed a significant increase in triglyceride levels (p=0.014). The modulating influence of APOE ε-4 on the relationship between alcohol intake and BMI as well as total cholesterol levels was also dependent on the presence or absence of AD family history (p<0.05). This study resulted in the addition of a family history of AD as a novel component to the prescreen algorithm developed for selection of at-risk individuals prior to APOE genotyping performed as part of a chronic disease screening program. The lifestyle questionnaire used in this study furthermore facilitated interpretation of the clinical relevance of variation detected in the APOE gene. This is important to prioritize the use of lipid-lowering medication towards patients with severe subtypes of dyslipidaemia such as familial hypercholesterolaemia (FH), which remains largely undiagnosed and untreated in the highrisk South African population. Incorporating the research findings into clinical practice would suggest that physical activity may be the most effective risk reduction strategy in carriers of the APOEε-4 allele, as supported by international studies.
- ItemEvaluation of high-throughput methodology for multi-gene screening in patients with Non-Alcoholic Fatty Liver Disease (NAFLD)(Stellenbosch : Stellenbosch University, 2011-12) Fisher, Leslie Reginald; Kotze, Maritha J.; Kruger, F. C.; Stellenbosch University. Faculty of Health Sciences. Dept. of Pathology.ENGLISH ABSTRACT: Non-Alcoholic Fatty Liver Disease (NAFLD) is the most prevalent chronic liver disease in Western countries and is considered the hepatic manifestation of the Metabolic Syndrome (MetS). Its heterogeneous nature ranges from hepatic steatosis through steatohepatitis to advanced fibrosis and cirrhosis where the ingestion of significant amounts of alcohol has been excluded. The disease profile of NAFLD and its necro-inflammatory subset Nonalcoholic Steatohepatitis (NASH) were described in the parent study, which provided a clinically well-characterised patient cohort for the present investigation. South African patients with NASH had significantly higher mean serum cholesterol and triglyceride levels than those with fatty liver only. The objective of this study was to implement a high-throughput real-time polymerase chain reaction (PCR) method in our laboratory to enable the assessment of cardiovascular genetic risk factors in NAFLD patients. The specific aims were to determine the clinical utility and perform analytical validation of each mutation included in the multi-gene cardiovascular disease (CVD) screening assay. The Pathology Supported Genetic Testing (PSGT) concept developed at our department provides a practical approach to personalized medicine. The CVD multi-gene screen analyses key metabolic pathways relating to atherogenic dyslipidaemia, chronic inflammation, hypercoagulation and iron dysregulation implicated in insulin resistance, which is known to be a universal factor in the pathogenesis of NAFLD. Deleterious low-penetrance mutations in the APOE (APOE2 and E4 alleles), MTHFR (677C>T and 1298A>C), F2 (20210G>A), FV (1691G>A, Leiden) and HFE (C282Y and H63D) genes were included for analysis due to their important role as genetic contributors to these biological processes. A total of 178 patients diagnosed with NAFLD and 75 controls were studied using direct DNA sequencing and a RT-PCR system for mutation detection. In addition, two patients with high ferritin levels were included as case studies. A significant association was found between HFE mutations and elevated Alanine Transaminase (ALT) levels in the NAFLD population (p = 0.04). This discovery is interpreted as the identification of a subset of patients at greater risk of developing progressive liver damage who would benefit most from genetic testing to direct more aggressive therapy at an earlier stage. The necessity of an integrative, systems-based network approach was demonstrated to more accurately distinguish between Hereditary Haemochromatosis (HH) and Insulin Resistance-associated Hepatic Iron Overload (IR-HIO) syndrome in obese patients. The PSGT approach to personalized medicine facilitates diagnosis of CVD subtypes, prevention of cumulative risk and the formulation of gene-based intervention programs tailored to the needs of the patient. These findings support the clinical utility of the CVD multi-gene test to guide chronic disease risk management in patients with NAFLD. The HFE mutation detection component of this test is of particular relevance in directing an effective treatment strategy in patients with a medical history of CVD and/or high iron stores.
- ItemGenetic aspects of pre-eclampsia : mutation screening of the low-density lipoprotein receptor, methylenetetrahydrofolate reductase, prothrombin and factor V candidate genes(Stellenbosch : Stellenbosch University, 2001-03) Gebhardt, G. S.; Odendaal, H. J.; Hillermann, R.; Kotze, Maritha J.; Stellenbosch University. Faculty of Medicine & Health Sciences. Dept. of Pathology. Anatomical Pathology.ENGLISH ABSTRACT: Pre-eclampsia is a condition unique to pregnancy and primarily affects the maternal and placental vascular endothelium. It has significant morbidity and mortality consequences for both mother and infant. Despite global research into the aetiology of the condition, the cause for this condition remains unknown. Several factors, including a strong family history of hypertension in pregnancy point to a familial or genetic component in the pathophysiology of this complication. The purpose of this research project was to investigate candidate genes implicated in endothelial damage. Common methylene-tetra-hydrofolate reductase (MTHFR) gene mutations C677T and A1298C, factor V Leiden mutation R506Q and prothrombin mutation A20210G were investigated in 50 patients with an uncomplicated pregnancy outcome (controls) and 350 patients with various clinical manifestations of preeclampsia, including severe, early onset forms and abruptio placentae. Fasting homocystein levels were determined biochemically on all participants. In addition, 126 consecutive pregnant patients were recruited at booking, fasting lipograms were performed on them as well as mutation screening of 7 common mutations in the low-density lipoprotein receptor gene. This was correlated with eventual pregnancy outcome, and those with an uncomplicated outcome were selected as an additional control group. A significant association between hyperhomocysteinaemia and early onset severe pre-eclampsia could be demonstrated. Mutant allele T of the C677T mutation could be associated with hyperhomocysteinaemia but not with pre-eclampsia whilst mutant allele C of mutation A1298C demonstrated a significant correlation with diastolic blood pressure. In addition, combined heterozygosity for these mutations may serve as a marker for abruptio placentae.
- ItemIdentification of clinically-informative biomarkers within the spectrum of gastro-oesophageal reflux disease in the South African population(Stellenbosch : University of Stellenbosch, 2006-03) Van Rensburg, C. J.; Kotze, Maritha J.; Wright, C.; De Jong, G.; University of Stellenbosch. Faculty of Health Sciences. Dept. of Pathology. Anatomical Pathology.Patients with chronic gastro-oesophageal reflux disease are predisposed to Barrett’s metaplasia and oesophageal adenocarcinoma. The availability of molecular markers that can objectively identify patients with Barrett’s oesophagus at increased risk of carcinoma is highly desirable. A literature search was conducted to identify potentially useful biomarkers for genotype-phenotype correlation studies in South African patients with Barrett’s oesophagus. The COX-2, c-myb and c-myc genes selected for mRNA expression analysis were analysed in 26 patients with Barrett’s metaplasia (BM) without dysplasia; 14 with Barrett’s oesophagus and dysplasia (BD); 2 patients with Barrett’s adenocarcinoma (BAC); 19 with erosive oesophagitis (ERD); 25 with non-erosive oesophagitis (NERD) and 19 control individuals with a normal gastroscopy and no gastro-oesophageal reflux disease (GORD) symptoms. In the BD/BAC group, 69% (11/16) showed increased c-myb mRNA expression compared with 35% (9/26) in the BM group (p = 0.03). A statistically significant difference (p = 0.002) in c-myb expression was also observed between Barrett’s patients (20/42, 48%) and the control groups (9/63, 14%). In the BD patients, 21% (3/14) had increased c-myc mRNA expression compared with none in those with BM (p < 0.05) and BAC. No significant differences in mRNA expression levels were observed between ethnic groups for the genes analysed. In an attempt to determine whether the low expression level of c-myc in the study cohort may be related to possible gene-gene interaction, DNA samples of 199 individuals were subjected to genotyping of the functional GT-repeat polymorphism in the promoter region of the NRAMP1/SLC11A1 gene. Both these genes are involved in iron metabolism and c-myc is known to repress NRAMP1/SLC11A1. Genotype and allele frequencies were similar in all the groups studied with the 3/3 genotype being the most common. However, none of the three above-mentioned BD patients with increased c-myc mRNA expression had the 3/3 genotype. Therefore, although small in number, c-myc-NRAMP1/SLC11A1 interaction may be of adverse significance in patients with allele 2. TP53 mutation analysis was performed on 68 Barrett’s patients, and TP53 immuno-staining on oesophageal biopsy specimens of 55 subjects. Sporadic TP53 mutations were not identified in any of the patients with BM or dysplasia without BAC. Immuno-histochemistry staining of 2+ and 3+ intensity was similar in patients with metaplasia and dysplasia (58%). The low mutation frequency and relative non-specificity of TP53 immunostaining observed in Barrett’s patients seem to preclude its widespread use as a screening tool. TP53 mutation detection may however be useful for risk stratification once dysplasia has been diagnosed, as mutations G245R and D281Y were identified in two patients with BAC. Of the genes studied in the South African population, c-myb represents the most useful marker for early detection of an increased cancer risk in Barrett’s patients. In future, patients with Barrett’s oesophagus may benefit from genetic assessment to complement existing cancer surveillance and treatment strategies.
- ItemThe MED-PED project : presymptomatic diagnosis in families with disease- related LDL receptor gene mutations(Stellenbosch : Stellenbosch University, 2000-03) Vergotine, Joseph Vincent; Kotze, Maritha J.; De Jong, G.; Stellenbosch University. Faculty of Medicine & Health Sciences . Dept. of Pathology.ENGLISH ABSTRACT: Familial hypercholesterolaemia (FH) contributes significantly to the high death rate from cardiovascular disease worldwide. FH is a common autosomal co-dominant disease characterised by raised cholesterol levels and premature coronary heart disease (CHD). Whilst these features usually are very prominent in homozygotes the clinical diagnosis of heterozygotes is complicated by variable phenotypic expression. Specific founder genes in the low-density lipoprotein receptor (LDLR) gene have increased the prevalence of FH in South African Afrikaners, Indians, Jews and Coloureds, and screening for these known mutations allows unequivocal diagnosis of FH-affected individuals. The systematic molecular analysis of FH resulted in the identification of at least ten founder-type LDLR gene mutations among the 56 different gene defects described to date in the diverse South African population. DNA screening of 792 at-risk family members for the FH-related mutations identified in 379 index cases, allowed accurate disease diagnosis in an additional 340 relatives and exclusion of the relevant mutation in 452 individuals. This effort forms part of the MED PED FH initiative, a collaborative project to "Make Early Diagnosis and Prevent Early Deaths in MEDical PEDigrees with FH". Evaluation of clinical criteria versus DNA diagnosis of three founder-related mutations (D154N, D206E and V408M) in the South African population demonstrated that the sensitivity and specificity of diagnoses, based on total cholesterol values measured in family members of index cases recruited for this study, were 88% and 77%, respectively. A population-directed DNA diagnosis of FH is therefore justified in South Africa on a routine basis, since expression of the defective gene measured in biochemical tests does not allow accurate diagnosis of FH in all cases. The application of mutation detection was illustrated by prenatal diagnosis of FH performed for a couple who are both heterozygous for the most common Afrikaner mutation, D206E. The mutation was absent in the foetus and a normocholesterolaemic infant was born. Prenatal diagnosis of FH, aimed at the detection of homozygous cases, is particularly applicable in populations and families with molecularly defined LDLR gene mutations. The MED-PED approach resulted in accurate diagnosis and subsequent treatment of FH in more patients, and referral to lipid clinics where they could receive the intensive care their condition justifies. Molecularly diagnosed FH patients will be the first to benefit from future treatment approaches based on mutation type.
- ItemMolecular-genetic analysis of Hirschsprung's disease in South Africa(Stellenbosch : Stellenbosch University, 2000-03) Julies, Monique G.; Moore, S. W.; Kotze, Maritha J.; Du Plessis, L.; Stellenbosch University. Faculty of Medicine & Health Sciences. Dept. of Pathology.ENGLISH ABSTRACT: Hirschsprung's disease, or aganglionic megacolon, is a common cause of intestinal obstruction in neonates and is associated with the congenital absence of intrinsic ganglion cells in the myenteric and submucosal plexuses of the gastrointestinal tract. The affected area is usually restricted to the distal part of the colon (short segment disease), but total colonic or intestinal involvement occurs in some patients (long segment disease). DNA analysis was performed on samples from 53 unrelated sporadic HSCR patients to search for mutations in RET proto-oncogene, endothelin-B receptor (EDNRB) and endothelin-3 (EDN3) genes. The patients were from different ethnic groups in South Africa, including 29 coloured, 14 white (Caucasian) and 9 black individuals. The origin of 1 patient was unknown. PCR HEX-SSCP analysis of the RET protooncogene revealed one previously described (P973L) and five novel mutations (V202M, E480K, IVS10-2A1G, D771N, IVS19-9Crr), likely to cause or contribute to the HSCR phenotype. Nine polymorphisms were also identified in the RET protooncogene, of which four were novel (IVS6+56deIG, IVS13-29Crr, IVS16-38deIG, X1159) and five previously described (A45, A432, L769, S904, R982). All the mobility shifts detected in the EDNRB gene represented polymorph isms (A60T, S184, 1187, V234, L277, IVS3-6Crr, IVS4+3A1G). No sequence variants were identified in the EDN3 gene. The majority of mutations in the RET proto-oncogene (28.6%) were identified in coloured patients while no mutations were identified in black patients. A mutation in RET was identified in two of 14 patients (14%) presenting with HSCR and Down's syndrome compared to 6 mutations identified in 9 of 39 patients (23%) with only HSCR. The fact that Down's syndrome patients have a high chance of developing HSCR, implies the involvement of modifier gene(s) in a HSCR/Oown's syndrome phenotype. This study demonstrated that, within the South African HSCR patient population, the RET proto-oncogene is the major susceptibility gene, whereas EDNRB and EDN3 may contribute only to a minority of cases. In 81% of patients no disease-causing mutation could be identified, which is in keeping with the heterogeneous nature of HSCR. The identification of mutations in HSCR patients would in future lead to improved and accurate counselling of South African HSCR patients and their families.