Cytogenetics at the University of Cape Town: A 45-year journey.

This article is a brief record of the cytogenetics laboratory from its birth in 1971, under the auspices of the University of Cape Town, throughout its development within the Department of Human Genetics, under the leadership of Professor Peter Beighton, to its present position at Groote Schuur Hospital, as a multidisciplinary unit run by the National Health Laboratory Service.

Inherited retinal disorders in South Africa and the clinical impact of evolving technologies.

Retinal degenerative disorders (RDDs) encompass a group of inherited diseases characterised by vision loss. The genetic and clinical complexity poses a challenge in unravelling the molecular genetic aetiology of this group of disorders. Furthermore, the population diversity in South Africa (SA) presents researchers with a particularly complicated task. Rapid advances in the development of cutting-edge technological platforms over the past two decades, however, have assisted in overcoming some of the challenges. The RDD research team has utilised these escalating technologies, which has facilitated a corresponding increase in molecular diagnoses. A biorepository has been established and comprises ~3 200 patient DNA samples archived with many forms of RDD (including retinitis pigmentosa, macular dystrophies, Stargardt disease, Leber congenital amaurosis, Usher syndrome and Bardet Biedl syndrome). A comprehensive review is presented of the SA journey spanning 25 years, into elucidating the molecular genetic basis of various forms of RDD in SA.

The hereditary ataxias: Where are we now? Four decades of local research.

The hereditary ataxias have been studied at the University of Cape Town for more than 40 years, following from initial clinical investigations by Beighton and colleagues in the early 1970s. This group of inherited disorders is characterised by progressive neurodegeneration and associated symptoms, including the inability to coordinate movement. Following initial local and international linkage studies, and the discovery of the genes responsible for the key dominant and recessive inherited ataxias in the 1990s, a local molecular testing service was established at Groote Schuur Hospital. More than 1 600 individuals have been referred through this testing service (now offered by the National Health Laboratory Service), leading to the molecular diagnosis of 253 families with spinocerebellar ataxia types 1, 2, 3, 6 or 7, and 30 families with Friedreich's ataxia. This is likely to be an under-representation of the number of South Africans affected with hereditary ataxia, and future research efforts will focus on increasing the awareness of this group of disorders, both locally and throughout the rest of Africa. Next-generation technologies will be beneficial in identifying additional genes underlying inherited ataxia in indigenous patients to enable more appropriate management and treatment of individuals with molecularly undiagnosed forms of the disease.

The rise of developmental genetics - a historical account of the fusion of embryology and cell biology with human genetics and the emergence of the Stem Cell Initiative.

Genetics and cell biology are very prominent areas of biological research with rapid advances being driven by a flood of theoretical, technological and informational knowledge. Big biology and small biology continue to feed off each other. In this paper, we provide a brief overview of the productive interactions that have taken place between human geneticists and cell biologists at UCT, and credit is given to the enabling environment created led by Prof. Peter Beighton. The growth of new disciplines and disciplinary mergers that have swept away division of the past to make new exciting syntheses are discussed. We show how our joint research has benefitted from worldwide advances in developmental genetics, cloning and stem cell technologies, genomics, bioinformatics and imaging. We conclude by describing the role of the UCT Stem Cell Initiative and show how we are using induced pluripotent cells to carry out disease-in-the- dish studies on retinal degeneration and fibrosis.

Duchenne muscular dystrophy in the Western Cape, South Africa: Where do we come from and where are we going?

Duchenne muscular dystrophy (DMD) is one of the most common and severe of the inherited dystrophies, with an incidence of 1 in 3 500 live, male births worldwide. Becker muscular dystrophy (BMD) has a lower incidence of 1:14 000 - 18 000 boys and a milder progression and longer life expectancy. Over the last two decades, better understanding of the underlying disease aetiology as well as major advances in medical technology have brought about significantly improved genetic diagnosis and clinical care for B/DMD patients. Exciting developments in the field of gene-based therapies have once again put B/DMD in the limelight, with renewed focus on the importance of comprehensive genetic testing protocols. We present a historical overview of the medical and molecular service for B/DMD offered over the last three decades in South Africa, specifically in the Western Cape, from a clinical as well as a laboratory perspective.

Spinocerebellar ataxia type 7 in South Africa: Epidemiology, pathogenesis and therapy.

Disorders of the nervous system represent a significant proportion of the global burden of non-communicable diseases, due to the trend towards ageing populations. The Department (now Division) of Human Genetics at the University of Cape Town (UCT) has been involved in pioneering research into these diseases since the appointment of Prof. Peter Beighton as Head of Department in 1972. Beighton's emphasis on understanding the genetic basis of disease laid the groundwork for investigations into several monogenic neurodegenerative conditions, including Huntington's disease and the polyglutamine spinocerebellar ataxias (SCAs). In particular, SCA7, which occurs at an unusually high frequency in the South African (SA) population, was identified as a target for further research and therapeutic development. Beginning with early epidemiological surveys, the SCA7 project progressed to molecular genetics-based investigations, leading to the identification of a founder effect in the SA SCA7 patient population in the mid-2000s. Capitalising on the founder haplotype shared by many SCA7 patients, UCT researchers went on to develop the first population-specific gene-silencing approach for the disease. More recently, efforts have shifted to the development of a more accurate model to decipher the precise mechanisms of neurodegeneration, using induced pluripotent stem cells derived from SA SCA7 patients. In many ways, the SA SCA7 journey reflects the legacy and vision of Prof. Peter Beighton, and his efforts to establish world-class, collaborative research into diseases affecting the African continent.

Duchenne muscular dystrophy: High-resolution melting curve analysis as an affordable diagnostic mutation scanning tool in a South African cohort.

BACKGROUND: Duchenne/Becker muscular dystrophy (D/BMD) is an X-linked recessive muscle disorder affecting 1/3 500 live male births worldwide. Up to 70% of all D/BMD cases are caused by exonic deletions or duplications routinely identified in diagnostic laboratories worldwide.The remaining patients harbour other sequence alterations for which testing availability is limited owing to the expense of interrogating the large DMD gene. Genetic screening for D/BMD in South Africa currently includes multiple ligase-dependent probe amplification (MLPA) for exonic deletions and duplications and linkage analysis. No genetic testing for small mutations in the DMD gene is offered, leaving a third of D/BMD families without genetic closure. The advent of potential mutation-specific therapies for DMD necessitates comprehensive testing protocols. OBJECTIVE: To investigate the effectiveness and affordability of high-resolution melting curve analysis (hrMCA) for detection of small/point mutations in the DMD gene, for possible inclusion into the local public health-funded diagnostic service. METHODS: DNA from 24 patients who had previously tested deletion-negative with multiplex polymerase chain reaction (mPCR) was analysed by MLPA and hrMCA. RESULTS: MLPA revealed eight previously undetected exonic rearrangements: five deletions and three duplications. HrMCA of the remaining samples revealed three nonsense, four frameshifts, one splice-site, one missense and one single-base substitution in the Dp427promoter/exon1 of the DMD gene. In addition, 41 polymorphisms and three changes of uncertain significance were detected. CONCLUSION: These findings identify hrMCA as an affordable and effective mutation scanning tool for incorporation into the local diagnostic setting, allowing for better genetic counselling of more DMD families and selection of potential candidates for future therapies.

A new class of stem cells in South Africa: iPS cells.

Created from adult rather than embryonic cells, induced pluripotent stem (iPS) cells represent a breakthrough in stem cell science, and their pioneers have been recognised with the 2012 Nobel Prize in Medicine. These cells offer new hope in the treatment of pathogenetic diseases, but there is still a way to go on the road to effective therapeutic applications.

Inherited polyglutamine spinocerebellar ataxias in South Africa.

OBJECTIVE: To determine the frequency and distribution of polyglutamine spinocerebellar ataxias (SCAs) from referrals over a 24-year period to the National Health Laboratory Service (NHLS) in South Africa (SA). METHODS: Paper-based clinical reports in the University of Cape Town laboratory and the NHLS electronic patient record database spanning a 24-year period were mined for information regarding the molecular diagnosis, ethnicity and CAG repeat length for individuals referred for molecular genetic testing for the polyglutamine SCAs. RESULTS: SCA1 and 7 are the most frequent types of polyglutamine SCA in the SA patient population, followed by SCA2, 3 and 6. SCA1 is the most common type in the coloured, white and Indian populations, whereas the majority of indigenous black African patients are affected with SCA7 and 2. Of individuals tested, 22% were found to be positive for one of the polyglutamine SCAs. CONCLUSION: Although trends in the frequency and distribution of the polyglutamine SCAs in SA have not changed significantly since our previous study in 2003, they differ remarkably from those reported elsewhere, and reflect the unique genetic and demographic background of SA. The provision of accurate and complete patient information and family history is crucial to the diagnostic process, to enable comprehensive epidemiological studies and assist in developing therapeutic and patient management strategies.

Predictive testing for Huntington disease in a developing country.

Predictive testing for Huntington disease (HD), by means of direct mutation analysis, has been offered at the Division of Human Genetics, University of Cape Town, from 1995. The aim of this study was to compile a comprehensive profile of the participants who had undergone predictive testing in the Western Cape from 1995 to 2005. The sociodemographic data, uptake and outcome of tests were analyzed to inform changes to improve the current genetic counseling services. A retrospective cross-sectional design using a 'multi-method' approach of both qualitative and quantitative methods was used. Data were gathered from the participants' hospital files and genetic database. Psychosocial data were obtained by face-to-face interviews with the participants in their homes or venues of choice. A total of 36 predictive tests were performed. The uptake for predictive testing was approximately 4.5% of the estimated at-risk population. The cohort of 27 individuals comprised 16 females and 11 males. Their mean age was 35.3 years; 6 were mixed ancestry and 21 were White people (European ancestry); 11 tested gene positive, 15 gene negative and 1 was in the reduced penetrance range. The most important issue identified was that the uptake of individuals classified as mixed ancestry was substantially lower than that of the White people possibly due to limited access to the predictive testing program because of the low levels of income and education in the general population of families with HD. Strategies to address these aspects have been incorporated into the program and will be reassessed after 1 year.

Arg120stop nonsense mutation in the RP2 gene: mutational hotspot and germ line mosaicism?

Mutations in the RP2 gene account for up to 20% of X-linked recessive retinitis pigmentosa (RP). Arg120stop is to date the most frequently reported mutation found in RP2. Mutation screening was performed during the course of a large screening program of retinal degenerative disorders (RDDs) in South Africa using exon 1 and 2 of RP2 in 20 unrelated families with an X-linked mode of retinal degenerative inheritance. Direct sequencing analysis revealed a C-->T transition at position 358 in the proband in a family of German origin. Subsequent analysis revealed that this Arg120stop mutation cosegregated with the disease in an additional affected family member. The nonsense mutation, Arg120stop, could not however, be detected in the somatic cells of the obligate carrier female. This, the first report of a germ line mutation for a family with RP, has many implications for genetic counseling of retinal degeneration (RD). To avoid inaccurate risk assessment for RP due to epigenetic events, such as the rare occurrence of germ line mosaicism, genetic counseling in families with XLRP should always be guided by molecular testing.

A computer-based register for inherited retinal dystrophies in Southern Africa.

A genetic register for inherited retinal degenerative disorders (RDDs) has been established at the Division of Human Genetics, UCT Medical School, Cape Town, South Africa. The primary role of the register is to monitor the progress of molecular research and to facilitate the efficient delivery of services, including genetic counselling, to respective family members and new patients. The database currently holds information on 1829 subjects. The RDD-specific breakdown of the data are presented.

Ethnic variation and in vivo effects of the -93t-->g promoter variant in the lipoprotein lipase gene.

Recently, a (t-->g) transition at nucleotide -93 in the lipoprotein lipase (LPL) gene promoter has been observed in Caucasians. Here, we have compared the frequency of the -93g carriers in three distinct populations (Caucasians, South African Blacks, and Chinese). The carrier frequency in the Caucasian population was 1.7% (4/232), which was in contrast to the South African Black population, which had a frequency for this allele of 76.4% (123/161) of the individuals tested. This transition was not observed in the Chinese population under study. Near complete linkage disequilibrium between the -93g and the previously described D9N mutation was observed in the Caucasian population but not in South African Blacks. To further assess the ancestral origins of these DNA changes, DNA haplotyping using a CA repeat 5' to these substitutions was performed. The -93t allele was associated with only a few specific dinucleotide repeat sizes. In contrast, the -93g allele occurred on chromosomes with many different repeat lengths. The broad distribution of repeats on -93g carrying chromosomes, their high frequency in the South African Black population, and the conservation of the -93g allele among different species may suggest that the -93g allele is the ancestral allele on which a transition to t and the D9N mutations arose. The very high frequency of the -93g allele distinct from the N9 allele in a cohort of Black South Africans allowed us to specifically assess the phenotypic effects of the -93g allele on lipids. Individuals homozygous for the g allele at -93 showed mildly decreased triglycerides compared with individuals homozygous for the t allele (1.14 +/- 0.66 mmol/L versus 0.82 +/- 0.3; P = .04). Thus, the -93g allele in this cohort is associated with low plasma triglyceride levels.

Genetic linkage between Huntington disease and the D4S10 locus in South African families: further evidence against non-allelic heterogeneity.

A study of genetic linkage between Huntington disease (HD) and the D4S10 locus (G8) has been undertaken in 10 South African (SA) families originating from the black, white and mixed acestry population groups. Allele frequencies at the D4S10 locus have been established in the non-Caucasoid population groups. There are significant differences in the allele frequencies at the D4S10 locus between the various SA populations. Clearly, information about population-specific frequencies for all polymorphisms is essential prior to the implementation of predictive testing in different population groups. Linkage has been demonstrated within this mixed group of HD families in SA using the HindIII, EcoRI and MspI polymorphisms, detected by G8. A maximum lod score of 8.14 at a recombination fraction of 0.00 (confidence limit 0-0.058) has been calculated using a combined haplotype of the HindIII and MspI polymorphisms. Taking into account the diverse ethnic backgrounds of the different SA population groups in this investigation, the data obtained from the study provide further evidence that there is probably only a single HD locus.

Tumor marker kinetics in the monitoring of breast cancer.

Controversy exists in using carcinoembryonic antigen (CEA) for monitoring the clinical course of breast cancer. In this study, the kinetics of two plasma tumor markers, CEA and CA15-3, immediately after the initiation of chemotherapy were assessed in 30 patients with advanced breast cancer. Four distinct kinetic patterns were seen. Two patterns fitted the expected relationship where the plasma marker increased during tumor progression (nine patients), and declined in tumor regression (five patients). The third pattern was paradoxical in that objective tumor regression in eight patients was associated with an acute surge of these markers followed by a steady decline. The doubling times for both CEA and CA15-3 were immediately shortened four-fold after therapy suggesting tumor cytolysis in treatment responders. Equally paradoxical was the fourth pattern where tumor progression in eight patients was associated with a rapid and transient decline of markers followed by rebounds. Such a rapid decline may be due to a suppression of marker release, as demonstrated in an in vitro study. Adequate knowledge of these putative paradoxical patterns will permit their effective use in monitoring the disease course and perhaps in the early prediction of the therapeutic response.