225 research outputs found
Identifying Genes and Novel Variants Involved in Nonsyndromic Hearing Impairment, and Assessment of the Psychosocial Burden of Hearing Impairment in Cameroon
No full textBackground Hearing impairment (HI) is the most common sensory disability and occurs in about 1 per 1000 live births in high-income countries, with a much higher incidence of up to 6 per 1000 live births in sub-Saharan Africa (SSA). HI can be due to environmental or genetic causes, and in many cases, it is not possible to establish a definite aetiology. Hereditary HI contributes to 30% to 50% of HI cases in SSA. Hereditary HI can be syndromic or non-syndromic, depending on whether it is associated with additional abnormalities in other organs or not. Non-syndromic HI (NSHI) accounts for 70% of hereditary hearing loss, and is genetically highly heterogeneous, with approximately 170 loci and 121 genes identified to date. Studies in European and Asian populations have identified pathogenic variants in GJB2 (MIM: 121011), and GJB6 (MIM: 604418) genes as the major contributors to autosomal recessive NSHI (ARNSHI). The genetic aetiology of HI in Cameroon is unclear, as previous studies have found no contribution of GJB2 and GJB6 genes to NSHI in Cameroon. However, patients included in those studies consisted of both familial and isolated cases, therefore, underlying environmental/multifactorial causes in some cases cannot be excluded (especially for the isolated cases). Six loci for X-linked HI have been described to date, including DFNX3 (Xp21.2), where DMD is located. Variants in DMD in humans are known to be responsible for Duchenne muscular dystrophy (DMD; MIM: 310200), and Becker muscular dystrophy (BMD; MIM: 300376), an Xlinked recessive disorder. Previous studies have demonstrated that mdx mice, (an animal knockout model for DMD), have an increased threshold for hearing when compared to wildtype mice. However, the contribution of DMD to HI in humans has not been extensively studied. Besides, most of the previous studies on DMD were conducted in Caucasians, Asians, and Arabs; therefore, little is known about the features of this condition in Africans. Parents of children with HI tend to face challenges of parenting especially in terms of communication and social interaction. In Africa, parent's perceived causes of deafness vary from environmental factors to mysterious (“evil forces”) or superstitious beliefs. Also, the attitude of the society towards people with HI does not encourage their participation and involvement in the community, as they face overt discrimination. Aim and methods The aim of this project was to examine the genetic aetiologies of HI in the Cameroonian population, and undercover the challenges faced by persons with HI in Cameroon and their understanding of the causes of HI. This was addressed by 1) Establishing the current status of knowledge on HI in Africa (in terms of prevalence, aetiologies, and genetics aspects) with a particular focus on Cameroon, and assessing the contribution of connexin genes to HI in humans at a global level, through systematic literature reviews; 2) Revisiting the contribution of GJB2 and GJB6 genes to NSHI in 29 multiplex Cameroonian families with NSHI and with strong evidence of non-environmental causes, through targeted gene sequencing and specific multiplex polymerase chain reaction (PCR); 3) Using multiplex ligand-dependent probe amplification (MLPA) technique to investigate the most common variants associated with DMD in Cameroon and assess their possible implication in HI in humans; 4) Performing whole exome sequencing (WES) on 2 Cameroonian multiplex families with NSHI and who tested negative for pathogenic variants in GJB2 and GJB6, to identify the underlying causative genes; 5) Performing in-depth interviews to gain an understanding of the challenges faced by people with HI in Cameroon, their understanding of the causes of hearing impairment (HI), and how challenges could be remedied to improve the quality of life of persons with HI. Results Literature reviews Our first systematic review showed that HI is a public health issue in Cameroon, especially in the elder population where the prevalence of HI is 14.8% in people aged 50 years and more. Environmental factors, including meningitis, impacted wax, and age-related disorders are the leading aetiologies of HI in Cameroon as in many other SSA countries, contributing 52.6% to 62.2% of HI cases. Hereditary HI comprises 0.8% to 14.8% of all cases in Cameroon, and in 32.6% to 37% of HI cases, the origin remains unknown. This contrasts with findings from highincome countries where hereditary HI constitutes the main aetiology of HI, contributing to approximately 50% of cases. NSHI is the most frequent clinical entity and accounts for 86.1% to 92.5% of cases of hereditary HI in the Cameroonian population. No pathogenic variant was described in GJB6 gene, and the prevalence of pathogenic variants in GJB2 ranged from 0% to 0.5%. The prevalence of pathogenic variants in other known NSHI genes was with type 2 Waardenburg syndrome, and three cases of type 2 Usher syndrome were identified in one family. By direct gene sequencing of the coding region of GJB2, no variants were found in any of the 29 families with NSHI. Additionally, through a specific multiplex PCR, the GJB6- D3S1830 deletion which contributes to 9.7% of NSHI cases in Europeans was not identified in any of the patients with HI. Subsequently, a total of 17 males with DMD from 14 families were recruited, aged 14 ± 5.1 (8–23) years. The mean age at onset of symptoms was 4.6 ± 1.5 years, and the mean age at diagnosis was 12.1 ± 5.2 years. Proximal muscle weakness was noted in all patients and calf hypertrophy in the large majority of them (88.2%; 15/17). Flexion contractures were particularly frequent on the ankle (85.7%; 12/14). Wasting of the shoulder girdle and thigh muscles was present in 50% (6/12) and 46.2% (6/13) of patients, respectively. No patient presented with HI. The MLPA found that deletions of at least one exon in DMD occurred in 45.5% of patients (5/11), while duplications were observed in 27.3% (3/11). Both variant types were clustered between exons 45 and 50, and the proportion of de novo variant was estimated at 18.2% (2/11). Whole exome sequencing We submitted DNA samples from five members of a multiplex non-consanguineous Cameroonian family segregating prelingual and progressive ARNSHI for WES. We identified novel bi-allelic compound heterozygous pathogenic variants in CLIC5 (MIM: 607293). The variants identified, i.e. the missense [NM_016929.5:c.224T>C; p.(Leu75Pro)] and the splicing (NM_016929.5:c.63+1G>A), were validated using Sanger sequencing in all seven available family members and co-segregated with HI in the three family members with HI. The three affected individuals were compound heterozygous for both variants, and all unaffected individuals were heterozygous for one of the two variants. Both variants classify as pathogenic by the American College of Medical Genetics (ACMG) guidelines for classification of variants and are absent from the genome aggregation database (gnomAD), UK10K, Greater Middle East (GME) database, and the Single Nucleotide Polymorphism Database (dbSNP), as well in 122 healthy controls from Cameroon. We also did not identify these pathogenic variants in 118 unrelated sporadic cases of NSHI from Cameroon. A second multiplex family was also screened through the use of WES, followed by direct Sanger sequencing in additional patients and control participants. We identified a heterozygous novel missense variant [NM_001174116.2:c.918G>T; p.(Gln306His)] in DMXL2 (MIM:612186) which was transmitted in an autosomal dominant manner, and co-segregates with congenital/prelingual profound to total non-syndromic sensorineural HI in a family from Cameroon. The described family showed a variable expressivity of the HI phenotype. The p.(Gln306His) variant which substitutes a highly conserved glutamine residue is predicted deleterious by various bioinformatics tools and is absent from several genome databases including genome aggregation database (gnomAD), and trans-omics for precision medicine (TOPMed) database. This variant was neither found in 121 healthy controls without personal or family history of HI, nor 112 sporadic cases of NSHI from Cameroon. Our study identified novel variants in CLIC5 and DMXL2 in two Cameroonian families, and provided only the second report of variants in these genes worldwide; thus, strengthening the case for these two genes as candidate genes for NSHI in humans. The psychosocial burden of HI We performed in-depth interviews with 10 HI professionals (healthcare workers, and educationists), and 10 persons affected by HI (persons with HI, and caregivers). The results show that in this study population, the cause of HI is attributed to a variety of causes, including genetics, environmental factors, and a spiritual curse. There were reported cases of stigma and discrimination with persons with HI in the Cameroonian population sometimes seen as having a “mental disorder”. Our participants also highlighted the difficulty that persons with HI have in accessing the necessary education and healthcare services, and suggested the need for policymakers and researchers to develop strategies to improve the social integration of persons with HI and their access to basic social services. This includes 1) Increased awareness amongst the general population, 2) the establishment of more special schools, and 3) building and equipping facilities for proper management of HI. Conclusions Our project confirms that variants in GJB2 and GJB6 genes do not contribute significantly to NSHI in the Cameroonian population. Also, variants in DMD that were shown to be associated with an increased hearing threshold in mice, do not seem to be implicated in HI in Cameroon, neither in previous human studies (although they did not objectively assess hearing using standardized testing methods). Despite the first symptoms of DMD occurring in infancy, the diagnosis is frequently made later in adolescence, indicating an underestimation of the number of cases of DMD in Cameroon. Future screening of deletions and duplications in patients from Cameroon should focus on the distal part of the DMD gene. Subsequently, this study successfully identified the candidate genes in two Cameroonian multiplex families with NSHI through the use of WES, and thus highlights the efficacy of next-generation sequencing techniques in resolving HI cases in Cameroonians and in cases where no pathogenic variants are found in common HI-genes. Additionally, our project which confirms that CLIC5 and DMXL2 genes are associated with HI in humans advocate for the inclusion of these two genes in diagnostic gene panels for NSHI in clinical settings. Last, this study shows the difficult social interaction and access to proper management faced by persons with HI in Cameroon, and highlights the need to educate populations on the causes of HI for a better acceptance of persons with HI in the Cameroonian society
Biomedical Research, A Tool to Address the Health Issues that Affect African Populations.
No full textTraditionally, biomedical research endeavors in low to middle resources countries have focused on communicable diseases. However, data collected over the past 20 years by the World Health Organization (WHO) show a significant increase in the number of people suffering from non-communicable diseases (e.g. heart disease, diabetes, cancer and pulmonary diseases). Within the coming years, WHO predicts significant decreases in communicable diseases while non-communicable diseases are expected to double in low and middle income countries in sub-Saharan Africa. The predicted increase in the non-communicable diseases population could be economically burdensome for the basic healthcare infrastructure of countries that lack resources to address this emerging disease burden. Biomedical research could stimulate development of healthcare and biomedical infrastructure. If this development is sustainable, it provides an opportunity to alleviate the burden of both communicable and non-communicable diseases through diagnosis, prevention and treatment. In this paper, we discuss how research using biomedical technology, especially genomics, has produced data that enhances the understanding and treatment of both communicable and non-communicable diseases in sub-Saharan Africa. We further discuss how scientific development can provide opportunities to pursue research areas responsive to the African populations. We limit our discussion to biomedical research in the areas of genomics due to its substantial impact on the scientific community in recent years however, we also recognize that targeted investments in other scientific disciplines could also foster further development in African countries
Genomics of sickle cell disease and fetal hemoglobin in African populations
No full textBackground More than 300,000 babies are born with sickle cell disease (SCD) each year. About 79% of these births occurs in sub-Saharan Africa where the sickle variant is known to have originated in the genetic background of the ancestors of Agriculturalist populations. Although the variant is highly lethal, the protection it confers against severe malaria in its heterozygous form has resulted in its persistence in sub-Saharan Africa where malaria is endemic. Without intervention, 50–90% of affected children in many sub-Saharan African countries die before their fifth birthday. The search for a definitive cure or effective disease-modifying therapy is therefore an imperative. Fetal hemoglobin (HbF) has long been recognized to ameliorate SCD severity whereby patients harboring natural genetic variations that lead to the persistence of high HbF levels in their blood tend to live longer with fewer complications. The HbF quantitative trait is highly heritable (89%), and is therefore the focus of enormous research for therapeutic purposes. Genetic polymorphisms influencing HbF level have been identified in three major loci; BCL11A, HBS1LMYB, and HBG2. However, these loci jointly explain less than 30% of HbF variability in African sickle cell anemia (SCA) patients as compared to ~50% in African Americans and non-anemic Europeans. Genome-wide association studies have been employed to replicate two of the three major loci (BCL11A and HBS1L-MYB) in African SCA patients, as well as to identify new loci including BACH2 involving a meta-analysis of African Americans and Tanzanians, SLC28A3, TICRR, and PIEZO2 in Nigerians, and FRMPD4 in Tanzanians. While BCL11A and HBS1L-MYB have been replicated in Cameroonian SCD patients through candidate genotyping studies, there has yet to be a genome-wide investigation. Moreover, the long survival of SCA patients in Africa despite a higher disease severity and mortality in the region hints at the enrichment of African genomes with ‘protective' polymorphisms potentially impacting the level of HbF, the strongest modifier of SCD severity. A better understanding of the genetic architecture of HbF in African SCA patients is therefore needed to foster research into potentially novel sickle cell disease therapeutic targets. Aims and Methods This thesis project aimed to: 1) present an in-depth description of the evolutionary history of the sickle cell mutation and its implication for global genetic medicine through a synthesis of publicly available data. This included updating and summarising the global georeferenced databases of the sickle gene and the HBB gene cluster haplotypes, systematically and critically evaluating the age and place of origin of the sickle cell mutation by incorporating information about malaria with which its evolution is highly intertwined, summarising bibliographic information on the genetic modifiers of SCD severity, and, importantly, examining other gene variants that co-occur with the sickle gene in sub-Saharan Africa that might hint at possible co-evolution and effect on SCD severity; 2) provide a comprehensive overview of SCD in Sub-Saharan Africa, bringing out transferable strategies and recommendations for prevention and care, through bibliographic searches; and 3) to investigate the missing heritability of HbF in SCA patients of African ancestry from Cameroon, Tanzanian, and the USA through a multiple imputation panel and genome-wide association approach, with fine-mapping and functional analysis. Briefly, the performance of different reference haplotype panels on genotype imputation accuracy for African SCA populations from Cameroon and Tanzania was assessed. Genome-wide association analyses for the two populations using all the imputation panels was performed, and then meta-analyses of the two populations, as well as with summary statistics from the USA-based cohorts. Statistical fine mapping and extensive in silico functional analyses were next performed to determine the functional relevance of significant associations, while extensive haplotype structure analysis was performed to illuminate the reason for substantial heterogeneity in association signals across different populations of the same ancestry. Finally, gene-based and gene set enrichment analyses were undertaken to identify additional significant loci and significantly enriched pathways, while heritability analysis was performed to further appreciate the observations of multiple significant signals and to better understand the genetic architecture of HbF in African SCA patients. Results Evolutionary history of sickle cell mutation In this first part, we successfully updated the global georeferenced databases of the sickle gene and the HBB gene cluster haplotypes. We showed that changes in population dynamics, contributed by migration and gene flow, might be introducing some HBB haplotypes in regions where they were previously absent, reflecting changes in regional SCD severity. Through our systematic and critical evaluation of the origin of the sickle cell mutation, we identified limitations to the models that have been used to estimate the age of the mutation, and we determined that the mutation is likely older than its currently held age of 22,000 years. Using data on the emergence of malaria, we determined that the mutation is most likely to have originated somewhere in West-Central Africa. Importantly, we showed overlapping distribution of the sickle gene and other gene variants that are under natural selection in sub-Saharan Africa. Data suggest that some of these gene variants impact SCD severity, while others have known modifying effects on the disease severity. Overview of sickle cell disease in sub-Saharan Africa We provided and overview of SCD in sub-Saharan Africa with transferable strategies for prevention and care as part of the Lancet Haematology's 2021 series on hematological care priorities in sub-Saharan Africa. We touched on aspects of SCD such as epidemiology, burden, mortality and life expectancy, hematological parameters, diagnosis, pathophysiology, biological and genetic modifiers of severity, management, environmental determinants, and psychosocial effects. We also presented challenges and proposed recommendations for SCD research in subSaharan Africa. Impact of reference haplotype panels on genotype imputation in African sickle cell anemia populations To use the genomics of HbF to search for other, perhaps more effective, targets of gene editing for better management and perhaps cure of SCD, we first assessed the impact of imputing missing genotypes into genome-wide single nucleotide polymorphism (SNP) data using different reference haplotype panels. We used six different panels including one which we created ourselves from whole genome sequencing data of fifty Cameroonians. The key observations included: i) different imputation performance for different African populations; ii) different imputation performance among different imputation panels within each African population, indicating that one variant can be imputed with vastly different accuracies across different panels, reflecting differences in haplotype structures across the panels resulting from different tagging schemes. This underscoresthe complementary use of the panels, with an expectation of different patterns of association (panel-specific significant signals). Genome-wide association analysis, statistical and functional fine mapping As expected from our assessment of imputation performance, we observed multiple panelspecific significant signals. We replicated the major known loci including BCL11A and HBS1L-MYB, and uncovered fourteen novel loci. The most significant of the novel loci, FLT1, which was observed in Cameroonians and replicated in a meta-analysis with Tanzanians, has known role in hematopoiesis. Fine-mapping and in silico functional analyses suggest an important role in HbF induction. Gene-based, gene set enrichment, and heritability analyses Gene-based analysis confirmed significant signals in the three major HbF-influencing loci, BCL11A, HBS1L-MYB, and HBG2. Gene set enrichment analysis revealed an overwhelming enrichment of hematopoiesis related pathways, as well as hemoglobin as the major enriched biological component, while blood traits were the most enriched phenotypes. Consistent with these, we estimated HbF heritability in a joint cohort of Cameroonian and Tanzanian SCA patients at 94% suggesting an enrichment of these populations with HbF-influencing loci in a way that has probably been underappreciated, and that might be better revealed through whole-genome sequencing. Conclusion and perspectives Our study presented data with overwhelmingly support for a single African origin of the sickle cell mutation, with opportunity for further research on determining the true age of the mutation. Tracking the movement of the mutation through the distribution of the HBB gene cluster haplotypes highlighted changing population dynamics that are important for public health. Calling attention to gene variants whose distribution in sub-Saharan Africa overlaps with the sickle gene is important because the co-occurrence could mean co-evolution which might suggest an impact on SCD severity. Therefore, further evolutionary genetic studies are warranted to understand the interactions of these gene variants and the sickle gene. We equally showed that progress in implementing newborn screening and comprehensive care in some sub-Saharan African countries has been encouraging. Early diagnosis and family education on management can reduce morbidity, while immunisation and hydroxyurea therapy initiated in affected children as young as 9 months old can greatly improve quality and quantity of life. Research in sub-Saharan Africa is urgently needed to establish the exact prevalence, mortality, and morbidity, environmental and genetic factors affecting clinical complications, and life expectancy of patients with sickle cell disease, to facilitate the design of future risk models and to investigate novel routes for therapeutic options with the ultimate aim of improving the clinical outcomes of patients with SCD in all parts of the world. Finally, our discovery of new genes that are associated with blood levels of HbF which is the strongest modifier of SCD severity and the target of gene editing means that our work has tremendous potential towards discovery of novel SCD therapeutic targets, but also in improving our understanding of the genetic architecture of HbF in understudied African populations
GJB2 and GJB6 mutations in hereditary recessive non-syndromic hearing impairment in Cameroon
No full textThis study aimed to investigate GJB2 (connexin 26) and GJB6 (connexin 30) mutations associated with familial non-syndromic childhood hearing impairment (HI) in Cameroon. We selected only families segregating HI, with at least two affected individuals and with strong evidence of non-environmental causes. DNA was extracted from peripheral blood, and the entire coding region of GJB2 was interrogated using Sanger sequencing. Multiplex PCR and Sanger sequencing were used to analyze the prevalence of the GJB6-D3S1830 deletion. A total of 93 patients, belonging to 41 families, were included in the analysis. Hearing impairment was sensorineural in 51 out of 54 (94.4%) patients. Pedigree analysis suggested autosomal recessive inheritance in 85.4% (35/41) of families. Hearing impairment was inherited in an autosomal dominant and mitochondrial mode in 12.2% (5/41) and 2.4% (1/41) of families, respectively. Most HI participants were non-syndromic (92.5%; 86/93). Four patients from two families presented with type 2 Waardenburg syndrome, and three cases of type 2 Usher syndrome were identified in one family. No GJB2 mutations were found in any of the 29 families with non-syndromic HI. Additionally, the GJB6-D3S1830 deletion was not identified in any of the HI patients. This study confirms that mutations in the GJB2 gene and the del(GJB6-D13S1830) mutation do not contribute to familial HI in Cameroon.Edmond Tingang Wonkam, Emile Chimusa, Jean Jacques Noubiap,
Samuel Mawuli Adadey, Jean Valentin F. Fokouo and Ambroise Wonka
Recommended from our members
Sex-based differences in the manifestations and complications of sickle cell disease: Report from the Sickle Cell Disease Implementation Consortium
No full textIntroductionSex-based clinical outcome differences in sickle cell disease (SCD) remain largely unknown despite evidence that female sex is associated with an increased lifespan. To better characterize sex-based differences in SCD, we assessed pain, treatment characteristics, laboratory measures and complications among males and females currently enrolled in the Sickle Cell Disease Implementation Consortium (SCDIC) registry.MethodsThe SCDIC consists of eight comprehensive SCD centers and one data coordinating center that received funding from the National Heart Lung and Blood Institute to improve outcomes for individuals with SCD. Eligibility criteria included: 15 to 45 years of age and a confirmed diagnosis of SCD. Self-report surveys were completed and data were also abstracted from the participants' medical records.ResultsA total of 2,124 participants were included (mean age: 27.8 years; 56% female). The majority had hemoglobin SS SCD genotype. Females had worse reports of pain severity (mean (SD) T-score 51.6 (9.6) vs 49.3 (10), p<0.001), more vaso-occlusive episodes (p = 0.01) and a higher occurrence of 3 or more hospital admissions in the past year (30.9% vs. 25.5, p = 0.03). On multivariable analysis, males had higher odds of acute chest syndrome (odds ratio (OR) 1.4, p = 0.002), cardiovascular (OR 1.70, p<0.001) and musculoskeletal (OR 1.33, p = 0.0034) complications and lower odds of depression (OR 0.77, p = 0.0381). Females had higher fetal hemoglobin levels with and without hydroxyurea use (9.6% vs 8.5%, p = 0.03 and 3% vs 2.2%, p = 0.0005, respectively).ConclusionOur data suggests that sex differences in clinical outcomes do occur among individuals with SCD. Future research needs to explore the mechanisms underlying these differences
The genetics of non-syndromic hearing impairment in South Africa
No full textHearing impairment (HI) is a sensory disorder resulting in the partial or complete disability to perceive sound in the better-hearing ear. It is defined as the inability to hear better than 25dB in the better-hearing ear. Subsequently, it is considered disabling when a child cannot perceive sound better the 30dB, in the better hearing ear, and an adult cannot perceive sound better than 40dB, in the better hearing ear. Hearing impairment may result from genetic, environmental, or unknown factors. The connexin gene, GJB2, is the prevalent gene resulting in congenital HI in most children with European, North American, and East Asian ancestry. Apart from the founder mutations present in GJB2 in Morocco, Ghana and Senegal, the prevalent causative genes resulting in congenital HI in African populations are yet to be fully elucidated. Congenital Hearing impairment in South Africa (RSA), has been estimated to have an incidence rate of 5.5 per 1000 live births, which is 5 times higher than the birth incidence in high-income countries (approximately 1 to 2 per 1000 live births). Patients are generally diagnosed late with HI, at approximately 3 years old, and the most prevalent environmental factors associated with HI in RSA are middle ear infections, with several reports implicating ototoxicity as a cause of HI. Variants in connexin genes i.e. GJB2 associated with HI have been shown to be irrelevant in the Black South African populations, and the limited genetic studies have identified private mutations in selected families. However, the full extent of prevalent genes associated with HI in the South African populations is still to be investigated. Methods and results Through a systematic review, we investigated the state of HI research in South Africa was established. Though studies have been performed since the 1960s, the results showed that genetics of HI in South Africa was not well explored. Universal new-born hearing screening is ideal in detecting congenital HI, but it is currently not standard practice in the country. However, with the advent of modern technology, HI screening may be more accessible to patients through community health workers. Patients who fail the repeated screenings may then be referred for further audiometry testing. This may positively impact the identification of patients with HI and assist with the necessary interventions. We also collaboratively worked to establish the first disease ontology for HI to further allow standardised and harmonised language surrounding HI. This provides the scalability and interoperability of research going forward. It will allow for all stakeholders in HI research to use the same terminology when discussing HI. In order to address the dearth of genetics research regarding non-syndromic HI, patients presenting with putative genetic HI were recruited from schools of the deaf across South Africa and two hospitals in Cape Town. The patients were recruited along with their family members, both with and without HI, and their DNA was extracted from whole blood. Twenty-seven families segregating non-syndromic HI, consisting of 65 affected and 35 unaffected individuals were subjected to whole exome sequencing (WES). The HI was resolved in 20 families (74%), and pathogenic variants were identified in the genes: WFS1 (c.A2141), MITF (cT918A), ADGRV1(c.G564T, c.A17450G, c.A11298C), PDSS1(c.C641T, NEU1(c.C1069T, c.G754C), c.G727A), TBC1D24(c.G1514A), MYO15A(c.C1378T, TMPRSS3(c.205+6t>A), c.9303+5G>A, c.G6634A), USH2A(c.T9437A, c.G2990T, c.G101A), STRC(c.G225A, c.C4057T, c.G4655C, c.C4351T, c.G4403A), P2RX2(c.G1064A, c.C1187G), OTOG(c.C2525A, c.G3143A, c.G916A), LHFPL5(c.621delC), TRIOBP(c.C3133T, c.C4298T), SLC26A4(c.T94C, c.T716A), GJB2(c.35delG), REST(c.G1244C), CRYM(c.*6_*2delACAAA), CDH23(c.T1585C, c.G8230A), FGFR2(c.1297+10G>C), MYO7A(c.6255delC). The pathogenic variants presented 8 autosomal dominant alleles and 12 autosomal recessive alleles Five families presented with pathogenic or likely pathogenic variations associated with Usher Syndrome and the remaining 14 families presented with pathogenic variations associated with non-syndromic HI. One family presented with putative pathogenic variations in NEU1, which is a gene associated with Sialidosis. We specifically investigated, in greater detail, a dominant novel variation in REST, present in one family, which encodes a transcription factor, that was identified using whole exome sequencing. This gene was previously suspected to be associated with hearing impairment only once, in an American family. The variation was absent in the unaffected South African family members, unrelated patients, and unaffected controls. In vitro cell-based studies indicated that the variation results in the loss of nuclear exclusivity of REST. Luciferase assays indicated that the mutant was unable to repress the expression of one of its target genes, whereas the wild-type effectively inhibited the expression of the target. Conclusions This thesis successfully performed the following investigations: 1) development of the first Hearing Impairment Ontology worldwide, 2) review the genetic profile of HI in South Africa, 3) used WES to find known and novel variants in established HI genes, and 4) confirmed REST as a novel HI gene. Future work will focus on sequencing all the remaining samples and identifying their putative causative mutations. Further work includes feedbacking the results of the genetic testing to the patients and their families. The data will contribute to improving the HI-genes pairs' curation in Africa, and globally
Genetics of hearing impairment and peripheral neuropathy in Mali
No full textBackground Hearing impairment (HI), the most common sensory disturbance, affects about 1 in 1000 living newborns globally. Its incidence is reported higher in sub-Saharan African (SSA) populations. HI is caused by environmental and genetics factors. In many developing countries, environmental factors are reported to be the most prevalent aetiologies while genetic causes are predominant in the developed countries. Over 50% of congenital HI has a genetic origin with more than 120 genes identified to date. Despite this large number of known genes, only GJB2 (OMIM: 121011) and GJB6 (OMIM: 604418) are systematically studied in SSA populations for which the prevalence of HI-causal variants is insignificant. Charcot-Marie-Tooth disease (CMT), is the most common inherited peripheral neuropathy (IPN) with a high clinical and genetic heterogeneity and over 100 genes are related to CMT, mostly in populations of Caucasian ancestry. Yet, despite being described more than 130 years ago, there remains a paucity of information on its global prevalence and genetic epidemiology due largely to challenges in diagnosis, especially in countries with limited resources. Over 90% of CMT are caused by mutations in PMP22 (OMIM: 601097), GJB1 (OMIM: 304040), MFN2 (OMIM: 608507), MPZ (OMIM: 159440) genes. HI is the common audiological symptom associated with CMT and is caused by several genes including PMP22 and GJB1. HI and IPN are inherited in autosomal (dominant and recessive), X-linked, and mitochondrial transmission. However, the genetic epidemiology of these diseases are largely unknown in Africa, and have not been investigated in Mali where consanguineous marriage is a common practice that may increase recessive conditions
Risk factors associated with blood pressure variation in sickle cell disease in Cameroon
No full textBackground: In SCD patients, studies have shown that increased Blood Pressure (BP) is associated with higher risk of stroke and mortality, even in a range of systolic BP(SBP) and diastolic BP(DBP) that are considered relatively normal for the general population (i.e., lower than 140 mmHg). SCD patients, generally, have lower systolic, diastolic, and mean BP compared to age and sex-matched controls. Relative Systemic Hypertension (RSH) for SCD patients is defined as BP ranging from 120–139/70–89 mmHg, and Systemic Hypertension for SCD patients with BP above 140/90 mmHg. BP may be a potential modulator of clinical severity in SCD patients. BP is a heritable trait with estimates indicating that 30–70% of the trait variance is attributable to genetic variation. A recurrent deleterious and loss of function mutations in genes associated with lowering BP, e.g., variants in CLCN6, have been recently associate with long survival in SCD in Cameroon. Additionally, demographic, biological, and anthropometric factors have been reported to be associated with BP in SCD patients. Understanding the aetiology of BP variation in SCD patients and assessment of up-to-date evidence is key to early prediction of potential BP-related complications in SCD patients. Early prediction of BP-related complications in SCD patients could lead to better prevention and treatment of SCD and its associated causes of mortality. Herein, we investigated (clinical, genetic, and epidemiological) risk factors associated with RSH or Systemic Hypertension in SCD patients in Cameroon to gain insight into the pathophysiology of BP variation in this disease in an African setting. Objectives: 1) A systematic review and meta-analysis on BP variation (Normal, Relative Systemic Hypertension(RSH) and Hypertension) among SCD patients; 2) Investigate clinical and epidemiological risk factors for RSH and Systemic Hypertension in 815 SCD patients in Cameroon; 3) Identification of genetic variants associated to BP variation in SCD patients in Cameroon using a Genome-wide Association study; 4) Functional analysis on pathways and mechanisms underlying BP variation in SCD patients. Methods: Systematic review and meta-analysis: A protocol was developed and registered with the International Prospective Register of Systematic Reviews (PROSPERO), registration number CRD42020168798. A search was conducted on the following electronic databases: PubMed, Scopus, and Web of Science. The Preferred Reporting Items for Systematic Reviews and Meta- analysis (PRISMA) served as a template for reporting the review and meta-analysis. Independently, in collaboration with another reviewer, we screened studies, extracted relevant data, assessed methodological quality and risk of bias on each included study. A retrospective analysis to investigate risk factors associated with RSH or Systemic Hypertension in SCD patients in Cameroon. Participants who had incomplete/out of range BP readings were excluded from this analysis. Categorical variables were compared using Chi squared test or Fisher exact test if the expected count in a cell was less than five while continuous variables were compared according to BP category with the Kruskal–wallis test. Multivariate multinomial logistic regression modelling was used to examine the effects of the demographic, anthropometric, clinical, and laboratory factors to determine the potential independent risk factors for RSH and Systemic Hypertension. A final model was created that included all the predictors and interactions that were significantly associated at the level of P 18 years (P < 0.001), weight (P < 0.001), height (P < 0.001), BMI (P < 0.001), pulse pressure (P = 0.046), creatinine (P < 0.001), eGFR (P < 0.001) and haemoglobin (P = 0.020) . Multivariate analyses found that age [OR = 1.02, (95% CI = 1.01–1.05), P = 0.021], creatinine [OR = 1.310, 95% CI = 1.05–1.63, P = 0.016], BMI [OR = 1.09, (95% CI = 1.03– 1.16), P = 0.002] were independent risk factors for high BP values in SCD patients with RSH compared with SCD patients with normal BP values. Genome-wide analysis and meta-analysis: For SBP, we found two genetic loci including independent and suggestive SNPs. The first locus was found in RP11-727A23.5 on chromosome 11 (rs146072506, P=4.88x10-08), the second locus was found on DLGAP1 gene on chromosome 18 (rs35715722, P=3.408x10-08). Two other SNPs were found in the intergenic regions. For DBP, the only significant SNP was in the intergenic region. Additionally, we found one SNP (rs186536474, p=8.90x10-09) located on GPR39 gene on chromosome 2 with a statistically significant p-value associated with BMI. Identified variants in intergenic regions may play a role in specific genes regulating BP or BMI. The meta-analysis of the Cameroon SCD cohort and Silent Infarct Transfusion Trial (SITT) cohorts, and African American cohort of SCD patients, did not find significant SNPs. After performing genetic analysis and Gene-set analysis with none of the identified genes or gene-set reached a significant threshold. Additionally, tissue expression analysis did not reach significant thresholds. Conclusion: This is the first in-depth investigation of the non-genetic and genetic risk factors associated with BP variation (RSH or Systemic Hypertension) in SCD patients in Africa, i.e., Cameroon. Age, male gender, BMI was found to be statistically significant independently associated factors of RSH and Hypertension in the SCD patients in Cameroon. The GWAS analysis identified a few SNPs which were statistically associated with BP and BMI in Cameroonian SCD patients. These SNPs require follow up studies in larger SCD cohorts. Tailored interventions that consider both genetic and non-genetic risk factors have potential to lead to better management of BP pressure in SCD patients and prevent possibly prevent development of severe SCD complications
Pharmacogenomics of sickle cell disease therapeutics: pain and drug metabolism associated gene variants and hydroxyurea-induced post-transcriptional expression of miRNAs
No full textSickle cell disease (SCD) is a common blood disease caused by a single nucleotide substitution (c.20T>A, p.Glu6Val) in the beta globin gene on chromosome 11. The prevalence of the disease is high throughout large areas in sub-Saharan Africa, the Mediterranean basin, the Middle East, and India due to the level of protection that the sickle cell trait, provides against severe malaria. Approximately 300,000 infants are born per year with sickle cell anemia, which is defined as homozygosity for the sickle hemoglobin (HbS). The majority (nearly 75%) of these births occur in sub-Saharan Africa, particularly in two countries: Nigeria, and the Democratic Republic of the Congo where there are poorly resourced healthcare systems. Early diagnosis, penicillin prophylaxis, blood transfusions, hydroxyurea, and hematopoietic stem-cell transplantation can dramatically improve survival and quality of life for patients with SCD. However, our understanding of the role of genetic and clinical factors in explaining the complex phenotypic diversity of this disease is still limited. Early prediction of the severity, and patients' responses to specific therapeutics of SCD could lead to more precise treatment and management. Beyond well-known modifiers of disease severity, such as fetal hemoglobin (HbF) levels and αthalassemia, other genetic variants might influence specific sub-phenotypes. New treatments and management strategies accounting for these genetic and nongenetic factors could substantially and rapidly improve the quality of life and reduce health care costs for patients with SCD. Patients with SCD are subjected to long term administration of drugs and there is a limited data on pharmacogenomics of SCD therapeutics. Vaso-occlusive crisis (VOC) are the main clinical events of SCD and are associated with recurrent and long-term use of antalgics/opioids and HU. This project aimed to investigate the clinical and genetic predictors of painful vaso-occlusive crisis (VOC) among SCD Cameroon patients by exploring pharmacokinetic determinants of treatment responses as well as post-transcriptional signatures triggered by hydroxyurea treatment, particularly, miRNA expression. SCD patients were recruited from Yaounde Central Hospital and Laquintinie Hospital in Douala (Wonkam et al., 2018, Mnika et al., 2019 (b)), and recent migrants SCD patients from the DRC, recruited at the Haematology Clinic, Groote Schuur Hospital in Cape Town, South Africa (Mnika et al., 2019 (a) and Mnika et al., 2019 (b)). Sociodemographic and clinical data were collected by means of a structured questionnaire. Patients' medical records were reviewed to extract their clinical features over the past 3 years. Specifically, the occurrences of VOC, hematological parameters, hospital outpatient visits, hospitalisation, overt strokes, blood transfusions, and administration of hydroxyurea were recorded. Height, weight, body mass index (BMI), systolic and diastolic blood pressures (SBP and DBP) were measured. Detailed descriptions of patients and sampling methods used in the Cameroonian patients have been reported previously (Wonkam et al., 2018 Mnika et al., 2019 (a) and Mnika et al., 2019 (b)). For the purpose of comparing frequencies of variants, ethnically matched Cameroonian controls were randomly recruited from apparently healthy blood donors in Yaounde for participation in the study. All blood samples were collected for genomic characterisation and analysis. DNA was extracted from peripheral blood, following instructions on the available commercial kit [QIAamp DNA Blood Maxi Kit ® (Qiagen, United States)]. Genotyping (TaqMan and MassArray) was performed for 40 variants in 17 pain-related genes, three fetal haemoglobin (HbF)-promoting loci, two kidney dysfunction-related genes, and HBA1/HBA2 genes for 436 patients. A subset of these samples was also genotyped to analyse 32 core and 267 extended pharmacogenes using commercially available PharmacoScan® platform for characterisation of pharmacokinetic determinant of response. We also compared the pharmacogenes variants from these African groups, to data extracted from the 1000 genomes Project. Moreover, association studies were carried out on pharmacogenes variants with SCD clinical variability. Additionally, protein-protein interaction (PPI) network and enriched biological processes and pathways were investigated. For association studies, statistical models using regression frameworks to analyse 40 variants were performed in R®. For miRNA expression, total RNA was isolated using the miRNeasy kit according to protocol of the Manufacturer (QIAGEN, Hilden, Germany); and sequenced by the Genomic and RNA Profiling Core at Baylor College of Medicine, United States, using the NanoString Platform (NanoString Technologies, Inc., Seattle, WA, United States), according to manufacturer's instructions. Genes with statistically significant changes in expression were analysed using the significance analyses of microarrays (SAM) tools. Female sex, body mass index, Hb/HbF, blood transfusions, leucocytosis and consultation or hospitalisation rates significantly correlated with VOC. Three painrelated gene variants correlated with VOC (CACNA2D3-rs6777055, P = 0·025; DRD2- rs4274224, P = 0·037; KCNS1-rs734784, P= 0·01). Five pain-related gene variants correlated with hospitalization/consultation rates (COMT-rs6269, P = 0·027; FAAHrs4141964, P = 0·003; OPRM1- rs1799971, P = 0·031; ADRB2-rs1042713; P < 0·001; UGT2B7-rs7438135, P = 0·037). The 3·7 kb HBA1/HBA2 deletion correlated with increased VOC (P = 0·002). HbF-promoting loci variants correlated with decreased hospitalisation (BCL11A-rs4671393, P = 0·026; HBS1L-MYB-rs28384513, P = 0·01). APOL1 G1/G2 correlated with increased hospitalisation (P = 0·048). A commercial genotyping array platform (PharmacoScan®) with 4627 markers located in 1191 genes was used to investigate 299 pharmacogenes (32 ADME core and 267 extended pharmacogenes). Based on the PharmacoScan analyses, no statistically significant differences in allele frequencies were detected between SCD cases and controls from Cameroon. A principal component analysis (PCA) revealed that Cameroonians' data clustered with other Africans, but this population is significantly distinct from American, European and Asian populations data. Variant allele frequencies in 21/32 core pharmacogenes were significantly different between the two SCD groups (Cameroon vs. Congo). No correlation between clinical variability and variants in the core genes was detected for both populations under study. An association study of the core and extended PharmacoScan variants to VOC identified statistically significant associations between two single nucleotide polymorphisms (SNPs) to VOC after correction of multiple testing. These two SNPs mapped to 50 genes, with two SNPs located in core pharmacogenes (SLCO4A1- rs118042746, p=1.21e-07; UGT1A10, UGT1A8- rs10176426, p=1.22e-07). Functional enrichment analyses revealed that these 50 genes are involved in three biological processes and four pathways relevant to SCD pathophysiology, including xenobiotic glucuronidation (GO:0052697, p = 2.3e-03), and drug metabolism - other enzymes (p = 2.1e-02). Further analyses of the 50 genes, identified key genes in human proteinprotein networks: NTSR1, LRMDA, SMAD SMAD4 and CDH2. These four genes also interacted with three core pharmacogenes associated with VOC: UGT1A8, UGT1A10 and SLCO4A1. We found 22/798 miRNAs to be differentially expressed under HU treatment, with the majority (13/22) being functionally associated with HbF-regulatory genes, including BCL11A (miR-148b-3p, miR-32-5p, miR-340-5p, miR-29c-3p), MYB (miR-105-5p), KLF-3 (miR-106b-5), and SP1 (miR-29b-3p, miR-625-5p, miR-324-5p, miR-125a-5p, miR-99b-5p, miR-374b-5p, miR-145-5p). The present thesis started by highlighting the scarcity of studies investigating variable responses to pain in SCD patients and then proceeded to addressing this research gap. To our knowledge this is the first body of from Africa to provide evidence supporting the possible development of a genetic risk model for pain in SCD. This is also the first body of work to report an association between these two SNPs and VOC in core and extended pharmacogenes. Our data reveals that the commercial pharmacogenes arrays investigated might need additional evidence for appropriateness among Africans. Therefore, it advocates the need to invest in research exploring population-specific arrays, drug design, targeting, and efficacy, for improved clinical management of patients of African descent. Previous studies have investigated various mechanisms to understand the genomic variations affecting responses to HU, but full understanding of the variable HU-mediated HbF production among individuals affected by SCD remains elusive. The present study showed that mechanisms of HbF production in response to HU, could particularly be mediated through miRNA regulation. The data reveals some alternative perspectives and routes towards identifying new therapeutic targets and approaches for SCD. However, this study needs to be replicated in larger samples in multiple African populations
- …
