Genetic factors associated with the cerebrovascular complications of Sickle Cell Disease

Student thesis: Doctoral ThesisDoctor of Philosophy


Patients with sickle cell disease (SCD) carry a significantly increased risk of cerebrovascular disease (CVD) throughout their life. This risk includes overt ischaemic stroke (OIS), which is typically preceded by a large vessel vasculopathy (LVD), and silent cerebral infarcts (SCI), which affect a patient’s IQ and confer an increased risk of OIS. Clinical studies based on sibling analysis suggest a strong underlying genetic susceptibility and, while co-inheritance of alpha thalassaemia is recognised to offer protection against OIS and LVD, genetic factors beyond this are still not clearly delineated.
I used two approaches in an attempt to identify novel genetic variants associated with CVD in SCD. First, I used linear mixed modelling (LMM) in a large cohort of patients (n=516) with well-defined cerebrovascular outcomes and genomic data based on SNParray genotyping and imputation, to perform a genome wide analysis (GWAS) on three cerebrovascular outcomes: OIS, LVD, and SCI. I used this analysis to also evaluate cerebrovascular-trait association of genetic variants implicated in previously published studies. Second, I used whole exome sequencing (WES) in a smaller (n=19), but more severely affected cohort of patients who had an OIS occurring before their 4th birthday, looking to identify rarer but stronger effect variants that may be in common. I used variant filtering pipelines to guide variant discovery first under a mono/oligogenic variant hypothesis, and then under a polygenic variant hypothesis. I then used the large cohort established in the first project for validation testing of the candidate variants identified. Finally, I used a large cohort of patients with sickle cell disease to investigate the various effects of alpha thalassaemia on the cerebrovascular, clinical and laboratory measurements of patients with sickle cell disease.
This work yielded a number of important and novel findings. Through the analysis of patients with a severe CVD phenotype using WES, I found the apolipoprotein E (APOE) haplotypes ξ2/ξ4 or ξ4/ξ4 (resulting from variants rs7412.C>T and rs429358.T>C) confer a 4.35-fold increased risk of overt stroke occurring in childhood (95% CI 1.85-10.0, p=0.0011). Additionally, rs2297518.G>A in Nitric Oxide Synthase 2 (NOS2) and rs2230123.T>C in Signal Transduction and Activator of Transcription 5a (STAT5a) also increase the risk of OIS in childhood (OR=2.25, 95%CI 1.21-4.19, p=0.0139, and OR=2.60, 95%CI 1.30-5.20, p=0.009 respectively). Through genome wide association analysis in the larger patient cohort, two variants were identified; rs73598466.G>A lying intronically within C-maf inducing protein (CMIP), OROIS=3.81, p=2.45x10-9, and rs13338739.T>G, lying within an intron of Cadherin 13 (CDH13), ORLVD=2.17, p=8.72x10-08, although these two have not yet been validated in a separate cohort. I also replicated the association of three variants, in two genes, previously implicated with OIS in SCD: rs267201.A>G, (OR=1.26, p=0.039) and rs449853.C>T (OR=0.4, p=0.005) in Bone morphogenetic protein 6 (BMP6) and rs662.T>C, (OR=2.70, p=0.007) in Paroxonase 1 (PON1), but I could not reproduce any other previously published associations with CVD in SCD. Additionally, I confirmed that alpha thalassemia is strongly protective against both OIS and LVD (OR=0.56, 95%CI 0.36-0.88, p=0.00743, and OR=0.52, 95%CI 0.36-0.75, p=0.00028 respectively), but also that it is protective against SCI (OR=0.65, 0.46-0.92, p=0.0128).
I suggest that an emerging theme from these genetic variants and their related pathways is the implication that aberrant lipid metabolism as an important exacerbating mechanism of both neuroinflammation and vascular endothelial inflammation. The variants identified by this research would therefore represent genetic risk factors that are potentially modifiable by both dietary and pharmaceutical manipulation of their dyslipidaemic effects.
Date of Award1 Oct 2021
Original languageEnglish
Awarding Institution
  • King's College London
SupervisorDavid Rees (Supervisor) & Stephan Menzel (Supervisor)

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