Haematological Pathology

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Browsing Haematological Pathology by browse.metadata.advisor "Swanepoel, Carmen"

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    Imatinib resistance : the role of pharmacogenetic variability in a South African chronic myeloid leukemia cohort
    (Stellenbosch : Stellenbosch University, 2023-03) De Long, Chantal; Swanepoel, Carmen; Stellenbosch University. Faculty of Medicine and Health Science. Dept. of Pathology. Haematological Pathology.
    ENGLISH SUMMARY: Drug-resistant cancers are often associated with poor patient outcomes and the underlying mechanism is poorly understood. Chronic Myeloid Leukemia (CML) serves as a disease model for studying cancer drug resistance, specifically to Imatinib, a tyrosine kinase inhibitor. Approximately 20-30% of patients become resistant to Imatinib. Variability in patient drug response could be due to single nucleotide variants (SNVs) in genes that encode for Imatinib-metabolizing enzymes and transporters. The overall aim of the present study is to determine whether selected SNVs located within genes CYP3A4/3A5, SLCO1A2, SLC22A4, and SLC22A1 that encode selected drug transporters (Cytochrome P450, OATP1A2, OCTN1, hOCT1), respectively, contribute to an alternative mechanism leading to Imatinib resistance in a South African cohort. A maximum of 45 samples from Imatinib-resistant CML patients were analysed along with 44 non-resistant CML patients (controls). The selected SNVs were analysed using PCR-based genotyping assays. Baseline allelic and genotypic frequencies within our CML cohort was determined and compared between Imatinib good responders and Imatinib resistant groups. In our study we observed that there were differences in allele frequencies for the following SNVs in genes SLC22A4, SLCO1A2, CYP3A4 and CYP3A5 when compared to the global/African frequencies. Furthermore, obtained results showed that the observed and expected genotype frequencies were comparable for genes SLC22A1, SLC22A4, and SLCO1A2, however, our observed genotype frequencies were different from the expected genotype frequencies for the following genes SLCO1A2, CYP3A4 and CYP3A5. Interesting findings include, the rs35191146 (ATG>AT: delG) that was linked poor Imatinib treatment outcome, however, the simultaneous presence of rs628031 (A>G: M408V) circumvented this effect. All patients within our cohort have both Met420 and Met408Val. Another interesting finding is the coincidental finding of variants SLC22A4 rs11568500 (c.616_617delinsCC), and SLC22A1 rs35191146 (c.1258_1260delATG and g.160139876_160139883delGTAAGTTG), which would be explored in future studies. Even though the selected SNVs do not affect Imatinib resistance in our cohort, our study to the best of our knowledge is the only study to determine baseline allelic and genotypic frequencies for CML patients treated with Imatinib in South Africa. Therefore, the data obtained from our study can serve as a useful tool to further investigate the pharmacogenetic variability in South Africa. In conclusion, our study adds to the body of knowledge out there related to SNV and its potential clinical relevance related to imatinib resistance especially within our diverse African cohort. This in turn highlights the need for future studies focusing on larger cohorts, with a larger selection of SNVs at more health care institutions across South Africa.
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    Screening and characterisation of BCR::ABL1 kinase domain mutations in chronic myeloid leukaemia participants at Tygerberg Hospital, South Africa
    (Stellenbosch : Stellenbosch University, 2023-03) Shareefa, Isaacs; Swanepoel, Carmen; Chapanduka, Zivanai; Stellenbosch University. Faculty of Medicine and Health Science. Dept. of Pathology. Haematological Pathology.
    ENGLISH SUMMARY: To date, an increased number of drug resistant cancers have been observed. The underlying mechanisms are not well understood; therefore, we use Chronic Myeloid Leukaemia (CML) as a disease model to explore the type and frequencies of potential drug resistant causing mutations and the effect on survival at Tygerberg Hospital (TBH). The underlying genetic abnormality is a t(9::22) chromosomal translocation, and consequent fusion between the Breakpoint Cluster Region and the Non-Receptor Tyrosine Kinase Abelson genes (BCR::ABL1). Although Tyrosine Kinase Inhibitors (TKD), such as Imatinib (Gleevec), effectively decrease BCR::ABL1 mRNA transcript levels whereby the drug acts on the protein and inhibits its mode of action, point mutations within the kinase domain (KD) have shown to confer resistance to treatments. A retrospective audit was conducted on a CML cohort from 2013 to 2020, and 20 of these participants were recruited for detection of KD mutations within the ABL1 oncogene. Peripheral blood from routine diagnostic screening by the NHLS Molecular Haematology diagnostic laboratory were obtained for the mutation detection assay via bi-directional Sanger Sequencing. Identified sequence variants were then subjected to various bioinformatics tools to investigate variant, protein, and consequent effect on pathways. For the audit, a total of 165 patients with confirmed CML treated at TBH was captured. Descriptive statistics showed 46.1% of CML patients were female, and 53.9% were male. The patients were from 69 different areas in the Northern part of the Cape Metropole and surrounding Boland areas in Western Cape, South Africa. Quantitative analysis showed the youngest patient to be two years old and the oldest 88 years old, x̄ = 45-46 years. We found a weak linear correlation between age and BCR::ABL1 mRNA transcript levels. A significant difference in BCR::ABL1 mRNA transcript levels (p < 0.000) was seen in CML patients on treatment after 1 year. Of interest’s sake, 104 of the initial 165 patients who were diagnosed were still undergoing treatment. Moreover, 57 out of the 165 patients presented with possible resistance; 75% were categorized as failure and 25%at warning stage according to European Leukaemia Net guidelines. In the second part of the study, 20 participants’ samples were screened for variants. The screening sample group had a mortality rate of 20% who had a relative survival rate of less than five years. Sanger sequence analysis showed potential variants of interest in Exon 4 and of the ABL1 gene, although bioinformatics analysis alludes to the fact that these variants are not likely to play a role in resistance. In conclusion, the TBH CML cohort has a younger age at diagnosis which puts a greater strain on the public sector, and a significant number of patients are lost to follow up. More so, the age relative to CML deaths, as well as relative survival greatly differs from literature. Additionally, although synonymous mutations have recently shown to be pathological, the two variants found within our cohort has not been published to do so. CML could be a poster child for personalised medicine as the survival rate of CML patients is nearing that of the general population and treatment-free remission is attainable. Unfortunately, the paucity of CML data, drug availability and monitoring limitations are some of the obstacles that hinders South Africa from achieving these goals.