Browsing by Author "Smith, Muneerah"

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    Open Access
    Age, Disease Severity and Ethnicity Influence Humoral Responses in a Multi-Ethnic COVID-19 Cohort
    (2021-04-28) Smith, Muneerah; Abdesselem, Houari B; Mullins, Michelle; Tan, Ti-Myen; Nel, Andrew J. M.; Al-Nesf, Maryam A Y; Bensmail, Ilham; Majbour, Nour K; Vaikath, Nishant N; Naik, Adviti; Ouararhni, Khalid; Mohamed-Ali, Vidya; Al-Maadheed, Mohammed; Schell, Darien T.; Baros-Steyl, Seanantha S; Anuar, Nur D; Ismail, Nur H; Morris, Priscilla E; Mamat, Raja N R; Rosli, Nurul S M; Anwar, Arif; Ellan, Kavithambigai; Zain, Rozainanee M; Burgers, Wendy A; Mayne, Elizabeth S; El-Agnaf, Omar M A; Blackburn, Jonathan M
    The COVID-19 pandemic has affected all individuals across the globe in some way. Despite large numbers of reported seroprevalence studies, there remains a limited understanding of how the magnitude and epitope utilization of the humoral immune response to SARS-CoV-2 viral anti-gens varies within populations following natural infection. Here, we designed a quantitative, multi-epitope protein microarray comprising various nucleocapsid protein structural motifs, including two structural domains and three intrinsically disordered regions. Quantitative data from the microarray provided complete differentiation between cases and pre-pandemic controls (100% sensitivity and specificity) in a case-control cohort (n = 100). We then assessed the influence of disease severity, age, and ethnicity on the strength and breadth of the humoral response in a multi-ethnic cohort (n = 138). As expected, patients with severe disease showed significantly higher antibody titers and interestingly also had significantly broader epitope coverage. A significant increase in antibody titer and epitope coverage was observed with increasing age, in both mild and severe disease, which is promising for vaccine efficacy in older individuals. Additionally, we observed significant differences in the breadth and strength of the humoral immune response in relation to ethnicity, which may reflect differences in genetic and lifestyle factors. Furthermore, our data enabled localization of the immuno-dominant epitope to the C-terminal structural domain of the viral nucleocapsid protein in two independent cohorts. Overall, we have designed, validated, and tested an advanced serological assay that enables accurate quantitation of the humoral response post natural infection and that has revealed unexpected differences in the magnitude and epitope utilization within a population.
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    An immunoproteomic approach to identifying cancer-associated autoantibody biomarkers
    (2018) Smith, Muneerah; Blackburn, Jonathan
    Cancer is a heterogenous disease capable of forming and spreading in most tissues of the human body. Cancer screening and diagnosis can be performed through medical procedures, which are highly invasive, requiring an intensive infrastructure. It is therefore important to create cost-effective, non-invasive cancer diagnostic tools that also gives an indication of disease prognosis. With this in mind, the Blackburn lab previously created a cancer-testis antigen microarray (CT100plus) functionalised with tumour-associated and tumour-specific antigens, capable of detecting plasma- or serum-derived autoantibodies in the picogram per millilitre (pg/ml) range. In this thesis, a newly established statistical pipeline was used to analyse colorectal cancer (CRC) patient-derived CT100plus data. Using the pipeline, the 10 antigens with the highest receiver operator characteristic (ROC)-derived area under the ROC curve (AUC)-values were identified as potential autoantibody-based biomarkers. The top 10 antigen biomarker candidates include CEACAM 1, COL6A1, GRWD1, MAGEA1, MAGEA5, MAGEA10, NY-CO-1, SGY-1, SPANXB1 and THEG. Using these biomarker candidates, distinct clusters of healthy controls (HCs) and CRC patients were observed using both unsupervised hierarchical clustering and principle component analysis (PCA) analysis. Combinatorial ROC analysis indicates that CEACAM1 and GRWD1 as the top autoantigen combination for CRC diagnosis, together producing sensitivity-, and specificity-, and AUC-values of 1.00, 0.77 and 0.94, respectively. Furthermore, other top autoantigens, including COL6A1, THEG and CEACAM7, a homologue of CEACAM1, were also identified in this thesis by affinity purification-mass spectrometry (AP-MS) for patients from the same cohort, providing supporting evidence that these antigens are associated with CRC. The CT100plus microarray content was enzymatically modified to include citrullinated proteins, with the subsequent assessment of CRC patient autoantibody response. Significantly (p-value ≤ 0.05; adjusted p-value ≤ 0.05) higher signal intensities were detected in CRC patients versus HCs for citrullinated CDK7, MAGEB1, MAGEB5, MAGEB6 and SYCP1, whereas no significant (adjusted p-value > 0.05) difference in autoantibody signal was detected for these autoantigens on the noncitrullinated microarray for the same patient cohort. ROC analyses of these antigens resulted in 2 an AUC-, sensitivity- and specificity-values of 0.91, 0.87 and 0.89, respectively. Together, this thesisshowsfor the first time that cancer patients elicit an autoantibody response to citrullinated proteins, resulting in potential novel CRC biomarkers. A novel AP-MS assay was developed to detect autoantibody responses to autologous native CRC tissue proteins. Using the optimised methodology, proteins or homologues of proteins that were significantly (> cut-off value) detected on the CT100plus microarray for the same 5 patients were re-identified by the orthogonal AP-MS method. Using the methodology, PAD2, an enzyme that catalyses the conversion of arginine to citrulline was also identified. In addition, citrullinated antigens associated with cancer were identified, including homologues of CDK7 and MAGEB supporting the conclusion that citrullinated homologues of these proteins induce an autoantibody response in CRC patients. Finally, serum and/or plasma samples of a cohort melanoma patients were analysed using the CT100plus microarray, and autoantibody signals were compared to those of healthy control (HC) samples. Using the established statistical pipeline, the 10 antigens with the highest ROC-derived AUC-values were identified as potential biomarkers. The top 10 biomarker autoantigen candidates for melanoma included CEACAM 1, DPPA2, FGFR2, ITGB1, MAGEA10, NANOG, PIM1, SPANXB1, THEG and XAGE1B. Using these biomarker candidates, distinct clusters of HCs and melanoma patients were identified in both unsupervised hierarchical clustering and PCA analysis. Combinatorial ROC analysis indicates that CEACAM1 and FGFR2 were identified as the top antigens for melanoma diagnosis, together producing sensitivity-, and specificity-, and AUCvalues of 0.96, 0.94 and 0.93, respectively. In conclusion, a statistical pipeline was established for microarray data, enabling the identification of potential antigens associated with cancer diagnosis, and the potential to determine disease prognosis. Using the established pipeline, cancer antigens associated with CRC and melanoma were identified. In addition, an AP-MS assay was developed for the identification of known and novel cancer antigen that can be added to the customisable CT100plus microarray.
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    Epitope Coverage of Anti-SARS-CoV-2 Nucleocapsid IgA and IgG Antibodies Correlates with Protection against Re-Infection by New Variants in Subsequent Waves of the COVID-19 Pandemic
    (2023-02-20) Mullins, Michelle O.; Smith, Muneerah; Maboreke, Hazel; Nel, Andrew J. M.; Ntusi, Ntobeko A. B.; Burgers, Wendy A.; Blackburn, Jonathan M.
    The COVID-19 pandemic continues to affect individuals across the globe, with some individuals experiencing more severe disease than others. The relatively high frequency of re-infections and breakthrough infections observed with SARS-CoV-2 highlights the importance of extending our understanding of immunity to COVID-19. Here, we aim to shed light on the importance of antibody titres and epitope utilization in protection from re-infection. Health care workers are highly exposed to SARS-CoV-2 and are therefore also more likely to become re-infected. We utilized quantitative, multi-antigen, multi-epitope SARS-CoV-2 protein microarrays to measure IgG and IgA titres against various domains of the nucleocapsid and spike proteins. Potential re-infections in a large, diverse health care worker cohort (N = 300) during the second wave of the pandemic were identified by assessing the IgG anti-N titres before and after the second wave. We assessed epitope coverage and antibody titres between the ‘single infection’ and ‘re-infection’ groups. Clear differences were observed in the breadth of the anti-N response before the second wave, with the epitope coverage for both IgG (p = 0.019) and IgA (p = 0.015) being significantly increased in those who did not become re-infected compared to those who did. Additionally, the IgG anti-N (p = 0.004) and anti-S titres (p = 0.018) were significantly higher in those not re-infected. These results highlight the importance of the breadth of elicited antibody epitope coverage following natural infection in protection from re-infection and disease in the COVID-19 pandemic.
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    An investigation into the anti-cancer mechanism of garlic-related organosulfur compounds
    (2014) Smith, Muneerah; Parker, Iqbal; Kaschula, Catherine Hart
    Crushed garlic contains organosulfur compounds (OSC), which are reported to have cancer chemotherapeutic properties both in vitro and in vivo. A library of 15 organosulfur analogues were obtained as mechanistic probes in WHCO1 oesophageal cancer cells. Structure-activity studies showed a positive correlation between the anti-proliferative-IC50 of disulfides and the relative stability of their anion leaving groups, as assessed through resonance and quantified by predictive pKa-values.
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    Longitudinal IgA and IgG Response, and ACE2 Binding Blockade, to Full-Length SARS-CoV-2 Spike Protein Variants in a Population of Black PLWH Vaccinated with ChAdOx1 nCoV-19
    (2023-02-06) Smith, Muneerah; Kwatra, Gaurav; Izu, Alane; Nel, Andrew; Cutland, Clare; Ahmed, Khatja; Baillie, Vicky; Barnabas, Shaun; Bhorat, Qasim; Briner, Carmen; Lazarus, Erica; Dheda, Keertan; Fairlie, Lee; Koen, Anthonet; Madhi, Shabir; Blackburn, Jonathan M.
    Vaccines against SARS-CoV-2 have been pivotal in overcoming the COVID-19 pandemic yet understanding the subsequent outcomes and immunological effects remain crucial, especially for at-risk groups e.g., people living with human immunodeficiency virus (HIV) (PLWH). In this study we report the longitudinal IgA and IgG antibody titers, as well as antibody-mediated angiotensin converting enzyme 2 (ACE2) binding blockade, against the SARS-CoV-2 spike (S) proteins after 1 and 2 doses of the ChAdOx1 nCoV-19 vaccine in a population of Black PLWH. Here, we report that PLWH (N = 103) did not produce an anti-S IgA response after infection or vaccination, however, anti-S IgG was detected in response to vaccination and infection, with the highest level detected for infected vaccinated participants. The anti-IgG and ACE2 blockade assays revealed that both vaccination and infection resulted in IgG production, however, only vaccination resulted in a moderate increase in ACE2 binding blockade to the ancestral S protein. Vaccination with a previous infection results in the greatest anti-S IgG and ACE2 blockade for the ancestral S protein. In conclusion, PLWH produce an anti-S IgG response to the ChAdOx1 nCoV-19 vaccine and/or infection, and ChAdOx1 nCoV-19 vaccination with a previous infection produced more neutralizing antibodies than vaccination alone.