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Development of an Autologous Human Dendritic Cell Vaccine against Mycobacterium tuberculosis in Patients with Extensively Drug-Resistant Tuberculosis
Introduction: Extensively drug-resistant tuberculosis (XDR-TB), and resistance beyond XDR-TB (often untreatable), is an increasing public health concern globally. Drug resistance has outpaced the drug development pipeline. Therefore, alternative immunotherapeutic approaches are urgently needed. In t...
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| Format: | Thesis |
| Language: | English |
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Department of Medicine
2023
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| _version_ | 1867613559351934976 |
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| access_status_str | Open Access |
| author | Londt, Rolanda Sabrina |
| author2 | Dheda, Keertan |
| author_browse | Dheda, Keertan Londt, Rolanda Sabrina |
| author_facet | Dheda, Keertan Londt, Rolanda Sabrina |
| author_sort | Londt, Rolanda Sabrina |
| collection | Thesis |
| description | Introduction: Extensively drug-resistant tuberculosis (XDR-TB), and resistance beyond XDR-TB (often untreatable), is an increasing public health concern globally. Drug resistance has outpaced the drug development pipeline. Therefore, alternative immunotherapeutic approaches are urgently needed. In this proof-of-concept study, and based on precedents in breast and prostate cancer, we aimed to develop an autologous therapeutic dendritic cell (DC) vaccine by evaluating two TB antigen-specific multi-peptide pools in combination with different adjuvants (such a cellular vaccine would require ex-vivo manipulation in a clean room and reinfusion back into the patient). Vaccine efficacy was evaluated using an in vitro mycobacterial containment model. Methods: DCs were derived from monocytes isolated from the peripheral blood of patients with XDR-TB (n=30) and participants with presumed latent TB infection (LTBI; n=15). DCs were matured with a differential combination of cytokines and pattern-recognition receptor agonists (maturation cocktail) together with different TB antigen combinations. The complete cocktail contained interferon-, interferon-, CD40L, IL-1 and TLR-3, TLR-7 and TLR-8 agonists, whilst in the limited cocktail the TLR agonists were absent. Two M.tb-specific multi peptide pools suited to GMP-grade vaccine manufacture were evaluated: (i) an immunodominant peptide pool (ESAT6 + CFP10 + Ag85B + TB10.4; referred to as ECAT) and (ii) a PE/PPE peptide pool. PPD and the lysate of a clinical M.tb strain (HN878), though not amenable to GMP-grade vaccine development, served as controls representing the entire antigenic repertoire of M.tb. Thus, there were four major comparator groups (unstimulated DCs, limited cocktail-only stimulated DCs, antigen-only stimulated DCs, and antigen + complete cocktail stimulated DCs). As the two latter groups were interrogated using the two multi-peptide pools and the two broad-spectrum antigen controls, a total of 10 different experimental groups were generated (see overview figure 3.2). DCs were assessed for the expression of key maturation markers using flow cytometry and the secretion of Th1-polarising cytokines by ELISA. The ability of DC-primed peripheral blood mononuclear cells (PBMCs) to restrict the growth of M.tb-infected monocyte derived-macrophages was evaluated using an in vitro validated mycobacterial containment assay. Results: In patients with XDR-TB, DCs matured with any M.tb-antigen + complete cocktail, compared to DCs matured with M.tb-antigen only, showed significantly higher upregulation of CD80, CD83, CD86, and CCR7 (p< 0.001 for all comparisons), and higher secreted levels of IL12p70 (0.67 versus 0.01 ng/mL per 106 cells; p< 0.001). A similar pattern was seen in the containment experiments: mycobacterial stasis within the XDR-TB group was significantly better with antigen + complete cocktail versus antigen alone (p≤0.0002 for PE/PPE and PPD), and the limited cocktail did not show this effect. Furthermore, PE/PPE + complete cocktail matured DCs achieved a higher magnitude of mycobacterial containment compared to ECAT + complete cocktail-matured DCs (50%, IQR:39-75, versus 46%, IQR: 15-62, p= 0.02). Using PPD and the HN878 lysate did not improve the containment effect. Furthermore, the improved containment effect of the PE/PPE + complete cocktail, versus ECAT + complete cocktail, was only seen in the XDR-TB and not in the LTBI group. Conclusion: In patients with XDR-TB, an effector response primed by PE/PPE antigen and complete cocktail-matured DCs was able to better restrict the growth of M.tb in vitro. These data indicate proof-of-concept feasibility to generate a DC-based immunotherapeutic intervention for therapeutically destitute patients with DR-TB. Further mechanistic studies and future phase 1 and 2 human clinical studies are warranted. |
| format | Thesis |
| id | oai:open.uct.ac.za:11427/37467 |
| institution | University of Cape Town (South Africa) |
| language | eng |
| last_indexed | 2026-06-10T12:38:04.526Z |
| license_str | Not specified — see source repository |
| provenance_str_mv | Harvested via OAI-PMH from UCTD — University of Cape Town Open Access Repository |
| publishDate | 2023 |
| publishDateRange | 2023 |
| publishDateSort | 2023 |
| publisher | Department of Medicine |
| publisherStr | Department of Medicine |
| record_format | dspace |
| source_str | UCTD — University of Cape Town Open Access Repository |
| spelling | oai:open.uct.ac.za:11427/37467 Development of an Autologous Human Dendritic Cell Vaccine against Mycobacterium tuberculosis in Patients with Extensively Drug-Resistant Tuberculosis Londt, Rolanda Sabrina Dheda, Keertan Tomasicchio, Michele Clinical Science and Immunology Introduction: Extensively drug-resistant tuberculosis (XDR-TB), and resistance beyond XDR-TB (often untreatable), is an increasing public health concern globally. Drug resistance has outpaced the drug development pipeline. Therefore, alternative immunotherapeutic approaches are urgently needed. In this proof-of-concept study, and based on precedents in breast and prostate cancer, we aimed to develop an autologous therapeutic dendritic cell (DC) vaccine by evaluating two TB antigen-specific multi-peptide pools in combination with different adjuvants (such a cellular vaccine would require ex-vivo manipulation in a clean room and reinfusion back into the patient). Vaccine efficacy was evaluated using an in vitro mycobacterial containment model. Methods: DCs were derived from monocytes isolated from the peripheral blood of patients with XDR-TB (n=30) and participants with presumed latent TB infection (LTBI; n=15). DCs were matured with a differential combination of cytokines and pattern-recognition receptor agonists (maturation cocktail) together with different TB antigen combinations. The complete cocktail contained interferon-, interferon-, CD40L, IL-1 and TLR-3, TLR-7 and TLR-8 agonists, whilst in the limited cocktail the TLR agonists were absent. Two M.tb-specific multi peptide pools suited to GMP-grade vaccine manufacture were evaluated: (i) an immunodominant peptide pool (ESAT6 + CFP10 + Ag85B + TB10.4; referred to as ECAT) and (ii) a PE/PPE peptide pool. PPD and the lysate of a clinical M.tb strain (HN878), though not amenable to GMP-grade vaccine development, served as controls representing the entire antigenic repertoire of M.tb. Thus, there were four major comparator groups (unstimulated DCs, limited cocktail-only stimulated DCs, antigen-only stimulated DCs, and antigen + complete cocktail stimulated DCs). As the two latter groups were interrogated using the two multi-peptide pools and the two broad-spectrum antigen controls, a total of 10 different experimental groups were generated (see overview figure 3.2). DCs were assessed for the expression of key maturation markers using flow cytometry and the secretion of Th1-polarising cytokines by ELISA. The ability of DC-primed peripheral blood mononuclear cells (PBMCs) to restrict the growth of M.tb-infected monocyte derived-macrophages was evaluated using an in vitro validated mycobacterial containment assay. Results: In patients with XDR-TB, DCs matured with any M.tb-antigen + complete cocktail, compared to DCs matured with M.tb-antigen only, showed significantly higher upregulation of CD80, CD83, CD86, and CCR7 (p< 0.001 for all comparisons), and higher secreted levels of IL12p70 (0.67 versus 0.01 ng/mL per 106 cells; p< 0.001). A similar pattern was seen in the containment experiments: mycobacterial stasis within the XDR-TB group was significantly better with antigen + complete cocktail versus antigen alone (p≤0.0002 for PE/PPE and PPD), and the limited cocktail did not show this effect. Furthermore, PE/PPE + complete cocktail matured DCs achieved a higher magnitude of mycobacterial containment compared to ECAT + complete cocktail-matured DCs (50%, IQR:39-75, versus 46%, IQR: 15-62, p= 0.02). Using PPD and the HN878 lysate did not improve the containment effect. Furthermore, the improved containment effect of the PE/PPE + complete cocktail, versus ECAT + complete cocktail, was only seen in the XDR-TB and not in the LTBI group. Conclusion: In patients with XDR-TB, an effector response primed by PE/PPE antigen and complete cocktail-matured DCs was able to better restrict the growth of M.tb in vitro. These data indicate proof-of-concept feasibility to generate a DC-based immunotherapeutic intervention for therapeutically destitute patients with DR-TB. Further mechanistic studies and future phase 1 and 2 human clinical studies are warranted. 2023-03-16T10:14:39Z 2023-03-16T10:14:39Z 2022 2023-03-16T09:51:05Z Doctoral Thesis Doctoral PhD http://hdl.handle.net/11427/37467 eng application/pdf Department of Medicine Faculty of Health Sciences |
| spellingShingle | Clinical Science and Immunology Londt, Rolanda Sabrina Development of an Autologous Human Dendritic Cell Vaccine against Mycobacterium tuberculosis in Patients with Extensively Drug-Resistant Tuberculosis |
| thesis_degree_str | Doctoral |
| title | Development of an Autologous Human Dendritic Cell Vaccine against Mycobacterium tuberculosis in Patients with Extensively Drug-Resistant Tuberculosis |
| title_full | Development of an Autologous Human Dendritic Cell Vaccine against Mycobacterium tuberculosis in Patients with Extensively Drug-Resistant Tuberculosis |
| title_fullStr | Development of an Autologous Human Dendritic Cell Vaccine against Mycobacterium tuberculosis in Patients with Extensively Drug-Resistant Tuberculosis |
| title_full_unstemmed | Development of an Autologous Human Dendritic Cell Vaccine against Mycobacterium tuberculosis in Patients with Extensively Drug-Resistant Tuberculosis |
| title_short | Development of an Autologous Human Dendritic Cell Vaccine against Mycobacterium tuberculosis in Patients with Extensively Drug-Resistant Tuberculosis |
| title_sort | development of an autologous human dendritic cell vaccine against mycobacterium tuberculosis in patients with extensively drug resistant tuberculosis |
| topic | Clinical Science and Immunology |
| url | http://hdl.handle.net/11427/37467 |
| work_keys_str_mv | AT londtrolandasabrina developmentofanautologoushumandendriticcellvaccineagainstmycobacteriumtuberculosisinpatientswithextensivelydrugresistanttuberculosis |