Intravesical instillations of Mycobacterium bovis bacillus Calmette-Guerin (BCG) is the preferred treatment in patients with non-muscle-invasive bladder cancer (NMIBC) after tumour resection. M. bovis BCG has demonstrated efficacy in reducing the recurrence and tumor progression due to the activation of the patient immune system, which enhances the survival rate of individuals with NMIBC. However, intravesical treatment with this immunotherapeutic agent is associated with adverse events in more than 50% of treated patients, being able to develop serious complications in 5% of cases, including lung or disseminated M. bovis BCG infections. Furthermore, up to 30-40% of patients do not respond to M. bovis BCG therapy and, consequently, they have to finish the treatment. One of the alternatives proposed to replace the M. bovis BCG treatment is the use of Mycobacterium brumae, an environmental non-pathogenic mycobacterium with an antitumor and immunomodulatory effect demonstrated in vitro, ex vivo and in vivo in a murine model of bladder cancer. With the aim of developing new therapeutic alternatives that are effective for all patients, the objective of this thesis is to decipher into the phenotypic characteristics and genotypes of different strains of M. brumae and compare them with M. bovis BCG; to evaluate their interaction with different tumour cells; to study the immunemodulatory effect of both mycobacteria in the specific context of older people; and to characterize its potential ability to induce trained immunity. Initially, the genome of the reference strain and three environmental isolates of M. brumae were sequenced and analysed. The comparison with the genome of M. bovis BCG and M. tuberculosis demonstrated the non-pathogenicity of M. brumae, as well as the presence of genes involved in the activation of the host immune system demonstrating its immunogenicity. No differences were observed at the phenotypic level of the different strains of M. brumae and very few at the genotypic level. However, it was observed that the CR-103 strain could inhibit some tumour cell proliferation more efficiently. Besides, the CR-142 strain induced an increase in cytokine production in infected macrophages, compared with the rest of the strains. The ability of M. brumae to inhibit the tumour proliferation was demonstrated in tumour cell lines from locations other than the bladder, such as colon, liver, or pancreas. On the other hand, the ability of M. brumae and M. bovis BCG to activate the immune system from young and old healthy populations, as well as from patients with bladder neoplasia was evaluated. Using peripheral blood mononuclear cells (PBMC) a significant difference between the evaluated populations in terms of triggering the production of proinflammatory cytokines and the antitumor capacity of the activated PBMCs. Finally, the capacity of M. brumae to induce trained immunity, compared to M. bovis BCG, was evaluated in different in vitro and ex vivo cultures using an in vitro model with THP-1 human monocytes and PBMC-derived monocytes, respectively. The results indicated that the initial stimulation of monocytes with M. brumae followed by a secondary non-specific stimulus induce an elevated production of IL-1[Beta], IL-6, and TNF-[Alfa], and an enhanced lactate production. This indicates a change in the metabolic profile of the cells and shows the potential of M. brumae to induce trained immunity.
Estudio in silico, in vitro y ex vivo de la actividad antitumoral de diferentes cepas de Mycobacterium Brumae y su papel en la activación y modulación del sistema inmune
Herrero Abadia, P. (Author). 2 Nov 2023
Student thesis: Doctoral thesis
Student thesis: Doctoral thesis