Studies on new tuberculosis vaccine candidates in animal models

Abstract

Background: Tuberculosis (TB) is a major health problem in many countries, especially in low income countries. Globally, TB causes more than 2 million human deaths annually and 1/5 of all adult deaths in developing countries. WHO estimates that one third of the world's population i.e. 1.9 billion people, are infected with M. tuberculosis (Mtb). The most cost effective way to combat infectious diseases is the preventive vaccination. Most of the world's population is vaccinated with the only available TB vaccine, the Bacillus Calmette-Guérin (BCG) vaccine that was developed a century ago. Even though the BCG vaccine protects the young child against disseminated TB disease it has none or little protective effect against adult pulmonary TB (PTB). PTB is the major disease manifestation of TB in adults and it causes death at the most productive age, further adding to poverty in already impoverished countries. Hence, the development of a new more efficient TB vaccine(s) is the highest priority in TB research.

Aims: The aim of this thesis was to evaluate protective efficacy of new TB vaccine candidates with the long term goal to improve the efficacy of primary BCG vaccination against adult PTB by boosting with new vaccine(s).

Results: New vaccine candidates and adjuvant/delivery systems for mucosal applications were developed and tested in animal models. Lipoarabinomannan (LAM) was purified from a virulent Mtb strain and used to prepare novel oligosaccharide-protein conjugate vaccines. To avoid the immuno-suppressive effects of the intact LAM molecule while taking advantage of its carbohydrate antigen epitopes, LAM was de-lipidated and split into oligosaccharides by partial chemical degradation. The arabinomannan oligosaccharides (AM) were used to prepare AM-protein conjugate (AM-Prot) vaccines and to generate a large number of monoclonal antibodies (MoAbs). Adjuvants can augment the immune response to many vaccine antigens and for some sub-cellular vaccines e.g. Diphteria and Tetanus vaccine, they are essential. There is a vast number of experimental adjuvants but most of them are intrinsically toxic and only few can be considered for use in man. Aluminum salts are so far the only adjuvants approved for large scale human use. In this thesis a new adjuvant L3 was investigated. L3 is non-toxic and approved by the Swedish FDA for human phase I/II trials. The different AM-Prot vaccines and a vaccine based on heat killed whole-cell BCG (H-kBCG) were formulated with L3 and studied for their ability to protect against virulent Mtb challenge in the mouse and guinea pig models - both as primary- and boost-vaccines. Both types of vaccines, when given nasally, evoked specific and robust cellular and humoral immune responses. The vaccines also conferred significant protection, at least equivalent to that induced by the BCG vaccine. Presently the general consensus is that cell mediated immunity (CMI) is decisive for protection against Mtb infections. This, however, has led investigators to neglect the role of the humoral immunity. Importantly, high antibody titers in particular towards LAM are seen in most TB patients but the role of these antibodies is unknown. Therefore, in this thesis the role of antibodies in Mtb infection was re-evaluated in passive protection experiments using an AM-specific MoAb. Mice were infected intravenously with virulent Mtb and the MoAb was added intravenously either prior to or together with the bacteria. The MoAb protected against the infection in terms of a dosedependent reduction in bacterial load in spleens and lungs, reduced weight loss and, importantly, enhanced long-term survival.

Conclusions: 1) AM-Prot vaccines can, when formulated with the L3 adjuvant and given nasally, provide as good protection as live BCG, ii) The protection is also conferred by conjugates containing Mtb-irrelevant carrier protein (i.e. tetanus toxoid) showing that antigenic epitopes of AM alone are protective. Therefore we propose that AM epitopes should be considered as part of a future new multi-sub-component vaccine, iii) The AM-Prot vaccine affords protection both when used as a primary vaccine and as a boost vaccine, iv) A H-kBCG vaccine did not protect when administered alone but was protective when given in the L3 adjuvant. The pre-clinical studies of this vaccine candidate are concluded and the vaccine is considered ready for phase 1/11 trials in man, v) None of the vaccines ameliorated the course of TB when used as a therapeutic regimen. Hence, caution is advised in the use of therapeutic vaccines, and in the design of phase 1-111 trials great care must be taken not to include individuals with active TB disease, vi) Certain antiAM antibodies may confer protection, most likely aiding to the early containment of infection and thus giving time for a protective cellular immune response to develop.