Understanding immune response to tuberculosis infection to help design new vaccines

Background

This project sought to determine if the protein antigens in two candidate tuberculosis (TB) vaccines that are protective against experimental TB infection are recognised by the T cell immune responses in Indonesian TB patients. This is an essential step in the selection of these vaccines for development as new TB vaccines. This was a collaboration between medical researchers at the University of Sydney and Universitas Gadjah Mada (UGM) to facilitate the transfer of immunological assays from Sydney to UGM.

Project objectives were:

• To establish immunological assay for the T cell recognition of M. tuberculosis protein antigens by TB patients in the TB laboratory at the Faculty of Medicine, UGM.
• To hold a workshop on tuberculosis research at UGM, focusing on the immune response to M. tuberculosis and development of new TB vaccines.
• To test the immune responses to M. tuberculosis protein antigens in TB patients and healthy endemic controls from the Provincial Lung Clinic.
• To determine if HIV co-infection in tuberculosis patients reduces the I cell response to the M. tuberculosis protein antigens.

The aim of the study was to understand the human immune response to TB vaccine antigen candidates in the setting of a high TB burden country. This project successfully initiated a collaboration on TB research between the Sydney Medical School at the University of Sydney and the Center for Tropical Medicine, Faculty of Medicine, UGM. The transfer of two immunology assays, ELISA and ELISpot, was conducted by sending an Indonesian PhD student from the University of Sydney, Heni Muflihah, to establish the assays and train Indonesian scientists in UGM, Yogyakarta. This was different to the planned activity of training Indonesian scientists in Sydney. This modification was required after initial assessment of the facilities and unexpected changes of safety requirements in the Indonesian laboratories. The last was also the reason for the inability to recruit HIV-TB patients and perform immunological assays in these patients.

A study to understand the human immune response to TB and TB vaccine antigens was conducted in Yogyakarta. Two protein antigens, CysD and MPT83, were developed in The University of Sydney. Studies by Professor Britton and Triccas have demonstrated that TB vaccines based on these protein antigens induce partial protective immunity against M. tuberculosis infection in aerosol model of TB infection in mice. This study was to test the human responses to these proteins, including three additional M. tuberculosis recombinant protein antigens. The protocol ethics for this study were approved by the University of Sydney Human Ethics Committee (Project No.2015/346), and by the Medical and Health Research Ethics Committee, Faculty of Medicine UGM. The application for protocol ethics was prolonged, contributing to delayed initiation of the study.

The study started in October 2015 and was finished by August 2016. Of 76 recruited participants, 33 subjects were active TB patients, and 43 subjects were healthy controls. Venous blood was collected and peripheral blood mononuclear cells were isolated for testing response to TB vaccine antigen candidates by the two immunology assays, ELISA and ELISpot. The M. tuberculosis recombinant protein antigens tested in this study were CysD, MPT83, Ag85B, ESAT-6, and CFP-10.

Findings

Generally, this study was able to measure the human immune response to TB vaccine candidates by ELISA. The responses to the antigens in active TB patients were similar to the response from healthy controls. Overall responses in the ELISpot assay were variable because reading the ELISpot plates by microscope in the field setting proved unreliable. The ELISpot technique requires an automated plate reader for accurate results.

Nevertheless, all five proteins tested were recognised by T cells from 40-50 per cent of active TB patients, including the two vaccine antigens, CysD and MPT83, developed at The University of Sydney. Future TB vaccines should use a combination of these antigens.