Periodic Reporting for period 4 - RespiriTB (Progress new assets (one pre-new molecular entity and one first-time-in-human start) for tuberculosis that act synergistically with bedaquiline, cytochrome bc or cytochrome bd inhibitors)
Reporting period: 2022-05-01 to 2023-10-31
Mycobacterium tuberculosis (Mtb) causes over 1.5 million deaths each year and has recently reclaimed the position of number one infectious disease killer worldwide. The currently recommended treatment regimens are composed of multiple drugs that need to be taken for a minimum of 6 months, with antibiotic resistance complicating treatment even further. Despite the recent advancements in MDR-TB treatment standards of care, there is an urgent need for novel antibiotics with a shorter treatment duration that can address emerging resistance and simplify treatment. The RespiriTB consortium aims to exploit the respiratory pathway as a source of novel antitubercular leads or new regimens that can be effectively combined with other antibiotics, to quickly reduce bacteremia and drive sterilization. Beyond this specific area of focus, the project will also aim to exploit additional vulnerable and druggable targets in the bacteria or the host that are critical for Mtb survival in humans.
RespiriTB aims to achieve its goals through a parallel three-pronged approach. First, we focus on two essential elements of the mycobacterial electron transport chain, the cytochrome bc and the cytochrome bd complexes. The second is to target mycothione reductase, a critical scavenger of reactive oxygen species that arise naturally in the life of a Mtb cell. Finally, given the intracellular lifestyle of Mtb and the close interplay between bacteria and host during infection, we aim to identify and exploit host and bacterial factors that could drive bacterial clearance by creating conditions under which the bacterium cannot survive. This 'host-directed therapy' has the major advantage that the inhibitors are aimed at human targets, for which the bacterium has no means to generate resistance.
Results 1 - Cytochromes. We have progressed one of the selected cytochrome inhibitors through pre-clinical studies to prepare it for Phase-I clinical trials. Through several rounds of optimization we have increased the potency and safety profile of the lead compound that will soon be submitted for new molecular entity definition. In addition, we have extracted and purified the cytochrome bc and cytochrome bd complexes to study their interaction with the inhibitors using biochemical and structural methods. Multiple cryo-EM structures were determined of cytochrome complexes with different inhibitors bound, to aid in the optimization of the early lead compounds. In addition, an enzymatic assay was implemented to measure the impact of resistance inducing mutations on the activity of the enzyme and impact of different variants of the lead inhibitor.
Results 2 - Mycothione reductase. Since the beginning of the project, we extracted and purified the mycothione reductase protein and used it to identify novel inhibitors. These novel inhibitors are currently being evaluated to select a small set to enter the drug development pipeline. Subsequent optimization of the hit compounds has led to a high-affinity inhibitor with nM IC50. In addition, the crystals structure of the Mycothione reductase bound to this inhibitor was determined. Unfortunately, deletion of the Mycothione reductase gene did not result in immediate killing of mycobacterial cells, while testing of the hit compound on bacterial cultures revealed that the inhibitor acted on a different cellular target. Therefore, this work was discontinued in favor of the development of the cytochrome inhibitors.
Results 3 - Host-directed therapies. We have identified a list of potential host-directed therapies and characterized them in novel host primary cell assay that maximizes the translational value of the results. Based on these results, a shortlist of compounds was selected, and their DMPK and Safety properties were compiled to evaluate their potential for progression to animal models of infection. During the evaluation it was determined that the most potent compound required concentrations that would not be compatible for animal studies. Further cellular work is being conducted to determine the host target of the most potent compound to enable the search for novel, more potent inhibitors that act at concentrations with a beneficial safety profile.
Results 4 - Long acting injectable bedaquiline
Results 1 - Cytochromes. We have progressed one of the selected cytochrome inhibitors through pre-clinical studies to prepare it for Phase-I clinical trials. Through several rounds of optimization we have increased the potency and safety profile of the lead compound that will soon be submitted for new molecular entity definition. In addition, we have extracted and purified the cytochrome bc and cytochrome bd complexes to study their interaction with the inhibitors using biochemical and structural methods. Multiple cryo-EM structures were determined of cytochrome complexes with different inhibitors bound, to aid in the optimization of the early lead compounds. In addition, an enzymatic assay was implemented to measure the impact of resistance inducing mutations on the activity of the enzyme and impact of different variants of the lead inhibitor.
Results 2 - Mycothione reductase. Since the beginning of the project, we extracted and purified the mycothione reductase protein and used it to identify novel inhibitors. These novel inhibitors are currently being evaluated to select a small set to enter the drug development pipeline. Subsequent optimization of the hit compounds has led to a high-affinity inhibitor with nM IC50. In addition, the crystals structure of the Mycothione reductase bound to this inhibitor was determined. Unfortunately, deletion of the Mycothione reductase gene did not result in immediate killing of mycobacterial cells, while testing of the hit compound on bacterial cultures revealed that the inhibitor acted on a different cellular target. Therefore, this work was discontinued in favor of the development of the cytochrome inhibitors.
Results 3 - Host-directed therapies. We have identified a list of potential host-directed therapies and characterized them in novel host primary cell assay that maximizes the translational value of the results. Based on these results, a shortlist of compounds was selected, and their DMPK and Safety properties were compiled to evaluate their potential for progression to animal models of infection. During the evaluation it was determined that the most potent compound required concentrations that would not be compatible for animal studies. Further cellular work is being conducted to determine the host target of the most potent compound to enable the search for novel, more potent inhibitors that act at concentrations with a beneficial safety profile.
Results 4 - Long acting injectable bedaquiline
The project aims to identify one novel drug candidate for progression to first-in-human studies. One of the lead compounds of the cytochrome bc inhibitors is being prepared for a new molecular entitity declaration, an essential step in preparation for phase 1 clinical trials. Additionally, a new asset was introduced into the program, the long acting injectable bedaquiline. Given the successful world-wide use of bedaquiline in the treatment of multi-drug resistant TB, this new long-acting injectable administration would provide a more effective administration for the patient and increase the adherence to the drug regimen. As a seminal step forward for this new approach, phase 1 clinical trials are being prepared that are to be initiated mid 2024.