New approaches for challenging therapeutic targets.

Despite successes with new drug approvals over the past two decades through conventional drug development approaches, many human diseases remain intractable to current therapeutic interventions. Possible barriers may be that the complexity of the target, and disease biology, are impervious to such conventional drug development approaches. The US National Institutes of Health hosted a workshop with the goal of identifying challenges and opportunities with alternative modalities for developing treatments across diseases associated with historically undruggable targets. This report highlights key issues discussed during the workshop that, if addressed, could expand the pool of therapeutic approaches for treating various diseases.

Preclinical murine models for the testing of antimicrobials against Mycobacterium abscessus pulmonary infections: Current practices and recommendations.

Mycobacterium abscessus, a rapidly growing nontuberculous mycobacterium, is increasingly recognized as an important pathogen of the human lung, disproportionally affecting people with cystic fibrosis (CF) and other susceptible individuals with non-CF bronchiectasis and compromised immune functions. M. abscessus infections are extremely difficult to treat due to intrinsic resistance to many antibiotics, including most anti-tuberculous drugs. Current standard-of-care chemotherapy is long, includes multiple oral and parenteral repurposed drugs, and is associated with significant toxicity. The development of more effective oral antibiotics to treat M. abscessus infections has thus emerged as a high priority. While murine models have proven instrumental in predicting the efficacy of therapeutic treatments for M. tuberculosis infections, the preclinical evaluation of drugs against M. abscessus infections has proven more challenging due to the difficulty of establishing a progressive, sustained, pulmonary infection with this pathogen in mice. To address this issue, a series of three workshops were hosted in 2023 by the Cystic Fibrosis Foundation (CFF) and the National Institute of Allergy and Infectious Diseases (NIAID) to review the current murine models of M. abscessus infections, discuss current challenges and identify priorities toward establishing validated and globally harmonized preclinical models. This paper summarizes the key points from these workshops. The hope is that the recommendations that emerged from this exercise will facilitate the implementation of informative murine models of therapeutic efficacy testing across laboratories, improve reproducibility from lab-to-lab and accelerate preclinical-to-clinical translation.

Identification of 14-3-3zeta by chemical affinity with salicylanilide inhibitors of interleukin-12p40 production.

Salicylanilides were found as selective inhibitors of interleukin-12p40 production in stimulated dendritic cells. The conversion of one of these bioactive salicylanilides into a comparably bioactive, chemically labeled derivative was achieved using a facile and systematic functional group derivatization strategy. This resulted in a tool reagent that was then employed in an affinity chromatography approach that resulted in the identification of the protein 14-3-3zeta as having selective affinity for the chromatography matrix that was derivatized with a salicylanilide that inhibited IL-12p40 production.

Salicylanilides: selective inhibitors of interleukin-12p40 production.

Interleukin (IL)-12p40, a subunit component of both IL-12 and IL-23, is being widely studied for its role in inflammatory disease. As part of an effort to profile cellular signaling pathways across different cell types, we report salicylanilide inhibitors of IL-12p40 production in stimulated dendritic cells. Based on a hypothesis that a desirable therapeutic profile is one that could block IL-12p40 but not IL-6 production, we engaged in directed analoging. This resulted in salicylanilides with similar IL-12p40 related potency but enhanced selectivity relative to IL-6 production.

Target-based identification of whole-cell active inhibitors of biotin biosynthesis in Mycobacterium tuberculosis.

Biotin biosynthesis is essential for survival and persistence of Mycobacterium tuberculosis (Mtb) in vivo. The aminotransferase BioA, which catalyzes the antepenultimate step in the biotin pathway, has been established as a promising target due to its vulnerability to chemical inhibition. We performed high-throughput screening (HTS) employing a fluorescence displacement assay and identified a diverse set of potent inhibitors including many diversity-oriented synthesis (DOS) scaffolds. To efficiently select only hits targeting biotin biosynthesis, we then deployed a whole-cell counterscreen in biotin-free and biotin-containing medium against wild-type Mtb and in parallel with isogenic bioA Mtb strains that possess differential levels of BioA expression. This counterscreen proved crucial to filter out compounds whose whole-cell activity was off target as well as identify hits with weak, but measurable whole-cell activity in BioA-depleted strains. Several of the most promising hits were cocrystallized with BioA to provide a framework for future structure-based drug design efforts.