High-Content Screening Technology Combined with a Human Granuloma Model as a New Approach To Evaluate the Activities of Drugs against Mycobacterium tuberculosis

Abstract

f Tuberculosis remains a major health problem due to the emergence of drug-resistant strains of Mycobacterium tuberculosis. Some models have provided valuable information about drug resistance and efficacy; however, the translation of these results into effective human treatments has mostly proven unsuccessful. In this study, we adapted high-content screening (HCS) technology to investigate the activities of antitubercular compounds in the context of an in vitro granuloma model. We observed significant shifts in the MIC 50 s between the activities of the compounds under extracellular and granuloma conditions. T uberculosis (TB) is a respiratory disease that is one of the major causes of mortality and morbidity worldwide. Almost 20 people develop TB and four patients die from the disease every minute somewhere in the world (1). The disease can be cured by drug treatment involving a regimen of several drugs. The World Health Organization (WHO) estimates that there were around 0.5 million new cases of multidrug-resistant TB (MDR-TB) in 2012 (2, 3). A recent report confirmed that MDR-TB is a continuing worldwide health problem, even in high-income countries with a low incidence of TB (4).The immune system undoubtedly plays a major role in TB control (5). Signaling events of the immune system lead to the formation of a granuloma, the hallmark of TB. Granulomas are an immune microenvironment in which the infection can be controlled but also a niche in which bacilli can thrive and modulate immune responses to ensure their survival for long periods without causing damage (6, 7). Granulomas are cell aggregates that form tridimensional and heterogeneous structures (8, 9). We previously described an in vitro granuloma model involving human blood cells either infected with mycobacteria or incubated with mycobacterial antigens (8, 10). Either of these models resulted in the formation of a typical epithelioid granuloma with morphological characteristics and properties of cellular differentiation similar to those of natural granulomas (8,10,11). This model may help bridge the gap between the in vitro evaluation of MICs and costly in vivo efficacy studies in guinea pigs. In particular, compounds that are effective in the in vitro granuloma model could be prioritized for guinea pig studies.High-content screening (HCS) technology has been used to identify anti-TB compounds (12)(13)(14). This technology has several advantages over traditional phenotypic screening approaches. This technology has mostly been used in single-cell experiments, because it is adapted to identify and analyze images (12, 13); however, in this paper, we focus on the use of granulomas instead of single cells. In this report, we describe the development of a novel HCS method to evaluate the activities of reference compounds against a green fluorescent protein (GFP)-expressing H37Rv strain of Mycobacterium tuberculosis (MTB-GFP) within 5 days, and we compared the MIC 50 values with those of classical liquid cultures (extracellular), intracellular growth (singl...

A Macrophage Infection Model to Predict Drug Efficacy AgainstMycobacterium Tuberculosis

“…48 This would suggest that our model is indeed reflective of or even comparable to other intracellular Mtb drug susceptibility infection models. In addition to these valuable and informative high-content screening methods against intracellular Mtb, 10-12,48 recent development of an in vitro granuloma-based Mtb drug susceptibility phenotyping confirms that drug concentration requirements are altered within granulomas, 49 which further supports findings from our simplified Mtb/macrophage aggregate model. Therefore, that our model is able to produce drug efficacy data consistent with several intracellular Mtb infection models shows its promise as an alternative method to quickly and reliably assess drug efficacy against Mtb during the early drug development process.…”

Hit and lead criteria in drug discovery for infectious diseases of the developing world

“…However, these assays do not fully capture the little-understood mechanisms whereby TB bacteria move in and out of latency [42][43][44] . In addressing this need, a new assay was recently developed for agents that kill M. tuberculosis bacteria that reside in macrophages 45 , resulting in the discovery of Q208 (REF. 46).…”

Translational research in oncology: Implications for palliative care

“…[ 60 ] Novel research techniques might help avoid untoward implications of such necessary practice. Use of predictive models such as isolated, paced Langendorff-perfused heart model in female rabbit to understand drug-induced electrophysiological effects,[ 61 ] “high content screening technology” in humans,[ 62 ] programs such as Simcyp ® 63 and pharmacokinetic-pharmacodynamic modeling[ 64 ] can be contemplated. The extrapolation of these model outputs can help in better planning, efficacy, and reduced side effects in drug trials Biomarkers are used for diagnosis, selection of therapy, prognosis, and for providing insights in disease progression in oncology.…”