An excerpt from the  PLOS blog of Sheetal Gandotra

The journey from compound to drug to resistance

Little did Hans Meyer and Joseph Malley know in 1912 that their doctoral work on the synthesis of isoniazid (INH) would 40 years later become the reason for a patent struggle between three pharma companies that would discover its potency against the tubercle bacillus – Mycobacterium tuberculosis (Mtb). In 1951 INH replaced streptomycin for which resistance and toxicity already raised problems for treatment. The first publication describing the emergence of INH resistant Mtb in clinic appeared in 1952. Thanks to years of research from Robert Koch in late 19th century and to Dubos, Lowenstein and Middlebrook in mid-20th century, culturing the tubercle bacilli was now possible but it still took three weeks or more until bacterial colonies formed and allowed determining if the strain was resistant to INH. The patient, meanwhile, was treated with a drug that not only lacked any effect on the bacteria, destroying the once healthy lung tissue and compromised the patient’s ability to breathe, but was also challenged with toxic side effects of the drug. Genetic tools borne out of laborious research on mycobacteriophages by William Jacobs and Graham Hatful have been crucial in positioning TB research where it is today. In 1992, Stewart Cole applied genetics to identify the first gene responsible for INH resistance in Mtb clinical isolates. This information soon translated into genotyping for antibiotic resistance in clinical settings. Today, with the development of rapid whole genome sequencing, we are at the helm of understanding at the global genomic level how bacteria retain their fitness despite their ability to mutate and develop drug resistance. It’s been over 100 years in our battle against TB but mechanisms underlying the interactions of the bacillus and its host are far from understood. Work done at the Hinduja hospital in Mumbai by Dr. Camilla Rodriguez showed that bacterial antibiotic sensitivity can vary from lesion to lesion within the same individual. So clinically, multi-drug resistance may be an outcome of several mono-drug resistant strains in addition to being a case of MDR at the strain level.

Gaining perspective of the bacillus

Given that drug resistance exists against all current TB drugs, the need for new drugs has been a focus of many research quests ranging from target based and whole cell screening approaches, to the identification and validation of new targets. Many exciting routes have been laid out by researchers in the last twenty years in their attempts to find new targets for anti-TB therapy.  This has included screening genome-wide transposon mutant libraries for “essential” genes of the bacilli, developing novel gene manipulation methods to generate and characterize mutants of essential genes, and gaining knowledge of the pathogen’s response to what may be a hostile environment created by the host’s immune system and to drug treatment. Gene expression data can be computed to predict adaptations made by the bacterium; these models will only be complete when individual genes and their biochemical interaction networks in the context of infection or drug treatment will be understood. Research is an ongoing endeavor and existing paradigms have to be revisited with new approaches and information that enhances our ability to understand the pathogenesis of TB. A personal favorite in this light is the transcriptional regulator DevR, first identified in the laboratory of Jaya Tyagi in India. Her lab identified the gene encoding this protein in early 1990s as a gene that was more strongly expressed in virulent Mtb compared to an avirulent strain. It was later re-discovered by the groups of David Sherman in the USA and Thomas Dick in Singapore as DosR, a transcriptional regulator important for adaptation to hypoxia, a condition that became associated with bacterial dormancy. The genes induced in response to DevR activation became the signature for “dormant” bacteria while controversy persisted on its role in virulence. In 2008, Michael Barer’s group in the UK discovered mycobacteria harbouring this “signature” in the sputum of actively infected individuals. However, phenotypically distinct subpopulations of bacilli can account for differences in virulence, fitness, and drug response. Tackling this variability and its origin is a long term challenging feat but is likely to provide new means to tackle this pathogen.

Focus on the human host

Koch’s postulates came into existence primarily because of his research on tuberculosis; he showed that the pathogen could be isolated from diseased human tissue and that it could be inoculated into a healthy host to generate disease. With years of research in various model organisms including mouse, rabbit, guinea pig, and macaque, we have learnt of various facets of the host response to infection and how these shape the outcome of infection. The TB mouse model has been the most genetically tractable to investigate the role of the host immune response in controlling disease; however, not all factors associated with murine susceptibility have been proved risk factors for human TB. However, if it were not for in vivo and macrophage models of infection, we would probably not have pyrazinamide as a front line drug against TB. The rabbit, guinea pig and in particular non-human primate models offer insights into TB pathogenesis that more closely mimics that of humans. While most published research has focused on pulmonary models of animal infection, little is known about extrapulmonary manifestations in the presence or absence of pulmonary involvement. Extrapulmonary TB in humans has its own problems – debilitating tissue damage, a two year or even longer treatment regimen, the pain and cost of invasive examinations, and the side effects of antibiotics. These are saddening realities that we basic researchers must not lose sight of when using a model system. Tools of modern biology including stem cell therapy may advance treatment methods and result in a better understanding of cellular and tissue level dynamics of this host-pathogen relationship. However, for treating the large number of patients – 2.3 million/year in India – we will need solutions that will work in resource poor settings as well.

Basic research has provided and continues to provide new solutions, awareness, and novel paradigms for treatment. Yes, the bacterium has outsmarted us, but we are today better prepared than ever to decode its mantra. Implementation of new solutions will require sustained public-private partnerships but without continued investment in high quality integrative research based on ideas that haven’t been tackled before, our preparedness is not going to yield. As a PhD student in the US, working on TB taught me not only the rigors of high quality research but also the patience, diligence, and team work required to address a novel paradigm in host-pathogen interplay. Today I am working at the CSIR-Institute of Genomics and Integrative Biology in New Delhi, India, with a mission not only to advance our understanding of the complex host-pathogen interplay in TB but also to seek answers to its success. Being in India, the sheer number of people affected by TB astounds me. While my nation seeks implementation of better TB control programs and doctors struggle to treat patients with drug resistance, scientists like me work around the clock looking for solutions. Combining perspectives from genomics, computational biology, immunology, cell biology, microbial diversity, and biochemistry, we must challenge our enemy TB with full force.


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