Kasturi Shikshan Sanstha College of Pharmacy, DBATU.
The evolution between Mycobacterium tuberculosis infection and active tuberculosis is multi factorial and involves different biological scales. The synthesis of ESAT-6 or the induction of alveolar macrophage necrosis are key, but to understand it, it is necessary to consider the dynamics of endogenous and exogenous reinfection, drainage of lung parenchyma and respiratory mechanics, local fibrosis processes and blood supply. Severe immunosuppression can only explain 10% of active tuberculosis cases, while the remainder are attributable to comorbidities, a proinflammatory environment and an unknown genetic propensity. The pathogenic capacity of environmental mycobacteria is discrete linked to deficits in the innate and acquired immune response. The ability to generate biofilms and the ability of M. ulcer to generate the exotoxin mycolactone is remarkable. The recommendation of the world health organization and the renewed guideline of the Hungarian board of pulmonology include all criteria necessary to the effective prevention, diagnosis and therapy of multidrug, and extensively drug resistant tuberculosis. This incidence corresponds to the epidemiological data of western European countries.
In 1882, Robert Koch identified the tubercles bacillus, also known as Mycobacterium tuberculosis as the etiologic agent of tuberculosis. The TB epidemic seems to be unabated, spreading in every corner of the globe. Tb is a highly contagious airborne disease and one of the top causes of the death worldwide. When the stalement is broken TB reactivation occurs and the bacterial burden soars, wherefore the disease become symptomatic. .M .tb can stay dormant for years and persist in the body without any indication of illness, in which many people become asymptomatic carriers.
Epidemiology: According to the WHO, in 2014, an estimated 9.6 million people developed active TB disease, of whom 1.5 million died1. The burden of TB is heterogeneously distributed . For example, TB incidence is >250?fold higher in South Africa (834 cases per 100,000 population per year) than in the United States (3 cases per 100,000 population per year)1. Rates of developing throughout their lifetime18. In many settings, up to 50% of all people with culture-positive active TB disease do not have a prolonged productive (phlegm or mucus-producing) cough, and at least 25% have no symptoms whatsoever19. Thus, the progression from LTBI to active TB disease can be clinically subtle, despite the fact that individuals with subclinical TB can transmit the organism to others20.
Figure 1 The spectrum of TB — from Mycobacterium tuberculosis infection to active (pulmonary) TB disease.
Types of tuberculosis:
Broadly classified, there are two types of tuberculosis infections
In active tuberculosis , the individual who is carrying the organism has active symptoms and can transmit the infection to the other people.
In latent tuberculosis, the individual carries the bacteria but does not exihibit any symptoms whatsoever. This is because immunity fights the infection and is able to suppress it to an extent. The bacteria can get reactived and the infection can become active tuberculosis.
Figure 3 Mycobacterium tuberculosis infection.
a | Infection begins when Mycobacterium tuberculosis enters the lungs via inhalation, reaches the alveolar space and encounters the resident alveolar macrophages. If this first line of defence fails to eliminate the bacteria, M. tuberculosis invades the lung interstitial tissue, either by the bacteria directly infecting the alveolar epithelium or the infected alveolar macrophages migrating to the lung parenchyma. Subsequently, either dendritic cells or inflammatory monocytes transport M. tuberculosis to pulmonary lymph nodes for T cell priming. This event leads to the recruitment of immune cells, including T cells and B cells, to the lung parenchyma to form a granuloma.
b | The bacteria replicate within the growing granuloma. If the bacterial load becomes too great, the granuloma will fail to contain the infection75 and bacteria will disseminate eventually to other organs, including the brain. At this phase, the bacteria can enter the bloodstream or re?enter the respiratory tract to be released — the infected host is now infectious, symptomatic and is said to have active TB disease.
Figure 2 Global incidence of active TB disease (pulmonary and extrapulmonary).
High-income countries — including most countries in western Europe, Canada, the United States, Australia and New Zealand — have the lowest rates of active tuberculosis (TB) disease, typically <10>
Histological:
Robert Koch identified and described the bacillus causing tuberculosis on 24 march 1882. In 1905, he was awarded the nobel prize in physiology or medicine for this discovery. The subsequents resurgence of tuberculosis resulted in the declaration of the global health emergency by the world health organization in 1993.
Granulomatous inflammation forms both caseating and noncaseating tubercles.
TB Pathogenesis
The pathogenic life cycle of M.tb is transmitted via M. tb containing aerosol droplets, propelled by active TB patients when they cough,sneeze or talk. The granuloma is the cardinal feature of pulmonary TB, which is amorphous collection of macrophages and other immune cell aimed at restricting the bacterial spread. In immune competent individuals, although the granuloma is unable to eliminate the pathogen, it restrain the bacilli and halts the progression to the active disease. As the granuloma matures, macrophages differentiate into foamy macrophages and other various morphotypes. The centre of the granuloma may necrotisc as a results of the necrotic lysis of the host immune cells forming what is referred to as caseum. Indeed, the accumulating soft necrotic debris, located in the core of the granuloma, resembles cheese. Foamy macrophages, which are charaterised by accumulated lipid droplets, distribute around the necrotic foci of the granuloma. In this case, the patient is still non infectious and asmptomatic. One of the challenges facing the current TB therapy is targeting this tenacious pathogen inside the granuloma. At the this point, the hosts innate immune system comes into play to quell the infection, the tubercle bacilli, are internalized by alveolar macrophages. When the macrophages fail to inhibit or destroy the bacillus , the bacteria multiply within their intracellular environment , get released, then phagocytosed by other alveolar macrophages and the cycle continues. Lymphocytes are the recruited to the infection sites, initiaging a cell-mediated immune response, in which a pile of immune cells arrives, attempting to sequester the bacterial and limit further multiplication.
Quality of life
Several studies have documented lower self-reported health-related quality of life among patients with active TB disease198 than healthy individuals or those with LTBI. Impairment of lung function with chronic pulmonary disability, bronchiectasis, aspergillomas and chronic pulmonary aspergillosis are known complications and are more frequent in patients with drug-resistant TB than in patients with drug-sensitive TB199. Patients with impaired lung function might require long?term pulmonary rehabilitation and chest physiotherapy. If patients are untreated, the prognosis for individuals affected by drug-resistant TB is similar to the prognosis for individuals with drug-sensitive TB (10?year case fatality rates of approximately 70%)16. The current WHO-recommended MDR?TB regimen has an approximate 50% cure rate, whereas the cure rate in endemic settings of extensively drug-resistant TB in the absence of drugs such as bedaquiline, delamanid and linezolid is approximately 207,200. Thus, TB (and drug-resistant TB in particular) poses a grave threat to human health and quality of life.
Mechanisms of drug resistance
TB is the infectious disease in which the phenomenon of drug resistance was first described in 1948, during the very first human trial of TB therapy86. As each new anti-TB drug has been introduced into clinical practice, widespread emergence of resistant strains has been described, usually within a decade. M. tuberculosis develops drug resistance through genetic mutations (there are no reports of resistance developed by the acquisition of new DNA). Although there is an ever-expanding list of genes that have been linked to resistance, allelic exchange experiments have confirmed the causality between mutation and drug resistance for only a subset of mutated genes87. In these genes, the two major mechanisms of drug resistance are target modification (for example, a mutant bacterial RNA polymerase that eludes the action of rifampicin) or a defective enzyme that converts a pro-drug into an active drug (for example, a mutant bacterial catalase that fails to activate isoniazid).
Current treatment regimen for drug-sensitive (DS) TB
The current recommended treatment for DS-TB involves a combination of four antibiotics isoniazid(INH), rifampicin(RIF), pyrazinamide(PZA) and ethambutol (EMB), which were all discovered nearly 60 year ago. This four drug cocktail should be administered for at least 6 months under directly observed treatment (DOT)to ensure high rates of treatment success and cure. The treatment involves two phases: the initial phase, which comprises administering the aforemention four drugs for two months, and the continuation phase treatment with INH and RIF for two months to kill the dormant bacteria.
The four drugs target M.tb via different mechanism of the action. Briefly , INH is a prodrug that upon activation inhibits the enoly-acyl carrier protein reductase, which is the drug enzyme in the Mas biosynthestic process. MAs are the primary mediators of the hydrophobic attributes and lack of permeability of the mycobacterial outer coating. RIF binds to the beta-subunit of the bacterial RNA polymerase and exerts its bactericidals activity by inhibiting the early steps of gene transcription. Like INH,PZA is a prodrug that gets activated after diffusing into the tb granuloma by the pyrazinamidase enzyme to pyrazinoic acid (POA), which is subsequently kills the M. tb bacillus inside the granuloma. The mode of action of PZA is still enigmatic. EMB is a bacteriostatic drug that inhibits the synthesis of arabinogalactan and lipoarabinomannan, two essential components of the mycobacterial cell wall, by targeting the three arabinosyltransferes EmbA, EmbB and EmbC.
Challenges to the global control of TB
Drug-resistance(DR) TB Crisis
The therapeutic approach for DR-TB and the prognosis thereof is significantly correlated to the resistance pattern; however, the clinical management of DR-TB is generally complicated. Multidrug-resistant TB (MDR-TB) is defined as resistance to INH and RIF, the two most powerful front-line anti-TB drugs. In 2021, there were an estimated 450,000 MDR-TB incident cases. The cure rates for MDR-TB are typically significantly lower than DS-TB. The 2019 WHO recommended second-line regimen for MDR-TB is an 18–20 months treatment protocol, contingent on the patient’s response to therapy. The MDR-TB medication regimen consists of at least four drugs in the intensive phase: three drugs from group A [linezolid, bedaquiline (BDQ) and moxifloxacin/levofloxacin] and one drug from group B (clofazimine, or terizidone/cycloserine). At least three of these drugs should be prescribed for the rest of the treatment (continuation phase) after BDQ is stopped. Two drugs in group B should be prescribed if only one or two drugs from group A are used. from group C [delamanid (DLM), streptomycin/amikacin, EMB, PZA, 4-aminosalicylic acid, imipenem, meropenem, ethionamide/prothionamide, high dose INH] should be added to the regimen.
TB and HIV Co-Infection.
HIV infection is considered the main predisposing risk factor for patients falling ill with M. tb, increasing the likelihood of disease progression into the active stage by 18-fold. TB is known to exacerbate the HIV infection and is considered the leading cause of death in HIV patients. In co-infected individuals, both pathogens have profound effects on the immune system, disarming the host’s immune responses and the decline of the immunological functions. The main interactions between TB and HIV antibiotics are correlated to RIF-induced elevated expression of hepatic cytochrome P450 (CYP) system. This induction of the CYP enzymes increases the metabolism of several HIV co-medications, such as protease inhibitors, and accordingly decreases their therapeutic concentrations.
The Coronavirus 2019 (COVID-19) Pandemic and TB
TB has long been the world’s leading cause of death from a single infectious disease (surpassing HIV/AIDS since 2007) until the COVID-19 pandemic . Indeed, according to the WHO, COVID-19 caused the deaths of more than 6.7 million people worldwide so far since the start of the pandemic. Indeed, the WHO indicated that, in 2020 and 2021, an increase in TB deaths was seen for the first time in more than a decade, reversing years of progress made up to 2019. The estimated surge in TB deaths globally was mostly located in four countries, namely, India, Indonesia, the Philippines and Myanmar . The WHO is expecting TB to regain the lead as the deadliest single infectious disease in the near future, replacing COVID-19, which means that the global TB targets have been thrown off track.
Figure 4 Imaging tools for active TB disease.
a | Conventional chest X?ray. The image shows typical features of active pulmonary tuberculosis (TB) disease: a large cavity in the right upper lobe of the lung (arrow) with surrounding infiltrates or consolidation (owing to inflammation and oedema). An abnormal chest X?ray is suggestive of TB, but not confirmatory. b | High-resolution CT scan. Three-dimensional rendering using 18F-fluorodeoxyglucose (FDG) PET-CT scan of the posterior half of the thoracic cavity of a person who was newly diagnosed with bilateral pulmonary TB. The orange colour depicts FDG uptake in regions with abnormalities with standardized uptake values ranging from 5 to 9. A 1–2 cm air-filled cavity in the right upper lobe (arrow) is embedded within an area of nodular disease with intense uptake.
TB Drug Targets
Overview
In 1998, the complete genome sequencing of M. tb (approximately 4000 genes) was unveiled, which advanced our understanding of the molecular biology of the bacterium. Knowledge of the whole-genome M. tb sequence enabled researchers to identify a subset of genes that are essential in vitro and in vivo. This revelation in turn contributed to the discovery of new targets for novel compounds via identifying the mutated genes of the strains resistant to these compounds. The gene knockdown techniques, whereby the gene of a specific target is depleted, has also facilitated the validation process of several M. tb drug targets. The genome-derived target-based approach (targetto- drug) involves the identification of a specific cellular target in advance but without giving any information about its druggability (drug penetration or efflux). In addition, several inhibitors, which were identified against essential targets, were lacking drug-like properties. Therefore, no anti-TB drug has emerged from this strategy to date. Indeed, it has been a difficult conundrum to translate a good bacterial enzyme inhibition into a potent whole-cell M. tb inhibitory activity because of the difficulty to penetrate the highly impermeable waxy cell wall of M. tb.
Current Hot Targets in M. tb Drug Discovery and Their Corresponding TB Drug Candidates
DNA gyrase is a highly conserved type II topoisomerase enzyme that is essential for
DNA transcription, replication and recombination in M. tb Therefore, inhibiting DNA gyrase results in impaired DNA replication and permanent double strands breaks, which leads to cytotoxic accumulation of cleaved double-strand DNA fragments, inducing bacterial death. The GyrA subunit carries the breakage-reunion active site and is a clinically validated drug target of the fluoroquinolone family of antibiotics, such as moxifloxacin. On the other hand, the GyrB subunit (ATPase) promotes ATP hydrolysis and has been relatively less exploited, thereby representing a new avenue for tackling M. tb strains that are resistant to fluoroquinolones. Indeed, various chemical entities have been developed as GyrB inhibitors, showing potent activity against DR-TB. In particular, a novel class of aminobenzimidazoles was found to target the ATPase subunit, which upon further optimisation led to the discovery of SPR720 (VXc-486).
QcrB
The cytochrome b subunit (QcrB) of the cytochrome bc1 complex has recently emerged as an interesting target in M. tb. The cytochrome bc1 complex is a key component of the respiratory electron transport chain required for ATP synthesis. Therefore, the inhibition of this complex disrupts the M. tb ability to generate energy. A phenotypic screening of a library encompassing more than 100,000 compounds as antimycobacterial agents led to the identification of imidazopyridine amides (IPAs) as a promising class that blocks the M. tb growth by targeting QcrB. An optimised IPA derivative Q203 showed potent growth inhibition against DS M. tb H37Rv strain (MIC50 = 2.7 nM) and numerous MDR and XDR M. tb clinical isolates in vitro (MIC90 < 0>
DprE1
Decaprenylphosphoryl-_-D-ribose 20-epimerase 1 (DprE1), also called decaprenylphosphoryl- -D-ribose oxidase, is a key enzyme implicated in the mycobacterial cell wall biosynthesis. In 2009, a ground-breaking report identified DprE1 as the target of a novel class of inhibitors, namely 1,3-benzothiazin-4-ones (BTZs), that were discovered in a phenotypic screening of a drug library. This new class of compounds is endowed with potent antimycobacterial activities, demonstrating bactericidal activities against M. tb in the nanomolar range. DprE1 is a flavoprotein that works in concert with decaprenylphosphoryl- D-2-keto erythro pentose reductase (DprE2) to generate an arabinose precursor that plays a fundamental role in the synthesis of the mycobacterial cell wall polysaccharides arabinogalactan and lipoarabinomannan. The extracytoplasmic localisation of DprE1 makes it more accessible to drugs that contribute to its vulnerability. It was demonstrated that inhibiting DprE1 abolishes the formation of DPA, thereby provoking cell lysis and mycobacterial death.
The fatty acyl-AMP ligase 32 (FAAL32 or FadD32), which is also called fatty acid degradation protein D32, and polyketide synthase 13 (Pks13) are crucial enzymes that actin concert with each other, playing pivotal roles in the biosynthetic machinery of Mas. MAs are the major integral lipid components of the exceptionally fortified waxy cell synthase II (FAS-II) systems, respectively. These two fatty acids chains get activated before the final condensation takes place.wall of M. tb and the primary mediators of hydrophobicity and impermeability thereof. Briefly, in the M. tb cytoplasm, the C24–C26 _-alkyl branch of the MAs and the C50–C60 meromycolate chain are generated from the fatty acid synthase I (FAS-I) and fatty acid.
CONCLUSION
TB continues to cause morbidity and mortality at alarming rates worldwide, especially in developing countries. The tubercle bacilli are typically constrained by granulomas in immunocompetent individuals, a lifelong standoff between the bacteria and the host’s immune system takes place (latent TB infection). This covert (asymptomatic) TB infection can recrudesce when the host immunity is impaired, which results in a high bacterial burden and the progression of the disease, culminating in clinical manifestations and TB transmission. Treating DS M. tb infections is usually attainable with the first-line anti-TB drug regimen. However, managing DR-TB infections is more challenging and less promising, leading to the continued relentlessness of the TB pandemic. In addition, the TB control efforts are generally hampered by the HIV co-infection, COVID-19, poor patient compliance and suboptimal treatment approaches in different parts of the world. Since the whole genome sequencing of M. tb (_4000 genes) was revealed, a multitude of small molecules with potent activities against both DS and DR M. tb strains were discovered, and their targets were identified and validated. Indeed, many scientists have been focusing their research efforts on newly identified M. tb drug targets, diverting from the traditional targets of the currently used TB antibiotics to bypass the DR issue. The most prominent drug targets that have recently been attracting attention involve GyrA/B, ATP synthase, QcrB, DprE1, FadD32, Pks13 and MmpL3.
REFERENCES
Shrushti Uchale*, Chaitali Ingawale, Sandhya Khomane, Rupali kharat, Kadambari Ghatpande, Pathophysiology And Treatment of Tuberculosis According to Who, Int. J. of Pharm. Sci., 2024, Vol 2, Issue 12, 1764-1773. https://doi.org/10.5281/zenodo.14441455
10.5281/zenodo.14441455
