scholarly journals Discovery of Novel Antituberculosis Compounds Using an Intra-Macrophage Assay

2021 ◽  
Author(s):  
◽  
Rekha Veliyayi Murikoli
Keyword(s):  
Current Treatment ◽  
Infection Model ◽  
World Population ◽  
Macrophage Infection ◽  
Human Immuno Deficiency Virus ◽  
Significant Toxicity ◽  
The World ◽  
Current Treatment Regimen ◽  
Set Up

<p>The causative agent of tuberculosis (TB) is Mycobacterium tuberculosis, which affects 2 billion of the world population and kills 1.8 million people annually. It is among the top three infectious killers in the world human immuno deficiency virus, TB and Malaria. Every year among 300-400 new cases of TB are reported in New Zealand according to a recent WHO 2008 report. The current treatment regimen for TB is very long and results in significant toxicity, development of resistant strains and is unable to eliminate the latent bacilli. The above reasons demonstrate the growing need of research for novel antimycobacterial compounds and novel targets for the treatment of TB. Many in vitro and biochemical screens are available for testing against different mycobacterium strains but none of these screens can be considered comprehensive. The reason for this can be the lack of resemblance of the in vitro screen model with the biological systems. Hence we chose the intra-macrophage infection screening model to look for novel antimycobacterial prodrugs which are not active in an in vitro screen but selectively active inside macrophage cell lines. We were successful in establishing and validating such an intra-macrophage infection model using the non-pathogenic M. smegmatis. The model was validated using common anti-tuberculosis drugs. A preliminary high throughput screen was then set up using a mini-library demo model, followed by screening with an actual Lopac synthetic library.</p>

scholarly journals Discovery of Novel Antituberculosis Compounds Using an Intra-Macrophage Assay

2021 ◽  
Author(s):  
◽  
Rekha Veliyayi Murikoli
Keyword(s):  
Current Treatment ◽  
Infection Model ◽  
World Population ◽  
Macrophage Infection ◽  
Human Immuno Deficiency Virus ◽  
Significant Toxicity ◽  
The World ◽  
Current Treatment Regimen ◽  
Set Up

<p>The causative agent of tuberculosis (TB) is Mycobacterium tuberculosis, which affects 2 billion of the world population and kills 1.8 million people annually. It is among the top three infectious killers in the world human immuno deficiency virus, TB and Malaria. Every year among 300-400 new cases of TB are reported in New Zealand according to a recent WHO 2008 report. The current treatment regimen for TB is very long and results in significant toxicity, development of resistant strains and is unable to eliminate the latent bacilli. The above reasons demonstrate the growing need of research for novel antimycobacterial compounds and novel targets for the treatment of TB. Many in vitro and biochemical screens are available for testing against different mycobacterium strains but none of these screens can be considered comprehensive. The reason for this can be the lack of resemblance of the in vitro screen model with the biological systems. Hence we chose the intra-macrophage infection screening model to look for novel antimycobacterial prodrugs which are not active in an in vitro screen but selectively active inside macrophage cell lines. We were successful in establishing and validating such an intra-macrophage infection model using the non-pathogenic M. smegmatis. The model was validated using common anti-tuberculosis drugs. A preliminary high throughput screen was then set up using a mini-library demo model, followed by screening with an actual Lopac synthetic library.</p>

In Vitro, In Silico and Ex Vivo Studies of Dihydroartemisinin Derivatives as Antitubercular Agents

2019 ◽  
Vol 19 (8) ◽  
pp. 633-644 ◽  
Author(s):  
Komal Kalani ◽  
Sarfaraz Alam ◽  
Vinita Chaturvedi ◽  
Shyam Singh ◽  
Feroz Khan ◽  
...  
Keyword(s):  
Bone Marrow ◽  
In Silico ◽  
Ex Vivo ◽  
Virulent Strain ◽  
Infection Model ◽  
Mouse Bone Marrow ◽  
Significant Activity ◽  
Macrophage Infection ◽  
In Silico Studies

Introduction: As a part of our drug discovery program for anti-tubercular agents, dihydroartemisinin (DHA-1) was screened against Mtb H37Rv, which showed moderate anti-tubercular activity (>25.0 µg/mL). These results prompted us to carry out the chemical transformation of DHA-1 into various derivatives and study their antitubercular potential. Materials and Methods: DHA-1 was semi-synthetically converted into four new acyl derivatives (DHA-1A – DHA-1D) and in-vitro evaluated for their anti-tubercular potential against Mycobacterium tuberculosis H37Rv virulent strain. The derivatives, DHA-1C (12-O-(4-nitro) benzoyl; MIC 12.5 µg/mL) and DHA-1D (12-O-chloro acetyl; MIC 3.12µg/mL) showed significant activity against the pathogen. Results: In silico studies of the most active derivative (DHA-1D) showed interaction with ARG448 inhibiting the mycobacterium enzymes. Additionally, it showed no cytotoxicity towards the Vero C1008 cells and Mouse bone marrow derived macrophages. Conclusion: DHA-1D killed 62% intracellular M. tuberculosis in Mouse bone marrow macrophage infection model. To the best of our knowledge, this is the first-ever report on the antitubercular potential of dihydroartemisinin and its derivatives. Since dihydroartemisinin is widely used as an antimalarial drug; these results may be of great help in anti-tubercular drug development from a very common, inexpensive, and non-toxic natural product.

In vitro evaluation of the anti-mycobacterial properties of the extracts of Momordica Charantia

2021 ◽  
Vol 16 (7) ◽  
pp. 15-22
Author(s):  
Paul Giftson ◽  
Jerrine Joseph ◽  
Revathy Kalyanasundaram ◽  
V. Ramesh Kumar ◽  
Wilson Aruni
Keyword(s):  
Medicinal Plants ◽  
Fungal Pathogens ◽  
Communicable Disease ◽  
Antimycobacterial Activity ◽  
Bitter Gourd ◽  
World Population ◽  
Methanol Extracts ◽  
Crude Extracts ◽  
The World

Tuberculosis (TB) is a communicable disease and remains one of the top 10 causes of death worldwide. One fourth of the world population is infected with TB at a risk of developing disease. The increase in the incidence of drug resistant TB around the world urges the need to develop a new candidate to fight against the disease. Plants were considered as the rich source of bioactive components to be used as potential drugs. Medicinal plants are used in pure as well as crude materials for their medicinal properties. Our research aims in identifying the phyto-molecules which have anti- tuberculosis property. Four medicinal plants namely, Acalyphaciliata (Kuppaimeni), Solanumtrilobatum (Thuthuvalai), Momordicacharantia (Bitter Gourd) and Sennaauriculata (Avaram) were chosen to evaluate their antimicrobial activity focusing on anti-tubercular activity. The methanol extracts of the medicinal plants showed significant inhibitory activity against bacterial and fungal pathogens. Sennaauriculata methanol extracts showed activity against S. aureus, E. coli, P. aeruginosa and C. albicans. In the screening of antimycobacterial activity done by LRP assay, among the plant extracts tested, the hexane crude extracts of Momordicacharantia (Bitter Gourd) showed 82.2% and 81.03% of inhibition against M. tuberculosis H37Rv at 500µg/ml and 250µg/ml concentration respectively. Similarly, the methanol crude extracts of Momordicacharantia showed 87.14% and 63.55% of inhibition at 500µg/ml and 250µg/ml concentration respectively.

scholarly journals Kinase Inhibitor Library Screening Identifies the Cancer Therapeutic Sorafenib and Structurally Similar Compounds as Strong Inhibitors of the Fungal Pathogen Histoplasma capsulatum

Antibiotics ◽  
2021 ◽  
Vol 10 (10) ◽  
pp. 1223
Author(s):  
Charlotte Berkes ◽  
Jimmy Franco ◽  
Maxx Lawson ◽  
Katelynn Brann ◽  
Jessica Mermelstein ◽  
...  
Keyword(s):  
Fungal Pathogen ◽  
Kinase Inhibitors ◽  
Histoplasma Capsulatum ◽  
Kinase Inhibitor ◽  
Systemic Disease ◽  
Antifungal Drugs ◽  
Infection Model ◽  
Macrophage Infection ◽  
Pathogenic Yeast

Histoplasma capsulatum is a dimorphic fungal pathogen endemic to the midwestern and southern United States. It causes mycoses ranging from subclinical respiratory infections to severe systemic disease, and is of particular concern for immunocompromised patients in endemic areas. Clinical management of histoplasmosis relies on protracted regimens of antifungal drugs whose effectiveness can be limited by toxicity. In this study, we hypothesize that conserved biochemical signaling pathways in the eukaryotic domain can be leveraged to repurpose kinase inhibitors as antifungal compounds. We conducted a screen of two kinase inhibitor libraries to identify compounds inhibiting the growth of Histoplasma capsulatum in the pathogenic yeast form. Our approach identified seven compounds with an elongated hydrophobic polyaromatic structure, five of which share a molecular motif including a urea unit linking a halogenated benzene ring and a para-substituted polyaromatic group. The top hits include the cancer therapeutic Sorafenib, which inhibits growth of Histoplasma in vitro and in a macrophage infection model with low host cell cytotoxicity. Our results reveal the possibility of repurposing Sorafenib or derivatives thereof as therapy for histoplasmosis, and suggest that repurposing of libraries developed for human cellular targets may be a fruitful source of antifungal discovery.

scholarly journals Sign Language Recognition using Smart Glove

2021 ◽  
Vol 9 (VII) ◽  
pp. 328-333
Author(s):  
Prof. Namrata Ghuse
Keyword(s):  
American Sign Language ◽  
Sign Language ◽  
Local Area ◽  
World Population ◽  
Language Recognition ◽  
Sign Language Recognition ◽  
The World ◽  
Impaired People ◽  
Indian Sign Language ◽  
Set Up

Gesture-based communication Recognition through innovation has been ignored idea even though an enormous local area can profit from it. There are more than 3% total population of the world who even can't speak or hear properly. Using hand Gesture-based communication, Every especially impaired people to communicate with each other or rest of world population. It is a way of correspondence for the rest of the world population who are neither speaking and hearing incompetency. Normal people even don't become intimate or close with sign language based communications. It's a reason become a gap between the especially impaired people & ordinary person. The previous systems of the project used to involve the concepts of image generation and emoji symbols. But the previous frameworks of a project are not affordable and not portable for the impaired person.The Main propaganda of a project has always been to interpret Indian Sign Language Standards and American Sign Language Standards and Convert gestures into voice and text, also assist the impaired person can interact with the other person from the remote location. This hand smart glove has been made with the set up with Gyroscope, Flex Sensor, ESP32 Microcontrollers/Micro bit, Accelerometer,25 LED Matrix Actuators/Output &, flex sensor, vibrator etc.

scholarly journals Zebrafish Embryo Model for Assessment of Drug Efficacy on Mycobacterial Persisters

2020 ◽  
Vol 64 (10) ◽  
Author(s):  
Susanna Commandeur ◽  
Nino Iakobachvili ◽  
Marion Sparrius ◽  
Mariam Mohamed Nur ◽  
Galina V. Mukamolova ◽  
...  
Keyword(s):  
Drug Screening ◽  
Zebrafish Embryo ◽  
Neutral Lipids ◽  
Drug Efficacy ◽  
Infection Model ◽  
Duration Of Treatment ◽  
Content Type ◽  
Set Up

ABSTRACT Tuberculosis continues to kill millions of people each year. The main difficulty in eradication of the disease is the prolonged duration of treatment, which takes at least 6 months. Persister cells have long been associated with failed treatment and disease relapse because of their phenotypical, though transient, tolerance to drugs. By targeting these persisters, the duration of treatment could be shortened, leading to improved tuberculosis treatment and a reduction in transmission. The unique in vivo environment drives the generation of persisters; however, appropriate in vivo mycobacterial persister models enabling optimized drug screening are lacking. To set up a persister infection model that is suitable for this, we infected zebrafish embryos with in vitro-starved Mycobacterium marinum. In vitro starvation resulted in a persister-like phenotype with the accumulation of stored neutral lipids and concomitant increased tolerance to ethambutol. However, these starved wild-type M. marinum organisms rapidly lost their persister phenotype in vivo. To prolong the persister phenotype in vivo, we subsequently generated and analyzed mutants lacking functional resuscitation-promoting factors (Rpfs). Interestingly, the ΔrpfAB mutant, lacking two Rpfs, established an infection in vivo, whereas a nutrient-starved ΔrpfAB mutant did maintain its persister phenotype in vivo. This mutant was, after nutrient starvation, also tolerant to ethambutol treatment in vivo, as would be expected for persisters. We propose that this zebrafish embryo model with ΔrpfAB mutant bacteria is a valuable addition for drug screening purposes and specifically screens to target mycobacterial persisters.

scholarly journals Fusidic Acid-Resistant Mutants of Salmonella enterica Serovar Typhimurium with Low Fitness In Vivo Are Defective in RpoS Induction

2003 ◽  
Vol 47 (12) ◽  
pp. 3743-3749 ◽  
Author(s):  
Mirjana Macvanin ◽  
Johanna Björkman ◽  
Sofia Eriksson ◽  
Mikael Rhen ◽  
Dan I. Andersson ◽  
...  
Keyword(s):  
Protein Synthesis ◽  
Fusidic Acid ◽  
Salmonella Enterica ◽  
Antimicrobial Agents ◽  
Relative Fitness ◽  
Infection Model ◽  
Macrophage Infection ◽  
Serovar Typhimurium

ABSTRACT Mutants of Salmonella enterica serovar Typhimurium resistant to fusidic acid (Fusr) have mutations in fusA, the gene encoding translation elongation factor G (EF-G). Most Fusr mutants have reduced fitness in vitro and in vivo, in part explained by mutant EF-G slowing the rate of protein synthesis and growth. However, some Fusr mutants with normal rates of protein synthesis still suffer from reduced fitness in vivo. As shown here, Fusr mutants could be similarly ranked in their relative fitness in mouse infection models, in a macrophage infection model, in their relative hypersensitivity to hydrogen peroxide in vivo and in vitro, and in the amount of RpoS production induced upon entry into the stationary phase. We identify a reduced ability to induce production of RpoS (σs) as a defect associated with Fusr strains. Because RpoS is a regulator of the general stress response, and an important virulence factor in Salmonella, an inability to produce RpoS in appropriate amounts can explain the low fitness of Fusr strains in vivo. The unfit Fusr mutants also produce reduced levels of the regulatory molecule ppGpp in response to starvation. Because ppGpp is a positive regulator of RpoS production, we suggest that a possible cause of the reduced levels of RpoS is the reduction in ppGpp production associated with mutant EF-G. The low fitness of Fusr mutants in vivo suggests that drugs that can alter the levels of global regulators of gene expression deserve attention as potential antimicrobial agents.

scholarly journals The anti-tubercular activity of simvastatin is mediated by cholesterol-driven autophagy via the AMPK-mTORC1-TFEB axis

2020 ◽  
Vol 61 (12) ◽  
pp. 1617-1628
Author(s):  
Natalie Bruiners ◽  
Noton K. Dutta ◽  
Valentina Guerrini ◽  
Hugh Salamon ◽  
Ken D. Yamaguchi ◽  
...  
Keyword(s):  
Molecular Mechanisms ◽  
Nuclear Translocation ◽  
Mevalonate Pathway ◽  
Biological Effects ◽  
Cholesterol Biosynthesis ◽  
Cholesterol Homeostasis ◽  
Infection Model ◽  
Protein Prenylation ◽  
Macrophage Infection

The rise of drug-resistant tuberculosis poses a major risk to public health. Statins, which inhibit both cholesterol biosynthesis and protein prenylation branches of the mevalonate pathway, increase anti-tubercular antibiotic efficacy in animal models. However, the underlying molecular mechanisms are unknown. In this study, we used an in vitro macrophage infection model to investigate simvastatin’s anti-tubercular activity by systematically inhibiting each branch of the mevalonate pathway and evaluating the effects of the branch-specific inhibitors on mycobacterial growth. The anti-tubercular activity of simvastatin used at clinically relevant doses specifically targeted the cholesterol biosynthetic branch rather than the prenylation branches of the mevalonate pathway. Using Western blot analysis and AMP/ATP measurements, we found that simvastatin treatment blocked activation of mechanistic target of rapamycin complex 1 (mTORC1), activated AMP-activated protein kinase (AMPK) through increased intracellular AMP:ATP ratios, and favored nuclear translocation of transcription factor EB (TFEB). These mechanisms all induce autophagy, which is anti-mycobacterial. The biological effects of simvastatin on the AMPK-mTORC1-TFEB-autophagy axis were reversed by adding exogenous cholesterol to the cells. Our data demonstrate that the anti-tubercular activity of simvastatin requires inhibiting cholesterol biosynthesis, reveal novel links between cholesterol homeostasis, the AMPK-mTORC1-TFEB axis, and Mycobacterium tuberculosis infection control, and uncover new anti-tubercular therapy targets.

scholarly journals Antibiotic Tolerance: Distinguishing between Classical Resistance and Persistence in a Macrophage Infection Model

Fine Focus ◽  
2018 ◽  
Vol 4 (1) ◽  
pp. 117-129
Author(s):  
Crystal Collins ◽  
Kara Mosovsky
Keyword(s):  
Burkholderia Pseudomallei ◽  
Infection Model ◽  
Intracellular Pathogen ◽  
Macrophage Infection ◽  
Persister Cells ◽  
Antibiotic Resistant ◽  
Related Organism ◽  
Infection Models ◽  
High Doses

Burkholderia pseudomallei is a Gram-negative bacillus and facultative intracellular pathogen. It causes the disease melioidosis, which is a potentially fatal human disease found throughout the world but particularly in Southeast Asia and Northern Australia. B. pseudomallei is inherently antibiotic resistant and therefore new therapies are needed to combat this pathogen. Previous studies with the related organism Burkholderia thailandensis have shown that the antibiotic ceftazidime does not eliminate all bacteria in an in vitro macrophage model, and the remaining bacteria could still pose a health threat to a potential host. Due to their survival in the presence of antibiotics, we hypothesized that the remaining bacteria were one of two types of antibiotic tolerant cells: classically antibiotic resistant cells or persister cells. To test our hypothesis we isolated the bacteria that had survived ceftazidime treatment in the macrophage infection model and performed additional in vitro experiments to show that the surviving bacteria are neither antibiotic resistant nor persister cells. Instead, they are still susceptible to high doses (200 μg/ml) of the antibiotic over a period of 48 hours (p<0.001). We believe the bacteria survive exposure to the antibiotic during the macrophage infection because of their ability to move between intracellular and extracellular compartments, thus avoiding the antibiotic and its deadly effects. Our results provide evidence to suggest that intracellular pathogens, through movement between intracellular and extracellular compartments, may be protected from the effects of antibiotics in similar macrophage infection models.

scholarly journals Candida albicans PEP12 Is Required for Biofilm Integrity and In Vivo Virulence

2009 ◽  
Vol 9 (2) ◽  
pp. 266-277 ◽  
Author(s):  
Suresh K. A. Palanisamy ◽  
Melissa A. Ramirez ◽  
Michael Lorenz ◽  
Samuel A. Lee
Keyword(s):  
Candida Albicans ◽  
Lipolytic Activity ◽  
Infection Model ◽  
Temperature Sensitive ◽  
Null Mutant ◽  
Macrophage Infection ◽  
Content Type ◽  
Vacuolar Acidification

ABSTRACT To investigate the role of the prevacuolar secretion pathway in biofilm formation and virulence in Candida albicans, we cloned and analyzed the C. albicans homolog of the Saccharomyces cerevisiae prevacuolar trafficking gene PEP12. C. albicans PEP12 encodes a deduced t-SNARE that is 28% identical to S. cerevisiae Pep12p, and plasmids bearing C. albicans PEP12 complemented the abnormal vacuolar morphology and temperature-sensitive growth of an S. cerevisiae pep12 null mutant. The C. albicans pep12 Δ null mutant was defective in endocytosis and vacuolar acidification and accumulated 40- to 60-nm cytoplasmic vesicles near the plasma membrane. Secretory defects included increased extracellular proteolytic activity and absent lipolytic activity. The pep12Δ null mutant was more sensitive to cell wall stresses and antifungal agents than the isogenic complemented strain or the control strain DAY185. Notably, the biofilm formed by the pep12Δ mutant was reduced in overall mass and fragmented completely upon the slightest disturbance. The pep12Δ mutant was markedly reduced in virulence in an in vitro macrophage infection model and an in vivo mouse model of disseminated candidiasis. These results suggest that C. albicans PEP12 plays a key role in biofilm integrity and in vivo virulence.

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