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DR. AMIT KUMAR PANDEY, Ph.D. 

Assistant Professor
+91-129-3087322  (911292876322)
amitpandey [at] thsti [dot] res [dot] in
Posdoctoral Fellow, University of Massachusetts Medical School, Worcester, Massachusetts, USA
Posdoctoral Fellow, University of Nebraska-Lincoln, Nebraska, USA
Ph.D. Indian Veterinary Research Institute, Izatnagar, U.P., India
M.Sc. (Animal Biotechnology), National Dairy Research Institute, Karnal, India
B.V.Sc and A.H, Orissa University of Agriculture and Technology, Bhubaneswar, India

I.      Understanding the biology of Mycobacterial Persistence

Globally, a third of human population is infected with Mycobacterium tuberculosis(Mtb), the causative agent of tuberculosis. Mtb, being an obligate intracellular pathogen, has lived to co-evolve with human for centuries. Unlike actively infected individuals, latently infected population harbors pathogen for decades without showing any overt symptoms. This interesting phenotype is attributed to a slow growing, metabolically altered sub set of heterogeneous Mtb population called ‘persisters’. These persisters are refractory to anti-mycobacterial insult and can only be targeted by a strict regimen comprising of a combination of drugs for an unusually extended period. The protracted regimen triggers non-compliance and contributes towards increase in the frequency of cases involving MDR and XDR tuberculosis.

A.   Bacterial metabolism, physiology and persistence

The ability of the pathogen to “adapt” to host environment is key to its persistence. Understanding bacterial metabolism and physiology is important to decipher the mechanism and pathways critical for persistence of mycobacteria inside host. Since Mtb is an obligate intracellular pathogen, it depends on host for its nutritional requirements. Inside the host, Mtb subsists mainly on host derived fatty acid and cholesterol as a preferred carbon source. Although Mtb ingests cholesterol throughout the infection process, cholesterol becomes essential only during the later stage of chronic infection. Genetic and molecular understanding of cholesterol utilization, its mechanism and relevance would contribute significantly in designing novel intervention strategies in the treatment of tuberculosis. The knowledge acquired on the genes involved in uptake and metabolism of cholesterol in Mtb is very likely to generate new and more efficient drug targets.  The role of cholesterol metabolism in mycobacterial persistence would also be better understood. The ultimate goal will be to generate an interactome map of the regulatory pathways of cholesterol utilization in Mtb.

B.   Cholesterol catabolic pathways as therapeutics target 

Current tuberculosis treatment regimen involves multiple drugs for a prolong period. The duration could be from three months to two years depending on the type of infection. Prolong treatment leads to non-compliance and emergence of newer drug resistance strains. Shortening the therapy would go a long way in alleviating this problem. It is widely perceived that the major culprits are the so-called non-replicating and metabolically inactive “persister” population. The importance of cholesterol metabolism during the persistence stage of Mtb infection and its potential role in generation of persisters is very intriguing. In light of the above facts and hypothesis the focus of the current proposal is to screen for chemical inhibitors that specifically target these pathways. The long-range goal would be identify novel anti-tubercular drugs that specifically targets “persisters”. These novel compounds in combination with the standard frontline anti-tubercular drugs would significantly enhance the success rate in tuberculosis therapy.

C.  Iron regulation in Mtb and its implications on mycobacterial persistence

Although, iron is essential for most of the bacteria, excess of intracellular free iron is toxic. Failure to do so might lead to death either due to iron deficiency or toxicity.  Hence, the acquisition and storage of iron in bacteria is tightly regulated. Since iron deprivation is also one of the anti-microbial strategies that the host adopts, both pathogen and the host compete for the limited iron during infection. We for the first time have demonstrated that the Mtb transcription repressor protein sufRTBregulates the ISC operon and has a role in controlling the intracellular iron homeostasis in Mtb. Disruption of the iron homeostasis in ΔsufRTBdecreased the fitness of the mutant strain to grow inside mouse bone marrow-derived macrophages. The transcription repressor protein sufRTBwas also required for growth of Mtb under oxidative and nitrosative stress conditions. The enhanced biofilm production phenotype observed in ΔsufRTBis intriguing and suggests a role of intracellular iron homeostasis in the generation of biofilms in mycobacteria. Finally, we demonstrated that the sufRTBprotein mediated regulation of Fe homeostasis in Mtb is required for Mtb to persist inside the host.

D.   Screening and Identifying new molecular scaffolds targeting persisters

We in our lab have successfully identified proteins critical for the generation and maintenance of mycobacterial persisters. The aim of the proposal is to shorten the treatment regimen by identifying novel compounds that specifically target these proteins thereby significantly decreasing the frequency of generation of persisters during Mtb infection. 

II.      Live recombinant M bovisBCG (rBCG) strain as a better vaccine candidate against tuberculosis

Although, Mycobacterium bovis derived BCG is widely been used as vaccine to prevent tuberculosis, its effectiveness towards controlling tuberculosis is questionable. BCG is shown to be good at controlling severe forms of tuberculosis in infants, but fails to do so in case of pulmonary tuberculosis in adults. The challenge is to design a vaccine strain that is a) safe, b) immunogenic, c) longlasting and d) protects against tuberculosis caused by all strains.We in our lab are working on strategies to generate  rBCG strains that we believe will be more immunogenic  and hence would generate better and long lasting protective immune response against Mtb.  

III.      Host-Directed Therapies against tuberculosis

We are also interested in identifying and targeting critical host proteins that facilitates mycobacterial survival inside host. We are in the currently developing protocols for CRISPR-cas9 based strategies to delete specific genes for development of knockout cell lines. The plan is to design an unbiased approach towards screening and identifying host genes that modulate growth of Mtb inside cells. The long-term goal is to identify inhibitors against these genes to be used as drug against tuberculosis. 

 
  1. Iron homeostasis in Mycobacterium tuberculosisis is essential for persistence. Manitosh Pandey, Sakshi Talwar, Sutapa Bose and Amit Kumar Pandey*Scientific Reports (2018). https://www.nature.com/articles/s41598-018-35012-3
  2. Diphenyleneiodonium chloride (DPIC) displays broad-spectrum bactericidal activity. Manitosh Pandey, Alok Kumar Singh, Ritesh Thakare, Sakshi Talwar, Pratiksha Karaulia, Arunava Dasgupta, Sidharth Chopra, and Amit Kumar Pandey*Scientific Reports (2017). https://doi:10.1038/s41598-017-11575-5
  3. Biological evaluation of Diphenyleneiodonium chloride (DPIC) as a potential drug candidate for treatment of non-tuberculous mycobacterial infections. Alok Kumar Singh, Pratiksha Karaulia, Ritesh Thakare, Swetarka Das, Manitosh Pandey, Amit Kumar Pandey, Sidharth Chopra and Arunava Dasgutpa*Journal of Antimicrobial Chemotherapy (2017).https://doi.org/10.1093/jac/dkx277
  4. Rational design of drug-like compounds targeting Mycobacterium marinumMelF Protein. RennuDharra, Sakshi Talwar, Jeffery D. Cirillo, Amit Kumar Pandey, Mahesh Kulharia, and Parmod K. Mehta*. PLoS One (2017).12(9):e0183060. https://doi.org/10.1371/journal.pone.0183060
  5. Identification of lipid metabolism-targeting compounds active against drug-resistant M. tuberculosis (2016). Alok Kumar Singh, Pratiksha Karaulia, Pragya Yadav, Tadigoppula Narender, Suriya P Singh, Koneni V Sashidhara, Amit K Pandey, Sidharth Chopra, Arunava Dasgupta*. Journal of global antimicrobial resistance (2016). Dec 1:(7):26-27
  6. Systematic Analysis of Mycobacterial Acylation Reveals First Example of Acylation-mediated Regulation of Enzyme Activity of a Bacterial Phosphatase.Anshika Singhal, Gunjan Arora, Richa Virmani, Parijat Kundu, Tanya Khanna, Andaleeb Sajid, Richa Misra, Jayadev Joshi, Vikas Yadav, Sintu Samanta, Neeru Saini, Amit K. Pandey*, Sandhya S. Visweswariah, Christian Hentschker, Dorte Becher, Ulf Gerth*and Yogendra Singh*J Biol Chem. (2015) Oct 23;290(43):26218-34. doi: 10.1074/jbc. M115.687269. Epub 2015 Sep 8.*co-corresponding authors
  7. Identification of Ser/Thr kinase and forkhead associated domains in Mycobacterium ulcerans: characterization of novel association between protein kinase Q and MupFHA.Gunjan Arora, Andaleeb Sajid, Anshika Singhal, Jayadev Joshi, Richa Virmani, Meetu Gupta, Nupur Verma, Abhijit Maji, Richa Misra, Gregory Baronian, Amit K. Pandey, Virginie Molle, Yogendra Singh. PLoS Negl Trop Dis. (2014) Nov 20;8(11):e3315. doi: 10.1371/journal.pntd.0003315. eCollection 2014 Nov
  8. Cholesterol catabolism by Mycobacterium tuberculosis requires transcriptional and metabolic adaptations. Jennifer E.Griffin*, Amit K. Pandey*, Sarah A. Gilmore, Valerie Mizrahi, John D. McKinney, Carolyn R. Bertozzi and Christopher M. Sassetti.Chemistry and Biology. (2012) Feb: 24,19(2):218-27 *contributed equally to this work
  9. NOD2, RIP2 and IRF5 Play a Critical Role in the Type I Interferon Response to Mycobacterium Tuberculosis. Amit K. Pandey, Yibin Yang, Zhaozhao Jiang, Sarah M. Fortune, Francois Coulombe, Marcel A. Behr, Katherine A. Fitzgerald, Christopher M. Sassetti and Michelle A. Kelliher. PLoS Pathogen, (2009) Jul;5(7):e1000500. Epub 2009 July3. 
  10. Increase NOD2-mediated recognition of N-glycolylated muramyl dipeptide. Francois Coulombe, Maziar Divangahi, Frederic Veyrier, James Gleason, Yibin Yang, Michelle A. Kelliher, Amit K. Pandey, Christopher M. Sassetti, Michael B. Reed and Marcel A. Behr. Journal of Expt. Med. (2009). Aug 3;206(8):1709-16. Epub 2009 Jul 6.
  11. Phthiocerol dimycocerosate transport is required for resisting IFN-γ-independent immunityJeffrey P. Murry, Amit K. Pandey, Christopher M. Sassetti, and Eric J. Rubin. 2009Journal of Infectious Diseases. (2009). Sep 1;200(5):774-82. 
  12. Role of Cholesterol in Mycobacterium tuberculosisinfection. Maurine D Miner, Jennifer C Chang, Amit K Pandey, Christopher M Sassetti and David R Sherman. Indian Journal of Exp. Biol(2009). June; 47: 407-11.
  13. igrgenes and Mycobacterium tuberculosischolesterol metabolism. Jennifer C Chang, Maurine D Miner, Amit K Pandey, Wendy P Gill, Nada S Harik, Christopher M  Sassetti and David R Sherman. J. Bacteriol. (2009). Aug;191(16):5232-9. 
  14. Nitrile-inducible gene expression in mycobacteria. Amit K. Pandey, Sahadevan Raman, Rose Proff, Swati Joshi, Choong-Min Kang, Eric J. Rubin, Robert N. Husson and Christopher M. Sassetti. Tuberculosis(Edinb) (2009).Jan;89(1):12-16
  15. Mycobacterial persistence requires the utilization of host cholesterol.Amit K. Pandey and Christopher M SassettiProc Natl Acad Sci. U S A (2008). March18;105(11):4376-80. http://www.pnas.org/content/105/11/4376.full.pdf+html
  16. NOD2 pathway activation by MDP or Mycobacterium tuberculosisinfection involves the stable polyubiquitination of Rip2. Yibin Yang, Catherine Yin, Amit Pandey, Derek Abbott, Christopher Sassetti and Michelle A. Kelliher. J. Biol. Chem.(2007)Dec 14; 282(50):36223-9. 
  17. Characterization of mycobacterial virulence genes through genetic interaction mapping. Swati Joshi,Amit K. Pandey, Nicole Capite, Sarah M. Fortune, Eric J. Rubin, and Christopher M. Sassetti. Proc Natl Acad Sci. U S A.(2006). Aug 1; 103(31):11760-5. 
  18. Identification of Mycobacteriummarinummacrophage infection mutants. Parmod K. Mehta*, Amit K. Pandey*, Selvakumar Subbian, Mustapha M. Samarkandi, Suat L.G. Cirillo and Jeffery D. Cirillo. Microb Pathog.(2006). Apr;40(4):139-51. *contributed equally to this work.

Projects in the laboratory:

1.     We have identified novel pathways that regulate cholesterol mediated mycobacterial persistence and we are looking for highly motivated graduate students who are interested in understanding the biology of mycobacterial persistence. More specifically, the research focus will be to characterize and understand the roles of few identified Mtb genes in mycobacterial persistence.

2.     My lab will be interested in generating a carbon source independent in-vitro model of mycobacterial persistence. Briefly, since we have some preliminary understanding of how persisters are generated, we would like to utilize our understanding to rewire Mtb signaling by generating recombinant Mtb that mimic “persisters”. The strain thus generated would be characterized by identifying persister specific transcriptional and proteomic signatures. This model would than be used for screening antimicrobial compounds specifically targeting persisters.

3.     Since Mtb is an obligate intracellular pathogen, it has to adapt very quickly to intracellular niche and neutralize any host-mediated insults. We would also be interested in exploring the contribution of host in helping long-term survival of the pathogen inside the host. Briefly, the potential host targets genes will be knocked out by CRISPR and the role of specific host genes would be deciphered by analysing the growth of mycobacteria in these KO cell lines. 

  • Ramalingaswami Fellowship, 2010-11

  • ICAR-SRF Fellowship

  • ICAR JRF Fellowship


MR. MANITOSH PANDEY
SENIOR RESEARCH FELLOW

MS. SAKSHI TALWAR
PH.D STUDENT

MS. SHAIFALI TYAGI
PH.D STUDENT