Hemanta K Majumder

Indian Institute of Chemical Biology, Kolkata, Bengal, India

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Publications (78)316.48 Total impact

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    ABSTRACT: In parasites, ATP binding cassette (ABC) transporters represent an important family of proteins related to drug resistance and other biological activities. Resistance of leishmanial parasites to therapeutic drugs continues to escalate in developing countries and in many instances it is due to overexpressed ABC efflux pumps. Progressively adapted baicalein (BLN)resistant parasites (pB25R) show overexpression of a novel ABC transporter, which was classified as ABCC2 or LdMrp2. The protein is primarily localized in the flagellar pocket region and in internal vesicles. Overexpressed LdABCC2 confers substantial BLN resistance to the parasites by rapid drug efflux. The BLN-resistant promastigotes when transformed into amastigotes in macrophage cells cannot be cured by treatment of macrophages with BLN. Amastigotes resistance is concomitant to the over expression of macrophage multidrug resistance protein 2 (MRP2) transporter. Reporter analysis and site directed mutagenesis assays demonstrate that antioxidant response element (ARE) 1 is activated upon infection. The expression of this phase II detoxifying gene is regulated by Nrf2 (NFE2 related factor 2) mediated antioxidant response element (ARE) activation. In view of the fact that the signaling pathway of phospho inositol 3 kinase controls microfilament rearrangement and translocation of actin associated proteins, the current study correlates with intricate pathway of PI3 kinase mediated nuclear translocation of Nrf2 which activates Mrp2 expression in macrophages upon infection by the parasites. In contrast, phalloidin, an agent that prevents actin filaments from depolymerization, inhibits Nrf2 translocation and Mrp2 activation by pB25R infection. Taken together, these results provide an insight into the mechanisms by which resistant clinical isolates of Leishmania donovani induce intracellular events relevant to drug resistance.
    Journal of Biological Chemistry 04/2014; · 4.65 Impact Factor
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    ABSTRACT: Poly(ADP-ribose) polymerases (PARP) attach poly(ADP-ribose) (PAR) chains to various proteins including themselves and chromatin. Topoisomerase I (Top1) regulates DNA supercoiling and is the target of camptothecin and indenoisoquinoline anticancer drugs, as it forms Top1 cleavage complexes (Top1cc) that are trapped by the drugs. Endogenous and carcinogenic DNA lesions can also trap Top1cc. Tyrosyl-DNA phosphodiesterase 1 (TDP1), a key repair enzyme for trapped Top1cc, hydrolyzes the phosphodiester bond between the DNA 3'-end and the Top1 tyrosyl moiety. Alternative repair pathways for Top1cc involve endonuclease cleavage. However, it is unknown what determines the choice between TDP1 and the endonuclease repair pathways. Here we show that PARP1 plays a critical role in this process. By generating TDP1 and PARP1 double-knockout lymphoma chicken DT40 cells, we demonstrate that TDP1 and PARP1 are epistatic for the repair of Top1cc. The N-terminal domain of TDP1 directly binds the C-terminal domain of PARP1, and TDP1 is PARylated by PARP1. PARylation stabilizes TDP1 together with SUMOylation of TDP1. TDP1 PARylation enhances its recruitment to DNA damage sites without interfering with TDP1 catalytic activity. TDP1-PARP1 complexes, in turn recruit X-ray repair cross-complementing protein 1 (XRCC1). This work identifies PARP1 as a key component driving the repair of trapped Top1cc by TDP1.
    Nucleic Acids Research 02/2014; · 8.28 Impact Factor
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    ABSTRACT: The unicellular organism Leishmania undergoes apoptosis-like cell death in response to external stress or exposure to antileishmanial agents. Here we showed that, triterpenoid disuccinyl betulin (DiSB), a potent topoisomerase IB inhibitor induced parasitic cell death by generating oxidative stress. The characteristic feature of the death process resembled programmed cell death (PCD) in higher eukaryotes. Generation of reactive oxygen species (ROS) followed by depolarization of mitochondrial membrane potential (ΔΨm) caused loss of ATP production in Leishmania parasites. This further gave positive feedback to produce high amount of ROS, which in turn caused oxidative DNA lesions and genomic DNA fragmentation. Treatment of promastigotes with DiSB induced high expression of metacaspase protein that led to cell death of this unicellular organism. The PCD was insensitive to zVAD-fmk, suggesting that the death process was not associated with activation of caspases. DiSB treatment translocated LdEndoG (Leishmania donovani Endonuclease G) from mitochondria to nucleus which was responsible for DNA degradation process. Conditional antisense knockdown of LdMC (Leishmania donovani Metacaspase) as well as EndoG subverted death of the parasite and rescued cell cycle arrest in G1 phase. The present study on effectors molecules associated with PCD pathway of the parasite should not only help to manifest the mechanisms of PCD but also could be exploited in antileishmanial chemotherapy.
    Antimicrobial Agents and Chemotherapy 01/2014; · 4.57 Impact Factor
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    ABSTRACT: Lignans are diphenyl propanoids with vast range of biological activities. The present study provides an important insight into the anti-leishmanial activities of two lignan glycosides viz. lyoniside and saracoside. These compounds inhibit catalytic activities of topoisomerase IB (LdTopIB) of Leishmania donovani in non-competitive manner and stabilize the LdTopIB mediated cleavage complex formation both in-vitro and in Leishmania promastigotes and subsequently inhibit the religation of cleaved strand. These two compounds not only poison LdTopIB but also can interact with the free enzyme LdTopIB. We have also shown that lyoniside and saracoside are cytotoxic to promastigotes and intracellular amastigotes. The protein-DNA complex formation leads to double strand breaks in DNA which ultimately triggers apoptosis-like cell death in the parasitie. Along with their cytotoxicity towards sodium antimony gluconate (SAG) sensitive AG83 strain, their ability to kill SAG resistant GE1 strain makes these two compounds potential anti-leishmanial candidates. Not only they effectively kill L. donovani amastigotes inside macrophages in vitro, lyoniside and saracoside demonstrated strong anti-leishmanial efficacies in BALB/c mice model of leishmaniasis. Treatment with these lignan glycosides produce nitric oxide and reactive oxygen species which result in almost complete clearance of the liver and splenic parasite burden. These compounds do not inhibit human topoisomerase IB upto 200μM concentrations and had poor cytotoxic effect on uninfected cultured murine peritoneal macrophages upto 100μM concentrations. Taken together it can be concluded that these compounds can be developed into excellent therapeutic agent against deadly disease leishmaniasis.
    Biochemical pharmacology 10/2013; · 4.25 Impact Factor
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    ABSTRACT: Abstract For the development of potent novel antileishmanial agents, 3,3'-(arylmethylene)bis(2-hydroxynaphthalene-1,4 dione) derivatives were synthesized from lawsone and evaluated for cytotoxicity on Leishmania donovani promastigotes as well as on leishmanial DNA topoisomerase-I. Enzyme inhibition studies were conducted with simultaneous and preincubation conditions. Total inhibition is compared to camptothecin (CPT), which was taken as positive control on both the systems of enzyme inhibition. The range of activity varied from 37.5 to 70 µM in simultaneous assay and 13-16 µM in preincubation assay. Furthermore, when evaluated against L. donovani promastigotes, the synthesized compounds exhibited the activity ranging from 2 to 14 µM. The results revealed that all the compounds exhibit promising antileishmanial activity.
    Journal of Enzyme Inhibition and Medicinal Chemistry 03/2013; · 1.50 Impact Factor
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    ABSTRACT: Niranthin, a lignan isolated from the aerial parts of the plant Phyllanthus amarus, exhibits a wide spectrum of pharmacological activities. In the present study, we have shown for the first time that niranthin is a potent anti-leishmanial agent. The compound induces topoisomerase I-mediated DNA-protein adduct formation inside Leishmania cells and triggers apoptosis by activation of cellular nucleases. We also show that niranthin inhibits the relaxation activity of heterodimeric type IB topoisomerase of L. donovani and acts as a non-competitive inhibitor interacting with both subunits of the enzyme. Niranthin interacts with DNA-protein binary complexes and thus stabilizes the 'cleavable complex' formation and subsequently inhibits the religation of cleaved strand. The compound inhibits the proliferation of Leishmania amastigotes in infected cultured murine macrophages with limited cytotoxicity to the host cells and is effective against antimony-resistant Leishmania parasites by modulating upregulated P-glycoprotein on host macrophages. Importantly, besides its in vitro efficacy, niranthin treatment leads to a switch from a Th2- to a Th1-type immune response in infected BALB/c mice. The immune response causes production of nitric oxide, which results in almost complete clearance of the liver and splenic parasite burden after intraperitoneal or intramuscular administration of the drug. These findings can be exploited to develop niranthin as a new drug candidate against drug-resistant leishmaniasis.
    EMBO Molecular Medicine 10/2012; 4(10):1126-43. · 7.80 Impact Factor
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    ABSTRACT: Chemical investigation of the stem bark of Saraca indica has resulted in the isolation of a new lignan glycoside, saracoside, along with four known lignan glycosides lyoniside, icariside E3, (+)5'-methoxyisolarciresinol-9'-O-beta-D-glucopyranoside and nudiposide, and a phenolic glucopyranoside, 3,4,5-trimethoxyphenyl-beta-D-glucopyranoside, which has been isolated for the first time from this species. The isolated lignan glycosides exhibit potent DNA topoisomerase IB inhibitory activity.
    Natural product communications 06/2012; 7(6):767-9. · 0.96 Impact Factor
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    ABSTRACT: Toward developing antileishmanial agents with mode of action targeted to DNA topoisomerases of Leishmania donovani, we have synthesized a large number of derivatives of betulin. The compound, a natural triterpene isolated from the cork layer of Betula spp. plants exhibits several pharmacological properties. Three compounds (disuccinyl betulin, diglutaryl dihydrobetulin, and disuccinyl dihydrobetulin) inhibit growth of the parasite as well as relaxation activity of the enzyme type IB topoisomerase [Leishmania donovani topoisomerase I (LdTOP1LS)] of the parasite. Mechanistic studies suggest that these compounds interact with the enzyme in a reversible manner. The stoichiometry of these compounds binding to LdTOP1LS is 1:1 (mole/mole) with a dissociation constant on the order of ∼10(-6) M. Unlike CPT, these compounds do not stabilize the cleavage complex; rather, they abrogate the covalent complex formation. In processive mode of relaxation assay condition, these compounds slow down the strand rotation event, which ultimately affects the relaxation of supercoiled DNA. It is noteworthy that these compounds reduce the intracellular parasite burden in macrophages infected with wild-type L. donovani as well as with sodium antimony gluconate resistant parasite (GE1). Taken together, our data suggest that these betulin derivatives can be exploited as potential drug candidates against threatening drug resistant leishmaniasis.
    Molecular Pharmacology 10/2011; 80(4):694. · 4.41 Impact Factor
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    ABSTRACT: Toward developing antileishmanial agents with mode of action targeted to DNA topoisomerases of Leishmania donovani, we have synthesized a large number of derivatives of betulin. The compound, a natural triterpene isolated from the cork layer of Betula spp. plants exhibits several pharmacological properties. Three compounds (disuccinyl betulin, diglutaryl dihydrobetulin, and disuccinyl dihydrobetulin) inhibit growth of the parasite as well as relaxation activity of the enzyme type IB topoisomerase [Leishmania donovani topoisomerase I (LdTOP1LS)] of the parasite. Mechanistic studies suggest that these compounds interact with the enzyme in a reversible manner. The stoichiometry of these compounds binding to LdTOP1LS is 1:1 (mole/mole) with a dissociation constant on the order of ∼10(-6) M. Unlike CPT, these compounds do not stabilize the cleavage complex; rather, they abrogate the covalent complex formation. In processive mode of relaxation assay condition, these compounds slow down the strand rotation event, which ultimately affects the relaxation of supercoiled DNA. It is noteworthy that these compounds reduce the intracellular parasite burden in macrophages infected with wild-type L. donovani as well as with sodium antimony gluconate resistant parasite (GE1). Taken together, our data suggest that these betulin derivatives can be exploited as potential drug candidates against threatening drug resistant leishmaniasis.
    Molecular pharmacology 07/2011; 80(4):694-703. · 4.53 Impact Factor
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    Bijoylaxmi Banerjee, Nilkantha Sen, Hemanta K Majumder
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    ABSTRACT: DNA topoisomerases of kinetoplastids represent a family of DNA processing enzymes that essentially solve the topological problems not only in nuclear DNA but also in kinetoplast DNA. We have, for the first time, identified a Leishmania donovani homologue of bacterial and eukaryotic IA type of topoisomerase III protein and termed as LdTopIIIβ. Complementation study of wild-type and mutant LdTopIIIβ with slow-growing topoisomerase III mutant yeast S. cerevisiae revealed the functional conservation of the leishmanial counterpart of topoisomerase IIIβ protein, the 327 tyrosine being the active site amino acid. A C-terminal deletion construct of LdTopIIIβ could not suppress the slow-growth phenotype of mutant yeast, indicating the requirement of C-terminal region for the enzyme function in vivo.LdTopIIIβ localized inside the nucleus and kinetoplast of the parasite. Taken together, our study indicates functional conservation and possible role of LdTopIIIβ in parasite DNA processing.
    Enzyme research. 01/2011; 2011:230542.
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    ABSTRACT: Leishmania donovani are the causative agents of visceral leishmaniasis worldwide. Lack of vaccines and emergence of drug resistance warrants the need for improved drug therapy and newer therapeutic intervention strategies against leishmaniasis. In the present study, we have investigated the effect of the natural indoloquinoline alkaloid cryptolepine on L. donovani AG83 promastigotes. Our results show that cryptolepine induces cellular dysfunction in L. donovani promastigotes, which leads to the death of this unicellular parasite. Interestingly, our study suggest that cryptolepine-induced cell death of L. donovani is counteracted by initial autophagic features elicited by the cells. For the first time, we show that autophagy serves as a survival mechanism in response to cryptolepine treatment in L. donovani promastigotes and inhibition of autophagy causes an early increase in the amount of cell death. This study can be exploited for designing better drugs and better therapeutic strategies against leishmaniasis in future.
    Molecular biology international. 01/2011; 2011:187850.
  • Molecular Biology International. 01/2011; doi:10.4061/2011/187850.
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    Hemanta K Majumder, Wanderley de Souza, Kwang Poo Chang
    Molecular biology international. 01/2011; 2011:185413.
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    ABSTRACT: The development of 3, 3'-diindolyl methane (DIM) resistant parasite Leishmania donovani (LdDR50) by adaptation with increasing concentrations of the drug generates random mutations in the large and small subunits of heterodimeric DNA topoisomerase I of Leishmania (LdTOP1LS). Mutation of large subunit of LdTOP1LS at F270L is responsible for resistance to DIM up to 50 µM concentration. In search of compounds that inhibit the growth of the DIM resistant parasite and inhibit the catalytic activity of mutated topoisomerase I (F270L), we have prepared three derivatives of DIM namely DPDIM (2,2'-diphenyl 3,3'-diindolyl methane), DMDIM (2,2'-dimethyl 3,3'-diindolyl methane) and DMODIM (5,5'-dimethoxy 3,3'-diindolyl methane) from parent compound DIM. All the compounds inhibit the growth of DIM resistant parasites, induce DNA fragmentation and stabilize topo1-DNA cleavable complex with the wild type and mutant enzyme. The results suggest that the three derivatives of DIM can act as promising lead molecules for the generation of new anti-leishmanial agents.
    PLoS ONE 01/2011; 6(12):e28493. · 3.73 Impact Factor
  • The Open Antimicrobial Agents Journal 01/2011; 3:23.
  • Nucleic Acids Research 01/2011; 39:3295. · 8.28 Impact Factor
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    ABSTRACT: Most type IB topoisomerases do not require ATP and Mg(2+) for activity. However, as shown previously for vaccinia topoisomerase I, we demonstrate that ATP stimulates the relaxation activity of the unusual heterodimeric type IB topoisomerase from Leishmania donovani (LdTOP1L/S) in the absence of Mg(2+). The stimulation is independent of ATP hydrolysis but requires salt as a co-activator. ATP binds to LdTOP1L/S and increases its rate of strand rotation. Docking studies indicate that the amino acid residues His93, Tyr95, Arg188 and Arg190 of the large subunit may be involved in ATP binding. Site directed mutagenesis of these four residues individually to alanine and subsequent relaxation assays reveal that the R190A mutant topoisomerase is unable to exhibit ATP-mediated stimulation in the absence of Mg(2+). However, the ATP-independent relaxation activities of all the four mutant enzymes remain unaffected. Additionally, we provide evidence that ATP binds LdTOP1L/S and modulates the activity of the otherwise ATP-independent enzyme. This study establishes ATP as an activator of LdTOP1L/S in the absence of Mg(2+).
    Nucleic Acids Research 12/2010; 39(8):3295-309. · 8.28 Impact Factor
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    ABSTRACT: Tyrosyl DNA phosphodiesterase 1 (Tdp1) is a member of phospholipase D superfamily, which cleaves a broad range of 3'-DNA adducts, the best characterized of which is the phosphodiester bond formed between DNA and topoisomerase IB. This study describes cloning and functional characterization of the enzyme, termed as LdTdp1 in the kinetoplastid parasite Leishmania donovani. Sequence analysis confirmed conservation of the active site motifs typical for all Tdp1 proteins. LdTdp1 activity was detected in the parasite nucleus as well as in the kinetoplast. The enzyme harbours a nuclear localization signal at its C-terminus. Overexpression of the active enzyme protected the parasites against topoisomerase IB inhibitor camptothecin (CPT) and oxidative agent H(2)O(2)-mediated cytotoxicity and its downregulation rendered the parasites hypersensitive to CPT. Trapping of mutant LdTdp1 on DNA takes place following CPT treatment in L. donovani cells. The expression level and associated activity of LdTdp1 were found to be higher in CPT-resistant L. donovani parasites. Altogether, this is the first report of Tdp1 from the kinetoplastid parasite L. donovani, which actively participates in topoisomerase I-mediated DNA damage repair process and thereby counteracts the cytotoxic effect of topoisomerase I inhibitors.
    Molecular Microbiology 10/2010; 78(1):119-37. · 4.96 Impact Factor
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    ABSTRACT: The geometrical optimization of (3,3')-diindolylmethane (DIM), an inhibitor of the bisubunit enzyme topoisomerase I from Leishmania donovani, a pathogenic protozoan parasite, mostly diffused in developing countries, has been carried out through quantum mechanical calculation. Using first-principle DFT restrained geometrical optimization, a potential energy surface has been constructed to identify a set of local minimum energy conformations of DIM. Starting from these conformations, the experimental UV-vis absorption spectrum in aqueous solution has been reproduced through TD-DFT calculations. A molecular mechanics classical force-field has been also parametrized and tested, verifying the correct coherence between the canonical ensemble obtained from molecular dynamics simulation and the potential energy surface calculation. The force field has been used to elucidate the interaction of DIM with a 22 bp DNA double strand. The best docked DIM-DNA complexes display a binding energy pretty similar to the experimental energy and are all located in the DNA minor groove, strongly suggesting that DIM is a minor groove binder.
    The Journal of Physical Chemistry A 07/2010; 114(26):7121-6. · 2.77 Impact Factor
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    ABSTRACT: Background and purpose:  New antileishmanials from natural products are urgently needed due to the emergence of drug resistance complicated by severe cytotoxic effects. 16α-Hydroxycleroda-3,13 (14)Z-dien-15,16-olide (Compound 1) from Polyalthia longifolia was found to be a potential antileishmanial and non-cytotoxic, as evidenced by long-term survival (>6 months) of treated animals. This prompted us to determine its target and, using molecular modelling, identify the interactions responsible for its specific antileishmanial activity.Experimental approach: In vitro activity of compound was assessed using intracellular transgenic green fluorescent protein-stably expressed Leishmania donovani parasites. In vivo activity and survival of animals post-treatment were evaluated in L. donovani-infected hamsters. Known property of clerodane diterpenes as potent human DNA topoisomerase inhibitors led us to evaluate the inhibition of recombinant L. donovani topoisomerase I using relaxation assay. Mode of cell death induced by Compound 1 was assessed by phosphotidylserine exposure post-treatment. Molecular modelling studies were conducted with DNA topoisomerase I to identify the binding interactions responsible for its activity.Key results:  Bioassay-guided fractionation led to isolation of Compound 1 as a non-cytotoxic, orally active antileishmanial. Compound 1 inhibited recombinant DNA topoisomerase I which, ultimately, induced apoptosis. Molecular docking studies indicated that five strong hydrogen-bonding interactions and hydrophobic interactions of Compound 1 with L. donovani DNA-topoisomerase are responsible for its antileishmanial activity.Conclusions and implications:  The data reveal Compound 1 is a potent and safe antileishmanial. The study further exploited the structural determinants responsible for its non-cytotoxic and potent activity, to raise the feasibility of specifically targeting the target enzyme responsible for its activity through rational drug design.
    British Journal of Pharmacology 02/2010; 159(5):1143 - 1150. · 5.07 Impact Factor

Publication Stats

1k Citations
316.48 Total Impact Points

Institutions

  • 1991–2011
    • Indian Institute of Chemical Biology
      • Infectious Diseases and Immunology Division (IICB)
      Kolkata, Bengal, India
  • 2004
    • University of Texas Medical Branch at Galveston
      Galveston, Texas, United States
  • 2002
    • University of Virginia
      • Department of Biochemistry, Molecular Biology and Genetics
      Charlottesville, VA, United States