Enhancing the copy number of Ldrab6 gene in Leishmania donovani parasites mediates drug resistance through drug‐thiol conjugate dependent multidrug resistance protein A (MRPA)

Chauhan, Indira Singh; Rao, Gita Subba; Singh, Neeloo

doi:10.1016/j.actatropica.2019.105158

Cited by 11 publications

(9 citation statements)

References 18 publications

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“…This protein was also more abundant in the LiSbR line [22,46]. It has been shown that RAB GTPases of L. major are highly immunogenic in individuals immune to cutaneous and visceral leishmaniasis [47]. L. donovani overexpressing RAB6 showed a resistant phenotype by allowing trans-dibenzalacetone-treated parasites to both increase internal thiol levels and enhance MRP pump activity [47].…”

Section: Discussionmentioning

confidence: 99%

Comparative transcriptomic analysis of antimony resistant and susceptible Leishmania infantum lines

Andrade¹,

Gonçalves

Liarte

et al. 2020

Parasites Vectors

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Background One of the major challenges to leishmaniasis treatment is the emergence of parasites resistant to antimony. To study differentially expressed genes associated with drug resistance, we performed a comparative transcriptomic analysis between wild-type and potassium antimonyl tartrate (SbIII)-resistant Leishmania infantum lines using high-throughput RNA sequencing. Methods All the cDNA libraries were constructed from promastigote forms of each line, sequenced and analyzed using STAR for mapping the reads against the reference genome (L. infantum JPCM5) and DESeq2 for differential expression statistical analyses. All the genes were functionally annotated using sequence similarity search. Results The analytical pipeline considering an adjusted p-value < 0.05 and fold change > 2.0 identified 933 transcripts differentially expressed (DE) between wild-type and SbIII-resistant L. infantum lines. Out of 933 DE transcripts, 504 presented functional annotation and 429 were assigned as hypothetical proteins. A total of 837 transcripts were upregulated and 96 were downregulated in the SbIII-resistant L. infantum line. Using this DE dataset, the proteins were further grouped in functional classes according to the gene ontology database. The functional enrichment analysis for biological processes showed that the upregulated transcripts in the SbIII-resistant line are associated with protein phosphorylation, microtubule-based movement, ubiquitination, host–parasite interaction, cellular process and other categories. The downregulated transcripts in the SbIII-resistant line are assigned in the GO categories: ribonucleoprotein complex, ribosome biogenesis, rRNA processing, nucleosome assembly and translation. Conclusions The transcriptomic profile of L. infantum showed a robust set of genes from different metabolic pathways associated with the antimony resistance phenotype in this parasite. Our results address the complex and multifactorial antimony resistance mechanisms in Leishmania, identifying several candidate genes that may be further evaluated as molecular targets for chemotherapy of leishmaniasis.

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Section: Discussionmentioning

confidence: 99%

Comparative transcriptomic analysis of antimony resistant and susceptible Leishmania infantum lines

Andrade¹,

Gonçalves

Liarte

et al. 2020

Parasites Vectors

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“…Because the MDR and MRP pumps are known to confer elevated Sb III and AmB resistance in Leishmania parasites ( 10 , 16 , 17 ), this study assessed their involvement in the activity of APs. For this purpose, the AP susceptibility of nonresistant strains in combination with reference ABC transport inhibitors was evaluated; probenecid is known to reverse the effects of MRP pumps ( 67 , 68 ), while verapamil and cyclosporine inhibit efflux mediated by both the MRP and MDR pumps ( 69 – 71 ). Treatment with Sb III was included as a positive control ( 21 , 72 – 75 ) and confirmed an increased susceptibility of extracellular and intracellular parasites in combination with verapamil and probenecid, although verapamil is considered not to efficiently inhibit the MDR pumps in Leishmania ( 76 ).…”

Section: Discussionmentioning

confidence: 99%

Antileishmanial Aminopyrazoles: Studies into Mechanisms and Stability of Experimental Drug Resistance

Kerkhof

Mabille

Hendrickx

et al. 2020

Antimicrob Agents Chemother

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Introduction: Current antileishmanial treatment is hampered by limitations such as drug toxicity and the risk of treatment failure, which may be related to parasitic drug resistance. Given the urgent need for novel drugs, the Drugs for Neglected Diseases initiative (DNDi) has undertaken a drug discovery program, which has resulted in the identification of aminopyrazoles - a highly promising antileishmanial chemical series. Multiple experiments have been performed to anticipate the propensity for resistance development. Methodology: Resistance selection was performed by successive exposure of Leishmania infantum promastigotes (in vitro) and intracellular amastigotes (both in vitro and in golden Syrian hamsters). The stability of resistant phenotypes was assessed after passage in mice and Lutzomyia longipalpis sand flies. Whole genome sequencing (WGS) was performed to identify mutated genes, copy number variations (CNVs) and somy changes. The potential role of efflux pumps (MDR/MRP) in the development of resistance was assessed by co-incubation of aminopyrazoles with specific efflux pump inhibitors (verapamil, cyclosporine A and probenecid). Results: Repeated drug exposure of amastigotes did not result in the emergence of drug resistance, either in vitro or in vivo. Selection on the promastigote stage, however, was able to select for parasites with reduced susceptibility (resistance index: 5-24). This phenotype proved to be unstable after in vivo passage in mice and sand flies, suggesting that non-fixed alterations are responsible for the elevated resistance. In line with this, single nucleotide polymorphisms and indels identified by whole genome sequencing could not be directly linked to the decreased drug susceptibility. Copy number variations were absent, whereas somy changes were detected, which may account for the transient acquisition of resistance. Finally, aminopyrazole activity was not influenced by the MDR and MRP efflux pump inhibitors tested. Conclusion: The selection performed does not suggest rapid development of resistance against aminopyrazoles in the field. Karyotype changes may confer elevated levels of resistance, but these do not seem to be stable in the vertebrate and invertebrate hosts. MDR/MRP efflux pumps are not likely to significantly impact the activity of the aminopyrazole leads.

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“…This protein was also more abundant in the LiSbR line [22,46]. It has been shown that RAB GTPases of L. major are highly immunogenic in immune individuals to cutaneous and visceral leishmaniasis [47]. L. donovani overexpressing RAB6 showed a resistant phenotype, by allowing trans-dibenzalacetone treated parasites to both increase internal thiol levels and enhance MRP pump activity [47].…”

Section: Discussionmentioning

confidence: 99%

Comparative transcriptomic analysis of antimony resistant and susceptible Leishmania infantum lines

Andrade

Gonçalves

Liarte

et al. 2020

Preprint

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Background: One of the major challenges for leishmaniasis treatment is the emergence of parasites resistant to antimony. In order to study differentially expressed genes associated with drug resistance we performed a comparative transcriptomic analysis between wild-type and potassium antimonyl tartrate (SbIII)-resistant Leishmania infantum lines using high-throughput RNA sequencing.Methods: All the cDNA libraries were constructed from promastigote forms of each line, sequenced and analyzed using STAR for mapping the reads against the reference genome (L. infantum JPCM5) and DESeq2 for differential expression statistical analyses. All the genes were functionally annotated using sequence similarity search.Results: The analytical pipeline considering an adjusted p-value lower than 0.05 and fold change greater than 2.0 identified 933 transcripts differentially expressed (DE) between wild-type and SbIII-resistant L. infantum lines. Out of 933 DE transcripts, 504 presented functional annotation and 429 were assigned as hypothetical proteins. A total of 837 transcripts were upregulated and 96 were downregulated in the SbIII-resistant L. infantum line. Using this DE dataset, the proteins were further grouped in functional classes according to the Gene Ontology database. The functional enrichment analysis for biological processes showed that the upregulated transcripts in the SbIII-resistant line are associated with protein phosphorylation, microtubule-based movement, ubiquitination, host-parasite interaction, cellular process and other categories. The downregulated transcripts in the SbIII-resistant line are assigned in the GO categories: ribonucleoprotein complex, ribosome biogenesis, rRNA processing, nucleosome assembly and translation.Conclusions: The transcriptomic profile of L. infantum showed a robust set of genes from different metabolic pathways associated with antimony resistance phenotype in this parasite. Our results address the complex and multifactorial antimony resistance mechanisms in Leishmania, identifying several candidate genes that may be further evaluated as molecular targets for chemotherapy of leishmaniasis.

show abstract

Enhancing the copy number of Ldrab6 gene in Leishmania donovani parasites mediates drug resistance through drug‐thiol conjugate dependent multidrug resistance protein A (MRPA)

Cited by 11 publications

References 18 publications

Comparative transcriptomic analysis of antimony resistant and susceptible Leishmania infantum lines

Comparative transcriptomic analysis of antimony resistant and susceptible Leishmania infantum lines

Antileishmanial Aminopyrazoles: Studies into Mechanisms and Stability of Experimental Drug Resistance

Comparative transcriptomic analysis of antimony resistant and susceptible Leishmania infantum lines

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