Towards an understanding of the mechanism of action of PZQ

 

aAnthony D. Aragon, aReza A. Imani, aVint R. Blackburn, aPauline M Cupit, aSandra Melman, bTinopiwa Goronga, bThomas Webb, aEric S. Loker, a*Charles Cunningham

 

aDepartment of Biology, University of New Mexico, Albuquerque, NM 87131, USA bChemical Biology & Therapeutics, St Jude Children's Research Hospital, Memphis, TN 38105, USA

 

Abstract

 

Although praziquantel (PZQ) has been used to treat schistosomiasis for over 20 years its mechanism of action remains unknown. We have developed an assay based on the transcriptional response of S. mansoni PR-1 to heat shock to confirm that while 6 week post infection (p.i.) schistosomes are sensitive to PZQ, 4 week p.i. schistosomes are not. Further, we have used this assay to demonstrate that in mice this sensitivity develops between days 37 and 40 p.i. When linked to the fluorophore BODIPY, a structurally modified PZQ which likely retains anti-helmenthic activity appears to enter easily the cells of intact 4 and 6 week p.i. schistosomes as well as mammalian NIH 3T3 cells suggesting that the differential effects of PZQ is not based on cell exclusion. A transcriptomal analysis of gene expression between 4 and 6 weeks p.i. revealed 607 up-regulated candidate genes whose products are potential PZQ targets. A comparison of this gene list with that of genes expressed by PZQ sensitive miracidia reduced this target list to 247 genes, including a number involved in aerobic metabolism and cytosolic calcium regulation. Finally, we also report the effect of an in vitro sub-lethal exposure of PZQ on the transcriptome of S. mansoni PR-1. Annotation of genes differentially regulated by PZQ exposure suggests that schistosomes may undergo a transcriptomic response similar to that observed during oxidative stress.

Supplemental files

 

Quantitative RT-PCR

 

Contraction assay (.pdf)

qRT-PCR primers (.txt)

qRT-PCR PZQ exposed (no heat shock) (.pdf)

qRT-PCR Female only (.pdf)

qRT-PCR Male only (.pdf)

qRT-PCR Recovery experiment (.pdf)

qRT-PCR PZQ exposed 4 hr (.pdf)

PZQ exposure Venn diagrams (.pdf)

 

PZQ compound

 

PZQ-BODIPY (.pdf)

 

Microarray files

 

Overlap of biological replicates (.pdf)

 

PZQ up gene list (.txt)

PZQ up gene list with sequences (.txt)

PZQ up Blast2GO output (.xls)

 

PZQ down gene list (.txt)

PZQ down gene list with sequences (.txt)

PZQ down Blast2GO output (.xls)

 

Immature gene list (.txt)

Immature gene list with sequences (.txt)

Immature Blast2GO (.xls)

 

Mature gene list (.txt)

Mature gene list with sequences (.txt)

Mature Blast2GO (.xls)

 

All microarray data generated in this study is available from the GEO database, using accession number GSE12850.

 

*Corresponding Author

 

Charlie Cunningham

University of New Mexico

Dept. of Biology

Castetter Hall 218

Albuquerque, NM

New Mexico 87131

Tel.: +1 505 277 5578

Fax +1 505 277 0304

E-mail address: ccunnin@unm.edu