Application of monoclonal antibodies to measure metabolism of an anti-trypanosomal compound in vitro and in vivo.
Human African trypanosomiasis (HAT), also called African sleeping sickness, is a neglected tropical parasitic disease indigenous to sub-Saharan Africa. Diamidine compounds, including pentamidine and CPD-0801, are potent anti-trypanosomal molecules. The latter is a potential drug in the development at the UNC based Consortium for Parasitic Drug Development. An orally bioavailable prodrug of CPD-0801, DB868, is metabolized primarily in the liver to the active form. A monoclonal antibody developed against a pentamidine derivative has shown significant reactivity with CPD-0801 (EC(50) 65.1 nM), but not with the prodrug (EC(50)>18,000 nM). An inhibitory enzyme-linked immunosorbent assay (IELISA) has been used to quantitatively monitor prodrug metabolism by detecting the production of the active compound over time in a sandwich culture rat hepatocyte system and in rats. These results were compared with the results of the standard LC/MS/MS assay. Spearman coefficients of 0.96 and 0.933 (in vitro and in vivo, respectively) indicate a high correlation between these two measurement methods. This novel IELISA provides a facile, inexpensive, and accurate method for drug detection that may aide in elucidating the mechanisms of action and toxicity of existing and future diamidine compounds.
Goldsmith
Rachel Beaulieu
RB
Department of Pathology and Laboratory Medicine, School of Medicine, University of North Carolina, Chapel Hill, North Carolina 27599-7525, USA.
Gray
Danny R
DR
Yan
Zhixia
Z
Generaux
Claudia N
CN
Tidwell
Richard R
RR
Reisner
Howard M
HM
D000818:Animals; D000911:Antibodies, Monoclonal / Q000096:biosynthesis / Q000276:immunology*; D000918:Antibody Specificity / Q000276:immunology; D001550:Benzamidines / Q000276:immunology; D003429:Cross Reactions / Q000276:immunology; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods; D005260:Female; D022781:Hepatocytes / Q000378:metabolism; D006801:Humans; D007074:Immunoglobulin G / Q000096:biosynthesis / Q000276:immunology; D008297:Male; D051379:Mice; D008807:Mice, Inbred BALB C; D010419:Pentamidine / Q000031:analogs & derivatives / Q000276:immunology; D011355:Prodrugs / Q000378:metabolism*; D051381:Rats; D017207:Rats, Sprague-Dawley; D017208:Rats, Wistar; D053719:Tandem Mass Spectrometry; D014344:Trypanocidal Agents / Q000032:analysis* / Q000097:blood / Q000276:immunology / Q000378:metabolism*; D014353:Trypanosomiasis, African / Q000188:drug therapy
D000911:Antibodies, Monoclonal; D001550:Benzamidines; D007074:Immunoglobulin G; D011355:Prodrugs; D014344:Trypanocidal Agents; D010419:Pentamidine; C033286:furamidine; C458513:pafuramidine