Document 1113
 DOCN  M94A1113
 TI    Measurement of the anti HIV agent -2',3'-didehydro-2',3'
       dideoxythymidine (D4T) by competitive ELISA.
 DT    9412
 AU    Guedj R; Ferrua B; Tran TT; Rotpin C; Quaranta JF; Sinet M; Durand J;
       Laboratoire de Chimie Bio-organique, Nice, France.
 SO    Int Conf AIDS. 1994 Aug 7-12;10(2):206 (abstract no. PB0836). Unique
       Identifier : AIDSLINE ICA10/94371462
 AB    OBJECTIVE: D4T, a thymidine analog with potent anti HIV activity in
       vitro, is currently investigated as therapy for patients with advanced
       HIV infection. D4T is a prodrug which is converted by cellular thymidine
       kinases to the active metabolite D4T triphosphate (D4T-TP), acting as
       competitive inhibitor of the HIV encoded reverse transcriptase. Because
       dose-limiting toxicity is associated with D4T therapy and level of
       intracellular D4T-TP is dependent on various factors such as the cell
       type, the activation state or the presence of other antiviral agents,
       measurement of D4T and its phosphate forms is essential. METHODS: In the
       present study, using anti-D4T rabbit antibodies raised against D4T
       hemisuccinate-bovine serum albumin conjugate, a D4T
       hemisuccinate-peroxidase as tracer and a microtiter plate coated with
       anti-rabbit IgG as separator system, a one-step convenient competitive
       ELISA method was developed for assaying D4T. RESULTS: The method was
       capable to specifically detect 2 ng/ml of D4T in ultrafiltrates from
       plasmas previously separated on microconcentrator devices. Thymidine,
       azido thymidine and D4T monophosphate which closely resembles D4T,
       cross-reacted at 0.04%, 0.25% and 1.8% respectively. The method was
       applied for quantitating intracellular D4T in CEM and Molt 4 cell lines
       and monitoring plasmatic D4T in patients with advanced HIV infection.
       Additionnally, this technique was extended to the indirect measurement
       of intracellular phosphorylated D4T metabolites following reverse phase
       separation of cell extracts and treatment of fractions with alkaline
       phosphatase. DISCUSSION AND CONCLUSIONS: The extrapolation of this
       method to another drugs and their metabolites, such as AZT, DDI, DDC,
       AZT-TP, DDI-TP, DDC-TP ..., will be investigated.
 DE    Cross Reactions  *Enzyme-Linked Immunosorbent Assay  Human
       Stavudine/*BLOOD  Thymidine/BLOOD  Zidovudine/BLOOD  MEETING ABSTRACT

       SOURCE: National Library of Medicine.  NOTICE: This material may be
       protected by Copyright Law (Title 17, U.S.Code).