Identification of cellular deoxyhypusine synthase as a novel target for antiretroviral therapy
Ilona Hauber, … , Thomas Harrer, Joachim Hauber
Ilona Hauber, … , Thomas Harrer, Joachim Hauber
Published January 3, 2005
Citation Information: J Clin Invest. 2005;115(1):76-85. https://doi.org/10.1172/JCI21949.
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Article AIDS/HIV

Identification of cellular deoxyhypusine synthase as a novel target for antiretroviral therapy

Abstract

The introduction of highly active antiretroviral therapy (HAART) has significantly decreased morbidity and mortality among patients infected with HIV-1. However, HIV-1 can acquire resistance against all currently available antiretroviral drugs targeting viral reverse transcriptase, protease, and gp41. Moreover, in a growing number of patients, the development of multidrug-resistant viruses compromises HAART efficacy and limits therapeutic options. Therefore, it is an ongoing task to develop new drugs and to identify new targets for antiretroviral therapy. Here, we identified the guanylhydrazone CNI-1493 as an efficient inhibitor of human deoxyhypusine synthase (DHS). By inhibiting DHS, this compound suppresses hypusine formation and, thereby, activation of eukaryotic initiation factor 5A (eIF-5A), a cellular cofactor of the HIV-1 Rev regulatory protein. We demonstrate that inhibition of DHS by CNI-1493 or RNA interference efficiently suppressed the retroviral replication cycle in cell culture and primary cells. We show that CNI-1493 inhibits replication of macrophage- and T cell–tropic laboratory strains, clinical isolates, and viral strains with high-level resistance to inhibitors of viral protease and reverse transcriptase. Moreover, no measurable drug-induced adverse effects on cell cycle transition, apoptosis, and general cytotoxicity were observed. Therefore, human DHS represents a novel and promising drug target for the development of advanced antiretroviral therapies, particularly for the inhibition of multidrug-resistant viruses.

Authors

Ilona Hauber, Dorian Bevec, Jochen Heukeshoven, Friedrich Krätzer, Florian Horn, Axel Choidas, Thomas Harrer, Joachim Hauber

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Figure 1

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Inhibition of hypusine formation in eIF-5A by interfering with DHS activ...
Inhibition of hypusine formation in eIF-5A by interfering with DHS activity. (A) Schematic representation of the pathway of eIF-5A biosynthesis. Hypusine is required for the biological activity of the protein. The conversion of a specific lysine residue (Lys50) into the unusual amino acid hypusine occurs in 2 posttranslational reactions that are mediated by the enzymes DHS and deoxyhypusine hydroxylase (DHH; see text for details). DHS is a target of experimental low–molecular weight drugs, such as the multivalent guanylhydrazone CNI-1493. (B) CNI-1493 inhibits hypusine formation in eIF-5A in vivo. Jurkat cells were metabolically labeled using either [14C]putrescine or [35S]cysteine in the presence of either DMSO (solvent control, lane 1) or CNI-1493 (1.0 μM, lane 2; 0.5 μM, lane 3). Equal amounts of the various radiolabeled cellular extracts were then subjected to eIF-5A–specific immunoprecipitation analysis and analyzed by SDS-PAGE and autoradiography. Molecular mass standards are indicated in kDa. (C) Two-dimensional electrophoresis of total proteins from PM1 cells that were metabolically labeled using [14C]spermidine in the presence of either DMSO (upper panel) or CNI-1493 (1.0 μM; lower panel). Protein separation was performed, using equal amounts of the radiolabeled extracts, by isoelectric focusing in the first dimension on an immobilized linear pH gradient (IPG) from pH 4.0 to 7.0, which was followed by SDS-PAGE separation in the second dimension. Only molecular mass of less than 25.0 kDa is shown in autoradiographs. No radioactive protein spots were detected elsewhere. Molecular mass standards are indicated in kDa on the right.