Discovery of a Small Molecule PDI Inhibitor That Inhibits Reduction of HIV-1 Envelope Glycoprotein gp120

Abstract

Protein disulfide isomerase (PDI) is a promiscuous protein with multifunctional properties. PDI mediates proper protein folding by oxidation or isomerization and disrupts disulfide bonds by reduction. The entry of HIV-1 into cells is facilitated by the PDI-catalyzed reductive cleavage of disulfide bonds in gp120. PDI is regarded as a potential drug target because of its reduction activity. We screened a chemical library of natural products for PDI-specific inhibitors in a high-throughput fashion and identified the natural compound juniferdin as the most potent inhibitor of PDI. Derivatives of juniferdin were synthesized, with compound 13 showing inhibitory activities comparable to those of juniferdin but reduced cytotoxicity. Both juniferdin and compound 13 inhibited PDI reductase activity in a dose-dependent manner, with IC(50) values of 156 and 167 nM, respectively. Our results also indicated that juniferdin and compound 13 exert their inhibitory activities specifically on PDI but do not significantly inhibit homologues of this protein family. Moreover, we found that both compounds can inhibit PDI-mediated reduction of HIV-1 envelope glycoprotein gp120.

Small molecule-induced oxidation of protein disulfide isomerase is neuroprotective

“…In this insulin aggregation assay (8,14,15,23), PDIa reduced the two disulfide bonds between the α-and β-chains of insulin, causing the β-chain to aggregate and precipitate, resulting in an increase in absorbance at 650 nm. Of eight hit compounds from the cell culture screen, two, LOC14 and LOC6, were able to almost completely inhibit PDIa enzymatic activity (Fig.…”

“…By matching the conditions reported by Kemmink et al (25), we were able to efficiently transfer the assignments of unaltered residues in the oxidized protein to the peaks of 1 H-15 N HSQC spectrum of reduced PDI protein. To validate the assignments, 15 N total correlated spectroscopy-HSQC and 15 N-nuclear Overhauser effect spectroscopy-HSQC were performed (Fig. S4).…”

“…One challenge has been the lack of available drug-like inhibitors, especially for in vivo evaluation in neurodegenerative disease models. Reported inhibitors of PDI are (i) irreversible binders to the catalytic site cysteines (1,8,13), (ii) not cell permeable, because they were designed for the inhibition of extracellular PDI (14,15), or (iii) nonselective hormones and antibiotics, such as estrone and bacitracin, that act broadly on multiple target proteins (15,16). Irreversible inhibitors, although having promise in ovarian cancer, have mechanism-based toxicity that is not likely well tolerated in neurons.…”

High-Throughput HIV–Cell Fusion Assay for Discovery of Virus Entry Inhibitors

“…[26][27][28][29][30][31][32][33][34][35] Multiple screens for HIV-1 inhibitors relied on in vitro assays, which used viral proteins or their fragments, or on HIV-1 infections/replication as a readout. In vitro screening has identified competitive inhibitors of assembly of the gp41 HR1-and HR2-derived peptides into the 6HB.…”

Rutin- potent natural thrombolytic agent

“… Agents like Juniferdin or Bacitracin which also inhibit PDI function and thus inhibit thrombus formation in vivo (Khan et al, 2011;Dickerhof et al, 2011;Cho et al, 2008) and are either cytotoxic or non-selective (Karala and Ruddock, 2010;Khan et al, 2011). When compared with these agents, rutin demonstrated selectivity towards extra-cellular PDI and is relatively non-toxic.…”