A small-molecule oxocarbazate inhibitor of human cathepsin L blocks severe acute respiratory syndrome and ebola pseudotype virus infection into human embryonic kidney 293T cells.
A tetrahydroquinoline oxocarbazate (PubChem CID 23631927) was tested as an inhibitor of human cathepsin L (EC 3.4.22.15) and as an entry blocker of severe acute respiratory syndrome (SARS) coronavirus and Ebola pseudotype virus. In the cathepsin L inhibition assay, the oxocarbazate caused a time-dependent 17-fold drop in IC(50) from 6.9 nM (no preincubation) to 0.4 nM (4-h preincubation). Slowly reversible inhibition was demonstrated in a dilution assay. A transient kinetic analysis using a single-step competitive inhibition model provided rate constants of k(on) = 153,000 M(-1)s(-1) and k(off) = 4.40 x 10(-5) s(-1) (K(i) = 0.29 nM). The compound also displayed cathepsin L/B selectivity of >700-fold and was nontoxic to human aortic endothelial cells at 100 muM. The oxocarbazate and a related thiocarbazate (PubChem CID 16725315) were tested in a SARS coronavirus (CoV) and Ebola virus-pseudotype infection assay with the oxocarbazate but not the thiocarbazate, demonstrating activity in blocking both SARS-CoV (IC(50) = 273 +/- 49 nM) and Ebola virus (IC(50) = 193 +/- 39 nM) entry into human embryonic kidney 293T cells. To trace the intracellular action of the inhibitors with intracellular cathepsin L, the activity-based probe biotin-Lys-C5 alkyl linker-Tyr-Leu-epoxide (DCG-04) was used to label the active site of cysteine proteases in 293T lysates. The reduction in active cathepsin L in inhibitor-treated cells correlated well with the observed potency of inhibitors observed in the virus pseudotype infection assay. Overall, the oxocarbazate CID 23631927 was a subnanomolar, slow-binding, reversible inhibitor of human cathepsin L that blocked SARS-CoV and Ebola pseudotype virus entry in human cells.
Shah
Parag P
PP
Department of Chemical and Biomolecular Engineering, Penn Center for Molecular Discovery, Institute for Medicine and Engineering, University of Pennsylvania, Philadelphia, Pennsylvania 19104-6383, USA.
Wang
Tianhua
T
Kaletsky
Rachel L
RL
Myers
Michael C
MC
Purvis
Jeremy E
JE
Jing
Huiyan
H
Huryn
Donna M
DM
Greenbaum
Doron C
DC
Smith
Amos B
AB
Bates
Paul
P
Diamond
Scott L
SL
D000998:Antiviral Agents / Q000494:pharmacology*; D056668:Cathepsin L / Q000037:antagonists & inhibitors*; D002460:Cell Line; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology*; D019142:Hemorrhagic Fever, Ebola / Q000517:prevention & control*; D006801:Humans; D007700:Kinetics; D011804:Quinolines / Q000494:pharmacology*; D015363:Quinolones / Q000494:pharmacology*; D045169:Severe Acute Respiratory Syndrome / Q000517:prevention & control*
C551180:2-(3,4-dihydro-2H-quinolin-1-yl)-2-oxo-ethyl ester; D000998:Antiviral Agents; D015853:Cysteine Proteinase Inhibitors; D011804:Quinolines; D015363:Quinolones; D056668:Cathepsin L
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