Discovery of Adamantane Carboxamides as Ebola Virus Cell Entry and Glycoprotein Inhibitors

Article abstract of DOI:10.1021/acsmedchemlett.0c00025

We identified and explored the structure-activity-relationship (SAR) of an adamantane carboxamide chemical series of Ebola virus (EBOV) inhibitors. Selected analogs exhibited half-maximal inhibitory concentrations (EC50 values) of ～10-15 nM in vesicular s

Full text of DOI:10.1021/acsmedchemlett.0c00025

pubs.acs.org/acsmedchemlett
Letter
Discovery of Adamantane Carboxamides as Ebola Virus Cell Entry
and Glycoprotein Inhibitors
Michael B. Plewe, Nadezda V. Sokolova, Vidyasagar Reddy Gantla, Eric R. Brown, Shibani Naik,
Alexandra Fetsko, Donald D. Lorimer, David M. Dranow, Hayden Smutney, Jameson Bullen, Rana Sidhu,
Arshil Master, Junru Wang, E. Adam Kallel, Lihong Zhang, Birte Kalveram, Alexander N. Freiberg,
Greg Henkel, and Ken McCormack*
Cite This: https://dx.doi.org/10.1021/acsmedchemlett.0c00025
Read Online
sı
*
Metrics & More
Article Recommendations
Supporting Information
ACCESS
ABSTRACT: We identified and explored the structure−activity-
relationship (SAR) of an adamantane carboxamide chemical series
of Ebola virus (EBOV) inhibitors. Selected analogs exhibited half-
maximal inhibitory concentrations (EC50 values) of ∼10−15 nM in
vesicular stomatitis virus (VSV) pseudotyped EBOV (pEBOV)
infectivity assays, low hundred nanomolar EC50 activity against
wild type EBOV, aqueous solubility >20 mg/mL, and attractive
metabolic stability in human and nonhuman liver microsomes. X-ray cocrystallographic characterizations of a lead compound with
the EBOV glycoprotein (GP) established the EBOV GP as a target for direct compound inhibitory activity and further provided
relevant structural models that may assist in identifying optimized therapeutic candidates.
KEYWORDS: Ebola virus, glycoprotein, cell entry, adamantane carboxamide
BOV is a single stranded RNA virus associated with severe
hemorrhagic fever exhibiting >50% lethality in humans.1,2
internalized into endosomes primarily via micropinocyto-
sis.3−5,7 Within acidified endosomes glycosylated GP1 is
cleaved of a “mucin-domain” into a smaller form by the pH-
dependent Cathepsin L and B proteases, priming it for
interactions with the Niemann Pick C1 (NPC1) protein, a
secondary intracellular receptor in endosomal/lysosomal
membranes. NPC1 binding, in combination with other events,
triggers GP conformational rearrangements including GP2-
mediated fusion of viral and intracellular organelle membranes
thereby releasing EBOV genetic material into the cell for
further replication.7−9
A number of antibody-based EBOV therapeutics bind to GP
blocking early cell entry steps while small molecule EBOV cell
entry inhibitors have primarily been reported to block later
steps in viral entry/fusion by interacting with a number of
intracellular mammalian host proteins regulating EBOV
transport, processing, and membrane fusion or, alternatively,
directly with the viral GP protein.3−5,8−13 We utilized a
biosafety level 2 (BSL2) nonreplicative vesicular stomatitis virus
(VSV) pseudotyped virus system in which VSV viral particles
express a luciferase reporter gene (in place of the VSV GP
E
Symptoms typically manifest with generalized fever, sore
throat, and muscular pain whereupon vomiting, diarrhea, and
both internal and external bleeding may follow; transmission of
EBOV is spread through contact with these bodily fluids. The
development of EBOV vaccines as well as intravenous (i.v.)
administered monoclonal antibody and small molecule
therapeutics, many of which focus on the EBOV glycoprotein
(GP) and/or viral cell entry, has recently drawn intensive
interest; the testing and utilization of these treatments was
accelerated in response to major EBOV outbreaks in West
Africa and the Democratic Republic of the Congo (DRC).3−5
However, despite the availability of effective vaccine and i.v.
administered therapeutics, the DRC Kivu outbreak continued
to escalate and spread geographically from August 2018
through August 2019 at which time the number of new cases
began to stabilize and subsequently decrease.6 Thus, a
significant need remains in some resource settings for the
development of more convenient and readily distributable
therapeutics including oral small molecule treatment options.
EBOV GP, the sole protein expressed on the EBOV
envelope membrane surface, is responsible for host cell
attachment, endosomal entry, and membrane fusion. The
EBOV GP is trimeric, with each monomer cleaved by furin
into two polypepides, GP1 and GP2. The entry of EBOV into
host cells is initiated by the interaction of sites on GP1
subunits with one or more receptors exposed on the host cell
external membrane surface, upon which the virus is
Received: January 15, 2020
Accepted: April 21, 2020
https://dx.doi.org/10.1021/acsmedchemlett.0c00025
© XXXX American Chemical Society
ACS Med. Chem. Lett. XXXX, XXX, XXX−XXX
A

ACS Medicinal Chemistry Letters
pubs.acs.org/acsmedchemlett
Letter
gene) while the EBOV GP is supplied in trans, to rapidly
screen a chemical library and characterize compound hits.14
Utilizing this approach we screened a collection of small
molecules representing ∼72,000 compounds in Vero cells at 5
μM to identify compounds inhibiting 50% of the luciferase
signal generated by infection with pEBOV without similar
activity against control pVSV infected cells.
We report the discovery and initial SAR of an adamantane
carboxamide chemical series of pEBOV inhibitor compounds
identified from our library screen as hit compound 1, which
exhibited >50% activity at 5 μM against pEBOV but not pVSV.
1 was then subjected to concentration−response experiments
and found to exhibit an EC50 of ∼3.9 μM. We subsequently
obtained commercially available analogs of 1, and identified 2,
which exhibited a ∼10-fold greater pEBOV potency (Figure
1). As 2 differed from 1 only by the addition of a phenyl group
bonding ability of terminal amine play an important role in
determining binding affinity.
We subsequently explored the addition of several sub-
stituents at the para-position of the phenyl group. Unfortu-
nately, both polar and nonpolar groups were found to decrease
compound potency as shown for compounds 9−13. Nonpolar
substituents such as Cl resulted in decreased activity (11 vs 5)
while more polar groups including OH caused a more
significant loss of activity (10 vs 5) indicating that para
substitutions on phenyl groups are not well tolerated.
Given the synthetic challenges associated with the
generation of similar ortho- and meta-phenyl substituted
analogs and further exploration of the phenyl group, we next
focused on modifications of the amide position, replacement of
the adamantane core with noradamantane, introduction of a
spacer between the adamantyl moiety and amide functionality,
and amide group replacements that are illustrated by
compounds 14−19 in Table 2 (see Supporting Information,
Schemes S1−S5 for the synthesis of intermediates and final
compounds). Transposition of the amide group to an adjacent
secondary carbon, giving compound 14, decreased potency
approximately 3-fold from the analogous tertiary carbon amide
5, whereas noradamantyl 15, acetamide 16, reverse amide 17,
urea 18, and diamine 19 analogs all exhibited even greater
decreases in pEBOV potency.
Figure 1. Screening hit compound 1 and its phenyl analog 2 with
Since each of the different amide group replacements
(compounds 16−19) decreased pEBOV potency, we next
explored the introduction of additional substituents into the
adamantane core (Supporting Information) and generated
novel phenyl adamantane carboxamides bearing alkyl groups at
the bridgehead positions. The addition of two methyl groups
into the adamantane core of compound 20 (Core D-1, Table
3) resulted in a ∼4-fold decrease in pEBOV potency compared
to compound 5, whereas in contrast, the addition of a single
methyl group (Core D-2, Table 3) improved potency about
∼2-fold as observed in pEBOV activity comparisons of
compounds 21 vs 5. We subsequently generated additional
analogs of Core D-2 by coupling 3-methyl-5-phenyladaman-
tane-1-carboxylic acid with a variety of different mono Boc-
protected diamines, followed by cleavage of the Boc-group
under acidic conditions (Table 3, Scheme 1, steps iii and iv).
Both cyclic and bicyclic amines of different ring sizes
containing an exocyclic or endocyclic nitrogen, as well as
acyclic amines have been investigated. Although substituted
aminopiperidines (compounds 22 and 23) have similar
potency to compound 21, other amines (compounds 24−27
and 29) exhibited decreased activity. Other novel amines
(compounds 28, 30, 31, and 33), particularly trans-cyclo-
hexyldiamine (compound 33) showed promising activity while
cis-cyclohexyldiamine (compound 32) is less active. These
trends further suggested the importance of the protonated
amine group to conform within a specified geometry in
association with its target protein, given the observed
improvement with addition of a single methyl substituent at
the adamantane bridgehead position protein. Given the
observed improvement with the addition of a single methyl
substituent at the adamantane bridgehead position (Core D-
2), we also explored replacement of the methyl group with the
larger and more lipophilic ethyl group (Core D-3, Table 3),
which further increased potency (compare 34 vs 21 and 35 vs
33) to provide compounds with EC50s ≤ 20 nM. Upon the
identification of analogs with improved compound potencies,
we subsequently assessed the SAR of prioritized compounds
indicated pEBOV EC50 values.
to the adamantane core, we were interested in further SAR
exploration of phenyl adamantane carboxamides. We sub-
sequently generated analogs with modifications of the phenyl
adamantane core as well as the amine portion as illustrated in
Scheme 1 (see Supporting Information for synthesis details
and analytical data).
a
Scheme 1. Synthesis of Aryl Adamantane Carboxamides
a
Reagents and conditions: (i) Br2, Fe, 0 °C for 30 min, then rt, 8 h;
(ii) AlCl3, Ar−H, reflux, 6 h; (iii) R3R4NH, EDC hydrochloride,
HOAt, DIEA, DCM, rt, 12 h; (iv) TFA, DCM, rt, 2 h.
A number of amine groups were thus initially explored as
shown by compounds 3−8 in Table 1. Comparing the
homologous series of compounds 2, 3, and 5 illustrated a
link between modifications of the terminal amine and
improved potency. In addition, comparison of pEBOV
activities of the tetrahydroisoquinoline compound 8 (a
commercially available compound) vs the 4-aminopiperidine
of compound 5 also suggested that the basicity and hydrogen
B
https://dx.doi.org/10.1021/acsmedchemlett.0c00025
ACS Med. Chem. Lett. XXXX, XXX, XXX−XXX

ACS Medicinal Chemistry Letters
pubs.acs.org/acsmedchemlett
Letter
Table 1. SAR of 3-Aryladamantane Carboxamides Exploring Various Amines and Aryl Groups
*
Data are average of three independent experiments, standard deviation (s.d.) ≤ 20%.
for potential metabolic liabilities. Compounds exhibiting EC50
values of ∼ ≤150 nM were evaluated for in vitro human liver
microsome (HLM) metabolic stability. Core D-2 compounds
21, 31, and 33 exhibited pEBOV EC50s < 50 nM and HLM
stability >75% at 1 h while core D-3 compounds 34 and 35
exhibited EC50s ≤ 20 nM and slightly lower HLM stabilities of
∼70%. These analogs were used as the basis of further
exploration and the 4-aminopiperidine (21), trans-4-
(aminomethyl)cyclohexylamine (31), and trans-cyclohexyldi-
amines (33 and 35) appeared to provide particularly attractive
lead candidates. Kinetic aqueous solubilities (Supporting
Information) were also assessed for all compounds described
in this report and each exhibited a kinetic aqueous solubility
>100 μM at pH 6.4 and 7.5 except compound 27, which
exhibited a solubility of ∼61 μM.
Because the adamantane nucleus forms a perfect tetrahe-
dron, adamantanes with four unique substituents at the
bridgehead positions are chiral. Hence, the addition of the
methyl and ethyl groups to the phenyladamantane carbox-
amide core (Cores D-2 and D-3) generates chiral compounds
with the chiral center residing in the center of the adamantane.
In light of this, we next generated the (1S,3R,5R,7S)-
(abbreviated as S) and (1R,3S,5S,7R)- (abbreviated as R)
enantiomers of the prioritized racemic analogs 21, 31, 33, and
35 (Supporting Information). As shown in Table 4, the S
enantiomers are eutomers exhibiting a greater potency than the
R enantiomers. The potency differences between the S and R
enantiomers of the ethyl (42 vs 43) and methyl derivatives (38
vs 39) were ∼30 and 10-fold, respectively while the rank order
SAR of pEBOV activity for racemic and S enantiomeric
compounds was similar.
as observed in the concentration−response relationships of the
S enantiomeric compounds plotted in Figure 2. Of note, the
Hill slopes are all substantially greater than 1 (Table 4)
indicating a positive cooperativity underlying the inhibitory
activity of each of the compounds.
In order to confirm that compound inhibitory activities
observed in the pEBOV assay format translate to activity
against replicative pathogenic EBOV, the prioritized S
enantiomers were characterized in plaque or viral yield
reduction (VYR) assays (Supporting Information) as shown
in Table 4. Comparison of the EBOV EC90 (from the VYR
assay) values vs the half-maximal cellular cytotoxicity or CC50
provides SI90 therapeutic selectivity indices ([CC50]/[EC90])
for compounds 36, 38, and 40 of 34, 77, and 40, respectively.
Compound 42 was only run in the plaque assay and exhibited
an SI50 value ([CC50]/[EC50]) of 221. These data confirm the
antiviral activity of the compounds against replicative EBOV
with potencies that translate in similar rank order to that found
against pEBOV. There is however, an offset in EBOV vs
pEBOV potencies, whereby EC50s against infectious EBOV
correlate more closely with EC90s against the pseudotyped
virus pEBOV. Nonetheless, the discovery and potency
optimization of our prioritized analogs from our initial
screening hit compound 1 vs 38, 40, and 42 (pEBOV EC50
improvement from ∼3.9 uM to ∼10 nM) further validates the
use of BSL2 pseudotyped viral systems for the discovery of
novel inhibitors of highly pathogenic BSL4 viruses.
For additional SAR comparison and compound prioritiza-
tion we further characterized the solubility of the S-
enantiomers 36, 38, 40, and 42 and found them all to exhibit
>20 mg/mL aqueous solubility at pH 7.4. To provide
additional assessments for advancement into one or more in
vivo animal model studies, we subsequently characterized 36,
In addition to pEBOV potency (EC50 values) the Hill slopes
and activity curves of the compounds are all roughly equivalent
C
https://dx.doi.org/10.1021/acsmedchemlett.0c00025
ACS Med. Chem. Lett. XXXX, XXX, XXX−XXX

ACS Medicinal Chemistry Letters
pubs.acs.org/acsmedchemlett
Letter
Table 2. SAR Exploration of Functional Group Spacers
Table 3. Adamantane Carboxamides Core SAR
*
Data are averages of three independent experiments, s.d. ≤ 20%.
38, 40, and 42 in multispecies liver microsome (MLM)
metabolic stability assays including human and the potential
efficacy and/or toxicology model species mouse, guinea pig,
rat, cynomolgus monkey, and dog. While each of the S analogs
exhibited suitable HLM metabolic stability, compounds 36 and
38 provided the most favorable MLM metabolic stability
profile. Further in vitro and in vivo ADMET characterizations
of these prioritized analogs will be assessed to identify lead
candidates and suitable species for in vivo efficacy and
toxicology studies.
*
Data are averages of three independent experiments, s.d. ≤ 20%.
**
Percent remaining at 60 min. HLM: human liver microsomes.
Herein, we have detailed the discovery and initial SAR of a
novel adamantane carboxamide chemical series of EBOV
inhibitors. Previous reports identifying inhibitors of EBOV cell
entry have provided a number of targets that include a variety
of potential mammalian host proteins regulating intracellular
EBOV processing, transport, and membrane fusion, as well as
the EBOV GP itself, which has been identified as the target for
several FDA-approved drugs.3−5,8−13,15,16 Interestingly, the
adamantane chemical series identified here is structurally
related to an adamantane dipeptide piperazine EBOV inhibitor
chemical series found to associate with the mammalian host
protein and intracellular GP receptor NPC1.8 Thus, it was
initially expected that NPC1 might also represent the target for
our adamantane carboxamide EBOV inhibitor chemical series.
To identify the target and potential mechanism of action of
our inhibitor compounds, we determined X-ray cocrystallo-
graphic structures of the EBOV GP protein with compound
38. In Figure 3 we provide structural representations generated
through crystallization of the apo protein with subsequent
soaking of the crystals with 38 similar to that previously
described (Supporting Information).13 As mentioned above,
the functional EBOV GP complex is comprised of a trimer of
GP1-GP2 dimers. In panels A−D (Figure 3) the GP binding
pocket, located at the interface of each GP1-GP2 dimer, and
specific amino acid interactions underlying association with 38
are depicted. The binding pocket is characterized by
hydrophobic residues defining a central hydrophobic cavity
to accommodate the adamantane core and phenyl group of 38
(analogs with a phenyl group showed increased potency of
∼10-fold) through lipophilic GP1 residues VAL66, LEU184,
and LEU186 as well as GP2 residues LEU515, TYR517,
D
https://dx.doi.org/10.1021/acsmedchemlett.0c00025
ACS Med. Chem. Lett. XXXX, XXX, XXX−XXX

ACS Medicinal Chemistry Letters
pubs.acs.org/acsmedchemlett
Letter
Table 4. Ebola Virus Inhibitory Activity, Cytotoxicity, and Liver Microsomal Stability of Enantiomeric Adamantane
Carboxamides
*
**
***
Data are average of three independent experiments, standard deviation ≤20%. n = 3, standard deviation <20%.
Percent remaining after 60
min, H: Human, M: Mouse, G: Guinea Pig, R: Rat, MK: Monkey, D: Dog.
MET548, and LEU558, while the carbonyl of the amide forms
an important hydrogen bond with ARG64.
In binding to the EBOV GP compound 38 displaces a GP1
polypeptide loop that is normally bound within this pocket in
the apo conformation. This GP1 polypeptide loop, previously
defined as comprising residues 192−194 (the “DFF lid”), was
previously found to be displaced by toremifene and a number
of other small molecules.13,15−17 The association of 38 with the
EBOV GP parallels that of other compounds although as
represented in Figure 3 (panel B) GP1 polypeptide residues
193−195 (residues FFS) appear to bind within the apo pocket
and overlap with bound compound 38 while D192 appears to
lie outside of a shared binding pocket. However the boundaries
of the apo polypeptide loop or “lid” may be defined, the
binding of a GP inhibitor to this site displaces the GP1 loop
and alters GP conformation thereby preventing fusion between
the viral and endosomal membranes. Such ligand induced
Figure 2. Concentration−response curves of prioritized S-enan-
tiomers against pEBOV s.d. (n = 3); the cooperative Hill slopes
ranged from 1.4 to 1.7 (Table 4).
E
https://dx.doi.org/10.1021/acsmedchemlett.0c00025
ACS Med. Chem. Lett. XXXX, XXX, XXX−XXX

ACS Medicinal Chemistry Letters
pubs.acs.org/acsmedchemlett
Letter
Figure 3. X-ray cocrystallographic structures of compound 38 bound to the EBOV GP. (Panel A) (top left) Compound 38 (yellow) binding
pocket located at the GP1 (dark green)−GP2 (dark blue) dimer interface. (Panel B) (top right) Overlay of the apo GP1 (light green) and GP2
(light blue) and 38-bound GP1 (dark green) and GP2 (dark blue) protein structures. Residues 193−195 (amino acids FFS) in the apo
conformation GP1 polypeptide are highlighted (purple) and exhibit overlap with compound 38 in a shared binding pocket. (Panel C) (bottom left)
GP binding pocket for 38 and residues determining important ligand−protein interactions including lipophilic GP1 residues VAL66, LEU184, and
LEU186 and GP2 residues LEU515, TYR517, MET548, and LEU558. (Panel D) (bottom right) Lipophilic (purple) and hydrophilic (green)
molecular surfaces of GP showing the central cavity accommodating the adamantane core, lipophilic phenyl pocket, and amide/cyclohexylamine
association sites with ARG64.
conformational changes may inhibit GP function through
compound-specific mechanisms including premature release of
GP2 and/or deficient GP processing, both of which could
disrupt engagement with its receptor NPC1.13,15−17 Regardless
of the potential for any compound-specific mechanisms for
these inhibitors, toremifene exhibits an EC50 of ∼80 nM in our
pEBOV assays while another recently identified EBOV GP
inhibitor (118a) has a reported pEBOV EC50 of ∼50 nM in
comparison to the adamantane carboxamide GP inhibitors
described here, of which some exhibit pEBOV EC50s in the
range of ∼10−14 nM.17
Given that the EBOV GP is composed of a trimer of GP1-
GP2 dimers with three potential inhibitor binding sites, along
with the positive cooperativity observed in the Hill slopes of
the prioritized compound inhibition curves (Table 4 and
Figure 2), it appears likely that the adamantane carboxamide
inhibitor molecules act allosterically whereby the binding of
one molecule to an initial GP1-GP2 dimer enhances the
binding of additional inhibitor molecules to other GP1-GP2
dimers within the same GP1-GP2 trimeric complex in order to
fully abrogate EBOV GP function and viral replication. GP
inhibitors exhibiting steeper Hill slopes would thereby be
expected to provide superior antiviral activity.
In summary, we have identified a novel adamantane
carboxamide chemical series and prioritized analogs exhibiting
potent activity against pEBOV and replicative EBOV, which
directly bind to and inhibit the EBOV GP. Selected lead
compounds also exhibit attractive aqueous (HCl salt) solubility
>20 mg/mL and multispecies in vitro liver microsome
metabolic stability to provide candidates for the further
development of novel small molecule therapeutics. The
determination of a binding site and relevant structural models
may also facilitate structure-based drug design of the
adamantane carboxamide chemical series to provide the
identification of improved therapeutic candidates.
F
https://dx.doi.org/10.1021/acsmedchemlett.0c00025
ACS Med. Chem. Lett. XXXX, XXX, XXX−XXX

ACS Medicinal Chemistry Letters
pubs.acs.org/acsmedchemlett
Letter
Lihong Zhang − Department of Pathology, The University of
Texas Medical Branch, Galveston, Texas 77555, United States
Birte Kalveram − Department of Pathology, The University of
Texas Medical Branch, Galveston, Texas 77555, United States
Alexander N. Freiberg − Department of Pathology and Center
for Biodefense and Emerging Infectious Diseases, The University
of Texas Medical Branch, Galveston, Texas 77555, United
States
ASSOCIATED CONTENT
■
sı
* Supporting Information
The Supporting Information is available free of charge at
https://pubs.acs.org/doi/10.1021/acsmedchemlett.0c00025.
Experimental procedures and synthetic schemes, ana-
lytical data, pseudotyped virus generation and assay,
infectious wild type Ebola virus plaque assay, virus yield
reduction (VYR) assay, liver microsome metabolic
stability assay, solubility and cytotoxicity assays; Ebola
GP protein expression, purification, crystallization, and
structure determination. The atomic coordinates and
structure factors have been deposited with the RCSB
Protein Data Bank under accession code 6NAE. (PDF)
Greg Henkel − Arisan Therapeutics, San Diego, California
92121, United States
Complete contact information is available at:
https://pubs.acs.org/10.1021/acsmedchemlett.0c00025
Funding
This work was funded in whole or in part with Federal funds
from the National Institute of Allergy and Infectious Diseases,
National Institutes of Health, Department of Health and
Human Services, under SIBR grants R43AI118207 and
R43AI138878 and Contract #HHSN272201700059C.
AUTHOR INFORMATION
■
Corresponding Author
Ken McCormack − Arisan Therapeutics, San Diego, California
92121, United States; orcid.org/0000-0003-1181-9137;
Phone: (858) 766-0495; Email: kenm@arisanthera.com
Notes
The authors declare no competing financial interest.
Authors
Michael B. Plewe − Arisan Therapeutics, San Diego, California
ACKNOWLEDGMENTS
■
92121, United States
Arisan Therapeutics has utilized the nonclinical and preclinical
services program offered by the National Institute of Allergy
and Infectious Diseases.
Nadezda V. Sokolova − Arisan Therapeutics, San Diego,
California 92121, United States
Vidyasagar Reddy Gantla − Arisan Therapeutics, San Diego,
California 92121, United States
Eric R. Brown − Arisan Therapeutics, San Diego, California
92121, United States
Shibani Naik − Arisan Therapeutics, San Diego, California
92121, United States
Alexandra Fetsko − Arisan Therapeutics, San Diego, California
92121, United States
Donald D. Lorimer − Seattle Structural Genomics Center for
Infectious Disease (SSGCID), Seattle, Washington 98105,
United States; UCB Pharma, Bainbridge Island, Washington
98110, United States
David M. Dranow − Seattle Structural Genomics Center for
Infectious Disease (SSGCID), Seattle, Washington 98105,
United States; UCB Pharma, Bainbridge Island, Washington
98110, United States
ABBREVIATIONS
■
EBOV, Ebola virus;; VSV, vesicular stomatitis virus; GP,
glycoprotein; pEBOV, pseudotyped VSV expressing the EBOV
GP; HLM, human liver microsomes; MLM, multispecies liver
microsomes; i.v., intravenous; SAR, structure−activity-relation-
ships; VYR, viral yield reduction; BSL4, biosafety level 4;
NPC1, Niemann Pick C1 protein; ADMET, absorption,
distribution, metabolism, excretion, and toxicity; DCM,
dichloromethane; DMF, N,N-dimethylformamide; TFA, tri-
fluoroacetic acid; DIEA, N,N-diisopropylethylamine; EDC, 1-
ethyl-3-(3-(dimethylamino)propyl)carbodiimide; RT, room
temperature; Me, methyl; Et, ethyl; HOAt, 1-hydroxy-7-
azabenzotriazole; Boc, tert-butoxycarbonyl.
Hayden Smutney − Seattle Structural Genomics Center for
Infectious Disease (SSGCID), Seattle, Washington 98105,
United States; UCB Pharma, Bedford, Massachusetts 01730,
United States
Jameson Bullen − Seattle Structural Genomics Center for
Infectious Disease (SSGCID), Seattle, Washington 98105,
United States; UCB Pharma, Bainbridge Island, Washington
98110, United States
Rana Sidhu − Seattle Structural Genomics Center for Infectious
Disease (SSGCID), Seattle, Washington 98105, United States;
UCB Pharma, Bedford, Massachusetts 01730, United States
Arshil Master − Seattle Structural Genomics Center for
Infectious Disease (SSGCID), Seattle, Washington 98105,
United States; UCB Pharma, Bedford, Massachusetts 01730,
United States
Junru Wang − Seattle Structural Genomics Center for Infectious
Disease (SSGCID), Seattle, Washington 98105, United States;
UCB Pharma, Bedford, Massachusetts 01730, United States
E. Adam Kallel − Victrix, San Diego, California 92128, United
States
REFERENCES
■
(1) Rojas, M.; Monsalve, D. M.; Pacheco, Y.; Acosta-Ampudia, Y.;
Ramírez-Santana, C.; Ansari, A. A.; Gershwin, M. E.; Anaya, J. M.
Ebola Virus Disease: An Emerging and Re-Emerging Viral Threat. J.
Autoimmun. 2020, 106, 102375. No. October.
(2) Burk, R.; Bollinger, L.; Johnson, J. C.; Wada, J.; Radoshitzky, S.
R.; Palacios, G.; Bavari, S.; Jahrling, P. B.; Kuhn, J. H. Neglected
Filoviruses. FEMS Microbiol. Rev. 2016, 40 (4), 494−519.
(3) Hoenen, T.; Groseth, A.; Feldmann, H. Therapeutic Strategies to
Target the Ebola Virus Life Cycle. Nat. Rev. Microbiol. 2019, 17 (10),
593−606.
(4) Furuyama, W.; Marzi, A. Ebola Virus: Pathogenesis and
Countermeasure Development. Annu. Rev. Virol. 2019, 6 (1), 435−
458.
(5) Edwards, M. R.; Basler, C. F. Current Status of Small Molecule
Drug Development for Ebola Virus and Other Filoviruses. Curr. Opin.
Virol. 2019, 35, 42−56.
(6) Oppenheim, B.; Lidow, N.; Ayscue, P.; Saylors, K.; Mbala, P.;
Kumakamba, C.; Kleinman, M. Knowledge and Beliefs about Ebola
Virus in a Conflict-Affected Area: Early Evidence from the North Kivu
Outbreak. J. Glob. Health 2019, 9 (2), 020311.
G
https://dx.doi.org/10.1021/acsmedchemlett.0c00025
ACS Med. Chem. Lett. XXXX, XXX, XXX−XXX

ACS Medicinal Chemistry Letters
pubs.acs.org/acsmedchemlett
Letter
(7) Rhein, B. A.; Maury, W. J. Ebola Virus Entry into Host Cells:
Identifying Therapeutic Strategies. Curr. Clin. Microbiol. Reports 2015,
2 (3), 115−124.
̂
́
(8) Cote, M.; Misasi, J.; Ren, T.; Bruchez, A.; Lee, K.; Filone, C. M.;
Hensley, L.; Li, Q.; Ory, D.; Chandran, K.; et al. Small Molecule
Inhibitors Reveal Niemann−Pick C1 Is Essential for Ebola Virus
Infection. Nature 2011, 477 (7364), 344−348.
́
́
́
(9) Feneant, L.; Szymanska-de Wijs, K. M.; Nelson, E. A.; White, J.
M. An Exploration of Conditions Proposed to Trigger the Ebola Virus
Glycoprotein for Fusion. PLoS One 2019, 14 (7), 1−20.
(10) Flint, M.; Chatterjee, P.; Lin, D. L.; McMullan, L. K.;
Shrivastava-Ranjan, P.; Bergeron, E.; Lo, M. K.; Welch, S. R.; Nichol,
́
S. T.; Tai, A. W.; et al. A Genome-Wide CRISPR Screen Identifies N-
Acetylglucosamine-1-Phosphate Transferase as a Potential Antiviral
Target for Ebola Virus. Nat. Commun. 2019, 10 (1), 285.
(11) Penny, C. J.; Vassileva, K.; Jha, A.; Yuan, Y.; Chee, X.; Yates, E.;
Mazzon, M.; Kilpatrick, B. S.; Muallem, S.; Marsh, M.; et al. Mining of
Ebola Virus Entry Inhibitors Identifies Approved Drugs as Two-Pore
Channel Pore Blockers. Biochim. Biophys. Acta, Mol. Cell Res. 2019,
1866 (7), 1151−1161.
(12) Salata, C.; Calistri, A.; Alvisi, G.; Celestino, M.; Parolin, C.;
̀
Palu, G. Ebola Virus Entry: From Molecular Characterization to Drug
Discovery. Viruses 2019, 11 (3), 274.
(13) Zhao, Y.; Ren, J.; Harlos, K.; Jones, D. M.; Zeltina, A.; Bowden,
T. A.; Padilla-Parra, S.; Fry, E. E.; Stuart, D. I. Toremifene Interacts
with and Destabilizes the Ebola Virus Glycoprotein. Nature 2016, 535
(7610), 169−172.
̈
(14) Garbutt, M.; Liebscher, R.; Wahl-Jensen, V.; Jones, S.; Moller,
̈
P.; Wagner, R.; Volchkov, V.; Klenk, H.-D.; Feldmann, H.; Stroher, U.
Properties of Replication-Competent Vesicular Stomatitis Virus
Vectors Expressing Glycoproteins of Filoviruses and Arenaviruses. J.
Virol. 2004, 78 (10), 5458 LP−5465.
(15) Ren, J.; Zhao, Y.; Fry, E. E.; Stuart, D. I. Target Identification
and Mode of Action of Four Chemically Divergent Drugs against
Ebolavirus Infection. J. Med. Chem. 2018, 61 (3), 724−733.
(16) Zhao, Y.; Ren, J.; Fry, E. E.; Xiao, J.; Townsend, A. R.; Stuart,
D. I. Structures of Ebola Virus Glycoprotein Complexes with Tricyclic
Antidepressant and Antipsychotic Drugs. J. Med. Chem. 2018, 61
(11), 4938−4945.
(17) Shaikh, F.; Zhao, Y.; Alvarez, L.; Iliopoulou, M.; Lohans, C.;
Schofield, C. J.; Padilla-Parra, S.; Siu, S. W. I.; Fry, E. E.; Ren, J.; et al.
Structure-Based in Silico Screening Identifies a Potent Ebolavirus
Inhibitor from a Traditional Chinese Medicine Library. J. Med. Chem.
2019, 62 (6), 2928−2937.
H
https://dx.doi.org/10.1021/acsmedchemlett.0c00025
ACS Med. Chem. Lett. XXXX, XXX, XXX−XXX