Structure-Property Basis for Solving Transporter-Mediated Efflux and Pan-Genotypic Inhibition in HCV NS5B Inhibitors

Article abstract of DOI:10.1021/acsmedchemlett.8b00379

In solving the P-gp and BCRP transporter-mediated efflux issue in a series of benzofuran-derived pan-genotypic palm site inhibitors of the hepatitis C virus NS5B replicase, it was found that close attention to physicochemical properties was essential. In these compounds, where both molecular weight (MW >579) and TPSA (>110 ?²) were high, attenuation of polar surface area together with weakening of hydrogen bond acceptor strength of the molecule provided a higher intrinsic membrane permeability and more desirable Caco-2 parameters, as demonstrated by trifluoroacetamide 11 and the benchmark N-ethylamino analog 12. In addition, the tendency of these inhibitors to form intramolecular hydrogen bonds potentially contributes favorably to the improved membrane permeability and absorption. The functional group minimization that resolved the efflux problem simultaneously maintained potent inhibitory activity toward a gt-2 HCV replicon due to a switching of the role of substituents in interacting with the Gln414 binding pocket, as observed in gt-2a NS5B/inhibitor complex cocrystal structures, thus increasing the efficiency of the optimization. Noteworthy, a novel intermolecular S=O?C=O n → π? type interaction between the ligand sulfonamide oxygen atom and the carbonyl moiety of the side chain of Gln414 was observed. The insights from these structure-property studies and crystallography information provided a direction for optimization in a campaign to identify second generation pan-genotypic NS5B inhibitors.

Full text of DOI:10.1021/acsmedchemlett.8b00379

Letter
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Cite This: ACS Med. Chem. Lett. XXXX, XXX, XXX−XXX
Structure−Property Basis for Solving Transporter-Mediated Efflux
and Pan-Genotypic Inhibition in HCV NS5B Inhibitors
Kap-Sun Yeung,*,† Brett R. Beno,† Kathy Mosure,† Juliang Zhu,† Katherine A. Grant-Young,†
Kyle Parcella,† Prakash Anjanappa,‡ Rajesh Onkardas Bora,‡ Kumaravel Selvakumar,‡ Ying-Kai Wang,†
Hua Fang,† Rudolph Krause,† Karen Rigat,† Mengping Liu,† Julie Lemm,† Steven Sheriff,§
Mark Witmer,§ Jeffrey Tredup,§ Adam Jardel,§ Kevin Kish,§ Dawn Parker,† Roy Haskell,†
Kenneth Santone,† Nicholas A. Meanwell,† Matthew G. Soars,† Susan B. Roberts,† and John F. Kadow†
†Bristol-Myers Squibb Research and Development, P.O. Box 5100, 5 Research Parkway, Wallingford, Connecticut 06492, United
States
‡Department of Discovery Chemistry, Biocon Bristol-Myers Squibb Research and Development Center, Biocon Park, Jigani Link
Road, Bommasandra IV, Bangalore 560099, India
§Bristol-Myers Squibb Research and Development, P.O. Box 4000, Princeton, New Jersey 08543, United States
S
* Supporting Information
ABSTRACT: In solving the P-gp and BCRP transporter-mediated efflux issue in a series of benzofuran-derived pan-genotypic
palm site inhibitors of the hepatitis C virus NS5B replicase, it was found that close attention to physicochemical properties was
essential. In these compounds, where both molecular weight (MW >579) and TPSA (>110 Å2) were high, attenuation of polar
surface area together with weakening of hydrogen bond acceptor strength of the molecule provided a higher intrinsic membrane
permeability and more desirable Caco-2 parameters, as demonstrated by trifluoroacetamide 11 and the benchmark N-
ethylamino analog 12. In addition, the tendency of these inhibitors to form intramolecular hydrogen bonds potentially
contributes favorably to the improved membrane permeability and absorption. The functional group minimization that resolved
the efflux problem simultaneously maintained potent inhibitory activity toward a gt-2 HCV replicon due to a switching of the
role of substituents in interacting with the Gln414 binding pocket, as observed in gt-2a NS5B/inhibitor complex cocrystal
structures, thus increasing the efficiency of the optimization. Noteworthy, a novel intermolecular SO···CO n → π* type
interaction between the ligand sulfonamide oxygen atom and the carbonyl moiety of the side chain of Gln414 was observed.
The insights from these structure−property studies and crystallography information provided a direction for optimization in a
campaign to identify second generation pan-genotypic NS5B inhibitors.
KEYWORDS: Hepatitis C virus, NS5B, benzofuran, P-gp, BCRP, efflux, TPSA, hydrogen bonding
chronic HCV infection, as measured by the sustained
hronic hepatitis C virus (HCV) infection, which affects
Caround 150 million people in the world, is a liver disease virological response, can be achieved with the new all oral
combination therapies that offer good safety profiles, broad
genotype activity, and abridged treatment duration.2,3 These
small molecule drugs inhibit one of the three HCV
that can progress into liver cirrhosis and life-threatening
hepatocellular carcinoma, and is responsible for about 700,000
deaths per year globally.1 Intensive drug discovery research in
the past quarter century since the characterization of the virus
in 1989 has resulted in the clinical application of HCV-targeted
direct-acting antiviral agents that have replaced the previous
standard of care, pegylated-interferon, and ribavirin. A cure for
Received: August 16, 2018
Accepted: November 5, 2018
Published: November 5, 2018
© XXXX American Chemical Society
A
DOI: 10.1021/acsmedchemlett.8b00379
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ACS Medicinal Chemistry Letters
Letter
nonstructural proteins, the NS3/4A serine protease, the NS5A
replication cofactor, and the NS5B replicase, all of which play
essential roles in the HCV replication cycle.4,5 The NS5B
replicase is an RNA-dependent RNA polymerase that is
responsible for the synthesis of HCV genomic RNA. Its tertiary
structure adopts a right-hand configuration with fingers, palm,
and thumb domains representative of polymerase enzymes. In
addition to the active site-directed nucleoside/nucleotide
inhibitors and their prodrugs, non-nucleoside inhibitors that
bind to three distinct allosteric sites (thumb-I, thumb-II, and
palm site) in HCV NS5B replicase have also been
developed.6,7
pre-existing gt-1bC316N mutant generally trended with an
increase in lipophilicity.8 A more desirable and balanced
potency profile was achieved with the phenylcyclopropyl amide
4, which is almost one cLogP9 unit more lipophilic than the 2-
pyridinyl matched pair analog 3 and has inhibitory EC50 values
below 10 nM across gt-1b, gt-1a, and gt-2a replicons. However,
initial in vitro profiling of these C6-N-(2-hydroxyethyl)-
methylsulfonamide substituted benzofuran compounds showed
that they had less than desirable microsomal stability, as
exemplified by the low percentage of unmetabolized 4
remaining in human and rat liver microsomes (HLM and
RLM, respectively) in a high throughput assay. The inadequate
metabolic stability of the compound was resolved by
incorporating a 2-pyrimidine ring, a metabolic stability
enhancing moiety identified during the lead optimization
leading to 1, in place of the phenyl ring in 4 and removing the
primary hydroxyl group in the hydroxyethyl side chain to avoid
any possible metabolic modifications of this potentially
vulnerable functionality. The derived analog 5 showed similar
lipophilicity to 4 but a significant improvement in metabolic
stability in both HLM and RLM compared to 4 while
maintaining the overall antiviral potency profile (Table 1).
Although the intrinsic membrane permeability of this series of
potent N-ethylmethylsulfonamide analogs was respectable, as
indicated by the PAMPA (parallel artificial membrane
permeability assay) values, a major deficiency in the in vitro
profile was the high susceptibility to efflux in Caco-2 cells, as
exemplified by the high efflux ratio exhibited by 5 and its close
4′-methoxy analog 6 (>8.6 and >12 (Table 2), respectively).
As a result, these compounds exhibited low oral bioavailability
with poor exposure in both the plasma and liver of rats
We have reported the discovery of BMS-929075 (1), a palm
site allosteric inhibitor of HCV NS5B replicase (Figure 1), that
Figure 1. Structure of the benzofuran-derived NS5B palm site
inhibitor 1.
was advanced into Phase 1 clinical studies.8 In cell-based
replicon assays, this benzofuran-based compound 1 exhibited
potent HCV inhibitory activity toward all genotypes with the
exception of gt-2 (EC50 > 100 nM).
During the subsequent second generation campaign to
identify pan-genotypic inhibitors from this series, it was
discovered that incorporation of a N-(2-hydroxyethyl)-
methylsulfonamide group at the C6 position of the benzofuran
ring led to a ∼20-fold enhancement in inhibitory activity
against gt-2a (JFH-1 strain) replicon cells, as illustrated by the
C6 matched molecular pair 2 and 3 (Table 1). During the lead
optimization resulting in 1, it was observed in the property−
activity relationship that potency toward gt-1b, gt-1a, and the
following oral dosing. In the presence of zosuquidar,10
a
selective P-glycoprotein (P-gp) efflux pump inhibitor, the
Table 2. In Vitro Profiling and Inhibitory Potency towards
HCV gt-2a Replicon (EC50) for Compounds 6 and 8−12
Table 1. Inhibitory Potency towards HCV Replicons (EC50)
and in Vitro Profiling for Compounds 2−5
a
Replicon EC50 (nM)
d
gt-
gt-
1a
gt-
HLM/RLM
%remain
b
f
Cpd 1b
1bC316N gt-2a
cLogP
c
2
3
4
5
3.8
52
6.2
8.8
36
35
5.7
7.7
75
385
64
515
22
8.6
5.4
91/54
99/8.7
63/0.6
4.93
2.71
3.56
3.48
a
HBA = hydrogen bonding acceptor, relative strength on the pKBHX
e
b
c
scale.12 TPSA = topological polar surface area.9 A-B = apical to
44
100/91
basolateral; B-A = basolateral to apical, Data for 5: efflux ratio >8.6, A-
a
b
d
Data are average values of n ≥ 2 independent experiments. JFH-1
B < 15, B-A = 130. Replicon, JFH-1 strain, data are average values of
c
e
strain. Data from a single experiment performed in duplicate; see SI
n ≥ 2 independent experiments; Data were obtained from a single
d
f
for CC50 and SD. HLM/RLM denotes human and rat liver
experiment performed in duplicate. pKBHX data for N-methylaniline.
microsomes, respectively. t1/2 (min) >120/49. Calculated values.9
See SI for CC50 and SD.
e
f
B
DOI: 10.1021/acsmedchemlett.8b00379
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ACS Medicinal Chemistry Letters
Letter
efflux of compound 6 in Caco-2 cells was dramatically
suppressed (efflux ratio = 1.3) and the apical to basolateral
(A to B) permeability was favorably increased to 168 nm/s
(Table 3). The same phenomenon was also observed in the
low metabolic stability in HLM and RLM (t1/2 = 13.9 and 26
min, respectively) and the unsatisfactory exposure in both the
plasma and liver (C5h = 277 and 7180 nM, respectively) of rats
following oral administration in a pharmacokinetic screen
prevented this compound from being further progressed.
Concurrently, a SAR exercise was conducted to evaluate the
effect of various sulfonamide surrogates that express differential
physicochemical properties related to hydrogen bonding on
the permeability profile (Table 2) to gain insight for resolving
the efflux issue.14,15 Interestingly, an examination of polar
surface area (represented by calculated topological polar
surface area (TPSA)9,16) of the molecule and hydrogen
bonding acceptor (HBA) strength (relative strength on the
pKBHX scale12) of the C6-functionality suggested that
modulation of either physicochemical property alone was not
sufficient to confer a more favorable membrane permeability
profile. As shown in Table 2, TPSA progressively decreases
from 6 to 8, 9, and 10, while the HBA strength increases in the
reverse order from 10 to 9, 8, and 6. However, neither 6 nor
10, which are at the lower end of the pKBHX and TPSA values,
respectively, provided a useful permeability profile. In these
analogs where the TPSA is >110 Å2 and MW >579, values that
are typically not conducive for membrane absorption, a
reduced polar surface area coupled with a weaker HBA
strength is required to provide a higher intrinsic permeability
as well as improved Caco-2 parameters. This was manifested in
the trifluoroacetamide analog 11 in which both the TPSA and
HBA strength values are the lowest when compared to
compounds 6−10, exhibiting the highest PAMPA value (835
nm/s) and a more favorable efflux ratio of 2.7 among these
analogs. These observations led to the assessment of the
weakly basic C6-N-ethyl aniline analog 12, which is also a very
weak HBA, representing a compromise between minimizing
both TPSA and HBA strength. Pleasingly, compound 12
maintained the high PAMPA permeability of 11. Importantly,
the apical to basolateral forward flux in Caco-2 cells was
dramatically improved in 12 with essentially no efflux
compared to 11 and in remarkable contrast to the profile of
the parent compound 6. These results are consistent with
studies that demonstrated that binding to the P-gp recognition
pocket is exclusively controlled by hydrogen bonding acceptors
present in the substrate.17 The favorable absorption profile of
Table 3. Caco-2 Permeability Behavior of Compound 6 in
the Presence of Transporter Inhibitors
Caco-2 Pc (nm/s)
Cpd
inhibitor
efflux ratio
A-B
B-A
6
Zosuquidar (P-gp)
KO143 (BCRP)
1.3
4.8
168
67
215
324
presence of KO143,11 a selective inhibitor of breast cancer
resistance protein (BCRP), although to a lesser extent. These
results suggested that the efflux of the sulfonamide analogs was
predominantly mediated by P-gp and to some extent by the
BCRP transporter as well.
Based on the SAR of the C6-unsubstituted series,8 the N-
ethylmethylsulfonamide group appeared to be responsible for
the potent activity against gt-2 but also a contributor to the
high efflux liability of this new series of pan-genotypic
inhibitors. One approach explored to address the efflux issue
was to enhance the intrinsic membrane permeability of the
molecule while retaining the sulfonamide moiety to preserve
potency toward gt-2 virus. This could be achieved empirically
by varying the substitution pattern at the 3′- and 4′-positions
on the C5-phenyl ring, with F, Cl, Me, and OMe groups
examined. In this regard, a substituent at the 4′-position that
could potentially engage the cyclopropyl amide NH in an
intramolecular hydrogen bond to facilitate permeability
appeared more favorable.13 As shown in Table 4, in general,
the PAMPA membrane permeability of these compounds
increased, and the apical to basolateral forward permeability in
Caco-2 cells was enhanced with increasing lipophilicity, as
suggested by their cLogP values. Although the 3′-methyl
analog 7c appeared to be an outlier, the A to B influx of this
compound was quantifiable compared to that of compound 6.
The 3′-Cl-4-OMe analog 7b emerged with the highest PAMPA
value in these C6-sulfonamide derivatives. Encouragingly, the
Caco-2 efflux ratio of this compound also decreased
substantially to a more desirable value of 2.2. However, the
Table 4. In Vitro Profiling and Inhibitory Potency towards HCV gt-2a Replicon (EC50) for Compounds 7a−d Compared with
6
b
c
Caco-2 Pc (nm/s)
gt-2a EC50 (nM)
a
Cpd
R4’
R3′
cLogP
PAMPA pH7.4 (nm/s)
Efflux Ratio
A-B
B-A
6
OMe
OMe
OMe
OMe
F
H
F
Cl
Me
F
3.27
3.41
3.82
3.76
3.77
487
659
821
409
731
>12.4
13.8
2.2
9.8
7.8
<15
29
52
33
79
185
406
114
322
623
13
7a
7b
7c
7d
8.8
6.6
16
9.0
a
b
c
Calculated values.9 A-B = apical to basolateral; B-A = basolateral to apical. Replicon, JFH-1 strain, data are average values of n ≥ 2 independent
experiments. See SI for CC50 and SD.
C
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Letter
12 translated into promising exposure in both the plasma and
the liver of rats with good oral bioavailability following oral
administration (Table 5). The plasma AUC and the
bond accepting moiety at C6 for activity toward gt-2 virus. The
optimization toward a pan-genotypic NS5B inhibitor in the
benzofuran and related 7-azabenzofuran series18,19 followed
the recognition of this key SAR point that a HBA was
dispensable. In the cocrystal structure of compound 5, a close
analog of 6, in complex with gt-2a NS5B L30S protein, the N-
ethylmethylsulfonamide moiety assumes a conformation that
appears to be partly driven by the electronic repulsion between
the sulfonamide oxygen atoms and the central C5-phenyl ring.
The sulfonamide methyl group projects into a hydrophobic
pocket formed by the side chains from the Pro197, Leu384,
and Tyr415 residues of the protein (Figure 2a). The ethyl
group is not in direct interaction with the protein, but rather in
intramolecular van der Waals contact with the C5-phenyl ring
within compound 5. One of the sulfonamide oxygen atoms
forms a hydrogen bond with the guanidine moiety in the side
chain of Arg200. In addition, the pyrimidine moiety π-stacks
with the phenyl ring of Tyr452 and the pyrimidinylcyclopropyl
amide carbonyl forms a network of water-bridged hydrogen
bonds to nearby residues including Tyr452 and Trp550. In the
absence of the SO2 group, the N-ethylamino side chain can
adopt a conformation in which the ethyl group flips to fill the
volume of the hydrophobic pocket, as revealed by a cocrystal
structure of the 5-fluoropyrimidine derivative 13, a close
analog of compound 12, bound to gt-2a NS5B L30S protein
(Figure 2b). In this cocrystal structure, the carbonyl oxygen
atom of the cyclopropylamide in compound 13 forms water-
bridged hydrogen bonds to the side chain of Arg200 and the
backbone carbonyl of Tyr195 through a single water molecule.
The N−H of the ethylamino moiety in compound 13 forms a
hydrogen bond with the carbonyl oxygen atom of the primary
amide in the side chain of Gln414 with a N−H···OC
Table 5. Pharmacokinetic Parameters in Rats for 6 and 12
a
(0 to 24 h)
IV
b
HLM/RLM t1/2
CL
Vss
(L/kg)
Cpd
(min)
(mL/min/kg)
IV t1/2 (h)
6
12
91/>120
36/39
60
11
1.6
4.7
14
4
PO
Cmax
(nM)
tmax
(h)
AUC
(nM·h)
C5 liver
(nM)
C24 liver
(nM)
Cpd
%F
c
6
12
223
776
1.9
5.3
740
7803
1177
5258
ND
391
20
57
a
b
IV, 2 mg/kg; PO, 6 mg/kg; n = 3. HLM/RLM denotes human and
c
rat liver microsomes, respectively. ND = not detected.
concentration in the liver at the 5 h time point obtained
after oral dosing of 12 were, respectively, 10- and 5-fold higher
than those from compound 6, which, in contrast to 12, was not
detected in either the plasma or the liver at the 24 h time point
postdosing. The undesirable high clearance after IV dosing
exhibited by 6 was also substantially improved in 12.
Compound 12 maintained potent activity toward gt-1b, 1a,
and gt-1bC316N mutant replicons (EC50 = 1.2, 2.3, and 3.6
nM, respectively); moreover, it exhibited the same level of
inhibitory potency against gt-2a replicon as the parent C6-
sulfonamide compound 6 (Table 2), a result that was not
anticipated based on the apparent requirement of a hydrogen
Figure 2. (a) Structure and close-up view of the Gln414 binding pocket region in gt-2a NS5B L30S/5 complex cocrystal structure with key
interacting residues highlighted. (b) Structure and close-up view of the Gln414 binding pocket region in gt-2a NS5B L30S/13 complex cocrystal
structure with key interacting residues highlighted. Hydrogen bonds are denoted with dashed yellow lines, π-stacking, and possible n → π*
interactions are shown as dashed blue and magenta lines, respectively, and water molecules are depicted as red spheres. Image generated with The
̈
PyMOL Molecular Graphics System (v. 2.0, Schrodinger, LLC).
D
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Letter
distance of 2.1 Å. Compound 5 also forms direct interactions
with the side chain of Gln414. In this case, one of the
pyrimidinyl nitrogen atoms hydrogen bonds to the Gln414
side chain amide NH2 moiety. Interestingly, one of the
sulfonamide oxygen atoms is positioned such that an n → π*
interaction20,21 with the Gln414 side chain amide carbonyl is
possible. The SO···CO distance is 3.5 Å, which is close to
the sum of the van der Waals radii of oxygen and carbon (3.22
Å), and the O···CO angle is 108.0°, a value consistent with
In conclusion, the poor absorption problem presented by P-
gp and BCRP transporter-mediated efflux of the benzofuran
class of pan-genotypic HCV NS5B replicase palm site
inhibitors, in which both the MW and TPSA are high, was
resolved by reduction of polar surface area in conjunction with
decreasing hydrogen bond acceptor strength. The permeability
of these inhibitors was also enhanced by their ability to form
intramolecular hydrogen bonds. The benchmark compound 12
emerging from this study exhibited promising pharmacokinetic
parameters and oral bioavailability in rats. The interactions of
the N-ethylmethylsulfonamide group in the gt-2a Gln414
binding pocket were fulfilled by a simpler N-ethylamino
moiety, enabling the preservation of potent activity toward gt-
2a in 12 and functional group efficiency in optimization. These
SAR insights and the structural information derived from
NS5B/inhibitor cocrystals provided a clear path for the
optimization of second generation pan-genotypic inhibi-
tors.18,19
Burgi−Dunitz trajectory (Figure 3). These compensatory
̈
ASSOCIATED CONTENT
* Supporting Information
■
S
The Supporting Information is available free of charge on the
ACS Publications website at DOI: 10.1021/acsmedchem-
lett.8b00379.
Anti-HCV activity profile of compound 12, compound
synthesis and characterization, biological methods, CC50
and assay standard deviation (SD) data, in vivo rat
pharmacokinetic studies, Caco-2 permeability assay,
production of the recombinant protein, crystallization
conditions, crystallographic methods, crystallographic
statistics for complexes of HCV gt-2a NS5B protein, and
a figure depicting the complexes with ligand electron
density contours (PDF)
Figure 3. Detailed view of intra- and intermolecular hydrogen bonds
involving the ligand pyrimidinylcyclopropyl amide and N-ethyl-
methylsulfonamide moieties of 5 and the putative n → π* interaction
formed between Gln414 and the ligand in the gt-2a NS5B L30S/5
complex. Hydrogen bonds and the n → π* interaction are depicted as
dashed yellow and magenta lines, respectively. Distances are in
angstroms and the interaction angle is reported in degrees. Image
created with The PyMOL Molecular Graphics System (v. 2.0,
̈
Schrodinger, LLC).
Accession Codes
The PDB codes for the X-ray crystal structures of HCV gt-2a
NS5B L30S in complex with 5 and 13 are 5QJ0 and 5QJ1,
respectively.
interactions in the vicinity of Gln414, a key residue that
differentiates gt-2 NS5B protein from that of gt-1, as observed
in the structures of the complexes of 5 and 13 with the gt-2a
NS5B L30S protein, may account for the improved coverage of
gt-2a observed for the C6-sulfonamide and C6-N-ethylamino
des-sulfonamide analogs.
AUTHOR INFORMATION
■
Corresponding Author
Interestingly, as depicted in the structure of the gt-2a NS5B
L30S/5 complex (Figure 2a), the inhibitor adopts a
conformation in which the pyrimidinylcyclopropyl amide N−
H engages in an intramolecular hydrogen bond with one of the
oxygen atoms of the C6-sulfonamide group (N−H···OS
distance = 2.1 Å) forming an unusual 11-membered ring
pseudoheterocycle. Without the strong hydrogen bonding
acceptor SO2 group, the pyrimidinylcyclopropyl amide N−H
points toward the 4′-methoxy oxygen atom, forming an
alternative intramolecular hydrogen bond, as represented in
compound 13 in the gt-2a NS5B L30S/13 complex structure
(Figure 2b). The propensity of both the sulfonamide and the
N-ethylamino compounds to form intramolecular hydrogen
bonds likely contributes to the good to high intrinsic
membrane permeability, as suggested by their PAMPA values
described above, that potentially facilitate the absorption
process.13,15,22,23 However, it is interesting to note that such
potential to form intramolecular hydrogen bonds in com-
pounds 6 to 10 did not alleviate the transporter-mediated
efflux. In this example, it is the combination of the TPSA and
HBA strength properties of the molecule that determines the
efflux susceptibility.
*E-mail: kapsun.yeung@bms.com. Phone: +1 (203) 677-6897.
ORCID
Kap-Sun Yeung: 0000-0003-3995-6555
Kyle Parcella: 0000-0003-0852-344X
Nicholas A. Meanwell: 0000-0002-8857-1515
Notes
The authors declare no competing financial interest.
ACKNOWLEDGMENTS
■
We would like to thank colleagues in Lead Profiling for
collecting in vitro profiling data, and the Discovery Chemistry
Synthesis group for scale-up synthesis of intermediates. Use of
the IMCA-CAT beamline 17-ID at the Advanced Photon
Source was supported by the companies of the Industrial
Macromolecular Crystallography Association through a con-
tract with Hauptman-Woodward Medical Research Institute.
This research used resources of the Advanced Photon Source,
a U.S. Department of Energy (DOE) Office of Science User
Facility operated for the DOE Office of Science by Argonne
National Laboratory under Contract No. DE-AC02-
06CH11357. X-ray data for gt-2a NS5B/5 were collected by
E
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Letter
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DOI: 10.1021/acsmedchemlett.8b00379
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