6′-Fluoro-3-deazaneplanocin: Synthesis and antiviral properties, including Ebola

Article abstract of DOI:10.1016/j.bmcl.2018.10.030

A convenient stereospecific synthesis of 6′-fluoro-3-deazaneplanocin (6) has been accomplished from D-ribose in 15 steps. It is reported to possess significant activity towards Ebola (Zaire, Vero, μM: EC₅₀ < 0.36; CC₅₀ 125; SI > 347)

Full text of DOI:10.1016/j.bmcl.2018.10.030

Accepted Manuscript
6'-Fluoro-3-deazaneplanocin: synthesis and antiviral properties, including Ebo-
la
Chong Liu, Qi Chen, Steven Cardinale, Terry L. Bowlin, Stewart W. Schneller
PII:
S0960-894X(18)30832-1
https://doi.org/10.1016/j.bmcl.2018.10.030
BMCL 26089
DOI:
Reference:
Bioorganic & Medicinal Chemistry Letters
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Received Date:
Revised Date:
Accepted Date:
19 September 2018
18 October 2018
20 October 2018
Please cite this article as: Liu, C., Chen, Q., Cardinale, S., Bowlin, T.L., Schneller, S.W., 6'-Fluoro-3-
deazaneplanocin: synthesis and antiviral properties, including Ebola, Bioorganic & Medicinal Chemistry Letters
(2018), doi: https://doi.org/10.1016/j.bmcl.2018.10.030
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S.W Schneller
1

S.W Schneller
6'-Fluoro-3-deazaneplanocin: synthesis and antiviral properties, including
Ebola
Chong Liua, Qi Chenb, Steven Cardinale c, Terry L. Bowlin c, and Stewart W.
Schneller*a
a Molette Laboratory for Drug Discovery, Department of Chemistry and
Biochemistry, Auburn University, Auburn, Alabama 36849-5312
bDepartment of Chemistry, Slippery Rock University, Slippery Rock, PA 16057
cMicrobiotix, Inc., Worcester, MA 01605
Abstract.
A convenient stereospecific synthesis of 6'-fluoro-3-deazaneplanocin (6) has
been accomplished from D-ribose in 15 steps. It is reported to possess significant
activity towards Ebola (Zaire, Vero, M: EC50 < 0.36; CC50 125; SI >347) with
moderate inhibition of the target enzyme (S-adenosylhomocysteine hydrolase),
which did not correlate directly with its anti-Ebola effects. Compound 6, with
limited cytotoxicity, also displayed activity against measles, H1N1 and Pichinde.
Keywords: carbocyclic nucleosides, 3-deazaneplanocin, Ebola, measles, H1N1,
Pichinde virus, cytomegalovirus.
S-Adenosylmethionine (SAM) is the methyl donor cofactor for numerous
biological methylations.1 As a consequence of this process, SAM is converted to S-
adenosylhomocysteine (SAH) that, subsequently, acts as a biofeedback inhibitor of
the SAM transformations, and, thus, moderating further methylations.2 S-Adenosyl-
*Corresponding author. Tel.: 334-844-6947; fax: 334-844-0239; e-mail: schnest@auburn.edu
2

S.W Schneller
homocysteine hydrolase (SAHase) serves to remove the product SAH permitting
further SAM methylations to occur.2 Thus, SAHase is a significant control agent in
this process and a site to target for affecting biomethylations.2
In this direction, adenine-based carbocyclic nucleosides based on
aristeromycin (1) and neplanocin (2) have arisen as a source of SAHase inhibitors to
control requisite methylations for antiviral drug discovery.3 This has provided the
impetus for our ongoing efforts of varying the known SAHase inhibitors 3-
deazaaristeromycin (3) and 3-deazaneplanocin (4) to achieve agents with non-
cytotoxic antiviral activity.4 In the evolution of our program, we were drawn to a
publication from Jeong's laboratory that, through extensive docking with human
SAHase, reported 6'-fluoroneplanocin (5)5 to have a dual mechanistic means of
inhibition of SAHase. This prompted us to seek the 3-deaza analog (6) as a
mechanistically-diverse target. This is described here.
For this objective, our retro-synthetic thinking was built around
cyclopentenyl iodide 7 (Scheme 1) whose corresponding alcohol (8, Scheme 2)
could, in a protected form, participate in an electrophilic vinyl fluorination with a
subsequent coupling reaction employing a protected 3-deazaadenine.
3

S.W Schneller
Few methods6 exist for preparing -iodo-,-unsaturated ketones (such as
7). For our purpose, the little known oxidative cleavage of alcohols possessing a
quaternary ammonium iodide at their 3-position drew our attention.7 To consider
this approach for obtaining 7 required accessing 9 from 10, which was obtained
from D-ribose by modifying a reported procedure7 where the benzyl group replaced
a trityl to improve the subsequent electrophilic fluorination reaction.8 Oxidation of
9 with pyridinium chlorochromate yielded 7 via oxidative elimination, iodination of
the resultant ,-ketone and then elimination of hydrogen iodide (Scheme 1).
I
MeHN
BnO
IMe3N
BnO
OH
OH
O
PDC/DCM
MeI
K2CO3
THF
BnO
62% for
2 steps
10 to 7
O
O
O
O
O
O
7
10
9
Scheme 1. Synthesis of the requisite vinyl iodide 7.
With 7 available, it was reduced under Luche conditions to alcohol 8
(Scheme 2) that was protected as the t-butyldiphenylsilyl derivative 11.
Fluorination with N-fluorobenzenesulfonimide (NFSI) gave 12. Removal of the
hydroxyl silyl group (to 13)9 followed by a Mitsunobu coupling with the N6,N6-
protected 3-deazaadenine (14)10 produced 15. Boron trichloride followed by
methanolic hydrochloric acid yielded the sought 6.11
4

S.W Schneller
I
F
NaBH4
OR
OR
NFSI
BnO
BnO
7
.
n-BuLi THF
50%
CeCl3 7H2O
O
O
O
O
MeOH
87%
TBAF
12, R=TBDPS
13, R=H
TBDPSCl
imidazole
DCM
97%
8, R=H
THF
88%
11, R=TBDPS
DIAD, PPh3
THF, 82%
N(Boc)2
N(Boc)2
N
N
N
H
14
NH2
N
N
N
F
N
N
N
F
i) BCl3, Et2O
BnO
HO
ii) HCl/MeOH
63%
O
O
HO
OH
15
6
Scheme 2. Synthesis of the target compound 6.
With 6 in hand, it was subjected to antiviral analysis.12 A most pronounced
activity was found versus Ebola (Zaire) as shown in Table 1with data for 2 and 4 and the
1',6'-isoneplanocins 16 and 17, benchmark compounds of our Ebola program. From
that data it can be seen that the anti-Ebola activity of 6 is comparable to 16 and 17
with an SI in line with 17. The only reported antiviral data for 5 is versus vesicular
stomatitis virus (VSV) (EC50 = 0.43 M; CC50 = 40 M). Since VSV and Ebola are (-)-
RNA viruses and VSV has been used as a pseudotype Ebola virus in gene product
studies13a and vaccine development,13b it is tempting to extrapolate the VSV results
for 5 to its potential towards Ebola and conclude that 6 is more promising than 5 as
an anti-Ebola candidate.
Table 1. In vitro activities against Ebola virus (Zaire) (M
Compounds
Cell lines
EC50 (µM)
CC50 (µM)
SI
6
214
Vero
4
414
Vero
2
1612b
Vero
0.38
1.3
174
Vero
<0.32
>100
>313
Vero
<0.36
125
60
15
>1700
>850
>347
3.5
5

S.W Schneller
In the other viral evaluations, 6 displayed activity12 versus (i) measles (Vero 76)
(EC50 < 0.36 M; CC50 115M; SI >320); (ii) influenza A H1N1 (MDCK) (EC50 20.2
M; CC50 133 M; SI 6.6); and (iii) Pichinde (Vero) (EC50 57.6 M; CC50 360 M; SI >
6.3).
To determine if the antiviral effects of 6 could be an outcome of its inhibition of
SAHase,12h this was investigated (Table 2). Its enzymatic IC50 was several fold less than
2, 4, 1612b and 174 (the latter two are lead compounds in our isoneplanocin Ebola studies,
Figure 1) but, surprisingly, 50 times more potent than 5. This latter observation is
problematic and requires further analysis of the reported data5 for 5. With the weaker
effect of 6 than expected based on 2, 4, 16 and 17 on SAHase but yet with its potent anti-
Ebola activity, it could be that 6 is acting by more than simply SAHase inhibition.
Table 2. Inhibitory results against SAHase
Compounds
IC50 (nM)
6
212b 412b
2.0
1612b
174
3.2
55
480
8.69 0.9
0.9
Compound 6 was inactive towards: Chikungunya (Vero 76), dengue virus 2
(Vero 76), polio virus 1 (Vero 76), Punta Toro (vero 76), Rift Valley fever (Vero 76),
Tacaribe (Vero), Venezuelan equine encepthalitis (Vero 76), West Nile (Vero 76),
Yellow fever (Vero), norovirus ( HG 23), human cytomegalovirus (HFF); hepatitis B
virus (2.2.15), adenovirus 5(A549), Utusu (vero 76), and Japanese equine
encephalitis (Vero 76).
6

S.W Schneller
In conclusion, by considering the antiviral properties of 3-deazaneplanocin
(4) and 6'-fluoroneplanocin (5) and combining their structural features, 6'-fluoro-3-
deazaneplanocin (6) arose as a target for our antiviral program. Thus, an efficient
and practical synthesis of 6 is reported in 15 steps from D-ribose. While the
antiviral effects of 6 were not as broad as desired, its potential versus Ebola,
measles, H1N1 and Pichinde (all RNA viruses) suggest 6 as a prototype compound
for antiviral agent design (for example, by combining the features of 6 and 17 as
represented by 18). This is being evaluated in our laboratory at Auburn together
with designing experiments to clarify the mechanism of antiviral action of 6 beyond
SAHase, also, for analog development.
Acknowledgments
We are grateful to the Molette Fund and Auburn University for support of
this research. We are indebted to the NIAID in vitro assay team for the viral data
presented herein: Don Smee, Utah State University; Brent Korba, Georgetown
University; and, Mark Prichard, University of Alabama – Birmingham.
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7

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8

S.W Schneller
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