Syntheses of 5′-amino-2′,5′-dideoxy-2′,2′- difluorocytidine derivatives as novel anticancer nucleoside analogs

Article abstract of DOI:10.1016/j.tetlet.2013.11.018

A novel class of 5′-amino-2′,5′-dideoxy-2′, 2′-difluorocytidine derivatives has been synthesized in order to identify anticancer nucleoside analogs. Several synthetic routes were devised and implemented which relied upon either S_N2 displacement

Full text of DOI:10.1016/j.tetlet.2013.11.018

Tetrahedron Letters 55 (2014) 598–602
Contents lists available at ScienceDirect
Tetrahedron Letters
journal homepage: www.elsevier.com/locate/tetlet
Syntheses of 50-amino-20,50-dideoxy-20,20-difluorocytidine derivatives
as novel anticancer nucleoside analogs
c
c
c
Marc A. Labroli a, Michael P. Dwyer b, , Ruichao Shen , Janeta Popovici-Muller , Qinglin Pu ,
⇑
Judson Richard c, Kristen Rosner c, Kamil Paruch a, Timothy J. Guzi c
a Merck Research Laboratories, 2000 Galloping Hill Road, Kenilworth, NJ 07033, USA
b Merck Research Laboratories, PO Box 2000, 126 E. Lincoln, Rahway, NJ 07065, USA
c Merck Research Laboratories, 33 Louis Pasteur Avenue, BMB-3, Boston, MA 02115, USA
a r t i c l e i n f o
a b s t r a c t
A novel class of 50-amino-20,50-dideoxy-20,20-difluorocytidine derivatives has been synthesized in order to
identify anticancer nucleoside analogs. Several synthetic routes were devised and implemented which
relied upon either SN2 displacement or reductive amination to provide the desired derivatives.
Ó 2013 Elsevier Ltd. All rights reserved.
Article history:
Received 19 October 2013
Accepted 5 November 2013
Available online 14 November 2013
Keywords:
Nucleoside
Gemcitabine
Reductive amination
Anticancer
NH2
HCl
NHBz
iii
20-Deoxy-20,20-difluorocytidine, the antimetabolite nucleoside
known as gemcitabine (1), is approved for the treatment of pancre-
atic, breast, and non-small cell lung cancers either as a single agent
or in combination with other chemotherapeutic agents.1 Gemcita-
bine acts as a prodrug which is intracellularly phosphorylated to its
active diphosphate and triphosphate intermediates.2 Gemcitabine
triphosphate competes with deoxycytidine triphosphate for incor-
poration into DNA which results in termination of DNA polymeri-
zation.3 The diphosphate intermediate effectively inhibits
ribonucleotide reductase (RRM1) which leads to the depletion of
the deoxynucleotide pool and halt of DNA synthesis.4 Since the dis-
covery of gemcitabine by Hertel et al.,5 numerous gemcitabine
derivatives have been synthesized in search for new anticancer
or antiviral agents. For example, the base-modified gemcitabine
derivatives include adenosine, guanine, and uracil analogs.6 The
ribose-modified derivatives include 40-azido analogs,7 40-allene
substituted analogs,8 30-deoxy analogs,9 and thio/aza/carbocyclic
analogs.10 However, to our knowledge, the 50-amino derivatives
of gemcitabine, a novel and potentially biologically interesting
class of compounds, have not been actively investigated. As part
of our program to identify novel and selective anticancer com-
pounds, we initiated efforts to develop efficient syntheses of 50-
amino-20,50-dideoxy-20,20-difluorocytidine derivatives in order to
profile this novel series. Herein, we describe our chemistry efforts
to prepare 50-amino derivatives of gemcitabine.11
O
O
N
N
N
N
i, ii
HO
HO
TBSO
O
O
O
F
F
HO
BzO
F
F
2
1
NHBz
NHBz
O
O
N
N
N
N
TsO
iv
O
F
F
BzO
BzO
F
F
3
4
v or vi
vii, viii
NH2
N
NH2
R1
O
N
O
N
O
N
N
R
v
I
O
F
HO
F
HO
F
F
6a-i
5
Scheme 1. Reagents and conditions: (i) TBSCl, imidazole, DMF, rt, 12 h, 92%; (ii)
BzCl, DMAP, pyridine, rt, 12 h, 90%; (iii) TBAF, THF, 0 °C, 3.5 h then AcOH, 85%; (iv)
TsCl, pyridine, Et3N, 85%; (v) HNRR1, DMF, 90–100 °C; (vi) NH3, MeOH, 58%; HNRR1,
DMF; (vii) MeP(OPh)3I, DMF, 60 °C, 84%; (viii) NH3, MeOH, 92%.
Our initial route to the 50-amino-20,50-dideoxy-20,20-dif-
luorocytidine analogs is outlined in Scheme 1. The synthesis began
⇑
Corresponding author. Tel.: +1 (732) 594 1733.
E-mail address: michael.dwyer@merck.com (M.P. Dwyer).
0040-4039/$ - see front matter Ó 2013 Elsevier Ltd. All rights reserved.
http://dx.doi.org/10.1016/j.tetlet.2013.11.018

M. A. Labroli et al. / Tetrahedron Letters 55 (2014) 598–602
599
with protection of the 50-hydroxy group of gemcitabine hydrochlo-
ride (1) as the tert-butyldimethylsilyl (TBDMS) ether followed by
global benzoyl protection to afford compound 2. Desilylation of
the silyl ether was achieved with TBAF at 0 °C to provide alcohol
312 which was converted to tosylate 4 under standard conditions.
Treatment of intermediate 4 with excess amine coupling partner
at 100 °C promoted both tosylate displacement and benzoyl group
cleavage to afford products 6aꢀd,i (Table 1). Alternatively, this
protocol could be carried out in a step-wise fashion by first remov-
ing the benzoyl groups of 4 by treatment with 7 N ammonia in
methanol to provide the penultimate product. Treatment of the
resulting tosylate with excess amine provided the desired com-
pounds 6e,f (Table 1). While the displacement reaction with the
amine coupling afforded modest yields of the desired products
(Table 1), multiple purifications were required to completely
remove the resultant TsOH from desired product. In order to
Table 1
Preparation of 6a–i by SN2 substitution
Entry
Amines
Product
Yield (%)
20a
NH2
NH2
NH2
NH2
NH2
NH2
O
O
O
O
O
N
N
N
N
H
6a
O
O
F
HO
HO
HO
F
F
H
N
N
O
O
54a
32a
19a
84a,c
13a
53b
55b
N
6b
6c
6d
6e
6f
F
NH2
O
O
N
N
N
H
O
F
F
N
NH2
NH2
O
N
N
N
N
N
H
O
F
HO
HO
F
F
O
N
N
H
O
N
F
NH2
O
N
NH2
N
H
O
O
O
N
F
HO
F
Me
NH2
Me
Me
N
Me
N
O
N
N
NH2
N
H
6g
6h
N
O
F
HO
F
NH2
NH
N
N
O
N
N
N
N
O
N
F
HO
F
O
O
N
NH2
NH2
O
N
N
N
6i
N
H
53a
O
F
HO
F
a
b
c
Yield for one step.
Combined yield for two steps.
Rxn heated at 40 °C.

600
M. A. Labroli et al. / Tetrahedron Letters 55 (2014) 598–602
NHBz
NHBz
NHBz
iii
amines at 90 °C provided 50-amino-20,50-dideoxy-20,20-difluorocyti-
dine 6g,h in respectable yields (Table 1). The use of iodide 5 al-
lowed us to scale back the number of equivalents of amine for
the displacement reaction while making the purification of the fi-
nal analogs much easier.
O
3
O
7
N
N
N
N
N
HO
N3
i, ii
O
O
F
F
BzO
BzO
R1
F
F
While the SN2 strategy depicted in Scheme 1 represented a con-
cise preparation of 50-amino-20,50-dideoxy-20,20-difluorocytidine
derivatives, we decided to pursue other alternatives due to the
poor to modest yields for the final compounds. Toward this end,
a reductive amination approach was explored not only to improve
the yield of the final 50-amino-20,50-dideoxy-20,20-difluorocytidine
analogs but to broaden the scope of coupling partners. As shown
in Scheme 2, alcohol 3 was converted to the mesylate followed
by treatment with NaN3 to afford the corresponding azide 7. Treat-
ment of azide 7 with trimethylphosphine provided amine interme-
diate 8 which was used directly without silica gel purification to
prevent benzoyl transfer to the 50-amino group. Treatment of 8
with various aldehydes under reductive amination conditions
(NaBH3CN) afforded the N-alkylated products which were treated
with 7 M NH3 in MeOH to afford 50-substituted amino analogs
NH2
O
N
O
O
N
N
N
H2N
iv, v
R
O
F
HO
F
F
BzO
F
9a−f
8
Scheme 2. Reagents and conditions: (i) MsCl, Et3N, pyridine, 89%; (ii) NaN3, DMF,
70 °C, 98%; (iii) Me3P, MeCN/H2O, 85%; (iv) RCHO, NaBH3CN, MeOH, AcOH, 35–75%;
(v) 7 M NH3/MeOH, 45–85%.
circumvent this issue, alcohol 3 was converted to the iodide inter-
mediate 5 using methyltriphenoxyphosphonium iodide followed
by benzoyl deprotection. Treatment of iodide 5 with 3 equiv of
Table 2
Reductive amination protocol/deprotection from 8 to afford 9a–f (Scheme 2)
Entry
Aldehydes
Product
Yield (%) of step iva/step v
O
NH2
NC
F
O
NC
H
N
N
N
H
8a
72/82
66/70
68/45
69/84
O
F
F
HO
F
Me
O
NH2
Me
O
F
N
N
H
N
O
N
H
b
c
N
O
O
F
HO
F3C
F3C
NH2
O
O
N
N
N
H
N
H
O
F
HO
O
F
O
NH2
H
N
O
N
H
d
O
O
F
F3C
Me
HO
F3C
F
O
NH2
Me
N
N
O
O
H
N
N
N
N
N
H
O
e
59b/50
F
HO
F
F
F
F
F
NH2
Me
Me
f
35/85
O
N
N
N
N
O
N
N
N
O
O
F
HO
F
10
a
Amine 8 was used as the crude product of reduction of azide 7.
Prepared by one-pot stepwise reductive amination.
b

M. A. Labroli et al. / Tetrahedron Letters 55 (2014) 598–602
601
NH2
NHBoc
O
O
O
O
N
N
N
N
BzO
BzO
i, ii
HO
3 steps
O
O
F
F
F
BzO
BzO
HO
F
F
F
13
12
11
NHBoc
NHBoc
+
O
O
N
N
N
N
iii
I
I
O
O
F
F
HO
HO
F
15
F
14
iv, v
NHBoc
NH2
Me
O
O
N
N
F
N
N
H2N
vi, vii
R
N
H
O
O
F
F
HO
HO
F
17a−d
16
Scheme 3. Reagents and conditions: (i) Boc2O, DMAP, THF, 75%; (ii) Et3N/MeOH/H2O, rt, 12 h, 92%; (iii) MeP(OPh)3I, DMF, rt, 1 h, 31% (14), 45% (15); (iv) NaN3, DMF, 40 °C,
quant.; (v) Me3P, MeCN/H2O, 73%; (vi) RCOMe, Ti(i-OPr)4, then NaBH3CN, 51–68%; (vii) TFA, CH2Cl2, 71–98%.
Table 3
Reductive amination/deprotection protocol from 16 to afford 17a–d (Scheme 3)
Entry
Ketones
Product
Yield (%) of step via/step vii
Me
NH2
Me
N
O
O
N
O
N
N
O
N
H
17a
68/71
O
F
HO
F
Me
NH2
Me
O
N
O
O
N
O
N
N
N
H
17b
17c
60/95
50/71
O
Me
F
HO
Me
F
Me
NH2
Me
S
N
O
S
N
O
N
N
N
H
O
F
HO
Me
F
Me
NH2
O
N
O
N
O
N
H
17d
51/98
F
HO
F
9aꢀf shown in Table 2. The reductive amination/deprotection
protocol worked well for both aromatic and aliphatic aldehydes
(entries 9aꢀd) with respectable yields observed for both steps
(Table 2). Additionally, this protocol was adapted to a stepwise
format by employing two distinct aldehydes to afford 9e or by
employing a dicarbonyl substrate 10 to afford bicyclic derivative
9f.
increased steric hindrance of the ketone coupling partners, it was
rationalized that a strong Lewis acid may be required to promote
the initial imine formation for these substrates. It was rationalized
that one might need an alternative 50-amino intermediate with a
suitable protecting group for harsher imine formation conditions
due to potentially labile 30-benzoate of 8. Additionally, it was desir-
able to develop a flexible route which could allow for rapid base
modification (other than cytosine) while at the same time
circumventing the need to use expensive gemcitabine as a starting
While the reductive amination approach utilizing amine 8
proceeded well with aldehydes, this protocol yielded very low
yields of
a-substituted amine products when ketones were used
material. Based on the above considerations,
a
second
as coupling partners. Owing to the reduced electrophilicity and
reductive amination synthetic route was developed to prepare

602
M. A. Labroli et al. / Tetrahedron Letters 55 (2014) 598–602
50-amino-20,50-dideoxy-20,20-difluorocytidine derivatives derived
from ketone coupling partners.
sulfonamides, and ureas. The biological activity associated with
this novel class of 50-amino-20,50-dideoxy-20,20-difluorocytidine
derivatives will be reported in due course.16
The synthesis started with commercially available 2-deoxyl-
2,2-difluoro-D-erythro-pentafuranous-1-ulose-3,5-dibenzoate (11)
(Scheme 3). Following the protocol described by Chou et al.,13 lac-
tone 11 was converted to nucleoside 12 in three steps as a mixture
References and notes
of both anomers (b:a = 1:1.5). The 4-amino group of the cytosine
1. (a) Burris, H. A., III; Moore, M. J.; Andersen, J.; Green, M. R.; Rothenberg, M. L.;
Modiano, M. R.; Cripps, M. C.; Portenoy, R. K.; Storniolo, A. M.; Tarassoff, P.;
Nelson, R.; Dorr, F. A.; Stephens, C. D.; Von Hoff, D. D. J. Clin. Oncol. 1997, 15,
2403; (b) Manegold, C. Expert Rev. Anticancer Ther. 2004, 4, 345; (c) Heinemann,
V. Expert Rev. Anticancer Ther. 2005, 5, 429.
2. Heinemann, V.; Hertel, L. W.; Grindey, G. B.; Plunkett, W. Cancer Res. 1988, 48,
4024.
3. Huang, P.; Chubb, S.; Hertel, L. W.; Grindey, G. B.; Plunkett, W. Cancer Res. 1991,
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4. (a) van der Donk, W. A.; Yu, G.; Pérez, L.; Sanchez, R. J.; Stubbe, J.; Samano, V.;
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Yokoyama, K.; Yu, G.; Griffin, R. G.; Bar, G.; Stubbe, J. Biochemistry 2009, 48,
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5. Hertel, L. W.; Kroin, J. S.; Misner, J. W.; Tustin, J. M. J. Org. Chem. 1988, 53, 2406.
6. (a) Hertel, L. W.; Grossman, C. S.; Kroin, J. S.; Mineishib, S.; Chubb, S.; Nowak,
B.; Plunkett, W. Nucleos. Nucleot. 1989, 8, 951; (b) Fahrig, R.; Lohmann, D.; Rolfs,
A.; Dieks, H.; Teubner, J.; Heinrich, J.-C. WO 2008017515; Chem. Abstr. 2008,
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was protected as the Boc derivative followed by hydrolysis of the
30,50-dibenzoate under mild basic conditions to provide diol 13.
Based on the selective iodination protocol of thymidine reported
by Verheyden and Moffatt,14 treatment of diol 13 with methyltri-
phenoxyphosphonium iodide afforded only the 50-iodo nucleoside
anomers 14 and 15. Presumably, the selectivity of this transforma-
tion is driven by the reduced nucleophilicity of the 30-hydroxyl
caused by the geminal di-fluorine atoms which yields only the
50-iodo products. Fortunately, the 50-iodo anomers 14 and 15 could
be separated by regular silica gel chromatography. Treatment of
b-anomer 14 with sodium azide followed by reduction furnished
the 50-amino intermediate 16. Treatment of amine 16 with aro-
matic ketones in the presence of Ti(i-OPr)415 followed by treatment
with NaBH3CN and Boc deprotection leads to a
-methylated 50-ami-
no compounds 17a–d as a mixture of diastereomers in moderate to
good yields (Scheme 3). The specific yields for the reductive amina-
tion step as well as deprotection are detailed in Table 3.
9. Hertel, L. W.; Grossman, C. S.; Kroin, J. S. EP 329348, 1989; Chem. Abstr. 1989,
112, 56592.
In summary, we have described three unique synthetic routes
for the syntheses of a novel class of 50-amino-20,50-dideoxy-20,20-
difluorocytidine derivatives. The first route relied upon a SN2 dis-
placement of either a 50-tosylate or 50-iodide intermediate using
excess amine as coupling partners. To circumvent the modest
yields and challenging purifications of final products using the first
route, a second route was developed which relied upon a reductive
amination of a 50amino derivative with aldehydes to afford a wide
variety of 50-N-alkylated derivatives. Finally, the final route relies
upon a reductive amination step with ketones but also offers the
ability to change the base with fewer manipulation of protecting
groups than the previous routes. While the 50-amino intermediates
8 and 16 were critical components for the reductive amination
protocols described, one could envision the utility of these materi-
als for preparation of such non-basic analogs such as amides,
10. (a) Qiu, X.-L.; Xu, X.-H.; Qing, F.-L. Tetrahedron 2010, 66, 789; (b) Devos, R.;
Dymock, B. W.; Hobbs, C. J.; Jiang, W.-R.; Martin, J. A.; Merrett, J. H.; Najera, I.;
Shimma, N.; Tsukuda, T. WO 2002018404; Chem. Abstr. 2002, 136, 217007.
11. Guzi, T. J.; Parry, D. A.; Labroli, M. A.; Dwyer, M. P.; Paruch, K.; Rosner, K. E.;
Shen, R.; Popovici-Muller, J. WO 2009061781; Chem. Abstr. 2009, 150, 515402.
12. It was necessary to maintain the desilylation reaction of 2 at low temperature
and higher dilution while quenching the reaction with acetic acid to suppress
gemcitabine 30,50,4-tribenzoate formation as a side product. This side product
could be formed by intermolecular benzoyl transfer from the less stable 30-
benzoate to 50-hydroxyl group.
13. Chou, T. S.; Heath, P. C.; Patterson, L. E.; Poteet, L. M.; Lakin, R. E.; Hunt, A. H.
Synthesis 1992, 565.
14. Verheyden, J. P. H.; Moffatt, J. G. J. Org. Chem. 1970, 35, 2319.
15. Mattson, R. J.; Pham, K. M.; Leuck, D. J.; Cowen, K. A. J. Org. Chem. 1990, 55,
2552.
16. Labroli, M. A.; Dwyer, M. P.; Shen, R.; Popovici-Muller, J.; Pu, Q.; Wyss, D.;
McCoy, M.; Barrett, D.; Davis, N.; Seghezzi, W.; Shanahan, F.; Taricani, L.; Parry,
D.; Guzi, T. J. Bioorg. Med. Chem. submitted for publication.