Efficient synthesis of 2′-C-α-aminomethyl-2′- deoxynucleosides

Article abstract of DOI:10.1039/c2cc34556k

Starting from methyl 3,5-di-O-benzyl-2-keto-α-d-ribofuranoside, a convergent, six-step synthesis is developed to give efficiently all four 2′-C-α-aminomethyl-2′-deoxynucleosides (U, C, A, G) in 38%, 42%, 12%, 12% yield, respectively. Convergence is achieved by the glycosylation of persilylated nucleobases with methyl 2-α-phthalimidomethyl ribofuranoside.

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COMMUNICATION
Efficient synthesis of 20-C-a-aminomethyl-20-deoxynucleosideswzy
Nan-Sheng Li*a and Joseph A. Piccirilli*ab
Received 25th June 2012, Accepted 12th July 2012
DOI: 10.1039/c2cc34556k
Starting from methyl 3,5-di-O-benzyl-2-keto-a-D-ribofuranoside,
a convergent, six-step synthesis is developed to give efficiently all
four 20-C-a-aminomethyl-20-deoxynucleosides (U, C, A, G) in
38%, 42%, 12%, 12% yield, respectively. Convergence is
achieved by the glycosylation of persilylated nucleobases with
methyl 2-a-phthalimidomethyl ribofuranoside.
efficiently. We converted methyl 3,5-di-O-benzyl-2-keto-a-D-
ribofuranoside (1)11 in two steps (via Wittig and hydroboration
reactions) to 2-a-hydroxymethyl ribofuranose derivative 2 in
52% yield (Scheme 1). Under Mitsunobu conditions,12 2 reacts
to form 2-a-phthalimidomethyl ribofuranoside 3 in 90% yield.
Deprotection of 3 with methylamine in ethanol13 followed by
reaction with S-ethyl a,a,a-trifluoroacetate gives 2-a-(a,a,a-
trifluoroacetyl)-aminomethyl ribofuranoside 4 in 68% yield.
Glycosylation of persilylated pyrimidine nucleobases (uracil
and N4-acetylcytosine) with 20-aminomethyl ribofuranoside
derivatives 3 and 4 in the presence of SnCl4 gives the corres-
ponding uridine and cytidine derivatives with high b-selectivity
(91 : 9 to 98 : 2) (Scheme 2). Although yield and b-selectivity
were higher using 2-a-phthalimidomethyl ribofuranoside 3,
the high b-selectivity of the glycosylation reaction indicates
that both phthalyl and trifluoroacetyl groups can stabilize the
1-oxocarbenium ion intermediates and direct the nucleobase
to attack the b face of the ribofuranoside.
The chemical synthesis of modified nucleosides and the
incorporation of nonnatural nucleosides into RNA have become
increasingly important in medical applications and as reagents for
structure and function investigations of RNAs.1,2 A new class of
nucleoside analogues, 20-C-a-aminomethyl-20-deoxynucleosides
(Fig. 1, IV), which hold both amino and 20-alkyl functionalities,
have features common to both 20-a-amino-20-deoxynucleosides3
and 20-C-branched nucleosides,4–8 and therefore might possess
interesting antiviral and anticancer activities. Additionally,
oligonucleotides containing 20-aminomethyl nucleosides provide
a new opportunity for regiospecific chemoligation reaction with
a variety of amine-reactive probes to attach desired reporter
groups.9,10 Here we describe an efficient approach to all of the
four potentially important nucleoside analogues: 20-C-a-amino-
methyl-20-deoxynucleosides (A, C, G, U).
Glycosylation of purine nucleobases generally requires nonpolar
solvent and the presence of TMSOTf to achieve the optimized
yield and b-9 selectivity.14,15 Glycosylation of persilylated
We designed a convergent approach (glycosylation of various
nucleobases with 20-aminomethyl ribose derivatives) to prepare
the four 20-C-a-aminomethylnucleosides (U, C, A and G)
Scheme 1 Synthesis of glycosylation reagents 3 and 4.
Fig. 1 Structures of ribonucleosides (I) and modified nucleosides
(II–IV).
a Department of Biochemistry & Molecular Biology, University of
Chicago, 929 East 57th Street, Chicago, Illinois 60637, USA
b Department of Chemistry, University of Chicago, 929 East 57th
Street, Chicago, Illinois 60637, USA. E-mail: nli@uchicago.edu,
jpicciri@uchicago.edu; Fax: +1 773 702 0438;
Tel: +1 773 702 5236, +1 773 702 9312
w Dedicated to the memory of Professor H. Gobind Khorana.
z This article is part of the ‘Nucleic acids: new life, new materials’
web-themed issue.
y Electronic supplementary information (ESI) available: Experimental
procedures and characterization of all new compounds. 1H and 13C
NMR spectra of 15a–d. See DOI: 10.1039/c2cc34556k
Scheme 2 Glycosylation of persilylated pyrimidine nucleobases with
3 and 4.
c
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Table 1 Synthesis of 8 by the glycosylation of 7 with 3
Table 2 Synthesis of 10 by the glycosylation of 9 with 3
Entry
Solvent
Reaction conditions
Yield (%)
Entry Solvent
Reaction conditions
Yield (%) b/a
1
2
ClCH2CH2Cl
Toluene
ClCH2CH2Cl
CH2Cl2
ClCH2CH2Cl
rt, 62 h; reflux, 1 h
80 1C, 3 h
reflux, 4 h
reflux, 4 h
reflux (80 1C), 1.5 h
45
15
—
—
58
1
ClCH2CH2Cl Reflux, 2 h
ClCH2CH2Cl rt, 48 h; reflux, 1 h
71
68
—
87 : 13
2
91 : 9
—
96 : 4
3a
4b
5
3a
4
CH3CN
Reflux, 1.5 h
ClCH2CH2Cl 50 1C, 3.5 h; reflux, 2 h 68
a
Lower yield and lower selectivity.
a
Product is decomposed significantly and the yield is not determined.
Reaction is incomplete and the yield is not determined.
b
N6-benzoyladenosine with 3 or 4 in the presence of TMSOTf
in refluxing 1,2-dichloroethane for two hours yields the corres-
ponding products in low yields (only B35%) and moderate
selectivity, with 3 giving better b-selectivity than with 4
(b/a 80 : 20 and 65 : 35, respectively). N6-Octanoyladenosine
has been used in the glycosylation with 1-O-acetyl-2,3,5-tri-
O-benzoyl-b-D-ribofuranose for its better solubility than
N6-benzoyladenosine.16 It also has been used in the transgly-
cosylation of 20-a-trifluoroacetylaminouridine.17 We then found
that use of persilylated N6-octanoyladenosine 7 significantly
improved yield and b-selectivity of the glycosylation with 3. The
results are shown in Table 1. Glycosylation in nonpolar solvent
(1,2-dichloroethane) generally yields the glycosylation product
in high b-selectivity. In contrast, glycosylation in the polar
solvent, acetonitrile, gave the product in low yield and low
selectivity (Table 1, Entry 3). The optimized results with 68%
yield and B96 : 4 b-selectivity were achieved using 1,2-dichloro-
ethane at 50 1C for 3.5 hours, then at reflux for 2 hours (Table 1,
entry 4).
Table
glycosylation products
3
Removal of the 30,50-di-O-benzyl groups of various
Yield
Pb (%)
Entry Sa Method Reaction conditions
1
2
5a A
5a B
20% Pd(OH)2/C, H2, CH3CO2C2H5, 11a 62
rt, 22 h
1. BCl3/CH2Cl2, ꢀ78 1C, 4 h;
2. MeOH/CH2Cl2 (v/v 1 : 1),
ꢀ78 1C-rt, 30 min
11a 90
3
4
5b A
6a B
20% Pd(OH)2/C, H2, CH3CO2C2H5, 11b 81
rt, 17 h
1. BCl3/CH2Cl2, ꢀ78 1C, 2.5 h; 0 1C, 12 99
30 min;
2. MeOH, 0 1C-rt, 12 h
20% Pd(OH)2/C, H2, CH3CO2C2H5, —
5c
6d
7e
6b A
—
—
—
rt, 6 h
20% Pd(OH)2/C, H2, CH3CO2C2H5, —
rt, 15 h
8
8
A
B
1. BCl3/CH2Cl2, ꢀ78 1C, 1 h;
2. MeOH/CH2Cl2 (v/v 1 : 1),
ꢀ78 1C-rt, 30 min
—
8
8
8
B
B
1. BCl3/CH2Cl2, ꢀ78 1C, 1 h;
2. Et3N/MeOH/CH2Cl2, 78 1C-rt,
30 min
13 47
13 52
The glycosylation of persilylated N2-acetylguanosine (2 equiv.)
with glycosylating agent 3 in the presence of TMSOTf (2 equiv.)
gave complex results even in nonpolar solvents such as
1,2-dichloroethane, toluene and benzene. Reactions gave all
of four possible products (b-9, b-7, a-9 and a-7) in low yields
and in low selectivity. However, using persilylated N2-acetyl-
O6-(diphenylcarbamoyl)-guanine (9)18,19 glycosylation in the
presence of TMSOTf gave the b-9 isomer product 10 exclusively.
The glycosylation of persilylated N2-acetyl-O6-(diphenylcarba-
moyl)-guanine (9) with 3 under various conditions are shown
in Table 2. The optimized yield (58%) is obtained if the
glycosylation of N2-acetyl-O6-(diphenylcarbamoyl)-guanine
(9) with 3 is carried out in refluxing 1,2-dichloroethane
(80 1C oil bath) for 1.5 hours (Table 2, entry 5).
9
1. BCl3/CH2Cl2, ꢀ78 1C, 1 h;
2. NaHCO3 (aq.), 78 1C-rt, 30 min
10
10 A
20% Pd(OH)2/C, H2, CH3CO2C2H5, 14 49
rt, 15 h
5% Pd/C, H2, CH3CO2C2H5, rt, 15 h 14 49
1. BCl3/CH2Cl2, ꢀ78 1C, 1 h;
2. Et3N/MeOH/CH2Cl2, 78 1C-rt,
30 min
11
12
10 A
10 B
14 19
a
b
S: Starting materials (substrates). P: Products. Cytosine ring also
c
d
reduced. The results are complicated. Product is decomposed.
e
derivatives (6a and 6b), Method A in addition to the debenzyla-
tion, reduced the 5,6-double bond of the cytosine ring (entry 5).
In contrast, Method B removes the benzyl and acetyl groups
from cytidine derivative 6a without affecting the 5,6-double
bond to generate 12 in 99% yield (entry 4).
To remove the 30,50-di-O-benzyl groups from the glycosylation
products, we tested two debenzylation methods (Method A:
Pd(OH)2 or Pd catalyzed hydrogenation,20 and Method B: boron
trichloride (Lewis acid) induced benzyl ether cleavage21–23
)
Attempts to remove the benzyl groups from adenosine
derivative 8 by Method A gave complex results (entry 6).
However, for Method B, if a base such as triethylamine or aqueous
sodium bicarbonate was added to quench the reaction (entries 8, 9),
the adenosine derivative 8 undergoes selective debenzylation to give
compound 13 in 47–52% yield, Without the base, the product
(Table 3). Method A removed the benzyl groups from uridine
derivatives 5a and 5b to give compounds 11a and 11b in 62% and
81% yield, respectively (Table 3, entries 1, 3). The uridine
derivative 5a can also be debenzylated by Method B to give
compound 11a in 90% yield (entry 2). However, for the cytidine
c
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convergently synthesized by the glycosylation of persilylated
nucleobases with 2-a-phthalimidomethyl ribofuranoside 3 or
2-a-trifluoroacetylaminomethyl ribofuranoside 4. As glycosylating
reagent, 2-a-phthalimidomethyl ribofuranoside 3 gives better
results (yield and selectivity) than 2-a-trifluoroacetylamino-
methyl ribofuranoside 4. Debenzylation of the glycosylation
products was accomplished either by hydrogenation (Method A)
for guanosine derivatives or by boron trichloride induced
benzyl ether cleavage (Method B) for uridine, cytidine
and adenosine derivatives. These analogues will enable new
strategies for investigating structure-function relationships
in RNA.
This work was supported by an N.I.H. grant to J.A.P.
(1R01AI081987).
Notes and references
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Scheme 3 Synthesis of 20-a-aminomethyl-20-deoxynucleosides.
decomposes (entry 7). For the guanosine derivative 10 under the
same conditions, Method B gives the corresponding debenzylated
product 14 in only 19% yield (entry 12). In contrast, Method A
converts the guanosine derivative 10 to compound 14 in 49%
yield (entries 10, 11). To complete the synthesis, 11a, 11b, 12, 13
and 14 were treated with n-butylamine in ethanol at 55 1C to give
the final 20-C-a-aminomethyl-20-deoxynucleosides 15 (A, C, G,
U) in 71–97% yields (Scheme 3).
NOESY experiments confirmed the structures of b-anomers.
For the uridine derivative 5a, we observed strong NOE between
20-H (d 3.02) and 30-H (d 4.22), between 10-H (d 6.14) and
CH2N (d 4.00) but no NOE between 10-H and 20-H. For
cytidine derivative 6a, we observed strong NOE between
between 10-H (d 6.04) and CH2N (d 4.00) but no NOE between
10-H and 20-H (d 2.97). For adenosine derivative 8, we also
observed strong NOE between 10-H (d 6.34) and CH2N (d 4.16)
but only weak NOE between 10-H and 20-H (d 3.84). For 20-C-
a-aminomethyl-20-deoxyguanosine (15d), we observed strong
NOE between 10-H (d 5.81) and 40-H (d 4.08), but no NOE
between 10-H (d 5.81) and 30-H (d 4.48); we also observed
stronger NOE between 10-H (d 5.81) and CH2N (d 3.30) but
weak NOE between 10-H (d 5.81) and 20-H (d 3.12). These data
suggest that 10-H and 20-CH2N reside on the same face of
the ribose ring and therefore the glycosylation products have
the b-configuration.
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Chem. Commun.