Synthesis of glycose carbamides and evaluation of the induction of erythroid differentiation of human erythroleukemic K562 cells

Article abstract of DOI:10.1016/j.ejmech.2008.05.001

A series of carbamides derived from 1,2:5,6-di-O-isopropylidene-d-gluco- (1) and d-allofuranose (3) as well as their 5,6-O-deprotected analogues (2 and 4) and methyl 3,4-O-isopropylidene-α- and β-d-galactopyranosides (5 and 6) have been prepared in order to evaluate their ability to induce erythroid differentiation of human erythroleukemic K562 cells. Twenty out of 51 carbamides tested exhibit an appreciable activity as inducers of erythroid differentiation and have been fully characterized and described.

Full text of DOI:10.1016/j.ejmech.2008.05.001

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European Journal of Medicinal Chemistry 44 (2009) 745e754
http://www.elsevier.com/locate/ejmech
Original article
Synthesis of glycose carbamides and evaluation of the induction of
erythroid differentiation of human erythroleukemic K562 cells
a
b
Martina Landi a, Giorgio Catelani a, , Felicia D’Andrea , Eleonora Ghidini ,
*
Gabriele Amari b, Puccini Paola b, Nicoletta Bianchi c, Roberto Gambari
c,**
a Dipartimento di Chimica Bioorganica e Biofarmacia, Universita` di Pisa, Via Bonanno 33, I-56126 Pisa, Italy
b Chiesi Farmaceutici S.p.A., Via Palermo 26/A, I-43100 Parma, Italy
c Dipartimento di Biochimica e Biologia Molecolare, Universita` di Ferrara, Via Borsari 46, I-44100 Ferrara, Italy
Received 17 November 2007; received in revised form 2 May 2008; accepted 2 May 2008
Available online 10 May 2008
Abstract
A series of carbamides derived from 1,2:5,6-di-O-isopropylidene-D-gluco- (1) and D-allofuranose (3) as well as their 5,6-O-deprotected an-
alogues (2 and 4) and methyl 3,4-O-isopropylidene-a- and b-D-galactopyranosides (5 and 6) have been prepared in order to evaluate their ability
to induce erythroid differentiation of human erythroleukemic K562 cells. Twenty out of 51 carbamides tested exhibit an appreciable activity as
inducers of erythroid differentiation and have been fully characterized and described.
Ó 2008 Elsevier Masson SAS. All rights reserved.
Keywords: Carbamides; Monosaccharides; Erythroid differentiation; K562 cells
1. Introduction
K562 cells, leading to a cytoplasmic accumulation of Hb Port-
land (z2g2) and Hb Gower 1 (z2e2) [4e7].
The K562 cell line, isolated and characterized by Lozzio and
Lozzio [1] from a patient with chronic myelogenous leukemia
in blast crisis, has been proposed as a very useful experimental
model system to identify inducers of g-globin gene expression
of possible interest in the therapy of several haematological
diseases, including b-thalassemia and sickle cell anaemia [2].
K562 cells exhibit a low proportion of hemoglobin-synthe-
sizing cells under standard cell growth conditions, but are able
to undergo erythroid differentiation when treated with a variety
of compounds, including short fatty acids, 5-azacytidine, mi-
thramycin, and chromomycin, cisplatin and cisplatin ana-
logues, tallimustine, rapamycin, everolimus, psoralens and
resveratrol [3]. Following erythroid induction, a sharp increase
of expression of human e- and g-globin genes is observed in
Among possible biological response modifiers, one of the
most studied classes of compounds is represented by short
fatty acids, especially butyric and pivalic acids and related
esters. Glycide esters have been initially proposed merely as
convenient prodrugs, able to gradually release the pharmaco-
phore following the in vivo action of esterases [8].
In recent papers regarding the synthesis and evaluation of
some partially acetonated monosaccharide esters of several
fatty acids we have highlighted the role of the structure of ei-
ther the ester or the carbohydrate residue in the antiprolifera-
tive effects and erythroid differentiation activity of K562 cells
[9]. Furthermore, it was observed that some glycose isobuty-
rates and pivaloates stimulate erythroid differentiation to
higher levels than the corresponding free fatty acids [9]. Con-
sequently, we assumed that, at least in the case of isobutyrates
and pivaloates, the biological activity of monosaccharide es-
ters might be related to the whole structure prior to the occur-
rence of the hydrolysis, rather than to the release of the free
fatty acid acting as biologically active component.
* Corresponding author. Tel.: þ39 050 2219678; fax: þ39 050 2219660.
** Corresponding author. Tel.: þ39 0532 424443; fax: þ39 0532 202723.
E-mail addresses: giocate@farm.unipi.it (G. Catelani), gam@unife.it (R.
Gambari).
0223-5234/$ - see front matter Ó 2008 Elsevier Masson SAS. All rights reserved.
doi:10.1016/j.ejmech.2008.05.001

746
M. Landi et al. / European Journal of Medicinal Chemistry 44 (2009) 745e754
Unfortunately, fast in vivo ester hydrolysis on rabbit [10]
and 16 was easily achieved starting from the corresponding
anomeric diols 9 [13] and 10 [13] and employing the same
reaction sequence. Reactions of both 9 and 10 with a slight
excess of p-toluenesulphonyl choride caused the expected
selective [14,15] tosylation of primary OH group. The resulting
6-sulphonates were subjected to nucleophilic displacement
with NaN3, followed by reduction of resulting azides to furnish
15 and 16 in satisfactory overall yield (39 and 45%,
respectively).
The glycose carbamides of type 1, 3, 5 and 6 were prepared
through parallel synthesis by treatment of the four aminated
scaffolds 7, 8, 15 and 16 with an excess of the appropriate
commercial acyl chlorides in methylene chloride in the pres-
ence of PS-piperidinomethyl resin (Scheme 2).
precludes the possibility of any therapeutic use of these glycide
esters. Therefore, we turned our efforts to investigate some
more stable isosters of active glycose esters. Among various
possibilities, we decided to focus on the substitution of the ester
group with an amide one, which would offer the advantage of
a simple synthetic access.
The present paper deals with the synthesis and the biolog-
ical evaluation of part of a library of glycose carbamides 1e6
(Chart 1) in which the carbohydrate scaffoldings were the
same in the previously described biologically active glycide
esters [9]. Besides butyrates and pivaloates, we have selected
a wide set of acyl residues classified into three groups
(Chart 1): the first one includes linear and branched fatty acids
(group A), the second one contains residues characterized by
the presence of aromatic or heteroaromatic rings (group B)
and the third one is a miscellaneous group (group C) including
residues with heteroatoms or unsaturations within the chain.
A subset of 51 compounds, out of 174 glycose carbamides
represented in Chart 1, were synthesized and tested toward for
biological activity on the K562 cellular system, a subset, 20 of
them showed an appreciable erythroid differentiation inducing
activity on K562 cells. Synthesis, characterization and ability
to induce erythroid differentiation of these biologically active
compounds are herein presented.
Partially protected furanosic carbamides of type 2 and 4
(Scheme 2) were obtained from protected ones (1 and 3)
through selective hydrolytic removal of 5,6 acetonide group
with 80% aqueous AcOH. In the case of the D-allose series
(3) acid hydrolysis was conducted under milder temperature
conditions (45 ꢀC) with respect to that used for D-glucose an-
alogues (60 ꢀC), in order to prevent complete removal of the
two acetal groups. As pointed out by Collins [16], in fact,
the hydrolysis rate of the 1,2-O-isopropylidene functionality
in the D-allose series is higher with respect to that observed
for analogous acetonide functionality in the D-glucose series
1
[16]. All the compounds were fully characterized by H and
13C NMR and standard analytical parameters (see Section 4).
2. Chemistry
3. Biological activity
3-Amino-3-deoxy-1,2:5,6-di-O-isopropylidene-a-D-hexofuranose
scaffolds 7 [11] and 8 [12] (Scheme 1) were prepared according
to the literature procedures. The synthesis of methyl a- and b-
6-amino-6-deoxy-3,4-O-isopropylidene-D-galactopyranosides 15
The human leukemia K562 cell line [17] was kept in a hu-
midified atmosphere of 5% CO2/air in RPMI 1640 medium
(Sigma, St. Louis, MO, USA) supplemented with 10% fetal
bovine serum (FBS; Celbio, MI, Italy), 50 units/mL penicillin
and 50 mg/mL streptomycin [18]. In order to determine the
ability of the tested compounds to inhibit the cell growth
and induce the erythroid differentiation, K562 cells (30,000
cells/mL) were cultured both in the absence and presence of
the indicated concentrations of compounds and the cell num-
ber/mL was determined with an ZF Coulter Counter (Counter
Electronics, Hialeah, FL, USA) at different days from the
culture set-up. In order to verify possible effects on erythroid
differentiation, the proportion of benzidine-positive K562 cells
was determined and compared to the values obtained employ-
ing other known inducers of erythroid differentiation, includ-
ing cytosine arabinoside (ara-C) [19], mithramycin [3],
rapamycin [3] and butyric acid [3].
Among the tested carbamides (1Ca, 1Ac, 1Al, 2Aa, 2Ab,
2Ac, 2Ad, 2Ae, 2Af, 2Ag, 2Ah, 2Ai, 2Aj, 2Ak, 2Am, 2An,
2Ao, 2Be, 2Ca, 2Cb, 3Ab, 3Ad, 3Af, 3Ah, 3Al, 3An, 3Ba,
3Bb, 3Bc, 3Bd, 3Bf, 3Bg, 3Bi, 3Bj, 3Ca, 4Ac, 4Ac, 4Ai,
4Aj, 4Al, 4An, 4Bb, 4Bg, 4Bh, 4Bi, 4Bj, 4Cc, 4Cd, 5Ac,
6Al, 6Ac) 14 (1Ca, 2Ab, 2Ac, 2Ad, 2Ae, 2Af, 2Ah, 3Ab,
3An, 4Ac, 4Aj, 4Bi, 5Ac, 6Al) were found to exhibit apprecia-
ble (ꢁ15%) erythroid differentiation effect, while nine (2Ac,
2Aj, 3Bb, 3Bf, 3Bd, 4Ac, 4Ai, 4Bh, 4Aj), three of which
(2Ac, 4Ac and 4Aj) active on their own, were the compounds
O
HO
HO
O
O
Me2C
Me2C
O
O
O
O
O
O
NH
NH
O
O
O
R
R
CMe2
CMe2
CMe2
R
NH
O
3
O
O
1
2
HO
HO
O
O
O
NHCOR
O
O
NHCOR
O
Me2C
Me2C
OMe
O
O
O
OH
CMe2
OH
R
NH
O
OMe
4
5
6
O
GROUP A
a: R = CH3, b: R = CH2CH3, c: R = (CH2)2CH3, d: R = (CH2)3CH3,
e: R = (CH2)4CH3, f: R = CH2CH(CH3)2, g: R = CH2CH(CH3)CH2C(CH3)3,
h: R = CH2C(CH3)3, i: R = CH(CH3)2, j: R = C(CH3)3,
k: R = CH2CH2(CH2)5,l: R = (CH2)3, m: R = (CH2)4,
n: R = (CH2)5, o: R = (CH2)6
GROUP B
a: R = CH2-C6H5, b: R = CH2-(3-OMe)C6H4, c: R = CH2-(4-OMe)C6H4,
d: R = CH2-(2,5-di-OMe)C6H3, e: R = CH(C6H5)2, f: R = CH2CH2-C6H5,
g: R = C6H5, h: R = (4-C6H5)-C6H4, i: R = (4-OMe)C6H4, j: R = C6H4N
GROUP C
a: R = C CCH3, b: R = CH2OCH3, c: R = CH2OC6H5,
d: R = CH2OCH2(C6H5)
Chart 1.

M. Landi et al. / European Journal of Medicinal Chemistry 44 (2009) 745e754
747
O
O
O
O
Me2C
Me2C
O
O
NH2
O
O
CMe2
CMe2
NH2
O
O
7
8
OH
OTs
N3
NH2
O
O
O
O
i
ii
iii
O
O
O
O
Me2C
Me2C
Me2C
Me2C
R
R
R
R
O
O
O
O
HO
HO
HO
HO
R'
R'
R'
R'
9: R= H, R'=OMe
10: R=OMe, R'=H
11: R=H, R'=OMe
12: R=OMe, R'=H
13: R= H, R'=OMe
14: R=OMe, R'=H
15: R= H, R'=OMe
16: R=OMe, R'=H
Conditions: i) TsCl, Py. ii) NaN3, DMF. iii) LiAlH4, Et2O
Scheme 1. Conditions: (i) TsCl, Py; (ii) NaN3, DMF; (iii) LiAlH4, Et2O.
resulted active in synergism, potentiating erythroid induction
of K562 cells treated with sub-optimal concentrations of ara-
C. The data obtained on erythroid differentiation are shown
in Tables 1 and 2.
As far as structureeactivity relationship (SAR) analysis, we
like to underline that compounds of type 2 display the highest
probability (35% in our set) to induce differentiation (6/17,
compared to 2/15 and 3/13 of compounds of type 3 and 4, re-
spectively). Compounds carrying residues of group A display
the highest probability (36%) to induce erythroid differentia-
tion (11/31, compared to 1/14 and 1/6 of compounds carrying
residues of group B and C, respectively).
More in detail, all the compounds carrying the residue
Ab (R ¼ CH2CH3) and three out of five compounds carrying
residue Ac [R ¼ (CH2)2CH3] were found able to induce dif-
ferentiation (two of them, 2Ac and 4Ac, also in synergism
with ara-C). With respect to synergism with ara-C, only
the compounds carrying the residue R ¼ C(CH3)3 (2Aj and
4Aj) were found to be active in inducing high level of
differentiation.
When the analysis of the structure of carbamides active in
the stimulation of erythroid differentiation was performed, it
was found that seven of those exhibited linear saturated fatty
acid residues, three branched saturated fatty acid residues
and two alicyclic residues. However, we underline that many
amides of linear, branched and most of alicyclic fatty acid res-
idues are present in the list of inactive tested amide (i.e. 2Ak,
3Af, 5Ac, 2An). In addition, it should be noted that, despite
the well-known activity of phenylbutyrates and phenylacetates
[20], only one of the 14 aromatic rings containing the tested
amides exhibits activity. In contrast, among the nine carbam-
ides active in synergism with ara-C, four bear aromatic rings.
With respect to the role of sugar scaffold, we did not recog-
nise any clear structureeactivity relationship either related to
the protection/deprotection of glycide [compare couples 1Ac
(inactive)/2Ad (active) and 1Ca (active)/2Ca (inactive)] or
to the stereochemistry of carbon bearing the pharmacophore
[compare couples 2Ac (active)/4Ac (active), 2Ai (inactive)/
4Ai (active in synergism) and 2Aj (active in synergism)/4Aj
(active)].
In conclusion, the majority of active glycose carbamides act
with a mechanism resembling ara-C (no synergism observed
with ara-C); three carbamides (2Ac, 4Ac and 4Aj) display
a mechanism of action presumably different compared to that
exhibited by sub-optimal concentrations of ara-C; six (2Aj,
3Bf, 3Bb, 3Bd, 4Ai and 4Bh), which are not active by them-
selves, are, however, able to enhance ara-C mediated erythroid
induction. Whether they induce a part of the erythroid differen-
tiation program complementary to that stimulated by sub-
optimal concentrations of ara-C remains to be investigated.
Table 1
Effects of active carbamides on in vitro growth and erythroid differentiation of
human leukemic K562 cells
Compound
IC50 value
Erythroid inductiona
(% of benzidine-positive cells)
1Ca
2Ab
0.5 mM
30.0 mM
0.5 mM
5.0 mM
10.0 mM
5.0 mM
5.0 mM
7.5 mM
1.0 mM
20.0 mM
20.0 mM
2.5 mM
0.1 mM
0.25 mM
500 nM
100 nM
1.0 mM
2.0 mM
19 ꢂ 3.5
20 ꢂ 4.4
20 ꢂ 4.5
18 ꢂ 3.2
28 ꢂ 6.5
25 ꢂ 5.2
15 ꢂ 3.3
19 ꢂ 4.5
21 ꢂ 3.2
20 ꢂ 3.7
25 ꢂ 7.2
24 ꢂ 4.7
18 ꢂ 4.4
27 ꢂ 5.5
78 ꢂ 24.5
86 ꢂ 48.3
75.5 ꢂ 7.5
32.5 ꢂ 3.4
2Ac
2Ad
2Ae
2Af
2Ah
3Ab
3An
4Ac
4Aj
4Bi
5Ac
6Al
Ara-C
Mithramycin
Rapamycin
Butyric acid
a
O
O
CH2Cl2
80% aq. AcOH
2 and 4
Sugar
N
H
C R
Sugar-NH +
R
C Cl
2
1, 3,
Results are presented as average ꢂ SD (three independent experiments per-
formed) of % of benzidine-positive (hemoglobin-containing) cells after 6 days
induction period at the indicated concentrations of the tested compounds.
5 and 6
Scheme 2.

748
M. Landi et al. / European Journal of Medicinal Chemistry 44 (2009) 745e754
Table 2
Synergism between ara-C and carbamides on in vitro growth and erythroid dif-
ferentiation of human leukemic K562 cells
HPLCeMS analyses (Waters Acquity UPLC, with Waters
Acquity PAD detector and Micromass ZQ 2000 mass analyzer,
controlled by PC with MassLynx TM 4.1, column Waters
Acquity UPLC BEH C18 2.1 ꢃ 50 mm, 1.7 mm, eluent:
H2O/CH3CN/HCOOH 95/5/0,05 v/v/v). Solvents were dried
by distillation according to standard procedures [22], and
Compound
Concentration
Erythroid inductiona
(% of benzidine-positive cells)
2Ac
2Aj
3Bf
3Bb
3Bd
4Ac
4Ai
4Aj
4Bh
Ara-C
a
0.5 mM
5.0 mM
0.75 mM
1.0 mM
0.5 mM
20 mM
8.0 mM
20 mM
0.1 mM
500 nM
35 ꢂ 3.8
68 ꢂ 5.5
49 ꢂ 6.1
43 ꢂ 4.4
50 ꢂ 4.3
40 ꢂ 3.5
66 ꢂ 7.8
40 ꢂ 5.6
33 ꢂ 4.4
22 ꢂ 2.8
˚
storage over 4 A molecular sieves activated for at least 24 h
at 250 ꢀC. MgSO4 was used as the drying agent for solutions.
Acyl chlorides were purchased from Aldrich, with the
exception of cyclopropanecarbonyl chloride prepared from
corresponding acid according to the literature procedure
[23]. PS-piperidinomethyl resin and polyamine resin were
purchased from Novabiochem.
Results are presented as average ꢂ SD (three independent experiments per-
formed) of % of benzidine-positive (hemoglobin-containing) cells after 6 days
induction period at the indicated concentrations of the tested compounds. Sub-
optimal concentrations of ara-C (200 nm) were used in combination with the
tested compounds.
4.2. Amino scaffolds
3-Ammino-3-deoxy-1,2:5,6-di-O-isopropylidene-a-D-
glucofuranose [11] (7) and 3-ammino-3-deoxy-1,2:5,6-di-O-
isopropylidene-a-D-allofuranose [12] (8) were synthesized
as reported.
Despite being limited, this SAR analysis suggests that the
most promising molecules able to induce differentiation are
glycose carbamides of type 2 or those displaying residues
Ab (R ¼ CH2CH3) and Ac [R ¼ (CH2)2CH3]. Residue Aj
[R ¼ C(CH3)3] appears to be involved in the property to act
in synergism with ara-C. Obviously, this hypothesis is not con-
clusive, due to our choice of a diversity oriented selection of
compounds within the complete library of carbamides. This
choice has allowed to explore some representatives of each
class of acyl residue, but it has prevented the performance of
a systematic sight on the effect of glycide scaffold in biolog-
ical activity when a same acyl residue is considered. The
SAR analysis here discussed should be considered a starting
point for further synthetic activity toward the generation of
other oriented set of analogues, thus helping to verify this
hypothesis.
4.2.1. Methyl-3,4-O-isopropylidene-6-O-methylsulphonyl-a-
D-galactopyranoside (11)
A solution of 9 [13] (1.00 g, 4.27 mmol) in pyridine
(25 mL) was treated at 0 ꢀC under stirring with commercial
methylsulphoyl chloride (896 mg, 4.70 mmol). The solution
was allowed to warm to room temperature and stirred until
the TLC analysis (EtOAc) showed the complete disappearance
of the starting material (74 h) and the formation of one com-
ponent (Rf 0.51). The reaction mixture was repeatedly coeva-
porated with toluene (5 ꢃ 15 mL) under diminished pressure.
The crude residue was partitioned between CH2Cl2 (60 mL)
and H2O (30 mL), the aqueous phase was extracted with
CH2Cl2 (4 ꢃ 40 mL) and the organic ones were collected,
dried (MgSO4) and concentrated under diminished pressure.
Flash chromatography on silica gel (petroleum ethereEtOAc
2:1) of the crude solid led to pure 15 (1.07 g, 64%) as a white
solid. Rf 0.51 (EtOAc); mp (EtOAc) 127e129 ꢀC; lit. [24]
4. Experimental section
1
129 ꢀC. H NMR (CDCl3, 200 MHz): d 7.82 (AA0XX0, 2H,
4.1. General methods
Ar-H), 7.35 (AA0XX0, 2H, Ar-H), 4.70 (d, 1H, J ¼ 3.9 Hz,
H-1), 4.28e4.12 (m, 5H, H-6a, H-6b, H-2, H-3, H-4), 3.79
(m, 1H, H-5), 3.41 (s, 3H, OMe), 2.45 (s, 3H, MePh), 1.42,
1.29 (2s, each 3H, CMe2). 13C NMR (CDCl3, 50 MHz):
d 133.2, 144.8 (2 ꢃ Ar-C), 128.0, 129.8 (4 ꢃ Ar-CH), 110.0
(CMe2), 97.8 (C-1), 75.6 (C-3), 72.4 (C-4), 68.7, 69.2 (C-2,
C-5), 66.5 (C-6), 55.5 (OMe), 25.7, 27.4 (CMe2), 21.6
(MePh). Compound 11 was stored at 4 ꢀC or lower tempera-
ture to prevent degradation.
Melting points were determined with a Kofler hot-stage ap-
paratus and are uncorrected. Optical rotations were measured
on a PerkineElmer 241 polarimeter at 20 ꢂ 2 ꢀC. H NMR
1
spectra were recorded in appropriate solvents (internal stan-
dard Me4Si) with a Bruker AC 200 instrument at 200 MHz
and with a Bruker AvanceII operating at 250 MHz. 13C
NMR spectra were recorded with the spectrometers operating
at 50 and 62.9 MHz. Assignments were made with the aid of
DEPT, HETCOR and COSY experiments and by comparison
with values for known compounds and applying the known
additivity rules [21]. All reactions were followed by TLC on
Kieselgel 60 F254 (E. Merck) with detection by UV light
and/or with ethanolic 10% phosphomolybdic or sulphuric
acid and heating. Kieselgel 60 (E. Merck, 70e230 and
230e400 mesh, respectively) was used for column and flash
chromatography. Parallel reactions were followed by
4.2.2. Methyl-3,4-O-isopropylidene-6-O-methylsulphonyl-b-
D-galactopyranoside (12)
This compound was prepared starting from 10 [13] (3.10 g,
13.3 mmol) by a procedure analogous to that of 11. White solid,
3.61 g (70%). Rf 0.57 (EtOAc); mp (EtOAc) 157e159 ꢀC; lit.
[15] 154e155 ꢀC; optical rotation (c 1.0, CHCl3): [a]D þ1.0;
lit. [15] (c 2.3, CHCl3): [a]D 1.0. 1H NMR (CDCl3,
200 MHz): d 7.81 (AA0XX0, 2H, Ar-H), 7.35 (AA0XX0, 2H,

M. Landi et al. / European Journal of Medicinal Chemistry 44 (2009) 745e754
749
Ar-H), 4.28 (d, 1H, J1,2 ¼ 8.2 Hz, H-1), 4.23, 4.07 (2m, 5H,
4.2.5. Methyl-6-amino-6-deoxy-3,4-O-isopropylidene-a-D-
galactopyranoside (15)
H-3, H-4, H-5, H-6a, H-6b), 3.47 (dd, 1H, J2,3 ¼ 6.5 Hz, H-
2), 3.47 (s, 3H, OMe), 2.46 (s, 3H, MePh), 1.44, 1.29 (2s,
each 3H, CMe2). 13C NMR (CDCl3, 50 MHz): d 145.1, 132.1
(2 ꢃ Ar-C), 127.9, 128.9 (4 ꢃ Ar-CH), 110.5 (CMe2), 103.0
(C-1), 78.5 (C-3), 73.4, 72.9, 71.0 (C-2, C-4, C-5), 68.6 (C-
6), 57.0 (OMe), 27.9, 26.2 (Me2C), 21.6 (MePh). Compound
12 has to be stored at 4 ꢀC or lower temperature to prevent
degradation.
To a suspension of LiAlH4 (93 mg, 2.5 mmol) in dry Et2O
(10 mL) at 0 ꢀC a solution of 13 (260 mg, 1.0 mmol) in dry
Et2O (6 mL) was added drop wise and under stirring. The
suspension was warmed to reflux and stirred until the TLC
analysis (petroleum ethereEtOAc 1:1) showed the complete
disappearance of the starting material (30 min) and the forma-
tion of one component (Rf 0). The reaction mixture was cooled
to 0 ꢀC, diluted with Et2O and treated in sequence with 1 mL
of water, 2 mL of 10% aqueous NaOH and 3 mL of water, ob-
taining the formation of a white precipitate. The mixture was
filtered, the solid was washed with CH2Cl2 (4 ꢃ 5 mL) and the
organic ones were collected, dried (MgSO4) and concentrated
under diminished pressure to provide crude 15 (184 mg, 79%).
Crystallization (EtOAc) of the crude solid furnished 15 as
a white crystalline solid. Mp 152e157 ꢀC; optical rotation (c
1.0, CHCl3): [a]D þ145.7. 1H NMR (CDCl3, 200 MHz):
d 4.76 (d, 1H, J1,2 ¼ 3.8 Hz, H-1), 4.21 (m, 2H, H-4, H-5),
3.91 (m, 1H, H-2), 3.79 (t, 1H, J2,3 ¼ J3,4 ¼ 5.7 Hz, H-3),
3.45 (s, 3H, OMe), 3.05 (dd, 1H, J5,6b ¼ 7.0 Hz, H-6b), 2.91
(dd, 1H, J6a,6b ¼ 13.3 Hz, J5,6a ¼ 4.9 Hz, H-6a), 1.35, 1.50
(2s, each 3H, CMe2). 13C NMR (CDCl3, 50 MHz): d 109.5
(CMe2), 98.7 (C-1), 76.4 (C-3), 74.0 (C-4), 69.3, 69.7 (C-5,
C-2), 55.4 (OMe), 42.8 (C-6), 25.9, 27.8 (CMe2). Anal. for
C10H19NO5. Calcd (%): C, 51.49; H, 7.95; N, 6.00. Found
(%): C, 50.97; H, 8.25; N, 5.77.
4.2.3. Methyl-6-azido-6-deoxy-3,4-O-isopropylidene-a-D-
galactopyranoside (13)
A solution of 11 (1.00 g, 2.58 mmol) in DMF (35 mL)
was treated with commercial sodium azide (355 mg,
5.15 mmol). The suspension was warmed to 120 ꢀC and stirred
until the TLC analysis (petroleum ethereEtOAc 1:1) showed
the complete disappearance of the starting material (28 h)
and the formation of one component (Rf 0.35). The reaction
mixture was allowed to cool to room temperature and concen-
trated under diminished pressure. The crude residue was par-
titioned between CH2Cl2 (35 mL) and H2O (30 mL), the
aqueous phase was extracted with CH2Cl2 (5 ꢃ 25 mL) and
the organic ones were collected, dried (MgSO4) and concen-
trated under diminished pressure. Flash chromatography on
silica gel (petroleum ethereEtOAc 1:1) of the crude syrup
led to pure 13 (677 mg, 77%) as a syrup. Rf 0.35 (petroleum
ethereEtOAc 1:1); optical rotation (c 1.2, CHCl3) [a]D
þ89.4. 1H NMR (CDCl3, 200 MHz): d 4.78 (d, 1H,
J1,2 ¼ 3.9 Hz, H-1), 4.29 (t, 1H, J3,4 ¼ 6.1 Hz, H-3), 4.14 (m,
2H, H-5, H-4), 3.85 (m, 1H, H-2), 3.59 (dd, 1H,
J5,6b ¼ 8.3 Hz, H-6b), 3.50 (s, 3H, OMe), 3.33 (dd, 1H,
J6a,6b ¼ 12.9 Hz, J5,6a ¼ 4.3 Hz, H-6a), 2.53 (d, 1H,
J2,OH ¼ 5.7 Hz, OH), 1.51, 1.35 (2s, each 3H, CMe2). 13C
NMR (CDCl3, 50 MHz): d 109.9 (CMe2), 98.1 (C-1), 75.6
(C-3), 73.1 (C-4), 68.8, 67.9 (C-5, C-2), 55.6 (OMe), 51.2
(C-6), 27.4, 25.7 (CMe2). Anal. for C10H17N3O5. Calcd (%):
C, 46.33; H, 6.61; N, 16.21. Found (%): C, 44.66; H, 6.47;
N, 15.66.
4.2.6. Methyl-6-amino-6-deoxy-3,4-O-isopropylidene-b-D-
galactopyranoside (16)
This compound was prepared starting from 14 (630 mg,
2.44 mmol) by a procedure analogous to that of 15. White solid
463 mg (81%); mp (hexane) 146e153 ꢀC; optical rotation (c
1.0, CHCl3): [a]D ꢄ18.9. 1H NMR (CDCl3, 200 MHz):
d 4.11 (d, 1H, J ¼ 8.3 Hz, H-1), 4.08 (m, 2H, H-3, H-4), 3.73
(ddd, 1H, J4,5 ¼ 1.9 Hz, H-5), 3.55 (s, 3H, OMe), 3.49 (t, 1H,
0
J ¼ 7.8 Hz, H-2), 3.14 (dd, 1H, J5.6 ¼ 7.6 Hz, H-6b), 2.96
0
(dd, 1H, J6,6 ¼ 13.3 Hz, J5,6 ¼ 4.7 Hz, H-6a), 2.48 (br s, 1H,
OH), 1.50, 1.34 (2s, each 3H, CMe2). 13C NMR (CDCl3,
50 MHz): d 110.0 (CMe2), 103.4 (C-1), 79.3 (C-3), 74.4,
74.1, 73.3 (C-2, C-5, C-4), 56.8 (OMe), 42.6 (C-6), 28.1,
26.2 (CMe2). Anal. for C10H19NO5. Calcd (%): C, 51.49; H,
7.95; N, 6.00. Found (%): C, 50.93; H, 7.83; N, 5.85.
4.2.4. Methyl-6-azido-6-deoxy-3,4-O-isopropylidene-b-D-
galactopyranoside (14)
This compound was prepared starting from 12 (1.50 g,
3.87 mmol) by a procedure analogous to that of 13. White
solid, 792 mg (79%). Rf 0.22 (petroleum ethereEtOAc
1:1); mp (hexane) 80e82 ꢀC; optical rotation (c 1.0,
4.3. General method for the parallel synthesis of amides
of type 1, 3, 5 and 6
1
CHCl3): [a]D ꢄ16.6. H NMR (CDCl3, 200 MHz): d 4.14
(d, 1H, J1,2 ¼ 8.0 Hz, H-1), 4.10 (m, 2H, H-3, H-4), 3.94
A 0.2 M solution of amine scaffold in CH2Cl2 and a 0.4 M
solution of the selected acyl chloride in CH2Cl2 were prepared
under anhydrous conditions.
In each reactor 1.00 g of resin PS-piperidinomethyl was put
and in sequence 5 mL of the solution containing the amine and
5 mL of the solution containing the acyl chloride were added.
The reaction mixture was stirred at room temperature for 12 h.
The reaction proceeding was checked by HPLCeMS. A
1.00 g of resin polyamine (loading ¼ 3.2 mmol/g) to extract
the excess acyl chloride was added after dilution with
(ddd, 1H, J4.5 ¼ 1.7 Hz, H-5), 3.73 (dd, 1H, J5,6b
¼
8.3 Hz, H-6b), 3.57 (s, 3H, OMe), 3.54 (m, 1H, H-2),
3.33 (dd, 1H, J6a,6b ¼ 12.9 Hz, J5,6a ¼ 4.2 Hz, H-6a), 2.85
(br s, 1H, OH), 1.35, 1.53 (2s, each 3H, CMe2). 13C
NMR (CDCl3, 50 MHz): d 110.4 (CMe2), 103.1 (C-1),
78.6 (C-3), 73.5, 73.7, 72.9 (C-2, C-5, C-4), 57.0 (OMe),
51.0 (C-6), 26.2, 27.9 (CMe2). Anal. for C10H17N3O5.
Calcd (%): C, 46.33; H, 6.61; N, 16.21. Found (%): C,
44.28; H, 6.32; N, 16.32.

750
M. Landi et al. / European Journal of Medicinal Chemistry 44 (2009) 745e754
CH2Cl2 (5 mL) and vortically stirred for 2 h. The suspension
was filtered on paper and the clear solutions were evaporated
under diminished pressure to give oils that were purified by
a Combi Flash chromatographic system (ISCO, using columns
Redisep of 10 g eluting with a CH2Cl2eMeOH gradient from
98:2 to 95:5). After evaporation the solvent pure products were
obtained.
4.5.2. 3-N-Propanoyl-3-deoxy-1,2-O-isopropylidene-a-D-
glucofuranose (2Ab)
From 7 (778 mg, 3.00 mmol) by acylation with propanoyl
chloride and subsequent hydrolysis; colourless syrup 504 mg
(61%); Rf 0.27 (EtOAceMeOH 9:1); optical rotation (c 1.0,
MeOH): [a]D þ15.4; 1H NMR (250 MHz, CD3CN): see
Table 3 and d 2.30 (br s, 2H, OH-5, OH-6), 2.25 (t, 2H,
J ¼ 7.5 Hz, CH2CO), 1.45, 1.27 (2s, each 3H, CMe2), 1.08
(t, 3H, J ¼ 7.5 Hz, Me); 13C NMR (62.9 MHz, CD3CN): see
Table 4 and d 176.6 (CO), 112.4 (CMe2), 29.7 (CH2CO),
26.7, 26.3 (CMe2), 10.2 (Me). Anal. for C12H21NO6. Calcd
(%): C, 52.35; H, 7.69; N, 5.09. Found (%): C, 52.16; H,
7.72; N, 5.07.
4.4. General method of the synthesis of selectively
deprotected furanosic amides of type 1 and 3
The fully protected amides of type 1 and 3 (1.5e3.0 mmol),
prepared following the general procedure reported above, were
dissolved in 80% aqueous AcOH (20 mL) and the resulting
solutions were warmed to 60 ꢀC in the case of hydrolysis of
amides of type 1 and to 45 ꢀC in the case of amides of type
3. Reaction mixtures were maintained under stirring until
the TLC analysis (HexaneeEtOAc) revealed the disappear-
ance of starting material (1e4 h). The solution was repeatedly
coevaporated with toluene (5 ꢃ 10 mL) under diminished
pressure. The crude residues were purified by a Combi Flash
chromatographic system (ISCO, using columns Redisep of
10 g and as eluted a CH2Cl2eMeOH gradient from 98:5 to
90:10). After evaporation the solvent pure products were
obtained.
4.5.3. 3-N-Butanoyl-3-deoxy-1,2-O-isopropylidene-a-D-
glucofuranose (2Ac)
From 7 (778 mg, 3.00 mmol) by acylation with butanoyl
chloride and subsequent hydrolysis; colourless syrup 564 mg
(65%); Rf 0.43 (EtOAceMeOH 9:1); optical rotation (c 1.0,
1
MeOH): [a]D þ19.8; H NMR (200 MHz, CDCl3): see Table
3 and d 3.60 (br s, 2H, OH-5, OH-6), 2.23 (t, 2H, J ¼ 7.0 Hz,
CH2CO), 1.58 (m, 2H, CH2Me), 1.50, 1.30 (2s, each 3H,
CMe2), 0.91 (t, 3H, J ¼ 7.2 Hz, Me); 13C NMR (50 MHz,
CDCl3): see Table 4 and d 175.2 (CO), 111.7 (CMe2), 37.8
(CH2CO), 26.2, 25.8 (CMe2), 18.9 (CH2Me), 13.5 (Me).
Anal. for C13H23NO6. Calcd (%): C, 53.97; H, 8.01; N,
4.84. Found (%): C, 54.18; H, 8.04; N, 4.86.
4.5. Active carbamides
4.5.1. 3-N-(2-Butynoyl)-3-deoxy-1,2:5,6-di-O-
isopropylidene-a-D-glucofuranose (1Ca)
4.5.4. 3-N-Pentanoyl-3-deoxy-1,2-O-isopropylidene-a-D-
glucofuranose (2Ad)
From 7 (778 mg, 3.00 mmol) by acylation with 2-butynoyl
chloride, white solid 705 mg (72%); Rf 0.32 (hexaneeEtOAc
3:7); mp (chrom) 160e162 ꢀC; optical rotation (c 1.0,
CHCl3): [a]D ꢄ47.3; 1H NMR (250 MHz, CD3CN): see
Table 5 and d 1.93 (s, 3H, MeC^), 1.44, 1.36, 1.28, 1.26 (4s,
each 3H, 2 ꢃ CMe2); 13C NMR (62.9 MHz, CD3CN): see
Table 6 and d 153.7 (CO), 112.6, 110.0 (2 ꢃ CMe2), 84.9
(^CCO); 75.2 (MeC^), 27.9, 27.8, 26.3, 25.4 (2 ꢃ CMe2),
3.5 (MeC^); Anal. for C16H23NO6. Calcd (%): C, 59.07; H,
7.13; N, 4.30. Found (%): C, 58.87; H, 7.16; N, 4.28.
From 7 (778 mg, 3.00 mmol) by acylation with pentanoyl
chloride and subsequent hydrolysis; colourless syrup 574 mg
(52%); Rf 0.25 (EtOAc); optical rotation (c 1.4, MeOH):
1
[a]D þ25.7; H NMR (250 MHz, CD3CN): see Table 3 and
d 4.30, 2.38 (2br s, each 1H, OH-5, OH-6), 2.20 (t, 2H,
J ¼ 7.4 Hz, CH2CO), 1.55 (m, 2H, CH2CH2CO), 1.34 (m,
2H, CH2Me), 1.45, 1.27 (2s, each 3H, CMe2), 0.90 (t, 3H,
J ¼ 7.3 Hz, Me); 13C NMR (62.9 MHz, CD3CN): see Table 4
and d 176.0 (CO), 112.5 (CMe2), 36.3 (CH2CO), 28.6
(CH2CH2CO), 26.7, 26.4 (CMe2), 23.0 (CH2Me), 14.1 (Me).
Table 3
1H NMR data (d, ppm; J, Hz) for compounds of type 2 and 4
Compound
Solvent
H-1
H-2
H-3
H-4
H-5
H-6a
H-6b
NH
J1,2
J2,3
J3,4
J3,NH
J4,5
J5,6a
J5,6b
J6a,6b
2Ab
2Ac
2Ad
2Ae
2Af
2Ah
2Aj
4Ac
4Ai
4Aj
4Bh
4Bi
CD3CN
CDCl3
CD3CN
5.84
5.89
5.83
5.83
5.83
5.84
5.89
5.76
5.76
5.79
5.84
5.81
4.59
4.58
4.50
4.49
4.50
4.50
4.56
4.59
4.59
4.62
4.76
4.72
4.20
4.36
4.21
4.21
4.20
4.21
4.31
4.23
4.23
4.17
4.47
4.45
4.01
4.10
4.01
4.02
4.00
4.02
4.15
3.77
3.80
3.82
4.04
4.08
3.50
3.70
3.49
3.55
3.53
3.48
3.90
3.65
3.66
3.66
3.79
3.80
3.50
3.70
3.49
3.55
3.53
3.48
3.70
3.42
3.40
3.40
3.48
3.44
3.64
3.70
3.66
3.62
3.53
3.64
3.70
3.51
3.51
3.52
3.58
3.55
6.87
7.49
6.96
e
3.7
3.7
3.7
3.6
3.6
3.7
3.7
3.8
3.8
3.8
3.8
3.8
0
3.2
3.2
3.2
3.2
3.1
3.2
3.4
9.2
9.2
9.2
9.4
9.7
7.6
7.1
7.7
e
8.4
7.3
8.3
7.9
8.2
8.3
6.4
4.8
4.6
4.9
4.7
4.1
n.d.
n.d.
n.d.
n.d.
n.d.
n.d.
n.d.
6.6
2.2
n.d.
n.d.
2.7
n.d.
2.6
n.d.
4.0
3.8
4.0
4.2
4.2
10.4
n.d.
0
0
n.d.
CD3CNeD2O
CD3CN
CD3CN
0
10.7
n.d.
6.84
6.90
6.84
6.62
6.56
6.46
7.15
e
0
7.8
7.4
6.4
8.9
9.0
8.2
8.1
e
0
10.3
n.d.
CDCl3
CD3CN
0
5.1
5.1
5.3
5.0
4.9
11.3
11.5
11.2
11.4
11.5
CD3CN
CD3CN
6.7
6.7
CD3CN
CD3CNeD2O
6.6
7.1

M. Landi et al. / European Journal of Medicinal Chemistry 44 (2009) 745e754
751
Table 4
13C NMR data (d, ppm) for compounds of type 2 and 4
C14H25NO6. Calcd (%): C, 55.43; H, 8.31; N, 4.62. Found
(%): C, 55.30; H, 8.34; N, 4.60.
Compound Solvent
C-1
C-2
C-3
C-4
C-5
C-6
2Ab
2Ac
2Ad
2Ae
2Af
2Ah
2Aj
4Ac
4Ai
4Aj
4Bh
4Bi
CD3CN
CDCl3
CD3CN
105.6 84.8 57.0 79.8 70.0 64.4
104.2 83.7 56.3 78.5 69.4 63.7
105.5 84.8 57.1 79.8 70.1 64.4
4.5.7. 3-N-(3,3-Dimethylbutanoyl)-3-deoxy-1,2-O-
isopropylidene-a-D-glucofuranose (2Ah)
CD3CNeD2O 105.5 84.8 56.7 79.4 70.0 64.1
From 7 (778 mg, 3.00 mmol) by acylation with 3,3-dime-
thylbutanoyl chloride and subsequent hydrolysis; colourless
syrup, 505 mg (53%); Rf 0.47 (EtOAceMeOH 95:5); optical
rotation (c 1.0, CHCl3): [a]D þ35.4; 1H NMR (250 MHz,
CD3CN): see Table 3 and d 4.31, 2.85 (2br s, each 1H, OH-
5, OH-6), 2.08 (m, 2H, CH2CO), 1.45, 1.27 (2s, each 3H,
CMe2), 1.01 (s, 9H, CMe3); 13C NMR (62.9 MHz, CD3CN):
see Table 4 and d 174.5 (CO), 112.5 (CMe2), 50.0 (CH2CO),
31.4 (CMe3), 30.1 (CMe3), 26.8, 26.4 (CMe2). Anal. for
C15H27NO6. Calcd (%): C, 56.77; H, 8.57; N, 4.41. Found
(%): C, 56.59; H, 8.55; N, 4.43.
CD3CN
CD3CN
CDCl3
105.6 84.8 57.1 79.9 70.2 64.5
105.6 84.8 57.1 79.7 70.2 64.4
104.0 84.1 56.7 78.1 69.9 63.5
105.1 80.6 53.3 80.5 73.2 63.8
105.2 80.7 53.3 80.5 73.1 63.8
105.2 80.9 53.6 80.3 73.1 63.8
105.3 80.4 54.1 80.2 73.1 63.8
CD3CN
CD3CN
CD3CN
CD3CN
CD3CNeD2O 105.2 80.3 53.5 79.6 72.7 63.4
Anal. for C14H25NO6. Calcd (%): C, 55.43; H, 8.31; N, 4.62.
Found (%): C, 55.21; H, 8.34; N, 4.58.
4.5.5. 3-N-Hexanoyl-3-deoxy-1,2-O-isopropylidene-a-D-
glucofuranose (2Ae)
4.5.8. 3-N-Pivaloyl-3-deoxy-1,2-O-isopropylidene-a-D-
glucofuranose (2Aj)
From 7 (778 mg, 3.00 mmol) by acylation with hexanoyl
chloride and subsequent hydrolysis; colourless syrup 628 mg
(66%); Rf 0.42 (EtOAceMeOH 95:5); optical rotation (c
1.0, MeOH): [a]D þ23.2; 1H NMR (250 MHz, CD3CNe
D2O): see Table 3 and d 2.18 (t, 2H, J ¼ 6.4 Hz, CH2CO),
1.54 (m, 2H, CH2CH2CO), 1.27 [m, 4H, (CH2)2Me], 1.44,
1.26 (2s, each 3H, CMe2), 0.89 (t, 3H, J ¼ 6.7 Hz, Me); 13C
NMR (62.9 MHz, CD3CNeD2O): see Table 6 and d 176.5
(CO), 112.7 (CMe2), 36.5 (CH2CO), 32.0 (CH2CH2CO),
26.6, 26.2 (CMe2), 26.1, 23.0 [(CH2)2Me], 14.2 (Me). Anal.
for C15H27NO6. Calcd (%): C, 56.77; H, 8.57; N, 4.41. Found
(%): C, 57.02; H, 8.61; N, 4.39.
From 7 (778 mg, 3.00 mmol) by acylation with 2,2-dime-
thylbutanoyl chloride and subsequent hydrolysis; white solid,
821 mg (90%); Rf 0.44 (EtOAceMeOH 95:5); mp (chrom)
161e163 ꢀC; optical rotation (c 1.0, CHCl3): [a]D þ22.3;
compound 2Aj 1H NMR (200 MHz, CDCl3): see Table 3
and d 4.35, 3.02 (2br s, each 1H, OH-5, OH-6), 1.51, 1.31
(2s, each 3H, CMe2), 1.21 (s, 9H, CMe3); 13C NMR
(50 MHz, CDCl3): see Table 4 and d 180.3 (CO), 111.9
(CMe2), 38.8 (CMe3), 27.4 (CMe3), 26.4, 26.2 (CMe2). Anal.
for C14H25NO6. Calcd (%): C, 55.43; H, 8.31; N, 4.62. Found
(%): C, 55.62; H, 8.35; N, 4.60.
4.5.6. 3-N-(3-Methylbutanoyl)-3-deoxy-1,2-O-
isopropylidene-a-D-glucofuranose (2Af)
4.5.9. 3-N-Propanoyl-3-deoxy-1,2:5,6-di-O-isopropylidene-
a-D-allofuranose (3Ab)
From 7 (778 mg, 3.00 mmol) by acylation with 3-methyl-
butanoyl chloride and subsequent hydrolysis; white solid
501 mg (55%); Rf 0.28 (EtOAc); mp (hexane) 115e116 ꢀC;
optical rotation (c 0.4, CHCl3): [a]D þ46.3; 1H NMR
(200 MHz, CD3CN): see Table 3 and d 4.23 (br s, 1H, OH-
5), 2.77 (br t, 1H, OH-6), 2.06 (m, 2H, CH2CO), 2.02 (m,
1H, CHMe2), 1.45, 1.27 (2s, each 3H, CMe2), 0.92 (d, 6H,
J ¼ 6.2 Hz, 2 ꢃ Me); 13C NMR (50 MHz, CD3CN): see Table
4 and d 175.2 (CO), 112.5, (CMe2), 45.7 (CH2CO), 26.9
(CHMe2), 26.8, 26.3 (CMe2), 22.6 (2 ꢃ Me). Anal. for
From 8 (778 mg, 3.00 mmol) by acylation with propanoyl
chloride; white solid, 833 mg (88%); Rf 0.60 (EtOAce
MeOH 9:1); mp (hexane) 93e95 ꢀC; optical rotation (c 1.0,
CHCl3): [a]D þ71.1; 1H NMR (200 MHz, CD3CN): see Table
5 and d 2.15 (q, 2H, J ¼ 7.5 Hz, CH2CO), 1.50, 1.34, 1.29,
1.27 (4s, each 3H, 2 ꢃ CMe2), 1.05 (t, 3H, Me); 13C NMR
(50 MHz, CD3CN): see Table 6 and d 174.3 (CO), 112.9,
109.9 (2 ꢃ CMe2), 29.6 (CH2CO), 26.9, 26.6, 26.5, 25.4
(2 ꢃ CMe2), 10.1 (Me). Anal. for C15H25NO6. Calcd (%): C,
57.13; H, 7.99; N, 4.44. Found (%): C, 57.39; H, 8.02; N, 4.42.
Table 5
1H NMR data (d, ppm; J, Hz) for compounds of type 1, 3, 5 and 6
Compound
Solvent
H-1
H-2
H-3
H-4
H-5
H-6a
H-6b
NH
J1,2
J2,3
J3,4
J3,NH
J4,5
J5,6a
J5,6b
J6a,6b
1Ca
3Ab
3An
3Bd
3Bf
CD3CN
CD3CN
CD3CN
CD3CN
CD3CN
CD3CN
CD3CN
CD3CN
5.81
5.75
5.75
5.75
5.75
5.75
4.59
4.02
4.46
4.55
4.54
4.53
4.52
4.56
3.57
3.28
4.33
4.13
4.19
4.09
4.10
4.13
4.02
3.94
4.10
3.90
3.91
3.84
3.86
3.87
4.15
4.10
4.20
4.13
4.11
4.12
4.07
4.11
3.96
3.80
3.83
3.90
3.91
3.79
3.82
3.81
3.23
3.28
4.05
3.90
3.91
3.94
3.95
3.96
3.48
3.53
7.05
6.43
6.43
6.60
6.48
6.59
6.62
6.90
3.7
3.7
3.6
3.8
3.8
3.8
3.6
8.2
0
3.7
n.d.
9.5
9.3
9.9
9.8
5.5
5.5
8.8
8.1
8.8
8.4
8.8
8.4
e
7.2
n.d.
n.d.
3.5
3.4
4.0
2.4
2.1
5.5
n.d.
n.d.
6.8
6.3
6.6
7.8
8.4
6.1
n.d.
n.d.
6.6
6.6
6.6
4.4
4.2
8.4
n.d.
n.d.
8.0
4.3
4.3
4.6
4.7
4.9
7.5
7.0
7.9
8.2
3Bb
5Ac
6Al
13.7
13.8
e

752
M. Landi et al. / European Journal of Medicinal Chemistry 44 (2009) 745e754
Table 6
13C NMR data (d, ppm) for compounds of type 1, 3, 5 and 6
4.5.13. 3-N-Hydrocinnammoyl-3-deoxy-1,2:3,4-di-O-
isopropylidene-a-D-allofuranose (3Bf)
Compound
Solvent
C-1
C-2
C-3
C-4
C-5
C-6
From 8 (778 mg, 3.00 mmol) by acylation with hydrocin-
nammoyl chloride; colourless syrup, 846 mg (72%); Rf 0.72
(EtOAceMeOH 9:1); optical rotation (c þ1.0, CHCl3): [a]D
þ66.6; 1H NMR (200 MHz, CD3CN): see Table 5 and
d 7.25 (m, 5H, Ar-H), 2.89, 2.47 (2t, each 2H, J ¼ 8.1 Hz,
2 ꢃ CH2), 1.49, 1.34, 1.28, 1.28 (4s, each 3H, 2 ꢃ CMe2);
13C NMR (50 MHz, CD3CN): see Table 6 and d 172.9 (CO),
142.3 (Ar-C), 129.3e127.0 (Ar-CH), 113.0, 110.0
(2 ꢃ CMe2), 38.2, 32.1 (2 ꢃ CH2), 26.9, 26.6, 26.5, 25.5
(2 ꢃ CMe2); Anal. for C21H29NO6. Calcd (%): C, 64.43; H,
7.47; N, 3.58. Found (%): C, 64.38; H, 7.50; N, 3.57.
1Ca
3Ab
3An
3Bd
3Bf
CD3CN
CD3CN
CD3CN
CD3CN
CD3CN
CD3CN
CD3CN
CD3CN
105.6
105.3
105.3
105.3
105.4
105.4
100.4
104.4
85.2
80.1
80.1
80.0
80.0
80.0
70.8
74.2
56.4
53.6
53.7
53.5
53.7
54.0
77.4
80.3
79.5
78.9
79.0
79.2
78.8
79.1
74.7
75.1
73.7
76.4
76.5
76.3
76.3
76.5
66.8
72.1
67.5
65.4
65.6
65.2
65.3
65.6
40.7
41.1
3Bb
5Ac
6Al
4.5.10. 3-N-Cyclopentanecarbonyl-3-deoxy-1,2:5,6-di-O-
isopropylidene-a-D-allofuranose (3An)
From 8 (778 mg, 3.00 mmol) by acylation with cyclopenta-
necarbonyl chloride; white solid, 960 mg (90%); Rf 0.33 (hex-
aneeEtOAc 1:1); mp (chrom) 102e104 ꢀC; optical rotation (c
4.5.14. 3-N-Butanoyl-3-deoxy-1,2-O-isopropylidene-a-D-
allofuranose (4Ac)
From 8 (778 mg, 3.00 mmol) by acylation with butanoyl
chloride and subsequent hydrolysis; white solid, 556 mg
(64%); Rf 0.32 (EtOAc); mp (chrom) 123e124 ꢀC; optical ro-
tation (c 1.0, CHCl3): [a]D þ4.4; 1H NMR (200 MHz,
CD3CN): see Table 3 and d 3.76 (d, 1H, J5,OH ¼ 4.6 Hz,
OH-5), 2.80 (dd, 1H, J6a,OH ¼ 5.4 Hz, J6b,OH ¼ 6.5 Hz, OH-
6), 2.17 (t, 2H, J ¼ 7.2 Hz, CH2CO), 1.53 (sestetto, 2H,
CH2Me); 1.50, 1.30 (2s, each 3H, CMe2), 0.90 (t, 3H,
J ¼ 7.4 Hz, Me); 13C NMR (50 MHz, CD3CN): see Table 4
and d 175.0 (CO), 113.0 (CMe2), 38.5 (CH2CO), 26.9, 26.7
(CMe2), 19.7 (CH2Me), 13.9 (Me); Anal. for C13H23NO6.
Calcd (%): C, 55.97; H, 8.01; N, 4.84. Found (%): C, 55.88;
H, 8.04; N, 4.86.
1
1.0, CHCl3): [a]D þ65.5; H NMR (200 MHz, CD3CN): see
Table 5 and d 2.61 (m, 1H, CH ), 1.63 (m, 8H, cyclopentyl
H), 1.50, 1.34, 1.29, 1.26 (4s, each 3H, 2 ꢃ CMe2); 13C
NMR (50 MHz, CD3CN): see Table 6 and d 176.8 (CO),
113.0, 110.0 (2 ꢃ CMe2), 45.8 (CH); 31.3, 30.7 (2 ꢃ CH2),
26.9, 26.7, 26.6, 25.6 (2 ꢃ CMe2), 26.7 (2 ꢃ CH2). Anal. for
C18H29NO6. Calcd (%): C, 60.83; H, 8.22; N, 3.94. Found
(%): C, 60.58; H, 8.25; N, 3.94.
4.5.11. 3-N-(3-Methoxyphenylacetyl)-3-deoxy-1,2:3,4-di-O-
isopropylidene-a-D-allofuranose (3Bb)
From 8 (778 mg, 3.00 mmol) by acylation with 3-methox-
yphenylacetyl chloride; colourless syrup, 1004 mg (82%); Rf
0.50 (EtOAc); optical rotation (c 1.0, CHCl3): [a]D þ70.1;
1H NMR (250 MHz, CD3CN): see Table 5 and d 7.23 (dd,
4.5.15. 3-N-(2-Methylpropanoyl)-3-deoxy-1,2-O-
isopropylidene-a-D-allofuranose (4Ai)
From 8 (778 mg, 3.00 mmol) by acylation with 2-methyl-
propanoyl chloride and subsequent hydrolysis; white solid,
460 mg (53%); Rf 0.26 (EtOAceMeOH 95:5); mp (chrom)
105e107 ꢀC; optical rotation (c 1.0, CHCl3): [a]D þ4.6; H
0
0
0
0
0
0
1H, J6 ,5 ¼ 8.0 Hz, J4 ,5 ¼ 8.2 Hz, H-5 ), 6.86 (m, 2H, H-2 ,
H-60), 6.82 (ddd, 1H, J ¼ 2.5 Hz, J ¼ 1.0 Hz, H-40), 3.77 (s,
3H, OMe), 3.47 (s, 2H, PhCH2), 1.48, 1.31, 1.28, 1.25 (4s,
each 3H, 2 ꢃ CMe2); 13C NMR (62.9 MHz, CD3CN): see
Table 6 and d 171.3 (CO), 160.8 (C-30), 138.3 (C-10), 130.5
(C-50), 122.4, 115.8, 113.1 (C-20, C-40, C-60), 113.0, 110.0
(2 ꢃ CMe2), 55.8 (OMe), 43.4 (PhCH2), 26.7, 26.6, 26.5,
25.5 (2 ꢃ CMe2); Anal. for C21H29NO7. Calcd (%): C,
61.90; H, 8.31; N, 3.44. Found (%): C, 61.87; H, 8.35; N, 3.43.
1
NMR (250 MHz, CD3CN): see Table 3 and d 3.73 (d, 1H,
J5,OH ¼ 4.8 Hz, OH-5), 2.84 (dd, 1H, J6a,OH ¼ 5.3 Hz,
J6b,OH ¼ 6.6 Hz, OH-6), 2.45 (ept, 1H, J ¼ 6.8 Hz, CHMe2),
1.51, 1.30 (2s, each 3H, CMe2), 1.06, 1.07 (2d, each 3H,
J ¼ 6.8 Hz, CHMe2); 13C NMR (62.9 MHz, CD3CN): see
Table 4 and d 178.8 (CO), 113.0 (CMe2), 35.6 (CHMe2),
26.9, 26.7 (CMe2), 19.7 (CHMe2). Anal. for C13H23NO6.
Calcd (%): C, 53.97; H, 8.01; N, 4.84. Found (%): C, 53.99;
H, 7.99; N, 4.82.
4.5.12. 3-N-(2,5-Dimethoxyphenylacetyl)-3-deoxy-1,2:3,4-
di-O-isopropylidene-a-D-allofuranose (3Bd)
From 8 (778 mg, 3.00 mmol) by acylation with 2,5-dime-
thoxyphenylacetyl chloride; colourless syrup, 998 mg (76%);
Rf 0.39 (hexaneeEtOAc 1:9); optical rotation (c 1.2,
CHCl3): [a]D þ57.1; 1H NMR (200 MHz, CD3CN): see Table
5 and d 6.91e6.78 (m, 3H, H-30, H-40, H-60), 3.78, 3.72 (2s,
each 3H, 2 ꢃ OMe), 3.44 (s, 2H, CH2CO), 1.47, 1.29, 1.28,
1.25 (4s, each 3H, 2 ꢃ CMe2); 13C NMR (50 MHz, CD3CN):
see Table 6 and d 171.4 (CO), 154.5, 152.4 (C-30, C-50),
126.0 (C-10), 118.1, 113.4, 112.6 (C-30, C-40, C-60), 113.0,
110.0 (2 ꢃ CMe2), 56.6, 56.2 (2 ꢃ OMe), 38.9 (CH2CO),
26.8, 26.6, 26.5, 25.5 (2 ꢃ CMe2); Anal. for C22H31NO8.
Calcd (%): C, 60.40; H, 7.14; N, 3.20. Found (%): C, 60.37;
H, 7.14; N, 3.19.
4.5.16. 3-N-Pivaloyl-3-deoxy-1,2-O-isopropylidene-a-D-
allofuranose (4Aj)
From 8 (778 mg, 3.00 mmol) by acylation with pivaloyl
chloride and subsequent hydrolysis; white solid, 510 mg
(56%); Rf 0.36 (EtOAceMeOH 95:5); mp (chrom) 107e
1
108 ꢀC; optical rotation (c 1.0, CHCl3): [a]D þ8.5; H NMR
(200 MHz, CD3CN): see Table 3 and d 3.58 (br s, 1H, OH-
5), 2.76 (br t, 1H, OH-6), 1.51, 1.31 (2s, each 3H, CMe2),
1.16 (s, 9H, CMe3); 13C NMR (50 MHz, CD3CN): see Table 4
and d 172.6 (CO), 113.0, (CMe2), 39.4 (CMe3), 27.5 (CMe3),

M. Landi et al. / European Journal of Medicinal Chemistry 44 (2009) 745e754
753
26.8, 26.7 (CMe2). Anal. for C14H25NO6. Calcd (%): C, 55.43;
H, 8.31; N, 4.62. Found (%): C, 55.47; H, 8.32; N, 4.64.
Table 4 and d 174.6 (CO), 110.3 (CMe2), 56.9 (OMe), 28.4,
26.6 (CMe2), 14.6 (CH ), 7.1 (CH2CH2). Anal. for
C14H25NO6. Calcd (%): C, 55.43; H, 8.31; N, 4.62. Found
(%): C, 55.45; H, 8.33; N, 4.62.
4.5.17. 3-N-(Biphenyl-4-carbonyl)-3-deoxy-1,2-O-
isopropylidene-a-D-allofuranose (4Bh)
From 8 (778 mg, 3.00 mmol) by acylation with biphenyl-4-
carbonyl chloride and subsequent hydrolysis; white solid,
685 mg (57%); Rf 0.29 (EtOAc); mp (chrom) 176e178 ꢀC; op-
tical rotation (c 1, CHCl3): [a]D þ107.0; 1H NMR (250 MHz,
CD3CN): see Table 3 and d 7.94 (m, 2H, Ar-H), 7.71 (m, 4H,
Ar-H), 7.45 (m, 3H, Ar-H), 3.65 (d, 1H, J5,OH ¼ 4.7 Hz, OH-
5), 2.88 (dd, 1H, J6a,OH ¼ 5.7 Hz, J6b,OH ¼ 6.3 Hz, OH-6),
1.55, 1.33 (2s, each 3H, CMe2); 13C NMR (62.9 MHz,
CD3CN): see Table 4 and d 168.8 (CO), 145.1, 140.7, 133.7
(3 ꢃ Ar-C), 130.0e128.0 (Ar-CH), 113.1 (CMe2), 27.0, 26.7
(CMe2). Anal. for C22H25NO6. Calcd (%): C, 66.15; H, 6.31;
N, 3.51. Found (%): C, 66.17; H, 6.32; N, 3.50.
Acknowledgements
R.G. is granted by Associazione Italiana Ricerca sul Cancro
(AIRC), Fondazione Cariparo (Cassa di Risparmio di Padova e
Rovigo), Associazione Veneta per la Lotta alla Talassemia e
Rovigo, Italy (AVLT), Telethon (grant GGP07257) and
STAMINA Project. We thank the EU Project ITHANET
(eInfrasctructure for Thalassemia Research Network) for sup-
port. M.L. was supported by a fellowship from AVLT and from
‘‘Associazione per la Lotta alla Talassemia di Ferrara e
Ferrara, Italy’’. Partial support from Universities of Pisa and
Ferrara is also acknowledged.
4.5.18. 3-N-(4-Methoxybenzoyl)-3-deoxy-1,2-O-
isopropylidene-a-D-allofuranose (4Bi)
References
From 8 (778 mg, 3.00 mmol) by acylation with 4-methox-
ybenzoyl chloride and subsequent hydrolysis; white solid,
583 mg (55%); Rf 0.35 (EtOAceMeOH 9:1); mp (chrom)
[1] B.B. Lozzio, C.B. Lozzio, Blood 45 (1975) 321e334.
[2] (a) I. Lampronti, N. Bianchi, M. Borgatti, E. Fibach, E. Prus, R. Gambari,
Eur. J. Haematol. 71 (2001) 189e195;
1
(b) R. Gambari, Minerva Biotechnol. 15 (2003) 123e128.
[3] (a) E. Fibach, N. Bianchi, M. Borgatti, E. Prus, R. Gambari, Blood 102
(2003) 1276e1281;
180e182 ꢀC; optical rotation (c 1.0, CHCl3): [a]D þ26.7; H
NMR (250 MHz, CD3CNeD2O): see Table 3 and d 7.70
(AA0XX0, 2H, H-20, H-60), 6.97 (AA0XX0, 2H, H30, H-50),
3.81 (s, 3H, OMe), 1.51, 1.28 (2s, each 3H, CMe2); 13C
NMR (62.9 MHz, CD3CN): see Table 4 and d 168.7 (CO),
163.5 (C-40), 126.8 (C-10), 130.3 (C-20, C-60), 114.8 (C-30,
C-50), 113.3 (CMe2), 56.2 (OMe), 26.9, 26.6 (CMe2). Anal.
for C17H23NO7. Calcd (%): C, 57.78; H, 6.56; N, 3.96. Found
(%): C, 67.72; H, 6.59; N, 3.95.
(b) C. Zuccato, N. Bianchi, M. Borgatti, I. Lampronti, F. Massei,
C. Favre, R. Gambari, Acta Haematol. 117 (2007) 168e176;
(c) E. Fibach, N. Bianchi, M. Borgatti, C. Zuccato, A. Finotti,
I. Lampronti, E. Prus, C. Mischiati, R. Gambari, Eur. J. Haematol. 77
(2006) 437e441;
(d) R. Gambari, E. Fibach, Curr. Med. Chem. 14 (2007) 199e212.
[4] P. Jeannesson, R. Lahlil, B. Chenais, L. Devy, R. Gillet, A. Aries,
F. Morceau, C. Trentesaux, Leuk. Lymphoma 26 (1997) 575e587.
[5] N. Bianchi, F. Osti, C. Rutigliano, F. Ginanni Corradini, E. Bosetti,
M. Tommasetti, C. Mischiati, G. Feriotto, R. Gambari, Br. J. Haematol.
104 (1999) 258e265.
[6] N. Bianchi, F. Ongaro, C. Chiarabelli, L. Gualandi, C. Mischiati,
P. Bergamini, R. Gambari, Biochem. Pharmacol. 60 (2000) 31e40.
[7] N. Bianchi, C. Chiarabelli, M. Borgatti, C. Mischiati, E. Fibach,
R. Gambari, Br. J. Haematol. 113 (2001) 951e961.
4.5.19. Methyl-6-N-butanoyl-6-deoxy-3,4-O-isopropylidene-
a-D-galactopyranoside (5Ac)
From 15 (700 mg, 3.00 mmol) by acylation with butanoyl
chloride; colourless syrup, 737 mg (81%); Rf 0.30 (EtOAce
MeOH 95:5); optical rotation (c 1.0, CHCl3): [a]D þ111.0;
1H NMR (250 MHz, CD3CNeD2O): see Table 5 and d 3.32
(s, 3H, OMe), 2.70 (br s, 1H, OH), 2.10 (t, 2H, J ¼ 7.2 Hz,
CH2CO), 2.02 (m, 2H, CH2CH2CO), 1.43, 1.29 (2s, each
3H, CMe2), 0.89 (t, 3H, J ¼ 7.4 Hz, Me); 13C NMR
(62.9 MHz, CD3CN): see Table 4 and d 173.9 (CO), 109.8
(CMe2), 55.7 (OMe), 38.7 (CH2CO), 28.3, 26.5 (CMe2),
19.9 (CH2CH2CO), 14.0 (Me). Anal. for C10H19NO5. Calcd
(%): C, 51.49; H, 8.21; N, 6.00. Found (%): C, 51.50; H,
8.23; N, 5.58.
[8] P. Pouillart, G. Ronco, I. Cerutti, J.H. Trouvin, F. Pieri, P. Villa, J. Pharm.
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[9] (a) G. Catelani, F. Osti, N. Bianchi, M.C. Bergonzi, F. D’Andrea,
R. Gambari, Bioorg. Med. Chem. Lett. 9 (1999) 3153e3158;
(b) M.C. Bergonzi, N. Bianchi, G. Catelani, F. D’Andrea, R. Gambari, M.
Lipucci di Paola, F. Osti, Eur. Patent 1,004,590 A2, 2000. Chem. Abstr.
133 (2000) 4898;
(c) G. Catelani, F. D’Andrea, E. Mastrorilli, N. Bianchi, C. Chiarabelli,
M. Borgatti, D. Martello, R. Gambari, Bioorg. Med. Chem. 10 (2002)
347e353;
(d) G. Catelani, D’Andrea, M. Landi, C. Zuccato, N. Bianchi,
R. Gambari, Carbohydr. Res. 341 (2006) 538e544.
[10] Unpublished results from Chiesi Laboratories.
4.5.20. Methyl-6-N-cyclopropanecarbonyl-6-deoxy-3,4-O-
isopropylidene-b-D-galctopyranoside (6Al)
[11] H. Kunz, P. Schmidt, Liebigs, Ann. Chem. (1982) 1245e1260.
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(b) J. Fernandez, C. Mellet, J. Jimenez, J. Fuentes, J. Org. Chem. 59
(1994) 5565e5572.
From 16 (700 mg, 3.00 mmol) by acylation with cyclopropa-
necarbonyl chloride; white solid, 679 mg (75%); Rf 0.40
(EtOAceMeOH 9:1); mp (chrom) 65e67 ꢀC; optical rotation
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[14] M.L. Wolfrom, F. Shafizadeh, R.K. Armstrong, T.M. Shen Han, J. Am.
Chem. Soc. 81 (1959) 3716e3718.
1
(c 1.1, CHCl3): [a]D þ37.2; H NMR (200 MHz, CD3CNe
D2O): see Table 5 and d 3.47 (s, 3H, OMe), 3.28 (br s, 1H,
OH), 1.48 (m, 1, CH ), 1.43, 1.30 (2s, each 3H, CMe2), 0.85e
0.66 (m, 4H, CH2CH2); 13C NMR (50 MHz, CD3CN): see
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[17] B.B. Lozzio, C.B. Lozzio, Int. J. Cancer 19 (1977) 136.

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M. Landi et al. / European Journal of Medicinal Chemistry 44 (2009) 745e754
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G. Raschella, A. Fantoni, Cell Differ. 14 (1984) 87e97.
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Mol. Pharmacol. 27 (1985) 683e688.
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66.
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[20] (a) E. Fibach, L.P. Burke, A.N. Schechter, C.T. Noguchi, G.P. Rodgers,
Blood 81 (1993) 1630e1635;
[23] F.J. Manganiello, L.W. Christensen, W.M. Jones, J. Organomet. Chem.
235 (1982) 327e334.
[24] P.A. Rao, F. Smith, J. Chem. Soc. (1944) 229e232.
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2203e2209.