Synthesis of Iminosugar-Containing KRN7000 Analogues

Article abstract of DOI:10.1002/cjoc.201600862

Three new iminosugar-containing KRN7000 (also referred to as α-GalCer) analogues were designed and synthesized. In the design, the galactose moiety of KRN7000 was replaced by iminosugars and the iminosugar structures were connected with ceramide in differ

Full text of DOI:10.1002/cjoc.201600862

FULL PAPER
DOI: 10.1002/cjoc.201600862
Synthesis of Iminosugar-Containing KRN7000 Analogues
Lei Zhang and Xin-Shan Ye*
State Key Laboratory of Natural and Biomimetic Drugs, School of Pharmaceutical Sciences, Peking University,
Xue Yuan Road No. 38, Beijing 100191, China
Three new iminosugar-containing KRN7000 (also referred to as α-GalCer) analogues were designed and syn-
thesized. In the design, the galactose moiety of KRN7000 was replaced by iminosugars and the iminosugar struc-
tures were connected with ceramide in different manners with a C-glycosidic bond instead of the O-glycosidic bond.
To our knowledge, this is the first report in which iminosugars are incorporated in KRN7000 structure modifica-
tions. The synthetic compounds were evaluated for their ability to stimulate cytokine release. The results may bene-
fit better understanding of structure-activity relationships and facilitate future design of more KRN7000 derivatives.
Keywords KRN7000 analogue, glycolipid, α-GalCer, iminosugar, synthesis
Introduction
KRN7000 (Figure 1), a synthetic α-galactosylcera-
mide (α-GalCer) derived from structural modifications
of the marine natural product named Agelasphin-9b, has
a strong ability to stimulate natural killer T (NKT) cells,
in a CD1d-restricted and T cell antigen receptor (TCR)-
mediated manner.[1,2] KRN7000 binds to CD1d mole-
cule and after being presented to NKT cells by CD1d,
stimulates rapid production of Th1 and Th2 cytokines
by NKT cells. However, the therapeutic applications of
KRN7000 were limited by the antagonizing effect be-
tween Th1 and Th2 cytokines. Therefore, it is essential
to develop more effective analogues that can produce
polarizing cytokine release profile towards either Th1 or
Th2, by structural modifications or derivations based on
KRN7000.[3,4]
A large number of KRN7000 analogues and related
compounds have been synthesized and their activity
towards NKT cell-activation has been tested.[3-10] Modi-
fications of KRN7000 are roughly based on three modi-
fiable parts: sugar head moiety, ceramide part, and sug-
ar-ceramide linkage. Sugar moiety modification studies
suggested that: (1) α-glycosidic bond is essential for
NKT cell-stimulation activity; (2) minor changes on
sugar head can result in activity loss except that C-6
position can tolerate small substitutions.[11-15] Regarding
the sugar-ceramide linkage modifications, a C-galac-
toside analogue of KRN7000, α-C-GalCer, induced a
potent Th1-type response in mice in vivo, due to the
stability of C-glycosidic bond which can resist enzy-
matic degradation at low pH.[16-18] Interestingly, re-
placement of galactose ring oxygen by carbon (carba-
Figure 1 The structures of KRN7000 and its designed ana-
logues.
α-D-galactopyranosyl analogue of KRN7000)[19] or sul-
fur
(5-thio-α-D-galactopyranosyl
analogue
of
KRN7000)[20] remarkably induced Th1-biased cytokine
production in vivo as well. Although many studies have
been performed regarding KRN7000 modifications,
accumulation of structure-activity-relationship infor-
mation is still essential to decipher the critical structural
factors of KRN7000 that induced biased cytokine re-
lease profiles.
Iminosugars are carbohydrate analogues in which
the ring oxygen is replaced by nitrogen. Iminosugars are
frequently found to be inhibitors of glycosidases that are
involved in important biological systems.[21-23] Imino-
sugar-based compounds are promising medicine candi-
dates against various diseases such as tumor metastasis,
diabetes, viral infection, and lysosomal storage disor-
ders.[23-25] Our group discovered that certain iminosugar
derivatives possess immunosuppressive activities while
*
E-mail: xinshan@bjmu.edu.cn; Tel.: 0086-010-82805736, Fax: 0086-010-82802724
Received December 5, 2016; accepted February 2, 2017; published online XXXX, 2017.
Supporting information for this article is available on the WWW under http://dx.doi.org/10.1002/cjoc.201600862 or from the author.
In Memory of Professor Enze Min.
Chin. J. Chem. 2017, XX, 1—8
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1

Zhang & Ye
FULL PAPER
minor structure changes lead to immunostimulating ac-
tivities, showing the potential as the lead compounds
with various biological activities.[26-28] Two castano-
spermine (an iminosugar) analogues also showed
up-regulation of the Th1/Th2 cytokine ratio, with ap-
parent bias towards Th1 cytokines.[29] Based on these
observations, and considering that very few examples
for iminosugar analogues of GalCer have been report-
ed,[30] it is possible to create new iminosugar-containing
compounds and explore their biological activities.
Therefore, three KRN7000 analogues were designed, in
which the galactose moiety was replaced by imino-
sugars and the iminosugars were connected with
ceramide in different manners (Figure 1, compounds 1
－3). Due to the stability of α-C-GalCer and its potent
Th1-biased responses, all the three structures adopted
C-glycosidic bonds instead of O-glycosidic bonds. Also
the linker between galactose and phytosphingosine was
elongated by different number of carbons with the aim
of studying how linker elongation affects the bioactivity
of α-GalCer.
mL of dry THF was added Ph3P＝CHCOOMe (142.5
mg, 0.415 mmol) at room temperature under nitrogen.
The reaction mixture was stirred overnight at room
temperature and then evaporated in vacuo. The residue
was purified by column chromatography with petroleum
ether/EtOAc (5∶1 to 3∶1) to give the desired product
8 as colorless oil (147.6 mg, 96%). 1H NMR (300 MHz,
CDCl3) δ: 7.31－7.19 (m, 25H), 6.00 (d, J＝16.2 Hz,
1H), 5.20－5.06 (m, 2H), 4.90－4.67 (m, 6H), 4.50－
4.20 (m, 2H), 4.07－4.00 (m, 3H), 3.73 (s, 3H), 3.49－
3.46 (m, 2H). MS (m/z): calcd for C46H47NO8: 741;
found 742 [M＋H]＋. Anal. calcd for C21H41N3O3 (%): C
74.47, H 6.39, N 1.89; found C 74.52, H 6.35, N 1.86.
Benzyl (2R,3S,4R,5S,6R)-3,4,5-tris(benzyloxy)-2-
((benzyloxy)methyl)-6-(3-hydroxy-3-oxoprop-1-en-
1-yl)piperidine-1-carboxylate (9)
A mixture of 8 (147.6 mg, 0.199 mmol) and 2 mol/L
NaOH (3.0 mL, 6.0 mmol) in MeOH (3.0 mL) was
stirred at 40 ℃ for 4 h. The mixture was cooled down
to room temperature, the H＋ resin was added to the
reaction mixture to neutralize the solution to pH＝7.
The mixture was filtered, the filtrate was concentrated.
The residue was dissolved in EtOAc, the solution was
washed with sat. NaHCO3 and brine, dried over Na2SO4,
and concentrated to give compound 9 which was di-
rectly used for the next step reaction.
Experimental
General
Air- and/or moisture-sensitive reactions were carried
out under protection of nitrogen and standard sy-
ringe/septa techniques. All chemicals were purchased as
reagent grade and used without further purification. Di-
chloromethane (CH2Cl2) was distilled over calcium hy-
dride (CaH2). Methanol was distilled from magnesium.
dimethylformamide (DMF) was stirred with CaH2 and
distilled over sodium/benzophenone. Tetrahydrofuran
(THF) was distilled over sodium/benzophenone. Ana-
lytical TLC was performed on silica gel 60-F254 pre-
coated on aluminum plates (E. Merck), with detection
by UV (254 nm) and/or by staining with acidic ceric
ammonium molybdate. Solvents were evaporated under
reduced pressure and below 35 ℃ (water bath). Or-
ganic solutions of crude products were dried over anhy-
drous Na2SO4. Column chromatography was performed
employing silica gel (200－300 mesh). 1H NMR spectra
were recorded on a JEOL AL-300, Varian INOVA-500
or Avance DRX Bruker-400 spectrometers at 25 ℃.
Chemical shifts (in parts per million) were referenced to
tetramethylsilane (TMS, δ 0) in deuterated chloroform.
13C NMR spectra were obtained by using the same
NMR spectrometers and were calibrated with CDCl3 (δ
77.00) or CD3OD (δ 49.00). Mass spectra were recorded
using a PE SCLEX QSTAR spectrometer. High-
resolution mass spectrometry (HRMS) was performed
on a Bruker APEX IV. Elemental analysis data were
recorded on a Vario EL-III element analyzer.
(2S)-2-Azido-2-((4S)-2,2-dimethyl-5-tetradecyl-1,3-
dioxolan-4-yl)ethan-1-ol (13)
To a stirred solution of 12[31] (0.80 g, 2.33 mmol) in
10 mL of DMP, catalytic amount of TsOH (19.8 mg, 5%)
was added at 0 ℃ under nitrogen. Then the reaction
was allowed to continue at room temperature for 1.5 h
under nitrogen. TLC test showed the reaction was com-
plete. The reaction was quenched by adding MeOH (15
mL) and stirred for another 1 h. The solvent was re-
moved in vacuo, and the residue was purified by column
chromatography with petroleum ether/EtOAc (8∶1 to
1
5∶1) to afford 13 as white foams (0.55g, 62%). H
NMR (500 MHz, CDCl3) δ: 4.20－4.16 (m, 1H), 4.02－
3.95 (m, 2H), 3.89－3.85 (m, 1H), 3.47 (m, 1H), 2.10
(br, 1H), 1.62－1.53 (m, 2H), 1.43 (s, 3H), 1.34 (s, 3H),
1.40－1.26 (m, 24H), 0.88 (t, J＝7.0 Hz, 3H); 13C NMR
(125 MHz, CDCl3) δ: 108.42, 77.73, 76.69, 63.96, 61.16,
31.91, 29.66, 29.57, 29.52, 29.38, 29.34, 28.01, 26.51,
25.54, 22.68, 14.10. MS (m/z): calcd for C21H41N3O3:
383; found 356 [M＋HN2]＋, 401 [M＋NH4]＋. Anal.
calcd for C21H41N3O3 (%): C 65.76, H 10.77, N 10.96;
found C 66.02, H 10.63, N 10.91.
2-((2S)-2-Azido-2-((4S)-2,2-dimethyl-5-tetradecyl-
1,3-dioxolan-4-yl)ethyl)isoindoline-1,3-dione (14)
To a solution of 13 (123.3 mg, 0.321 mmol) in dry
toluene under nitrogen was added Ph3P (168.4 mg,
0.642 mmol), phthalimide (94.6 mg, 0.642 mmol) and
DEAD (106 L, 0.642 mmol). The reaction mixture was
stirred at room temperature under nitrogen for 1.5 h.
The solvent was removed in vacuo. The residue was
Benzyl(2R,3S,4R,5S,6R)-3,4,5-tris(benzyloxy)-2-
((benzyloxy)methyl)-6-(3-methoxy-3-oxoprop-1-en-1-
yl)piperidine-1-carboxylate (8)
To a solution of 6[26] (142.2 mg, 0.208 mmol) in 8
2
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Chin. J. Chem. 2017, XX, 1—8

Synthesis of Iminosugar-Containing KRN7000 Analogues
Benzyl (2S,3S,4S,5S,6R)-2-(((2S,3S,4R)-2-azido-3,4-
dihydroxyoctadecyl)carbamoyl)-3,4,5-tris(benzyl-
oxy)-6-((benzyloxy)methyl)piperidine-1-carboxylate
(17)
purified by column chromatography with petroleum
ether/EtOAc (10∶1) to provide 14 as white foams
1
(157.5 mg, 96%). H NMR (300 MHz, CDCl3) δ: 7.88
(dd, J＝5.4, 3.0 Hz, 2H), 7.75 (dd, J＝5.4, 3.3 Hz, 2H),
4.20 (dd, J＝13.2, 5.4 Hz, 1H), 4.09－3.94 (m, 3H),
3.77 (td, J＝9.0, 4.2 Hz, 1H), 1.64－1.54 (m, 3H), 1.50
(s, 3H), 1.35 (s, 3H), 1.38－1.21 (m, 23H), 0.87 (t, J＝
6.6 Hz, 3H); 13C NMR (75 MHz, CDCl3) δ: 168.22,
134.16, 131.87, 123.43, 108.65, 77.84, 58.60, 39.59,
31.89, 29.65, 29.55, 29.50, 29.33, 29.11, 27.81, 26.57,
25.54, 22.66, 14.10. MS (m/z) calcd for C29H44N4O4:
512.3; found 530.2 [M ＋ NH4] ＋ . Anal. calcd for
C29H44N4O4 (%): C 67.94, H 8.65, N 10.93; found C
68.07, H 8.54, N 10.83.
To compound 16 (174.4 mg, 0.164 mmol) in 7 mL of
MeOH/THF (V∶V＝1∶1) was added 0.2 mL of con-
centrated HCl, and the reaction mixture was stirred
overnight at room temperature. TLC test showed that
the reaction was complete. The solvent was removed in
vacuo, and the residue was purified using column
chromatography with petroleum ether/EtOAc (2∶1) to
1
afford 17 as colorless oil (158.9 mg, 95%). H NMR
(500 MHz, CDCl3) δ: 7.37－7.19 (m, 25H), 5.08 (d, J＝
12.0 Hz, 1H), 4.90 (d, J＝12.0 Hz, 1H), 4.74－4.60 (m,
6H), 4.48－4.28 (m, 3H), 4.24－4.18 (m, 1H), 4.10－
4.02 (m, 2H), 3.94－3.83 (m, 3H), 3.69－3.65 (m, 2H),
3.50 (brs, 1H), 3.43 (brs, 1H), 3.23 (brs, 1H), 2.38 (d,
J＝6.5 Hz, 1H), 1.74 (d, J＝2.5 Hz, 1H), 1.54－1.40 (m,
(2S,3S,4R)-2-Azido-3,4-isopropylidene ketal-octade-
cane-1-amine (15)
A mixture of 14 (166.0 mg, 0.324 mmol) and
NH2NH2•H2O (3.0 mL) in MeOH (6.0 mL) was heated
at 70 ℃ for 0.5 h. The mixture was cooled down to
room temperature. The reaction mixture was concen-
trated, and the residue was dissolved in EtOAc. The
solution was washed with sat. brine, dried over Na2SO4,
and concentrated to dryness, yielding 15 which was di-
rectly used for the next step reaction.
13
3H), 1.31－1.26 (m, 23H), 0.88 (t, J＝7.0 Hz, 3H); C
NMR (125 MHz, CDCl3) δ: 170.79, 138.37, 138.12,
138.00, 137.45, 136.02, 128.49, 128.41, 128.36, 128.33,
128.25, 128.05, 127.99, 127.70, 127.66, 127.61, 127.54,
78.95, 75.71, 74.22, 73.77, 73.34, 72.96, 72.86, 71.95,
67.80, 67.32, 62.44, 56.71, 56.58, 39.91, 31.90, 31.53,
29.67, 29.34, 25.85, 22.67, 14.10. MS (m/z): calcd for
C61H79N5O9: 1025.6; found 1026.3 [M＋H]＋, 1048.3
[M＋Na]＋. Anal. calcd for C61H79N5O9 (%): C 71.39, H
7.76, N 6.82; found C 71.51, H 7.93, N 6.80.
Benzyl (2S,3S,4S,5S,6R)-2-(((2S)-2-azido-2-((4S)-
2,2-dimethyl-5-tetradecyl-1,3-dioxolan-4-yl)ethyl)-
carbamoyl)-3,4,5-tris(benzyloxy)-6-((benzyloxy)-
methyl)piperidine-1-carboxylate (16)
Benzyl (2R,3S,4S,5S,6S)-3,4,5-tris(benzyloxy)-2-
((benzyloxy)methyl)-6-(((2S,3S,4R)-2-hexacosan-
amido-3,4-dihydroxyoctadecyl)carbamoyl)piperi-
dine-1-carboxylate (18)
To a mixture of 7[26] (150 mg, 0.214 mmol) and 15
(91.3 mg, 0.238 mmol) in dry THF, HBTU (99.2 mg,
0.262 mmol) and DIEA (91.4 L, 0.523 mmol) were
added under nitrogen. The reaction mixture was stirred
overnight at room temperature under nitrogen. TLC de-
tection showed the reaction was complete, then the sol-
vent was removed in vacuo, and the residue was puri-
fied by column chromatography with petroleum ether/
EtOAc (15∶1 to 6∶1) to afford 16 as colorless oil
To a solution of 17 (37.6 mg, 0.0366 mmol) in 7 mL
of MeOH were added NiCl2•6H2O (17.4 mg, 0.0733
mmol) and NaBH4 (6.8 mg, 0.183 mmol) at 0 ℃. The
reaction mixture was allowed to warm up to room tem-
perature and stirred overnight. A few drops of ammo-
nium hydroxide were added to the reaction mixture and
the solvent was removed in vacuo. The residue was
re-dissolved in EtOAc, and washed by saturated Na-
HCO3 and NaCl solution. The organic layer was dried
over anhydrous Na2SO4, filtered and concentrated. The
crude product was directly used for the next step. The
crude product was dried by vacuum and dissolved in 5
mL of dry THF. C25H51COOH (29.0 mg, 0.0733 mmol),
HBTU (27.7 mg, 0.0733 mmol) and DIEA (19.2 L,
0.110 mmol) were added under nitrogen, and the reac-
tion mixture was stirred overnight at room temperature.
TLC test showed the reaction was complete. The solvent
was removed in vacuo. Column chromatography with
petroleum/EtOAc (1∶1) provided 18 as colorless oil
(47.4 mg, 94%). 1H NMR (500 MHz, CDCl3) δ: 7.31－
7.20 (m, 25H), 6.78 (brs, 1H), 6.44 (brs, 1H), 5.10 (d,
J＝12.5 Hz, 1H), 4.90 (d, J＝12.5 Hz, 1H), 4.66－4.62
(m, 3H), 4.56 (d, J＝11.5 Hz, 2H), 4.52－4.38 (m, 5H),
4.18 (dd, J＝6.0, 3.5 Hz, 1H), 4.00 (brs, 3H), 3.87 (brs,
1H), 3.76 (dd, J＝6.5, 3.5 Hz, 1H), 3.65 (brs, 1H), 3.49
1
(207.5 mg, 91%). H NMR (500 MHz, CDCl3) δ:
7.29－7.20 (m, 25H), 5.09 (d, J＝11.5 Hz, 1H), 4.88 (d,
J＝12.0 Hz, 2H), 4.76－4.65 (m, 6H), 4.52－4.31 (m,
3H), 4.25 (dd, J＝7.5, 4.5 Hz, 1H), 4.11－4.03 (m, 3H),
3.98－3.80 (m, 2H), 3.76 (d, J＝5.0 Hz, 1H), 3.68 (dd,
J＝8.5, 5.5 Hz, 1H), 3.47 (brs, 1H), 3.28－2.98 (m, 1H),
1.56－1.48 (m, 3H), 1.40 (s, 3H), 1.34－1.22 (m, 26H),
1.25 (s, 3H), 0.85 (t, J＝7.0 Hz, 3H); 13C NMR (125
MHz, CDCl3) δ: 169.04, 138.56, 138.35, 138.20, 137.72,
136.11, 128.44, 128.39, 128.31, 128.21, 127.97, 127.91,
127.57, 127.48, 108.38, 79.97, 77.63, 77.11, 76.12,
74.17, 73.38, 73.08, 72.92, 67.68, 59.92, 56.64, 41.18,
31.91, 29.69, 29.65, 29.60, 29.34, 29.28, 27.96, 26.56,
25.52, 22.68, 22.37, 16.52, 14.11. MS (m/z): calcd for
C64H83N5O9: 1065.6; found 1066.4 [M＋H]＋, 1089.3
[M＋Na]＋. Anal. calcd for C64H83N5O9 (%): C 72.08, H
7.85, N 6.57; found C 71.97, H 7.72, N 6.52.
Chin. J. Chem. 2017, XX, 1—8
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Zhang & Ye
FULL PAPER
(brs, 1H), 3.44 (brs, 1H), 3.30－3.25 (m, 1H), 2.35 (brs,
1H), 2.07－1.97 (m, 2H), 1.57－1.51 (m, 3H), 1.50－
1.44 (m, 1H), 1.40－1.18 (m, 68H), 0.88 (t, J＝7.0 Hz,
6H); 13C NMR (75 MHz, CDCl3) δ: 173.85, 170.71,
137.96, 137.74, 135.91, 128.49, 128.41, 128.36, 128.15,
127.93, 127.85, 127.72, 127.60, 75.24, 74.35, 73.50,
73.15, 73.00, 72.77, 67.73, 66.68, 56.47, 52.14, 39.95,
36.68, 32.37, 31.92, 29.70, 29.56, 29.43, 29.36, 25.95,
25.62, 22.69, 14.13. HRMS: calcd for C87H132N3O10
C66H85N5O9 (%): C 72.56, H 7.84, N 6.41; found C
72.41, H 7.76, N 6.28.
Benzyl (3S,4R,5S,6R)-2-(3-(((2S,3S,4R)-2-azido-3,4-
dihydroxyoctadecyl)amino)-3-oxoprop-1-en-1-yl)-
3,4,5-tris(benzyloxy)-6-((benzyloxy)methyl)piperi-
dine-1-carboxylate (20)
Treatment of 19 (62.1 mg, 0.0568 mmol) with con-
centrated HCl as described in the synthesis of 17, led to
20 (52.2 mg, 87%) as colorless oil. The product was a
mixture of separable E- and Z-isomers with a ratio of
E-/Z-configuration 5∶1. E-Isomer: 1H NMR (300 MHz,
CDCl3) δ: 7.34－7.21 (m, 25H), 5.96－5.77 (m, 1H),
5.21 (t, J＝6.0 Hz, 1H), 5.08 (d, J＝12.3 Hz, 1H), 4.93
(d, J＝11.7 Hz, 1H), 4.80－4.16 (m, 10H), 4.10－4.00
(m, 1H), 3.98 (s, 1H), 3.91－3.60 (m, 5H), 2.70－2.43
(m, 1H), 1.83 (s, 1H), 1.51－1.47 (m, 3H), 1.46－1.25
[M ＋ H] ＋ 1378.9907, found 1378.9975; calcd for
＋
C87H131N3O10Na [M ＋ Na]
1400.9727, found
1400.9764.
(2S,3S,4S,5S,6R)-N-((2S,3S,4R)-2-Hexacosanamido-
3,4-dihydroxyoctadecyl)-3,4,5-trihydroxy-6-(hydro-
xymethyl)piperidine-2-carboxamide (1)
To a solution of 18 (20.0 mg, 0.0145 mmol) in 6 mL
of THF/H2O (V∶V＝2∶1) was added concentrated
HCl (0.6 mL), catalytic amount of Pd/C, and the mix-
ture was stirred for 3 d under H2 atmosphere. The reac-
tion mixture was filtered and the filtrate was concen-
trated, and the residue was purified by column chroma-
tography with CH2Cl2/MeOH (15∶1 to 3∶1) to afford
1 as white foams (12.8 mg, 100%). 1H NMR (500 MHz,
CDCl3/CD3OD) δ: 4.16 (dd, J＝10.0, 6.0 Hz, 1H), 4.00
(d, J＝6.0 Hz, 1H), 3.86 (s, 1H), 3.74－3.70 (m, 2H),
3.63 (dd, J＝13.5, 3.5 Hz, 1H), 3.57－3.51 (m, 3H),
3.39 (d, J＝9.5 Hz, 1H), 2.72 (brs, 1H), 2.28－2.18 (m,
2 H), 1.61－1.57 (m, 4H), 1.42－1.40 (m, 2H),
1.31－1.27 (m, 67H), 0.89 (t, J＝7.0 Hz, 6H); 13C NMR
(125 MHz, CDCl3/CD3OD) δ: 175.24, 75.41, 72.22,
71.21, 69.51, 68.96, 62.65, 57.66, 56.08, 53.72, 50.02,
30.01, 29.93, 29.78, 29.71, 29.60, 26.21, 22.91, 14.20.
HRMS: calcd for C51H102N3O8 [M＋H]＋ 884.7661,
found 884.7681.
1
(m, 23H), 0.88 (t, J＝6.6 Hz, 3H). Z-Isomer: H NMR
(500 MHz, CDCl3) δ: 7.37－7.05 (m, 25H), 6.82 (s, 1H),
5.84－5.71 (m, 1H), 5.24 (d, J＝12.5 Hz, 1H), 5.15 (dd,
J＝10.5, 2.0 Hz, 1H), 5.08 (d, J＝12.0 Hz, 1H), 4.86
(brs, 1H), 4.79 (d, J＝8.0 Hz, 1 H), 4.65 (dd, J＝12.0,
9.5 Hz, 1H), 4.60－4.44 (m, 3H), 4.35 (dd, J＝16.0,
11.5 Hz, 1H), 4.26 (d, J＝3.5 Hz, 1H), 4.18 (t, J＝11.5
Hz, 1H), 4.03 (dd, J＝6.0, 3.0 Hz, 1H), 3.96 (dd, J＝6.0,
3.0 Hz, 1H), 3.88－3.86 (m, 1H), 3.82 (d, J＝10.5 Hz,
1H), 3.75－3.68 (m, 1H), 3.68－3.52 (m, 1H), 3.44－
3.41 (m, 1H), 3.39－3.34 (m, 1H), 3.32－3.28 (m, 1H),
3.25－3.14 (m, 2H), 3.12－3.04 (m, 1H), 2.70－2.61
(m, 1H), 2.33 (dd, J＝13.0, 7.5 Hz, 1H), 1.56 (s, 3H),
1.48－1.40 (m, 2H), 1.31－1.26 (m, 22H), 0.88 (t, J＝
7.0 Hz, 3H); 13C NMR (125 MHz, CDCl3) δ: 173.09,
172.75, 155.00, 138.32, 138.11, 137.72, 136.39, 131.75,
128.65, 128.53, 128.42, 128.34, 128.18, 128.03, 127.97,
127.74, 127.65, 127.43, 127.36, 81.13, 74.08, 73.93,
73.16, 72.72, 72.37, 72.24, 71.80, 71.48, 67.90, 67.07,
65.86, 62.47, 56.72, 56.35, 55.79, 52.70, 40.06, 39.88,
36.31, 36.22, 31.92, 31.37, 31.21, 29.69, 29.36, 25.87,
22.68, 14.12. MS (m/z): calcd for C63H81N5O9: 1051.6,
found 1052.4 [M＋H]＋. Anal. calcd for C63H81N5O9 (%):
C 71.90, H 7.76, N 6.65; found C 71.74, H 7.89, N6.79.
Benzyl (3S,4R,5S,6R)-2-(3-(((2S)-2-azido-2-((4S)-2,2-
dimethyl-5-tetradecyl-1,3-dioxolan-4-yl)ethyl)-
amino)-3-oxoprop-1-en-1-yl)-3,4,5-tris(benzyloxy)-
6-((benzyloxy)methyl)piperidine-1-carboxylate (19)
The synthetic procedure was the same as that de-
scribed in the synthesis of compound 16, using 9 (24.8
mg, 0.0341 mmol) and 15 (13.0 mg, 0.0341 mmol),
HBTU (25.8 mg, 0.0682 mmol), and DIEA (12.0 L,
0.0682 mmol) to afford 19 (34.9 mg, 80%) as colorless
oil. The product was a mixture of separable E- and
Z-isomers with a ratio of E-/Z-configuration 5∶1.
Benzyl (2R,3S,4R,5S)-3,4,5-tris(benzyloxy)-2-((ben-
zyloxy)methyl)-6-(3-(((2S,3S,4R)-2-hexacosanami-
do-3,4-dihydroxyoctadecyl)amino)-3-oxopropyl)pi-
peridine-1-carboxylate (21)
Compound 21 was synthesized by the same proce-
dure as that in the preparation of 18. Compound 20
(102.1 mg, 0.0970 mmol) was reduced and conjugated
with C25H51COOH. Following column chromatography
compound 21 was obtained (45.0 mg, 33%) as colorless
1
E-isomer: H NMR (300 MHz, CDCl3) δ: 7.36－7.18
(m, 25H), 7.08－7.06 (m, 1H), 6.60－6.55 (m, 1H),
5.84－5.79 (m, 1H), 5.30－5.19 (m, 2H), 4.92 (d, J＝
12.0 Hz, 1H), 4.78 (d, J＝10.5 Hz, 1H), 4.66－4.03 (m,
8H), 3.90－3.63 (m, 4H), 3.49－3.46 (m, 1H), 3.25－
3.04 (m, 3H), 2.77－2.61 (m, 1H), 1.60－1.26 (m, 32H),
1
oil. H NMR (500 MHz, CDCl3) δ: 7.31－7.14 (m,
25H), 6.86 (brs, 1H), 6.27 (brs, 1H), 5.02 (brs, 1H), 4.88
(brs, 1H), 4.69－4.47 (m, 5H), 4.37－4.35 (m, 2H),
4.25－4.18 (m, 2H), 4.12－3.94 (m, 3H), 3.81－3.71
(m, 2H), 3.56 (d, J＝10.0 Hz, 2H), 3.45 (brs, 4H), 2.83
(brs, 1H), 2.23 (brs, 1H), 2.14－2.09 (m, 4H), 1.76 (brs,
2H), 1.63－1.56 (m, 3H), 1.50－1.48 (m, 1H), 1.38－
1.13 (m, 68H), 0.88 (t, J＝7.0 Hz, 6H); 13C NMR (125
1
0.88 (t, J＝7.2 Hz, 3H). Z-Isomer: H NMR (300 MHz,
CDCl3) δ: 7.33－7.16 (m, 25H), 5.88－5.67 (m, 1H),
5.27－5.05 (m, 3H), 4.78－4.30 (m, 6H), 4.01－3.48
(m, 8H), 3.48－3.02 (m, 3H), 1.58－1.25 (m, 32H),
0.88 (t, J＝6.6 Hz, 3H). MS (m/z): calcd for C66H85N5O9:
1091.6; found 1092.6 [M ＋ H] ＋ . Anal. calcd for
4
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Chin. J. Chem. 2017, XX, 1—8

Synthesis of Iminosugar-Containing KRN7000 Analogues
MHz, CDCl3) δ: 174.67, 174.26, 157.79, 138.59, 138.31,
137.80, 135.85, 130.85, 128.49, 128.42, 128.33, 128.18,
128.05, 127.85, 127.78, 127.63, 127.52, 127.44, 74.74,
74.36, 73.97,73.21, 72.95, 68.14, 67.85, 55.14, 54.02,
52.54, 40.84, 36.74, 32.90, 31.91, 30.34, 29.70, 29.55,
29.42, 29.36, 28.91, 26.01, 25.70, 23.73, 22.97, 22.67,
14.10. HRMS: calcd for C89H136N3O10 [M ＋ H] ＋
1407.0220, found 1407.0231.
23 (60.7 mg, 92%) colorless oil. 1H NMR (500 MHz,
CDCl3) δ: 7.86－7.53 (m, 1H), 7.33－7.19 (m, 25H),
5.18 (d, J＝8.0 Hz, 1H), 5.11 (d, J＝11.5 Hz, 1H), 4.90
(brs, 1H), 4.72 (s, 2H), 4.55 (s, 2H), 4.46 (d, J＝9.5 Hz,
2H), 4.38 (d, J＝12.0 Hz, 2H), 4.32 (t, J＝7.5 Hz, 1H),
4.25－4.11 (m, 1H), 3.92 (t, J＝3.5 Hz, 2H), 3.58 (d,
J＝4.5 Hz, 3H), 3.48 (brs, 1H), 3.35 (s, 1H), 3.23 (brs,
1H), 3.00 (brs, 1H), 2.32 (s, 1H), 1.66 (s, 1H), 1.54－
1.50 (m, 1H), 1.42 (brs, 2H), 1.32－1.25 (m, 23H), 0.88
(t, J＝7.0 Hz, 3H); 13C NMR (125 MHz, CDCl3) δ:
172.37, 156.47, 138.16, 137.86, 137.76, 137.47, 136.18,
129.78, 128.53, 128.50, 128.41, 128.34, 128.16, 128.05,
127.99, 127.75, 127.67, 127.63, 74.72, 74.14, 73.08,
72.68, 72.52, 71.82, 71.64, 68.45, 67.87, 62.73, 57.84,
55.04, 39.93, 31.91, 31.48, 29.68, 29.64, 29.34, 25.90,
22.67, 14.11; MS (m/z): calcd for C61H79N5O9: 1025.6;
found 1026.3 [M＋H]＋, 1048.2 [M＋Na]＋. Anal. calcd
for C61H79N5O9 (%): C 71.39, H 7.76, N 6.82; found C
71.14, H 7.86, N 6.70.
N-((2S,3S,4R)-3,4-Dihydroxy-1-(3-((3S,4R,5S,6R)-
3,4,5-trihydroxy-6-(hydroxymethyl)piperidin-2-yl)-
propanamido)octadecan-2-yl)hexacosanamide (2)
Compound 21 (20.0 mg, 0.0142 mmol) was depro-
tected and the product was purified as in the preparation
1
of 1, providing 2 (13.0 mg, 100%) as white foams. H
NMR (500 MHz, CDCl3/CD3OD) δ: 4.19－4.11 (m,
3H), 3.75－3.71 (m, 1H), 3.68－3.50 (m, 5H), 3.27 (brs,
1H), 3.00 (brs, 1H), 2.76－2.66 (m, 1H), 2.52 (ddd, J＝
15.5, 8.0, 5.0 Hz, 1H), 2.33－2.23 (m, 3H), 2.07－1.98
(m, 1H), 1.61－1.55 (m, 4H), 1.45－1.14 (m, 69H),
0.88 (t, J ＝ 7.0 Hz, 6H); 13C NMR (150 MHz,
MeOD/CDCl3) δ: 175.90, 175.67, 75.92, 72.61, 70.83,
68.56, 66.33, 51.15, 40.91, 37.15, 33.99, 32.71, 32.70,
30.53, 30.52, 30.48, 30.47, 30.42, 30.40 30.36, 30.26,
30.13, 30.10, 26.76, 26.67, 23.39, 14.37. HRMS: calcd
for C53H106N3O8 [M＋H]＋ 912.7974, found 912.7993.
Benzyl (2S,3S,4S,5S,6S)-3,4,5-tris(benzyloxy)-2-
((benzyloxy)methyl)-6-(((2S,3S,4R)-2-hexacosan-
amido-3,4-dihydroxyoctadecyl)carbamoyl)piperi-
dine-1-carboxylate (24)
Compound 24 was synthesized by the same proce-
dure as the preparation of 18. Compound 23 (31.8 mg,
0.0310 mmol) was reduced and conjugated with
C25H51COOH. Following column chromatography
compound 24 was obtained (30.3 mg, 71%) as colorless
Benzyl (2S,3S,4S,5S,6S)-2-(((2S)-2-azido-2-((4S)-2,2-
dimethyl-5-tetradecyl-1,3-dioxolan-4-yl)ethyl)car-
bamoyl)-3,4,5-tris(benzyloxy)-6-((benzyloxy)methyl)-
piperidine-1-carboxylate (22)
1
oil. H NMR (500 MHz, CDCl3) δ: 7.34－7.15 (m,
25H), 6.17 (brs, 2H), 5.14 (s, 2H), 4.83 (d, J＝6.5 Hz,
1H), 4.67 (s, 2H), 4.57－4.50 (m, 4H), 4.42 (d, J＝12.0
Hz, 2H), 4.35 (t, J＝7.5 Hz, 2H), 3.92 (s, 4H), 3.64 (dd,
J＝10.0, 7.0 Hz, 2H), 3.57 (brs, 1H), 3.38 (s, 2H), 3.23
(dd, J＝11.5, 6.0 Hz, 2H), 2.44 (d, J＝8.0 Hz, 1H), 1.93
(brs, 3H), 1.74 (s, 1H), 1.51－1.41 (m, 6H), 1.31－1.13
(m, 62H), 0.88 (t, J＝7.0 Hz, 6H); 13C NMR (125 MHz,
CDCl3) δ: 173.58, 172.18, 156.56, 138.10, 137.83,
137.62, 136.07, 128.59, 128.57, 128.43, 128.36, 128.17,
128.10, 127.99, 127.78, 127.70, 127.62, 74.80, 74.25,
74.01, 73.06, 72.77, 72.61, 71.69, 68.80, 67.91, 58.47,
55.35, 51.73, 40.22, 36.56, 32.37, 31.91, 29.70, 29.57,
The synthetic procedure was the same as that de-
scribed in the synthesis of compound 16, using 10[26]
(97.8 mg, 0.140 mmol), 15 (59.5 mg, 0.155 mmol),
HBTU (64.6 mg, 0.170 mmol), and DIEA (89.3 L,
0.511 mmol) to afford 22 (146.7 mg, 99%) as colorless
1
oil. H NMR (500 MHz, CDCl3) δ: 7.36－7.17 (m,
25H), 5.15 (dd, J＝19.5, 12.5 Hz, 2H), 4.98 (brs, 1H),
4.79 (brs, 1H), 4.68 (brs, 1H), 4.52 (dd, J＝32.0, 11.5
Hz, 5H), 4.39－4.36 (m, 3H), 4.03 (d, J＝28.5 Hz, 2H),
3.83 (brs, 1H), 3.61 (s, 3H), 3.40－3.25 (m, 1H), 2.95
(brs, 1H), 2.68 (brs, 1H), 1.49 (brs, 3H), 1.38 (s, 3H),
1.31－1.24 (m, 23H), 1.23 (s, 3H), 0.88 (t, J＝7.0 Hz,
3H); 13C NMR (125 MHz,CDCl3) δ: 170.05, 138.30,
138.03, 137.79, 136.32, 128.46, 128.31, 128.30, 128.09,
127.91, 127.66, 127.55, 108.26, 77.58, 74.76, 74.44,
73.39, 72.74, 72.50, 71.74, 68.90, 67.77, 60.41, 59.10,
57.38, 55.29, 40.80, 31.91, 29.68, 29.60, 29.34, 29.18,
27.97, 26.59, 25.54, 22.68, 14.11. MS (m/z): calcd for
C64H83N5O9: 1065.6; found 1066.4 [M＋H]＋, 1088.3
[M＋Na]＋. Anal. calcd for C64H83N5O9 (%): C 72.08, H
7.85, N 6.57; found C 71.96, H 7.81, N 6.68.
29.43, 29.35, 26.01, 25.55, 22.67, 14.10. HRMS: calcd
＋
for C87H132N3O10 [M ＋ H]
1378.9907, found
1378.9975.
(2S,3S,4S,5S,6S)-N-((2S,3S,4R)-2-Hexacosanamido-
3,4-dihydroxyoctadecyl)-3,4,5-trihydroxy-6-(hydro-
xymethyl)piperidine-2-carboxamide (3)
Compound 24 (42.4 mg, 0.0308 mmol) was depro-
tected and the product was purified as in the synthesis of
compound 1, providing 3 (27.1 mg, 100%) as white
1
foams. H NMR (500 MHz, CDCl3/CD3OD) δ: 4.27－
Benzyl (2S,3S,4S,5S,6S)-2-(((2S,3S,4R)-2-azido-3,4-
dihydroxyoctadecyl)carbamoyl)-3,4,5-tris(benzyl-
oxy)-6-((benzyloxy)methyl)piperidine-1-carboxylate
(23)
4.22 (m, 2H), 4.02－4.00 (m, 3H), 3.97－3.95 (m, 2H),
3.70 (dd, J＝14.0, 3.0 Hz, 1H), 3.56－3.53 (m, 1H),
3.50 (t, J＝6.0 Hz, 1H), 3.21－3.15 (m, 2H), 2.27 (t,
J＝7.5 Hz, 2H), 1.70－1.50 (m, 6H), 1.39－1.23 (m,
66H), 0.89 (t, J＝7.0 Hz, 6H); 13C NMR (100 MHz,
CDCl3/CD3OD) δ: 175.03, 74.67, 71.22, 69.54, 69.26,
Treatment of 22 (68.8 mg, 0.0645 mmol) with con-
centrated HCl as described in the synthesis of 17, led to
Chin. J. Chem. 2017, XX, 1—8
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www.cjc.wiley-vch.de
5

Zhang & Ye
FULL PAPER
62.48, 55.68, 54.15, 49.42, 39.54, 35.85, 31.33, 29.13,
29.07, 28.93, 28.84, 28.75, 25.32, 22.05, 13.21. HRMS:
calcd for C51H102N3O8 [M ＋ H] ＋ 884.7661, found
884.7690.
Scheme 1 Synthetic pathway of iminosugars
Spleen cell proliferation assay
The splenocytes (8×105 cells/well) were plated in
96-well flat-bottom tissue culture plates with synthetic
compound (100 ng/mL, 100 μL/well) diluted in 200 μL
of medium. After 48 h at 37 ℃, CCK-8 (20 μL) was
added to the cultured cell and the colorimetric values
were measured by a microplate reader with 600 nm as
reference.
In vivo stimulation with synthetic compounds
Stock solutions of KRN 7000 and synthetic com-
pounds were prepared in 100% DMSO at a concentra-
tion of 1 mg/mL. Before use, the solutions were diluted
with phosphate buffered saline (pH 7.4) to obtain a final
concentration of 10 μg/mL. Mice were injected intraper-
itoneally with 5 μg of compound or with diluted DMSO
alone. Sera were collected at 2 time points, and the lev-
els of IFN-γ (at 16 h) and IL-4 (at 2 h) were measured
by a standard sandwich ELISA using purified capture
and biotin-conjugated detection monoclonal antibodies
and standards. ELISAs were developed with TMB sub-
strate, followed by evaluation using a microplate reader.
Regents and conditions: (a) TEMPO, DAIB, CH2Cl2, r.t., 99%; (b)
NaClO2 (aq.), NaH2PO4 (aq.), 30% H2O2, r.t., 83%; (c) Ph3P＝
CHCO2Me, THF, r.t., 96%; (d) 2 mol/L NaOH (aq.), MeOH, 40 ℃,
quantitative; (e) TEMPO, DAIB, CH2Cl2, r.t. 94%
Scheme 2 Synthetic pathway of phytosphingosine chain
Results and Discussion
The synthesis was divided into three parts: the
iminosugar synthesis, the phytosphingosine chain syn-
thesis, and the conjugation. The iminosugar synthesis
started from galactose. As shown in Scheme 1, com-
pounds 4 and 5 were prepared according to our previ-
ously reported procedures.[26-28] The oxidation of 4 with
TEMPO/DAIB led to aldehyde 6, which was further
oxidized by NaClO2 and H2O2 to give carboxylic acid 7.
The Witting reaction between 6 and ylide Ph3P＝
CHCO2Me followed by ester hydrolysis afforded com-
pound 9. Unlike compound 4, oxidation of 5 with
TEMPO/DAIB at room temperature overnight directly
provided carboxylic acid 10.
The synthetic route of phytosphingosine chain began
with mannose. As shown in Scheme 2, following the
method developed previously,[31] 2,3;5,6-di-O-iso-pro-
pylidene-D-mannofuranose (11) was converted to
sphingosine 12 through the seven-step manipulation in a
total yield of 69%. Protection of 12 with 2,2-dimetho-
xypropane (DMP) in the presence of a catalytic amount
of TsOH gave isopropylidene ketal protected compound
13. This step could lead to the side product in which
1,3-hydroxyl groups were protected instead. The side
product could be converted to 12 after acidic deprotec-
tion and then be reused. The Mitsunobu reaction on the
terminal hydroxyl group in 13, which was followed by
the hydrazine deprotection of phthalimide 14, afforded
amine 15.
Reagents and conditions: (a) acetone, H2SO4, r.t., then NaOH (aq.),
86%; (b) TsOH, DMP, 62%; (c) Phthalimide, DEAD, Ph3P, toluene,
96%; (d) NH2NH2•H2O, MeOH, 70 ℃, quantitative
The target compounds 1－3 were assembled via the
coupling reactions between sugar heads and sphingosine
chain, which was followed by lipid chain (hexacosanoic
acid) conjugation (Scheme 3). The carboxylic acid on
sugar heads was conjugated to the terminal amine pre-
sent in sphingosine 15 to form the corresponding amide
compounds. The isopropylidene protective group was
removed by concentrated hydrochloric acid using
MeOH/THF as solvent. It was found that the removal of
isopropylidene ketal is necessary for the next-step azide
reduction. Azide reduction was not successful in the
6
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Chin. J. Chem. 2017, XX, 1—8

Synthesis of Iminosugar-Containing KRN7000 Analogues
Scheme 3 Synthesis of final compounds 1－3
OBn
BnO
OBn
N
BnO
BnO
CBz
CBz
c
a
N
N3
O
b
N3 OH
C14H29
H
N
H
N
+
15
BnO
7
BnO
C14H29
BnO
C
C
O
OH
O
O
17
16
O
C25H51
NH OH
O
C25H51
NH OH
OBn
OH
NH
BnO
BnO
HO
HO
CBz
d
N
H
N
H
N
BnO
C14H29
HO
C14H29
C
C
OH
OH
O
O
18
1
OBn
N
BnO
OBn
BnO
CBz
CBz
N
b
c
a
BnO
O
BnO
O
+
N3 OH
9
15
N3
O
BnO
BnO
NH
NH
C14H29
C14H29
OH
O
20
19
OH
NH
HO
HO
OBn
N
BnO
C25H51
NH OH
CBz
C25H51
O
O
O
BnO
d
O
NH OH
HO
BnO
NH
NH
C14H29
C14H29
OH
OH
N3
21
2
BnO
BnO
CBz
CBz
N
N
c
BnO
BnO
a
b
BnO
BnO
O
N3 OH
15
H
N
10 +
BnO
O
H
N
HO
BnO
O
C14H29
C14H29
O
23
22
HO
BnO
O
O
CBz
C25H51
OH
NH
C25H51
OH
N
d
HO
HO
BnO
BnO
HN
HN
H
HO
H
N
BnO
O
N
C14H29
O
C14H29
OH
OH
3
24
Reagents and conditions: (a) HBTU, DIEA, THF, 91% for 16, 80% for 19, 99% for 22; (b) HCl, MeOH/THF, 95% for 17, 87% for 20, 92% for 23;
(c) i) NaBH4, NiCl2•6H2O, MeOH; ii) C25H51COOH, HBTU, DIEA, THF; 94% for 18, 33% for 21, 71% for 24; (d) Pd/C, H2, THF/H2O/HCl, quan-
titative
presence of isopropylidene ketal probably due to steric
hindrance. After removal of the isopropylidene group,
the azido group on sphingosine chain was reduced by
NaBH4 and NiCl2•6H2O to yield the crude product
which was conjugated to hexacosanoic acid through an
amide bond. In this step, if the reaction time was elon-
gated to overnight, the double bond in 20 (as a mixture
of Z- and E-configurations) was also reduced (as in 21).
Finally, the benzyl (Bn) groups and the benzyloxycar-
bonyl (Cbz) group were removed smoothly over cata-
lytic hydrogenolysis to afford the final products 1－3.
The synthetic compounds 1－3 were tested for their
NKT cell-stimulation activity using KRN7000 as a pos-
itive control. It turned out that the analogues induced no
detectable cytokine or proliferative response.
α-GalCer are inserted into two hydrophobic pockets of
CD1d while the sugar head is extended above the li-
pid-binding groove surface for recognition by the TCR
of NKT cells.[32-34] The crystal structure of ternary hu-
man CD1d-αGalCer-TCR complex revealed that the
galactose ring is sandwiched between CD1d and NKT
TCR.[35] TCR displays a rigid “lock and key” type in-
teraction with the glycolipid. TCR interacts with not
only the 2', 3', and 4' hydroxyl groups of galactose ring
but also the 3-hydroxyl group of the sphingosine chain.
One reasonable explanation to our bioassay results is
that the elongated sphingosine chain in compounds 1－
3 could probably break the rigid spatial relationship
between the sugar head and the ceramide moiety, thus
interrupting the interactions of α-GalCer with NKT and
CD1d. Concurring with our speculation, Tashiro and
Franck et al. respectively discovered that the elongated
The X-ray crystallographic analysis of CD1d-α-
GalCer complex showed that the two lipid chains of
Chin. J. Chem. 2017, XX, 1—8
© 2017 SIOC, CAS, Shanghai, & WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim
www.cjc.wiley-vch.de
7

Zhang & Ye
FULL PAPER
Fukushima, H.; Natori, T.; Koezuka, Y. Bioorg. Med. Chem. 1997, 5,
1447.
[14] Uchimura, A.; Shimizu, T.; Morita, M.; Ueno, H.; Motoki, K.; Fu-
kushima, H.; Natori, T.; Koezuka, Y. Bioorg. Med. Chem. 1997, 5,
2245.
C- and O-glycoside analogues of KRN7000 with an
extra methylene unit (CH2) between the sugar and
ceramide showed no bioactivity at all.[5,36]
Conclusions
[15] Liu, Y.; Goff, R. D.; Zhou, D.; Mattner, J.; Sullivan, B. A.; Khurana,
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In summary, three novel KRN7000 analogues, in
which the ring oxygen of the galactopyranose residue is
replaced by the nitrogen atom along with the variation
on the ceramide, were designed and synthesized. These
synthetic compounds were evaluated for their ability to
stimulate cytokine release. Although the compounds
displayed no activity on inducing the cytokine release,
the results may benefit design of the next generation
KRN7000 analogues. Since iminosugars have shown
many applications in drug discovery, the disclosed
iminosugar-substitution strategy may facilitate the prep-
aration of more KRN7000 derivatives with biological
importance.
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Acknowledgement
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This work was financially supported by the National
Natural Science Foundation of China (No. 21232002).
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(Lu, Y.)
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