N-Oxyamide-linked glycoglycerolipid coated AuNPs for receptor-targeting imaging and drug delivery

Article abstract of DOI:10.1039/c5cc09749e

The synthesis of a series of new N-oxyamide-linked glycoglycerolipids and their assembly with gold nanoparticles for receptor-targeting imaging and drug delivery are reported.

Full text of DOI:10.1039/c5cc09749e

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DOI: 10.1039/C5CC09749E
Journal Name
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N-Oxyamide-linked glycoglycerolipid coated AuNP for
receptor-targeting imaging and drug delivery
Cite this: DOI: 10.1039/x0xx00000x
Na Chen,a Zhi-Hao Yu,b Dan Zhou,b Xi-Le Hu,b Yi Zang,c Xiao-Peng He,*b Jia Li*c
and Juan Xie*a
Received 00th January 2015,
Accepted 00th January 2015
DOI: 10.1039/x0xx00000x
www.rsc.org/
Synthesis of a series of new
N-oxyamide-linked glycoglycer-
olipids and their assembly with gold nanoparticles for the
receptor-targeting imaging and drug delivery are reported.
Glycolipids are known to play crucial roles in a variety of im-
portant biological processes such as cell-cell interactions, viral
and bacterial infections, activation and modulation of the im-
mune system, signal transduction and cell proliferation.1 Glyco-
lipids are composed of one or several monosaccharide residues
linked by a glycosidic bond to a hydrophobic moiety, such as
an acylglycerol (termed glycoglycerolipids – GGLs) or a
ceramide (termed glycosphingolipids – GSLs). While the de-
sign of glycolipid mimics has been a useful strategy in drug
discovery,2-4 we5 and others6-8 have also demonstrated the in-
teresting biological activities of synthetic glycolipids. Modifi-
cations have been made on the substitution and variation of the
glycosyl moiety, the configuration and nature of the anomeric
bond, the polar moieties of the ceramide or glycerol, and the
length and location of the lipid chains.
Fig. 1 Structure of the
and schematic illustration of the self-assembly of a SH-PEG@AuNP with 17 for
the receptor-targeting fluorophore/drug delivery.
N-oxyamide-linked glycoglycerolipids used for bio-tests
The galactoglycerolipids were prepared from
pentaacetate and ( )-1,2-di- -benzyl-glycerol according to
our previous strategy.11 The
-galactose pentaacetate was con-
verted to galactosyl bromide, which was then treated with ben-
D-galactose
S
O
1
D
zylated glycerol
trile to give the
1
catalysed by HgBr2 and Hg(CN)2 in acetoni-
-galactopyranoside in 75% yield (Scheme
Recently, we have been interested in the
N-oxyamide-
β
-
D
2
modified compounds because of their improved metabolic
stability than the amide-linked counterparts,9 and their interest-
ing secondary structures produced by intramolecular H-bonding
1). Hydrogenolysis followed by a selective silylation gave
alcohol . Then, a Mitsunobu reaction with PhthN-OH
smoothly produced the key -phthaloyl oxyamine in 97%
4
N
-oxyamide-linked peptides.10 We have developed a meth-
N
5
in
odology for the synthesis of
olipids, by replacing the ester group in GGLs by an
oxyamide, as new analogues of both GGL and GSL.11 With
continuing interest in
-oxyamide-modified biomolecules,12-20
yield. Subsequently, desilylation was carried out with catalytic
AcCl in MeOH (instead of TBAF which led to partial
deacetylation products). Then, a coupling with palmitic acid
N-oxyamide-linked glucoglycer-
N-
gave galactolipid
controlled hydrazinolysis (1.1 equiv. at 0 °C) to avoid
deacetylation. Acylation of oxyamine with different fatty
acids produced the -oxyamide-linked galactolipids 9-13 in
7, and the phthaloyl group was removed by a
N
we report here the synthesis of a new series of galactoglycer-
olipids and glucoglycerolipids. Considering their amphiphilic
properties and cellular targeting ability, the synthesized glyco-
glycerolipids have been used to self-assemble with a gold na-
noparticle (AuNP). The produced AuNPs coated with galacto-
glycerolipids have been proven effective for receptor-targeting
hepatocellular imaging and drug delivery (Fig. 1).
8
N
good yields. The NH signals appeared in the range of 8.81-8.87
ppm on the 1H NMR spactra in CDCl3 for these compounds.
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DOI: 10.1039/C5CC09749E
OAc
OAc
O
1. HBr, AcOH
CH Cl
AcO
AcO
ONH2
OAc
O
ONPhth
OTBS
AcO
AcO
OAc
O
AcO
AcO
O
AcO
AcO
NH2NH2 (1.1 eq.)
TBSCl, I
2
2
OR
2
O
O
OAc
O
2. 1, HgBr
N-methylimidazole
MeOH, 20 min, 0 °C
92%
2
OAc
OAc
Hg(CN) , MeCN
OTBS
2
OAc
90%
OR
OAc
22
23
2 R = Bn, 75%
3 R = H, 70%
10% Pd/C, H , EtOH
2
2) TBAF, THF
1) RCO2H, HOBt
EDC, Et3N
CH2Cl2
EtOAc, AcOH
AcO
AcO
AcO
AcO
OAc
O
OAc
O
O
O
ONPhth
PhthN-OH (3 eq.)
OH
OAc
OH
R
O
R
NH
O
O
NH
O
PPh (3 eq.), DIAD (3 eq.)
3
PhMe, 0 °C to RT
Na (0.3 eq.) AcO
AcO
O
HO
HO
O
O
OAc
O
OH
OH
OAc
OTBS
MeOH
OTBS
5
4
97%
OAc
OH
24 R= -(CH2)6CH3 56%
25 R= -(CH2)14CH3 47%
26 R= -(CH2)7CH=CH(CH 2)7CH3 49%
27 R= -(CH2)6CH3 89%
28 R= -(CH2)14CH3 100%
29 R= -(CH2)7CH=CH(CH 2)7CH3 99%
AcO
AcO
OAc
O
ONPhth
OR
AcCl (0.15 eq.)
MeOH
NH NH (1.1 eq.)
2
2
O
MeOH, 20 min, 0 °C
Scheme 2 Synthesis of glucoglycerolipids.
OAc
6
R = H, 77%
palmitic acid, EDC, DMAP
CH Cl , Et N, 0 °C to RT
7 R = CO(CH ) Me, 77%
2
2
3
2 14
With the N-oxyamide-linked glycoglycerolipids in hand, we tested
their ability to self-assemble with an SH-PEG coated gold nanoparti-
cle (AuNP),21 producing a new class of glycoglycerolipid-based
functional AuNPs. The glyconanoparticle formation was probably
because of the insertion of amphiphilic glycolipids to the SH-PEG
coated core-shell AuNP.21 Galactolipid 17 was used because of its
suitable lipid chain length and the targeting ability of galactose for
hepatocellular receptors.22-26 Shown in Fig. 2a are the scanning
electron microscopic images of the AuNP and its ensemble with 17
(CNAu). Both AuNP and CNAu were observed to be well dispersed,
whereas the addition of a galactose-selective peanut agglutinin (PNA)
caused aggregation of CNAu. This is in agreement with previous
reports on the aggregation of glyco-AuNPs caused by sugar-lectin
recognition.27-30 CNAu was determined to have good stability by a
long-term incubation in a serum system (Fig. S1).
O
NH
AcO
AcO
AcO
2
OAc
OAc
O
RCO H
R
NH
O
2
O
.
EDC HCl, HOBt
O
O
O
O
AcO
Et N, CH Cl
3
2
2
14
OAc
O
OAc
O
14
9 R= -(CH ) CH 88%
10 R= -(CH ) CH 81%
2 4
3
8
O
2 6
3
11 R= -(Et)CH(CH ) CH 78%
2 3
3
NH NH (12 eq.)
2
2
12 R= -(CH ) CH 90%
2 14 3
85% EtOH, 50 °C
O
13 R= -(CH ) CH=CH(CH ) CH 92%
2 7
2 7
3
HO
OH
NH
Na (0.3 eq.)
MeOH, RT
R
O
O
for 9 - 10
O
O
HO
14
O
OH
R
O
14
15
16
R= -(CH ) CH 47%
R= -(CH ) CH 50%
OH
OH
O
2 4
3
N
O
2 6
3
14
R= -(Et)CH(CH ) CH 52%
O
2 3
3
O
HO
17 R= -(CH ) CH 58%
18 R= -(CH ) CH=CH(CH ) CH 55%
2 14
3
OH
2 7
2 7
3
OH
19 R= -(CH ) CH 96%
2 4
3
20 R= -(CH ) CH 99%
2 6
3
OH
H
OH
N
Na (0.3 eq.)
O
14
O
12
O
O
MeOH, RT
68%
HO
OH
OH
21
Scheme 1 Synthesis of galactoglycerolipids.
To obtain the deprotected galactoglycerolipids, we have tried dif-
ferent deacetylation conditions. The acetyl groups can be removed
with 12 equivalents of hydrazine at 50 °C in EtOH, leading to the
desired galactolipids 14-18. Surprisingly, deacetylation of com-
pounds 9 and 10 under Zemplan condition (Na/MeOH) gave the
intramolecular transacylation products 19 and 20 in excellent yields,
but this condition cleaved all the ester bonds of compound 12, af-
fording galactolipid 21.
Fig. 2 (a) Scanning electron microscopic (SEM) images of AuNP, CNAu and
CNAu with PNA (0.3 µM) (Scale bar: 50 µm). UV-vis absorption spectra of
CNAu in the presence of (b) increasing PNA (0-0.3 µM) and (c) PNA or other
proteins including the mannose-selective lens culinaris lectin, GalNAc-selective
soybean agglutinin, GlcNAc-selective wheat germ agglutinin, bovine serum
albumin and pepsin (0.05 µM each). (d) Dynamic light scattering of (1) AuNP, (2)
CNAu and (3) CNAu with PNA (0.3 µM).
Using the similar protocol, we also prepared glucoglycerolipids
with a single lipid chain by hydrazinolysis of a previously pre-
pared
pling with fatty acids followed by removal of TBS and Zem-
O . A subsequent cou-
-phthalimido glucoglycerol 22 11
plan deacetylation afforded the glucoglycerolipids 27-29
(Scheme 2).
The aggregation was also corroborated by dynamic light scattering;
the size of CNAu drastically increased in the presence of PNA (Fig.
2d and Fig. S2). A large difference in Zeta potential was observed
between AuNP and CNAu (Fig. S3), suggesting the presence of 17
on the gold nanoparticles. The loading concentration of galactoglyc-
erolipid 17 was determined to be 21.6 µg/mg AuNP by an an-
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throne/sulfuric acid method (Fig. S4). UV-vis spectroscopy suggests used to load CNAu for the fluorescence imaging of Hep-G2 (human
that the absorption band of CNAu gradually red-shifted (which is a liver cancer cell line) using HeLa (human cervix cancer cell line)
signature of AuNP aggregation)27-30 in the presence of increasing and A549 (human lung cancer cell line) as control. We observed that,
PNA (Fig. 2b), but not other unselective proteins (Fig. 2c). These while DCM alone (Fig. 3a and 3b) and DCM@AuNP (i.e. DCM
data suggest the biospecificity of the CNAu. To test the generality of loaded with bare SH-PEG coated AuNP, Fig. S6) produced a similar
the glyco-AuNP formation with N-oxyamide-linked glycoglycer- level of fluorescence after incubation with different cells, association
olipids, the mono-lipid galactoglycerolipid 21, mono-lipid gluco- of DCM with CNAu (DCM@CNAu) largely enhanced the fluores-
glycerolipid 28 and a previously synthesized di-lipid glucoglycer- cence imaging of the resulting material for Hep-G2 (Fig. 3a and 3c).
olipid 3011 (Fig. 1) were used. Likewise, we observed that the ab- This result is in agreement with the ASGPr expression level of the
sorption band of the formed glyco-AuNPs red-shifted with a selec- cells as determined by real-time quantitative polymerase chain reac-
tive lectin in a concentration-dependent manner (Fig. S5). The limit tion (Fig. 3d). Then, we used an anticancer drug, hydrocamptothecin
of detection of the di-lipid AuNPs was found to be lower than the (HCPT), to test the receptor-targeting drug delivery ability of CNAu.
mono-lipid counterparts (Fig. S1).
Interestingly, while a short-term (15 min) incubation of HCPT (Fig.
3e) or CNAu (Fig. 3g) alone did not induce cell death, the associa-
tion of the drug with glyconanoparticle led to an evident suppression
of the cell viability of Hep-G2, but not that of HeLa and A549 (Fig.
3f). We also determined that the receptor-targeting HCPT delivery
was both concentration- and time-dependent (Fig. S7). These data
suggest the promise of galactoglycerolipid-coated AuNPs for target-
specific theranostics.
In summary, we have synthesized a series of N-oxyamide-linked
glycoglycerolipids, which could be used to form a new type of gly-
co-AuNP capable of receptor-targeting cell imaging and drug deliv-
ery. This study provides an insight into the construction of glyco-
materials for target-specific disease theranostics.32
This research is supported by the 973 project (2013CB733700),
the National Natural Science Foundation of China (21572058,
21576088), the Key Project of Shanghai Science and Technology
Commission (13NM1400900) and the Fundamental Research Funds
for Central Universities (222201414010). N. Chen gratefully
acknowledges China Scholarship Council (CSC) for a doctoral
scholarship.
Notes and references
a
PPSM, ENS Cachan, CNRS, Université ParisꢀSaclay, Cachan, 94235
France, Email: joanne.xie@ensꢀcachan.fr (J. Xie)
b Key Laboratory for Advanced Materials & Institute of Fine Chemicals,
East China University of Science and Technology, 130 Meilong Rd.,
Shanghai 200237, PR China, Email: xphe@ecust.edu.cn (X.ꢀP. He)
c National Center for Drug Screening, State Key Laboratory of Drug
Research, Shanghai Institute of Materia Medica (SIMM), Chinese Acadꢀ
emy of Sciences (CAS),189 Guo Shoujing Rd., Shanghai 201203, PR
China, jli@simm.ac.cn (J. Li)
Electronic Supplementary Information (ESI) available: [Experimental
section, additional figures and original spectral copies]. See
DOI: 10.1039/c000000x/
Fig. 3 (a) Fluorescence imaging of Hep-G2 (human liver cancer), HeLa (human
cervix cancer) and A549 (human lung cancer) cells with DCM (1 µM) and
DCM@CNAu (1/0.1 µM) (Scale bar: 100 µm; cell nucleus were stained by
Hoechst 33342). Fluorescence quantification of different cells treated with (b)
DCM and (c) DCM@CNAu. (d) Relative mRNA level of ASGPr of different
1
2
3
K. Brandenburg and O. Holst. 2015, Glycolipids: Distribution and
Biological Function. eLS. 1.
J. L. Daniotti, A. A. Vilcaes, V. T. Demichelis, F. M. Ruggiero and
cells determined by real-time quantitative polymerase chain reaction (***
P <
0.001). Normalized viability of different cells treated with (e) HCPT alone (1
µM), (f) HCPT@CNAu (1/0.1 µM) and (g) CNAu alone (0.1 µM).
M. Rodriguez-Walker, Front. Oncolo., 2013,
3, 306.
T. Wennekes, R. J. van den Berg, R. G. Boot, G. A. van der Marel,
H. S. Overkleeft and J. M. Aerts, Angew. Chem. Int. Ed., 2009, 48
8848.
K. D. McReynolds and J. Gervay-Hague, Chem. Rev., 2007, 107,
1533.
,
Next, we tested the ability of CNAu for hepatocellular imaging
and drug delivery considering the selective recognition between
galactose and the asialoglycoprotein receptor (ASGPr) on hepato-
cytes.21-25 A red-emitting dicyanomethylene (DCM, Fig. S6)31 was
4
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5
(
a
) X.-P. He, J. Xie, Y. Tang, J. Li and G.-R. Chen, Curr. Med. 20 N. Chen and J. Xie, Org. Biomol. Chem.
Chem., 2012, 19, 2399; ( ) X.-L. He, D. Li, L. Shao, X. Dong, X.-P. 10.1039/C5OB02328A.
He, G.-R. Chen and D. Chen, ACS Med. Chem. Lett., 2015, , 793; 21 X.-L. Hu, H.-Y. Jin, X.-P. He, T. D. James, G.-R. Chen and Y.-T.
) H.-L. Zhang, X.-P. He, L. Sheng, Y. Yao, W. Zhang, X.-X. Shi, J. Long, ACS Appl. Mater. Interfaces, 2015, , 1874.
Li and G.-R. Chen, Mol. Divers., 2011, 15, 889; ( ) S.-S. Deng, C. 22 H.-L. Zhang, X.-L. Wei, Y. Zang, J.-Y. Cao, S. Liu, X.-P. He, Q.
Zhang, H. Wang, Y. Zang, J. Li, X.-P. He and G.-R. Chen, Carboꢀ
hydr. Res., 2015, 408, 114; ( ) S.-X. Song, M.-L. Wu, X.-P. He, Y.-B.
Zhou, L. Sheng, J. Li and G.-R. Chen, Bioorg. Med. Chem. Lett.
2012, 22, 2030; ( ) S.-X. Song, H.-L. Zhang, C.-G. Kim, L. Sheng,
, 2016, DOI:
b
6
(c
7
d
Chen, Y.-T. Long, J. Li, G.-R. Chen and K. Chen, Adv. Mater., 2013,
25, 4097.
23 W. Ma, H.-T. Liu, X.-P. He, Y. Zang, J. Li, G.-R. Chen, H. Tian and
Y.-T. Long, Anal. Chem., 2014, 86, 5502.
e
,
f
X.-P. He, Y.-T. Long, J. Li and G.-R. Chen, Tetrahedron, 2010, 66
9974.
,
24 K.-B. Li, Y. Zang, H. Wang, J. Li, G.-R. Chen, T. D. James, X.-P. He
and H. Tian, Chem. Commun., 2014, 50, 11735.
6
7
(
a
) M. Vetro, B. Costa, G. Donvito, N. Arrighetti, L. Cipolla, P. 25 D.-T. Shi, D. Zhou, Y. Zang, J. Li, G.-R. Chen, T. D. James, X.-P.
Perego, F. Compostella, F. Ronchetti and D. Colombo, Org. Biomol. He and H. Tian, Chem. Commun., 2015, 51, 3653.
Chem., 2015, 13, 1091; ( ) J. Bi, J. Wang, K. Zhou, Y. Wang, M. 26 L. Dong, Y. Zang, D. Zhou, X.-P. He, G.-R. Chen, T. D. James and J.
Fang and Y. Du, ACS Med. Chem. Lett., 2015, , 476. Li, Chem. Commun., 2015, 51, 11852.
S. Aspeslagh, Y. Li, E. D. Yu, N. Pauwels, M. Trappeniers, E. Gir- 27 For recent reviews, see: ( ) M. J. Marín, C. L. Schofield, R. A. Field
and D. A. Russell, Analyst, 2015, 140, 59; ( ) Y. Zhang, Y. Guo, Y.
b
6
a
ardi, T. Decruy, K. van Beneden, K. Venken, M. Drennan, L. Ley-
baert, J. Wang, R. W. Franck, S. van Calenbergh, D. M. Zajonc and
D. Elewaut, EMBO J., 2011, 30, 2294.
b
Xianyu, W. Chen, Y. Zhao and X. Jiang, Adv. Mater., 2013, 25
,
3802.
8
9
A. Banchet-Cadeddu, E. Henon, M. Dauchez, J. H. Renault, F. Mon- 28 M. Marín, A. Rashid, M. Rejzek, S. A. Fairhurst, S. A. Wharton, S.
neaux and A. Haudrechy, Org. Biomol. Chem., 2011,
F. Chen, B. Ma, Z.-C. Yang, G. Lin and D. Yang, Amino Acids, 2012,
43, 499.
9
, 3080
R. Martin, J. W. McCauley, T. Wileman, R. A. Field and D. A. Rus-
sell, Org. Biomol. Chem., 2013, 11, 7101.
29 J. Wei, L. Zheng, X. Lv, Y. Bi, W. Chen, W. Zhang, Y. Shi, L. Zhao,
X. Sun, F. Wang, S. Cheng, J. Yan, W. Liu, X. Jiang, G. F. Gao and
10 X. Li, Y.-D. Wu and D. Yang, Acc. Chem. Res., 2008, 41, 1428.
11 N. Chen and J. Xie, J. Org. Chem., 2014, 79, 10716.
X. Li, ACS Nano, 2014, 8, 4600.
12 A. Malapelle, R. Ramozzi and J. Xie, Synthesis, 2009, 888.
13 Y. C. Gong, H. B. Sun and J. Xie, Eur. J. Org. Chem., 2009, 6027.
14 Y. Gong, S. Peyrat, H. Sun and J. Xie, Tetrahedron, 2011, 67, 7114.
15 Z. Song, X.-P. He, G.-R. Chen and J. Xie, Synthesis, 2011, 2761.
16 S. Peyrat and J. Xie, Synthesis, 2012, 1718.
17 O. Noel and J. Xie, Synthesis, 2013, 134.
18 S. Peyrat, K. Cheng and J. Xie, Synthesis, 2013, 2737.
19 H.-L. Zhang, Y. Zang, J. Xie, G.-R. Chen, X.-P. He and H. Tian, Sci.
30 S. Gatard, L. Salmon, C. Deraedt, J. Ruiz, D. Astruc and S. Bouquil-
lon, Eur. J. Inorg. Chem., 2014, 2671-2677.
31 Y. Cui, J. Yu, J. Gao, Z. Wang and G. Qian, J. SolꢀGel Sci. Technol.
,
2009, 52, 362.
32 (
Rev., 2015, 44, 4239 ; (
Chen, J. Mater. Chem. B, 2015,
Zang, W. Liu, X. Zhang, J. Li, G.-R. Chen, T. D. James and X.-P. He,
J. Mater. Chem. B, 2015, , 9182.
a
) X.-P. He, Y. Zang, T. D. James, J. Li and G.-R. Chen, Chem. Soc.
) D.-K. Ji, Y. Zhang, X.-P. He and G.-R.
, 6656; ( ) D.-K. Ji, Y. Zhang, Y.
b
3
c
Rep., 2014,
4
, 5513.
3
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