2-Aminoimidazole facilitates efficient gene delivery in a low molecular weight poly(amidoamine) dendrimer

Article abstract of DOI:10.1039/c8ob00953h

Functional groups have shown great potential in gene delivery. However, a number of the reported functional groups can only overcome one certain physiological barrier, resulting in limited transfection efficiencies. Based on the structure-activity relationships of both imidazolyl and guanidyl, we designed a novel multifunctional group, 2-aminoimidazole (AM), for gene delivery. On modifying with the AM group, the transfection efficiency of low molecular weight poly(amidoamine) (G2) was 200 times greater than the parent dendrimer in vitro. In contrast, the transfection efficiency of G2 showed a decreasing trend when it was grafted with imidazole. Assays revealed that the AM group played multiple roles in gene delivery, including condensing DNA into monodisperse nanoparticles of 80-90 nm in diameter, achieving nearly ten times higher cellular-uptake efficacy, and enhancing the abilities of endosome/lysosome escape and nuclear localization. What's more, AM showed low toxicity. These results demonstrate that the AM group could be a promising tool in non-viral gene delivery.

Full text of DOI:10.1039/c8ob00953h

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2-Aminoimidazole facilitates efficient gene delivery in low
molecular weight poly(amidoamine) dendrimer
Jing Wang,a Xuefeng Hu,a Dongli Wang,a Cao Xie,a Weiyue Lu,a Jie Song,a Ruifeng Wang,a Chunli
Gao,b and Min Liu*a
Received 00th January 20xx,
Accepted 00th January 20xx
DOI: 10.1039/x0xx00000x
Functional groups have shown great potential in gene delivery. However a number of the reported functional groups can
only overcome one certain physiological barrier, resulting in limited transfection efficiencies. Based on the structure-
activity relationships of both imidazolyl and guanidyl, we designed a novel multifunctional group, 2-aminoimidazole (AM),
for gene delivery. Modifying with AM group, the transfection efficiency of low molecular weight poly(amidoamine) (G2)
was 200 times greater than the parent dendrimer in vitro. In contrast , the transfection efficiency of G2 showed a trend of
decrease when it grafted with imidazole. Assays revealed that AM group played multiple roles in gene delivery, including
condensing DNA into monodisperse nanoparticles of 80～90 nm in diameter, achieving nearly ten times higher cellular-
uptake efficacy, enhancing abilities of endosome/lysosome escape and nuclear localization. What’s more, AM showed low
www.rsc.org/
toxicity. These results demonstrate that AM group could be
a promising tool in non-viral gene delivery.
functional groups listed, we are particularly interested in
imidazolyl and guanidyl. Derived from histidine, imidazolyl can
facilitate the endosomal escape of DNA complexes via a
“proton sponge” mechanism, due to its high buffering capacity
at endosomal pH. Guanidyl, from arginine, can be beneficial
not only to DNA condensation, but also membrane
transportation, due to forming bidentate hydrogen bonds with
phosphate groups both in the DNA and the cell membrane.21
Based on the structure-activity relationships of both imidazolyl
and guanidyl, we predicted that integrating imidazolyl and
guanidyl into one functional group could produce a cumulative
effect. Therefore, we designed a novel multifunctional group,
2-aminoimidazole (AM), for DNA delivery, which contains an
amino group at position 2 of the imidazole ring. It was
reported that the pKa of AM was 8.46, which was about 1.5
units higher than imidazole (7.01) and 5 units lower than
guanidine (13.4).22,23 We speculated that AM group would
increase DNA condensation, cytomembrane penetration,
endosome/lysosome escape and nuclear localization
concurrently. To verify this hypothesis, we grafted PAMAM
generation 2 (G2) with AM to develop a cationic polymer (G2-
AM) as a non-viral vector. G2 was chosen as the basis for
modification with AM for the following reasons: first, G2 has a
well-defined number of amine groups on the surface, and is
easy to be chemically modified; second, G2 is relatively well
tolerated; third, G2 has a poor transfection efficiency due to its
low density of primary and tertiary amine groups, making it
easier to assess improvements in performance resulting from
AM modification. Interestingly, AM modification significantly
enhanced the transfection efficiency of G2 without introducing
additional toxicity. G2-AM was found to have a transfection
efficiency ~200 times greater than that of G2 alone in vitro,
Introduction
Non-viral vectors have attracted increasing attention due to
their low immunogenicity, flexible DNA loading capacity and
tailorable properties.1-5 However, their applications are limited
by low transfection efficiency.6,7 Non-viral vectors need to
overcome the following physiological barriers to achieve DNA
delivery: (1) potential degradation by endonucleases and
phagocytosis by the reticuloendothelial system in the blood,
(2) cellular internalization across the amphipathic
cytomembrane with negatively charged regions, (3)
degradation by lysosomal enzymes and transportation into the
nucleus.8-10 Each of these physiological barriers has the
potential to significantly reduce the transfection efficiency of
non-viral DNA delivery.
These barriers might be overcome owing to the
developments in material sciences, which have yielded new
functional groups as gene delivery enhancer, such as
hydrophobic molecules, targeting molecules, cell penetrating
peptides, guanidyl, imidazolyl, fusion proteins and nuclear
11-20
localization signal peptides.
to enhance DNA
These functional groups aim
cellular uptake,
condensation,
endosome/lysosome escape and nuclear localization. Of the
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and was more efficient than Lipofectamine 2000. Imidazolyl- packaging, buffering capacity, cell uptake and nuclear
modified G2 (G2-M) was also synthesized as a control, and localization. The data showed that AM could contribute to
showed no significant improvement compared with the parent overcoming the multiple physiological barriers that non-viral
dendrimer. Finally, we investigated the transfection vectors encounter. Our results demonstrate that the AM group
mechanism of G2-AM as a gene vector, including DNA could have potential applications in non-viral gene delivery.
Scheme 1. Effects of AM in efficient gene delivery.
Results and discussion
Design and Synthesis of the materials.
We synthesized G2-AM through the reaction of G2 and tert-butyl
(4-formyl-1H-imidazol-2-yl) carbamate. The modification ratio of
1
AM was verified by H NMR spectrum (Figure S4-S7). The resulting
materials were termed G2-AM-2, G2-AM-5, G2-AM-8 and G2-AM-
11 respectively. As controls, a series of G2 modified with M were
obtained (Figure S8-S10), which termed G2-M-3, G2-M-7 and G2-M-
11 respectively. Transfection efficiency of above dendrimers were
Fig. 1 Synthesis of compound 4 (a), G2-AM-11 (b) and G2-M-11 (c)
evaluated in HEK293T cells using an enhanced green fluorescent
protein (EGFP) plasmid. As shown in Figure S11, G2-AM-11
performed best in HEK293T cells. Therefore, G2-AM-11 was
investigated in detail in further studies.
AM modification condenses DNA into smaller nanoparticles.
Nanoparticle size is a key parameter affecting the cellular
uptake efficiency. In general, particles in the 40–200 nm range
24
exhibit cellular uptake in vitro. Fig. 2a and 2b show that the
G2-AM-11/pGL3
complex
assembled
into
spherical
nanostructures of 80–90 nm in diameter in water at the mass
ratio of 12. The control complexes, G2-M-11/pGL3 and
G2/pGL3, assembled into irregular shaped nanostructures of
400–1500 nm in diameter, which appeared aggregated. Gel
retardation assays were performed to verify whether G2-AM-
11 could condense DNA, with G2 and G2-M-11 used as
controls. The mass ratios of G2-AM-11 and G2 for complete
retardation were both ~4, whereas that of G2-M-11 was ~8
(Fig. 2c). Since G2-AM-11 could condense DNA into smaller
nanostructures than G2 and G2-M-11, we speculate that there
might be other interactions such as hydrogen bonding
between G2-AM-11 and DNA.
Fig. 2 Formation and characterization of the complexes prepared from G2,
G2-M-11, G2-AM-11 and free DNA. (a) Size and zeta potential of the
nanoparticles (n=3). (b) TEM images of the complexes, with fixed weight
ratio of polymer/pGL3 of 12:1. The scale bar in the top right image is 100
nm, all others are 200 nm. (c) Gel retardation assay of three complexes,
using free pGL3 as a negative control (far left), the number shows the mass
ratio of polymer/pGL3.
AM modification greatly improved transfection efficiency in vitro
and in vivo.
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Due to its high sensitivity, the luciferase assay was conducted
to quantitatively evaluate the transfection efficiency. Fig. 3a
showed that the transfection efficiency of G2-AM-11/pGL3
was ~200 times greater than that of G2/pGL3 and ~10000
times greater than that of G2-M-11/pGL3 at a mass ratio of
12:1. What’s more, it even performed better than
Lipofectamine 2000. The EGFP assay (Fig. 3a) showed similar
results. We also investigated the relationship between
modification ratio and transfection efficiency. The EGFP and
Luciferase assays (Fig. S11) showed that the transfection
efficiency increased with the modification ratios of AM, which
revealed that AM played an important role in gene delivery. All
of the complexes prepared from G2-M showed low
transfection efficiency.
Additionally we investigated whether G2-AM-11 could
increase transfection efficiency in HeLa cells. Flow cytometry
assay showed that G2-AM-11 had higher transfection
efficiency in HeLa cells and performed better than the positive
control (Fig. S12). The findings described confirmed that AM
modification could improve the transfection efficiency of G2.
It is well understood that cationic polymers with high
molecular weight show high transfection efficiency
25
accompanied with high cytotoxicity. To investigate whether
Fig. 4 Mechanism Assays. (a) Buffering Index (β) of G2, G2-M-11 and G2-
AM-11. (b) Cellular uptake efficiency of the complexes prepared using G2,
G2-M-11 and G2-AM-11 in HEK 293T cells. pGL3 plasmid was stained with
TOTO-3 and the result is shown as the mean fluorescence of DNA. (c)
Cellular uptake images in HEK 293T cells. Cell nuclei were stained with DAPI
and pGL3 was stained with TOTO-3. The scale bar is 5 μm.
AM modification could lead to G2 cytotoxicity, we conducted
MTT assay to evaluate the cytotoxicity of the polymers and
complexes in HEK 293T cells. As shown in Fig. 2b, all complexes
prepared from G2, G2-M-11 and G2-AM-11 showed no
significant cytotoxicity, with cell viabilities of more than 90%,
even when the mass ratio was as great as 16:1. In contrast, the
viability of the cells that were co-incubated with the
complexes prepared from Lipofectamine was lower than 80%,
and decreased to nearly 30% at higher concentration. MTT
assay of the polymers was conducted at the concentrations of
0.5 mg/mL, 1 mg/mL and 2 mg/mL (Fig. S14). The results
showed that all G2 derivatives had good compatibility with
HEK 293T and HeLa cells.
Mechanism Studies.
When it comes to the design of non-viral vectors,
overcoming only selected obstacles to delivery will not lead to
systems capable of improving transfection efficiencies. Due to
sophisticated physiological barriers, it appears particularly
important to coordinate various functional groups to address
more of the challenges presented by the body. Derived from
histidine and arginine, imidazolyl and guanidyl have been
widely investigated for gene delivery. Joon et al developed
imidazolyl and guanidyl grafted G4, and the resulting
26a,b
dendrimers showed significant gene delivery potency.
contrast to that work,
In
we integrated imidazolyl and guanidyl into one functional
group, AM, in line with atom economy. Interestingly, a
significant distinction between the transfection efficiency of
G2-M-11 and G2-AM-11 was observed. To probe the
transfection mechanism, we studied the buffering capability
and cell uptake of G2-AM-11.
High buffering capacity of non-viral vectors is widely
considered as a prerequisite for efficient endosome/lysosome
escape. We therefore investigated the buffering capacity of
the polymers using acid-base titration.27-29 To make the
results clearer, we adopted buffering index as an indicator of
buffering capacity. We compared the buffering index of G2,
G2-M-11 and G2-AM-11 in the pH range 2–10. As shown in Fig.
4a, the buffering index of G2 was poorest in the pH range 4–8.
This is possibly due to the low density of tertiary amine groups
on G2. G2-M-11 and G2-AM-11 performed well across the pH
Fig. 3 In vitro transfection and cytotoxicity studies in HEK 293T cells. (a)
Luciferase activities, mass ratios were showed in figure legends, using
lipofectamine 2000 as positive controls at the recommended
concentration. (b) Cytotoxicity of the complexes, mass ratios were showed
in figure legends, compared with lipofectamine 2000. (c) EGFP images,
mass ratios were fixed at 12:1.
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range. G2-M-11 possessed a higher buffering index in the pH United States). TOTO-3 fluorescence dye was bought from Life
range 5.8–7.0. G2-AM-11 performed better in the pH range Technologies (Karlsruhe, Germany). Gelred was purchased
4.2–5.8, with a highest buffering index at pH ~5.2, which was 3 from Bitoium (Fremont, United States). The Luciferase Assay
times higher than that of G2. The pH values most frequently Kit and the luciferase reporter gene plasmid (pGL3) were
found in early endosome, late endosome and lysosome products of Promega (Beijing, China). The BCA Protein Assay
compartments are 6.2–6.3, 5.0–5.5 and 4.5–5.0, respectively. Kit were purchased from Beyotime (Shanghai, China). The
30-32
Therefore, we speculate that G2-AM-11 might facilitate plasmid encoding EGFP (pEGFP-N2) was from Genechem Co.
escape of DNA from both the late endosome and lysosome.
After incubation for 4 h, strong TOTO-3 labeled red
(Shanghai, China).
Human Embryonic Kidney 293T cells (HEK293T cells) was
fluorescence was present in the cells incubated with G2-AM- obtained from Shanghai Institute of Cell Biology. Human cervix
11/pGL3 (Fig. 4c), and it could be observed that the uptake carcinoma cell lines (HeLa cells) were obtained from the
efficiency of G2-AM-11 was nearly 10 times as great as for G2 American Type Culture Collection (ATCC). All of the cells lines
(Fig. 4b). In addition, there was clear co-localization of TOTO-3 were maintained in Dulbecco's Modified Eagle Medium (Gibco)
labeled pGL3 and the nucleus in the cells incubated with the supplemented with 10% FBS (Gibco), 100 μg/mL penicillin, and
G2-AM-11/pGL3 complex, which suggests AM modification 100 μg/mL streptomycin at 37°C under
improved nucleus localization. Most polymers can successfully atmosphere containing 5% CO2.
pass though the cell membrane but fail to reach the nucleus Synthesis of G2-AM and G2-M.
due to the presence of the lysosome and karyotheca. AM
a humidified
To obtain G2-AM, tert-butyl (4-formyl-1H-imidazol-2-yl)
carbamate was synthesized by the synthetic route in Figure 1.
The detailed steps were as follows.
modification could overcome these two barriers and ultimately
achieve a robust transfection efficiency.
The only difference between M and AM is the additional
amino group in the molecular structure of AM. However, M
conjugated to G2 had a negative effect on the transfection
efficiency. Despite the lower pKa of AM leads to some debate
concerning the ability of it to directly mimic a guanidine, 22,33
our results verified AM made some effects similar to guanidyl.
The gel retardation assay showed that the minimum mass ratio
that could completely condense DNA was larger for G2-M-11
than for G2 and G2-AM-11. TEM images also showed that G2-
M-11/pDNA had large particle sizes and irregular shapes, and
G2-AM-11 could condense DNA into monodisperse
Synthesis of 3-bromo-1,
Methylglyoxal 1,1-dimethyl acetal (20 g, 0.169 mol) was
dissolved in 400 mL CCl4, then the solution was cooled to 0
1-dimethoxypropan-2-one.
℃
and Br2 (32.45 g, 0.202 mol) was added dropwise into the
reaction solution. Subsequently, the reaction solution was
stirred at room-temperature for 12 h under nitrogen
atmosphere. Then, the solution was cooled to 0
℃ and 100 mL
sodium bicarbonate saturated solution was dropped into the
system. The organic phase was separated by separating funnel
and washed with saturated salt solution for 3 times
(3×100mL), dried with anhydrous sodium sulfate. Compound 1
(37.26 g) was obtained by evaporating the organic solvent and
directly used for next step without further purification.
nanoparticles of 80～90 nm in diameter. We speculate that
the hydrogen bonding between G2-AM-11 and DNA might
result in these differences. Moreover, there are many
researches on the enhanced effect of combining hydrogen
bonding and cationic elements in gene delivery systems.34,35
Structure-activity studies showed that the optimal functional
groups depends on the parent polymer,34 which indicated that
G2 might not be the best polymer to be modified with AM. We
will further investigate more non viral vectors.
Synthesis of tert-butyl 2-amino-4-(dimethoxymethyl)-1H-
imidazole-1-carboxylate. Boc-guanidine (10 g, 63 mmol) and
compound 1 (18.5 g, 76 mmol) were dissolved in 200 mL
tetrahydrofuran, and isopropyl titanate (5.96 g, 21 mmol) was
added into the reaction solution. The reaction solution was
stirred at room-temperature for 48 h. Subsequently, the crude
product was obtained by filtration and evaporating the organic
solvent. The crude product was purified by column
chromatography
(eluent:
Experimental
CH2Cl2/MeOH/Triethylamine=3/1/0.5) to obtain compound 2
(13.68 g) as a white solid in 84.5% yield. The structure of
compound 2 was ascertained by 1H NMR spectrum (Varian 400
MHz, Palo Alto, USA). (Figure S1)
Materials.
Sodium borohydride and boc-guanidine were bought from
Sinopharm Chemical Reagent Co. Ltd (Shanghai, China).
Imidazole-4-carboxaldehyde and methylglyoxal 1, 1-dimethyl
acetal were purchased from J&K Scientific Ltd (Beijing, China).
Poly (amidoamine) generation 2 (G2, MW = 3256, 16 NH2
groups per molecule) was bought from Weihai CY Dendrimer
Technology Co. Ltd. (Shandong, China). High molecular weight
polyethyleneimine (PEI, branched, MW= 25000) was obtained
from Sigma-Aldrich Co. (St. Louis, United States). 4’, 6-
Diamidino-2-phenylindole (DAPI) was purchased from Dalian
Meilun Biology Technology Co. Ltd. (Dalian, China).
Lipfectamine 2000 was purchased from Invitrogen (Carlsbad,
Synthesis of tert-butyl 2-((tert-butoxycarbonyl) amino)-4-
(dimethoxymethyl)-1H-imidazole-1-carboxylate. Compound 2
(5.0 g, 19.45 mmol) was dissolved in 50mL tetrahydrofuran
under nitrogen atmosphere. Then N, N-diisopropylethylamine
(3.01 g, 23.34 mmol) and di-tert-butyl pyrocarbonate (5.08 g,
23.34 mmol) were added into the flask subsequently and they
were stirred at room temperature for 12 h. Then the organic
solvent was evaporated under reduced pressure and the
mixture was re-dissolved with CH2Cl2, washed with saturated
salt solution and dried with anhydrous sodium sulfate. The
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crude product was purified by column chromatography replaced with 0.5 mL serum-free medium and 0.05 mL
(eluent: CH2Cl2/CH3COOCH2CH3=5/1) to get compound 3 as a complex, and each well included 2 μg pEGFP-N2. The medium
white solid (6.42 g) in 92.1% yield. The structure of compound was changed to culture medium containing 10% serum after
3 was ascertained by 1H NMR spectrum. (Figure S2)
Synthesis of tert-butyl (4-formyl-1H-imidazol-2-yl)
carbamate. Compound 3 (2.0 g, 5.6 mmol) was dissolved in determined with an inverted fluorescence microscope (Leica,
20mL CH2Cl2 and the reaction was cooled to 0 . Then pyridine DMI4000B, Germany) and quantitatively measured by flow
incubation at 37
℃ for 4 h. Cells were further incubated for 44
h
before determination. Transfection efficiency was
℃
hydrochloride was added into the reaction solution and the cytometry.
mixture was stirred at room-temperature overnight. The Luciferase Assay.
reaction process was detected by TLC. Following the
Cells were seeded in a 48-well plate at a density of 2×104 /well
completion of the reaction, 10mL water was poured into the
solution and the mixture was washed with saturated salt
solution for 3 times (3×10mL). The organic phase was dried
with anhydrous sodium sulfate. The crude product was
purified by column chromatography (eluent: CH2Cl2/MeOH
=10/1) to get compound 4 as a white solid (0.99 g) in 84.0%
yield. The structure of compound 4 was ascertained by 1H NMR
spectrum. (Figure S3)
and incubated 24 h before transfection, then the medium was
replaced with 0.5 mL serum-free medium and 0.05 mL
complex, and each well included 2 μg pGL3. The pGL3
encoding luciferase was used as the reporter gene in this
assay. The medium was changed to culture medium containing
10% serum after incubation at 37
℃ for 4 h. Cells were further
incubated for 44 h before determination. The transfection
efficiency was determined according to the manufacturer’s
protocols (Promega). Luciferase activity was determined by an
Ultra-Weak Luminescence Analyzer (Chuanghe, China). And
the total protein concentration of transfected cell lysate was
Synthesis of G2-AM and G2-M. G2 and compound 4 were
dissolved in methanol at different molar ratios and stirred for
24 h under nitrogen atmosphere. Then equimolar amount
(compared with compound 4) of sodium borohydride was
added and the mixture was stirred at room-temperature
overnight. The solvent was evaporated under reduced
pressure to get a yellow solid. Then the solid was dissolved in
CH2Cl2 which contained 20% trifluoroacetic acid and the
solution was stirred at room-temperature for 8 h. The mixture
was purified by dialysis (MWCO 500-1000 Da) against distilled
water which contained a little ammonium hydroxide. The
solution was then freeze-dried and their modification ratios
measured by
a BCA protein assay kit following the
manufacturer’s instructions. Data were expressed as RLU/mg
protein (relative light units per milligram cellular protein).
Cytotoxicity assay.
The cytotoxicity of complex and polymer were determined by
MTT assay. Cells were seeded in a 96-well plate at a density of
3×103 /well and incubated for 24 h, then the media was
replaced with 0.2 mL serum-free medium and 0.02 mL
complex, and each well included 0.8 μg pGL3. The medium was
changed to culture medium containing 10% serum after
1
were ascertained by H NMR spectrum (Varian 400 MHz, Palo
Alto, USA). (Figure S4-S7) To be a control, imidazolyl modified
G2 were synthesized by a similar method between G2 and
imidazole-4-carboxaldehyde. The synthesis method was the
same as G2-AM unless the lack of taking off the protection
incubation at 37
℃ 4 h. Cells were further incubated for 44 h
and 20 μL MTT (5 mg/mL) was added to each well and reacted
for 4 h at 37 °C. Then the media in each well was replaced by
0.2 mL DMSO and incubated for 4 h. The absorbance at 490
nm was recorded by a microplate reader (Power Wave XS, Bio-
TEK, United States). The relative cell viability (%) of the
1
base. The products were ascertained by H NMR spectrum.
(Figure S8-S10)
Formation and characterization of the complex.
complexes
and
materials
was
calculated
by
To get the complex, different polymers and pGL3 plasmid were
dissolved in double distilled water respectively, and equal
volume of the polymer and pGL3 solutions were vortexed for
30 s, then incubated for 30 min at room temperature. The final
pGL3 concentration of the complex was 40 μg/mL. All of the
complexes in this article were prepared in this method unless
otherwise noted.
A
sample/Acontrol×100%, the control group received the same
treatments except adding gene complexes or materials.
Cell uptake assay.
Cells were seeded in a 24-well plate at a density of 1×105 /well
and incubated for 24 h, then the media was replaced with 0.5
mL serum-free medium and 0.05 mL complex, and each well
included 2 μg TOTO-3 labeled pGL3. Result was analyzed by
flow cytometry 4 h later.
The size distribution and zeta potential of the complexes
prepared from different polymers were determined by
dynamic light scattering (DLS). The formed complexes were
stained by phosphotungstic acid and their morphology
structures were characterized using transmission electron
microscopy (TEM). To investigate the DNA condensation ability
of the polymers, the complexes were prepared and analyzed
To investigate its delivery ability to cell nucleus, cells were
incubated with TOTO-3 labeled complex for 4 h, then the cell
nucleus was stained with a blue fluorescent dye (DAPI). The
fluorescent images were visualized with a laser scanning
confocal microscope.
Buffering capability.
by 1% agarose gel electrophoresis (120V
，40min).
EGFP Assay.
We study the buffering capacity of G2-AM-11 by acid-base
titration, with G2 and G2-M-11 as controls. Each polymer was
dissolved in 30 mL double distilled water. All of their
Cells were seeded in a 48-well plate at a density of 2×104 /well
and incubated 24 h before transfection, then the medium was
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K. Charisse, M. W. Kieran, K. Fitzgerald, M. Nahrendorf, D.
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Panigrahy, A. Schroeder, V. Koteliansky, R. Langer, D. G.
concentration was 0.2 mg/mL. Each solution was adjusted to
about pH 10 by 0.1 M NaOH solution and then was titrated by
0.1 M HCl in 20 μL increments under vigorous stirring
condition at room temperature. The pH was measured by a
same pH meter and data were recorded when the numerical
reading was stable.
Conclusions
In summary, we designed a novel multifunctional group, 2-
aminoimidazole
(AM)
for
gene
delivery
.
Using
Anderson, Nat. Nanotechnol., 2014, 9, 648-655.
14 M. Wang, H. Liu, L. Li, Y. Cheng, Nat. Commun., 2014, 5,
poly(amidoamine) generation 2 grafted with AM as a gene
vector, the transfection efficiency was ~200 times greater than
3053.
for the parent dendrimer and performed better than 15 L. H. Wang, D. C. Wu, H. X. Xu, Y.Z. You, Angew Chem. Int. Ed.
Engl., 2016, 55, 755-759.
Lipofectamine 2000. Assays revealed that AM could be
conducive to condensing DNA into small, monodisperse
16 S. Y. Sun, S. Shen, C. F. Xu, H. J. Li, Y. Liu, Z. T. Cao, X. Z. Yang,
J. X. Xia, J. Wang, J. Am. Chem. Soc., 2015, 137, 15217-15224.
17 J. L. Choi, J. K. Tan, D. L. Sellers, H.Wei, P. J. Horner, S. H. Pun,
Biomaterials, 2015, 54, 87-96.
nanoparticles,
enhancing
cellular
penetration,
endosome/lysosome escape and nuclear localization. More
importantly, unlike other cationic non-viral vectors with high 18 T. I. Kim, T. Rothmund, T. Kissel, S. W. Kim, J. Controlled
Release, 2011, 152, 110-119.
19 Z. Salmasi, W. T. Shier, M. Hashemi, E. Mahdipour, H. Parhiz,
transfection efficiency, G2-AM has a low toxicity. In general,
these studies highlight the effects of the AM group in gene
delivery and support further investigation into its applications.
K. Abnous, M. Ramezani, Eur. J. Pharm. Biopharm., 2015, 96
76-88.
,
20 K. Chen, L. Guo, J. Zhang, Q. Chen, K. Wang, C. Li, W. Li, M.
Qiao, X. Zhao, H. Hu, D. Chen, Acta Biomater., 2017, 48, 215-
226.
21 E. G. Stanzl, B. M. Trantow, J. R. Vargas, P. A. Wender, Acc.
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Conflicts of interest
22 T. S. Bayard, W. S. Walter, L. M. Calvin, J. Org. Chem., 1964,
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23 J.D. Sullivan, R.L. Giles, R.E. Looper, Curr. Bioactive Comp.,
There are no conflicts to declare.
2009, 5, 39-98.
24 M. J. Ernsting, M. Murakami, A. Roy, S. D. Li, J. Controlled
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25 Yang, J.; Zhang, Q.; Chang, H.; Cheng, Y. Surface-Engineered
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26 (a) Y. Bae, E. S. Green, G. Kim, S. J. Song, J. Y. Mun, S. Lee, J.
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We are grateful for the financial support from the National
Science Foundation of China (Grant no. 81473148 and
81690263) and the Foundation Program of Key Laboratory of
Smart Drug Delivery of the Ministry of Education.
27 H. Tian, W. Xiong, J. Wei, Y. Wang, X. Chen, X. Jing, Q. Zhu,
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