A Synthetic Bioinspired Carbohydrate Polymer with Mucoadhesive Properties

Article abstract of DOI:10.1002/anie.201911720

Mucoadhesive polymers are of significant interest to the pharmaceutical, medical device, and cosmetic industries. Polysaccharides possessing charged functional groups, such as chitosan, are known for mucoadhesive properties but suffer from poor chemical d

Full text of DOI:10.1002/anie.201911720

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Accepted Article
Title: A Synthetic Bioinspired Carbohydrate Polymer with
Mucoadhesive Properties
Authors: Mark W Grinstaff, Anant S. Balijepalli, Robert C. Sabatelle,
Mingfu Chen, and Bela Suki
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10.1002/anie.201911720
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A Synthetic Bioinspired Carbohydrate Polymer with
Mucoadhesive Properties
Anant S. Balijepalli[a], Robert C. Sabatelle[a], Mingfu Chen[a], Bela Suki[a], and Mark W. Grinstaff[a],[b],[c],
*
chemical characteristics of polysaccharides for biological
applications (pyranose backbone, numerous hydroxyl
Abstract: Mucoadhesive polymers are of significant interest to the
pharmaceutical, medical device, and cosmetic industries.
Polysaccharides possessing charged functional groups, such as
chitosan, are known for mucoadhesive properties but suffer from
poor chemical definition and solubility, while the chemical synthesis
of polysaccharides is challenging with few reported examples of
synthetic carbohydrate polymers with engineered-in ionic
functionality. We report the design, synthesis, and evaluation of a
synthetic, cationic, enantiopure carbohydrate polymer inspired by the
structure of chitosan. These water-soluble, cytocompatible polymers
are prepared via an anionic ring-opening polymerization of a bicyclic
β-lactam sugar monomer. The synthetic method provides control
over the site of amine functionalization and the length of the polymer
while providing narrow dispersities. These well-defined polymers are
mucoadhesive as documented in single molecule scale (AFM), bulk
solution phase (FRAP), and ex vivo tissue experiments. Polymer
length and functionality affects bioactivity as long, charged polymers
display higher mucoadhesivity than long, neutral polymers or short,
charged polymers.
functionalities, defined stereochemistry) necessitate rigorous
and lengthy synthesis to prepare even oligosaccharides (<10
repeating units) [18–22]. Consequently, efforts are ongoing to
develop carbohydrate-inspired polymers with non O-glycosidic
linkages[23,24]
such
as
amide[25]
,
carbonate[26,27]
,
and
phosphodiester[28] linkages. These novel materials, however, fail
to recapitulate all of the key physicochemical and biointerfacial
properties of polysaccharides. Indeed, previously reported
carbohydrate-inspired polymers largely suffer from poor
aqueous solubility to complete insolubility, a lack of deprotected
functional groups important for biological interaction (e.g.,
hydroxyls, amines, phosphates, carboxylates)[23,24,26–28], and/or
rapid degradation in acidic or basic pH[23,24,29]. Moreover, the
rigid pyranose ring backbone absent in some carbohydrate-
mimic[25] and glyco-polymers[30] plays an essential role in
polymer structure and resulting macroscale properties[31–33]
.
Thus, there is a significant unmet need for well-defined materials
that closely resemble polysaccharides and possess their highly
desirable properties, such as water solubility and deprotected
functional groups, to harness in biomedical applications.
Mucus is
a dense network comprised of macromolecular
Herein, we describe a novel synthetic route and resulting
carbohydrate polymer, inspired by the composition of chitosan,
which shares many of its characteristics including a rigid
pyranose backbone, hydroxyl functionalities, stereoregularity,
components including enzymes, extracellular nucleic acids, and
high molecular weight proteins, including heavily glycosylated
mucins[1].
Mucoadhesive
pharmaceutical, medical device, and cosmetic applications,
adhere to this matrix through combination of chain
entanglement, ionic interactions, and hydrogen bonding[2–5]
materials,
of
interest
for
and
a
regioselectively-introduced amine. The one-day
a
polymerization of the protected sugar-β-lactam monomer yields
polymers from 10 to 50 repeat units with low dispersity. In
.
Polysaccharides are uniquely suited to mucoadhesive
applications due to their high degree of polymerization, rigid
backbone structure, dense functionality with hydrogen bond-
participating groups, and potential for ionic functionality.
contrast to polysaccharides, these carbohydrate polymers,
termed poly-amido-saccharides (PASs)
[31,33–35]
[36], possess an
amide linkage between the carbohydrate repeat units. The
polymers adhere to mucin with a length and charge dependency
as demonstrated through a combination of molecular-based
biophysical techniques as well as ex vivo analysis on mucosal
tissue.
Chitosan, in particular, is
polysaccharide widely used as a mucoadhesive material[5–10] as
it adheres through combination of charge interactions,
hydrogen bonding, and hydrophobic effects[2]. Despite their
advantages, well-defined polysaccharides for detailed
a
cationic naturally-obtained
a
We are cognizant of the need to avoid the loss of chemical
definition associated with post-polymerization modification as
well as the need for facile, controlled and reproducible chemical
synthesis of carbohydrate polymers, and, thus, we introduce an
amine functionality at the monomer level followed by anionic
ring-opening polymerization. As shown in Figure 1A, we first
prepared a regioselectively amine-functionalized glycal from D-
glucal (1) suitable for subsequent monomer synthesis and
polymerization. Using the bulky triisopropylsilyl chloride group
(TIPS-Cl), we regioselectively protected the primary 6’-OH in
good yield (2), followed by introduction of O-benzyl groups at the
3’- and 4’-OH positions (3). With these positions protected, we
subsequently removed the TIPS group with tetrabutylammonium
fluoride (TBAF) to yield 4, thus regenerating the 6’-OH for
functionalization. It should be noted that intermediate 4 serves
as an important branching point for the synthesis of new
regioselectively functionalized PAS monomers. Next, we
converted the 6’-OH of 4 to a N-nosyl-N-boc protected 6’-N
functionality using the Mitsunobu reaction with di-2-methoxyethyl
azodicarboxylate (DMEAD) and triphenylphosphine (PPh3)
(Figure 1B). The reaction of 5 with chlorosulfonyl isocyanate at -
68 oC proceeded with significantly slower kinetics than
previously observed with our benzyl-protected glycals (Figure
S1). As an alternative strategy, we utilized the room
experimental studies and for translational applications are
difficult to obtain as they are mixtures of polymers of differing
molecular weight and/or degree of branching, contaminated by
chemically similar entities [11–13], and compositionally variable
due to batch-to-batch variation as a result of being isolated from
natural sources [14–17]. Alternatively, purely synthetic routes to
such polymers are equally challenging as many of the desirable
[a]
A.S.B., R.C.S., M.C., B.S., M.W.G.
Department of Biomedical Engineering
Boston University
44 Cummington Mall, Boston, MA, 02215
E-mail: mgrin@bu.edu
[b]
[c]
M.W.G
Department of Chemistry
Boston University
712 Beacon Street, Boston, MA, 02215
M.W.G
School of Medicine
Boston University
72 East Concord Street, Boston, MA, 02118
Supporting information and the ORCID identification number(s) for
the author(s) of this article can be found under:
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10.1002/anie.201911720
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Figure 1. Small molecule and polymer synthesis. (A) Synthesis of regioselectively-modified amine glycal; (B) Monomer synthesis, nosyl
deprotection, and polymerization of amine-modified monomer; (C) Synthesis of thiol-containing initiator; (D) Mn vs [M]o/[I] for protected AmPAS
(P1’’-P3’’); (E) Mn vs [M]o/[I] for deprotected AmPAS (P1-P3). Pink – P1, Green – P2, Gold – P3.
temperature-stable trichloroacetyl isocyanate (TCAI), facilitating
the 8-10 day cycloaddition reaction to form monomer 6. As we
observed no polymer formation over 24 hours, we reduced the
steric constraints of 6 by removing the N-nosyl group using
group using trifluoroacetic acid (TFA) followed by a Birch type
reduction to remove the O- and S-benzyl groups (revealing the
terminal thiol). The resulting water-soluble polymers were
purified by dialysis against pure water and lyophilized to obtain
K2CO3 and PhSH to yield monomer 7, which polymerizes readily. white, cotton-like solids. All polymers possess molecular weights
in good agreement with theoretical values and low dispersity
<1.3 (NMR, IR, and GPC traces in S4-6, S8, S9B). IR spectra
reveal a broad peak at 3300 cm-1 indicating deprotection of O-
To obtain a single amine-orthogonal thiol functionality for post-
polymerization conjugation to a fluorescent dye, we prepared the
pentafluorophenyl ester of S-benzyl-thioglycolic acid (9) via a
carbodiimide coupling reaction (Figure 1C). Using this Pfp ester
9, we synthesized the polymers in good yield (Figure 1B; Table
1), and the polymers exhibit molecular weights ranging from 5.6
to 26.6 kDa as [M]o/[I] is varied from 12 to 50 with dispersity <1.2
as determined by THF size exclusion chromatography with
polystyrene standards (Figure 1D, GPC traces in S9A). The
relatively straightforward synthetic methodology of PAS is
especially attractive for the synthesis of amine-functional
carbohydrate polymers, where there are relatively few literature
examples and many of these suffer from low degrees of
polymerization, poor aqueous solubility, and harsh reaction
benzyl groups (Figure S4) and CD spectra display a minimum at
220 nm ([θ]MRE ~ -7 x 103 deg*cm2*dmol-1) and maximum at 180
nm ([θ]MRE ~ 30 x 103 deg*cm2*dmol-1) (Figure S5) consistent
with a helical structure.
Table 1. Size exclusion chromatography of AmPAS
polymers.
Polymer [M]o/[I] Mn,theor
(kDa)
Mn,GPC
(kDa)
Mw,GPC
(kDa)
Đ
Yield
conditions[18]
.
P’’1
P’’2
P’’3
P1
12
25
50
12
25
50
5.6
11.7
23.4
2.6
5.6
11.2
26.6
4.1
6.6
12.1
29.2
4.7
1.2 72%
1.1 81%
1.1 79%
1.1 41%
1.2 79%
1.3 73%
The NMR spectra of P1’’-P3’’ confirm polymerization with the
presence of broad peaks (Figure S2) and IR spectra show the
amide bonds (~3200 cm-1 and ~1800 cm-1) and aromatic groups
(~1100 cm-1) present on the polymer (Figure S3). To deprotect
these polymers, as shown in Scheme 1, we removed the Boc
P2
5.2
5.6
6.5
P3
10.3
9.8
13.2
Scheme 1. Deprotection of P1’’-P3’’.
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Figure 3. (A) Schematic of polymers P1-P3 and their estimated average
lengths[31]; AmPAS12SH – P1, AmPAS25SH – P2, AmPAS50SH – P3.
Magnified: structure of repeating polymer subunit. (B) Schematic of SMFS
experiment. Magnified: structure of sialic acid. (C) Representative retraction
curves obtained from SMFS. D. Average number of adhesion events per
curve comparing 2-mercaptoethanol (2ME), GlcPAS, DP 50, and P1-P3 tips
over: unmodified mica (negative control), amine silanized (negative control),
and mucin functionalized. Data obtained by randomly selecting 5 sets of 20
curves to tabulate average number of events. This was repeated three times
to generate an average and standard deviation (error bars) (n.s., p> 0.05; *,
p < 0.05; **, p < 0.01, ***, p < 0.001).
IC50 3.8 µg/mL) (Figure S11). Next, we performed multiplexed
cytokine assays on RAW 264.7 cells[38] (see SI methods for
detailed explanation of chemiluminescence assay) with P1 and
P3, at a non-cytotoxic concentration of 10 µg/mL, to assess their
potential to elicit an immune response. Following a 24-hour
incubation period, the supernatant of wells exposed to P1 and
P3 as well as those exposed to media alone (negative control)
display nearly identical chemokine secretion profile (Figure 2D,
2E). In contrast, wells exposed to LPS (positive control)
additionally show upregulation and secretion of the cytokines IL-
6 and TNF-α[39]. In particular, the lack of IL-6 and TNF-α
expression in RAW 264.7 cells incubated with P1 and P3 is
highly encouraging and indicates that no component of P1 or P3
activates pattern recognition receptors on these cells.
Figure 2. (A-C) MTS cell proliferation assay conducted on A549, BEAS-2B,
and HLF-1 cells, respectively, exposed to P1-P3 for 24 hours (n=18,
biological replicates). Hill functions fit to each curve to extract IC50 shown
below corresponding graph. (D) Chemiluminescence images obtained (all 90
s exposure) for array panel membranes incubated with media of RAW 264.7
cells exposed to either media alone (negative control), P1, P3, or LPS
(positive control). (E) Densitometric quantification of chemiluminescence
signal.
As a first step toward assessing the biocompatibility of P1-P3,
we performed in vitro MTS proliferation assays with a set of
respiratory tract cell lines. Following 24 hours of exposure to P1-
P3, we observe <20% inhibition of proliferation for all cell lines at
concentrations as high as 10 µg/mL (Figure 2A-C). Furthermore,
the inhibitory effect on cell proliferation is length-dependent, with
IC50 values for P1 across all cell lines between 68 and 77 µg/mL
while the longer polymers P2 and P3 display IC50 values
between 14 and 34 µg/mL. While exhibiting greater cytotoxicity
than biocompatible PEG and Chitosan (both neutral at
physiological pH), P3 exhibits >4 fold higher tolerability than the
biocompatible[37] and similarly cationic linear PEI (MW 10 kDa,
We hypothesize that polymers P1-P3 (Table 1), like other ionic
carbohydrate polymers, will adhere to mucus as a result of their
cationic charge, hydrogen bonding capability, and potential for
chain entanglement with sialylated mucins. We selected single-
molecule force spectroscopy (SMFS), which utilizes the
optically-determined deflection of an AFM cantilever with known
spring constant in order to calculate the force experienced by a
probe conjugated to the tip during a molecular interaction event.
This technique is ideal for biopolymer analyses as it is non-
destructive, may be performed in solution, and can measure
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transient electrostatic or hydrogen-bonding interactions with
samples of heterogenous molecular weight that would otherwise
be lost in population averaged assays such as fluorescence
polarization[40,41]. Thus, SMFS allows for the direct quantification
of adhesive forces between molecules covalently attached to
AFM tips and species covalently attached to smooth surfaces
(e.g., silanized mica). For further background, we direct the
reader to two reviews (Newman, 2008[40] and Hughes, 2016[41]
)
on the SMFS technique. We conjugated P1-P3, 2-
mercaptoethanol (2ME), and a thiol-terminated 50-mer neutral
PAS (GlcPAS50) to gold-coated AFM tips via thiol-gold
chemistry (Figure 3B) and performed repeated extension and
retractions from a porcine gastric mucin (PGM) coated mica
surface in 10 mM PBS buffer pH 7.4 (Figure S12). Hysteresis
between the extension and retraction curves corresponds to
adhesion events, which reflect intermolecular interactions at
rupture lengths greater than 20 nm[42] (Figure 3C). When
comparing between tips on mucin-functionalized surfaces, all
four polymer functionalized tips exhibit significantly higher
adhesion event probability than 2ME functionalized tips (0.5
average events per curve) as expected (Figure 3D). Additionally,
P3 displays a significantly higher adhesion event probability on
mucin-coated surfaces (1.88 average events per curve) than all
other tips due to its combination of positive charge and relatively
higher chain length. The observation of no statistically significant
difference between P1 and P2, which are short, positively-
charged polymers, and the longer, uncharged GlcPAS50
support the hypothesis that the increased likelihood of
interaction for P3 is a result of the combination of these two
independently acting factors.
Adhesion events at the single molecule scale are stochastic,
occurring with varying rupture lengths, rupture forces, and
adhesion energies (area under curve)[43]
.
Analyzing the
statistical distribution of these three parameters, computationally
extracted from all recorded retraction curves, allows for the
detailed characterization of the nature of adhesion events and
discrimination between weaker and stronger events. Due to the
heterogeneity of polymer-mucin interactions, histograms of each
parameter were fit to mixed model Gaussians that are a
weighted linear combination of 2 independent Gaussians each
with its own mean and standard deviation (Table S14). As
shown in Figure 4A, this statistical analysis quantifies the
observed elongation of GlcPAS50 and P3 rupture length
histograms, each with a significant number of counts above 200
nm, in comparison to 2ME. While the mixed model Gaussian
distribution of 2ME is described by a single mean of 87 nm, the
distributions of GlcPAS50 and P3 include contributions with 35%
and 47% weight from means of 340 and 390 nm, respectively.
The modeled rupture length distributions of P1 and P2 similarly
include significant weight (>28%) from means greater than 200
nm (Figure S15). In contrast, the rupture force distributions
(Figure 4B) for all tips exhibit minimal differences with all
possessing mixed model Gaussian means of 60 and 230 pN;
total counts, however, are higher for P3 versus 2ME.
Figure 4. Left, 2-mercaptoethanol (2ME); middle – GlcPAS50; right – P3
(AmPAS50). Mixed model Gaussian fits were applied to generate probability
distribution functions and the integral was scaled to match the histogram
area. (A-C) Histograms of computationally extracted rupture length, rupture
force, and adhesion energy, respectively. (D) Scatter plots of maximum
rupture force (pN) vs rupture length (nm) for all adhesion events on mucin-
functionalized surfaces. Point area is scaled by the adhesion energy of that
event (smaller circle – less adhesion energy).
significant (>23%) contribution from a 22 aJ mean (Figure S16).
Importantly, the adhesion energy mixed model means for P1-P3
are all larger than those calculated for GlcPAS50 (0.9 and 13 aJ).
Finally, as evident in scatter plots combining all 3 parameters
(Figure 4D), P3 exhibits higher event frequency, rupture length,
and adhesion energy in comparison to 2ME and GlcPAS50. This
is particularly apparent at rupture lengths above 200 nm where
P3 rupture force distributions include weight from a mean of 200
pN (Figure S17) while those of GlcPAS50 are below 80 pN. This
observed mean rupture force of ~200 pN agrees with previously
reported values for Chitosan and PGM at an identical tip
retraction speed of 1 µm/s[42]. Taken together, these data show
that while P1-P3 and GlcPAS50 are all capable of binding with
mucin, P3 possesses a unique combination of positive charge
and chain length that translates to significant mucoadhesivity.
Despite comparable rupture forces, the observed force-
deflection curves (e.g., Figure 3C) of 2ME are generally sharp,
narrow peaks unlike the broad and exponentially increasing
peaks of P3. These differences manifest in the adhesion energy
(Figure 4C) histograms and mixed model Gaussians for each
functionalized tip. While the 2ME adhesion energy histogram is
modeled by a single distribution with a mean of 0.83 aJ, the
modeled distribution for P3 is described by significantly larger
means of 2.2 and 33 aJ (68% and 32% weight, respectively).
Mixed model Gaussians of P1 and P2 similarly include a
Next, we performed fluorescence recovery after photobleaching
(FRAP) experiments on TAMRA-labeled P3 and FITC-labeled
dextran in water and in dilute (0.5 mg/mL) and concentrated (10
mg/mL) mucin solutions. We additionally examined low
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While the single molecule and bulk measurements presented
hitherto provide insight into the mechanism and factors
associated with interactions between P1-P3 and mucin, their
effectiveness in mucoadhesive nanoformulations is best
characterized by their adhesion to animal tissue. Thus, we
incubated samples of trachea (porcine, Figure 5A), esophagus
(ovine, Figure 5B), and duodenum (porcine, Figure 5C), each
possessing either a significant number of mucus-producing
goblet cells on their inner lumen or glandular mucus secretion, in
PBS solutions of either TAMRA-labeled P3 or FITC-dextran for
30 min and then subjected the samples to repeated washing
with PBS to remove the bound polymer. The average
fluorescence intensity measured on each tissue sample
increases after incubation with both FITC-dextran and TAMRA-
P3 (Figure 5A, B). Samples incubated with TAMRA-P3, however,
demonstrate a significantly higher increase in fluorescence
intensity post-incubation compared to FITC-dextran. In the case
of porcine trachea and ovine esophageal tissue (Figure 5A-D),
FITC-dextran adheres slightly (40 units above baseline) and this
signal is readily lost upon washing the sample 3 times in PBS. In
contrast, the TAMRA-P3 soaked tissue shows significant loading
(120-130 units above baseline) and a large portion of this
loading (>75%) remains after 7 washes. While the increase in
signal relative to baseline is still higher for TAMRA-P3 on
porcine duodenum tissue (Figure 5E-F), compared to FITC-
dextran, both polymers remain with little loss of signal until wash
7. To quantify initial tissue loading of P3 in comparison to
dextran and the mucoadhesive, charged chitosan and alginate,
we incubated fresh porcine trachea, duodenum, and esophagus
samples with fluorescently labeled polymers, followed by a quick
PBS wash and vigorous sonication for 1 h to remove all bound
polymer. We only observed high loading for alginate on trachea
samples and poor loading for chitosan on all tissues, despite
their known mucoadhesivity. Nevertheless, significantly higher
loading of P3 occurs on duodenum and esophagus samples
compared to dextran, chitosan, and alginate. These data support
our hypothesis that TAMRA-P3 strongly adheres to biological
tissue and resists clearance (i.e., enhanced residence time), in
agreement with results obtained through AFM and FRAP
experiments.
Figure 5. (A-C) Images under UV irradiation (fixed illumination distance) of
porcine trachea (A), ovine esophagus (B), and porcine duodenum (C)
In summary, we report the synthesis of enantiopure cationic
carbohydrate polymers via anionic ring-opening polymerization
of a bicyclic β-lactam sugar monomer. Our synthetic method
imparts regioselective amine functionalization and control over
the length of the polymer with narrow dispersities. Results from
single molecular force spectroscopy demonstrate that tuning
polymer length impacts nanoscale interactions, as polymers with
degree of polymerization of 50 (P3) interact with mucin with
significantly higher frequency and adhesion energy than 2-
mercaptoethanol, GlcPAS (degree of polymerization 50, non-
cationic control), P1 (degree of polymerization 12), and P2
(degree of polymerization 25). Additionally, in contrast to the
non-mucoadhesive dextran, P3 adheres to ex vivo mucin coated
tissues. Mucosal routes for drug delivery are underutilized in
comparison to parenteral administration, in spite of the large
available surface area and minimal invasiveness, due to the lack
of well-defined polymeric materials available for tunable
adhesion, prolonged retention time, and modification of viscosity.
The highly controlled synthetic methodology and associated
tunability in biointerfacial properties of these cytocompatible and
bioinspired PASs expands the formulation scientist’s toolkit for
mucosal drug delivery.
collected from
a freshly sacrificed animal (0) and incubated with either
TAMRA-P3 or FITC-dextran for 30 minutes (1) and washed up to 7 times
afterwards (2-8). Images are of the same sample over each wash. (D-F)
Quantification of images, split into RGB channels and followed by
densitometry, obtained for each tissue sample (n=3). Values obtained relative
to samples in PBS only to account for background. Error bars represent the
variability in densitometry between independent tissue samples. (G) Tissue
loading of FITC-dextran, FITC-alginate, FITC-chitosan, or TAMRA-P3. Tissue
samples incubated with polymer for 30 minutes followed by vigorous
sonication to remove bound polymer; fluorescence quantified via plate reader
against standard curve (n=3 tissue samples). Error bars are standard deviation
(n.s., p> 0.05; *, p < 0.05; **, p < 0.01, ***, p < 0.001).
molecular weight FITC-labeled alginate and chitosan, to
compare P3 to other charged, mucoadhesive polymers. In a 20
mM HEPES solution, the TAMRA-labeled P3 diffuses slower
than FITC-dextran with a diffusion coefficient of 1.7 x 10-7 versus
1.3 x 10-6 cm2/s. Moreover, when dissolved in a 10 mg/mL mucin
solution, the diffusion coefficient of TAMRA-labeled P3
decreases by nearly 70% to 5.3 x 10-8 cm2/s (Figure S18A) while
remaining relatively unchanged for the uncharged and non-
mucoadhesive FITC-dextran (Figure S18B). FITC-alginate
exhibits a similar 80% reduction in diffusion coefficient with
increasing mucin concentration, while only a modest 50% drop
is observed for FITC-chitosan (Figure S18C-D).
Acknowledgements
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10.1002/anie.201911720
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This work was supported in part by BU and DOD USU
(HU0001810012). The authors would like to acknowledge Dr.
Anderson Chen of the BU BME Micro/Nano imaging facility, Dr.
Clemens Anklin of Bruker BioSpin, and Dr. Paul Ralifo of the BU
Chemical Instrumentation Center for assistance.
[20]
[21]
O. Calin, S. Eller, P. H. Seeberger, Angew. Chemie Int. Ed. 2013,
52, 5862–5865.
P. Stallforth, B. Lepenies, A. Adibekian, P. H. Seeberger, J. Med.
Chem. 2009, 52, 5561–5577.
[22]
[23]
P. H. Seeberger, D. B. Werz, Nature 2007, 446, 1046–1051.
L. Li, K. Knickelbein, L. Zhang, J. Wang, M. Obrinske, G. Z. Ma, L.-
M. Zhang, L. Bitterman, W. Du, Chem. Commun. 2015, 51, 13078–
13081.
Author Information
Corresponding Author: mgrin@bu.edu
ORCID numbers
[24]
[25]
[26]
[27]
[28]
[29]
[30]
[31]
[32]
[33]
[34]
[35]
[36]
[37]
L. Li, J. Wang, M. Obrinske, I. Milligan, K. O’Hara, L. Bitterman, W.
Du, Chem. Commun. 2015, 51, 6972–6975.
M. Metzke, N. O’Connor, S. Maiti, E. Nelson, Z. Guan, Angew.
Chemie Int. Ed. 2005, 44, 6529–6533.
Anant S. Balijepalli
Robert C. Sabatelle 0000-0002-4335-5384
Mingfu Chen
Bela Suki
Mark W. Grinstaff
0000-0003-4686-8123
T. P. Gustafson, A. T. Lonnecker, G. S. Heo, S. Zhang, A. P. Dove,
K. L. Wooley, Biomacromolecules 2013, 14, 3346–3353.
A. T. Lonnecker, Y. H. Lim, K. L. Wooley, ACS Macro Lett. 2017, 6,
748–753.
0000-0002-0169-613X
0000-0002-9720-1006
0000-0002-5453-3668
Y.-Y. T. Tsao, K. L. Wooley, J. Am. Chem. Soc. 2017, 139, 5467–
5473.
Keywords: Biocompatibility • Carbohydrate Polymer • Cationic
polymers • Mucoadhesive • Single Molecule Force Spectroscopy
S. E. Felder, M. J. Redding, A. Noel, S. M. Grayson, K. L. Wooley,
Macromolecules 2018, 51, 1787–1797.
[1]
[2]
B. K. Rubin, Respir. Care 2002, 47, 761–8.
V. Ladmiral, E. Melia, D. M. Haddleton, Eur. Polym. J. 2004, 40,
431–449.
I. A. Sogias, A. C. Williams, V. V. Khutoryanskiy,
Biomacromolecules 2008, 9, 1837–1842.
S. L. Chin, Q. Lu, E. L. Dane, L. Dominguez, C. J. McKnight, J. E.
Straub, M. W. Grinstaff, J. Am. Chem. Soc. 2016, 138, 6532–6540.
E. L. Dane, S. L. Chin, M. W. Grinstaff, ACS Macro Lett. 2013, 2,
887–890.
[3]
[4]
Y. Y. Wang, S. K. Lai, C. So, C. Schneider, R. Cone, J. Hanes,
PLoS One 2011, 6, e21547.
A. Sosnik, J. Das Neves, B. Sarmento, Prog. Polym. Sci. 2014, 39,
2030–2075.
R. Xiao, E. L. Dane, J. Zeng, C. J. McKnight, M. W. Grinstaff, J. Am.
Chem. Soc. 2017, 139, 14217–14223.
[5]
[6]
[7]
J. D. Smart, Adv. Drug Deliv. Rev. 2005, 57, 1556–1568.
C. Valenta, Adv. Drug Deliv. Rev. 2005, 57, 1692–1712.
H. Takeuchi, H. Yamamoto, Y. Kawashima, Adv. Drug Deliv. Rev.
2001, 47, 39–54.
E. L. Dane, M. W. Grinstaff, J. Am. Chem. Soc. 2012, 134, 16255–
16264.
S. E. Stidham, S. L. Chin, E. L. Dane, M. W. Grinstaff, J. Am. Chem.
Soc. 2014, 136, 9544–9547.
[8]
R. Jayakumar, D. Menon, K. Manzoor, S. V Nair, H. Tamura,
Carbohydr. Polym. 2010, 82, 227–232.
E. L. Dane, S. L. Chin, M. W. Grinstaff, ACS Macro Lett. 2013, 2,
887–890.
[9]
M. George, T. E. Abraham, J. Control. Release 2006, 114, 1–14.
L. Casettari, L. Illum, J. Control. Release 2014, 190, 189–200.
S. K. Tam, J. Dusseault, S. Polizu, M. Ménard, J.-P. Hallé, L. Yahia,
Biomaterials 2006, 27, 1296–1305.
[10]
[11]
I. Y. Perevyazko, M. Bauer, G. M. Pavlov, S. Hoeppener, S.
Schubert, D. Fischer, U. S. Schubert, Langmuir 2012, 28, 16167–
16176.
[12]
R. Lieder, V. S. Gaware, F. Thormodsson, J. M. Einarsson, C.-H.
Ng, J. Gislason, M. Masson, P. H. Petersen, O. E. Sigurjonsson,
Acta Biomater. 2013, 9, 4771–4778.
[38]
[39]
M. Derive, Y. Bouazza, C. Alauzet, S. Gibot, Intensive Care Med.
2012, 38, 1040–1049.
Y. Liu, H. Yin, M. Zhao, Q. Lu, Clin. Rev. Allergy Immunol. 2014, 47,
136–147.
[13]
[14]
[15]
G. A. Paredes-Juarez, B. J. de Haan, M. M. Faas, P. de Vos, Mater.
(Basel, Switzerland) 2014, 7, 2087–2103.
[40]
[41]
[42]
K. C. Neuman, A. Nagy, Nat. Methods 2008, 5, 491–505.
M. L. Hughes, L. Dougan, Reports Prog. Phys. 2016, 79, 076601.
K. E. Haugstad, A. G. Håti, C. T. Nordgård, P. S. Adl, G. Maurstad,
M. Sletmoen, K. I. Draget, R. S. Dias, B. T. Stokke, Polymers
(Basel). 2015, 7, 161–185.
B. Sabrina, B. Antonella, T. Giangiacomo, B. Donata, M.
Terbojevich, Biomacromolecules 2005, 6, 168–173.
R. H. Chen, J. R. Chang, J. S. Shyur, Carbohydr. Res. 1997, 299,
287–294.
[16]
[17]
[18]
[19]
E. Kabat, A. Bezer, Arch. Biochem. Biophys. 1958, 78, 306–318.
M. Stefansson, Anal. Chem. 1999, 71, 2373–2378.
R. Xiao, M. W. Grinstaff, Prog. Polym. Sci. 2017, 74, 78–116.
D. B. Werz, P. H. Seeberger, Chem. - A Eur. J. 2005, 11, 3194–
3206.
[43]
A. Janshoff, M. Neitzert, Y. Oberdörfer, H. Fuchs, Angew. Chemie
2000, 39, 3212–3237.
This article is protected by copyright. All rights reserved.

10.1002/anie.201911720
Angewandte Chemie International Edition
COMMUNICATION
COMMUNICATION
We describe the synthesis of a novel
Anant S. Balijepalli, Robert C.
Poly-Amido-Saccharide
regioselectively-introduced
with
amine
Sabatelle, Mingfu Chen, Bela Suki and
Mark W. Grinstaff
functionality, inspired by chitosan.
These well-defined cationic polymers
Page No. – Page No.
are
biocompatible
and
non-
A Synthetic Bioinspired
Carbohydrate Polymer with
Mucoadhesive Properties
immunogenic. Single molecule force
spectroscopy
experiments
and
reveal
ex
vivo
significant
mucoadhesivity.
This article is protected by copyright. All rights reserved.