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Polyacetals: Water-Soluble, pH-Degradable Polymers with Extraordinary Temperature Response Sanjoy Samanta,† Danielle R. Bogdanowicz,‡ Helen H. Lu,‡ and Jeffrey T. Koberstein*,† †

Department of Chemical Engineering and ‡Department of Biomedical Engineering, Columbia University, New York, New York 10027, United States S Supporting Information *

ABSTRACT: Temperature-responsive polymers exhibit a drastic change in solubility upon change in temperature, a property responsible for their increased use in a wide variety of emerging smart material applications. Here, we describe a flexible new family of polyacetals with lower critical solution temperature (LCST) behavior, falling out of aqueous solution upon increase in temperature. These are the first synthetic LCST polymers to be intrinsically biodegradable, forming neutral degradation products in acidic environments. The temperature response of the polyacetals is extraordinary. Their LCST cloud point temperatures can be predicted and tuned to high precision over a temperature range of ca. 6−80 °C as they are linearly dependent on the number of methylene and ethylene oxide units in the diol and divinyl ether monomers. We further demonstrate that the LCST temperatures are insensitive to salt and/or polymer concentration, properties that are important to in vivo applications, and that they are biocompatible. nanoparticle−polymer hybrids.23,24 In biomedical applications, it is desirable to tune the LCST so that the polymer loses solubility at a specific, biologically relevant temperature. For example, in hypothermia-targeted drug delivery,23,24 a drug delivery vehicle is designed to be stable in solution at physiological temperature but to fall out of solution upon encountering a tumor with a slightly elevated temperature associated with hyperthermia.25 In this fashion, drug-laden micelles can be concentrated and immobilized within a tumor. Controlled release of the drug is possible if the drug delivery vehicle comprises a mechanism for biodegradation within the tumor microenvironment, which is subject to acidosis with a pH of 5−6.5.26−28 Because extant synthetic LCST homopolymers are intrinsically nondegradable, sites for biodegradation must be added by copolymerization with monomers containing degradable (e.g., acid-labile) backbone linkages29−33 or with non-LCST biodegradable macromonomers.34 Elastin-like polypeptides have recently been prepared by genetic engineering techniques and offer both intrinsic biodegradability and an LCST temperature that can be tuned to temperatures relevant for a variety of drug delivery applications.35 We report herein the synthesis and characterization of an exciting new family of water-soluble, temperature-responsive acetal polymers with remarkable LCST temperature response. While structurally different polyacetals have been prepared previously, they have not been shown to manifest LCST behavior. The new polyacetals are the first synthetic LCST

1. INTRODUCTION The fundamental nature of polymer applications has changed markedly over the years. While early applications called for polymers that were strong and light, resistant to chemicals, and environmentally inert, today’s applications demand smart polymers that can shrink, expand, thicken, or change effectively any physical, optical, or electronic property in response to a host of different external stimuli. Smart polymers have been developed to respond to almost every conceivable stimulus:1,2 temperature, pH, light, magnetic and electric fields, ultrasound, guest−host interactions, electron transfer (oxidation−reduction), the presence of compounds such as sugars, salt, antigens, enzymes, and even CO2. The stimuli-responsive polymers that have received the most attention to date are those that respond to temperature. Temperature-responsive polymers (TRPs)3,4 exhibit a drastic and discontinuous change in some physical property with change in temperature. While in principle the term temperature-responsive can apply to any property, in common use it has come to refer to polymers that show a drastic change in solubility with temperature. Polymers with an upper critical solution temperature (UCST) lose solubility on cooling whereas polymers with a lower critical solution temperature (LCST) lose solubility on heating. TRPs have a myriad of potential smart material applications such as stationary phases in liquid chromatography,5 lithography,6 microfuidics,7 and molecular imprinting.8 Water-soluble TRPs are particularly important due to their relevance in biomedical and green technologies. Medical applications include9−11 bioseparations,12−15 tissue scaffolds,16 actuators/ artificial muscles,17 and drug delivery vehicles18 such as block copolymer micelles,19,20 hydrogels,21 nanoparticles,22 and © XXXX American Chemical Society

Received: October 20, 2015 Revised: February 3, 2016

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= 0) with ethylene glycol (i.e., n2 = 2, m2 = 0), which has vinyl ether end groups and a molecular weight of 13 500 Da. 2.2. NMR. The purities and compositions of the polymers were verified by 1H NMR (Bruker instruments, 400 MHz) in CDCl3 and in D2O (if necessary). Figure S.2 shows a typical NMR spectrum for vinyl ether-terminated polyacetal. The vinyl ether end groups, denoted as a and b in the structural schematic, give rise to signals at (4 ppm, 4.25 ppm) and (6.5 ppm), respectively. These signals are absent in the NMR spectrum for a typical hydroxyl-terminated polyacetal, as shown in Figure S.3 and in the expanded NMR spectrum shown in Figure S.4. The inability to detect vinyl ether protons indicates that the conversion for the hydroxyl-terminated materials is essentially 100%. 2.3. GPC. Solutions for GPC analysis were prepared by dissolving the polymer in THF followed by filtration through a 0.2 μm PTFE syringe filter. Solutions were injected manually into the GPC (2 Polymer Laboratories ResiPore Columns in a Shimadzu HPLC system: LC-10ATvp pump, CTO-10ACvp column oven, SPD-10a UV−vis detector, RID-10A RI detector, SCL-10Avp system controller and a Wyatt miniDAWN TREOS static light scattering detector). Data analysis was performed using custom MATLAB programs and Origin 8.0. GPC data were calibrated using linear PS standards [EasiVial PSM(2 mL), Varian]. The degree of polymerization, DP, can be manipulated by adjusting the reactant ratio, r, according to

polymers to be intrinsically biodegradable, and their degradation mechanism is unique, producing neutral degradation products in acidic environments. Most other degradable polymers, in contrast, produce acidic degradation products that can cause inflammation.36 The LCST behavior of aqueous polyacetal solutions is extraordinary: the LCST cloud point transitions are sharp, occurring over a range of 3−5 °C, show almost no hysteresis between heating and cooling cycles, and the cloud point temperatures (CPTs) can be predicted directly from the molecular structure of the monomers and tuned to high precision anywhere within a range of about 6−80 °C because they are linearly dependent on the number of methylene and/or ethylene oxide units comprising the monomers.

2. EXPERIMENTAL PROCEDURE 2.1. Polyacetal Synthesis. Polyacetals (PA) are synthesized by a simple step growth polymerization of divinyl ethers with diols (Figure 1) to produce a polymer that is linked by pH degradable acetal

DP = (1 + r )/(1 − r )

(1)

where r ≡ [MLim]/[MEx], [MLim] is the moles of limiting reactant, and [MEx] is the moles of excess reactant. This form of Carother’s equation assumes that the conversion is 100%. Macromonomers terminated with either vinyl ether or hydroxyl functional groups can be readily prepared by using an appropriate stoichiometric excess of either the vinyl ether or diol monomer, respectively. Properties of the polyacetals synthesized for this work are shown in Table 1. The theoretical and apparent experimental molecular weights, determined by GPC, agree well for the hydroxyl-terminated macromonomers, but the experimental values are consistently higher by a factor of about 2 for the vinyl ether-terminated macromonomers. The reported experimental molecular weights are based upon polystyrene standards, and the values are therefore relative to the hydrodynamic volume of polystyrene. The discrepancies between theoretical and experimental molecular weights may be due to the effects of end groups on the hydrodynamic volume or due to interactions of the end groups with the column packing. A universal calibration to obtain absolute molecular weights was not attempted. The fact that protons associated with vinyl ether end groups are not detectable in the hydroxyl-terminated polyacetals (see Figures S.3 and S.4) and that the GPC molecular weights correlate well with (1) as shown in Figure S.5 support the conclusion that the conversion obtained in the polymerizations is effectively 100%. 2.4. UV Transmittance. Cloud point temperatures (CPTs) for freshly prepared polymer solutions (DI water, Millipore Milli-Q, 5 g L−1) were determined by turbidity measurements (transmittance mode, 500 nm) as a function of temperature (cell path length, 10 mm; one heating/cooling cycle at rate of 1 °C min−1) using a UV−vis spectrometer (Agilent 8453) equipped with a temperature controller (Quantum Northwest, TC1). The CPTs were taken as the midpoint in the transmittance versus temperature curve obtained upon heating. 2.5. Cells and Cell Culture. Human anterior cruciate ligament (ACL) fibroblasts were isolated from explant culture of tissues obtained from a patient (male, 21 years old) undergoing ACL reconstruction. Cells (passage 4) were seeded at a density of 3 × 104 cells/cm2 in 24 well tissue culture plates (Corning, Tewksbury, MA) and maintained in Dulbecco’s Modified Eagle Medium (DMEM) supplemented with 10% fetal bovine serum (FBS, Atlanta Biologicals, Atlanta, GA), 1% nonessential amino acids, 1% penicillin− streptomycin, 0.1% amphotericin B, and 0.1% gentamicin sulfate at 37 °C and 5% CO2. All medium and supplements were purchased from Cellgro-Mediatech (Manassas, VA) unless otherwise noted. Cells were allowed to attach and spread on tissue culture plastic for 3 days prior to incubation with polyacetal.

Figure 1. Synthesis of polyacetals and their degradation to produce neutral products upon acid hydrolysis. The acetal linkages are shown in red, and the end groups are shown in blue. bonds.37 Polymerization reactions were carried out on a 10 mmol (1 equiv) scale. In brief, divinyl ether and diol in stoichiometric imbalance were allowed to react in the presence of an acid catalyst, pyridinium ptoluenesulfonate (PPTS, 0.05 equiv), in anhydrous dichloromethane (DCM, 1 mL). Because of its exothermic nature, the reaction was held initially at 0 °C for 30 min, after which the temperature was raised to room temperature, 25 °C, for the remainder of the reaction. After 2.5 h, DCM was removed by rotary evaporation under high vacuum for 45 min. The resultant polymer was extracted with ethyl acetate (EtOAc, 40 mL) and washed three times with dilute aqueous solutions of potassium carbonate (K2CO3). Each time the aqueous part was saturated by adding excess sodium chloride, NaCl, an important step, particularly for hydrophilic polyacetals. The organic part was dried over anhydrous sodium sulfate (Na2SO4) and passed through a short basic-Al2O3 column. The polymer was finally isolated by removing solvent using rotary evaporation followed by drying under vacuum at room temperature for 48 h. The polymerization reaction goes to completion within about 2 h (see Figure S.1 in the Supporting Information). The composition of each PA can conveniently be described by adopting the nomenclature PAijklEMn, where PA denotes polyacetal, the indices i through l denote the values of the four compositional variables n1, m1, n2, and m2, respectively (see Figure 1), the letter E denotes the end group type (V for vinyl ether or H for hydroxyl groups), and Mn designates the molecular weight in kilodaltons. A polymer designated as PA4020V13.5 therefore refers to polyacetal synthesized by the reaction of butanediol divinyl ether (i.e., n1 = 4, m1 B

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Macromolecules Table 1. Polyacetal Sample Designations, Compositions, Reactant Ratios, and Molecular Weights sample PA4020V13.5 PA4020V5.5 PA4020V2.8 PA4021V12.1 PA4022V14.9 PA4022V6.1 PA4022V3.1 PA4022V1.5 PA4022H4.6 PA4022H2.7 PA4022H1.8 PA4022H1.0 PA4023V17.3 PA2120V8.0 PA2220V9.2 PA2130V9.6 PA2230V11.5 PA2140V10.0 PA2240V11.9 PA2150V10.6 PA2250V12.3 a

divinyl ether (V) n1 n1 n1 n1 n1 n1 n1 n1 n1 n1 n1 n1 n1 n1 n1 n1 n1 n1 n1 n1 n1

= = = = = = = = = = = = = = = = = = = = =

4, 4, 4, 4, 4, 4, 4, 4, 4, 4, 4, 4, 4, 2, 2, 2, 2, 2, 2, 2, 2,

m1 m1 m1 m1 m1 m1 m1 m1 m1 m1 m1 m1 m1 m1 m1 m1 m1 m1 m1 m1 m1

= = = = = = = = = = = = = = = = = = = = =

0 0 0 0 0 0 0 0 0 0 0 0 0 1 2 1 2 1 2 1 2

diol (H) n2 n2 n2 n2 n2 n2 n2 n2 n2 n2 n2 n2 n2 n2 n2 n2 n2 n2 n2 n2 n2

= = = = = = = = = = = = = = = = = = = = =

2, 2, 2, 2, 2, 2, 2, 2, 2, 2, 2, 2, 2, 2, 2, 3, 3, 4, 4, 5, 5,

m2 m2 m2 m2 m2 m2 m2 m2 m2 m2 m2 m2 m2 m2 m2 m2 m2 m2 m2 m2 m2

= = = = = = = = = = = = = = = = = = = = =

0 0 0 1 2 2 2 2 2 2 2 2 3 0 0 0 0 0 0 0 0

V:H (mol/mol)

end groupa

Mn (theor) (kg/mol)

Mnb (GPC) (kg/mol)

1.034:1 1.083:1 1.18:1 1.04:1 1.04:1 1.09:1 1.19:1 1.48:1 1:1.06 1:1.11 1:1.21 1:1.515 1.04:1 1.05:1 1.05:1 1.05:1 1.05:1 1.05:1 1.05:1 1.05:1 1.05:1

V V V V V V V V H H H H V V V V V V V V V

6.1 2.56 1.23 5.7 7.15 3.36 1.68 0.73 5.0 2.8 1.53 0.71 8.25 4.29 5.15 4.56 5.4 4.83 5.7 5.1 5.95

13.5 5.50 2.80 12.1 14.9 6.10 3.10 1.50 4.60 2.70 1.80 1.00 17.3 8.00 9.20 9.60 11.5 10.0 11.9 10.6 12.3

Vinyl ether (V) or hydroxyl (H). bBy GPC using polystyrene standards.

2.6. Cytotoxicity Testing. On the day of the experiment, polyacetal was added to FBS-free, supplemented DMEM at the desired concentrations (0.1, 1, and 10% w/v). The solution was then filtered through a sterile filter with 0.22 μm pores, and 10% FBS was added. Afterward, the polyacetal solution (1 mL) was added to each well containing fibroblast monolayers, and cells were incubated at 37 °C and 5% CO2. Cell viability (n = 2/group) was evaluated at 3, 12, 24, and 72 h using Live/Dead staining (Molecular Probes, Eugene, OR). Briefly, samples were rinsed twice with phosphate buffered saline (PBS, Sigma-Aldrich, St. Louis, MO) and then stained, following the manufacturer’s suggested protocol. The samples were imaged by confocal microscopy (Olympus Fluoview, Center Valley, PA) at excitation wavelengths of 473 nm (live cells, indicated in green) and 559 nm (dead cells, indicated in red).

3. RESULTS The divinyl ether (1) and diol (2) monomers used each comprise a hydrophobic part containing n1 or n2 methylene units, respectively, and a hydrophilic part with m1 or m2 ethylene oxide units, respectively. This flexible system of amphiphilic monomers allows an entire family of versatile PAs to be prepared wherein the hydrophilic/hydrophobic balance can be tuned or adjusted by simply altering the ratio of methylene to ethylene oxide moieties in either of the two monomers. The CPTs of aqueous PA solutions were determined by analysis of UV transmittance experiments (λ = 500 nm), an example of which is shown in Figure 2. PA solutions are clear at low temperatures but become cloudy above a critical temperature, indicating that the polymers fall out of solution upon heating. The solubility transition is sharp, occurring over about 4 °C, and the heating and cooling cycles superimpose indicating a lack of hysteresis. The assignment of this transition to a cloud point associated with a lower critical temperature is confirmed by the dynamic light scattering results shown in Figure 3. The hydrodynamic radius shows an initial decrease with temperature as is expected when there is a decrease in solubility with temperature.

Figure 2. UV transmission measurements of the CPT for PA4022V14.9. The closed circles (solid line) were obtained upon heating while the open circles (dashed line) were obtained upon cooling.

The polymer chains should, as observed, gradually collapse as the solvent quality decreases with temperature during an LCST transition. As the cloud point is reached, chains begin to first aggregate and then to eventually phase separate as the temperature is increased above the cloud point temperature. The process of chain collapse and aggregation is observed to be reversible upon heating and cooling. The thermal response of the PAs is truly remarkable: the CPT is linearly dependent on three of the four composition variables defined in Figure 1. The dependence of the CPT on the average number of ethylene oxide repeats in the diol, m2,av, is shown Figure 4a for PAs prepared using butanediol divinyl ether (i.e., n1 = 4, m1 = 0). The circles denote polymers prepared using a single diol, while triangles denote polymers C

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Figure 3. Temperature dependence of the hydrodynamic radius of a polyacetal determined by dynamic light scattering.

prepared from a mixture of diols. m2 = 1.5 refers to a polymer prepared from a 50:50 mixture of m2 = 1.0 and m2 = 2.0 diols. Regression of the combined data yields a linear relationship between the CPT and m2,ave CPT(PA120m2,ave) = 6.50( ±0.54) + 12.2(± 0.29)m2,ave (2)

The error bars denote one standard deviation. This result indicates that the CPT can be predicted and tuned precisely to any temperature within the range 6−80 °C by simply using mixtures of two diol monomers to provide the appropriate m2,ave value. Each hydrophilic ethylene oxide unit added to the diol monomer (averaged over each of the diols used) increases the CPT by 12.2 °C. The CPT is also linearly dependent on the number of hydrophobic methylene groups in the diol monomer, n2 (Figure 4b). Regression of the two data sets in Figure 4b yields the relationships CPT(PA23n2 0) = 96.3( ±0.88) − 15.01(± 0.24)n2

(3)

CPT(PA22n2 0) = 87.6( ±1.15) − 15.66(± 0.31)n2

(4)

The addition of each hydrophobic methylene unit within the diol decreases the CPT by 15.3 °C (within error), regardless of the number of ethylene oxide units in the vinyl ether monomer. Increasing the number of hydrophilic ethylene oxide units in the vinyl ether monomer, m1, increases the CPT, with each additional unit adding 8.7 °C to the CPT. The linear dependence of CPTs on the molecular composition of the monomers is found, in all cases, when the diol and divinyl ether used are complementary, that is, when one is hydrophobic and the other is hydrophilic. When both monomers are hydrophilic, the CPT is not linearly dependent on m2, the number of ethylene oxide units in the diol, as shown in Figure 4c for two sets of data. All of the data in Figure 4 are based upon polymers with the same theoretical DP; that is, they were synthesized using the same reactant ratio of 1.04 mol of vinyl ether per mole of diol. At this reactant ratio, the theoretical DP is 51 and all polymers have vinyl ether end groups.

Figure 4. CPT temperatures as a function of polyacetal composition: (a) effect of the number of ethylene oxide units in the diol monomer (n1 = 4, m1 = 0, m2,ave, n2 = 2); (b) effect of the number of methylene units in the diol monomer (open squares, n1 = 2, m1 = 2, n2, m2 = 0), (open circles, n1 = 2, m1 = 1, n2, m2 = 0); (c) effect of the number of ethylene oxide units in the diol monomer (open squares, m1 = 1, m2, n1 = 2, n2 = 2), (open circles, m1 = 2, m2, n1 = 2, n2 = 2).

The molecular weights and the end group type can be systematically varied by altering the reactant ratio and choosing which monomer is in excess. The dependence of the CPT on the molecular weight and end group type for the series of PA4022 polymers is shown in Figure 5 for hydroxyl and vinyl ether end groups. The CPTs of vinyl-ether-terminated PAs increase with apparent molecular weight while those of D

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Figure 5. Inverse apparent molecular weight dependence of CPTs for hydroxyl-terminated (triangles) and vinyl ether-terminated (circles) polyacetals of composition PA4022. Molecular weights were determined by gel permeation chromatography and are relative to polystyrene standards.

Figure 6. Degradation profiles for PA4023V17.3 as a function of time and pH. The polymer concentration in all cases was 5 mg/mL while the concentration of the buffer solution was 0.2 M.

PAs are relatively stable under physiological pH. The molecular weight decreases by about 70% after 3 days at pH = 6.5, and the polymer is completely degraded after 3 days at pH = 5.5 or after 30 min at a pH of 3. For in vivo applications, it is optimal that the CPT be insensitive to environmental changes upon introduction into the body, more specifically, to changes in polymer and/or salt concentration. The CPTs for PAs do not depend strongly on either the salt or polymer concentration, as shown in Figure 7. The latter result indicates that the LCST phase diagram for the polyacetals is flat at the bottom. Such behavior is not uncommon for polymers with a low molecular weight and relatively broad molecular weight distribution (i.e., Mw/Mn = 2 in the present case), for which apparent double critical points have even observed.40 Finally, to be useful biomaterials, the PAs must also be safe for in vivo applications. Preliminary results of biocompatibility tests for PAs by extended exposure to cell cultures (human ACL fibroblasts) indicate that the PAs are not toxic. Figure 8 shows photomicrographs of fibroblasts exposed for 72 h to various concentrations of aqueous PA2222V14.7 solutions. Dead cells would be indicated by a red color in these images. The fact that no red (i.e., dead) cells are observed indicates that the PAs are biocompatible over an extended period of time.

hydroxyl-terminated PAs decrease with apparent molecular weight. End group/molecular weight effects on polymer properties were originally treated by Fox and Flory,38 who showed that the values of a number of colligative properties scaled inversely with the molecular weight. In previous work, we have shown that LCST phase behavior of polymer blends39 is also subject to end group effects. The results presented in Figure 5 suggest that the molecular weight (MW) dependence of the CPT is also due to an end group effect. CPTs scale inversely with MW and extrapolate to a similar infinite MW value for both end groups, as would be expected for an end group dependent property. The CPTs for the series of PA4022 polymers follow the relationship CPT(PA4022EM n) = CPT(PA4022E∞) + K (PA4022E)M n−1

(5)

For the vinyl ether-terminated polyacetals, regression of the data yields CPT(PA4022V∞) = 34.4 ± 0.4 °C and K(PA4022V) = −(2.5 ± 0.1) × 104 °C-Da, while for the hydroxyl-terminated polyacetals, regression of the data yields CPT(PA4022H∞) = 31.7 ± 1.2 °C and K(PA4022H) = (3.4 ± 0.2) × 104 °C-Da. The effect of chain ends (i.e., molecular weight) on the CPT temperature can therefore be quantitatively accounted for and included in the design of a particular polymer for a given application. The fact that the cloud point temperatures follow the Fox−Flory relationship for two different end groups provides additional support for the previous conclusions that the reaction conversion is essentially 100% and that both diol-terminated macromonomers and divinyl ether-terminated macromonomers can be efficiently synthesized with controlled molecular weight. PAs are relatively stable under physiological pH but degrade under acidic conditions to produce the neutral products shown in Figure 1. The degradation rate can be characterized by determining the average molecular weight as a function of degradation time by GPC measurements. GPC-based degradation profiles are shown in Figure 6 for a PA4023V17.3 with a CPT of 42.6 °C. After 3 days at pH = 7.4, the average molecular weight decreases by about 10%, indicating that the

4. CONCLUSIONS The novel family of polyacetals described herein are intrinsically pH-degradable, exhibit highly predictable and tunable LCST behavior in aqueous solutions, and manifest a sharp temperature transition (i.e., breadth of 3−5 °C) with little hysteresis upon heating and cooling. In contrast, poly(Nisopropylacrylamide),41−43 the gold standard of synthetic TRPs, is not biodegradable and is reported to have a broad temperature transition and to exhibit significant hysteresis between heating and cooling cycles.44 Because synthetic TRPs developed to date lack an intrinsic biodegradation mechanism, degradable polymers with LCST behavior have typically been prepared by copolymerizing nondegradable LCST monomers and macromonomers with monomers containing a degradation site, a process that leads to concentration fluctuations that broaden the LCST transition.45 Polyacetals, in contrast, have sharp LCST transitions since copolymerization is not necessary to introduce degradation sites. Moreover, the CPTs of PAs made from mixtures of diols fall on the same lines as those E

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made from a single diol, indicating that the CPT may be predicted and manipulated to high precision simply by using predesignated mixtures of diol reactants. The expected influence of increased concentration fluctuations is evident in PAs prepared from mixtures of diols, as reflected by slightly broader temperature transitions with breadths of the order of 6−8 °C, as can be seen from Figure S.6. The control over the CPT of the polyacetals within a broad temperature range that conveniently brackets body temperature is remarkable. The cloud point temperatures of PAs can be predicted with high precision for essentially any integer or noninteger values of the structural parameters n1, m1, and m2 as well as for any molecular weight. The CPTs of the PAs are tunable over a temperature range of about 6−80 °C. To our knowledge, no other TRP offers such exquisite structural control over the LCST. This range encompasses any temperatures that may be biologically relevant including those of normal (37 °C) and malignant tissue, the latter of which is generally characterized by mild hyperthermia, with temperatures 1−2 °C above that of normal tissue.25 The latter characteristic is important to potential applications in hyperthermia-targeted drug delivery vehicles for cancer where TRPs are designed to fall out of solution when they encounter the elevated temperature inside of a tumor. The exceptional temperature responsiveness of this new family of polyacetals can be attributed to the unique molecular structure of the two monomer cores. The divinyl ether (1) and diol (2) monomers each comprise a hydrophobic part containing n1 or n2 methylene units, respectively, and a hydrophilic part with m1 or m2 ethylene oxide units, respectively. The hydrophilic/hydrophobic balance can be finely tuned by simply adjusting the ratio of methylene to ethylene oxide moieties in either of the two monomers. The PAs are biocompatible and degrade in acidic environments. Acid-catalyzed hydrolysis of PAs produces neutral degradation products, two diols and acetaldehyde, as indicated by the last reaction in Figure 1. In vivo, acetaldehyde is known to be oxidized by aldehyde dehydrogenase enzymes to form acetic acid which leaves the liver and is metabolized by muscle tissue.46,47 In contrast, the degradation products from almost all other degradable polymers are acidic in nature and can cause inflammation.36 Properly designed PAs are therefore expected to biodegrade in the acidic environment encountered within tumors and some intracellular compartments. The pH of tumor microenvironments ranges from 6.5 to 7.226−28 while that within endosomes is reported to be 5−6.5.48−51 The temperature responsiveness and pH degradation of PA materials are therefore well suited to a myriad of potential biological applications, especially in drug delivery for cancer treatment. In summary, the new polyacetals show a number of unique properties and extraordinary temperature response that collectively distinguishes them from any other existing temperature responsive or pH-degradable polymers.

Figure 7. Dependence of the LCST of PA4022V14.9 on (a) polymer concentration without salt and (b) salt concentration at a polymer concentration of 5 g/L.

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ASSOCIATED CONTENT

S Supporting Information *

Figure 8. Fibroblast cell cultures after 72 h exposure to PA2222V14.7 solutions: (a) control, polymer concentration = 0.0 mg/mL; (b) polymer concentration = 0.1 mg/mL; (c) polymer concentration = 1 mg/mL; (d) polymer concentration = 10 mg/mL. Live cells are green while dead cells would be red.

The Supporting Information is available free of charge on the ACS Publications website at DOI: 10.1021/acs.macromol.5b02304. Figures S1−S6 (PDF) F

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AUTHOR INFORMATION

Corresponding Author

*E-mail: [email protected] (J.T.K). Notes

The authors declare no competing financial interest.

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ACKNOWLEDGMENTS The research was supported by the US Army Research Office (Grants W911NF1010184 and W911NF1110372) and by the National Science Foundation (Grant DMR1206191 from the Polymers Program).

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DOI: 10.1021/acs.macromol.5b02304 Macromolecules XXXX, XXX, XXX−XXX