In Vivo Albumin Traps Photosensitizer

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In vivo albumin traps photosensitizer monomers from self-assembled phthalocyanine nanovesicles: A facile and switchable theranostic approach Xingshu Li, Sungsook Yu, Yoonji Lee, Tian Guo, Nahyun Kwon, Dayoung Lee, Su Cheong Yeom, Yejin Cho, Gyoungmi Kim, Jian-Dong Huang, Sun Choi, Ki Taek Nam, and Juyoung Yoon J. Am. Chem. Soc., Just Accepted Manuscript • DOI: 10.1021/jacs.8b12167 • Publication Date (Web): 19 Dec 2018 Downloaded from http://pubs.acs.org on December 19, 2018

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Journal of the American Chemical Society

In Vivo Albumin Traps Photosensitizer Monomers from SelfAssembled Phthalocyanine Nanovesicles: A Facile and Switchable Theranostic Approach Xingshu Li,1,2 Sungsook Yu,3 Yoonji Lee,4 Tian Guo,2 Nahyun Kwon,2 Dayoung Lee,2 Su Cheong Yeom,5 Yejin Cho,3 Gyoungmi Kim,2 Jian-Dong Huang,*,1 Sun Choi,*,4 Ki Taek Nam,*,3 and Juyoung Yoon*,2 1 College

of Chemistry, State Key Laboratory of Photocatalysis on Energy and Environment, Fujian Provincial Key Laboratory of Cancer Metastasis Chemoprevention and Chemotherapy, Fuzhou University, Fuzhou 350108, China. 2 Department

of Chemistry and Nano Science, Ewha Womans University, Seoul 03760, Republic of Korea.

3 Severance

Biomedical Science Institute, Brain Korea 21 PLUS Project for Medical Science, College of Medicine, Yonsei University, Seoul 03760, Republic of Korea. 4 College

of Pharmacy and Graduate School of Pharmaceutical Sciences, Ewha Womans University, Seoul 03760, Republic of Korea. 5 Graduate

School of International Agricultural Technology, Seoul National University, 1447 Pyeongchang-Ro, Daewha, Pyeongchang, Gangwon 25354, Republic of Korea ABSTRACT: Albumin is a promising candidate as a biomarker for potential disease diagnostics and has been extensively used as a drug delivery carrier for decades. In these two directions, many albumin-detecting probes and exogenous albuminbased nanocomposite delivery systems have been developed. However, there are only a few cases demonstrating the specific interactions of exogenous probes with albumin in vivo, and nanocomposite delivery systems usually suffer from tedious fabrication processes and potential toxicity of the complexes. Herein, we demonstrate a facile “one-for-all” switchable nanotheranostic (NanoPcS) for both albumin detection and cancer treatment. In particular, the in vivo specific binding between albumin and PcS, arising from the disassembly of injected NanoPcS, is confirmed using an inducible transgenic mouse system. Fluorescence imaging and antitumor tests on different tumor models suggest that NanoPcS has superior tumor-targeting ability and the potential for time-modulated, activatable photodynamic therapy.

INTRODUCTION Theranostics that have the ability to indicate the expression of biomarkers and simultaneously transport therapeutic agents play a key role in basic biological studies as well as in treatment applications.1-3 With increasing insights into the delivery processes and biological responses, theranostics are guiding the medical field toward a new era of precise treatments.4-6 However, it is still difficult to develop switchable theranostics with a biomarker-driven signal output that can be “turned on” and an on-demand therapeutic delivery/activation that can be fabricated in a facile way. Currently, several switchable strategies, such as molecular beacon,7-10 aggregation-induced emission,11-13 structure-switching aptamer14,15 and supramolecular

approaches,16-21 have been developed to probe biomarkers and deliver therapeutic agents. Upon the specific recognition of target biomarkers through affinity labeling, specific peptide fragment recognition, electrostatic interactions, hydrophobic ligand binding, etc., the diagnostic signals and therapeutic activities could be switched on. Although these theranostic systems permit both diagnosis and treatment by using novel approaches, most of these systems suffer from the requirements of multiple components (e.g. a system requiring dye Cy5.5 for fluorescent signal and Au nanoparticle for photothermal therapy), extra materials (e.g. a non-biodegradable graphene oxide as carrier) and multistep fabrication. In addition, the low concentration of reagent loading causes poor therapeutic outcome and the potential toxicity of

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system complexes may hamper the clinical translation of these “all-in-one” systems.22 Therefore, we herein describe a “one-for-all” switchable strategy based on the dynamic assembly- and disassembly-driven nanotheranostics that are constructed using an inherently multifunctional molecule as the building block. In our system, a versatile phthalocyanine derivative bearing a hydrophilic group is designed as the building block (Fig. 1). This amphiphilic chemical structure facilitates spontaneous assembly to form a uniform nanovesicle dispersion in aqueous solutions. Interestingly, albumin can trap the phthalocyanine molecule from this nanovesicle and induce its disassembly, leading to switchable photoactivity. Thus, our nanovesicle assembly possesses several highly competitive characteristics: (1) it is self-assembled from one pure component without complex processes; (2) the targeted protein-driven “turn on” fluorescent signal permits the potential application of our nanovesicle assembly for in vivo albumin labeling and tumor imaging with relatively high signal-to-background ratios; (3) the in vivo latent albumin-triggered reactive oxygen species (ROS) generation provides a high possibility for time-modulated, activatable photodynamic therapy (PDT) with minimal side effects.

therapeutic agents for PDT, zinc(II) phthalocyanines can generally serve as good imaging agents owing to their ability of fluorescence emission in the far-red and nearinfrared region;25,26 and (3) the hydrophobic phthalocyanine macrocycle  system conjugated with a hydrophilic group makes it possible to control supramolecular formation.27,28 The synthesis details are shown in the Supporting Information. Interestingly, PcS can self-assemble in aqueous solutions to form nearly regular vesicle shapes with a diameter of approximately 15 nm (Fig. 2a, Fig. 2b and Fig. 2c). Considering the size of three-dimensional PcS (Fig. S1), we speculate that the nanovesicle structure (NanoPcS) most likely corresponds to two monolayers of PcS. We also found that NanoPcS shows good stability in dispersions (Fig. 2d and Fig. S2), while zinc(II) phthalocyanine without the substituent of 4-sulfonatophenoxyl, as a control, did not form uniform and stable nanostructures. Superquenching was observed for the photoactivity of NanoPcS in water compared to the PcS monomers in DMSO (Fig. 2e). This self-quenching photoactivity is attractive because it leads to photostability and easy storage. Comparing NanoPcS to the clinically approved molecular dye indocyanine green (ICG) or one of the representative nonquenched zinc(II) phthalocyaninebearing octa-sulphonates (PcS8)29 indicated that NanoPcS has higher photostability (Fig. 2f).

Fig. 1 Schematic illustration of the dynamic PcS assembly and disassembly processes as well as the switchable photoactivities for in vivo fluorescence imaging and tumortargeted PDT.

RESULTS AND DISCUSSION Synthesis of NanoPcS and its self-quenching photoactivity. We first designed a new multifunctional building block consisting of zinc(II) phthalocyanine mono- substituted with 4-sulfonatophenoxyl (PcS, see Fig. 1). PcS was selected because (1) zinc(II) phthalocyanines are high-potential, second-generation photosensitizers for PDT owing to their strong absorption in the phototherapeutic window (650–800 nm) and high efficiency of generating ROS as monomer, and one zinc(II) phthalocyanine derivative, Photocyanine®, is currently under phase II clinical trials;23,24 (2) in addition to being

Fig. 2 Characterization of NanoPcS. a Morphology of NanoPcS determined by transmission electron microscopy (TEM). b The picture of NanoPcS (100 µM) in water, PBS, or RPMI 1640 culture medium after aging for 20 min. c Size distribution of NanoPcS (100 µM) in water, PBS, or RPMI 1640 culture medium after aging for 20 min detected by dynamic light scattering (DLS). d Size distribution of NanoPcS and zinc(II) phthalocyanine (ZnPc) in water (both at 30 µM) after aging for


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Journal of the American Chemical Society different times and then detected by DLS. e Fluorescence spectra (excited at 610 nm) of PcS (in DMSO) and NanoPcS (in water), both at 3 M. F. I.: Fluorescence Intensity. f Photostability of NanoPcS, ICG and PcS8 in water detected by absorption spectra. The samples (all at 3 μM) were continuously irradiated by a light source, which consisted of a halogen lamp, a water tank for cooling, and a colored glass filter with a cut-on wavelength of 610 nm. The power density ( > 610 nm) was 26 mW/cm2.

Albumin traps PcS from NanoPcS, leading to the recovery of photoactivity. Albumin, the most abundant plasma protein, is closely associated with the regulation of plasma colloidal osmotic pressure and the delivery of various endogenous and exogenous substances.30-32 The levels of albumin in body fluids play a crucial role in the progression and development of many diseases, such as sepsis, acute renal failure and liver injury.33-35 In addition, many studies have reported that albumin might be involved in the provision of nutrition to tumors.36-38 Overexpressed albumin receptors (e.g., glycoprotein 60) and albumin-binding protein SPARC (secreted protein, acidic and rich in cysteine) on cancer cells probably facilitate the accumulation and degradation of albumin in tumor tissues.30 Therefore, albumin is a promising candidate as a biomarker for potential disease diagnosis and tumor-specific drug delivery. Inspired by the fact that albumin allows hydrophobic organic anions to bind to the molecule and to be buried within it,39,40 we explored the albumin-responsive properties of NanoPcS. As shown in Fig. 3a and Fig. S3, the absorbance band at 678 nm, which indicates a monomer, increases with the concentration of bovine serum albumin (BSA) or human serum albumin (HSA). In addition, its fluorescence (Fig. 3b and Fig. S4 ) and ROS generation (Fig. 3c) are dramatically recovered upon albumin addition. Size distribution detections indicated that after adding albumin, the mean particle size of NanoPcS was reduced to approximately 6 nm, corresponding to the size of albumin (Fig. 3d). Moreover, the nanovesicle structures disappeared upon albumin addition as detected by TEM (Fig. S5). Based on these results, we propose that albumin trapped the PcS molecule and induced the disassembly of NanoPcS, leading to the signal to “turn on” photoactivity (Fig. S6). To determine the binding mechanism, we first used fluorescence titration analysis to study the binding constant and binding site. The results showed that PcS has a strong binding affinity with albumin with binding constants higher than 107 M-1 and binding stoichiometry of 1:1 (Fig. S7 and Fig. S8). Next, we used molecular modeling to simulate the binding between PcS and HSA. As shown in Fig. S9, the structure of HSA consists of three homologous domains (DI, DII, and DIII), including several pockets for fatty acid binding (FA1-7). The most wellknown drug binding sites are near FA7, where several drugs such as warfarin or phenylbutazone bind, and FA3-4 for the binding of diazepam, ibuprofen, propofol, etc.33 Among the FA binding sites, the largest pocket is the heme binding site (FA1), which is also the binding site of bilirubin. Other FA binding sites seem to be quite small for the

binding of large molecules such as PcS. One more cavity where large molecules can bind is the cleft region between two domains (DI and DIII). To predict the binding mechanism of PcS on HSA, we performed molecular modeling studies. We searched 500 binding conformations per compound using a grid that covers most of the binding cavities in HSA (Fig. S10). The PcS docking study suggested two different binding modes: one at the heme binding site (Fig. 3e) and the other bound to the cleft region (Fig. S11). In the binding mode at the heme site, which is the conformation with the lowest energy score, the flat phthalocyanine-zinc ring nicely fits into the pocket with a Zn atom, coordinating with an oxygen atom of the Y161 residue. The sulfonate group forms ionic interactions with K519. Another binding mode, the most populated conformer, is found in the cleft region, forming a dative bond with D187. The flat phthalocyanine-zinc ring penetrates the cleft region, and the substituent’s sulfonate group participates in the ionic interactions with K190 and R114. These results indicate that albumin efficiently trapped a PcS monomer into its specific pocket from NanoPcS, leading to the disassembly of NanoPcS and the recovery of photoactivity.

Fig. 3 Albumin-responsive properties of NanoPcS and the working mechanisms. Absorption spectra (a) and fluorescence spectra (excited at 610 nm) (b) of NanoPcS (3 M) with different concentrations of BSA in water. c ROS generation of NanoPcS (3 M) in water with and without BSA (30 M) detected by using 2,7-dichlorofluorescin diacetate (12 μM) as a fluorescent probe. d Size distribution of NanoPcS (3 µM) in water before and after adding BSA (30 µM) detected by DLS. BSA in water is used as a control. e Binding mode of PcS at the heme binding site of HSA. The protein surface is colored



to indicate the electrostatic potential, and the bound compound is shown in spheres with the carbon atoms in magenta. In the enlarged view, the compound is displayed in a ball-and-stick model, and the interacting residues are shown as thin sticks with their carbon atoms in light blue. The dative bond between the Zn2+ (shown in purple sphere) and Y161 and the ionic interactions are depicted by black dashed lines.

transgenic mouse system represents a useful method for tracing specific proteins in vivo.

In vivo albumin labeling. Although several fluorescent probes have been used to detect albumin in vitro through specific binding,33,41,42 the in vivo applications or the direct evidence demonstrating the specific binding of exogenous probes with albumin in vivo are limitedly reported.43-45 Therefore, we attempted to study the interaction between NanoPcS and albumin in vivo and identify the specific binding of PcS with albumin using an inducible transgenic mouse system that produces fluorescent albumin in the body. To determine the function of a particular gene product in the given situation, genetic techniques have been developed.46,47 The most well-known reporter mouse line, the Rosa 26R that produces yellow fluorescent protein (YFP) after Cre-mediated excision of a STOP cassette on the expressed Rosa 26 locus,48 and a mouse line with albumin-CreERT2 carrying CreERT2 recombinase sites in the gene of albumin (Fig. S12) were crossed to generate inducible reporter mice (Fig. 4a). CreER recombinases are inactive but can be activated by the estrogen receptor ligand tamoxifen. Experimental mice with a single copy of both the Cre and the reporter were treated by daily intraperitoneal injection three times with vehicle or 1 mg/kg tamoxifen to label albumin with YFP by Cre-loxP recombinase,49,50 and one week later, NanoPcS was administered via the tail vein to observe the binding of albumin-YFP protein to PcS. The expression of albuminYFP with or without Cre recombinase (tamoxifen) was confirmed by an animal optical imaging system in several organs, including the liver, heart, lung, kidney and spleen, after application of NanoPcS. The images were merged with the fluorescent signals of PcS, arising from the disassembly of injected NanoPcS (Fig. S13). After 1 h of NanoPcS treatment, plasma from blood was analyzed by native gel electrophoresis (Fig. 4b), and albumin was detected by an albumin antibody. YFP fluorescence and PcS fluorescence signals were observed at the same position, coinciding with the fact that signals of albumin and PcS appeared at the same position in the gel assay using fetal bovine serum (FBS) and purified BSA (Fig. S14). In confocal imaging using frozen sections (Fig. 4c), a yellow signal was observed in which the fluorescence of PcS and albumin-YFP was merged in the liver and lungs of mice treated with NanoPcS, while the untreated group showed only albumin-YFP fluorescence without PcS fluorescence. PcS fluorescence was also detected in the tissues of mice without the induction of albumin-YFP expression (Fig. S15). This result suggests that albumins trapped PcS from the injected NanoPcS in vivo. These data provide direct evidence supporting the binding of PcS with albumin without interference of the false signal by the use of exogenous probes. The application of the inducible

Fig. 4 Direct evidence indicates the albumin-selective binding of PcS using genetically engineered mice. a Schematic representation of the inducible transgenic mouse system for obtaining mice with YFP-labeled albumin expression. b Gel assays of plasma from mice with expression of albumin-YFP protein treated or untreated with NanoPcS (200 L, 300 M, injection through the tail vein). Plasma was collected from blood collected from the mice at 1 h posttreatment. Lanes 1, 2 and 3 indicate the plasma with protein amounts of 50, 100, 200 g, respectively, on 8% native gel electrophoresis. c Confocal imaging of organ sections from mice with expression of albumin-YFP protein treated or untreated with NanoPcS. The fluorescence of YFP was excited at 510 nm and monitored at 520-550 nm. The fluorescence of PcS was excited at 640 nm and monitored at 650-750 nm. Scale bars are 50 m.

In vivo tumor fluorescence imaging and tracking of PcS delivery. To demonstrate the tumor phototherapeutic utility of NanoPcS, time-dependent biodistribution was evaluated in four tumor models (HepG2, HeLa, SW620 and


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Journal of the American Chemical Society H640 xenograft-bearing mice). Because it shows similar optical spectra to PcS, the clinically used photosensitizer methylene blue (MB) was selected as the control for this test. Compared to PcS, MB did not show strong and stable binding with albumin (Fig. S16). After tail vein injection, MB quickly spread throughout the whole body of tumorbearing mice and then was quickly eliminated from the body without obvious tumor accumulation (Fig. 5 and Fig. S17). In contrast, NanoPcS-injected mice did not show obvious fluorescence signal in the first few hours. However, 24 h after NanoPcS injection, the xenograft tumors were clearly visualized via live imaging with low back-ground noise. The results of fluorescence imaging ex vivo confirmed that PcS, arising from the disassembly of NanoPcS, has a superior tumor-targeting ability. In particular, NanoPcS displayed a time-modulated signal to “turn on” fluorescence, which was probably the consequence of latent albumin-induced trapping and disassembly processes. We also prepared multicellular tumor spheroids from HepG2 cells and used them as an evaluation system to investigate the penetration of NanoPcS. After comparing the fluorescence images of HepG2 spheroids via confocal microscopy z-stack scanning, we observed that both NanoPcS with and without albumin pretreatment in the culture medium displayed excellent penetration into the spheroids, which were better than the zinc(II) phthalocyanine-bearing four 4-sulfonatophenoxy groups (PcS4)51 and MB (Fig. 6 and Fig. S18). The achieved penetration depth of NanoPcS is highly promising for improved PcS delivery to solid tumors after extravasation from the blood vessels. These results indicated that NanoPcS has a unique interaction with endogenous albumin, which in turn likely improves the pharmacokinetic characteristic of PcS and increases its tumor accumulation via both passive targeting (enhanced permeability and retention effect)52 and active targeting (e.g., recognition of glycoprotein 60).30,31

Fig. 5 In/ex vivo fluorescence images of HepG2 xenograftbearing mice after intravenous injection of MB and NanoPcS (same dose: 200 L, 300 M). The red dotted circles indicate tumor sites. H: heart, Lu: lungs, Li: liver, K: kidneys, S: spleen, T: tumor. Fluorescence images were acquired by excitation at 640 nm and monitoring at 750 nm with an IVIS Lumina II imaging system. Color scales indicate the radiant efficiency ( 107 p s1 cm1 sr1 W1 cm2).

Photodynamic antitumor efficacy of NanoPcS. To evaluate the phototherapeutic efficacy of NanoPcS in vivo, HepG2 and HeLa tumor-bearing mice were treated with saline (as a control), only laser irradiation, only NanoPcS or NanoPcS combined with laser irradiation. As shown in Fig. 7a, tumor growth in HepG2 tumor-bearing mice treated with NanoPcS and then with laser irradiation was significantly inhibited compared with that of the control group. However, neither laser irradiation nor NanoPcS treatment induced any therapeutic effect. We also observed the phototherapeutic effect of NanoPcS on HeLa tumor-bearing mice, but its inhibition of tumor growth was lower than that of HepG2 tumors, which may be attributable to the faster proliferation of HeLa cells. In addition, histological analysis of HepG2 and HeLa tumors obtained from mice on the 18th day after treatment confirmed the phototherapeutic outcome of NanoPcS (Fig. 7b and Fig. S19). We believe that future systematic investigations focusing on the optimization of drug dose, drug-light interval, and light dose will further enhance the phototherapeutic efficacy. Finally, to evaluate the biocompatibility of these treatments, mouse lungs, liver, heart, kidneys, and spleen were collected for hematoxylin and eosin staining on the 18th day following treatment. As shown in Fig. 7c, no pathological or other adverse changes were observed in the tissues from the mice treated with NanoPcS or NanoPcS combined with laser irradiation. These results suggest that the facile, switchable approach to tumor treatment is both effective and biocompatible.



Fig. 6 Fluorescence images of HepG2 spheroids after different treatments. HepG2 spheroids were treated with MB, MB + BSA, NanoPcS, NanoPcS + BSA, PcS4 and PcS4 + BSA (photosensitizers: 5 μM, BSA: 50 μM) for 2 h in FBS-free culture media, and fluorescence images were acquired by confocal microscopy z-stack scanning (ex. 635 nm, em. 655755 nm). The surface of the spheroids was defined as 0 μm, and z-stack scanning was performed from the surface to equatorial cross-section. Scale bar: 200 μm.

CONCLUSION In summary, we have developed and successfully proved the concept of a facile, switchable theranostic strategy for the design of nanostructured self-assembly based on an intrinsically multifunctional molecule, PcS. The vesicleshaped nanoassembly displayed albumin-dependent disassembly. Consequently, NanoPcS is an effective theranostic agent for tumor-targeted fluorescence imaging and time-modulated, activatable PDT. In addition, using an inducible transgenic mouse system, we obtained direct evidence to demonstrate the specific interaction of exogenous probes with albumin in vivo.

indicates P < 0.05 compared to the control group. b Histological analysis of HeLa and HepG2 tumors acquired from mice on the 18th day after various treatments as indicated. Apoptosis was tested using the TUNEL assay. Green is a positive signal. Nuclei were visualized using DAPI (blue). c Histological analysis of the organs acquired from mice bearing HepG2 or HeLa tumors on the 18th day after various treatments as indicated.

ASSOCIATED CONTENT Supporting Information The Supporting Information is available free of charge on the ACS Publications website. Detailed materials and methods, synthesis, characterization, and 19 supplementary figures.

AUTHOR INFORMATION Corresponding Authors * [email protected] * [email protected] * [email protected]. * [email protected]

Notes The authors declare no competing ﬁnancial interest.

ACKNOWLEDGMENT J.Y. thanks the National Research Foundation of Korea (NRF), which was funded by the Korea government (MSIP) (No. 2012R1A3A2048814). J.D.H. thanks National Natural Science Foundation of China (Grant Nos. U1705282, 21473033). K.T.N. thanks the Korea Mouse Phenotyping Project (NRF2016M3A9D5A01952416) of the National Research Foundation, the Ministry of Food and Drug Safety (14182MFDS978) and the Brain Korea 21 PLUS Project for Medical Science, Yonsei University. S.C. thanks the National Research Foundation of Korea (NRF) grants, which were funded by the Korea government (MSIT) (No. 2018R1A5A2025286 and NRF2017R1A2B4010084). It was inspired by the international and interdisciplinary environments of the JSPS Asian CORE Program, “Asian Chemical Biology Initiative”.

REFERENCES

Fig. 7 Phototherapeutic efficacy of NanoPcS on mice bearing HepG2 or HeLa tumors. a Tumor growth of mice after various treatments as indicated. Tumor volumes were normalized to their initial values. At 72 h after treatment (same dose: 200 L, 300 M), tumors were laser irradiated at 670 nm (0.2 W/cm2 for 10 min). Data are expressed as the mean ± SEM (n = 5). 

1. Chen, H., Zhang, W., Zhu, G., Xie, J. & Chen, X. Rethinking cancer nanotheranostics. Nat. Rev. Mater. 2, 17024 (2017). 2. Srinivasarao, M., Galliford, C. V. & Low, P. S. Principles in the design of ligand-targeted cancer therapeutics and imaging agents. Nat. Rev. Drug Discovery 14, 203-219 (2015). 3. Weissleder, R. Molecular imaging in cancer. Science 312, 11681171 (2006). 4. Conway, J. R., Carragher, N. O. & Timpson, P. Developments in preclinical cancer imaging: innovating the discovery of therapeutics. Nat. Rev. Cancer 14, 314-328 (2014). 5. Li, Y., Lin, T. Y., Luo, Y., Liu, Q., Xiao, W., Guo, W., Lac, D., Zhang, H., Feng, C., Wachsmann-Hogiu, S., Walton, J. H., Cherry, S. R., Rowland, D. J., Kukis, D., Pan, C. & Lam, K. S. A smart and versatile theranostic nanomedicine platform based on nanoporphyrin. Nat. Commun. 5, 4712 (2014). 6. Li, X., Kim, J., Yoon, J. & Chen, X. Cancer-associated, stimulidriven, turn on theranostics for multimodality imaging and therapy. Adv. Mater. 29, 1606857 (2017).


Page 6 of 9

Page 7 of 9 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

Journal of the American Chemical Society 7. Chen, J., Stefflova, K., Niedre, M. J., Wilson, B. C., Chance, B., Glickson, J. D. & Zheng, G. Protease-triggered photosensitizing beacon based on singlet oxygen quenching and activation. J. Am. Chem. Soc. 126, 11450-11451 (2004). 8. Liu, T. W., Chen, J. & Zheng, G. Peptide-based molecular beacons for cancer imaging and therapy. Amino. Acids. 41, 11231134 (2011). 9. Li, X., Mu, J., Liu, F., Tan, E. W. P., Khezri, B., Webster, R. D., Yeow, E. K. L. & Xing, B. Human transport protein carrier for controlled photoactivation of antitumor prodrug and real-time intracellular tumor imaging. Bioconjugate. Chem. 26, 955-961 (2015). 10. Hu, J. -J., Liu, L. -H., Li, Z. -Y., Zhou, R. -X. & Zhang, X. -Z. MMP-responsive theranostic nanoplatform based on mesoporous silica nanoparticles for tumor imaging and targeted drug delivery. J. Mater. Chem. B 4, 1932-1940 (2016). 11. Yuan, Y., Kwok, R. T. K., Tang, B. Z. & Liu, B. Targeted theranostic platinum(IV) prodrug with a built-in aggregationinduced emission light-up apoptosis sensor for noninvasive early evaluation of its therapeutic responses in situ. J. Am. Chem. Soc. 136, 2546-2554 (2014). 12. Yuan, Y., Zhang, C. J., Gao, M., Zhang, R., Tang, B. Z. & Liu, B. Specific light-up bioprobe with aggregattoin-induced emission and activatable photoactivity for the targeted and image-guided photodynamic ablation of cancer cells. Angew. Chem. Int. Ed. Engl. 54, 1780-1786 (2015). 13. Shin, W. S., Lee, M. -G., Verwilst, P., Lee, J. H., Chi, S. -G. & Kim, J. S. Mitochondria-targeted aggregation indeuced emission theranostics: crucial importance of in situ activation. Chem. Sci. 7, 6050-6059 (2016). 14. Liu, K., Lin, B. & Lan, X. Aptamers: a promising tool for cancer imaging, diagnosis, and therapy. J. Cell. Biochem. 114, 250255 (2013). 15. Wang, Y. M., Wu, Z., Liu, S. J. & Chu, X. Structure-switching aptamer triggering hybridization chain reaction on the cell surface for activatable theranostics. Anal. Chem. 87, 6470-6474 (2015). 16. Li, S., Zou, Q., Li, Y., Yuan, C., Xing, R. & Yan, X. Smart peptide-based supramolecular photodynamic matollo-nanodrugs designed by multicomponent coordination self-assembly. J. Am. Chem. Soc. 140, 10794-10802 (2018). 17. Gao, J., Li, J., Geng, W. -C., Chen, F. -Y., Duan, X., Zheng, Z., Ding, D. & Guo, D. -S. Biomarker displacement activation: A general host-guest strategy for targeted phototheranostics in vivo. J. Am. Chem. Soc. 140, 4945-4953 (2018). 18. Li, X., Yu, S., Lee, D., Kim, G., Lee, B., Cho, Y., Zheng, B. -Y., Ke, M. -R., Huang, J. -D., Nam, K. T., Chen, X. & Yoon, J. ACS Nano 12, 681-688 (2018). 19. Huang, Y., Song, C., Li, H., Zhang, R., Jiang, R., Liu, X., Zhang, G., Fan, Q., Wang, L. & Huang, W. Cationic conjugated polymer/hyaluronan-doxorubicin complex for sensitive fluorescence detection of hyaluronidase and tumor-targeting drug delivery and imaging. ACS Appl. Mater. Interfaces 7, 2152921537 (2015). 20. Gao, S., Zhang, L., Wang, G., Yang, K., Chen, M., Tian, R., Ma, Q. & Zhu, L. Hybrid graphene/Au activatable theranostic agent for multimodalities imaging guided enhanced photothermal therapy. Biomaterials 79, 36-45 (2016). 21. Li, X., Zheng, B. -Y., Ke, M. -R., Zhang, Y., Huang, J. -D. & Yoon, J. A tumor-pH-responsive supramolecular photosensitizer for activatable photodynamic therapy with minimal in vivo skin phototoxicity. Theranostics 7, 2746-2756 (2017). 22. Huynh, E. & Zheng, G. Engineering multifunctional nanoparticles: all-in-one versus one-for-all. Wiley. Interdiscip. Rev. Nanomed. Nanobiotechnol. 5, 250-265 (2013). 23. Li, X., Zheng, B. -D., Peng, X. -H., Li, S. -Z., Ying, J. -W., Zhao, Y., Huang, J. -D. & Yoon, J. Phthalocyanines as medicinal

photosensitizers: Developments in the last five years. Coord. Chem. Rev. https://doi.org/10.1016/j.ccr.2017.08.003. 24. Li, X., Kolemen, S., Yoon, J. & Akkaya, E. U. Activatable photosensitizers: agents for selective photodynamic Ttherapy. Adv. Funct. Mater. 27, 1604053 (2017). 25. Josefsen, L. B. & Boyle, R. W. Unique diagnostic and therapeutic roles of porphyrins and phthalocyanines in photodynamic therapy, imaging and theranostics. Theranostics 2, 916-966 (2012). 26. Zhang, Y. & Lovell, J. F. Recent applications of phthalocyanines and naphthalocyanines for imaging and therapy. Wiley. Interdiscip. Rev. Nanomed. Nanobiotechnol. 2017, doi: 10.1002/wnan.1420. 27. Li, X., Lee, S. & Yoon, J. Supramolecular photosensitizers rejuvenate photodynamic therapy. Chem. Soc. Rev. 47, 1174-1188 (2018). 28. Li, X., Kim, C. -y., Lee, S., Lee, D., Chung, H. -M., Kim, G., Heo, S. -H., Kim, C., Hong, K. -S. & Yoon, J. Nanostructured phthalocyanine assemblies with protein-driven switchable photoactivities for biophotonic imaging and therapy. J. Am. Chem. Soc. 139, 10880-10886 (2017). 29. Li, X. -S., Ke, M. -R., Zhang, M. -F., Tang, Q. -Q., Zheng, B. -Y. & Huang, J. -D. A non-aggregated and tumour-associated macrophage-targeted photosensitizer for photodynamic therapy: a novel zine(II) phthalocyanine containing octa-sulphonates. Chem. Commun. 51, 4704-4707 (2015). 30. Liu, Z. & Chen, X. Simple bioconjugate chemistry serves great clinical advances: albumin as a versatile platform for diagnosis and precision therapy. Chem. Soc. Rev. 45, 1432-1456 (2016). 31. Chen, Q. & Liu, Z. Albumin carriers for cancer theranostics: A conventional platform with new promise. Adv. Mater. 28, 1055710566 (2016). 32. Liu, H., Moynihan, K. D., Zheng, Y., Szeto, G. L., Li, A. V., Huang, B., Van Egeren, D. S., Park, C. & Irvine, D. J. Structurebased programming of lymph-node targeting in molecular vaccines. Nature 507, 519-522 (2014). 33. Wang, Y. -R., Feng, L., Xu, L., Li, Y., Wang, D. -D., Hou, J., Zhou, K., Jin, Q., Ge, G. -B., Cui, J. -N. & Yang, L. A rapid-response fluorescent probe for the sensitive and selective detection of human albumin in plasma and cell culture supernatants. Chem. Commun. 52, 6064-6067 (2016). 34. Oettl, K., Stadlbauer, V., Petter, F., Greilberger, J., PutzBankuti, C., Hallström, S., Lackner, C. & Stauber, R. E. Oxidative damage of albumin in advanced liver disease. Biochim. Biophys. Acta. 1782, 469-473 (2008). 35. de Zeeuw, D., Parving, H. H. & Henning R. H. Miroalbuminuria as an early marker for cardiovascular disease. J. Am. Soc. Nephrol. 17, 21002105 (2006). 36. Stehle, G., Sinn, H., Wunder, A., Schrenk, H. H., Stewart, J. C. M., Hartung, G., Maier-Borst, W. & Heene, D. L. Plasma protein (albumin) catabolism by the tumor itself-implications for tumor metabolism and the genesis of cachexia. Crit. Rev. Oncol. Hematol. 26, 77-100 (1997). 37. Davidson, S. M., Jonas, O., Keibler, M. A., Hou, H. W., Luengo, A., Mayers, J. R., Wyckoff, J., Del Rosario, A. M., Whitman, M., Chin, C. R., Condon, K. J., Lammers, A., Kellersberger, K. A., Stall, B. K., Stephanopoulos, G., Bar-Sagi, D., Han, J., Rabinowitz, J. D., Cima, M. J., Langer, R. & Vander Heiden, M. G. Direct evidence for cancer-cell-autonomous extracellular protein catabolism in pancreatic tumors. Nat Med. 23, 235-241 (2017). 38. Hoogenboezem, E. N. & Duvall, C. L. Harnessing albumin as a carrier for cancer therapies. Adv. Drug Deliv. Rev.. 130, 73-89 (2018). 39. Zhu, G., Lynn, G. M., Jacobson, O., Chen, K., Liu, Y., Zhang, H., Ma, Y., Zhang, F., Tian, R., Ni, Q., Cheng, S., Wang, Z., Lu, N.,



Yung, B. C., Wang, Z., Lang, L., Fu, X., Jin, A., Weiss, I. D., Vishwasrao, H., Niu, G., Shroff, H., Klinman, D. M., Seder, R. A. & Chen, X. Albumin/vaccine nanocomplexes that assemble in vivo for combination cancer immunotherapy. Nat. Commun. 8, 1954 (2017). 40. Simard, J. R., Zunszain, P. A., Ha, C. E., Yang, J. S., Bhagavan, N. V., Petitpas, I., Curry, S. & Hamilton, J. A. Locating high-affinity fatty acid-binding sites on albumin by x-ray crystallography andNMR spectroscopy. Proc. Natl. Acad. Sci. USA 102, 17958-17963 (2005). 41. Fan, X., He, Q., Sun, S., Li, H., Pei, Y. & Xu, Y. Nanoparticles self-assembled from multiple interactions: a novel near-infrared fluorescent sensor for the detection of serum albumin in human sera and turn-on live-cell imaging. Chem. Commun. 52, 1178-1181 (2016). 42. Dey, G., Gaur, P., Giri, R. & Ghosh, S. Optical signaling in biofluids: a nondenaturing photostable molecular probe for serum albumins. Chem. Commun. 52, 1887-1890 (2016). 43. Schniering, J., Borgna, F., Siwowska, K., Benešová, M., Cohrs, S., Hasler, R., van der Meulen, N. P., Maurer, B., Schibili, R. & Müller, C. In vivo labeling of plasma proteins for imaging of enhanced vascular permeability in the lungs. Mol. Pharm. 15, 4995-5004 (2018). 44. Hahnenkamp, A., Alsibai, W., Bremer, C. & Höltke, C. Optimizing the bioavailability of small molecular optical imaging probes by conjugation to an albumin affinity tag. J. Control. Release 186, 32-40 (2014).

45. Fischer, C. R., Groehn, V., Reber, J., Schibli, R., Ametamey, S. M. & Müller, C. Improved PET imaging of tumors in mice using a nover (18)F-folate conjugate with an albumin-binding entity. Mol. Imaging Biol. 15, 649-654 (2013). 46. Branda, C. S. & Dymecki, S. M. Talking about a revolution: the impact of site-specific recombinases on genetic analyses in mice. Dev. Cell 6, 7-28 (2004). 47. Feil, R. Conditional somatic mutagenesis in the mouse using site-specific recombinases. Handb. Exp. Pharmacol. 178, 328 (2007). 48. Soriano P. Generalized lacZ expression with the ROSA26 Cre reporter strain. Nat. Genet. 21, 70-71 (1999). 49. Reinert, R. B., Kantz, J., Misfeldt, A. A., Poffenberger, G., Gannon, M., Brissova, M. & Powers, A. C. Tamoxifen-induced CreloxP recombination is prolonged in pancreatic islets of adult mice. PLoS One. 7, e33529 (2012). 50. Falke, L. L., Broekhuizen, R., Huitema, A., Maarseveen, E., Nguyen, T. Q. & Goldschmeding, R. Tamoxifen for induction of Cre-recombination may confound fibrosis studies in female mice. J. Cell Commun. Signal. 11, 205-211 (2017). 51. Li, X. -S., Ke, M. -R., Huang, W., Ye, C. -H. & Huang, J. -D. A pH-responsive layered double hydroxide (LDH)–phthalocyanine nanohybrid for efficient photodynamic therapy. Chem. Eur. J. 21, 3310-3317 (2015). 52. Maeda, H. Tumor-selective delivery of macromolecular drugs via the EPR effect: Backgroung and future prospects. Bioconjugate Chem. 21, 797-802 (2010).


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