Erythrocyte-Cancer Hybrid Membrane Camouflaged Hollow Copper

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Erythrocyte-Cancer Hybrid Membrane Camouflaged Hollow Copper Sulfide Nanoparticles for Prolonged Circulation Life and Homotypic-Targeting Photothermal/Chemotherapy of Melanoma Dongdong Wang, HaiFeng Dong, Meng Li, Yu Cao, Fan Yang, Kai Zhang, Wenhao Dai, Changtao Wang, and Xueji Zhang ACS Nano, Just Accepted Manuscript • DOI: 10.1021/acsnano.7b08355 • Publication Date (Web): 25 May 2018 Downloaded from http://pubs.acs.org on May 25, 2018

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Erythrocyte-Cancer Hybrid Membrane Camouflaged Hollow Copper Sulfide Nanoparticles for Prolonged Circulation Life and Homotypic-Targeting Photothermal/Chemotherapy of Melanoma Dongdong Wang,† Haifeng Dong,*, † Meng Li,‡ Yu Cao,† Fan Yang,† Kai Zhang,† Wenhao Dai,† Changtao Wang‡, Xueji Zhang.*, † †

Beijing Key Laboratory for Bioengineering and Sensing Technology, Research Center for

Bioengineering and Sensing Technology, School of Chemistry and Bioengineering, University of Science & Technology Beijing, Beijing 100083, P. R. China. ‡

Beijing Key Lab of Plant Resource Research and Development, School of Science, Beijing Technology and Business University, Beijing 100048, P. R. China.

Haifeng Dong*: E-mail: [email protected] Xueji Zhang*: E-mail: [email protected] KEYWORDS. cellular hybrid membrane, hollow copper sulfide nanoparticles, prolonged circulation lifetime, homotypic-targeting, synergistic photothermal/chemotherapy

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ABSTRACT. Cellular membrane-coated nanoparticles have increasingly been pursued to leverage the nature cell functions for enhancing biocompatibility and improved therapeutic efficacy. Taking advantage of specialized cells membrane or combining functions from different membrane types facilitate to strengthen their functionality. Herein, we fuse membrane materials derived from red blood cells (RBCs) and melanoma cells (B16-F10 cells) to create a hybrid biomimetic coating (RBC-B16), and RBC-B16 hybrid membrane camouflaged doxorubicin (DOX)-loaded hollow copper sulfide nanoparticles (DCuS@[RBC-B16] NPs) are fabricated for combination therapy of melanoma. The DCuS@[RBC-B16] NPs are comprehensively characterized, showing the inherent properties of the both source cells. Compared to the bare CuS NPs, the DCuS@[RBC-B16] NPs exhibit highly specific self-recognition to source cell line in vitro, and achieve markedly prolonged circulation lifetime and enhanced homogenous targeting abilities in vivo inherited from the source cells. Thus, the DOX-loaded [RBC-B16]coated CuS NP platform exhibits excellent synergistic photothermal/chemotherapy with about 100 % melanoma tumor growth inhibition (TGI) rate. The reported strategy may contribute to personalized nanomedicine of various tumors by combining the RBCs with homotypic cancer membrane accordingly on the surface of nanoparticle.


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Nanoparticles have gained widespread attention as probes or drug delivery vehicles toward personalized nanomedicine.1-4 A primary directive of nanotechnology is to design nano-delivery platforms with excellent therapeutic efficacy that can evade the immune system and cross various biological barriers and enable specific targeting to tumors.5-9 For example, the photothermal therapy (PTT) utilizing photothermal conversion agents (PTCAs) can convert optical energy into thermal energy to preferentially achieve tumor cells localized thermal ablation without affecting healthy tissue. However, prolonging the circulation lifetime and enhancing the tumor targeting of PTCAs to maximize the delivery and accumulate to tumor tissues in vivo are subject to a number of biological barriers that limit their applications.6,10-14 For the chemotherapy, encapsulating free drug in nanoparticles is advantageous, but avoiding opsonization and non-specific clearance is greatly challenged.15-18 However, the conversional chemical ligand-related surface modification approach is highly restricted by the limited effects and complicated chemical preparation. Recently, cell membrane-coated approaches have emerged as a way to endow the nanoparticle with extraordinary bio-interfacing ability.19-22 By combining the synthetic nanoparticles with natural or biomimetic materials, the resulting cell membrane coated nanoparticles preserve the highly controllable physicochemical properties of synthetic nanoparticle core while harnessing the versatile and complex cellular membrane functions that are hardly replicated by tailor-made materials.7,23-25 It could escape the clearance of immune system or promote specific targeting ability to the tumor cells.5,26-28 For examples, red blood cells (RBCs) membrane coating, also known as erythrocytes, significantly improves the immune evading capability of coated nanoparticles,29,30 stem cell-derived nanoparticles present good cancer targeting properties,15,31 macrophage cell membrane camouflaged nanoparticles could reduce opsonization and prolong


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circulation lifetime,32 leukocyte membrane cloaked nanoparticles possess the ability to traverse endothelium16 and cancer cell membrane coated nanoparticles present the excellent selfrecognition internalization by the source cancer cell lines.33,34 More recently, Zhang et al. reported approach for increasing the nanoparticle functionality by fusing of natural cell membranes from erythrocyte and platelet,35 representing a facile and effective method to produce new multi-membrane nanoparticle platforms with increasingly complex tasks. It is often desirable to introduce specialized functionality for specific application. However, to the best of our knowledge, it has not been explored so far by fusing membrane materials derived from RBCs and cancer cells for enhanced nanoparticle functionalization. Herein, we report on an approach for fusing RBCs and B16-F10 cells membrane to create a RBC-B16 hybrid biomimetic coating, and successfully coat them onto doxorubicin (DOX)loaded hollow copper sulfide nanoparticles (DCuS@[RBC-B16] NPs) for combination photothermal/chemotherapy of melanoma (Scheme 1A). The DCuS@[RBC-B16] NPs were thoroughly characterized, and it is shown that they preserved characteristic functionality derived from both of the source cells. The RBCs membrane coating can significantly improve the immune evading capability of coated DCuS since the surface makeup comprising a mass of “self-marker” protein inhibiting macrophage phagocytosis,5,10 while the B16 membrane coating enable enhanced melanoma homogenous targeting abilities due to the cancer cells expressing surface adhesion molecules with homologous adhesion domains.23,24 In comparison with the bare CuS NPs, the DCuS@[RBC-B16] NPs exhibited much higher specific self-recognition to B16F10 cell line in vitro, markedly prolonged circulation life and enhanced homogenous targeting abilities in vivo (Scheme 1B). The excellent immune evading and homogenous targeting tumor abilities combined with the high loading efficiency of DOX and inherent photothermal


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conversion property of CuS led to outstanding performance of DCuS@[RBC-B16] in synergistic photothermal/chemotherapy of melanoma in vivo. Scheme 1. (A) Schematic of membrane fusion and coating. Membrane materials are derived from RBCs and B16F10 cells and then fused together. The resulting hybrid membrane is used to camouflage DOX-loaded hollow copper sulfide

nanoparticles

(DCuS

NPs)

to

produce

DCuS@[RBC-B16]

NPs.

(B)

The

synergistic

photothermal/chemotherapy of melanoma.

RESULTS AND DISCUSSION RBC-B16 Hybrid Membrane Characterization. To demonstrate the fusion, a Förster resonance

energy

transfer

(FRET)

pair

dyes

of

1,1'-dioctadecyl-3,3,3',3'-

tetramethylindodicarbocyanine, 4-chlorobenzenesulfonate salt (DiD) and 1,1'-dioctadecyl3,3,3',3'-tetramethylindocarbocyanine perchlorate (DiI) were employed to label the B16-F10 cell membrane (Figure S1). The RBC membrane was increasingly added into the two dye-doped B16-F10 cell membrane for fusion. As observed in Figure 1A, with the amounts of RBC membrane increasing, there was a recovery of fluorescence at 565 nm, while the fluorescence at


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670 nm gradually decreased (inset in Figure 1A). It suggested the weakening of the FRET interaction in the original B16-F10 cell membrane due to the interspersing of the two membrane materials. Furthermore, SDS-PAGE protein analysis toward a series of membrane protein markers indicated the well retention of characteristic protein inherited from the B16-F10 membrane and RBC membrane in the protein profile of RBC-B16 hybrid membrane (Figure 1B). Western blotting analysis for analyzing specific protein markers was carried out (Figure 1C). CD47, an immunomodulatory protein responsible for inhibiting macrophage uptake that is expressed by RBC, was revealed on RBC membrane, RBC-B16 hybrid membrane and CuS@[RBC-B16] NPs. Meanwhile, gp100, a type of known membrane-bound tumor-associated antigen expressed on B16-F10 membrane, was found on RBC-B16 hybrid membrane and CuS@[RBC-B16] NPs. Additionally, immunogold staining and transmission electron microscope (TEM) imaging analysis provided visual evidence that RBC-B16 hybrid membrane and CuS@[RBC-B16] NPs simultaneously presented both RBC and B16-F10 membrane markers (Figure 1D), confirming formation of a homogenous mixture. Using a 1:1 protein weight ratio of B16-F10 cell membrane labeled with 3,3'-dioctadecyloxacarbocyanine, perchlorate (DiO): RBC membrane labeled with DiI as precursors, a RBC-B16 hybrid membrane was fabricated, and the resultant materials was coated onto the synthesized CuS NPs. Significant colocation of fluorescent signals derived from the DiI and DiO was observed for the CuS@[RBC-B16] NPs (Figure 1E). In stark contrast, the mixture of CuS@RBC NPs and CuS@B16 NPs constructed with the individual dye-labeled membrane showed the distinct red and green light spots (Figure 1F). These results suggested the successful fusion of the two types of natural cell membranes and achieving incorporation the hybrid membrane onto the CuS NPs.


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Figure 1. (A) B16-F10 membrane doped with DiD and DiI and mixed with increasing amounts of RBC membrane. The fluorescence recovery of the donor (DiI) was used to monitor the increasing amounts of fusion. (B) SDS-PAGE protein analysis of retention protein (1: marker. 2: RBC, 3: B16-F10, 4: RBC-B16 membrane, and 5: CuS@[RBCB16] NPs). (C) Western blot analysis of RBC, B16-F10, RBC-B16 membrane, and CuS@[RBC-B16] NPs for characteristic RBC membrane markers CD47, and characteristic B16-F10 membrane markers gp100. (D) Immunogold TEM images of B16-F10, RBC, RBC-B16 membrane, and CuS@[RBC-B16] NPs samples probed for CD47 (red arrows, large gold) and gp100 (yellow arrows, small gold), followed by negative staining with uranyl acetate (scale bars = 100 nm). Confocal fluorescent microscopy images of the CuS@[RBC-B16] NPs (E), and a mixture of CuS@RBC NPs and CuS@B16 NPs (F) (red = RBC membrane, green = B16-F10 membrane; scale bar = 5 µm).

Physicochemical Characterization of CuS@[RBC-B16] NPs. The TEM image of CuS@[RBC-B16] NPs (Figure 2B) and negatively stained with uranyl acetate (Figure 2C) depicted a characteristic core-shell structure with a hollow core and a uniform outer membrane shell compared to the TEM image of CuS NPs (Figure 2A) and negatively stained with uranyl acetate (Figure S2A). The average size of CuS@[RBC-B16] NPs was about 200 nm, and the thickness of the outer membrane shell was around 8 nm (Figure 2B, inset). Obvious highly


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parallel and ordered lattice fringe spacing of 0.306 nm and 0.281 nm, corresponding to the (102) and (103) faces of CuS crystals were observed from the High-resolution TEM (HRTEM) of CuS@[RBC-B16] NPs (Figure 2D). The nitrogen adsorption-desorption measurements showed the surface area, pore volume, and average pore diameter of CuS NPs was 22.837 m2/g, 0.059 cm3/g, and 4.2 nm, respectively, which validated the mesoporous nature of CuS NPs (Figure S2B, S2C). Dynamic light scattering (DLS) was employed to compare the CuS NPs and CuS@[RBC-B16] NPs. As shown in Figure 2E, the bare CuS NPs exhibited an original size of about 190 nm, increased uniformly by ≈ 20 nm after coating RBC-B16 membrane, consisting the size results (Figure 2B). Additionally, the surface zeta potential changed from -16 mV to -23 mV after the CuS NPs coated with RBC-B16 membrane (Figure 2F), indicating the shielding of negative CuS NPs by the more negative outer membrane surface. Next, the colloidal stability of CuS@[RBC-B16] NPs in phosphate buffered solution (PBS) was measured by DLS, negligible change of particle size was observed over a span of 72 h (Figure S3), which indicated the good colloidal stability of the CuS@[RBC-B16] NPs.


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Figure 2. (A) TEM image of CuS NPs and (B) CuS@[RBC-B16] NPs (scale bar = 200 nm), Inset: the magnification of the marked square (scale bar = 20 nm). (C) TEM images of CuS@[RBC-B16] NPs negatively stained with uranyl acetate (scale bar = 150 nm). (D) HRTEM image of CuS@[RBC-B16] NPs (scale bar = 5 nm). (E) Z-average size of bare CuS and CuS@[RBC-B16] NPs. (F) Surface zeta potential of CuS and CuS@[RBC-B16] NPs.

In Vitro Photothermal Properties of CuS@[RBC-B16] NPs. The synthesized hollow CuS NPs presented a strong photo-absorption at around 1064 nm, and the absorption of CuS@[RBCB16] NPs has barely altered (Figure S2D). The photothermal conversion experiments were performed to investigate the photothermal effect of CuS@[RBC-B16] NPs. The CuS, CuS@[RBC-B16], DCuS@[RBC-B16] NPs presented the similar temperature profiles, which suggested the negligible impact of the membrane coating and DOX loading to the photothermal effect of CuS (Figure 3A). The temperature increase of CuS@[RBC-B16] NPs presented a concentration-dependent pattern (Figure S4). The temperature of CuS@[RBC-B16] NPs increased rapidly to 75.4oC during the NIR-laser irradiation (power density of 1 W/cm2) for 10 min, while temperature of water free of nanoparticles only increased by about 13oC (Figure 3B). The advanced temperature rise could contribute to destroying the cancer cells due to the hyperthermia.27 Furthermore, it found that CuS@[RBC-B16] NPs remained a robust photothermal agent after four cycles of 1064 nm NIR laser irradiation, indicating superb photostability of CuS@[RBC-B16] NPs (Figure 3C). The PA mapping of CuS@[RBC-B16] NPs at various concentrations was also observed (Figure 3D), which exhibited a concentrationdependent pattern, indicating the potential of CuS@[RBC-B16] NPs for PA imaging.


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Figure 3. (A) Temperature increases of water, CuS, CuS@[RBC-B16], and DCuS@[RBC-B16] NPs (50 µg/mL). (B) Infrared thermal imaging of water and CuS@[RBC-B16] (100 µg/mL) during 1064 nm NIR laser irradiation (1 W/cm2) for 10 min. (C) Photostability of CuS@[RBC-B16] (100 µg/mL) under 1064 nm laser irradiation at (1 W/cm2). (D) PA mapping of CuS@[RBC-B16] NPs at various concentrations.

In Vitro DOX Loading of CuS@[RBC-B16] and Releasing. The CuS NPs were expected to be ideal carriers to delivery antitumor drugs owing to the hollow mesoporous structure. The DOX was chosen as antitumor drug model to be loaded into the CuS@[RBC-B16] NPs. with The loading capacity (LC) of DOX was extremely high, reaching up to 87.7 % in weight (Figure 4A), and the highest DOX loading efficiency (LE) is 95.5 % (Figure S5). The high drug loading efficiency was assigned to huge cavity and numerous mesopores as well as the electrostatic interaction between negative CuS NPs and positively charged DOX. The release of DOX from DCuS@[RBC-B16] NPs revealed a typical photo-effect (Figure 4B), and the pH also influenced


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the release of DOX. The pH-related release profile phenomenon was probably caused by the fact that the increased solubility of DOX at weak acidic environment, which enhanced the release of DOX absorbed on membrane surface. The DOX release rate at pH 7.4 was only 4.6 %, and at pH 5.0 was 10.3 % without NIR laser irradiating within 10 h. Conversely, in the presence of laser irradiation, the DOX release at pH 7.4 was increased to 64.2 %. Notably, the release rate was further increased to 75.2 % at pH 5.0. The phenomenon was probably caused by the fact that the increased solubility at weak acidic environment and the local hyperthermia generated by photothermal effect of CuS NPs, which could destroy of the outer cell membrane and enhance the diffusion of drug from the hollow interior in to surrounding medium.27,30 The cellular uptake of DCuS@[RBC-B16] NPs was explored on the B16-F10 cells. As control, the free DCuS NPs was added to B16-F10 cells and cultured in similar condition (Figure 4C). The DCuS@[RBC-B16] NPs exhibited much higher DOX fluorescence compared to the slight fluorescence intensity of DCuS NPs group, indicating the outer cell membrane significant improved the cellular internalization of DCuS NPs due to the retaining cell membranes preserved homotypic targeting ability.27 As expected, after exposure to1064 nm NIR laser irradiation for 5 min, the DCuS@[RBC-B16] NPs presented a markedly increased red fluorescence because of local hyperthermia generated from CuS, destroying of the outer cell membrane shelling and improving drug release from DCuS@[RBC-B16] NPs.


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Figure 4. (A) DOX loading capacity of CuS@[RBC-B16] when the DOX concentration was 200, 400, 600, 800, and 1000 µg/mL, respectively. (B) DOX release from DCuS@[RBC-B16] at pH 7.4 and 5.0 without or with NIR laser irradiation (1064 nm, 1.0 W/cm2). (C) Confocal fluorescent microscopy images of B16-F10 cells cultured with 1: DCuS, 2: DCuS@[RBC-B16], 3: DCuS@[RBC-B16] without or with NIR laser irradiation (1064 nm, 1.0 W/cm2) (scale bar = 50 µm).

B16-F10 Cell Self-Recognition Capability of CuS@[RBC-B16] NPs. The much stronger red fluorescence derived from DiI dyed CuS@B16 NPs in B16-F10 compared to A549 cells, and the flow cytometry revealed that the B16-F10 cells exhibited approximated to 8.1 folds higher mean fluorescence intensity (MFI) inside cells than that of A549 cells (Figure S6B). These results demonstrated the CuS@B16 NPs were preferentially internalized by B16-F10 over A549 cells (Figure S6A). The MFI in CuS@RBC-treated B16-F10 cells was much lower than that of CuS@B16 NPs-treated B16-F10 cells (Figure S6B, C), indicating preferable internalization of CuS@B16 NPs by B16-F10 cells. Furthermore, CuS@RBC-treated B16-F10 cells and CuS@RBC-treated A549 cells exhibited similar MFI value, which suggested B16-F10 cells and


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A549 cells possessed similar uptake efficiency. It excluded the possibility that B16-F10 cells uptake whatever nanoparticle at higher efficiency than A549 cells and indicated the preferable internalization of CuS@B16 by B16-F10 was as a result of B16 cell membrane-mediated homogenous targeting ability. To prove the assumption that the RBC-B16 coating would present cancer cell self-recognition by homotypic cancer cells, cellular uptake of CuS@[RBC-B16] NPs with three cell lines including Human HT1080 fibrosarcoma cells (HT1080 cells), normal human skin fibroblasts (NHDF cells) and B16-F10 was studied. The fluorescence intensity originated from CuS@[RBC-B16] NPs in B16-F10 and was stark superior to that in HT1080, NHDF (Figure 5A), which was approximated to 8.25 and 9.78 folds higher in terms of the mean fluorescence intensity, respectively. The specific self-recognition ability of CuS@[RBC-B16] NPs was further explored on co-incubated B16-F10/A549 cell lines. The B16-F10 cells with DiO (green fluorescence)-dyed membrane were co-cultured with A549 cells together (Figure 5B). After the DiI-dyed CuS@[RBC-B16] NPs (red fluorescence) were added to the cells and incubated for 4 h, amazing outcome was found that the red fluorescence originating from the B16-F10 cells was far stronger compared to barely visible red fluorescence was observed for the A549 cells (in the yellow square). The flow cytometry was employed to investigate the specificity of CuS@[RBC-B16] NPs to target homologous B16-F10 cells. It demonstrated that the B16-F10 cells exhibited approximated to 7-9 fold higher mean fluorescence intensity inside cells than that of other cells, indicating the highly specific self-recognition to B16-F10 cells (Figure 5C, D). These results indicated the internalization of CuS@[RBC-B16] by heterotypic cells was minimal during the observation period, while preserved highly specific self-targeting adhesive interaction to source cells.


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Figure 5. (A) Confocal fluorescent microscopy images of HT1080 cells, NHDF cells, and B16-F10 cells cultured with DiI dyed CuS@[RBC-B16] NPs and (B) dye-labeled CuS@[RBC-B16] NPs after incubation with co-incubated B16-F10/A549 cell lines (red = CuS@[RBC-B16] NPs, green = B16-F10 cells membrane, blue = nucleus; scale bar = 10 µm). (C) Flow cytometric profiles and (D) mean fluorescence intensity of the four cell lines B16-F10, HT1080, NHDF, A549 upon 4 h incubation with DiI dyed CuS@[RBC-B16] NPs.

In Vitro Anticancer Performance. The in vitro anticancer activity of DCuS@[RBC-B16] was evaluated using calcein-AM/propidium iodide (PI) (live/dead) staining coupling with MTT assay. As shown in Figure 6A, neither CuS@[RBC-B16] nor 1064 nm laser irradiation has significant cytotoxicity against B16-F10 cells. For DCuS@[RBC-B16], just about 24.2 % cells were killed without the NIR irradiation, while more than 94.5 % cells were dead for the


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DCuS@[RBC-B16]-treated B16-F10 cells under NIR irradiation. It was more effective than the DOX combined with or without NIR-irradiation (with cell viability around 65 %) or CuS@[RBC-B16] NPs under NIR irradiation (with cell viability around 12.3 %). The live/dead staining analysis was consistent with the MTT results (Figure 6B). Upon NIR irradiation, almost B16-F10 cells treated with DCuS@[RBC-B16] were destroyed to death and stained by PI with red fluorescence, and the CuS@[RBC-B16]-treated B16-F10 cells also present significant anticancer activity. While, loads of living cells with green fluorescence were observed in other control groups. Thus, the DCuS@[RBC-B16] NPs mediated photothermal/chemotherapy could result in a significant combinational antitumor efficacy.


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Figure 6. (A) The cell viabilities of B16-F10 cells with different treatments. (B) Confocal fluorescent microscopy images of live/dead staining of B16-F10 cells cultured with different materials without or with NIR laser irradiation (1064 nm, 1.0 W/cm2) (scale bar = 100 µm ).

In Vivo Antitumor Activity. Encouraged by the outstanding in vitro anticancer performance, we further performed the in vivo of DCuS@[RBC-B16] NPs antitumor experiments. The in vivo photothermal effect was first measured. The in vivo photothermal imaging was performed after the nude mice received the normal saline (NS) (100 µL), CuS@[RBC-B16] NPs, and DCuS@[RBC-B16] NPs intravenous injection from tail vein for 4 h. The mice treated with CuS@[RBC-B16] NPs and DCuS@[RBC-B16] presented obvious temperature rise, which increased to 52.6oC and 51.2oC under the l064 nm NIR laser (1064 nm, 1 W/cm2) irradiation irradiating for 5 min (Figure 7A). By contrast, for the mice injected with NS, the local temperature only increased to 40.1oC. It indicated the superior heat producing and highly targeting accumulation potential for the RBC-B16 hybrid membrane coated CuS NPs, which hold great potential to induce photothermal ablation of tumor tissues and specifically hyperthermia-responsive drug release in vivo. Meanwhile, the PA imaging analysis was consistent with the photothermal imaging. After injection of the CuS@[RBC-B16] NPs into nude mice for 4 h, strong local PA signal (in yellow circle) at the tumor tissue was observed (Figure 7B), indicating the highly specific self-targeting of CuS@[RBC-B16] NPs in vivo. In principle, the CuS@[RBC-B16] NPs retained the advantage of long circulation lifetime resulted from RBC-B16 hybrid membrane. To verify this, the blood retention of CuS NPs, CuS@RBC NPs, CuS@B16 NPs, and CuS@[RBC-B16] NPs over a span of 24 h after injection was studied. As shown in Figure 7C, after 24 h injection from tail vein, CuS@[RBC-B16] NPs exhibited blood retention of 20.2 % ID/g similar to that of the CuS@RBC NPs with blood retention of 22.9 % ID/g, while CuS@B16 NPs displayed blood retention of 14.5 % ID/g. By contrast, only 5.2 %


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ID/g of CuS NPs was found in blood circulation. Therefore, the CuS@[RBC-B16] NPs exhibited superior blood retention compared to bare CuS NPs and CuS@B16 NPs owing to the inherent properties of the RBC. The circulation half-time of the CuS@[RBC-B16] NPs and CuS@RBC NPs was calculated to be 9.6 h and 9.9 h, which was much longer that 1.0 h for the bare CuS NPs and 6.0 h for CuS@B16 NPs. The observed superior blood retention and circulation lifetime of CuS@[RBC-B16] NPs preferably facilitated to increase the accumulation of NPs at tumor. After 24 h, the tumors and major organs of mice were collected to further investigate the distribution of CuS NPs and CuS@[RBC-B16] NPs, respectively. After 24 h, all mice tumors and major organs were reaped and analyzed by ICP-OES after the injection for biodistribution study. As shown in Figure 7D, the CuS@[RBC-B16] NPs displayed a 1.5-fold, 1.4-fold, and 2.5-fold higher tumor accumulation than that of CuS@RBC NPs, CuS@B16 NPs, and CuS NPs. It revealed that the CuS@[RBC-B16] NPs possessed higher selftargeting capability derived from B16 membrane compared to the CuS@RBC NPs, and the long circulation time inherited from RBC membrane enhanced its tumor accumulation in comparison with CuS@B16 NPs. The amount of CuS@[RBC-B16] NPs accumulated in the liver and spleen reduced by 23.0 % and 34.6 % than that of bare CuS NPs, respectively. It demonstrated that CuS@[RBC-B16] NPs could be disguised as cells to decrease the interception by liver and spleen because of the RBC-B16 hybrid membrane. These results proved that CuS@[RBC-B16] NPs not only had a self-recognition of B16-F10 cell effect at cellular level but also demonstrated a self-recognition ability at animal level.


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Figure 7. (A) Infrared thermography of tumor-bearing mice which were tail vein injected of a: NS, b: CuS@[RBCB16] and c: DCuS@[RBC-B16]. (B) PA imaging in the B16-F10-tumor-bearing mice pre-injection or after-injection of CuS@[RBC-B16]. (C) Blood retention and (D) Biodistribution of CuS NPs, CuS@RBC NPs, CuS@B16 NPs and CuS@[RBC-B16] NPs over a span of 24 h after injection.

We investigated the in vivo toxicity of the CuS@[RBC-B16] NPs and DCuS@[RBC-B16] NPs systematically by blood biochemistry. Three important liver function indicators including alanine aminotransferase (ALT), aspartate aminotransferase (AST) and alkaline phosphatase (ALP), as well as one kidney function indicator of blood urea nitrogen (BUN) were all no significant differences between the control group and the treatment groups (Figure S8), indicating the good biocompatibility of the CuS@[RBC-B16] NPs and DCuS@[RBC-B16] NPs. To study the antitumor performance of DCuS@[RBC-B16] NPs, BALB/c nude mice (female, 6 weeks) bearing with melanoma (around 30 mm3) were divided into 7 groups (n=5) randomly. As shown in Figure 8A-C, neither DOX nor NIR-irradiation showed significant antitumor effect compared to NS control. The tumor-inhibition efficiency of CuS@[RBC-B16] was similar to


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free DOX, the tumor progression of DCuS@[RBC-B16]-treated mice was seriously inhibited upon NIR irradiation. For DCuS@[RBC-B16], the tumor growth inhibition (TGI) rate was around 14 % without the NIR irradiation, while the TGI rate almost reached up to 100 % for the mice treated with DCuS@[RBC-B16] and irradiated with NIR-irradiation. It was more effective than the DOX group (with the TGI rate around 10 %), NIR-irradiation group (with the TGI rate around 3 %) or CuS@[RBC-B16] NPs under NIR irradiation group (with the TGI rate around 86 %). Thus, the combination therapy induces an exciting effect in inhibiting tumor growth in vivo. Meanwhile, no obvious abnormality was observed for the body weight of the DCuS@[RBCB16] treated mice Figure 8D.

Figure 8. (A) Photographs of melanoma-bearing mice and (B) tumors for each tested groups (1: NS, 2: CuS@[RBC-B16], 3: DOX, 4: NIR laser (1064 nm, 1 W/cm2), 5: DCuS@[RBC-B16], 6: CuS@[RBC-B16] with NIR laser (1064 nm, 1 W/cm2), 7: DCuS@[RBC-B16] with NIR laser (1064 nm, 1 W/cm2). (C) Relative tumor volume and (D) Relative body weight of melanoma-bearing mice received different treatments.


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Furthermore, H&E and terminal-deoxynucleoitidyl transferase tediated nick end labeling (TUNEL) staining were performed after various treatments. A more marked destroyed of tumor cells in the DCuS@[RBC-B16] treated tumor with NIR laser irradiation groups than other groups (Figure 9A). It was observed that, significant antitumor effects and cell apoptosis in the CuS@[RBC-B16] and DCuS@[RBC-B16] with NIR laser irradiation groups, especially the later, almost the cell were apoptotic/dead, while no obvious antitumor behavior for the other groups. The H&E staining assay of the major organs after mice were scarified indicated the minimal systematic toxicity of DCuS@[RBC-B16] (Figure 9B), indicating the good biocompatibility of DCuS@[RBC-B16] with NIR irradiation.


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Figure 9. (A) H&E staining and TUNEL assay of the tumor tissues after different treatments (scale bar = 50 µm), (B) H&E staining of heart, liver, spleen, lung and kidney tissue slices from tumor-bearing mice received different treatments for 15 days. 1: NS, 2: CuS@[RBC-B16], 3: DOX, 4: NIR laser (1064 nm, 1 W/cm2), 5: DCuS@[RBCB16], 6: CuS@[RBC-B16] with NIR laser (1064 nm, 1 W/cm2), 7: DCuS@[RBC-B16] with NIR laser (1064 nm, 1 W/cm2) (scale bar = 200 µm).

CONCLUSION In summary, we have successfully created B16-F10s and RBCs hybrid biomimetic coating and fabricated RBC-B16 hybrid membrane camouflaged doxorubicin (DOX)-loaded hollow copper sulfide nanoparticles (DCuS@[RBC-B16] NPs) for combination therapy of melanoma. Compared to the bare CuS NPs, the DCuS@[RBC-B16] NPs exhibit infusive specific selfrecognition to B16-F10 cells in vitro, markedly prolonged circulation life and enhanced homogenous targeting abilities in vivo due to the preserved characteristic functionality derived from both of the source cells. The synergistic photothermal/chemotherapy of DCuS@[RBC-B16] NPs show excellent anti-cancer efficacy with about 100 % melanoma TGI rate. The proposed strategy may contribute to personalized nanomedicine of various tumors by combining the RBCs with homotypic cancer membrane accordingly to camouflage on the surface of nanoparticle. EXPERIMENTAL SECTION Materials. Poly(vinylpyrrolidone) (PVP-K30), cupric chloride (CuCl2), sodium sulfide (Na2S) and hydrazine anhydrous were obtained from Sinopharm Chemical Reagent Co., Ltd. (Beijing, China).

3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium

bromide

(MTT)

and

phenylmethanesulfonyl fluoride (PMSF) were purchased from Sigma-Aldrich (St. Louis, MO). Hydrazine anhydrous (H4N2·H2O) was obtained from Guanghua Sci-Tech Co., Ltd. (Guangdong, China). All other reagents were of analytical grade. Hoechst 33342, DiD, DiI, DiO, calcein-AM and PI were obtained from Yeasen Biotech. Co., Ltd. (Shanghai, China). Membrane protein


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extraction kit and bicinchoninic acid (BCA) protein assay kit were obtained from Beyotime Biotech. Co., Ltd. (Shanghai, China). Instrument. The morphologies of CuS@[RBC-B16] NPs were examined with a FEIF20 TEM (FEI, USA). The UV-visible (UV-vis) absorption analysis was recorded with an UV-1800 spectrophotometer (Shimadzu, Japan). Zeta potential analysis and DLS was performed on Nano ZS (Malvern, UK). All fluorescence photographs were obtained on a confocal laser scanning fluorescence microscope (LSM710META, Zeiss, Germany). The fluorescence spectra were measured with a fluorescence spectrophotometer (F-7000, Hitachi, Japan). The hematoxylin and eosin (H&E) slices were observed with a fluorescent inverted microscope (IX73, Olympus, Japan). A thermal imager (TiS65, Fluke, America) was employed to record infrared thermal imaging. Preparation of CuS NPs. The CuS NPs were prepared according to previous reports.36,37 Briefly, CuCl2 solution (100 µL, 0.5 mol/L) and PVP-K30 (240 mg) were added to 25 mL deionized water under magnetic stirring at room temperature. NaOH aqueous stock solution (25 mL, 0.01 mM, pH = 9.0) was then added in the mixture, and N2H4·H2O (6.4 µL) was added 2 min later. Afterwards, Na2S aqueous stock solution (200 µL, 320 mg/mL) was added to the suspension after 5 min. Keeping the reaction at 75oC for 2 h to generate CuS NPs. The CuS NPs were collected by centrifuging at 11000 r/min for 10 min and washing twice with deionized water. Preparation of RBC Membrane5,30 and B16-F10 Cell Membrane.23 The whole blood from BALB/c nude mice was centrifuged for 5 min at 3000 r/min (4oC) to remove the plasma, and the obtained RBCs precipitate was washed with NS thrice. Afterwards, adding deionized water to the RBCs precipitate for hemolysis in 4oC for 1 h, and the hemoglobin was removed by


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centrifugation at 13000 r/min for 5 min. The RBC membrane was further washed with deionized water until the supernatant became colorless. The RBC membrane was collected and stored at 80oC for further use. The preparation of B16-F10 cell membrane was according to the manufacturer’s instructions of membrane protein extraction kit. Briefly, B16-F10 cells were incubated in cell culture dishes with diameter of 10 cm containing Dulbecco's Modified Eagle's Medium (DMEM) supplemented with 10 % (v/v) fetal bovine serum (FBS), 1 % penicillin and streptomycin at 37 °C in a humidified atmosphere containing 5 % CO2. The cells were scraped off with cell scrapers after 48 h and collected by centrifugation at 1000 r/min for 1 min. The collected cells were washed thoroughly with PBS (10 mM, pH = 7.4) at 4oC and then suspended in membrane protein extraction reagent A containing PMSF (1 mM). The mixture was incubated in ice bath for 15 min and then centrifuged at 3000 r/min for 10 min. The supernatant was further centrifuged at 13000 r/min for 30 min to obtain the membrane, which was lyophilized and stored at -80oC for further use. Creation and Characterization of RBC-B16 Hybrid Membrane. BCA protein kit was used to quantify the concentration of RBC and B16-F10 membrane proteins, and the weight of membrane was the double of membrane protein weight. RBC membrane was added to B16-F10 membrane at RBC membrane to B16-F10 membrane protein weight ratios of 5:1, 3:1, 1:1, and 0:1, respectively, followed by sonicated at 37°C for 10 min to complete membrane fusion.35 FRET was employed to study the fusion process. Briefly, B16-F10 cell membrane was stained with DiD (excitation/emission=644/663 nm) and DiI (excitation/emission = 549/565 nm), and RBC membrane was then added to the DiD/DiI-dyed B16-F10 membrane at different weight ratios. The fluorescence spectrum of each sample was then read between 550 and 750 nm with


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excitation wavelength at 525 nm. The fluorescence recovery of the donor (DiI) was used to monitor the increasing amounts of fusion. Preparation of CuS@[RBC-B16] NPs. B16-F10 cell membrane solution (1 mL, 0.3 mg/mL) was added to RBC membrane solution (250 µL, 1.2 mg/mL) at RBC membrane to B16-F10 membrane weight ratio of 1:1, the mixture was then sonicated at 37 °C for 10 min to obtain the [RBC-B16] membrane. CuS NPs solution (0.5 mL, 0.2 mg/mL) was then added to RBC-B16 solution (0.5 mL, 0.3 mg/mL) and sonicated for 10 min to achieve the RBC-B16 coating. The mixture solution was centrifuged at 10000 r/min for 5 min to remove the excess membrane, and the resulting CuS@[RBC-B16] NPs were re-suspended with deionized water for future use. DOX Loading and Release in Vitro.30 1 mL PBS (10 mM, pH = 7.4) solution containing different concentrations of DOX (0.2 mg/mL, 0.4 mg/mL, 0.6 mg/mL, 0.8 mg/mL, 1.0 mg/mL) was added to 0.4 mg CuS NPs and stirred for 24 h. Afterwards, RBC-B16 membrane (1 mL, 0.8 mg/mL) was added to the mixture and sonicated for 10 min to generate DCuS@[RBC-B16] NPs. The DCuS@[RBC-B16] NPs were collected by centrifugation at 8000 r/min for 5 min. The DOX LE and LC was calculated according to the following equation using the UV-Vis absorbance spectrometer, respectively. LE =

− × 100%

LC =

− × 100% +

Where MT is the total mass of DOX for drug loading, and MU is the mass of unencapsulation DOX; MN is the mass of CuS, and MM was the mass of membrane in CuS@[RBC-B16] NPs. The DOX release study was performed in three groups including (1) pH = 7.4 without NIR irradiation, (2) pH = 5.0 without NIR irradiation, (3) pH = 7.4 with NIR irradiation, (4) pH=5.0


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with NIR irradiation (1064 nm, 1 W/cm2). PBS buffer (10 mM, pH = 7.4) and acetate buffer (10 mM, pH = 5.0) ware used. To study the DOX release efficiency, DCuS@[RBC-B16] NPs (1 mL, with CuS concentration at 0.4 mg/mL, the DOX LC was 87.7 % in weight) was dissolved in PBS buffer (10 mM, pH = 7.4) or acetate buffer (10 mM, pH = 5.0), respectively. 0.2 mL of solution was taken out at desired time points. The samples were spun down at 8000 r/min for 5 min, and DOX content in the supernatant was measured to determine the released DOX content using UV-vis spectrometer. To investigate the hyperthermia-triggered drug release, the solution was irradiated under 1064 nm laser (1 W/cm2) for 5-min and 0.2 mL of solution was taken out at desired time points, and the other process was similar to aforementioned. Intracellular DOX Release. The intracellular DOX release was divided into three groups according to the different treatments: (1) DCuS, (2) DCuS@[RBC-B16], (3) DCuS@[RBC-B16] with NIR irradiation (1064 nm, 1 W/cm2). B16-F10 cells were seeded into confocal dishes at the density of 5 × 104 cells per dish and incubated for 24 h. Then the medium was replaced by fresh medium containing DCuS NPs (with CuS concentration at 50 µg/mL) or DCuS@[RBC-B16] (with CuS concentration at 50 µg/mL) and incubated for 4 h at 37oC. Then the group (3) was irradiated under NIR for 5 min. The nucleus were stained with hoechst 33342 30 min later, and the cells were observed with a confocal fluorescence microscopy. Specific-Targeting B16-F10 Cell Line in Vitro. HT1080 cells, NHDF cells and B16-F10 cells were seeded into confocal dishes at the density of 1× 104 cells per dish and incubated for 12 h. Then the medium was replaced with the fresh medium containing CuS@[RBC-B16] NPs with DiI-dyed RBC-B16 (with CuS concentration at 50 µg/mL). The nucleus were dyed with hoechst 33342, and 4 h later all the groups were detected with a confocal fluorescence microscopy.


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In Vitro Anti-tumor Performance.30,38 B16-F10 cells were seeded into 96-well plate at the density of 1 × 104 cells per well and cultured for 24 h at 37oC. The media were replaced with fresh media containing DOX (50 µg/mL), CuS@[RBC-B16] (with CuS concentration at 50 µg/mL), DCuS@[RBC-B16] (with CuS concentration at 50 µg/mL) or PBS (10 mM, pH = 7.4), respectively. After 4 h incubation, the cells were irradiated with or without 1064 nm NIR laser (1 W/cm2) for 5 min. The cells were further incubated for 12 h, and the media were replaced with fresh medium containing MTT (0.5 mg/mL) and incubated for 4 h. Afterwards, the media were removed and 200 µL DMSO was added into each well. The absorbance at 490 nm was measured with a microplate reader. The cell viability was calculated according to the following equations. Cell viability =

⁄ !"#$ × 100% ⁄&'()'#

Calcein-AM/PI assay were also employed to study the in vitro anti-tumor of DCuS@[RBCB16]. The B16-F10 cells seeded in confocal dishes were similarly treated with NPs incubation and NIR-irradiation. The resulting cells were stained with calcein-AM/PI and observed using a confocal fluorescence microscopy. In Vivo Blood Circulation and Bio-distribution. BALB/c nude mice (female, 6 weeks old) with B16-F10 tumor xenografts were received tail vein injection of CuS NPs (25 mg/kg) or CuS@[RBC-B16] NPs (with CuS concentration at 25 mg/kg). The venous blood was collected at desired time points (0 h, 2 h, 4 h, 8 h, 12 h, 24 h) and the mice were then euthanized. The major organs and tumor tissues were gathered and dissolved in chloroazotic acid. The copper ion contents in blood, organs and tumors were measured with ICP-OES. In Vivo Melanoma Tumor Growth Inhibition.39 The BALB/c nude mice (female, 6 weeks old) with subcutaneous B16-F10 tumor xenografts were divided into seven groups according to the different treatments: (n = 5 per group): (1) NS, (2) CuS@[RBC-B16] NPs (with CuS


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concentration at 5 mg/kg), (3) DOX dissolved in NS (5 mg/kg), (4) NIR laser irradiation (1064 nm, 1 W/cm2), (5) DCuS@[RBC-B16] NPs (with CuS concentration at 5 mg/kg), (6) CuS@[RBC-B16] NPs (with CuS concentration at 5 mg/kg) with NIR laser irradiation, (7) DCuS@[RBC-B16] NPs (with CuS concentration at 5 mg/kg) with NIR laser irradiation. All the samples were intravenously injected into mice from tail vein. As for the photoacoustic (PA) imaging, the mice were observed after the samples were intravenously injected into mice for 4 h. Similarly, the photothermal imaging was performed during the NIR laser (1064 nm, 1 W/cm2) irradiation after the samples were injected for 4 h. To perform the in vivo tumor treatment, the BALB/c nude mice (female, 6 weeks old) with subcutaneous B16-F10 tumor xenografts were divided into seven groups according to the different treatments as mentioned above. (n = 5 per group): (1) NS, (2) CuS@[RBC-B16] NPs (with CuS concentration at 5 mg/kg), (3) DOX dissolved in NS (5 mg/kg), (4) NIR laser irradiation (1064 nm, 1 W/cm2), (5) DCuS@[RBC-B16] NPs (with CuS concentration at 5 mg/kg), (6) CuS@[RBC-B16] NPs (with CuS concentration at 5 mg/kg) with NIR laser irradiation, (7) DCuS@[RBC-B16] NPs (with CuS concentration at 5 mg/kg) with NIR laser irradiation. For NIR laser irradiation, the mice were irradiated for 5 min with a NIR laser (1064 nm, 1 W/cm2) after the samples were intravenously injected into mice for 4 h. The tumor volume and mice body weight were measured every day. After 15 days, all the mice were euthanized and the major organs and tumors were collected for H&E staining and TUNEL staining. The TGI rate was calculated according to the following equations. TGI =

-. − - × 100% -.

Where VC is the tumor volume of the control group (NS group), and VT is the tumor volume of the other six groups.


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ASSOCIATED CONTENT Supporting Information. The following files are available free of charge. Additional figures (PDF) AUTHOR INFORMATION Corresponding Author *E-mail: [email protected]. *E-mail: [email protected]. ACKNOWLEDGMENT The work was supported by Special Foundation for State Major Research Program of China (Grant Nos. 2016YFC0106602 and 2016YFC0106601); National Natural Science Foundation of China (Grant No. 21645005, 21475008); the Open Research Fund Program of Beijing Key Lab of Plant Resource Research and Development, Beijing Technology and Business University (PRRD-2016-YB2). REFERENCES (1) Fang, R. H.; Hu, C. J.; Luk, B. T.; Gao, W.; Copp, J. A.; Tai, Y.; O Connor, D. E.; Zhang, L. Cancer Cell Membrane-Coated Nanoparticles for Anticancer Vaccination and Drug Delivery. Nano Lett. 2014, 14, 2181−2188. (2) Kroll, A. V.; Fang, R. H.; Zhang, L. Biointerfacing and Applications of Cell MembraneCoated Nanoparticles. Bioconjugate Chem. 2017, 28, 23−32.


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(3) Wang, J.; Liu, J.; Liu, Y.; Wang, L.; Cao, M.; Ji, Y.; Wu, X.; Xu, Y.; Bai, B.; Miao, Q.; Chen, C.; Zhao, Y. Gd-Hybridized Plasmonic Au-Nanocomposites Enhanced Tumor-Interior Drug Permeability in Multimodal Imaging-Guided Therapy. Adv. Mater. 2016, 28, 8950−8958. (4) Zhang, Q.; Colazo, J.; Berg, D.; Mugo, S. M.; Serpe, M. J. Multiresponsive Nanogels for Targeted Anticancer Drug Delivery. Mol. Pharmaceut 2017, 14, 2624−2628. (5) Rao, L.; Bu, L.; Xu, J.; Cai, B.; Yu, G.; Yu, X.; He, Z.; Huang, Q.; Li, A.; Guo, S.; Zhang, W.; Liu, W.; Sun, Z.; Wang, H.; Wang, T.; Zhao, X. Red Blood Cell Membrane as a Biomimetic Nanocoating for Prolonged Circulation Time and Reduced Accelerated Blood Clearance. Small 2015, 11, 6225−6236. (6) Guo, L.; Yan, D. D.; Yang, D.; Li, Y.; Wang, X.; Zalewski, O.; Yan, B.; Lu, W. Combinatorial Photothermal and Immuno Cancer Therapy Using Chitosan-Coated Hollow Copper Sulfide Nanoparticles. ACS Nano 2014, 8, 5670−5681. (7) Li, S.; Cheng, H.; Xie, B.; Qiu, W.; Zeng, J.; Li, C.; Wan, S.; Zhang, L.; Liu, W.; Zhang, X. Cancer Cell Membrane Camouflaged Cascade Bioreactor for Cancer Targeted Starvation and Photodynamic Therapy. ACS Nano 2017, 11, 7006−7018. (8) Zhang, S.; Sun, C.; Zeng, J.; Sun, Q.; Wang, G.; Wang, Y.; Wu, Y.; Dou, S.; Gao, M.; Li, Z. Ambient Aqueous Synthesis of Ultrasmall PEGylated Cu2-xSe Nanoparticles as a Multifunctional Theranostic Agent for Multimodal Imaging Guided Photothermal Therapy of Cancer. Adv. Mater. 2016, 28, 8927−8936. (9) Gao, Y.; Ahiabu, A.; Serpe, M. J. Controlled Drug Release from the AggregationDisaggregation Behavior of pH-Responsive Microgels. ACS Appl. Mater. Interfaces 2014, 6, 13749−13756.


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(10) Piao, J.; Wang, L.; Gao, F.; You, Y.; Xiong, Y.; Yang, L. Erythrocyte Membrane Is an Alternative Coating to Polyethylene Glycol for Prolonging the Circulation Lifetime of Gold Nanocages for Photothermal Therapy. ACS Nano 2014, 8, 10414−10425. (11) Rao, L.; Bu, L.; Meng, Q.; Cai, B.; Deng, W.; Li, A.; Li, K.; Guo, S.; Zhang, W.; Liu, W.; Sun, Z.; Zhao, X. Antitumor Platelet-Mimicking Magnetic Nanoparticles. Adv. Funct. Mater. 2017, 27, 1604774. (12) Liu, J.; Zhang, L.; Lei, J.; Shen, H.; Ju, H. Multifunctional Metal-Organic Framework Nanoprobe for Cathepsin B-Activated Cancer Cell Imaging and Chemo-Photodynamic Therapy. ACS Appl. Mater. Interfaces 2017, 9, 2150−2158. (13) Fojtů, M.; Chia, X.; Sofer, Z.; Masařík, M.; Pumera, M. Black Phosphorus Nanoparticles Potentiate the Anticancer Effect of Oxaliplatin in Ovarian Cancer Cell Line. Adv. Funct. Mater. 2017, 27, 1701955. (14) Cheng, L.; Wang, C.; Feng, L.; Yang, K.; Liu, Z. Functional Nanomaterials for Phototherapies of Cancer. Chem. Rev. 2014, 114, 10869−10939. (15) Fang, R. H.; Jiang, Y.; Fang, J. C.; Zhang, L. Cell Membrane-Derived Nanomaterials for Biomedical Applications. Biomaterials 2017, 128, 69−83. (16) Parodi, A.; Quattrocchi, N.; van de Ven, A. L.; Chiappini, C.; Evangelopoulos, M.; Martinez, J. O.; Brown, B. S.; Khaled, S. Z.; Yazdi, I. K.; Enzo, M. V.; Isenhart, L.; Ferrari, M.; Tasciotti, E. Synthetic Nanoparticles Functionalized with Biomimetic Leukocyte Membranes Possess Cell-Like Functions. Nat. Nanotechnol. 2012, 8, 61−68. (17) Molinaro, R.; Corbo, C.; Martinez, J. O.; Taraballi, F.; Evangelopoulos, M.; Minardi, S.; Yazdi, I. K.; Zhao, P.; De Rosa, E.; Sherman, M. B.; De Vita, A.; Toledano Furman, N. E.;


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(39) Meng, X.; Liu, Z.; Cao, Y.; Dai, W.; Zhang, K.; Dong, H.; Feng, X.; Zhang, X. Fabricating Aptamer-Conjugated PEGylated-MoS2/Cu1.8S Theranostic Nanoplatform for Multiplexed Imaging Diagnosis and Chemo-Photothermal Therapy of Cancer. Adv. Funct. Mater. 2017, 27, 1605592.


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BRIEFS. The excellent immune evading and homogenous targeting tumor abilities combined with the high loading efficiency of DOX and inherent photothermal conversion property of CuS led to outstanding performance of DCuS@[RBC-B16] in synergistic photothermal/chemotherapy of melanoma in vivo. SYNOPSIS. RBC membrane and B16-F10 cells membrane materials are fused to create a hybrid biomimetic coating (RBC-B16), and RBC-B16 hybrid membrane camouflaged doxorubicin (DOX)-loaded hollow copper sulfide nanoparticles (DCuS@[RBC-B16] NPs) are fabricated for combination therapy of melanoma. The DCuS@[RBC-B16] NPs show the inherent properties of the both source cells. The DCuS@[RBC-B16] NPs exhibit highly specific self-recognition to source cell line in vitro, and achieve markedly prolonged circulation lifetime and enhanced homogenous targeting abilities in vivo inherited from the source cells. Thus, the DOX-loaded [RBC-B16]-coated CuS NP platform exhibits excellent synergistic photothermal/chemotherapy. Table of Contents (TOC) graphic


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Erythrocyte-Cancer Hybrid Membrane Camouflaged Hollow Copper

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