Environment-Responsive Dopamine Nanoplatform for Tumor Synergistic Therapy

Environment-Responsive Dopamine Nanoplatform for Tumor Synergistic Therapy | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Research Article Environment-Responsive Dopamine Nanoplatform for Tumor Synergistic Therapy Chunmin Deng, Hao Zhang, Li Song This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-3844904/v1 This work is licensed under a CC BY 4.0 License Status: Posted Version 1 posted You are reading this latest preprint version Abstract Nanoparticle-based photothermal therapy (PTT) has emerged as a promising approach in tumor treatment due to its high selectivity and low invasiveness. However, the penetration of near-infrared light (NIR) is limited, leading it fails to induce damage to the deep-seated tumor cells within the tumor tissue. Additionally, inefficient uptake of photothermal nanoparticles by tumor cells results in suboptimal outcomes for PTT. Based on the above-mentioned issues, this study utilized the adhesive properties of photothermal material, polydopamine (PDA), which can successfully load the photosensitizer indocyanine green (ICG) and chemotherapeutic drug doxorubicin (DOX) to achieve combined photothermal and chemotherapy treatment (PDA/DOX&ICG), aiming to compensate for the poor penetration of NIR in tumor tissues and the photothermal conversion performance of PDA. For the purpose of extending the blood circulation time of PDA/DOX&ICG nanoparticles, evading clearance by the body immune system and achieving targeted delivery to tumor tissues, a protective envelopment was created using erythrocyte membranes modified with folate acid (FA-EM). After reaching the tumor tissue, the obtained FA-EM@PDA/DOX&ICG nanoparticles can specific bind with folate acid receptors on the surface of tumor cells. This interaction facilitates improved uptake by tumor cells leading to the subsequent release of loaded DOX and ICG in response to the unique tumor microenvironment. DOX penetration ability can effectively compensate the limitation of NIR penetration at the tumor tissue. While ICG, as a typical photosensitizer, significantly enhances the photothermal conversion performance of FA-EM@PDA/DOX&ICG nanoparticles, thereby inducing tumor cells damage. In vitro and in vivo experimental results demonstrated that the coordinated NIR treatment with FA-EM@PDA/DOX&ICG not only effectively inhibits tumor growth but also exhibits superior biocompatibility, effectively mitigating DOX-induced tissue damage. Photothermal therapy combined therapy targeted delivery environment-responsive Figures Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 Introduction Tumor remains one of the primary threats to human health, current clinical treatments mainly revolve around chemotherapy, radiotherapy, immunotherapy, and surgical resection. 1–2 While these treatments effectively suppress tumor growth and extend patient survival, they lack precision in targeting tumors and often induce substantial toxic side effects in the body's normal tissues. 3–4 Therefore, the development of a novel type of treatment is urgently needed to enhance tumor treatment efficacy while minimizing damage to healthy tissues. In recent years, various innovative therapies and strategies, such as targeted drugs, photothermal therapy, and starvation therapy, have emerged. 5–6 These approaches not only enable precise and controllable treatment of tumors but also reduce the toxic side effects to body normal tissues and organs, significantly enhancing patients’ compliance. 7 Studies have revealed that tumor tissues lack sufficient blood supply, making them unable to regulate heat dissipation through blood flow and velocity. 8 Consequently, tumor tissues exhibit greater sensitivity to heat than normal tissues, leading to the term "natural heat reservoirs" for tumor tissues, presenting a natural advantage for photothermal therapy (PTT) in treating tumors. 8–9 Additionally, PTT offers temporal and spatial controllability, low invasiveness and high selectivity, making it an optimal approach for tumor treatment. 10 However, standalone PTT often requires high temperatures to induce tumor cells death, which risks damaging surrounding normal tissues. In recent years, with the advancement of photothermal nanoparticles, these limitations have been addressed. When these nanoparticles aggregate within tumor tissues, and then exposed under near-infrared light (NIR) irradiation. Nanoparticles can convert light energy into heat, and then rapidly elevating local tumor tissue temperatures to induce tumor cells apoptosis and reduce damage to surrounding normal tissues. 11 Nonetheless, nanoparticles are prone to clearance by body's immune system during their transport within the body, especially through phagocytosis by macrophages, resulting in shortened blood circulation periods and hindered them accumulated at tumor tissues. 12 To tackle this issue effectively, Fang et al. utilized folate acid-modified erythrocyte membranes to coat metal-organic frameworks and then loading with chemotherapeutic drug doxorubicin for tumor combination treatment. 13 This system of diagnosis and treatment not only effectively avoided phagocytosis by macrophages to prolonging their circulation in the bloodstream, but also with folate acid molecule (FA) modifications on their surfaces that can enhance phagocytosis of biomimetic nanoparticles by tumor cells and achieved controlled drug release according to the specific tumor microenvironment (TME). However, the limited penetration of NIR light resulting in its inability to eradicate deep-seated tumor tissues entirely. To achieve completely tumor cells eradication, researchers have exploited the inherent properties of photothermal nanoparticles to load chemotherapeutic drugs. 14 Upon reaching tumor tissues, these nanoparticles releasing the loaded chemotherapy drugs based on the specific TME, promoting penetration of the drugs into deep-seated tumor tissues while reducing their systemic toxicity during transport. 15 Consequently, a plethora of combined therapies based on PTT, such as PTT/chemotherapy and PTT/photodynamic therapy/chemotherapy, effectively compensate for the limitations of individual treatment methods and achieve efficient tumor clearance. 16–17 To enhance the biocompatibility of photothermal nanoparticles, this study ingenious utilized the neurotransmitter-dopamine is prone to oxidative polymerization to form the photothermal material polydopamine (PDA) under alkaline conditions, to achieve tumor photothermal therapy. 18 To improve the photothermal performance and therapeutic efficacy of PDA, doxorubicin (DOX), a chemotherapeutic drug, and indocyanine green (ICG), a photothermal material, are loaded into PDA surface (PDA/DOX&ICG) due to its surface adsorption properties. 19 And then PDA/DOX&ICG nanoparticles are then enveloped with folate-modified erythrocyte membranes (FA-EM) to obtain FA-EM@PDA/DOX&ICG, which can effectively avoid FA-EM@PDA/DOX&ICG nanoparticles being cleared by the body's immune system, thereby prolonging their blood circulation cycle (Scheme 1a ). Upon reaching at tumor tissues, they can readily bind with folate acid receptors specifically located on the surface of tumor cells, enhancing uptake by tumor cells and facilitating DOX and ICG in-site release based on the specific TME (Scheme 1b ). When NIR is irradiated, the ICG present in the FA-EM@PDA/DOX&ICG nanoparticles can effectively improve the photothermal conversion efficiency, and the penetration of DOX into tumor tissue can compensate for the lack of NIR tissue penetration, thereby inhibiting tumor growth and alleviate the systemic toxic effects of DOX. Materials and Methods Materials Dopamine, polyvinyl alcohol (PVA) and KMnO 4 were purchased from Beijing Guoyao (Beijing, China). Doxorubicin (DOX), indocyanine green (ICG), DAPI, and CCK-8 were obtained from Shanghai Aladdin Reagent Co., Ltd (Shanghai, China). PBS, 1640 culture medium, and fetal bovine serum were sourced from Sigma-Aldrich Chemicals (Madison, USA). All chemical reagents used in the experiments were not further purified, and ultrapure water was used throughout the entire experimental process. Preparation of Polydopamine The synthesis of polydopamine (PDA) followed by a modified version of the laboratory's previous procedure. 20–21 Initially, 5 mg/mL polyvinyl alcohol (PVA) was added into 10 mL ultrapure water and heated to 90°C to completely dissolve PVA. After cooling the PVA solution to room temperature, 0.2 g dopamine and 1 mL KMnO 4 (1 mg/mL) were separately added to the solution, followed by stirring for 2 h at room temperature. The solution was then centrifuged at 3500 rpm for 20 min to collect the supernatant, which was subsequently placed in a dialysis bag and dialyzed for 3 days. Finally, the resulting sample of PDA nanoparticles was freeze-dried and stored for later use. Preparation of PDA/DOX&ICG 20 mg PDA nanoparticles were dispersed in 10 mL ultrapure water to achieve thorough dispersion. 22 Subsequently, 10 mg of DOX and 10 mg of ICG were separately added to the dispersed solution under light-avoiding conditions and stirred for 24 h. The resulting mixed solution was collected, placed in a dialysis bag, and dialyzed for 3 days. Finally, the obtained PDA/DOX&ICG nanoparticles were freeze-dried and stored for later use. Preparation of FA-EM@PDA/DOX&ICG Fresh blood was collected from BALB/c mice through eye extraction and placed in an anticoagulant tube. After centrifugation (5000 rpm, 10 min) to remove unwanted cells and serum, the collected erythrocyte membranes (EM) were washed several times with PBS until the supernatant was colorless. The EM were gently mixed with ultrapure water and left to stand at 4°C for 1 h to release hemoglobin and other related components. The cells were then washed with PBS until the supernatant was colorless and transparent. Next, 20 mg of DSPE-PEG-FA was added to 5 mL of the EM solution and stirred for 24 h. The obtained DSPE-PEG-FA modified EM (FA-EM) were collected after centrifugation and washed three times with PBS. Finally, the collected FA-EM was resuspended in 1 mL PBS, and 1 mg of PDA/DOX&ICG was added to the solution. This mixture was extruded through a 220 nm polycarbonate porous membrane using a squeezing method to obtain FA-EM@PDA/DOX&ICG nanoparticles. The nanoparticles were collected by centrifugation, freeze-dried, and stored at -80°C. 23–24 Characterization of FA-EM@PDA/DOX&ICG Nanoparticles FA-EM@PDA/DOX&ICG nanoparticles were dispersed in deionized water and sonicated for 10 minutes. The dispersed solution was then dropped into a copper grid, air-dried, and observed under transmission electron microscopy (TEM) to determine the size and shape of the nanoparticles. Changes in the zeta potential and particle size of FA-EM@PDA/DOX&ICG nanoparticles before and after drug loading were analyzed using a particle size analyzer. The successful preparation of FA-EM@PDA/DOX&ICG nanoparticles was further validated by UV-spectrophotometry. Protein gel electrophoresis was used to analyze the expression of the EM protein profile on the surface of the prepared nanoparticles. Cell Culture Mouse 4T1 breast cancer cells, human macrophages (RAW264.7), and human gastric mucosal cells (GES-1) were all obtained from Nanjing University (Nanjing, China) and cultured in a 37°C, 5% CO 2 incubator. The cell culture medium used was RPMI 1640 medium (Gibco) containing 10% fetal bovine serum (FBS) (Gibco, Shanghai, China). All cell lines were approved by the Institutional Animal Care and Use Committee of Nanijing University. Photothermal Performance of FA-EM@PDA/DOX&ICG Nanoparticles Initially, 1 mg of FA-EM@PDA/DOX&ICG nanoparticles were dispersed in 1 mL PBS buffer solution and thoroughly mixed. Subsequently, varying volumes of FA-EM@PDA/DOX&ICG nanoparticles were aspirated and added to different volumes of PBS buffer solution to prepare suspensions with different concentrations of FA-EM@PDA/DOX&ICG dispersion solution. Then, the prepared dispersion solution of FA-EM@PDA/DOX&ICG nanoparticles were exposed under different NIR powers. At predetermined time points, the temperature changes for each group were recorded using an infrared thermal imaging instrument. 25 The photothermal conversion efficiency of FA-EM@PDA/DOX&ICG nanoparticles was evaluated according to the relevant methods. 26–27 Initially, 100 µl of FA-EM@PDA/DOX&ICG (200 µg/ml) dispersion solution was added to an Eppendorf tube (EP) and placed under 808 nm (1.5 W/cm 2 ) NIR irradiation for 10 minutes. After turning off the NIR light for 10 minutes, irradiation was resumed for another 10 minutes. The temperature changes at each time point were recorded using an infrared thermal image. The photothermal conversion efficiency of FA-EM@PDA/DOX&ICG nanoparticles was then assessed: θ = (T - T min )/(T max - T min ) ( 1 ) hS = - m*C p /ζ ( 2 ) η = hs (ΔT max,mix − ΔT max,H2O )/ I (1 − 10 − A808 ) ( 3 ) θ is the dimensionless driving force temperature, and defined as the ratio of (T - T min ) to (T max - T min ).ζis the slope of T and -Lnθ. Consequently, the m NPs and C p, NPs of NPs were neglected. m was 2×10 − 4 kg. C p was 4.2×10 3 J/(kg·℃). ΔT max,mix is the temperature change of the FA-EM@GO-MOF dispersion at the maximum steady-state temperature. ΔT max,H2O is the temperature change of water at the maximum steady-state temperature. I is the laser power, A 808 is the absorbance of FA-EM@GO-MOF at the wavelength of 808 nm in aqueous solution. Targeted Delivery and Immune Escape of FA-EM@PDA/DOX&ICG Nanoparticles Initially, 4T1 cells and GES-1 cells were separately seeded into cell culture dishes and incubated in a cell culture incubator (37°C, 5% CO 2 ) for 12 h. Subsequently, FA-EM@PDA/DOX&ICG nanoparticles were added to the 4T1 cells and GES-1 cells, and then incubated for 3 hours. Afterward, the supernatant was discarded, and fixation was performed using 4% paraformaldehyde for 15 minutes followed by DAPI staining for 20 minutes and several washes with PBS. Finally, the fluorescence intensity of 4T1 cells and GES-1 cells was observed using fluorescence confocal microscopy. 28 RAW264.7 cells were seeded into cell culture dishes and incubated in a cell culture incubator (37°C, 5% CO 2 ) for 12 hours. Subsequently, PDA/DOX&ICG, EM@PDA/DOX&ICG, and FA-EM@PDA/DOX&ICG nanoparticles were separately co-incubated with RAW264.7 cells for 3 hours. After removing the supernatant, fixation was conducted using 4% paraformaldehyde for 15 minutes, followed by DAPI staining for 20 minutes and multiple PBS washes. Finally, the fluorescence intensity of RAW264.7 cells in each group was observed using laser scanning confocal microscopy (LSCM). 29–30 Animal welfare In this study, all animal experiments were executed according to a protocol approved by the Animal Management Rules of the Ministry of Health of the People’s Republic of China and approved by the Institutional Animal Care and Use Committee of Nanijing University. Female 6- to 6-week-old BALB/c mice were purchased from Qinglong Mountain (Nanjing, People’s Republic of China) and maintained under specific pathogen-free-conditions. All efforts were made to minimize the animals’ suffering and to reduce the number of animals used. Construction and Treatment of Mouse 4T1 Subcutaneous Xenograft Model A suspension containing 2×10 7 4T1 cells were injected into the groin of male Balb/c mice to establish a mouse subcutaneous xenograft model. When the size of the 4T1 xenograft tumors reached approximately 100 mm 3 , the 4T1 xenograft mice were randomly divided into 6 groups (n = 6/group). These groups were treated with PBS, DOX (1 mg/kg), ICG (1 mg/kg), PDA (1 mg/kg), PDA/DOX&ICG (1 mg/kg), and FA-EM@PDA/DOX&ICG (1 mg/kg), respectively. The required drugs were intravenously injected into the mice via the tail vein, and administered every 3 days. After 24 h injection, the mice were exposed under NIR irradiation (1.5 W/cm 2 , 9 min). The mice's tumor growth and body weight were observed daily (as per animal ethics standards: a decrease in mouse body weight exceeding 15% of their initial weight was considered an indicator of death). The formula for calculating mouse xenograft tumor volume was as follows: V (mm 3 ) = 0.5 × a × b 2 , where V represents tumor volume, a represents the longer diameter of the tumor, and b represents the shorter diameter of the tumor. 15 days later, the mice were euthanized, and tumors from each treatment group were collected. 31–32 Histological Analysis Following the completion of the treatment, mice from each treatment group were euthanized using cervical dislocation. Major organs (heart, liver, spleen, lungs, and kidneys) and tumor tissues from the 4T1 xenograft mice were collected for histological analysis. The collected tissues were fixed in 4% formaldehyde and subsequently embedded in paraffin. Tissue sections were prepared and stained with hematoxylin and eosin (H&E) after embedding. For tumor tissues, in situ terminal deoxynucleotidyl transferase dUTP nick-end labeling (TUNEL) staining was also performed. The processed tissue sections were placed on a slide scanner to capture tissue images, and the prepared tissue sections were assessed by experienced pathologists. 33–34 Statistical Analysis The experimental data were analyzed using SPSS software (version 17.0) and expressed as mean ± standard deviation (X ± S). One-way analysis of variance (ANOVA) was employed for comparing means among multiple groups, while the t-test was further utilized for comparing means between two samples. A significance level of p < 0.05 was considered indicative of statistical differences. When p < 0.05, it was denoted as "*", p < 0.01 as "**", and p < 0.001 as "***". Results and Discussion Biological Characterization of FA-EM@PDA/DOX&ICG Nanoparticles In this study, the properties of dopamine undergoing oxidative polymerization to form PDA under alkaline conditions were utilized. 35 Therefore, we analyzed the biological characteristics of PDA, TEM results demonstrated that the prepared PDA nanoparticles were circular, with a particle size for about 70 nm (Fig. 1 a and 1 c). Besides, utilizing the adsorption properties of the PDA to load therapeutic drugs. Once, DOX and ICG were successfully adsorption into the PDA surface, the resulting PDA/DOX&ICG nanoparticles exhibited an increased particle size and a shift in the zeta potential from − 4.1 mV to -1.2 mV (Fig. 1 c and 1 d). However, when the prepared PDA/DOX&ICG nanoparticles underwent further encapsulation with FA-EM, a thin film covered the surface of PDA/DOX&ICG nanoparticles (Fig. 1 b). The size and zeta potential of the obtained FA-EM@PDA/DOX&ICG nanoparticles became 122 nm and − 6.3 mV, respectively (Fig. 1 b, 1 c and 1 d). Subsequently, UV-Vis spectroscopy results confirmed the absorption peaks of the prepared FA-EM@PDA/DOX&ICG nanoparticles, further verified the successful adsorption of DOX and ICG onto the PDA surface, with FA-EM effectively encapsulated on the surface of PDA/DOX&ICG nanoparticles (Fig. 1 e). More importantly, polyacrylamide gel electrophoresis was employed to analyze the retention of membrane proteins on the surface of FA-EM@PDA/DOX&ICG nanoparticles. As shown in Fig. 1 f, the protein profile of FA-EM@PDA/DOX&ICG nanoparticles matched well with EM proteins, indicated good retention of membrane proteins after FA-EM encapsulation. These results demonstrated that FA-EM@PDA/DOX&ICG nanoparticles were successfully fabricated and retaining the relevant proteins of EM. FA-EM@PDA/DOX&ICG Photothermal Conversion Performance PDA, as a typical photothermal material, can convert light energy into heat under NIR irradiation, thereby inducing photothermal ablation of tumors. 36 However, PDA exhibits suboptimal photothermal conversion performance, resulting in inadequate therapeutic outcomes. 37 Hence, in this study, the adsorption capability on the surface of PDA was utilized to load the photosensitizer ICG to enhance the photothermal conversion performance of FA-EM@PDA/DOX&ICG nanoparticles. 38 As illustrated in Fig. 2 a, 200 µg/mL of PDA, PDA/DOX&ICG, and FA-EM@PDA/DOX&ICG nanoparticles dispersion solution were separately placed under NIR irradiation (1.5 W/cm 2 , 9 min), and the temperature changes for each group were recorded at designated time points using an infrared thermal image (Fig. 2 b). After 9 min irradiation, the temperature of PDA, PDA/DOX&ICG, and FA-EM@PDA/DOX&ICG nanoparticles were respectively increased into 46.4 ℃, 55.1 ℃, and 54.8 ℃. These results indicated that the modification of FA-EM@PDA/DOX&ICG nanoparticles with FA-EM did not affect the photothermal conversion efficiency of PDA/DOX&ICG, while the incorporation of the photosensitizer ICG effectively enhanced PDA nanoparticles photothermal conversion performance. However, when 200 µg/mL of FA-EM@PDA/DOX&ICG nanoparticles were exposed under 2.0 W/cm 2 NIR for 9 min, the temperature increased to 59.8 ℃. These findings suggest a positive correlation between the photothermal conversion efficiency of FA-EM@PDA/DOX&ICG nanoparticles and NIR power, and duration. To verify the photothermal stability of FA-EM@PDA/DOX&ICG nanoparticles, a solution of 200 µg/mL FA-EM@PDA/DOX&ICG nanoparticles were exposed under 1.5 W/cm 2 NIR for 9 min, followed by turning on/off NIR irradiation for 9 minutes, and repeated four times. The temperature changes were recorded using an infrared thermal image (recorded every 3 minutes). As depicted in the Fig. 2 c, after four cycles of irradiation, there was no significant changes in the temperature of FA-EM@PDA/DOX&ICG nanoparticles, indicating sustained excellent photothermal conversion performance. And the photothermal conversion efficiency of FA-EM@PDA/DOX&ICG nanoparticles were measured for about 41.6% (Fig. 2 d), which is high than other photothermal materials,such as Cu 9 S 5 nanocrystals, copper selenide nanoparticles, WS 2 nanosheet and so on. 39–41 Thus, the prepared FA-EM@PDA/DOX&ICG nanoparticles exhibited superior photothermal performance and achieved multiple reversible conversion. Immune Evasion and Targeted Delivery of FA-EM@PDA/DOX&ICG Nanoparticles Due to the size of nanoparticles were very similar to viruses, when their entry into the body can trigger immune responses, causing nanoparticles are easily cleared by mononuclear phagocytes and hindering effective therapeutic delivery. 42–43 In this context, FA-EM@PDA/DOX&ICG nanoparticles were successfully developed, which were post-encapsulation with FA-EM, and retained relevant proteins from EM. Subsequently, co-incubation with macrophages (RAW264.7 cells) was conducted to evaluate the impact of FA-EM on the engulfment of FA-EM@PDA/DOX&ICG nanoparticles. When PDA/DOX&ICG nanoparticles were co-incubated with RAW264.7 cells, strong green and red fluorescence signals were observed through LSCM. Conversely, co-incubation with EM@PDA/DOX&ICG resulted in weak fluorescence signals. However, when FA-EM@PDA/DOX&ICG nanoparticles were co-incubated with RAW264.7 cells, the fluorescence signals partially recovered compared to EM@PDA/DOX&ICG, although remaining lower than the PDA/DOX&ICG group (Fig. 3 a). These findings suggest that EM imparts superior "stealth" properties to PDA/DOX&ICG nanoparticles, with folate acid (FA) modification causing a minor reduction in this stealth effect. 44 Nevertheless, FA-EM@PDA/DOX&ICG nanoparticles effectively evaded engulfment by RAW264.7 cells, indicating continued superior stealth performance for circulation and successful evasion of immune system clearance in vivo . For the FA-EM@PDA/DOX&ICG nanoparticles to achieve highly efficient anti-tumor effects, effective uptake by tumor cells is crucial. 45 The FA modification on the EM surface enables specific binding to folate acid receptors (FAR) on tumor cells, thereby enhancing the uptake of FA-EM@PDA/DOX&ICG by tumor cells. 46 Co-incubation of FA-EM@PDA/DOX&ICG nanoparticles with 4T1 and GES-1 cells revealed approximately fourfold higher fluorescence signal intensity in 4T1 cells compared to GES-1 cells (Fig. 3 b). This confirms the active targeting delivery capability of FA-EM@PDA/DOX&ICG nanoparticles. These results indicated that FA-EM@PDA/DOX&ICG nanoparticles could specifically bind with 4T1 cells, and then increasing their phagocytosis by 4T1 cells. Drug Release and In Vitro Combined Therapy of FA-EM@PDA/DOX&ICG Nanoparticles Although FA-EM@PDA/DOX&ICG nanoparticles can be effectively taken up by 4T1 cells, the efficient release of loaded drugs within 4T1 cells is a prerequisite for exerting anti-tumor effects. Therefore, we simulated the tumor microenvironment (TME) in vitro to analyze the release behavior of DOX and ICG from FA-EM@PDA/DOX&ICG nanoparticles. As shown in Fig. 4 a and 4 b, when FA-EM@PDA/DOX&ICG nanoparticles were immersed in mimicking normal physiological environment (pH = 7.4), the release of DOX and ICG after 24 h was represented for about 8.31% and 8.56%, respectively. However, when FA-EM@PDA/DOX&ICG nanoparticles immersed in acidic conditions (pH = 5.5) condition, approximately 95.1% and 95.9% of DOX and ICG were respectively released from FA-EM@PDA/DOX&ICG nanoparticles after 24 h. This release was attributed to drugs are prone to protonation under acidic conditions, leading to a weakened electrostatic interaction between drugs and FA-EM@PDA/DOX&ICG nanoparticles, and then leading drugs releasing. 47–48 Additionally, when FA-EM@PDA/DOX&ICG nanoparticles were immersed in GSH and H 2 O 2 solutions, a minor amount of drug release occurred due to the destabilization of PDA surface properties in an oxidative stress microenvironment. 49 When FA-EM@PDA/DOX&ICG nanoparticles were immersed in an in vitro simulated TME (pH = 5.5, 10 mM GSH, 30 µM H 2 O 2 ), drug can completely release occurred around 20 h. Therefore, FA-EM@PDA/DOX&ICG nanoparticles have environment-responsive behavior. Due to FAR on the surface of 4T1 cells can specifically bind with FA molecules on the surface of FA-EM@PDA/DOX&ICG nanoparticles, this effectively promoted the uptake of FA-EM@PDA/DOX&ICG nanoparticles by 4T1 cells. Using CCK-8 experiments to compare the viability of GES-1 cells and 4T1 cells, the combined chemo/photothermal anti-tumor effects of FA-EM@PDA/DOX&ICG nanoparticles were evaluated. The Fig. 4 c and 4 d shown that with an increase of FA-EM@PDA/DOX&ICG nanoparticles concentration, the viability of both GES-1 cells and 4T1 cells were gradually decreased. When GES-1 cells and 4T1 cells were exposed under NIR (9 min, 1.5 W/cm 2 ) irradiattion at the same concentration and environment, the cells viability of 4T1 cells was significantly lower than that of GES-1 cells. At the high concentration of 200 µg/mL FA-EM@PDA/DOX&ICG nanoparticles, the cells viability of GES-1 cells remained at around 42.6%, while that of 4T1 cells was only about 19.8%. This result indicated that the synergistic chemo/photothermal therapy of FA-EM@PDA/DOX&ICG nanoparticles have a good cytotoxic effect for tumor cells. This effect is mainly attributed to the specific binding of FA molecule on the surface of FA-EM@PDA/DOX&ICG nanoparticles with FAR on tumor cells, increasing the uptake of FA-EM@PDA/DOX&ICG nanoparticles by tumor cells and releasing loaded drugs. Therefore, under NIR irradiation, the constructed FA-EM@PDA/DOX&ICG nanoparticles successfully achieved combined chemo/photothermal therapy, effectively killing tumor cells and reducing damage to normal cells. Synergistic In Vivo Antitumor Effects of FA-EM@PDA/DOX&ICG Nanoparticles with NIR Treatment The synergistic therapy effect of FA-EM@PDA/DOX&ICG nanoparticles with NIR therapy has demonstrated effectively inhibition 4T1 cells growth, prompting us to further investigate their in vivo antitumor efficacy. Firstly, the in vivo photothermal conversion performance of FA-EM@PDA/DOX&ICG nanoparticles should be evaluated before animal experiments. Once different photothermal material (PBS, PDA, ICG, PDA/DOX&ICG and FA-EM@PDA/DOX&ICG nanoparticles) were injected into mice via the tail vein and then exposed under NIR irradiation (1.5 W/cm 2 , 9 min) after 24 h later. As shown in Fig. 5 a and 5 b, FA-EM@PDA/DOX&ICG nanoparticles have the highest photothermal conversion property among these materials as well as the temperature rising into 59.9 ℃, which was ascribed into the FA-EM@PDA/DOX&ICG nanoparticles have targeting properties and has ability to avoid being cleared by the immune system during the blood circulation. Subsequently, to further verify the anti-tumor performance of FA-EM@PDA/DOX&ICG nanoparticles in vivo synergistic therapy. 4T1 tumor-bearing mice were randomly divided into six groups and intravenously injected with PBS, DOX, ICG, PDA, PDA/DOX&ICG, and FA-EM@PDA/DOX&ICG nanoparticles. After 24 h, mice in each treatment group were exposed under NIR (1.5 W/cm 2 , 9 min) irradiation every 3 days for a total of 6 treatments. Mouse weight were recorded, and tumor changes were measured throughout the course of the treatment. As shown in Fig. 5 c, 5 d and 5 e, compared to the PBS treatment group, all treatment groups exhibited varying degrees of tumor size inhibition. Notably, the FA-EM@PDA/DOX&ICG + NIR treatment group displayed a more pronounced inhibitory effect on tumors than the PDA/DOX&ICG + NIR treatment group. This enhanced effect was attributed to the ability of FA-EM@PDA/DOX&ICG nanoparticles to effectively evade clearance by the body's immune system in the bloodstream. Upon reaching the tumor tissue, FA molecules on the surface of FA-EM@PDA/DOX&ICG nanoparticles bind specifically with FAR on 4T1 cells, facilitating the uptake of nanoparticles by 4T1 cells. Under NIR irradiation, the temperature of the tumor tissue rapidly increased, demonstrating superior antitumor efficacy of FA-EM@PDA/DOX&ICG nanoparticles. However, 4T1 tumor-bearing mice treated with DOX alone experienced a significant impact on mental status, including symptoms such as weight loss, disoriented gaze, dehydration, and the mice died completely on day 21 (Fig. 5 f and 5 d). In contrast, after FA-EM@PDA/DOX&ICG + NIR synergistic therapy, not only can effectively inhibit tumor growth, but also the mental state of the mice was maintained well and the survival time of the mice was effectively prolonged during the therapy process. Besides, tumor tissues and other major organ (heart, liver, spleen, lung, kidney) were collected and stained with H&E and TUNEL for histopathological analysis. As shown in Fig. 6a, in comparison to the PBS + NIR treatment group, where no tumor cells necrosis or apoptosis were observed, but the treatment groups with DOX, ICG + NIR, PDA + NIR, PDA/DOX&ICG + NIR, and FA-EM@PDA/DOX&ICG + NIR exhibited varying degrees of tumor cells death and apoptosis. While ICG + NIR and PDA + NIR treatments could inhibit tumor growth, a significant number of apoptotic tumor cells were only observed on the surface of the tumor tissue. Although DOX effectively killed tumor cells deep within the tumor tissue, it brought about substantial side effects to normal tissues, particularly causing damage to the normal structure of the heart (Fig. 6b). However, after the synergistic photothermal therapy with FA-EM@GO-MOF/DOX nanoparticles, abundant apoptotic tumor cells were observed both on the surface and deep within the tumor tissue. This was evidenced by H&E staining results showing extensive nuclear condensation and dissolution, and TUNEL staining results displaying widespread green fluorescence signals across the field of view. Importantly, major mouse organs did not exhibit pathological changes (Fig. 6a and 6b). These results underscore that the synergistic photothermal therapy with FA-EM@GO-MOF/DOX nanoparticles demonstrates significant inhibitory and cytotoxic effects on tumor tissues. Furthermore, the nanoparticles exhibit superior biocompatibility to normal tissues, and effectively mitigating the adverse effects brought about by the antitumor drug DOX during the treatment process were ameliorated. Conclusion The photo/thermal FA-EM@PDA/DOX&ICG nanoparticles designed in this study demonstrated superior photothermal conversion performance and enable precise and controllable drug release. Surface modification of FA-EM@PDA/DOX&ICG nanoparticles with FA-EM can effectively prevent their clearance by the body's immune system during in vivo transport, resulting in an extended blood circulation period. Upon reaching the tumor tissue, the nanoparticles exploit their surface FA molecules, forming selective bonds with FAR on tumor cells and consequently augmenting cellular uptake. In response to the specific TME, FA-EM@PDA/DOX&ICG nanoparticles can release loaded DOX and ICG, achieving on-demand and controlled drug release. Under NIR irradiation, FA-EM@PDA/DOX&ICG nanoparticles exhibit superior photothermal conversion efficiency. Moreover, the adsorption of ICG effectively enhances the photothermal conversion performance of FA-EM@PDA/DOX&ICG, and DOX's penetration into tumor tissues compensates for NIR penetration deficiencies, promoting the destruction of deep-seated tumor cells and effectively inhibiting tumor growth. Importantly, FA-EM@PDA/DOX&ICG nanoparticles, constructed based on the neurotransmitter dopamine, exhibit excellent biocompatibility and mitigate the toxic side effects of DOX. In conclusion, the environmentally responsive treatment platform developed in this study achieves combined photothermal therapy and chemotherapy through a one-step approach. This platform effectively addresses the shortcomings of individual treatments, thereby enhancing therapeutic efficacy. It also provides valuable insights for the design of more intelligent and efficient nanoplatforms for tumor treatment in the future. Declarations Ethics approval and consent to participate All experimental procedures were approved by the Institutional Animal Care and Use Committee of the Medical College of Nanjing University. Consent for publication Each coauthor has read the manuscript and approves its submission. This work is being submitted exclusively to your journal. Data Availability Statement The datasets supporting the results of this article are included within the article. Competing interests The authors declared that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper. Funding This research was supported by the “Jiangsu Provincial Postdoctoral Excellence Program” (2023ZB570). Authors' contributions Chunmin Deng responsible for the experimental operations, data collection and analysis. Hao Zhang data processing and image generation. Li Song is responsible for the experimental design and writing of the manuscript. All authors read and approved the final manuscript. Acknowledgements The acknowledged are included within the article. References Hyuna Sung, Jacques Ferlay, Rebecca L Siegel, et al. Global cancer statistics 2020: GLOBOCAN estimates of incidence and mortality worldwide for 36 cancers in 185 countries. CA. Cancer J. Clin., 2021.71: 209–249. 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Mitochondria-targeting polydopamine nanoparticles to deliver doxorubicin for overcoming drug resistance. ACS. Appl. Mater. Interfaces, 2017. 9: 16793–16802. Scott S Ruppel, Junfeng Liang. Tunable properties of polydopamine nanoparticles and coated surfaces. Langmuir, 2022. 38: 5020–5029. Qiwei Tian, Feiran Jiang, Rujia Zou, et al. Hydrophilic Cu 9 S 5 nanocrystals: a photothermal agent with a 25.7% heat conversion efficiency for photothermal ablation of cancer cells in vivo. ACS Nano., 2011. 5: 9761–9771. Xuemeng Wang, Shaoshan Pan, Lin Chen, et al. Biogenic copper selenide nanoparticles for near-infrared photothermal therapy application. ACS Appl. Mater. Interfaces, 2023. 15: 27638–27646. Yuan Yong, Liangjun Zhou, Zhanjun Gu, et al. WS 2 nanosheet as a new photosensitizer carrier for combined photodynamic and photothermal therapy of cancer cells. Nanoscale, 2014. 6: 10394–10403. Yuan Yong, Liangjun Zhou, Zhanjun Gu, et al. MnOx nanospikes as nanoadjuvants and immunogenic cell death drugs with enhanced antitumor immunity and antimetastatic effect. Angew. Chem. Int. Ed. Engl., 2020. 59: 16381–16384. Weiwei Gao, Cheming J Hu, Ronnie H Fang, et al. Surface functionalization of gold nanoparticles with red blood cell membranes. Adv. Mater., 2013. 25:3549–3553. Hui Ding, Yanlin Lv, Dezhi Ni, et al. Erythrocyte membrane-coated NIR-triggered biomimetic nanovectors with programmed delivery for photodynamic therapy of cancer. Nanoscale, 2015. 7: 9806–9815. Narsireddy Amreddy, Anish Babu, Ranganayaki Muralidharan, et al. Recent advances in nanoparticle-based cancer drug and gene delivery. Adv. Cancer Res., 2018. 137: 115–170. Wei Zhang, Zi-Li Yu, Min Wu, et al. Magnetic and folate functionalization enables rapid isolation and enhanced tumor-targeting of cell-derived microvesicles. ACS Nano., 2017. 11: 277–290. Jinfeng Zeng, Dongjian Shi, Yanglin Gu, et al. Injectable and near-infrared-responsive hydrogels encapsulating dopamine-stabilized gold nanorods with long photothermal activity controlled for tumor therapy. Biomacromolecules, 2019. 20: 3375–3384. Christopher E Grant, Amy L Flis, Bríd M Ryan. Understanding the role of dopamine in cancer: past, present and future. Carcinogenesis, 2022. 43: 517–527. Hong Li, Yi Jia, Haonan Peng, Junbai Li. Recent developments in dopamine-based materials for cancer diagnosis and therapy. Adv. Colloid Interface Sci., 2018. 252: 1–20. Scheme Scheme 1 is available in the Supplementary Files section Additional Declarations No competing interests reported. Supplementary Files floatimage1.png Scheme 1. (a) The construction of FA-EM@PDA/DOX&ICG nanoparticles through drug loading and erythrocyte membrane coating. (b) Proposed action mechanism of FA-EM@PDA/DOX&ICG nanoparticles in a mouse tumor model. Cite Share Download PDF Status: Posted Version 1 posted You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. We do this by developing innovative software and high quality services for the global research community. Our growing team is made up of researchers and industry professionals working together to solve the most critical problems facing scientific publishing. Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-3844904","acceptedTermsAndConditions":true,"allowDirectSubmit":true,"archivedVersions":[],"articleType":"Research Article","associatedPublications":[],"authors":[{"id":266367951,"identity":"5a453700-7caa-49a2-a80a-c61c8859f0bd","order_by":0,"name":"Chunmin Deng","email":"","orcid":"","institution":"Suzhou University of Science and Technology","correspondingAuthor":false,"prefix":"","firstName":"Chunmin","middleName":"","lastName":"Deng","suffix":""},{"id":266367952,"identity":"0111cc14-de23-433e-9a0c-85c2cf6249fa","order_by":1,"name":"Hao Zhang","email":"","orcid":"","institution":"Yancheng First People's Hospital","correspondingAuthor":false,"prefix":"","firstName":"Hao","middleName":"","lastName":"Zhang","suffix":""},{"id":266367953,"identity":"fa4a74f1-13c1-4e6e-8ee4-8ae69baae584","order_by":2,"name":"Li Song","email":"data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAZAAAAAyAQMAAABI0h/eAAAABlBMVEX///8AAABVwtN+AAAACXBIWXMAAA7EAAAOxAGVKw4bAAAA4klEQVRIie3RIQvCQBTA8Xcc3Mq51YMF/QLCk4EYxj7L4ODSwqJJhIFJsZr8DH6E4aEW0WowKIJ5ccHgNFiUnTbD/fP9ON57ADbbP0a986VEETEv06cCw8hMHE4DSHvSFWvVmaVKmonHmQ9Fn8whafm8WJKhSbSzRu6nKCiDLQQh5hQcvVrUka5242CGwmV0kl8SPLrAlTrUE8CYP3/ZxUGCVwqCd40krwgZQYJ+DzUZmgnvZC8C3xEmSTWLZI8lj1FJZpxlrzdlcRtEzWl1yvIWRp6j17XkPfbbc5vNZrN96g78kEdKDBPkgAAAAABJRU5ErkJggg==","orcid":"","institution":"Yancheng First People's Hospital","correspondingAuthor":true,"prefix":"","firstName":"Li","middleName":"","lastName":"Song","suffix":""}],"badges":[],"createdAt":"2024-01-08 08:37:11","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-3844904/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-3844904/v1","draftVersion":[],"editorialEvents":[],"editorialNote":"","failedWorkflow":false,"files":[{"id":49550590,"identity":"27858d48-bb4b-4c62-9eae-6777385c1e08","added_by":"auto","created_at":"2024-01-12 20:34:27","extension":"png","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":359627,"visible":true,"origin":"","legend":"\u003cp\u003eCharacterization of FA-EM@PDA/DOX\u0026amp;ICG nanoparticles. TEM image of (a) PDA and (b) FA-EM@PDA/DOX\u0026amp;ICG nanoparticles. (c) Size distribution of PDA, PDA/DOX\u0026amp;ICG and FA-EM@PDA/DOX\u0026amp;ICG nanoparticles. (d) Zeta potential of PDA, PDA/DOX\u0026amp;ICG and FA-EM@PDA/DOX\u0026amp;ICG nanoparticles. (e) UV-Vis spectroscopy of ICG, PDA, DOX, PDA/ICG\u0026amp;DOX, FA-EM, and FA-EM@PDA/DOX\u0026amp;ICG nanoparticles. (f) Protein analysis of protein marker (1), EM (2), PDA (3), FA-EM (4), PDA/ICG\u0026amp;DOX (5), and FA-EM@PDA/DOX\u0026amp;ICG nanoparticles (6) by using SDS-PAGE. Samples were stained with Coomassie Brilliant.\u003c/p\u003e","description":"","filename":"floatimage2.png","url":"https://assets-eu.researchsquare.com/files/rs-3844904/v1/348e602b354e72134c5142fc.png"},{"id":49550883,"identity":"25d73d90-f27a-4c0d-a1ef-aa936cee5ccd","added_by":"auto","created_at":"2024-01-12 20:42:28","extension":"png","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":271638,"visible":true,"origin":"","legend":"\u003cp\u003e(a) Thermal imaging and (b) corresponding temperature changes of PBS (1), 200 μg/mL PDA (2), 200 μg/mL PDA/ICG\u0026amp;DOX (3), and 200 μg/mL FA-EM@PDA/ICG\u0026amp;DOX (4) irradiated with 808 nm irradiation (1.5 W/cm\u003csup\u003e2\u003c/sup\u003e) for 9 min, and 200 μg/mL FA-EM@PDA/ICG\u0026amp;DOX (5) irradiated with NIR irradiation (2.0 W/cm\u003csup\u003e2\u003c/sup\u003e) for 9 min. (c) Temperature changes of 200 μg/mL FA-EM@PDA/ICG\u0026amp;DOX disperse solution irradiation with NIR irradiation (2.0 W/cm\u003csup\u003e2\u003c/sup\u003e) for four turn on/off cycles. (d) Photothermal response of FA-EM@PDA/ICG\u0026amp;DOX solution were treated with an NIR irradiation (808 nm, 1.5 W/cm\u003csup\u003e2\u003c/sup\u003e) for 600 s and then the irradiation was turned off.\u003c/p\u003e","description":"","filename":"floatimage3.png","url":"https://assets-eu.researchsquare.com/files/rs-3844904/v1/0f4e9e29052479e2e66f077c.png"},{"id":49550592,"identity":"dd24fcc6-40d5-4646-91c1-132fbea96a7d","added_by":"auto","created_at":"2024-01-12 20:34:28","extension":"jpeg","order_by":3,"title":"Figure 3","display":"","copyAsset":false,"role":"figure","size":373793,"visible":true,"origin":"","legend":"\u003cp\u003e(a) CLSM images of macrophage cells (RAW264.7 cells) incubated with EM@PDA/DOX\u0026amp;ICG, FA-EM@PDA/DOX\u0026amp;ICG, and PDA/DOX\u0026amp;ICG nanoparticles for 3 h. (b) CLSM images of 4T1 and GES-1 cells incubated with FA-EM@PDA/DOX\u0026amp;ICG nanoparticles for 3 h. Images show cell nuclei stained by DAPI (blue), ICG fluorescence in cells (green), DOX fluorescence in cells and the merged overlap of the three images. Scale bars: 30 μm\u003c/p\u003e","description":"","filename":"floatimage4.jpeg","url":"https://assets-eu.researchsquare.com/files/rs-3844904/v1/b5d32b638a9c60e3ee96d0ab.jpeg"},{"id":49550596,"identity":"45fc349d-dd0e-48a9-8ac4-8882404c52a4","added_by":"auto","created_at":"2024-01-12 20:34:28","extension":"png","order_by":4,"title":"Figure 4","display":"","copyAsset":false,"role":"figure","size":70279,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cem\u003eIn vitro\u003c/em\u003e profiles of (a) DOX and (b) ICG release behavior from FA-EM@PDA/DOX\u0026amp;ICG nanoparticles under conditions of pH 7.4, pH 5.5, H\u003csub\u003e2\u003c/sub\u003eO\u003csub\u003e2\u003c/sub\u003e, GSH and simulated TME (pH = 5.5, 10 mM GSH, 30 μM H\u003csub\u003e2\u003c/sub\u003eO\u003csub\u003e2\u003c/sub\u003e). Viability of (c) GES-1 cells and (d) 4T1 cells upon treated with PBS, PDA, PDA/DOX\u0026amp;ICG, and FA-EM@PDA/DOX\u0026amp;ICG nanoparticles under NIR irradiation (1.5 W/cm\u003csup\u003e2\u003c/sup\u003e, 9 min).\u003c/p\u003e","description":"","filename":"floatimage5.png","url":"https://assets-eu.researchsquare.com/files/rs-3844904/v1/184d1556759b9b15d0e59999.png"},{"id":49550594,"identity":"0adf14ea-f520-44cc-9306-a01ecbb0a166","added_by":"auto","created_at":"2024-01-12 20:34:28","extension":"png","order_by":5,"title":"Figure 5","display":"","copyAsset":false,"role":"figure","size":427057,"visible":true,"origin":"","legend":"\u003cp\u003e(a) Infrared thermal images of mice at different time points of varied treatments under NIR irradiation (1.5 W/cm\u003csup\u003e2\u003c/sup\u003e, 9 min). (b) Temperature change curves at tumor sites of mice in different groups under NIR irradiation. (c) The tumor volumes in six groups after different treatment. (d) Mean tumor weights on day 15 after the last treatment and (e) photographs of the tumors after 15 day different treatment. PBS (1), DOX (2), ICG + NIR (3), PDA + NIR (4), PDA/DOX\u0026amp;ICG + NIR (5), FA-EM@PDA/DOX\u0026amp;ICG + NIR (6). (f) Body weight of mice in different groups during 15 days treatment. (g) The survival curves of mice in the different treatment groups.\u003c/p\u003e","description":"","filename":"floatimage6.png","url":"https://assets-eu.researchsquare.com/files/rs-3844904/v1/10ab24eec8e86054b0944125.png"},{"id":49550591,"identity":"e9acdaeb-9664-4e6e-8a8f-ce21a9a009d2","added_by":"auto","created_at":"2024-01-12 20:34:27","extension":"png","order_by":6,"title":"Figure 6","display":"","copyAsset":false,"role":"figure","size":5713,"visible":true,"origin":"","legend":"\u003cp\u003eFigure legend not available with this version.\u003c/p\u003e","description":"","filename":"placeholderimage.png","url":"https://assets-eu.researchsquare.com/files/rs-3844904/v1/4366479b544963b2581a80e7.png"},{"id":54130577,"identity":"82a4ccf8-bd8c-43ae-8103-85c4d218b635","added_by":"auto","created_at":"2024-04-05 04:25:03","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":1759672,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-3844904/v1/eb46d4bc-b271-4b8d-90ff-34544af46ddb.pdf"},{"id":49550595,"identity":"bbf69b3d-dcfd-48d5-a124-bec15a8c44bf","added_by":"auto","created_at":"2024-01-12 20:34:28","extension":"png","order_by":1,"title":"","display":"","copyAsset":false,"role":"supplement","size":493129,"visible":true,"origin":"","legend":"\u003cp\u003eScheme 1. (a) The construction of FA-EM@PDA/DOX\u0026amp;ICG nanoparticles through drug loading and erythrocyte membrane coating. (b) Proposed action mechanism of FA-EM@PDA/DOX\u0026amp;ICG nanoparticles in a mouse tumor model.\u003c/p\u003e","description":"","filename":"floatimage1.png","url":"https://assets-eu.researchsquare.com/files/rs-3844904/v1/501641677d47680a0a2ab4d9.png"}],"financialInterests":"No competing interests reported.","formattedTitle":"Environment-Responsive Dopamine Nanoplatform for Tumor Synergistic Therapy","fulltext":[{"header":"Introduction","content":"\u003cp\u003eTumor remains one of the primary threats to human health, current clinical treatments mainly revolve around chemotherapy, radiotherapy, immunotherapy, and surgical resection.\u003csup\u003e1\u0026ndash;2\u003c/sup\u003e While these treatments effectively suppress tumor growth and extend patient survival, they lack precision in targeting tumors and often induce substantial toxic side effects in the body's normal tissues.\u003csup\u003e3\u0026ndash;4\u003c/sup\u003e Therefore, the development of a novel type of treatment is urgently needed to enhance tumor treatment efficacy while minimizing damage to healthy tissues.\u003c/p\u003e \u003cp\u003eIn recent years, various innovative therapies and strategies, such as targeted drugs, photothermal therapy, and starvation therapy, have emerged.\u003csup\u003e5\u0026ndash;6\u003c/sup\u003e These approaches not only enable precise and controllable treatment of tumors but also reduce the toxic side effects to body normal tissues and organs, significantly enhancing patients\u0026rsquo; compliance.\u003csup\u003e7\u003c/sup\u003e Studies have revealed that tumor tissues lack sufficient blood supply, making them unable to regulate heat dissipation through blood flow and velocity.\u003csup\u003e8\u003c/sup\u003e Consequently, tumor tissues exhibit greater sensitivity to heat than normal tissues, leading to the term \"natural heat reservoirs\" for tumor tissues, presenting a natural advantage for photothermal therapy (PTT) in treating tumors.\u003csup\u003e8\u0026ndash;9\u003c/sup\u003e Additionally, PTT offers temporal and spatial controllability, low invasiveness and high selectivity, making it an optimal approach for tumor treatment.\u003csup\u003e10\u003c/sup\u003e However, standalone PTT often requires high temperatures to induce tumor cells death, which risks damaging surrounding normal tissues. In recent years, with the advancement of photothermal nanoparticles, these limitations have been addressed. When these nanoparticles aggregate within tumor tissues, and then exposed under near-infrared light (NIR) irradiation. Nanoparticles can convert light energy into heat, and then rapidly elevating local tumor tissue temperatures to induce tumor cells apoptosis and reduce damage to surrounding normal tissues.\u003csup\u003e11\u003c/sup\u003e\u003c/p\u003e \u003cp\u003eNonetheless, nanoparticles are prone to clearance by body's immune system during their transport within the body, especially through phagocytosis by macrophages, resulting in shortened blood circulation periods and hindered them accumulated at tumor tissues.\u003csup\u003e12\u003c/sup\u003e To tackle this issue effectively, Fang et al. utilized folate acid-modified erythrocyte membranes to coat metal-organic frameworks and then loading with chemotherapeutic drug doxorubicin for tumor combination treatment.\u003csup\u003e13\u003c/sup\u003e This system of diagnosis and treatment not only effectively avoided phagocytosis by macrophages to prolonging their circulation in the bloodstream, but also with folate acid molecule (FA) modifications on their surfaces that can enhance phagocytosis of biomimetic nanoparticles by tumor cells and achieved controlled drug release according to the specific tumor microenvironment (TME). However, the limited penetration of NIR light resulting in its inability to eradicate deep-seated tumor tissues entirely. To achieve completely tumor cells eradication, researchers have exploited the inherent properties of photothermal nanoparticles to load chemotherapeutic drugs.\u003csup\u003e14\u003c/sup\u003e Upon reaching tumor tissues, these nanoparticles releasing the loaded chemotherapy drugs based on the specific TME, promoting penetration of the drugs into deep-seated tumor tissues while reducing their systemic toxicity during transport.\u003csup\u003e15\u003c/sup\u003e Consequently, a plethora of combined therapies based on PTT, such as PTT/chemotherapy and PTT/photodynamic therapy/chemotherapy, effectively compensate for the limitations of individual treatment methods and achieve efficient tumor clearance.\u003csup\u003e16\u0026ndash;17\u003c/sup\u003e To enhance the biocompatibility of photothermal nanoparticles, this study ingenious utilized the neurotransmitter-dopamine is prone to oxidative polymerization to form the photothermal material polydopamine (PDA) under alkaline conditions, to achieve tumor photothermal therapy.\u003csup\u003e18\u003c/sup\u003e To improve the photothermal performance and therapeutic efficacy of PDA, doxorubicin (DOX), a chemotherapeutic drug, and indocyanine green (ICG), a photothermal material, are loaded into PDA surface (PDA/DOX\u0026amp;ICG) due to its surface adsorption properties.\u003csup\u003e19\u003c/sup\u003e And then PDA/DOX\u0026amp;ICG nanoparticles are then enveloped with folate-modified erythrocyte membranes (FA-EM) to obtain FA-EM@PDA/DOX\u0026amp;ICG, which can effectively avoid FA-EM@PDA/DOX\u0026amp;ICG nanoparticles being cleared by the body's immune system, thereby prolonging their blood circulation cycle (Scheme \u003cspan refid=\"Sch1\" class=\"InternalRef\"\u003e1a\u003c/span\u003e). Upon reaching at tumor tissues, they can readily bind with folate acid receptors specifically located on the surface of tumor cells, enhancing uptake by tumor cells and facilitating DOX and ICG in-site release based on the specific TME (Scheme \u003cspan refid=\"Sch1\" class=\"InternalRef\"\u003e1b\u003c/span\u003e). When NIR is irradiated, the ICG present in the FA-EM@PDA/DOX\u0026amp;ICG nanoparticles can effectively improve the photothermal conversion efficiency, and the penetration of DOX into tumor tissue can compensate for the lack of NIR tissue penetration, thereby inhibiting tumor growth and alleviate the systemic toxic effects of DOX.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e"},{"header":"Materials and Methods","content":"\u003cp\u003eMaterials\u003c/p\u003e \u003cp\u003eDopamine, polyvinyl alcohol (PVA) and KMnO\u003csub\u003e4\u003c/sub\u003e were purchased from Beijing Guoyao (Beijing, China). Doxorubicin (DOX), indocyanine green (ICG), DAPI, and CCK-8 were obtained from Shanghai Aladdin Reagent Co., Ltd (Shanghai, China). PBS, 1640 culture medium, and fetal bovine serum were sourced from Sigma-Aldrich Chemicals (Madison, USA). All chemical reagents used in the experiments were not further purified, and ultrapure water was used throughout the entire experimental process.\u003c/p\u003e \u003cp\u003ePreparation of Polydopamine\u003c/p\u003e \u003cp\u003eThe synthesis of polydopamine (PDA) followed by a modified version of the laboratory's previous procedure.\u003csup\u003e20\u0026ndash;21\u003c/sup\u003e Initially, 5 mg/mL polyvinyl alcohol (PVA) was added into 10 mL ultrapure water and heated to 90\u0026deg;C to completely dissolve PVA. After cooling the PVA solution to room temperature, 0.2 g dopamine and 1 mL KMnO\u003csub\u003e4\u003c/sub\u003e (1 mg/mL) were separately added to the solution, followed by stirring for 2 h at room temperature. The solution was then centrifuged at 3500 rpm for 20 min to collect the supernatant, which was subsequently placed in a dialysis bag and dialyzed for 3 days. Finally, the resulting sample of PDA nanoparticles was freeze-dried and stored for later use.\u003c/p\u003e \u003cp\u003ePreparation of PDA/DOX\u0026amp;ICG\u003c/p\u003e \u003cp\u003e20 mg PDA nanoparticles were dispersed in 10 mL ultrapure water to achieve thorough dispersion.\u003csup\u003e22\u003c/sup\u003e Subsequently, 10 mg of DOX and 10 mg of ICG were separately added to the dispersed solution under light-avoiding conditions and stirred for 24 h. The resulting mixed solution was collected, placed in a dialysis bag, and dialyzed for 3 days. Finally, the obtained PDA/DOX\u0026amp;ICG nanoparticles were freeze-dried and stored for later use.\u003c/p\u003e \u003cp\u003ePreparation of FA-EM@PDA/DOX\u0026amp;ICG\u003c/p\u003e \u003cp\u003eFresh blood was collected from BALB/c mice through eye extraction and placed in an anticoagulant tube. After centrifugation (5000 rpm, 10 min) to remove unwanted cells and serum, the collected erythrocyte membranes (EM) were washed several times with PBS until the supernatant was colorless. The EM were gently mixed with ultrapure water and left to stand at 4\u0026deg;C for 1 h to release hemoglobin and other related components. The cells were then washed with PBS until the supernatant was colorless and transparent. Next, 20 mg of DSPE-PEG-FA was added to 5 mL of the EM solution and stirred for 24 h. The obtained DSPE-PEG-FA modified EM (FA-EM) were collected after centrifugation and washed three times with PBS. Finally, the collected FA-EM was resuspended in 1 mL PBS, and 1 mg of PDA/DOX\u0026amp;ICG was added to the solution. This mixture was extruded through a 220 nm polycarbonate porous membrane using a squeezing method to obtain FA-EM@PDA/DOX\u0026amp;ICG nanoparticles. The nanoparticles were collected by centrifugation, freeze-dried, and stored at -80\u0026deg;C.\u003csup\u003e23\u0026ndash;24\u003c/sup\u003e\u003c/p\u003e \u003cp\u003eCharacterization of FA-EM@PDA/DOX\u0026amp;ICG Nanoparticles\u003c/p\u003e \u003cp\u003eFA-EM@PDA/DOX\u0026amp;ICG nanoparticles were dispersed in deionized water and sonicated for 10 minutes. The dispersed solution was then dropped into a copper grid, air-dried, and observed under transmission electron microscopy (TEM) to determine the size and shape of the nanoparticles. Changes in the zeta potential and particle size of FA-EM@PDA/DOX\u0026amp;ICG nanoparticles before and after drug loading were analyzed using a particle size analyzer. The successful preparation of FA-EM@PDA/DOX\u0026amp;ICG nanoparticles was further validated by UV-spectrophotometry. Protein gel electrophoresis was used to analyze the expression of the EM protein profile on the surface of the prepared nanoparticles.\u003c/p\u003e \u003cp\u003eCell Culture\u003c/p\u003e \u003cp\u003eMouse 4T1 breast cancer cells, human macrophages (RAW264.7), and human gastric mucosal cells (GES-1) were all obtained from Nanjing University (Nanjing, China) and cultured in a 37\u0026deg;C, 5% CO\u003csub\u003e2\u003c/sub\u003e incubator. The cell culture medium used was RPMI 1640 medium (Gibco) containing 10% fetal bovine serum (FBS) (Gibco, Shanghai, China). All cell lines were approved by the Institutional Animal Care and Use Committee of Nanijing University.\u003c/p\u003e \u003cp\u003ePhotothermal Performance of FA-EM@PDA/DOX\u0026amp;ICG Nanoparticles\u003c/p\u003e \u003cp\u003eInitially, 1 mg of FA-EM@PDA/DOX\u0026amp;ICG nanoparticles were dispersed in 1 mL PBS buffer solution and thoroughly mixed. Subsequently, varying volumes of FA-EM@PDA/DOX\u0026amp;ICG nanoparticles were aspirated and added to different volumes of PBS buffer solution to prepare suspensions with different concentrations of FA-EM@PDA/DOX\u0026amp;ICG dispersion solution. Then, the prepared dispersion solution of FA-EM@PDA/DOX\u0026amp;ICG nanoparticles were exposed under different NIR powers. At predetermined time points, the temperature changes for each group were recorded using an infrared thermal imaging instrument.\u003csup\u003e25\u003c/sup\u003e\u003c/p\u003e \u003cp\u003eThe photothermal conversion efficiency of FA-EM@PDA/DOX\u0026amp;ICG nanoparticles was evaluated according to the relevant methods.\u003csup\u003e26\u0026ndash;27\u003c/sup\u003e Initially, 100 \u0026micro;l of FA-EM@PDA/DOX\u0026amp;ICG (200 \u0026micro;g/ml) dispersion solution was added to an Eppendorf tube (EP) and placed under 808 nm (1.5 W/cm\u003csup\u003e2\u003c/sup\u003e) NIR irradiation for 10 minutes. After turning off the NIR light for 10 minutes, irradiation was resumed for another 10 minutes. The temperature changes at each time point were recorded using an infrared thermal image. The photothermal conversion efficiency of FA-EM@PDA/DOX\u0026amp;ICG nanoparticles was then assessed:\u003c/p\u003e \u003cp\u003eθ = (T - T\u003csub\u003emin\u003c/sub\u003e)/(T\u003csub\u003emax\u003c/sub\u003e - T\u003csub\u003emin\u003c/sub\u003e) (\u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e)\u003c/p\u003e \u003cp\u003ehS\u0026thinsp;=\u0026thinsp;-\u0026thinsp;m*C\u003csub\u003ep\u003c/sub\u003e/ζ (\u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e)\u003c/p\u003e \u003cp\u003eη\u0026thinsp;=\u0026thinsp;hs (ΔT\u003csub\u003emax,mix\u003c/sub\u003e\u0026thinsp;\u0026minus;\u0026thinsp;ΔT\u003csub\u003emax,H2O\u003c/sub\u003e)/\u003cem\u003eI\u003c/em\u003e(1\u0026thinsp;\u0026minus;\u0026thinsp;10\u003csup\u003e\u0026minus;\u0026thinsp;A808\u003c/sup\u003e) (\u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e)\u003c/p\u003e \u003cp\u003eθ is the dimensionless driving force temperature, and defined as the ratio of (T - T\u003csub\u003emin\u003c/sub\u003e) to (T\u003csub\u003emax\u003c/sub\u003e - T\u003csub\u003emin\u003c/sub\u003e).ζis the slope of T and -Lnθ. Consequently, the m\u003csub\u003eNPs\u003c/sub\u003e and C\u003csub\u003ep, NPs\u003c/sub\u003e of NPs were neglected. m was 2\u0026times;10\u003csup\u003e\u0026minus;\u0026thinsp;4\u003c/sup\u003e kg. C\u003csub\u003ep\u003c/sub\u003e was 4.2\u0026times;10\u003csup\u003e3\u003c/sup\u003e J/(kg\u0026middot;℃). ΔT\u003csub\u003emax,mix\u003c/sub\u003e is the temperature change of the FA-EM@GO-MOF dispersion at the maximum steady-state temperature. ΔT\u003csub\u003emax,H2O\u003c/sub\u003e is the temperature change of water at the maximum steady-state temperature. I is the laser power, A\u003csub\u003e808\u003c/sub\u003e is the absorbance of FA-EM@GO-MOF at the wavelength of 808 nm in aqueous solution.\u003c/p\u003e \u003cp\u003eTargeted Delivery and Immune Escape of FA-EM@PDA/DOX\u0026amp;ICG Nanoparticles\u003c/p\u003e \u003cp\u003eInitially, 4T1 cells and GES-1 cells were separately seeded into cell culture dishes and incubated in a cell culture incubator (37\u0026deg;C, 5% CO\u003csub\u003e2\u003c/sub\u003e) for 12 h. Subsequently, FA-EM@PDA/DOX\u0026amp;ICG nanoparticles were added to the 4T1 cells and GES-1 cells, and then incubated for 3 hours. Afterward, the supernatant was discarded, and fixation was performed using 4% paraformaldehyde for 15 minutes followed by DAPI staining for 20 minutes and several washes with PBS. Finally, the fluorescence intensity of 4T1 cells and GES-1 cells was observed using fluorescence confocal microscopy.\u003csup\u003e28\u003c/sup\u003e\u003c/p\u003e \u003cp\u003eRAW264.7 cells were seeded into cell culture dishes and incubated in a cell culture incubator (37\u0026deg;C, 5% CO\u003csub\u003e2\u003c/sub\u003e) for 12 hours. Subsequently, PDA/DOX\u0026amp;ICG, EM@PDA/DOX\u0026amp;ICG, and FA-EM@PDA/DOX\u0026amp;ICG nanoparticles were separately co-incubated with RAW264.7 cells for 3 hours. After removing the supernatant, fixation was conducted using 4% paraformaldehyde for 15 minutes, followed by DAPI staining for 20 minutes and multiple PBS washes. Finally, the fluorescence intensity of RAW264.7 cells in each group was observed using laser scanning confocal microscopy (LSCM).\u003csup\u003e29\u0026ndash;30\u003c/sup\u003e\u003c/p\u003e \u003cp\u003eAnimal welfare\u003c/p\u003e \u003cp\u003e In this study, all animal experiments were executed according to a protocol approved by the Animal Management Rules of the Ministry of Health of the People\u0026rsquo;s Republic of China and approved by the Institutional Animal Care and Use Committee of Nanijing University. Female 6- to 6-week-old BALB/c mice were purchased from Qinglong Mountain (Nanjing, People\u0026rsquo;s Republic of China) and maintained under specific pathogen-free-conditions. All efforts were made to minimize the animals\u0026rsquo; suffering and to reduce the number of animals used.\u003c/p\u003e \u003cp\u003eConstruction and Treatment of Mouse 4T1 Subcutaneous Xenograft Model\u003c/p\u003e \u003cp\u003eA suspension containing 2\u0026times;10\u003csup\u003e7\u003c/sup\u003e 4T1 cells were injected into the groin of male Balb/c mice to establish a mouse subcutaneous xenograft model. When the size of the 4T1 xenograft tumors reached approximately 100 mm\u003csup\u003e3\u003c/sup\u003e, the 4T1 xenograft mice were randomly divided into 6 groups (n\u0026thinsp;=\u0026thinsp;6/group). These groups were treated with PBS, DOX (1 mg/kg), ICG (1 mg/kg), PDA (1 mg/kg), PDA/DOX\u0026amp;ICG (1 mg/kg), and FA-EM@PDA/DOX\u0026amp;ICG (1 mg/kg), respectively. The required drugs were intravenously injected into the mice via the tail vein, and administered every 3 days. After 24 h injection, the mice were exposed under NIR irradiation (1.5 W/cm\u003csup\u003e2\u003c/sup\u003e, 9 min). The mice's tumor growth and body weight were observed daily (as per animal ethics standards: a decrease in mouse body weight exceeding 15% of their initial weight was considered an indicator of death). The formula for calculating mouse xenograft tumor volume was as follows: V (mm\u003csup\u003e3\u003c/sup\u003e)\u0026thinsp;=\u0026thinsp;0.5 \u0026times; a \u0026times; b\u003csup\u003e2\u003c/sup\u003e, where V represents tumor volume, a represents the longer diameter of the tumor, and b represents the shorter diameter of the tumor. 15 days later, the mice were euthanized, and tumors from each treatment group were collected.\u003csup\u003e31\u0026ndash;32\u003c/sup\u003e\u003c/p\u003e \u003cp\u003eHistological Analysis\u003c/p\u003e \u003cp\u003eFollowing the completion of the treatment, mice from each treatment group were euthanized using cervical dislocation. Major organs (heart, liver, spleen, lungs, and kidneys) and tumor tissues from the 4T1 xenograft mice were collected for histological analysis. The collected tissues were fixed in 4% formaldehyde and subsequently embedded in paraffin. Tissue sections were prepared and stained with hematoxylin and eosin (H\u0026amp;E) after embedding. For tumor tissues, in situ terminal deoxynucleotidyl transferase dUTP nick-end labeling (TUNEL) staining was also performed. The processed tissue sections were placed on a slide scanner to capture tissue images, and the prepared tissue sections were assessed by experienced pathologists.\u003csup\u003e33\u0026ndash;34\u003c/sup\u003e\u003c/p\u003e \u003cdiv id=\"Sec3\" class=\"Section2\"\u003e \u003ch2\u003eStatistical Analysis\u003c/h2\u003e \u003cp\u003eThe experimental data were analyzed using SPSS software (version 17.0) and expressed as mean\u0026thinsp;\u0026plusmn;\u0026thinsp;standard deviation (X\u0026thinsp;\u0026plusmn;\u0026thinsp;S). One-way analysis of variance (ANOVA) was employed for comparing means among multiple groups, while the t-test was further utilized for comparing means between two samples. A significance level of p\u0026thinsp;\u0026lt;\u0026thinsp;0.05 was considered indicative of statistical differences. When p\u0026thinsp;\u0026lt;\u0026thinsp;0.05, it was denoted as \"*\", p\u0026thinsp;\u0026lt;\u0026thinsp;0.01 as \"**\", and p\u0026thinsp;\u0026lt;\u0026thinsp;0.001 as \"***\".\u003c/p\u003e \u003c/div\u003e"},{"header":"Results and Discussion","content":"\u003cp\u003eBiological Characterization of FA-EM@PDA/DOX\u0026amp;ICG Nanoparticles\u003c/p\u003e \u003cp\u003eIn this study, the properties of dopamine undergoing oxidative polymerization to form PDA under alkaline conditions were utilized.\u003csup\u003e35\u003c/sup\u003e Therefore, we analyzed the biological characteristics of PDA, TEM results demonstrated that the prepared PDA nanoparticles were circular, with a particle size for about 70 nm (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003ea and \u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003ec). Besides, utilizing the adsorption properties of the PDA to load therapeutic drugs. Once, DOX and ICG were successfully adsorption into the PDA surface, the resulting PDA/DOX\u0026amp;ICG nanoparticles exhibited an increased particle size and a shift in the zeta potential from \u0026minus;\u0026thinsp;4.1 mV to -1.2 mV (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003ec and \u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003ed). However, when the prepared PDA/DOX\u0026amp;ICG nanoparticles underwent further encapsulation with FA-EM, a thin film covered the surface of PDA/DOX\u0026amp;ICG nanoparticles (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eb). The size and zeta potential of the obtained FA-EM@PDA/DOX\u0026amp;ICG nanoparticles became 122 nm and \u0026minus;\u0026thinsp;6.3 mV, respectively (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eb, \u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003ec and \u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003ed). Subsequently, UV-Vis spectroscopy results confirmed the absorption peaks of the prepared FA-EM@PDA/DOX\u0026amp;ICG nanoparticles, further verified the successful adsorption of DOX and ICG onto the PDA surface, with FA-EM effectively encapsulated on the surface of PDA/DOX\u0026amp;ICG nanoparticles (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003ee). More importantly, polyacrylamide gel electrophoresis was employed to analyze the retention of membrane proteins on the surface of FA-EM@PDA/DOX\u0026amp;ICG nanoparticles. As shown in Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003ef, the protein profile of FA-EM@PDA/DOX\u0026amp;ICG nanoparticles matched well with EM proteins, indicated good retention of membrane proteins after FA-EM encapsulation. These results demonstrated that FA-EM@PDA/DOX\u0026amp;ICG nanoparticles were successfully fabricated and retaining the relevant proteins of EM.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cdiv id=\"Sec5\" class=\"Section2\"\u003e \u003ch2\u003eFA-EM@PDA/DOX\u0026amp;ICG Photothermal Conversion Performance\u003c/h2\u003e \u003cp\u003ePDA, as a typical photothermal material, can convert light energy into heat under NIR irradiation, thereby inducing photothermal ablation of tumors.\u003csup\u003e36\u003c/sup\u003e However, PDA exhibits suboptimal photothermal conversion performance, resulting in inadequate therapeutic outcomes.\u003csup\u003e37\u003c/sup\u003e Hence, in this study, the adsorption capability on the surface of PDA was utilized to load the photosensitizer ICG to enhance the photothermal conversion performance of FA-EM@PDA/DOX\u0026amp;ICG nanoparticles.\u003csup\u003e38\u003c/sup\u003e As illustrated in Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003ea, 200 \u0026micro;g/mL of PDA, PDA/DOX\u0026amp;ICG, and FA-EM@PDA/DOX\u0026amp;ICG nanoparticles dispersion solution were separately placed under NIR irradiation (1.5 W/cm\u003csup\u003e2\u003c/sup\u003e, 9 min), and the temperature changes for each group were recorded at designated time points using an infrared thermal image (Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003eb). After 9 min irradiation, the temperature of PDA, PDA/DOX\u0026amp;ICG, and FA-EM@PDA/DOX\u0026amp;ICG nanoparticles were respectively increased into 46.4 ℃, 55.1 ℃, and 54.8 ℃. These results indicated that the modification of FA-EM@PDA/DOX\u0026amp;ICG nanoparticles with FA-EM did not affect the photothermal conversion efficiency of PDA/DOX\u0026amp;ICG, while the incorporation of the photosensitizer ICG effectively enhanced PDA nanoparticles photothermal conversion performance. However, when 200 \u0026micro;g/mL of FA-EM@PDA/DOX\u0026amp;ICG nanoparticles were exposed under 2.0 W/cm\u003csup\u003e2\u003c/sup\u003e NIR for 9 min, the temperature increased to 59.8 ℃. These findings suggest a positive correlation between the photothermal conversion efficiency of FA-EM@PDA/DOX\u0026amp;ICG nanoparticles and NIR power, and duration.\u003c/p\u003e \u003cp\u003eTo verify the photothermal stability of FA-EM@PDA/DOX\u0026amp;ICG nanoparticles, a solution of 200 \u0026micro;g/mL FA-EM@PDA/DOX\u0026amp;ICG nanoparticles were exposed under 1.5 W/cm\u003csup\u003e2\u003c/sup\u003e NIR for 9 min, followed by turning on/off NIR irradiation for 9 minutes, and repeated four times. The temperature changes were recorded using an infrared thermal image (recorded every 3 minutes). As depicted in the Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003ec, after four cycles of irradiation, there was no significant changes in the temperature of FA-EM@PDA/DOX\u0026amp;ICG nanoparticles, indicating sustained excellent photothermal conversion performance. And the photothermal conversion efficiency of FA-EM@PDA/DOX\u0026amp;ICG nanoparticles were measured for about 41.6% (Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003ed), which is high than other photothermal materials,such as Cu\u003csub\u003e9\u003c/sub\u003eS\u003csub\u003e5\u003c/sub\u003e nanocrystals, copper selenide nanoparticles, WS\u003csub\u003e2\u003c/sub\u003e nanosheet and so on.\u003csup\u003e39\u0026ndash;41\u003c/sup\u003e Thus, the prepared FA-EM@PDA/DOX\u0026amp;ICG nanoparticles exhibited superior photothermal performance and achieved multiple reversible conversion.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec6\" class=\"Section2\"\u003e \u003ch2\u003eImmune Evasion and Targeted Delivery of FA-EM@PDA/DOX\u0026amp;ICG Nanoparticles\u003c/h2\u003e \u003cp\u003eDue to the size of nanoparticles were very similar to viruses, when their entry into the body can trigger immune responses, causing nanoparticles are easily cleared by mononuclear phagocytes and hindering effective therapeutic delivery.\u003csup\u003e42\u0026ndash;43\u003c/sup\u003e In this context, FA-EM@PDA/DOX\u0026amp;ICG nanoparticles were successfully developed, which were post-encapsulation with FA-EM, and retained relevant proteins from EM. Subsequently, co-incubation with macrophages (RAW264.7 cells) was conducted to evaluate the impact of FA-EM on the engulfment of FA-EM@PDA/DOX\u0026amp;ICG nanoparticles. When PDA/DOX\u0026amp;ICG nanoparticles were co-incubated with RAW264.7 cells, strong green and red fluorescence signals were observed through LSCM. Conversely, co-incubation with EM@PDA/DOX\u0026amp;ICG resulted in weak fluorescence signals. However, when FA-EM@PDA/DOX\u0026amp;ICG nanoparticles were co-incubated with RAW264.7 cells, the fluorescence signals partially recovered compared to EM@PDA/DOX\u0026amp;ICG, although remaining lower than the PDA/DOX\u0026amp;ICG group (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003ea). These findings suggest that EM imparts superior \"stealth\" properties to PDA/DOX\u0026amp;ICG nanoparticles, with folate acid (FA) modification causing a minor reduction in this stealth effect.\u003csup\u003e44\u003c/sup\u003e Nevertheless, FA-EM@PDA/DOX\u0026amp;ICG nanoparticles effectively evaded engulfment by RAW264.7 cells, indicating continued superior stealth performance for circulation and successful evasion of immune system clearance \u003cem\u003ein vivo\u003c/em\u003e.\u003c/p\u003e \u003cp\u003eFor the FA-EM@PDA/DOX\u0026amp;ICG nanoparticles to achieve highly efficient anti-tumor effects, effective uptake by tumor cells is crucial.\u003csup\u003e45\u003c/sup\u003e The FA modification on the EM surface enables specific binding to folate acid receptors (FAR) on tumor cells, thereby enhancing the uptake of FA-EM@PDA/DOX\u0026amp;ICG by tumor cells.\u003csup\u003e46\u003c/sup\u003e Co-incubation of FA-EM@PDA/DOX\u0026amp;ICG nanoparticles with 4T1 and GES-1 cells revealed approximately fourfold higher fluorescence signal intensity in 4T1 cells compared to GES-1 cells (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003eb). This confirms the active targeting delivery capability of FA-EM@PDA/DOX\u0026amp;ICG nanoparticles. These results indicated that FA-EM@PDA/DOX\u0026amp;ICG nanoparticles could specifically bind with 4T1 cells, and then increasing their phagocytosis by 4T1 cells.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003e \u003cb\u003eDrug Release and\u003c/b\u003e \u003cb\u003eIn Vitro\u003c/b\u003e \u003cb\u003eCombined Therapy of FA-EM@PDA/DOX\u0026amp;ICG Nanoparticles\u003c/b\u003e\u003c/p\u003e \u003cp\u003eAlthough FA-EM@PDA/DOX\u0026amp;ICG nanoparticles can be effectively taken up by 4T1 cells, the efficient release of loaded drugs within 4T1 cells is a prerequisite for exerting anti-tumor effects. Therefore, we simulated the tumor microenvironment (TME) \u003cem\u003ein vitro\u003c/em\u003e to analyze the release behavior of DOX and ICG from FA-EM@PDA/DOX\u0026amp;ICG nanoparticles. As shown in Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003ea and \u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003eb, when FA-EM@PDA/DOX\u0026amp;ICG nanoparticles were immersed in mimicking normal physiological environment (pH\u0026thinsp;=\u0026thinsp;7.4), the release of DOX and ICG after 24 h was represented for about 8.31% and 8.56%, respectively. However, when FA-EM@PDA/DOX\u0026amp;ICG nanoparticles immersed in acidic conditions (pH\u0026thinsp;=\u0026thinsp;5.5) condition, approximately 95.1% and 95.9% of DOX and ICG were respectively released from FA-EM@PDA/DOX\u0026amp;ICG nanoparticles after 24 h. This release was attributed to drugs are prone to protonation under acidic conditions, leading to a weakened electrostatic interaction between drugs and FA-EM@PDA/DOX\u0026amp;ICG nanoparticles, and then leading drugs releasing.\u003csup\u003e47\u0026ndash;48\u003c/sup\u003e Additionally, when FA-EM@PDA/DOX\u0026amp;ICG nanoparticles were immersed in GSH and H\u003csub\u003e2\u003c/sub\u003eO\u003csub\u003e2\u003c/sub\u003e solutions, a minor amount of drug release occurred due to the destabilization of PDA surface properties in an oxidative stress microenvironment.\u003csup\u003e49\u003c/sup\u003e When FA-EM@PDA/DOX\u0026amp;ICG nanoparticles were immersed in an \u003cem\u003ein vitro\u003c/em\u003e simulated TME (pH\u0026thinsp;=\u0026thinsp;5.5, 10 mM GSH, 30 \u0026micro;M H\u003csub\u003e2\u003c/sub\u003eO\u003csub\u003e2\u003c/sub\u003e), drug can completely release occurred around 20 h. Therefore, FA-EM@PDA/DOX\u0026amp;ICG nanoparticles have environment-responsive behavior.\u003c/p\u003e \u003cp\u003eDue to FAR on the surface of 4T1 cells can specifically bind with FA molecules on the surface of FA-EM@PDA/DOX\u0026amp;ICG nanoparticles, this effectively promoted the uptake of FA-EM@PDA/DOX\u0026amp;ICG nanoparticles by 4T1 cells. Using CCK-8 experiments to compare the viability of GES-1 cells and 4T1 cells, the combined chemo/photothermal anti-tumor effects of FA-EM@PDA/DOX\u0026amp;ICG nanoparticles were evaluated. The Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003ec and \u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003ed shown that with an increase of FA-EM@PDA/DOX\u0026amp;ICG nanoparticles concentration, the viability of both GES-1 cells and 4T1 cells were gradually decreased. When GES-1 cells and 4T1 cells were exposed under NIR (9 min, 1.5 W/cm\u003csup\u003e2\u003c/sup\u003e) irradiattion at the same concentration and environment, the cells viability of 4T1 cells was significantly lower than that of GES-1 cells. At the high concentration of 200 \u0026micro;g/mL FA-EM@PDA/DOX\u0026amp;ICG nanoparticles, the cells viability of GES-1 cells remained at around 42.6%, while that of 4T1 cells was only about 19.8%. This result indicated that the synergistic chemo/photothermal therapy of FA-EM@PDA/DOX\u0026amp;ICG nanoparticles have a good cytotoxic effect for tumor cells. This effect is mainly attributed to the specific binding of FA molecule on the surface of FA-EM@PDA/DOX\u0026amp;ICG nanoparticles with FAR on tumor cells, increasing the uptake of FA-EM@PDA/DOX\u0026amp;ICG nanoparticles by tumor cells and releasing loaded drugs. Therefore, under NIR irradiation, the constructed FA-EM@PDA/DOX\u0026amp;ICG nanoparticles successfully achieved combined chemo/photothermal therapy, effectively killing tumor cells and reducing damage to normal cells.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003e \u003cb\u003eSynergistic\u003c/b\u003e \u003cb\u003eIn Vivo\u003c/b\u003e \u003cb\u003eAntitumor Effects of FA-EM@PDA/DOX\u0026amp;ICG Nanoparticles with NIR Treatment\u003c/b\u003e\u003c/p\u003e \u003cp\u003eThe synergistic therapy effect of FA-EM@PDA/DOX\u0026amp;ICG nanoparticles with NIR therapy has demonstrated effectively inhibition 4T1 cells growth, prompting us to further investigate their \u003cem\u003ein vivo\u003c/em\u003e antitumor efficacy. Firstly, the \u003cem\u003ein vivo\u003c/em\u003e photothermal conversion performance of FA-EM@PDA/DOX\u0026amp;ICG nanoparticles should be evaluated before animal experiments. Once different photothermal material (PBS, PDA, ICG, PDA/DOX\u0026amp;ICG and FA-EM@PDA/DOX\u0026amp;ICG nanoparticles) were injected into mice via the tail vein and then exposed under NIR irradiation (1.5 W/cm\u003csup\u003e2\u003c/sup\u003e, 9 min) after 24 h later. As shown in Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003ea and \u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003eb, FA-EM@PDA/DOX\u0026amp;ICG nanoparticles have the highest photothermal conversion property among these materials as well as the temperature rising into 59.9 ℃, which was ascribed into the FA-EM@PDA/DOX\u0026amp;ICG nanoparticles have targeting properties and has ability to avoid being cleared by the immune system during the blood circulation. Subsequently, to further verify the anti-tumor performance of FA-EM@PDA/DOX\u0026amp;ICG nanoparticles \u003cem\u003ein vivo\u003c/em\u003e synergistic therapy. 4T1 tumor-bearing mice were randomly divided into six groups and intravenously injected with PBS, DOX, ICG, PDA, PDA/DOX\u0026amp;ICG, and FA-EM@PDA/DOX\u0026amp;ICG nanoparticles. After 24 h, mice in each treatment group were exposed under NIR (1.5 W/cm\u003csup\u003e2\u003c/sup\u003e, 9 min) irradiation every 3 days for a total of 6 treatments. Mouse weight were recorded, and tumor changes were measured throughout the course of the treatment. As shown in Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003ec, \u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003ed and \u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003ee, compared to the PBS treatment group, all treatment groups exhibited varying degrees of tumor size inhibition. Notably, the FA-EM@PDA/DOX\u0026amp;ICG\u0026thinsp;+\u0026thinsp;NIR treatment group displayed a more pronounced inhibitory effect on tumors than the PDA/DOX\u0026amp;ICG\u0026thinsp;+\u0026thinsp;NIR treatment group. This enhanced effect was attributed to the ability of FA-EM@PDA/DOX\u0026amp;ICG nanoparticles to effectively evade clearance by the body's immune system in the bloodstream. Upon reaching the tumor tissue, FA molecules on the surface of FA-EM@PDA/DOX\u0026amp;ICG nanoparticles bind specifically with FAR on 4T1 cells, facilitating the uptake of nanoparticles by 4T1 cells. Under NIR irradiation, the temperature of the tumor tissue rapidly increased, demonstrating superior antitumor efficacy of FA-EM@PDA/DOX\u0026amp;ICG nanoparticles. However, 4T1 tumor-bearing mice treated with DOX alone experienced a significant impact on mental status, including symptoms such as weight loss, disoriented gaze, dehydration, and the mice died completely on day 21 (Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003ef and \u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003ed). In contrast, after FA-EM@PDA/DOX\u0026amp;ICG\u0026thinsp;+\u0026thinsp;NIR synergistic therapy, not only can effectively inhibit tumor growth, but also the mental state of the mice was maintained well and the survival time of the mice was effectively prolonged during the therapy process.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003eBesides, tumor tissues and other major organ (heart, liver, spleen, lung, kidney) were collected and stained with H\u0026amp;E and TUNEL for histopathological analysis. As shown in Fig.\u0026nbsp;6a, in comparison to the PBS\u0026thinsp;+\u0026thinsp;NIR treatment group, where no tumor cells necrosis or apoptosis were observed, but the treatment groups with DOX, ICG\u0026thinsp;+\u0026thinsp;NIR, PDA\u0026thinsp;+\u0026thinsp;NIR, PDA/DOX\u0026amp;ICG\u0026thinsp;+\u0026thinsp;NIR, and FA-EM@PDA/DOX\u0026amp;ICG\u0026thinsp;+\u0026thinsp;NIR exhibited varying degrees of tumor cells death and apoptosis. While ICG\u0026thinsp;+\u0026thinsp;NIR and PDA\u0026thinsp;+\u0026thinsp;NIR treatments could inhibit tumor growth, a significant number of apoptotic tumor cells were only observed on the surface of the tumor tissue. Although DOX effectively killed tumor cells deep within the tumor tissue, it brought about substantial side effects to normal tissues, particularly causing damage to the normal structure of the heart (Fig.\u0026nbsp;6b). However, after the synergistic photothermal therapy with FA-EM@GO-MOF/DOX nanoparticles, abundant apoptotic tumor cells were observed both on the surface and deep within the tumor tissue. This was evidenced by H\u0026amp;E staining results showing extensive nuclear condensation and dissolution, and TUNEL staining results displaying widespread green fluorescence signals across the field of view. Importantly, major mouse organs did not exhibit pathological changes (Fig.\u0026nbsp;6a and 6b). These results underscore that the synergistic photothermal therapy with FA-EM@GO-MOF/DOX nanoparticles demonstrates significant inhibitory and cytotoxic effects on tumor tissues. Furthermore, the nanoparticles exhibit superior biocompatibility to normal tissues, and effectively mitigating the adverse effects brought about by the antitumor drug DOX during the treatment process were ameliorated.\u003c/p\u003e \u003c/div\u003e"},{"header":"Conclusion","content":"\u003cp\u003eThe photo/thermal FA-EM@PDA/DOX\u0026amp;ICG nanoparticles designed in this study demonstrated superior photothermal conversion performance and enable precise and controllable drug release. Surface modification of FA-EM@PDA/DOX\u0026amp;ICG nanoparticles with FA-EM can effectively prevent their clearance by the body's immune system during \u003cem\u003ein vivo\u003c/em\u003e transport, resulting in an extended blood circulation period. Upon reaching the tumor tissue, the nanoparticles exploit their surface FA molecules, forming selective bonds with FAR on tumor cells and consequently augmenting cellular uptake. In response to the specific TME, FA-EM@PDA/DOX\u0026amp;ICG nanoparticles can release loaded DOX and ICG, achieving on-demand and controlled drug release. Under NIR irradiation, FA-EM@PDA/DOX\u0026amp;ICG nanoparticles exhibit superior photothermal conversion efficiency. Moreover, the adsorption of ICG effectively enhances the photothermal conversion performance of FA-EM@PDA/DOX\u0026amp;ICG, and DOX's penetration into tumor tissues compensates for NIR penetration deficiencies, promoting the destruction of deep-seated tumor cells and effectively inhibiting tumor growth. Importantly, FA-EM@PDA/DOX\u0026amp;ICG nanoparticles, constructed based on the neurotransmitter dopamine, exhibit excellent biocompatibility and mitigate the toxic side effects of DOX. In conclusion, the environmentally responsive treatment platform developed in this study achieves combined photothermal therapy and chemotherapy through a one-step approach. This platform effectively addresses the shortcomings of individual treatments, thereby enhancing therapeutic efficacy. It also provides valuable insights for the design of more intelligent and efficient nanoplatforms for tumor treatment in the future.\u003c/p\u003e"},{"header":"Declarations","content":"\u003cp\u003e\u003cstrong\u003eEthics approval and consent to participate\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eAll experimental procedures were approved by the Institutional Animal Care and Use Committee of the Medical College of Nanjing University.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eConsent for publication\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eEach coauthor has read the manuscript and approves its submission. This work is being submitted exclusively to your journal.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eData Availability Statement\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe datasets supporting the results of this article are included within the article.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eCompeting interests\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe authors declared that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eFunding\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThis research was supported by the \u0026ldquo;Jiangsu Provincial Postdoctoral Excellence Program\u0026rdquo; (2023ZB570).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAuthors\u0026apos; contributions\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eChunmin Deng responsible for the experimental operations, data collection and analysis.\u0026nbsp;\u0026nbsp;Hao Zhang\u0026nbsp;data processing and image generation. Li Song is responsible for the experimental design and writing of the manuscript. All authors read and approved the final manuscript.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAcknowledgements\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe acknowledged are included within the article.\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\u003cli\u003e\u003cspan\u003eHyuna Sung, Jacques Ferlay, Rebecca L Siegel, et al. Global cancer statistics 2020: GLOBOCAN estimates of incidence and mortality worldwide for 36 cancers in 185 countries. CA. Cancer J. Clin., 2021.71: 209\u0026ndash;249.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eZhengzou Fang, Ziyu Zhu, Zijian Zhuang, et al. Cascade biomimetic intelligent nanotheranostic agents for imaging-guided tumor synergistic therapy. Nanomedicine (Lond), 2023. 18: 35\u0026ndash;52.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eYanping Ding, Shishuai Su, Ruirui Zhang, et al. 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Colloid Interface Sci., 2018. 252: 1\u0026ndash;20.\u003c/span\u003e\u003c/li\u003e\u003c/ol\u003e"},{"header":"Scheme ","content":"\u003cp\u003eScheme 1 is available in the Supplementary Files section\u003c/p\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":true,"highlight":"","institution":"","isAcceptedByJournal":false,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true},"keywords":"Photothermal therapy, combined therapy, targeted delivery, environment-responsive","lastPublishedDoi":"10.21203/rs.3.rs-3844904/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-3844904/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003eNanoparticle-based photothermal therapy (PTT) has emerged as a promising approach in tumor treatment due to its high selectivity and low invasiveness. However, the penetration of near-infrared light (NIR) is limited, leading it fails to induce damage to the deep-seated tumor cells within the tumor tissue. Additionally, inefficient uptake of photothermal nanoparticles by tumor cells results in suboptimal outcomes for PTT. Based on the above-mentioned issues, this study utilized the adhesive properties of photothermal material, polydopamine (PDA), which can successfully load the photosensitizer indocyanine green (ICG) and chemotherapeutic drug doxorubicin (DOX) to achieve combined photothermal and chemotherapy treatment (PDA/DOX\u0026amp;ICG), aiming to compensate for the poor penetration of NIR in tumor tissues and the photothermal conversion performance of PDA. For the purpose of extending the blood circulation time of PDA/DOX\u0026amp;ICG nanoparticles, evading clearance by the body immune system and achieving targeted delivery to tumor tissues, a protective envelopment was created using erythrocyte membranes modified with folate acid (FA-EM). After reaching the tumor tissue, the obtained FA-EM@PDA/DOX\u0026amp;ICG nanoparticles can specific bind with folate acid receptors on the surface of tumor cells. This interaction facilitates improved uptake by tumor cells leading to the subsequent release of loaded DOX and ICG in response to the unique tumor microenvironment. DOX penetration ability can effectively compensate the limitation of NIR penetration at the tumor tissue. While ICG, as a typical photosensitizer, significantly enhances the photothermal conversion performance of FA-EM@PDA/DOX\u0026amp;ICG nanoparticles, thereby inducing tumor cells damage. \u003cem\u003eIn vitro\u003c/em\u003e and \u003cem\u003ein vivo\u003c/em\u003e experimental results demonstrated that the coordinated NIR treatment with FA-EM@PDA/DOX\u0026amp;ICG not only effectively inhibits tumor growth but also exhibits superior biocompatibility, effectively mitigating DOX-induced tissue damage.\u003c/p\u003e","manuscriptTitle":"Environment-Responsive Dopamine Nanoplatform for Tumor Synergistic Therapy","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2024-01-12 20:34:23","doi":"10.21203/rs.3.rs-3844904/v1","editorialEvents":[{"type":"communityComments","content":0}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"d6997818-a2d1-4291-9f73-ebd7c76bb9f8","owner":[],"postedDate":"January 12th, 2024","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"posted","subjectAreas":[],"tags":[],"updatedAt":"2024-04-24T04:06:38+00:00","versionOfRecord":[],"versionCreatedAt":"2024-01-12 20:34:23","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-3844904","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-3844904","identity":"rs-3844904","version":["v1"]},"buildId":"qtupq5eGEP_6zYnWcrvyt","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}