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Biomedical applications of functionalized hollow mesoporous silica nanoparticles: focusing on molecular imaging Hollow mesoporous silica nanoparticles (HMSNs), with a large cavity inside each original mesoporous silica nanoparticle, have recently gained increasing interest owing to their tremendous potential for cancer imaging and therapy. The last several years have witnessed a rapid development in the engineering of functionalized HMSNs (i.e., f-HMSNs), with various types of inorganic functional nanocrystals integrated into the system for imaging and therapeutic applications. In this article, we summarize the recent progress in the design and biological applications of f-HMSNs, with a special emphasis on molecular imaging. Commonly used synthetic strategies for the generation of high quality HMSNs will be discussed in detail, followed by a systematic review of engineered f-HMSNs for optical, PET, MRI and ultrasound imaging in preclinical studies. Finally, we discuss the challenges and future research directions regarding the use of f-HMSNs for cancer imaging and therapy. KEYWORDS: cancer n hollow mesoporous silica nanoparticle n molecular imaging n nanomedicine n optical imaging n PET n theranostics
The last decade has witnessed rapid development in the design and synthesis of various types of multifunctional nanosystems that can potentially be used for cancer-targeted imaging and therapy [1–3]. Since silica is “generally recognized as safe” by the US FDA [101], silica-based nanomaterials have been extensively investigated because of the nontoxic nature and facile chemistry for surface modification [4,5]. Recently, dye-doped ultrasmall silica nanoparticles called Cornell dots [6] have entered clinical investigation in melanoma patients, which is an important milestone for the use of inorganic nanomaterials in the same fashion as a drug in humans. In comparison with pure silica nanoparticles, mesoporous silica nanoparticles (MSNs) possess many attractive properties, such as a large surface area, high pore volume, uniform and tunable pore size, and low mass density [7]. Although MSNs have been intensively investigated for drug delivery applications since 2000 [8], how to improve the drug loading capacity and in vivo targeting efficiency while minimizing the undesired side effects to healthy organs remains a major challenge. Hollow MSNs (HMSNs), with a large cavity inside each original MSN, have recently been developed to greatly enhance the drug loading capacity [4]. With the availability of well-established techniques for integrating various types of inorganic functional nanocrystals (e.g., iron oxide nanoparticles and gold [Au] nanoparticles, among others) inside or at the surface of HMSNs, in addition to surface
functionalization of HMSNs to confer biocompatibility, imaging capability and specific targeting, among others, such functionalized HMSNs (f‑HMSNs) (Figure 1) with a rattle-type (or yolk–shell) structure are highly attractive multifunctional nanoplatforms for future cancer imaging and therapeutic applications. This review summarizes the recent progress in the design and biomedical applications of f‑HMSNs, with a primary focus on molecular imaging since, to date, limited progress has been made in the use of f‑HMSNs for cancer therapy in vivo. First, commonly used synthetic strategies for the generation of HMSNs and f‑HMSNs will be discussed in detail. Second, the progress to date in the engineering of f‑HMSNs for optical imaging, PET, MRI and ultrasound (US) imaging in small animals will be reviewed. Finally, we discuss the challenges and future research directions in the use of f‑HMSNs for cancer imaging and therapeutic applications.
10.2217/NNM.13.177 © 2013 Future Medicine Ltd
Nanomedicine (2013) 8(12), 2027–2039
Sixiang Shi‡1, Feng Chen‡2 & Weibo Cai*1,2,3,4 Materials Science Program, University of Wisconsin – Madison, WI, USA 2 Department of Radiology, University of Wisconsin – Madison, WI, USA 3 Department of Medical Physics, University of Wisconsin – Madison, Room 7137, 1111 Highland Avenue, Madison, WI 53705-2275, USA 4 University of Wisconsin Carbone Cancer Center, Madison, WI, USA *Author for correspondence: Tel.: +1 608 262 1749 Fax: +1 608 265 0614 [email protected] ‡ Authors contributed equally 1
Templating methods for the synthesis of HMSNs & f-HMSNs Generally speaking, soft and hard templating are two of the most popular methods used for the synthesis of HMSNs. The soft-templating method uses certain surfactants (e.g., tetrapropylammonium hydroxide) and costructuredirecting agents (e.g., tetraethyl orthosilicate or 3-aminopropyltriethoxysilane) to form interior hollow structures and mesoporous silica shell simultaneously [9,10]. For example, a
part of
ISSN 1743-5889
2027
Review
Shi, Chen & Cai
Drug Functional nanoparticle Polymer Antibody Radioisotope
HMSN
Figure 1. Functionalized hollow mesoporous silica nanoparticle. HMSN: Hollow mesoporous silica nanoparticle.
hollow-structured aluminosilicate with a highly ordered 3D mesoporous shell and significantly improved hydrothermal stability was generated using this technique (Figure 2A) [9]. In another report, HMSNs with improved controllability in both morphology and size were synthesized by employing micelle and emulsion as dual soft templates [10]. However, reducing the aggregation of nanoparticles was a major challenge. In addition to HMSNs, a modified soft-templating method could also be used to prepare yolk/SiO2 shell structures with different cores inside, such as Au and Fe2O3 (Figure 2B) [11–14]. One major limitation of soft-templating methods is that owing to the difficulty in controlling the droplet size, it is quite challenging to obtain monodispersed HMSNs with well-controlled size, morphology and reproducibility. The hard-templating method has recently been demonstrated to be a better strategy for the synthesis of high quality HMSNs (or rattle-type f‑HMSNs) with good monodispersity and reproducibility, owing to the improved control over the synthesis of templates and well-established techniques for selective etching [4,15]. The basis of such an improved method relies on the compositional or structural difference between the core and shell of the nanoparticles, where typically the inner core could be selectively etched away to leave a large cavity inside the thin porous shell to form a HMSN. The selection of suitable hard templates and etchants is critical and highly dependent on the different etching mechanisms [9,16–19]. To date, various materials, including polystyrene spheres, dense silica 2028
Nanomedicine (2013) 8(12)
and iron oxide nanoparticles, have been used as hard templates, whereas agents such as sodium carbonate, sodium hydroxide, hydrofluoric acid, hydrochloric acid and even hot water have been successfully employed as the etchant [16,19–22]. For example, a flexible, scalable and robust method to synthesize tailored silica nanorattle structures was developed using an organic–inorganic hybrid solid silica sphere as the template and hydrofluoric acid as the etchant, which has a small silica core inside each volume-tunable hollow cavity with a mesoporous shell (Figure 2C) [16]. Similar etching mechanisms, such as structural difference-based selective etching (Figure 2D) and cationic surfactant-assisted self-templating method, have also been utilized to synthesize high-quality HMSNs (or rattle-type HMSNs) with Na 2CO3 as the etchant at an elevated temperature [19,23]. Besides the abovementioned selective etching of core/shell structured nanoparticles to form HMSNs, surface-protected etching using NaOH as the etchant is another means to prepare HMSNs [22,24,25], for which poly(vinylpyrrolidone) and poly(dimethyldiallylammonium chloride) have been demonstrated to be good surface protectors. In order to apply this strategy to the synthesis of silica-based HMSNs with particle sizes less than 100 nm, a milder etching strategy using hot water has also been developed [17,20,26], which has provided researchers with alternative techniques for the design and generation of HMSNs with suitable sizes. Overall, using the hard-templating method, the cavity volume, size and shell thickness of HMSNs could be readily controlled by selecting suitable templates and etching strategies [4]. More importantly, inorganic functional nanocrystals such as Au nanoparticles, Fe3O4, MnOx and upconversion nanoparticles (UCNPs) could also be encapsulated inside the HMSNs to fabricate f‑HMSNs with various molecular imaging capabilities [17,27–32]. The following section will be focused on the engineering of f‑HMSNs for in vivo imaging applications using different modalities.
Engineering of f-HMSNs for molecular imaging Molecular imaging, defined as the “visualization, characterization and measurement of biological processes at the molecular and cellular levels in humans and other living systems” [33], has greatly facilitated the investigation of complex biological events in both the preclinical and clinical setting [34]. Although clinical translation of novel imaging agents has so far been rather slow, molecular imaging does hold great potential in multiple future science group
Biomedical applications of functionalized hollow mesoporous silica nanoparticles
aspects such as drug development, disease diagnosis, monitoring therapeutic responses and understanding complex interactions between nanomedicine and living biological systems [35]. Over the last decade, engineered f‑HMSNs have been investigated with various imaging techniques such as optical, PET, MRI and US. Optical imaging Generally speaking, optical imaging is inexpensive, widely available, easy-to-handle and highly sensitive, and has been extensively used for monitoring various molecular/biological events in cells and small animal models [35–37]. Although various strategies (e.g., organic dye doping and fluorescent nanoparticle encapsulation) have
Review
been developed to engineer MSNs for optical imaging applications in vitro and in vivo [38,39], the design and synthesis of f‑HMSNs for in vivo optical imaging is still in its infancy and needs much more investigation in the future [40–42]. In one study, f luorescein isothiocyanate (FITC) and doxorubicin were coloaded into multishelled HMSNs by sequentially mixing HMSNs with aqueous solutions of FITC and doxorubicin [40]. Although a green fluorescence signal from FITC could be detected in mice after intraperitoneal injection of FITC-loaded HMSNs at various dosages (Figure 3A & B), other dyes with near-infrared (NIR; 700–900 nm) excitation and emission will be more desirable for future studies because of the much
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Figure 2. Soft- and hard-templating synthesis of hollow mesoporous silica nanoparticles. Representative transmission electron microscopy images of hollow mesoporous silica nanoparticles synthesized via (A & B) soft- or (C & D) hard-templating methods. (A) A high‑resolution transmission electron microscopy image of hollow spherical cubic mesoporous aluminosilicate. (B) Yolk/shell structure with spindle-like Fe2O3 particles inside the cavity. (C) Silica-based nanorattles and (D) hollow mesoporous silica nanoparticles synthesized by selective etching using hydrofluoric acid and Na2CO3 as the etchant, respectively. Reproduced with permission from [9,11,16,19] .
future science group
www.futuremedicine.com
2029
Review
Shi, Chen & Cai
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Figure 3. In vivo optical imaging of functionalized hollow mesoporous silica nanoparticles. (A) A transmission electron microscopy image of hollow double-shelled silica nanoparticles. (B) Optical imaging of fluorescein isothiocyanate-loaded hollow double-shelled silica nanoparticles at 55 min postinjection into mice. (C) A transmission electron microscopy image of Fe3O4 @ hollow@a‑NaYF4:Er/Yb nanoparticles. (Ci) A higher-magnification transmission electron microscopy image and (Cii) a selected area electron diffraction of a-NaYF4:Er/Yb shell. (D) In vivo UCL imaging of H22 xenograft tumor-bearing mice after intravenous injection of doxorubicin-loaded Fe3O4 @ hollow@a‑NaYF4:Er/Yb nanoparticles (Di) without or (Dii) with application of magnetic field for 1 h. (E) A transmission electron microscopy image of rattle-structured upconversion nanoparticle@hollow mesoporous silica nanoparticles. (F) UCL imaging and (G) in vivo T1-weighted MRI of upconversion nanoparticle@hollow mesoporous silica nanoparticles after intratumoral injection into mice. For color images see online at www.futuremedicine.com/doi/full/10.2217/NNM.13.177. UCL: Upconversion luminescence. Reproduced with permission from [40–42] .
better tissue penetration of light and significantly lower tissue autofluorescence in the NIR window [43,44]. 2030
Nanomedicine (2013) 8(12)
In addition to dye-doped HMSNs, inte gration of fluorescent nanoparticles, such as quantum dots (QDs) or UCNPs, inside or future science group
Biomedical applications of functionalized hollow mesoporous silica nanoparticles
at the surface of HMSNs, can also be highly desirable for in vivo optical imaging applications. Although PEG-modified and liposomecoated QD@MSN core/shell nanoparticles have been investigated for imaging studies in cell culture [45], the combination of QDs with HMSNs has not been reported to date, possibly owing to the difficulty of trapping a single QD inside HMSNs and toxicity-related concerns for cadmium-based QDs [46,47]. When compared with conventional organic dyes and QDs, UCNPs can avoid the potential ultraviolet photodamage of tissue (since the excitation wavelength of UCNPs is in the NIR window, typically 980 nm) and exhibit good biocompatibility in living systems [48]. Such upconversion luminescence (UCL) typically occurs when lowenergy light (e.g., NIR) is converted to higher energy light through the sequential absorption of multiple photons or energy transfer. In addition, UCNPs have many other attractive UCL features, such as sharp emission lines [49], superb photostability [50], high detection sensitivity [51], deep tissue penetration depth [52] and extremely low autofluorescence [53], which make them one of the best classes of nanoparticles for optical imaging applications. In a recent study, an ion-exchange process was utilized to fabricate nanorattles, each of which consists of a UCNP (i.e., a-NaYF4:Er/Yb)-containing shell and a loose magnetic nanoparticle core (Figure 3C) [42]. It was demonstrated that the multifunctional nanorattles could be directed by an external magnetic field for in vivo tumor targeting, which can also be noninvasively imaged using the visible luminescence (i.e., green and red emissions) under 980 nm excitation (Figure 3D). Instead of coating UCNPs at the surface of HMSNs, a different strategy was used in another report [41]. Core/shell structured NaYF4:Er/Yb@NaGdF4 UCNP was first encapsulated inside a dense silica shell, part of which was then selectively etched away via a protective etching strategy to form a multifunctional rattle-structured UCNP@ HMSN nanosystem (Figure 3E) [41]. With the presence of NaYF4:Er/Yb@NaGdF4 UCNP inside the nanosystem, in vivo UCL imaging after intratumoral injection into HeLa tumor-bearing mice was achieved (Figure 3F). In addition, with the doping of Gd3+ ions into the UCNP surface/matrix, this multifunctional nanosystem could also be detected by T1-weighted MRI (Figure 3G). However, neither in vivo optical nor MRI imaging after intravenous injection of the UCNP@ HMSN nanosystem was reported, possibly due to future science group
Review
very low tumor accumulation of the nanoparticle based on passive targeting alone. Although UCNP-f‑HMSNs possess attractive UCL features for in vivo optical imaging, to date only UCNP-f‑HMSNs emitting in the visible range have been reported [41,42]. With the rapid development in optimizing Tm3+/Yb3+ co-doped b-NaYF4 UCNP with strong NIR-to-NIR emission intensity [54,55], b-NaYF4:Tm/Yb UCNPf‑HMSNs will probably become a vibrant research area in the near future. One major limitation of optical imaging, even in the NIR range, is limited tissue penetration depth, which typically cannot reach beyond a few centimeters. In addition, optical imaging is qualitative in nature and semiquantitative at best. Therefore, the use of quantitative imaging techniques capable of deep tissue penetration is highly desirable for determining the fate of f‑HMSNs in vivo. PET is sensitive, quantitative, clinically relevant and has excellent tissue penetration [56–60], making it a highly desirable imaging technique for in vivo investigation of f‑HMSNs. PET Owing to the abovementioned advantages of PET over other imaging modalities (e.g., optical and MRI), labeling nanoparticles with positronemitting radionuclides has been generally recognized as the most accurate means for noninvasive evaluation of their biodistribution and pharmacokinetics [61,62]. Since radiolabeled nanoparticles (especially with PET isotopes) represent a newly emerged research area over the last decade, to the best of our knowledge, no reports on radiolabeled HMSNs exist in the literature to date. We have successfully developed novel antibodyconjugated and 64Cu (half-life = 12.7 h) labeled HMSNs, denoted as 64Cu-NOTA-HMSNTRC105 (Figure 4A & B), and demonstrated the proof-of-principle for in vivo active tumor vasculature targeting, which can be monitored with noninvasive serial PET imaging (Figure 4C). Since many nanoparticles suffer from poor extra vasation in the tumor tissue [61,62], tumor vasculature targeting was adopted where extravasation is not required to observe the tumor signal [63–65]. CD105 (also called endoglin) was chosen as the vascular target, which is almost exclusively expressed on proliferating tumor endothelial cells [66,67], and TRC105 (a human/murine chimeric IgG1 monoclonal antibody that binds to both human and murine CD105 [68]) was used as the targeting ligand. Significantly higher uptake in 4T1 murine breast tumors (which express high level of CD105 on the tumor vasculature [69,70]) www.futuremedicine.com
2031
Shi, Chen & Cai
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Figure 4. In vivo PET imaging of radiolabeled hollow mesoporous silica nanoparticles. (A) A transmission electron microscopy image of a HMSN. (B) 64Cu-NOTA-HMSN-TRC105. (C) Serial coronal PET images of 4T1 breast tumor‑bearing mice at different time points postinjection of 64 Cu-NOTA-HMSN-TRC105 or 64Cu-NOTA-HMSN. Tumors are indicated by arrows. HMSN: Hollow mesoporous silica nanoparticle. For color images see online at www.futuremedicine.com/doi/full/10.2217/NNM.13.177.
was observed for 64 Cu-labeled, TRC105conjugated HMSNs, which is approximately twofold that for the nontargeted HMSNs (Figure 4C). With demonstrated active tumor targeting efficiency in vivo and high drug loading capacity, 64 Cu-NOTA-HMSN-TRC105 is a promising platform for future image-guided, tumor-targeted drug delivery and more efficacious cancer therapy. Although PET imaging is highly sensitive and quantitative for in vivo applications, its spatial resolution (milimeter level) is significantly lower than that of MRI (typically
Biomedical applications of functionalized hollow mesoporous silica nanoparticles: focusing on molecular imaging Hollow mesoporous silica nanoparticles (HMSNs), with a large cavity inside each original mesoporous silica nanoparticle, have recently gained increasing interest owing to their tremendous potential for cancer imaging and therapy. The last several years have witnessed a rapid development in the engineering of functionalized HMSNs (i.e., f-HMSNs), with various types of inorganic functional nanocrystals integrated into the system for imaging and therapeutic applications. In this article, we summarize the recent progress in the design and biological applications of f-HMSNs, with a special emphasis on molecular imaging. Commonly used synthetic strategies for the generation of high quality HMSNs will be discussed in detail, followed by a systematic review of engineered f-HMSNs for optical, PET, MRI and ultrasound imaging in preclinical studies. Finally, we discuss the challenges and future research directions regarding the use of f-HMSNs for cancer imaging and therapy. KEYWORDS: cancer n hollow mesoporous silica nanoparticle n molecular imaging n nanomedicine n optical imaging n PET n theranostics
The last decade has witnessed rapid development in the design and synthesis of various types of multifunctional nanosystems that can potentially be used for cancer-targeted imaging and therapy [1–3]. Since silica is “generally recognized as safe” by the US FDA [101], silica-based nanomaterials have been extensively investigated because of the nontoxic nature and facile chemistry for surface modification [4,5]. Recently, dye-doped ultrasmall silica nanoparticles called Cornell dots [6] have entered clinical investigation in melanoma patients, which is an important milestone for the use of inorganic nanomaterials in the same fashion as a drug in humans. In comparison with pure silica nanoparticles, mesoporous silica nanoparticles (MSNs) possess many attractive properties, such as a large surface area, high pore volume, uniform and tunable pore size, and low mass density [7]. Although MSNs have been intensively investigated for drug delivery applications since 2000 [8], how to improve the drug loading capacity and in vivo targeting efficiency while minimizing the undesired side effects to healthy organs remains a major challenge. Hollow MSNs (HMSNs), with a large cavity inside each original MSN, have recently been developed to greatly enhance the drug loading capacity [4]. With the availability of well-established techniques for integrating various types of inorganic functional nanocrystals (e.g., iron oxide nanoparticles and gold [Au] nanoparticles, among others) inside or at the surface of HMSNs, in addition to surface
functionalization of HMSNs to confer biocompatibility, imaging capability and specific targeting, among others, such functionalized HMSNs (f‑HMSNs) (Figure 1) with a rattle-type (or yolk–shell) structure are highly attractive multifunctional nanoplatforms for future cancer imaging and therapeutic applications. This review summarizes the recent progress in the design and biomedical applications of f‑HMSNs, with a primary focus on molecular imaging since, to date, limited progress has been made in the use of f‑HMSNs for cancer therapy in vivo. First, commonly used synthetic strategies for the generation of HMSNs and f‑HMSNs will be discussed in detail. Second, the progress to date in the engineering of f‑HMSNs for optical imaging, PET, MRI and ultrasound (US) imaging in small animals will be reviewed. Finally, we discuss the challenges and future research directions in the use of f‑HMSNs for cancer imaging and therapeutic applications.
10.2217/NNM.13.177 © 2013 Future Medicine Ltd
Nanomedicine (2013) 8(12), 2027–2039
Sixiang Shi‡1, Feng Chen‡2 & Weibo Cai*1,2,3,4 Materials Science Program, University of Wisconsin – Madison, WI, USA 2 Department of Radiology, University of Wisconsin – Madison, WI, USA 3 Department of Medical Physics, University of Wisconsin – Madison, Room 7137, 1111 Highland Avenue, Madison, WI 53705-2275, USA 4 University of Wisconsin Carbone Cancer Center, Madison, WI, USA *Author for correspondence: Tel.: +1 608 262 1749 Fax: +1 608 265 0614 [email protected] ‡ Authors contributed equally 1
Templating methods for the synthesis of HMSNs & f-HMSNs Generally speaking, soft and hard templating are two of the most popular methods used for the synthesis of HMSNs. The soft-templating method uses certain surfactants (e.g., tetrapropylammonium hydroxide) and costructuredirecting agents (e.g., tetraethyl orthosilicate or 3-aminopropyltriethoxysilane) to form interior hollow structures and mesoporous silica shell simultaneously [9,10]. For example, a
part of
ISSN 1743-5889
2027
Review
Shi, Chen & Cai
Drug Functional nanoparticle Polymer Antibody Radioisotope
HMSN
Figure 1. Functionalized hollow mesoporous silica nanoparticle. HMSN: Hollow mesoporous silica nanoparticle.
hollow-structured aluminosilicate with a highly ordered 3D mesoporous shell and significantly improved hydrothermal stability was generated using this technique (Figure 2A) [9]. In another report, HMSNs with improved controllability in both morphology and size were synthesized by employing micelle and emulsion as dual soft templates [10]. However, reducing the aggregation of nanoparticles was a major challenge. In addition to HMSNs, a modified soft-templating method could also be used to prepare yolk/SiO2 shell structures with different cores inside, such as Au and Fe2O3 (Figure 2B) [11–14]. One major limitation of soft-templating methods is that owing to the difficulty in controlling the droplet size, it is quite challenging to obtain monodispersed HMSNs with well-controlled size, morphology and reproducibility. The hard-templating method has recently been demonstrated to be a better strategy for the synthesis of high quality HMSNs (or rattle-type f‑HMSNs) with good monodispersity and reproducibility, owing to the improved control over the synthesis of templates and well-established techniques for selective etching [4,15]. The basis of such an improved method relies on the compositional or structural difference between the core and shell of the nanoparticles, where typically the inner core could be selectively etched away to leave a large cavity inside the thin porous shell to form a HMSN. The selection of suitable hard templates and etchants is critical and highly dependent on the different etching mechanisms [9,16–19]. To date, various materials, including polystyrene spheres, dense silica 2028
Nanomedicine (2013) 8(12)
and iron oxide nanoparticles, have been used as hard templates, whereas agents such as sodium carbonate, sodium hydroxide, hydrofluoric acid, hydrochloric acid and even hot water have been successfully employed as the etchant [16,19–22]. For example, a flexible, scalable and robust method to synthesize tailored silica nanorattle structures was developed using an organic–inorganic hybrid solid silica sphere as the template and hydrofluoric acid as the etchant, which has a small silica core inside each volume-tunable hollow cavity with a mesoporous shell (Figure 2C) [16]. Similar etching mechanisms, such as structural difference-based selective etching (Figure 2D) and cationic surfactant-assisted self-templating method, have also been utilized to synthesize high-quality HMSNs (or rattle-type HMSNs) with Na 2CO3 as the etchant at an elevated temperature [19,23]. Besides the abovementioned selective etching of core/shell structured nanoparticles to form HMSNs, surface-protected etching using NaOH as the etchant is another means to prepare HMSNs [22,24,25], for which poly(vinylpyrrolidone) and poly(dimethyldiallylammonium chloride) have been demonstrated to be good surface protectors. In order to apply this strategy to the synthesis of silica-based HMSNs with particle sizes less than 100 nm, a milder etching strategy using hot water has also been developed [17,20,26], which has provided researchers with alternative techniques for the design and generation of HMSNs with suitable sizes. Overall, using the hard-templating method, the cavity volume, size and shell thickness of HMSNs could be readily controlled by selecting suitable templates and etching strategies [4]. More importantly, inorganic functional nanocrystals such as Au nanoparticles, Fe3O4, MnOx and upconversion nanoparticles (UCNPs) could also be encapsulated inside the HMSNs to fabricate f‑HMSNs with various molecular imaging capabilities [17,27–32]. The following section will be focused on the engineering of f‑HMSNs for in vivo imaging applications using different modalities.
Engineering of f-HMSNs for molecular imaging Molecular imaging, defined as the “visualization, characterization and measurement of biological processes at the molecular and cellular levels in humans and other living systems” [33], has greatly facilitated the investigation of complex biological events in both the preclinical and clinical setting [34]. Although clinical translation of novel imaging agents has so far been rather slow, molecular imaging does hold great potential in multiple future science group
Biomedical applications of functionalized hollow mesoporous silica nanoparticles
aspects such as drug development, disease diagnosis, monitoring therapeutic responses and understanding complex interactions between nanomedicine and living biological systems [35]. Over the last decade, engineered f‑HMSNs have been investigated with various imaging techniques such as optical, PET, MRI and US. Optical imaging Generally speaking, optical imaging is inexpensive, widely available, easy-to-handle and highly sensitive, and has been extensively used for monitoring various molecular/biological events in cells and small animal models [35–37]. Although various strategies (e.g., organic dye doping and fluorescent nanoparticle encapsulation) have
Review
been developed to engineer MSNs for optical imaging applications in vitro and in vivo [38,39], the design and synthesis of f‑HMSNs for in vivo optical imaging is still in its infancy and needs much more investigation in the future [40–42]. In one study, f luorescein isothiocyanate (FITC) and doxorubicin were coloaded into multishelled HMSNs by sequentially mixing HMSNs with aqueous solutions of FITC and doxorubicin [40]. Although a green fluorescence signal from FITC could be detected in mice after intraperitoneal injection of FITC-loaded HMSNs at various dosages (Figure 3A & B), other dyes with near-infrared (NIR; 700–900 nm) excitation and emission will be more desirable for future studies because of the much
200 nm
50 nm
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0.5 µm
Figure 2. Soft- and hard-templating synthesis of hollow mesoporous silica nanoparticles. Representative transmission electron microscopy images of hollow mesoporous silica nanoparticles synthesized via (A & B) soft- or (C & D) hard-templating methods. (A) A high‑resolution transmission electron microscopy image of hollow spherical cubic mesoporous aluminosilicate. (B) Yolk/shell structure with spindle-like Fe2O3 particles inside the cavity. (C) Silica-based nanorattles and (D) hollow mesoporous silica nanoparticles synthesized by selective etching using hydrofluoric acid and Na2CO3 as the etchant, respectively. Reproduced with permission from [9,11,16,19] .
future science group
www.futuremedicine.com
2029
Review
Shi, Chen & Cai
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Figure 3. In vivo optical imaging of functionalized hollow mesoporous silica nanoparticles. (A) A transmission electron microscopy image of hollow double-shelled silica nanoparticles. (B) Optical imaging of fluorescein isothiocyanate-loaded hollow double-shelled silica nanoparticles at 55 min postinjection into mice. (C) A transmission electron microscopy image of Fe3O4 @ hollow@a‑NaYF4:Er/Yb nanoparticles. (Ci) A higher-magnification transmission electron microscopy image and (Cii) a selected area electron diffraction of a-NaYF4:Er/Yb shell. (D) In vivo UCL imaging of H22 xenograft tumor-bearing mice after intravenous injection of doxorubicin-loaded Fe3O4 @ hollow@a‑NaYF4:Er/Yb nanoparticles (Di) without or (Dii) with application of magnetic field for 1 h. (E) A transmission electron microscopy image of rattle-structured upconversion nanoparticle@hollow mesoporous silica nanoparticles. (F) UCL imaging and (G) in vivo T1-weighted MRI of upconversion nanoparticle@hollow mesoporous silica nanoparticles after intratumoral injection into mice. For color images see online at www.futuremedicine.com/doi/full/10.2217/NNM.13.177. UCL: Upconversion luminescence. Reproduced with permission from [40–42] .
better tissue penetration of light and significantly lower tissue autofluorescence in the NIR window [43,44]. 2030
Nanomedicine (2013) 8(12)
In addition to dye-doped HMSNs, inte gration of fluorescent nanoparticles, such as quantum dots (QDs) or UCNPs, inside or future science group
Biomedical applications of functionalized hollow mesoporous silica nanoparticles
at the surface of HMSNs, can also be highly desirable for in vivo optical imaging applications. Although PEG-modified and liposomecoated QD@MSN core/shell nanoparticles have been investigated for imaging studies in cell culture [45], the combination of QDs with HMSNs has not been reported to date, possibly owing to the difficulty of trapping a single QD inside HMSNs and toxicity-related concerns for cadmium-based QDs [46,47]. When compared with conventional organic dyes and QDs, UCNPs can avoid the potential ultraviolet photodamage of tissue (since the excitation wavelength of UCNPs is in the NIR window, typically 980 nm) and exhibit good biocompatibility in living systems [48]. Such upconversion luminescence (UCL) typically occurs when lowenergy light (e.g., NIR) is converted to higher energy light through the sequential absorption of multiple photons or energy transfer. In addition, UCNPs have many other attractive UCL features, such as sharp emission lines [49], superb photostability [50], high detection sensitivity [51], deep tissue penetration depth [52] and extremely low autofluorescence [53], which make them one of the best classes of nanoparticles for optical imaging applications. In a recent study, an ion-exchange process was utilized to fabricate nanorattles, each of which consists of a UCNP (i.e., a-NaYF4:Er/Yb)-containing shell and a loose magnetic nanoparticle core (Figure 3C) [42]. It was demonstrated that the multifunctional nanorattles could be directed by an external magnetic field for in vivo tumor targeting, which can also be noninvasively imaged using the visible luminescence (i.e., green and red emissions) under 980 nm excitation (Figure 3D). Instead of coating UCNPs at the surface of HMSNs, a different strategy was used in another report [41]. Core/shell structured NaYF4:Er/Yb@NaGdF4 UCNP was first encapsulated inside a dense silica shell, part of which was then selectively etched away via a protective etching strategy to form a multifunctional rattle-structured UCNP@ HMSN nanosystem (Figure 3E) [41]. With the presence of NaYF4:Er/Yb@NaGdF4 UCNP inside the nanosystem, in vivo UCL imaging after intratumoral injection into HeLa tumor-bearing mice was achieved (Figure 3F). In addition, with the doping of Gd3+ ions into the UCNP surface/matrix, this multifunctional nanosystem could also be detected by T1-weighted MRI (Figure 3G). However, neither in vivo optical nor MRI imaging after intravenous injection of the UCNP@ HMSN nanosystem was reported, possibly due to future science group
Review
very low tumor accumulation of the nanoparticle based on passive targeting alone. Although UCNP-f‑HMSNs possess attractive UCL features for in vivo optical imaging, to date only UCNP-f‑HMSNs emitting in the visible range have been reported [41,42]. With the rapid development in optimizing Tm3+/Yb3+ co-doped b-NaYF4 UCNP with strong NIR-to-NIR emission intensity [54,55], b-NaYF4:Tm/Yb UCNPf‑HMSNs will probably become a vibrant research area in the near future. One major limitation of optical imaging, even in the NIR range, is limited tissue penetration depth, which typically cannot reach beyond a few centimeters. In addition, optical imaging is qualitative in nature and semiquantitative at best. Therefore, the use of quantitative imaging techniques capable of deep tissue penetration is highly desirable for determining the fate of f‑HMSNs in vivo. PET is sensitive, quantitative, clinically relevant and has excellent tissue penetration [56–60], making it a highly desirable imaging technique for in vivo investigation of f‑HMSNs. PET Owing to the abovementioned advantages of PET over other imaging modalities (e.g., optical and MRI), labeling nanoparticles with positronemitting radionuclides has been generally recognized as the most accurate means for noninvasive evaluation of their biodistribution and pharmacokinetics [61,62]. Since radiolabeled nanoparticles (especially with PET isotopes) represent a newly emerged research area over the last decade, to the best of our knowledge, no reports on radiolabeled HMSNs exist in the literature to date. We have successfully developed novel antibodyconjugated and 64Cu (half-life = 12.7 h) labeled HMSNs, denoted as 64Cu-NOTA-HMSNTRC105 (Figure 4A & B), and demonstrated the proof-of-principle for in vivo active tumor vasculature targeting, which can be monitored with noninvasive serial PET imaging (Figure 4C). Since many nanoparticles suffer from poor extra vasation in the tumor tissue [61,62], tumor vasculature targeting was adopted where extravasation is not required to observe the tumor signal [63–65]. CD105 (also called endoglin) was chosen as the vascular target, which is almost exclusively expressed on proliferating tumor endothelial cells [66,67], and TRC105 (a human/murine chimeric IgG1 monoclonal antibody that binds to both human and murine CD105 [68]) was used as the targeting ligand. Significantly higher uptake in 4T1 murine breast tumors (which express high level of CD105 on the tumor vasculature [69,70]) www.futuremedicine.com
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Figure 4. In vivo PET imaging of radiolabeled hollow mesoporous silica nanoparticles. (A) A transmission electron microscopy image of a HMSN. (B) 64Cu-NOTA-HMSN-TRC105. (C) Serial coronal PET images of 4T1 breast tumor‑bearing mice at different time points postinjection of 64 Cu-NOTA-HMSN-TRC105 or 64Cu-NOTA-HMSN. Tumors are indicated by arrows. HMSN: Hollow mesoporous silica nanoparticle. For color images see online at www.futuremedicine.com/doi/full/10.2217/NNM.13.177.
was observed for 64 Cu-labeled, TRC105conjugated HMSNs, which is approximately twofold that for the nontargeted HMSNs (Figure 4C). With demonstrated active tumor targeting efficiency in vivo and high drug loading capacity, 64 Cu-NOTA-HMSN-TRC105 is a promising platform for future image-guided, tumor-targeted drug delivery and more efficacious cancer therapy. Although PET imaging is highly sensitive and quantitative for in vivo applications, its spatial resolution (milimeter level) is significantly lower than that of MRI (typically
