Development of optical nanotheranostics for the capability of photodynamic therapy (PDT) provides opportunities for advanced cancer therapy. However, most nanotheranostic systems fail to regulate their generation levels of reactive oxygen species (ROS) according to the disease microenvironment, which can potentially limit their therapeutic selectivity and increase the risk of damage to normal tissues. We herein report the development of hybrid semiconducting polymer nanoparticles (SPNs) with self-regulated near-infrared (NIR) photodynamic properties for optimized cancer therapy. The SPNs comprise a binary component nanostructure: a NIR-absorbing semiconducting polymer acts as the NIR fluorescent PDT agent, while nanoceria serves as the smart intraparticle regular to decrease and increase ROS generation at physiologically neutral and pathologically acidic environments, respectively. As compared with nondoped SPNs, the NIR fluorescence imaging ability of nanoceria-doped SPNs is similar due to the optically inactive nature of nanoceria; however, the self-regulated photodynamic properties of nanoceria-doped SPN not only result in dramatically reduced nonspecific damage to normal tissue under NIR laser irradiation but also lead to significantly enhanced photodynamic efficacy for cancer therapy in a murine mouse model. This study thus provides a simple yet effective hybrid approach to modulate the phototherapeutic performance of organic photosensitizers.Keywords: cancer therapy; nanomedicine; near-infrared light; photodynamic therapy; polymer nanoparticles;

Photoacoustic (PA) imaging holds great promise for preclinical research and clinical practice. However, most studies rely on the laser wavelength in the first near-infrared (NIR) window (NIR-I, 650–950 nm), while few studies have been exploited in the second NIR window (NIR-II, 1000–1700 nm), mainly due to the lack of NIR-II absorbing contrast agents. We herein report the synthesis of a broadband absorbing PA contrast agent based on semiconducting polymer nanoparticles (SPN-II) and apply it for PA imaging in NIR-II window. SPN-II can absorb in both NIR-I and NIR-II regions, providing the feasibility to directly compare PA imaging at 750 nm with that at 1064 nm. Because of the weaker background PA signals from biological tissues in NIR-II window, the signal-to-noise ratio (SNR) of SPN-II resulted PA images at 1064 nm can be 1.4-times higher than that at 750 nm when comparing at the imaging depth of 3 cm. The proof-of-concept application of NIR-II PA imaging is demonstrated in in vivo imaging of brain vasculature in living rats, which showed 1.5-times higher SNR as compared with NIR-I PA imaging. Our study not only introduces the first broadband absorbing organic contrast agent that is applicable for PA imaging in both NIR-I and NIR-II windows but also reveals the advantages of NIR-II over NIR-I in PA imaging.Keywords: brain imaging; photoacoustic imaging; Polymer nanoparticles; second near-infrared window;

Upregulation of highly reactive oxygen species (ROS) such as hypochlorite (ClO–) is associated with many pathological conditions including cardiovascular diseases, neuron degeneration, lung injury, and cancer. However, real-time imaging of ClO– is limited to the probes generally relying on fluorescence with shallow tissue-penetration depth. We here propose a self-assembly approach to develop activatable and degradable photoacoustic (PA) nanoprobes for in vivo imaging of ClO–. A near-infrared absorbing amphiphilic oligomer is synthesized to undergo degradation in the presence of a specific ROS (ClO–), which integrates a π-conjugated but ClO– oxidizable backbone with hydrophilic PEG side chains. This molecular architecture allows the oligomer to serve as a degradable nanocarrier to encapsulate the ROS-inert dye and self-assemble into structurally stable nanoparticles through both π–π stacking and hydrophobic interactions. The self-assembled nanoprobe exhibits sensitive and specific ratiometric PA signals toward ClO–, permitting ratiometric PA imaging of ClO– in the tumor of living mice.Keywords: activatable probes; near-infrared dyes; photoacoustic imaging; polymer nanoparticles; reactive oxygen species; self-assembly; sensors;

Semiconducting polymer nanoparticles (SPNs) have emerged as an alternative class of optical nanoagents for imaging applications. However, the general preparation method of SPNs is nanoprecipitation, which is likely to encounter the issue of nanoparticle dissociation. We herein report nondissociable near-infrared (NIR)-absorbing organic semiconducting nanoparticles for in vivo photoacoustic (PA) and fluorescence imaging. The nanoparticles are self-assembled from an amphiphilic semiconducting oligomer (ASO) that has a hydrophobic semiconducting oligomer backbone attached by hydrophilic poly(ethylene glycol) (PEG) side chains. The ASO has a higher structural stability and brighter PA signals compared to those of its counterpart nanoparticles synthesized by nanoprecipitation. The small size and the PEG-passivated surface of the ASO allow it to passively target to and efficiently accumulate in the tumor of living mice, permitting tumor imaging with high signal-to-background ratios. Our study provides new NIR-absorbing organic nanoparticles for PA and fluorescence imaging, which also have the potential to be used as a drug carrier for theranostics.Keywords: fluorescence imaging; nanoparticles; photoacoustic imaging; self-assembly; semiconducting oligomer;

Protein sulfenic acids play a key role in oxidative signal transduction of many biological and pathological processes; however, current chemical tools rely on visible fluorescence signals, limiting their utility to in vitro assays. We herein report reaction-based semiconducting polymer nanoprobes (rSPNs) with near-infrared absorption for in vivo photoacoustic (PA) imaging of protein sulfenic acids. rSPNs comprise an optically active semiconducting polymer as the core shielded with inert silica and poly(ethylene glycol) corona. The sulfenic acid reactive group (1,3-cyclohexanedione) is efficiently conjugated to the surface of nanoparticles via click chemistry. Such a nanostructure enables the specific recognition reaction between rSPNs and protein sulfenic acids without compromising the fluorescence and PA properties. In addition to in vitro tracking of the production of protein sulfenic acids in cancer cells under oxidative stress, rSPNs permit real-time PA and fluorescence imaging of protein sulfenic acids in tumors of living mice. This study thus not only demonstrates the first reaction-based PA probes with submolecular level recognition ability but also highlights the opportunities provided by hybrid nanoparticles for advanced molecular imaging.Keywords: organic nanoparticles; photoacoustic imaging; reaction-based probes; sulfenic acids;

Afterglow or persistent luminescence eliminates the need for light excitation and thus circumvents the issue of autofluorescence, holding promise for molecular imaging. However, current persistent luminescence agents are rare and limited to inorganic nanoparticles. This study reports the design principle, synthesis, and proof-of-concept application of organic semiconducting nanoparticles (OSNs) with ultralong phosphorescence for in vivo afterglow imaging. The design principle leverages the formation of aggregates through a top-down nanoparticle formulation to greatly stabilize the triplet excited states of a phosphorescent molecule. This prolongs the particle luminesce to the timescale that can be detected by the commercial whole-animal imaging system after removal of external light source. Such ultralong phosphorescent of OSNs is inert to oxygen and can be repeatedly activated, permitting imaging of lymph nodes in living mice with a high signal-to-noise ratio. This study not only introduces the first category of water-soluble ultralong phosphorescence organic nanoparticles but also reveals a universal design principle to prolong the lifetime of phosphorescent molecules to the level that can be effective for molecular imaging.

AbstractRegulation of transgene systems is needed to develop innovative medicines. However, noninvasive remote control of gene expression has been rarely developed and remains challenging. We herein synthesize a near-infrared (NIR) absorbing dendronized semiconducting polymer (DSP) and utilize it as a photothermal nanocarrier not only to efficiently deliver genes but also to spatiotemporally control gene expression in conjunction with heat-inducible promoter. DSP has a high photothermal conversion efficiency (44.2 %) at 808 nm, permitting fast transduction of NIR light into thermal signals for intracellular activation of transcription. Such a DSP-mediated remote activation can rapidly and safely result in 25- and 4.5-fold increases in the expression levels of proteins in living cells and mice, respectively. This study thus provides a promising approach to optically regulate transgene systems for on-demand therapeutic transgene dosing.

AbstractRegulation of transgene systems is needed to develop innovative medicines. However, noninvasive remote control of gene expression has been rarely developed and remains challenging. We herein synthesize a near-infrared (NIR) absorbing dendronized semiconducting polymer (DSP) and utilize it as a photothermal nanocarrier not only to efficiently deliver genes but also to spatiotemporally control gene expression in conjunction with heat-inducible promoter. DSP has a high photothermal conversion efficiency (44.2 %) at 808 nm, permitting fast transduction of NIR light into thermal signals for intracellular activation of transcription. Such a DSP-mediated remote activation can rapidly and safely result in 25- and 4.5-fold increases in the expression levels of proteins in living cells and mice, respectively. This study thus provides a promising approach to optically regulate transgene systems for on-demand therapeutic transgene dosing.

Hypochlorite (ClO−) as a highly reactive oxygen species not only acts as a powerful “guarder” in innate host defense but also regulates inflammation-related pathological conditions. Despite the availability of fluorescence probes for detection of ClO− in cells, most of them can only detect ClO− in single cellular organelle, limiting the capability to fully elucidate the synergistic effect of different organelles on the generation of ClO−. This study proposes a nanoprobe cocktail approach for multicolor and multiorganelle imaging of ClO− in cells. Two semiconducting oligomers with different π-conjugation length are synthesized, both of which contain phenothiazine to specifically react with ClO− but show different fluorescent color responses. These sensing components are self-assembled into the nanoprobes with the ability to target cellular lysosome and mitochondria, respectively. The mixture of these nanoprobes forms a nano-cocktail that allows for simultaneous imaging of elevated level of ClO− in lysosome and mitochondria according to fluorescence color variations under selective excitation of each nanoprobe. Thus, this study provides a general concept to design probe cocktails for multilocal and multicolor imaging.

AbstractSmart molecular probes that emit deep-tissue penetrating photoacoustic (PA) signals responsive to the target of interest are imperative to understand disease pathology and develop innovative therapeutics. This study reports a self-assembly approach to develop semiconducting macromolecular activatable probe for in vivo imaging of reactive oxygen species (ROS). This probe comprises a near-infrared absorbing phthalocyanine core and four poly(ethylene glycol) (PEG) arms linked by ROS-responsive self-immolative segments. Such an amphiphilic macromolecular structure allows it to undergo an ROS-specific cleavage process to release hydrophilic PEG and enhance the hydrophobicity of the nanosystem. Consequently, the residual phthalocyanine component self-assembles and regrows into large nanoparticles, leading to ROS-enhanced PA signals. The small size of the intact macromolecular probe is beneficial to penetrate into the tumor tissue of living mice, while the ROS-activated regrowth of nanoparticles prolongs the retention along with enhanced PA signals, permitting imaging of ROS during chemotherapy. This study thus capitalizes on stimuli-controlled self-assembly of macromolecules in conjunction with enhanced heat transfer in large nanoparticles for the development of smart molecular probes for PA imaging.

Chemo-photothermal nanotheranostics has the advantage of synergistic therapeutic effect, providing opportunities for optimized cancer therapy. However, current chemo-photothermal nanotheranostic systems generally comprise more than three components, encountering the potential issues of unstable nanostructures and unexpected conflicts in optical and biophysical properties among different components. We herein synthesize an amphiphilic semiconducting polymer (PEG-PCB) and utilize it as a multifunctional nanocarrier to simplify chemo-photothermal nanotheranostics. PEG-PCB has a semiconducting backbone that not only serves as the diagnostic component for near-infrared (NIR) fluorescence and photoacoustic (PA) imaging, but also acts as the therapeutic agent for photothermal therapy. In addition, the hydrophobic backbone of PEG-PCB provides strong hydrophobic and π-π interactions with the aromatic anticancer drug such as doxorubicin for drug encapsulation and delivery. Such a trifunctionality of PEG-PCB eventually results in a greatly simplified nanotheranostic system with only two components but multimodal imaging and therapeutic capacities, permitting effective NIR fluorescence/PA imaging guided chemo-photothermal therapy of cancer in living mice. Our study thus provides a molecular engineering approach to integrate essential properties into one polymer for multimodal nanotheranostics.Download high-res image (233KB)Download full-size image

Development of photoacoustic (PA) imaging agents is crucial to advancing PA imaging in biology and medicine. In this study, we report the design and synthesis of near-infrared (NIR) absorbing amphiphilic semiconducting polymers that can spontaneously self-assemble into homogeneous water-soluble nanoparticles for PA imaging of tumor in living mice.

Molecular probes that change their signals in response to the target of interest have a critical role in fundamental biology and medicine. Semiconducting polymer nanoparticles (SPNs) have recently emerged as a new generation of purely organic photonic nanoagents with desirable properties for biological applications. In particular, tunable optical properties of SPNs allow them to be developed into photoluminescence, chemiluminescence, and photoacoustic probes, wherein SPNs usually serve as the energy donor and internal reference for luminescence and photoacoustic probes, respectively. Moreover, facile surface modification and intraparticle engineering provide the versatility to make them responsive to various biologically and pathologically important substances and indexes including small-molecule mediators, proteins, pH and temperature. This article focuses on recent advances in the development of SPN-based activatable molecular probes for sensing and imaging. The designs and applications of these probes are discussed in details, and the present challenges to further advance them into life science are also analyzed.

Optogenetics provides powerful means for precise control of neuronal activity; however, the requirement of transgenesis and the incapability to extend the neuron excitation window into the deep-tissue-penetrating near-infrared (NIR) region partially limit its application. We herein report a potential alternative approach to optogenetics using semiconducting polymer nanobioconjugates (SPNsbc) as the photothermal nanomodulator to control the thermosensitive ion channels in neurons. SPNsbc are designed to efficiently absorb the NIR light at 808 nm and have a photothermal conversion efficiency higher than that of gold nanorods. By virtue of the fast heating capability in conjunction with the precise targeting to the thermosensitive ion channel, SPNsbc can specifically and rapidly activate the intracellular Ca2+ influx of neuronal cells in a reversible and safe manner. Our study provides an organic nanoparticle based strategy that eliminates the need for genetic transfection to remotely regulate cellular machinery.

Although organic semiconducting polymer nanoparticles (SPNs) have emerged as an important category of optical imaging agents, their application in molecular imaging is still in its infancy and faces many challenges. We herein report a straightforward one-pot synthetic approach to construct multilayered near-infrared (NIR) fluorescent SPNs with enhanced fluorescence and optimized biodistribution for in vivo molecular imaging. In addition to the SP core, the multilayered SPNs have a middle silica protection layer and an outer poly(ethylene glycol) (PEG) corona, which play crucial roles in enhancing the NIR fluorescence by up to ∼100 fold and reducing nonspecific interactions, respectively. Their proof-of-concept imaging applications are demonstrated in cells, zebrafish and living mice. The multilayered nanoarchitecture not only permits in vivo lymph node tracking with an ultrahigh signal-to-noise ratio (∼85), but also allows for more sensitive in vivo imaging of tumors with a fluorescence intensity ratio of tumor to liver that is ∼8-fold higher compared to that of the counterpart silica SPN. Thus, this study provides a simple yet effective nanoengineering approach to facilitate the application of SPNs in molecular imaging.

Optical theranostic nanoagents that seamlessly and synergistically integrate light-generated signals with photothermal or photodynamic therapy can provide opportunities for cost-effective precision medicine, while the potential for clinical translation requires them to have good biocompatibility and high imaging/therapy performance. We herein report an intraparticle molecular orbital engineering approach to simultaneously enhance photoacoustic brightness and photothermal therapy efficacy of semiconducting polymer nanoparticles (SPNs) for in vivo imaging and treatment of cancer. The theranostic SPNs have a binary optical component nanostructure, wherein a near-infrared absorbing semiconducting polymer and an ultrasmall carbon dot (fullerene) interact with each other to induce photoinduced electron transfer upon light irradiation. Such an intraparticle optoelectronic interaction augments heat generation and consequently enhances the photoacoustic signal and maximum photothermal temperature of SPNs by 2.6- and 1.3-fold, respectively. With the use of the amplified SPN as the theranostic nanoagent, it permits enhanced photoacoustic imaging and photothermal ablation of tumor in living mice. Our study thus not only introduces a category of purely organic optical theranostics but also highlights a molecular guideline to amplify the effectiveness of light-intensive imaging and therapeutic nanosystems.Keywords: photoacoustic imaging; photothermal therapy; semiconducting polymer nanoparticles; theranostic

Photoacoustic (PA) imaging as a new hybrid imaging modality holds great promise for real-time in vivo monitoring of biological processes with deep tissue penetration and high spatial resolution. To endow PA imaging with the ability to provide real-time molecular information at disease sites, molecular probes that can change their PA signals responding to the target or event of interest have to be developed. This review focuses on the recent development of smart activatable PA probes for molecular imaging. A brief summary of PA imaging agents is given first, followed by the detailed discussion of the contemporary design approaches toward activatable PA probes for different imaging applications. At last, the current challenges are highlighted.

Detection of reactive oxygen species (ROS), a hallmark of many pathological processes, is imperative to understanding, detection and treatment of many life-threatening diseases. However, methods capable of real-time in situ imaging of ROS in living animals are still very limited. We herein report the development and optimization of chemiluminescent semiconducting polymer nanoparticles (SPNs) for ultrasensitive in vivo imaging of hydrogen peroxide (H2O2). The chemiluminescence is amplified by adjusting the energy levels between the luminescence reporter and the chemiluminescence substrate to facilitate intermolecular electron transfer in the process of H2O2-activated luminescence. The optimized SPN can emit chemiluminescence with the quantum yield up to 2.30 × 10–2 einsteins/mol and detect H2O2 down to 5 nM, which substantially outperforms the previous probes. Further doping of this SPN with a naphthalocyanine dye creates intraparticle chemiluminescence resonance energy transfer (CRET), leading to the near-infrared (NIR) luminescence responding to H2O2. By virtue of high brightness and ideal NIR optical window, SPN-NIR permits ultrasensitive imaging of H2O2 in the mouse models of peritonitis and neuroinflammation with the minute administration quantity. Thus, this study not only provides a category of optical probes that eliminates the need of external light excitation for imaging of H2O2, but also reveals the underlying principle to enhance the brightness of chemiluminescence systems.Keywords: chemiluminescence; molecular imaging; organic nanoparticles; reactive oxygen species

Development of photoacoustic (PA) imaging agents is crucial to advancing PA imaging in biology and medicine. In this study, we report the design and synthesis of near-infrared (NIR) absorbing amphiphilic semiconducting polymers that can spontaneously self-assemble into homogeneous water-soluble nanoparticles for PA imaging of tumor in living mice.

Although organic semiconducting polymer nanoparticles (SPNs) have emerged as an important category of optical imaging agents, their application in molecular imaging is still in its infancy and faces many challenges. We herein report a straightforward one-pot synthetic approach to construct multilayered near-infrared (NIR) fluorescent SPNs with enhanced fluorescence and optimized biodistribution for in vivo molecular imaging. In addition to the SP core, the multilayered SPNs have a middle silica protection layer and an outer poly(ethylene glycol) (PEG) corona, which play crucial roles in enhancing the NIR fluorescence by up to ∼100 fold and reducing nonspecific interactions, respectively. Their proof-of-concept imaging applications are demonstrated in cells, zebrafish and living mice. The multilayered nanoarchitecture not only permits in vivo lymph node tracking with an ultrahigh signal-to-noise ratio (∼85), but also allows for more sensitive in vivo imaging of tumors with a fluorescence intensity ratio of tumor to liver that is ∼8-fold higher compared to that of the counterpart silica SPN. Thus, this study provides a simple yet effective nanoengineering approach to facilitate the application of SPNs in molecular imaging.