Tumor Microenvironment Stimuli-Responsive Nanoparticles for Programmed Anticancer Drug Delivery.

It is well-known that large size nanoparticles stay for a long time in the circulation system, but show poor tissue penetration and low cellular uptake. In order to reconcile the conflicting needs for extended circulation time, extensive tumor tissue penetration, and enhanced cellular uptake for nanodrug delivery systems, we designed DOX-containing hypersensitive nanoparticles that responded to the tumor microenvironment for programmed DOX delivery. A supersensitive polymer material, poly(2-ethyl-2-oxazoline)-poly(methacryloyl sulfadimethoxine), was synthesized (PEOz-b-PSD, pKa = 6.96). At the physiological environment, PEOz-b-PSD and polyamidoamine/DOX (PAMAM/DOX) can form nanoparticles, PEOz-b-PSD/PAMAM/DOX (PEPSD/PAM/DOX), via electrostatic adsorption. The PEPSD/PAM/DOX has an intact structure, which can prolong circulation time. While in the tumor environment, the PEOz-b-PSD was rapidly protonated and showed charge reversal, leading the detachment of PEOz-b-PSD from the nanoparticles; then the large size nanoparticles with a negative charge (PEPSD/PAM/DOX) instantaneously turn into positively charged ultrafine nanoparticles. The sudden inversion of size and charge can effectively improve tumor accumulation and internal penetration. After entering tumor cells, nanoparticles can release drugs quickly through the action of a PAMAM proton sponge, resulting in enhanced tumor inhibition. Our results proved that the programmed nanoparticles could remarkably enhance the in vivo antitumor efficacy and reduce cardiotoxicity of DOX. This study designed ultrasensitive nanoparticles in the tumor microenvironment, which appear to be beneficial for enhancing the treatment efficacy of DOX in solid tumors.

Near-infrared Light-Triggered Size-Shrinkable theranostic nanomicelles for effective tumor targeting and regression.

Most nanomedicines with suitable sizes (normally 100-200 nm) exhibit favorable accumulation in the periphery of tumors but hardly penetrate into deep tumors. Effective penetration of nanomedicines requires smaller sizes (less than 30 nm) to overcome the elevated tumor interstitial fluid pressure. Moreover, integrating an efficient diagnostic agent in the nanomedicines is in high demand for precision theranostics of tumors. To this end, a near-infrared light (NIR) -triggered size-shrinkable micelle system (Fe3O4@AuNFs/DOX-M) coloaded antitumor drug doxorubicin (DOX) and biomodal imaging agent magnetic gold nanoflower (Fe3O4@AuNFs) was developed to achieve efficient theranostic of tumors. Upon the accumulation of Fe3O4@AuNFs/DOX-M in the tumor periphery, a NIR laser was irradiated near the tumor sites, and the loaded Fe3O4@Au NFs could convert the light energy to heat, which triggered the cleavage of DOX-M to the ultra-small micelles (∼5 nm), thus realizing the deep penetration of micelles and on-demand drug release. Moreover, Fe3O4@AuNFs in the micelles could also be used as CT/MRI dual-modal contrast agent to "visualize" the tumor. Up to 92.6 % of tumor inhibition was achieved for the developed Fe3O4@AuNFs/DOX-M under NIR irradiation. This versatile micelle system provided a promising drug carrier platform realizing efficient tumor dual-modal diagnosis and photothermal-chemotherapy integration.

Dismantlable Coronated Nanoparticles for Coupling the Induction and Perception of Immunogenic Cell Death.

Therapy-induced immunogenic cell death (ICD) can initiate both innate and adaptive immune responses for amplified anti-tumor efficacy. However, dying cell-released ICD signals are prone to being sequestered by the TIM-3 receptors on dendritic cell (DC) surfaces, preventing immune surveillance. Herein, dismantlable coronated nanoparticles (NPs) are fabricated as a type of spatiotemporally controlled nanocarriers for coupling tumor cell-mediated ICD induction to DC-mediated immune sensing. These NPs are loaded with an ICD inducer, mitoxantrone (MTO), and wrapped by a redox-labile anti-TIM-3 (αTIM-3) antibody corona, forming a separable core-shell structure. The antibody corona disintegrates under high levels of extracellular reactive oxygen species in the tumor microenvironment, exposing the MTO-loaded NP core for ICD induction and releasing functional αTIM-3 molecules for DC sensitization. Systemic administration of the coronated NPs augments DC maturation, promotes cytotoxic T cell recruitment, enhances tumor susceptibility to immune checkpoint blockade, and prevents the side effects of MTO. This study develops a promising nanoplatform to unleash the potential of host immunity in cancer therapy.

Applications and challenges of ultra-small particle size nanoparticles in tumor therapy.

With the development of nanotechnology, nanomedicines are widely used in tumor therapy. However, biological barriers in the delivery of nanoparticles still limit their application in tumor therapy. As one of the most fundamental properties of nanoparticles, particle size plays a crucial role in the process of the nanoparticles delivery process. It is difficult for large size nanoparticles with fixed size to achieve satisfactory outcomes in every process. In order to overcome the poor penetration of larger size, nanoparticles with ultra-small particle size are proposed, which are more conducive to deep tumor penetration and uniform drug distribution. In this review, the latest progresses and advantages of ultra-small nanoparticles are systematically summarized, the perspectives and challenges of ultra-small nanoparticles strategy for cancer treatment are also discussed.

Cellular Hypoxia Mitigation by Dandelion-like Nanoparticles for Synergistic Photodynamic Therapy of Oral Squamous Cell Carcinoma.

Hypoxia at the tumor site limits the therapeutic effects of photodynamic therapy (PDT) in oral squamous cell carcinoma (OSCC), which is an oxygen-consumption process. Inhibiting cellular oxygen consumption and reducing cellular ATP production are expected to enhance PDT. In this study, we designed and constructed dandelion-like size-shrinkable nanoparticles for tumor-targeted delivery of hypoxia regulator resveratrol (RES) and photodynamic agent chlorine e6 (CE6). Both drugs were co-encapsulated in small-sized micelles modified with EGFR targeting ligand GE11, which was further conjugated on hyaluronic nanogel (NG) to afford RC-GMN. After targeted accumulation in tumors mediated by GE11 and enhanced penetration and retention (EPR) effects, RC-GMN was degraded by hyaluronidase (HAase) and resulted in small-sized micelles, allowing for deep penetration and dual-receptor-mediated cellular internalization. Resveratrol inhibited cellular oxygen consumption and provided sufficient oxygen for PDT, which consequently activated PDT to produce reactive oxygen species (ROS). Notably, we found that autophagy was overactivated in PDT, which was further strengthened by the hypoxia regulator resveratrol, elevating autophagic cell death. The synergistic effects of resveratrol and CE6 promoted autophagic cell death and apoptosis in the enhanced PDT, resulting in stronger antitumor effects in the orthotopic OSCC model. Therefore, the facilitated delivery of hypoxia regulator enhanced PDT efficacy by elevating oxygen content in tumor cells and inducing autophagic cell death and apoptosis, which offers an alternative strategy for enhancing the PDT effects against OSCC.

Rapid pH-responsive self-disintegrating nanoassemblies balance tumor accumulation and penetration for enhanced anti-breast cancer therapy.

The dilemma of tumor accumulation and deep penetration has always been a barrier in antitumor therapy. Stimuli-responsive size changeable drug delivery systems provide possible solutions. Nevertheless, the low size-shrinkage efficiency limited the antitumor effects. In this study, an instant pH-responsive size shrinkable nanoassemblies named self-aggregated DOX@HA-CD (SA-DOX@HA-CD) was formulated using small-sized hyaluronic acid modified carbon dots (HA-CD) as monomers, which could self-aggregate into raspberry-like structure via hydrophobicity force in neutral pH and rapidly disassemble into shotgun-like DOX-loaded CD monomer in simulated tumor microenvironment (pH 6.5), owing to the transformation in electrical charge and hydrophobicity/hydrophilicity of this system. The transmission electron microscopy showed that the clustered SA-DOX@HA-CD had a diameter of ~150 nm, and thoroughly disassembled into ~30 nm nanoparticles in response to acidic environment. The disassemble efficiency was approximately 100%. Attributed to this property, SA-DOX@HA-CD led to enhanced cellular internalization and accumulation in 4T1 cells in simulated tumor microenvironment, as well as deep tumor penetration in 3D tumor spheroid model. Besides, the imine bond between DOX and HA-CD endowed DOX with pH-responsive release profile in the acidic lysosome environment. Furthermore, in the orthotopic 4T1 tumor-bearing mouse model, SA-DOX@HA-CD demonstrated higher tumor accumulation than non-aggregated DOX-HA-CD. Meanwhile, in response to the acid tumor microenvironment, the dissociated DOX-HA achieved deep tumor penetration, which consequently resulted in 2.5-fold higher antitumor efficiency. The formulation of self-aggregated SA-DOX@HA-CD provides a simple and effective alternative to prepare pH-responsive size-shrinkable nanodrug delivery systems. STATEMENT OF SIGNIFICANCE: The heterogeneity of tumor vasculature and the high tumor interstitial pressure lead to the barriers in tumor accumulation and deep penetration, which calls for opposite properties (e.g. size) of drug delivery systems. To address this dilemma, various size changeable nanoparticles have been developed utilizing special features of tumor microenvironment, such as pH, enzyme and reactive oxygen species. Nevertheless, the current strategies face the problems of incomplete hydrolysis of chemical bonds or insufficient enzyme degradation, which result in only partial size shrinkage, hindering the tumor deep penetration effects. Here we developed a self-assembled nanocluster, which could respond to acidic pH rapidly and thoroughly disassemble into small nanodots due to the alteration of hydrophobicity/hydrophilicity/charge, leading to approximately 100% dissociation. This strategy provides a new concept for design of size changeable drug delivery systems.

A Near-Infrared Laser-Triggered Size-Shrinkable Nanosystem with In Situ Drug Release for Deep Tumor Penetration.

The development of smart size-tunable drug delivery nanoplatform enables the solving of the paradox of inconsistent size-dependence of high tumor accumulation and deep penetration during its delivery process, thus achieving superior cancer treatment efficacy. Herein, we report a size-shrinkable nanomicelle complex system with an initial size of 101 nm enabling effective retention around the tumor periphery and could destruct to ultrasmall nanomicelles triggered by a near-infrared (NIR) laser to realize the deep tumor penetration. The nanomicelle system is consisted of an upper critical solution temperature (UCST)-type block copolymer poly(acrylamide-acrylonitrile)-polyethylene glycol-lipoic acid (p(AAm-co-AN)-g-PEG-LA) encapsulating gold nanorods. Upon the irradiation of the NIR laser at the tumor site, gold nanorods could convert the light energy to heat energy, realizing the photothermal ablation of superficial tumor tissue. Concurrently, the large micelles split into a cascade of ultrasmall micelles (∼7 nm), which could easily penetrate into the deep site of the tumor and achieve the in situ "on-demand" release of the loaded drug to exert superior combined photothermal-chemotherapy of cancer. By the precise manipulation of laser, the micelle complex system realized the hierarchical killing from the superficial-to-deep tumor and achieved almost complete tumor growth inhibition on the established xenograft liver tumor mice model.

Tumor-Microenvironment- Responsive Size-Shrinkable Drug-Delivery Nanosystems for Deepened Penetration Into Tumors.

Over the years, the manipulation and clinical application of drug-delivery nanosystems for cancer diseases have attracted a rapid growth of academic research interests, and some nanodrugs have been approved for clinic application. Although encouraging achievements have been made, the potency of nanomedicines in cancer treatment is far from satisfaction, and one significant reason is the inefficient penetration of nanoparticles into solid tumors. Particle size is one of the most significant features that influence diffusion ability of the drug-delivery system in tumors. Size-shrinkable drug-delivery nanosystems possess a size-switchable property that can achieve passive targeting via the enhanced permeability and retention (EPR) effect and transform into ultrasmall particles in tumors for deep penetration into tumors. The tumor microenvironment is characterized by acidic pH, hypoxia, upregulated levels of enzymes, and a redox environment. In this review, we summarize and analyze the current research progresses and challenges in tumor microenvironment responsive size-shrinkable drug-delivery nanosystems. We further expect to present some meaningful proposals and enlightenments on promoting deep penetration into tumors of nanoparticles.

A carbohydrate mimetic peptide modified size-shrinkable micelle nanocluster for anti-tumor targeting and penetrating drug delivery.

PURPOSE: To deliver the chemotherapeutics through the nanoparticles, the delivery system should accumulate at the tumor site first and then penetrate through the interstitium into the interior. The specific tumor-targeting pathway mediated via the receptor-ligand binding could achieve the desirable accumulation of nanoparticles, and the nanoparticles with smaller sizes were required for penetration. METHODS AND MATERIALS: We constructed a size-shrinkable nanocluster modified with a tumor-targeting motif IF-7 (IF-7-MNC) based on a pH-sensitive framework which could be disintegrated in an acid environment to release the micelles aggregated inside. The micelles were constructed by amphiphilic block copolymers PEG-PLA to encapsulate paclitaxel (PTX), while the cross-linked framework consisting of TPGS-PEI was used as a net to gather and release micelles. This nanoplatform could specifically bind with the tumor receptor Annexin A1 through the ligand IF-7 and then shrunk into small micelles with a desirable size for penetration. CONCLUSION: IF-7-MNC of 112.27±6.81 nm could shrink into micelles in PBS (0.01 M, pH 5.0) with sizes of 14.89±0.32 nm. The cellular-uptake results showed that IF-7-MNC could be significantly internalized by A549 cells and HUVEC cells, while the penetration of IF-7-MNC could be more prominent into the 3D-tumor spheroids compared with that of MNC. The biodistribution results displayed that the fluorescence of IF-7-MNC in the tumor site at 24 hrs was 4.5-fold stronger than that of MNC. The results of anti-tumor growth demonstrated that IF-7-MNC was more favorable for the tumor therapy than MNC, where the inhibitory rate of tumor growth was 88.29% in the PTX-loaded IF-7-MNC (IF-7-PMNC) treated group, significantly greater than PMNC treatment group (p<0.05).

Targeting tumor hypoxia with stimulus-responsive nanocarriers in overcoming drug resistance and monitoring anticancer efficacy.

Although existing nanomedicines have focused on the tumor microenvironment with the goal of improving the effectiveness of conventional chemotherapy, the penetration of a tumor's core still represents a formidable barrier for existing drug delivery systems. Therefore, a novel multifunctional hypoxia-induced size-shrinkable nanoparticle has been designed to increase the penetration of drugs, nucleic acids, or probes into tumors. This cooperative strategy relies on three aspects: (i) the responsiveness of nanoparticles to hypoxia, which shrink when triggered by low oxygen concentrations; (ii) the core of a nanoparticle involves an internal cavity and strong positive charges on the surface to deliver both doxorubicin and siRNA; and (iii) a reactive oxygen species (ROS) probe is incorporated in the nanoparticle to monitor its preliminary therapeutic response in real time, which is expected to realize the enhanced efficacy together with the ability to self-monitor the anticancer activity. A more effective inhibition of tumor growth was observed in tumor-bearing zebrafish, demonstrating the feasibility of this cooperative strategy for in vivo applications. This research highlights a promising value in delivering drugs, nucleic acids, or probes to a tumor's core for cancer imaging and treatment. STATEMENT OF SIGNIFICANCE: Hypoxia-induced chemoresistance of tumor cells still represents a formidable barrier, as it is difficult for existing drug delivery systems to penetrate the tumor hypoxia core. This study involves the hypoxia-responsive size-shrinkable nanoparticle co-delivery of DOX and siRNA to enhance the penetration of DOX deep within tumors and subsequently disturb crucial pathways of cancer development induced by hypoxia and to improve sensitization to DOX chemotherapy. Furthermore, the nanopreparation can combine the ROS probe as a self-reporting nanopreparation to realize the function of real-time feedback efficacy, which has a good application prospect in the diagnosis and treatment of cancer.

Coadministration of iRGD with Multistage Responsive Nanoparticles Enhanced Tumor Targeting and Penetration Abilities for Breast Cancer Therapy.

Limited tumor targeting and poor penetration of nanoparticles are two major obstacles to improving the outcome of tumor therapy. Herein, coadministration of tumor-homing peptide iRGD and multistage-responsive penetrating nanoparticles for the treatment of breast cancer are reported. This multistage-responsive nanoparticle, IDDHN, was comprised of an NO donor-modified hyaluronic acid (HN) shell and a small-sized dendrimer, namely, dendri-graft-l-lysine conjugated with doxorubicin and indocyanine (IDD). The results showed that IDDHN could be degraded rapidly from about 330 nm to a smaller size that was in a size range of 35 to 150 nm (most at 35-60 nm) after hyaluronidase (HAase) incubation for 4 h; in vitro cellular uptake demonstrated that iRGD could mediate more endocytosis of IDDHN into 4T1 cells, which was attributed to the overexpression of αvß3 integrin receptor. Multicellular spheroids penetration results showed synergistically enhanced deeper distribution of IDDHN into tumors, with the presence of iRGD, HAase incubation, and NO release upon laser irradiation. In vivo imaging indicated that coadministration with iRGD markedly enhanced the tumor targeting and penetration abilities of IDDHN. Surprisingly, coadministration of IDDHN with iRGD plus 808 nm laser irradiation nearly suppressed all tumor growth. These results systematically revealed the excellent potential of coadministration of iRGD with multistage-responsive nanoparticles for enhancing drug delivery efficiency and overcoming the 4T1 breast cancer.

Enzyme-triggered size shrink and laser-enhanced NO release nanoparticles for deep tumor penetration and combination therapy.

Chemotherapy remains restricted by poor drug delivery efficacy due to the heterogenous nature of tumor. Herein, we presented a novel nanoparticle that could not only response to the tumor microenvironment but also modulate it for deep tumor penetration and combination therapy. The intelligent nanoparticle (IDDHN) was engineered by hyaluronidase (HAase)-triggered size shrinkable hyaluronic acid shells, which were modified with NIR laser sensitive nitric oxide donor (HN), small-sized dendrimeric prodrug (IDD) of doxorubicin (DOX) as chemotherapy agent and indocyanine green (ICG) as photothermal agent into a single nanoparticle. IDDHN displayed synergistic deep penetration both in vitro and in vivo, owing to the enzymatically degradable HN shell mediated by HAase and laser-enhanced NO release triggered deep penetration upon strong hyperthermia effect of ICG under the NIR laser irradiation. The therapeutic effect of IDDHN was verified in 4T1 xenograft tumor model, and IDDHN showed a much better antitumor efficiency with few side effects upon NIR laser irradiation. Therefore, the valid of this study might provide a novel tactic for engineering nanoparticles both response to and modulate the tumor microenvironment for improving penetration and heterogeneity distribution of therapeutic agents in tumor.

Size shrinkable drug delivery nanosystems and priming the tumor microenvironment for deep intratumoral penetration of nanoparticles.

The penetration of nanomedicine into solid tumor still constitutes a great challenge for cancer therapy, which lead to the failure of thorough clearance of tumor cells. Aiming at solving this issue, lots of encouraging progress has been made in the development of multistage nanoparticles triggered by various stimuli in the past few years. Besides, the therapeutical effects of nanoagents are also greatly impacted by the complex tumor microenvironment, and remodeling tumor microenvironment has become another important approach for promoting nanoparticles penetration. In this review, we summarize and analyze recent research progress and challenges in promoting nanoparticle penetration based on two kinds of different strategies, which include size shrinkable nanoparticles and priming tumor microenvironments. Especially, many recent reported multi-strategy approaches based on particle size reduction in conjugated with other therapeutic strategies are discussed. And we expect to provide some useful enlightenments and proposals on nanotechnology-based drug delivery systems for more effective therapy of solid tumors.

A dual strategy to improve the penetration and treatment of breast cancer by combining shrinking nanoparticles with collagen depletion by losartan.

Although development of nanomedicines has been a promising direction in tumor treatment, the therapeutic outcome of current nanomedicines is unsatisfying, partly because of the poor retention and penetration in tumors. Recently, a kind of tumor microenvironment sensitive size shrinkable nanoparticles (DOX-AuNPs-GNPs) has been developed by our lab, which could enhance the tumor penetration and retention depending on the size shrinking. However, the further enhancement is still restricted by dense collagen network in tumors. Thus in this study, we combined DOX-AuNPs-GNPs with losartan to deplete tumor collagen (constituted up to 90% of extracellular matrix) to further improve tumor penetration. In vitro, DOX-AuNPs-GNPs can shrink from over 117.8nm to less than 50.0nm and release DOX-AuNPs under the triggering of tumor overexpressed matrix metalloproteinases-2 (MMP-2). In vivo, pretreatment with losartan significantly decrease the collagen level and improve the tumor penetration. In combination, losartan combined with DOX-AuNPs-GNPs showed the best drug delivery efficiency, striking penetration efficiency and best 4T1 breast tumor inhibition effect. In conclusion, this study provided a promising synergetic strategy to improve the tumor treatment efficiency of nanomedicines. STATEMENT OF SIGNIFICANCE: We have developed a dual strategy for deep tumor penetration through combining size shrinkable DOX-AuNPs-GNPs with depleting tumor collagen by losartan. Additionally, we demonstrate therapeutic efficacy in breast tumor bearing mouse model. DOX-AuNPs-GNPs co-administration with losartan is a novel and highly attractive strategy for anti-tumor drug delivery with the potential for broad applications in clinic.

A Novel Strategy through Combining iRGD Peptide with Tumor-Microenvironment-Responsive and Multistage Nanoparticles for Deep Tumor Penetration.

Despite the great achievements that nanomedicines have obtained so far, deep penetration of nanomedicines into tumors is still a major challenge in tumor treatment. The enhanced permeability and retention (EPR) effect was the main theoretical foundation for using nanomedicines to treat solid tumor. However, the antitumor efficiency is modest because the tumor is heterogeneous, with dense collagen matrix, abnormal tumor vasculature, and lymphatic system. Nanomedicines could only passively accumulate near leaky site of tumor vessels, and they cannot reach the deep region of tumor. To enhance further the tumor penetration efficiency, we developed a novel strategy of coadministering cell-homing penetration peptide iRGD with size-shrinkable and tumor-microenvironment-responsive multistage system (DOX-AuNPs-GNPs) to overcome these barriers. First, iRGD could specifically increase the permeability of tumor vascular and tumor tissue, leading to more DOX-AuNPs-GNPs leaking out from tumor vasculature. Second, the multistage system passively accumulated in tumor tissue and shrank from 131.1 to 46.6 nm to reach the deep region of tumor. In vitro, coadministering iRGD with DOX-AuNPs-GNPs showed higher cellular uptake and apoptosis ratio. In vivo, coadministering iRGD with DOX-AuNPs-GNPs presented higher penetration and accumulation in tumor than giving DOX-AuNPs-GNPs alone, leading to the best antitumor efficiency in 4T1 tumor-bearing mouse model.

Matrix metalloproteinase-sensitive size-shrinkable nanoparticles for deep tumor penetration and pH triggered doxorubicin release.

Nanocarriers are widely used for delivering drugs to tumors and are progressing in a stable trend. The enhanced permeability and retention (EPR) effect has been a key rationale for the development of stimulus-responsive nanocarriers to solid tumor. In this study, we developed a kind of novel nanocarrier, G-AuNPs-DOX-PEG, which was constructed with shrinkable gelatin nanoparticles coated, doxorubicin (DOX) tethered gold nanoparticles and long chain polyethylene glycol (PEG). The particle size of G-AuNPs-DOX-PEG was 186.5 nm with a zeta potential of -4.21 mV and the DOX loading capacity was 9.22%. In vitro, the G-AuNPs-DOX-PEG could be degraded by MMP-2 proteins with a size shrink from 186.5 nm to 59.3 nm. The release of DOX from G-AuNPs-DOX-PEG was in a pH- and time-dependent manner. At pH 5.0, the release of DOX was much quicker than that at high pH value and the cumulative release rate of DOX from G-AuNPs-DOX-PEG was approach 90.9%. Cellular uptake demonstrated that G-AuNPs-DOX-PEG could be internalized via the endosome-mediated pathway. Tumor spheroid penetration and collagen gel diffusion showed G-AuNPs-DOX-PEG with pre-incubation with MMP-2 could significantly enhance its penetrating efficiency. In vivo and ex vivo imaging exhibit that G-AuNPs-DOX-PEG could distribute into 4T1 and B16F10 tumor at a highest intensity. Correspondingly, 4T1 and B16F10 tumor bearing mice treated with G-AuNPs-DOX-PEG displayed the lowest tumor growth rate. In summary, the tumor microenvironment sensitive size-shrinkable G-AuNPs-DOX-PEG could deliver into deep tumor region and then release DOX, resulting in a best anti-tumor effect.