Application of near-infrared-activated and ATP-responsive trifunctional upconversion nano-jelly for in vivo tumor imaging and synergistic therapy.

Upconversion nanoparticles (UCNPs)-mediated in-situ imaging and synergistic therapy may be an effective approach against tumors. However, it remains a challenge to improve therapeutic index and reduce toxicity. Here, we investigated the construction process of a three-layer (core-shell-shell) upconversion nano-jelly hydrogels (UCNJs) coated with stimulus-responsive deoxyribonucleic acid chains, aiming to achieve selective recognition of tumor cells and controlled release of drugs. The UCNJs have a NaYF4: Yb, Er core with an outer silica shell with embedded methylene blue (MB). Then the outer layer was coated with mesoporous silica and loaded with doxorubicin (DOX). Finally, polyacrylamide chains containing anti-adenosine triphosphate (ATP) aptamer sequences were assembled layer-by-layer on the surface of particles to form DNA hydrogels to lock DOX. Under near-infrared irradiation, green light (540 nm) emitted by UCNJs can be used for imaging, while red light (660 nm) is absorbed by MB. The latter generates singlet oxygen, resulting in photodynamic therapy (PDT) effect to inhibit tumor growth. UCNJs also can recognize ATP in tumor cells, leading to hydrogel degradation and DOX release. The hydrogel coating can increase drug-carrying capacity of mesoporous materials and improve biocompatibility. Therefore, the UCNJs has great potential advantages for application in the field of cancer diagnosis and treatment.

Stimulus-responsive and dual-target DNA nanodrugs for rheumatoid arthritis treatment.

Tumor necrosis factor receptor-1 (TNFR1) and DEK are closely associated with the development of rheumatoid arthritis (RA). Taking advantage of the high adenosine triphosphate (ATP) in RA microenvironment and the interactions of DNA aptamers with their targets, an ATP-responsive DNA nanodrug was constructed that simultaneously targets TNFR1 and DEK for RA therapy. To this end, DEK target aptamer DTA and TNFR1 target aptamer Apt1-67 were equipped with sticky ends to hybridize with ATP aptamer (AptATP) and fabricated DNA nanodrug DAT. Our results showed that DAT was successfully prepared with good stability. In the presence of ATP, DAT was disassembled, resulting in the release of DTA and Apt1-67. In vitro studies demonstrated that DAT was superior to the non-responsive DNA nanodrug TD-3A3T in terms of anti-inflammation activity and ATP was inevitable to maximize the anti-inflammation ability of DAT. The superior efficacy of DAT is attributed to the more potent inhibition of caspase-3 and NETs formation. In vivo results further confirmed the anti-RA efficacy of DAT, whereas the administration routes (intravenous injection and transdermal administration via microneedles) did not cause significant differences. Overall, the present study supplies an intelligent strategy for RA therapy and explores a promising administration route for future clinical medication of RA patients.

Bioresponsive Polymers for Nanomedicine-Expectations and Reality!

Bioresponsive polymers in nanomedicine have been widely perceived to selectively activate the therapeutic function of nanomedicine at diseased or pathological sites, while sparing their healthy counterparts. This idea can be described as an advanced version of Paul Ehrlich's magic bullet concept. From that perspective, the inherent anomalies or malfunction of the pathological sites are generally targeted to allow the selective activation or sensory function of nanomedicine. Nonetheless, while the primary goals and expectations in developing bioresponsive polymers are to elicit exclusive selectivity of therapeutic action at diseased sites, this remains difficult to achieve in practice. Numerous research efforts have been undertaken, and are ongoing, to tackle this fine-tuning. This review provides a brief introduction to key stimuli with biological relevance commonly featured in the design of bioresponsive polymers, which serves as a platform for critical discussion, and identifies the gap between expectations and current reality.

Combating multidrug resistance and metastasis of breast cancer by endoplasmic reticulum stress and cell-nucleus penetration enhanced immunochemotherapy.

Rationale: Multidrug resistance (MDR) and metastasis of breast cancer remain major hurdles in clinical anticancer therapy. The unsatisfactory outcome is largely due to insufficient cytotoxicity of chemotherapeutic agents and limited immunogenic cell death (ICD). On the other hand, efflux proteins, especially P-glycoprotein (P-gp), can recognize and promote the efflux of drugs from tumor cells. Methods: In this study, silver nanoparticles (Ag NPs) and peptide- functionalized doxorubicin (PDOX) were used to prepare a theranostic nanocomposite (Ag-TF@PDOX), which induced organelle-mediated immunochemotherapy and drug efflux protein inhibition in drug-resistant breast cancer cells (MCF-7/ADR) via a strategy based on endoplasmic reticulum (ER) stress and cell-nucleus penetration. Results: The silver nanoparticle-triggered persistent activation of ER stress synergizes with chemotherapy to enhance cytotoxicity and stimulate the ICD effect. It has the potential to enhance chemosensitivity by downregulating of P-gp expression due to the increased production of ATP-consuming chaperones. In addition, the novel peptide (CB5005), which not only penetrates the cell membrane but also has a nuclear localization sequence, is conjugated to DOX to improve both cellular internalization and intranuclear accumulation. Moreover, surface TA-Fe3+ engineering endows the nanocomposite with ATP-responsive disassembly and ATP depletion properties to improve biocompatibility and decrease ATP-dependent drug efflux. Ag-TF@PDOX has potential as a dual-mode (PAI/MRI) contrast-enhanced agent for realizing theranostic guidance. Conclusion: This theranostic nanocomposite greatly restricts the growth of drug-resistant breast tumors and activates a strong immune response as well, providing an opportunity for the development of therapeutics that reverse tumor MDR and metastasis at the subcellular level.

Aptamer-based ATP-responsive delivery systems for cancer diagnosis and treatment.

In recent years, many stimuli-triggered drug delivery platforms have been designed to deliver drugs accurately to specific sites and reduce their side effects, improving "on-demand" therapeutic efficacy. Adenosine-5'-triphosphate (ATP)-responsive drug delivery methods are examples of these systems that use ATP molecules as a trigger for delivery of therapeutic agents. Since intra- and extra-cellular ATP concentrations are significantly different from each other (1-10 mM and <0.4 mM, respectively), the use of ATP can be a practical method for regulating drug release. Aptamers possess unique properties including, ligand-specific response, short sequence (~ 20-80 bases) and easy functionalization. Thus, their combination with ATP-responsive systems results in more accurate drug delivery systems and greater control of drug release. A wide range of nanoparticles, such as polymeric nanogels, liposomes, metallic nanoparticles, protein, or DNA nano-assemblies, have been employed in the fabrication of nanocarriers. In this review, we describe several ATP-responsive drug delivery systems based on the various carriers and discuss the challenges and strengths of each method.

mRNA loading into ATP-responsive polyplex micelles with optimal density of phenylboronate ester crosslinking to balance robustness in the biological milieu and intracellular translational efficiency.

Carriers for messenger RNA (mRNA) delivery require propensities to protect the mRNA from enzymatic degradation and to selectively release mRNA in the cytosol for smooth mRNA translation. To meet these requirements, we designed mRNA-loaded polyplex micelles (PMs) with ATP-responsive crosslinking in the inner core by complexing mRNA with poly(ethylene glycol)-polycation block copolymers derivatized with phenylboronic acid and polyol groups, which form crosslinking structures via spontaneous phenylboronate ester formation. PMs thus prepared are tolerable against enzymatic attack and, in turn, disintegrate in the cytosol to release mRNA when triggered by the cleavage of phenylboronate ester linkages in response to elevated ATP concentration. Two structural factors of the PM, including (i) the introduction ratios of phenylboronate ester crosslinkers and (ii) the structure and protonation degree of amino groups in the polycation segment, are critical for maximizing protein expression in cultured cells due to the optimized balance between the robustness in the biological milieu and the ATP-responsive mRNA release in the cytosol. The optimal PM formulation was further stabilized by installing cholesterol moieties into both the mRNA and ω-end of the block copolymer to elicit longevity in blood circulation after intravenous injection.

ATP-Responsive and ATP-Fueled Self-Assembling Systems and Materials.

Adenosine triphosphate (ATP) is a central metabolite that plays an indispensable role in various cellular processes, from energy supply to cell-to-cell signaling. Nature has developed sophisticated strategies to use the energy stored in ATP for many metabolic and non-equilibrium processes, and to sense and bind ATP for biological signaling. The variations in the ATP concentrations from one organelle to another, from extracellular to intracellular environments, and from normal cells to cancer cells are one motivation for designing ATP-triggered and ATP-fueled systems and materials, because they show great potential for applications in biological systems by using ATP as a trigger or chemical fuel. Over the last decade, ATP has been emerging as an attractive co-assembling component for man-made stimuli-responsive as well as for fuel-driven active systems and materials. Herein, current advances and emerging concepts for ATP-triggered and ATP-fueled self-assemblies and materials are discussed, shedding light on applications and highlighting future developments. By bringing together concepts of different domains, that is from supramolecular chemistry to DNA nanoscience, from equilibrium to non-equilibrium self-assembly, and from fundamental sciences to applications, the aim is to cross-fertilize current approaches with the ultimate aim to bring synthetic ATP-dependent systems closer to living systems.

A facile adenosine triphosphate-responsive nanoplatform for efficacious therapy of esophageal cancer.

Current chemotherapeutic agents against esophageal cancer (EC) are suboptimal. To improve treatment efficacy, a nanoplatform based on ATP-responsive drug release was developed for EC therapy. First, the chemotherapeutic agent epirubicin (EPI) was inserted into an ATP aptamer (Ap) to form double-stranded DNA ('DNA duplex'). Subsequently, polyethyleneimine (PEI) was employed to condense the EPI-loaded duplex to construct the final nanoplatform (PEI-Ap-EPI). Following internalization by cancer cells, the EPI-loaded DNA duplex could open and release EPI in an intracellular ATP-rich environment. An in vitro drug-release assay demonstrated that ~50% of EPI was released from PEI-Ap-EPI in an ATP-rich condition. However, only 15% of EPI was released in the presence of a low concentration of ATP. In vitro cytotoxicity and apoptosis assays demonstrated that PEI-Ap-EPI could enhance EPI efficiency against EC cells markedly compared with those in the control group. Therefore, this facile PEI-Ap-EPI nanoplatform may be a promising strategy to improve the efficacy of EPI treatment in EC.

Programing Assembling/Releasing Multifunctional miRNA Nanomedicine to Treat Prostate Cancer.

MicroRNAs (miRNAs) therapy has shown to have great promise for the treatment of androgen-independent prostate cancer (AIPC) due to the low efficiency of hormonal therapy. However, instability of RNA and inefficiency of RNA therapy limit the use of miRNAs in the treatment of AIPC. Here, we report a pH/ATP-activated nanocomplexes for increasing cytosolic delivery of miR146a which can effectively inhibit the expression of epidermal growth factor receptor (EGFR) in AIPC. The nanocomplexes show identical suppressing effect in invasion, colony formation, migration ability, and growth of DU145 cells compared with Lipofectamine 2000 (lipo). But for in vivo experiments, the nanocomplexes vigorously suppress the growth of tumor volumes comparing to lipo group after five weeks' treatment. These results demonstrate the potential of the pH/ATP-activated nanocarriers for AIPC gene therapy.

A smart ATP-responsive chemotherapy drug-free delivery system using a DNA nanostructure for synergistic treatment of breast cancer in vitro and in vivo.

This study demonstrated a chemotherapy drug-free delivery system for breast cancer treatment based on a simple DNA nanostructure composed of sequence 1 containing ATP and AS1411 aptamers and sequence 2 containing antimiR-21. The DNA nanostructure was used for co-delivery of KLA peptide and antimiR-21 as antiapoptotic agents. These therapeutic agents could not be internalised into eukaryotic cells freely which is one of the great features of this targeting platform. The presented delivery system was ATP-responsive, leading to disassembly of the DNA nanostructure in high ATP concentration of cancer cells and restoration of the function of antimiR-21 in these cells. The DNA nanostructure was associated with high cellular uptake by MCF-7 and 4T1 cells due to expression of nucleolin as target of AS1411 on their plasma membranes, while the developed targeting platform could not be internalised into CHO cells because of lack of the active targeting moiety on their surfaces. Furthermore, the results showed that co-delivery of antimiR-21 and KLA peptide using the DNA nanostructure could efficiently prohibit tumour growth in vitro and in vivo and induce a synergistic anticancer activity. Thus, this work provides a new ATP-responsive nanotargeting delivery system and synergistic chemotherapy drug-free regimen for cancer treatment.

Tissue-Specific Regulation of Reactive Oxygen Species by an ATP-Responsive Nanoregulator Enhances Anticancer Efficacy and Reduces Anthracycline-Induced Cardiotoxicity.

Chemotherapy plays an important role in cancer treatment, yet its clinical application is inhibited by side effects. Chemotherapeutic agents accumulate at nonspecific sites and induce oxidative stress damage in noncancer tissues. A selective approach would be ideal, which would not only enhance anticancer efficacy in the tumor sites but also reduce chemotherapy-induced adverse effects on normal tissues. Therefore, we reported an adenosine-5'-triphosphate (ATP)-responsive oxidative stress nanoregulator (DePQu-DOX) to achieve the tissue-specific therapy. The DePQu-DOX NPs coloading doxorubicin (DOX) and quercetin (Qu) enhanced oxidative stress in murine breast cancer cells and scavenged DOX-induced oxygen free radicals in normal cardiac myocytes and podocytes. The released Qu could accelerate free radical scavenging more efficiently in oxygen-rich myocardium than in hypoxic tumors. Additionally, the ATP-specific responsiveness of nanocarriers enable cargos to selectively accumulate at tumor sites and decline the accumulation amount at normal tissues, resulting in lower system toxicity and improved anticancer effects. In vitro and in vivo experiments showed that this oxidative stress nanoregulator could efficiently protect normal tissues and significantly inhibit tumor growth. This study suggests that nanomedicine-mediated oxidative stress regulation could provide selective tumor therapeutics and reduce anthracycline-induced system toxicity.

Polyplexes by Polymerized Dequalinium and Bifunctional Aptamer for Mitochondrial Targeting Drug Release to Overcome Drug Resistance.

Drug resistance is one of the major obstacles to the success of cancer chemotherapy. Mitochondrial targeting drugs are increasingly thought to be able to eradicate resistant cancer cells. However, immature drug release outside mitochondria and the absence of multifunctional targeting carriers against tumor mitochondria greatly limit the corresponding therapeutic benefits. Here, we synthesized polymerized dequalinium by integrating dequalinium, lysine, and poly(ethylene glycol) for mitochondrial targeting. The polymerized dequalinium exhibited lower cytotoxicity and stronger gene condensing ability than free dequalinium. We designed AS1411-ATP fusion aptamer to load doxorubicin (DOX) for both tumor targeting and ATP-responsive DOX release. The polyplexes by polymerized dequalinium and bifunctional aptamer can target tumor cells via AS1411 and show improved stability, mitochondrial targeting, DOX release in response to mitochondrial ATP, and enhanced apoptosis-inducing effect on DOX-resistant MCF-7/DOX cells. The present study highlights a promising application of the polyplexes in reversing drug resistance in tumor cells via tumor mitochondrial targeting drug release.

ATP-activated decrosslinking and charge-reversal vectors for siRNA delivery and cancer therapy.

Stimuli-responsive polycations have been developed for improved nucleic acid transfection and enhanced therapeutic efficacy. The most reported mechanisms for controlled release of siRNA are based on polyelectrolyte exchange reactions in the cytoplasm and the degradation of polycations initiated by specific triggers. However, the degradation strategy has not always been sufficient due to unsatisfactory kinetics and binding of cationic fragments to siRNA, which limits the gene silencing effect. In this study, a new strategy that combines degradation and charge reversal is proposed. Methods: We prepared a polycation (CrossPPA) by crosslinking of phenylboronic acid (PBA)-grafted 1.8k PEI with alginate. It was compared with 25k PEI, 1.8k PEI and 1.8k PEI-PBA on siRNA encapsulation, ATP-responsive behavior and mechanism, cytotoxicity, cell uptake, siRNA transfection, in vivo biodistribution and in vivo anti-tumor efficacy. The in vitro and in vivo experiments were performed on 4T1 murine breast cancer cells and 4T1 tumor model separately. Results: The crosslinking strategy obviously improve the siRNA loading ability of 1.8k PEI. We validated that intracellular levels of ATP could trigger CrossPPA disassembly and charge reversal, which resulted in efficient and rapid siRNA release due to electrostatic repulsion. Besides, CrossPPA/siRNA showed strong cell uptake in 4T1 cells compared with 1.8k PEI/siRNA. Notably, the cytotoxicity of CrossPPA was pretty low, which was owing to its biodegradability. Furthermore, the crosslinked polyplexes significantly enhanced siRNA transfection and improved tumor accumulation. The high gene silencing ability of CrossPPA polyplex led to strong anti-tumor efficacy when using Bcl2-targeted siRNA. Conclusion: These results indicated that the ATP-triggered disassembly and charge reversal strategy provided a new way for developing stimuli-responsive siRNA carriers and showed potential for nucleic acid delivery in the treatment of cancer.

Harnessing Phosphato-Platinum Bonding Induced Supramolecular Assembly for Systemic Cisplatin Delivery.

To improve the therapeutic index of cisplatin (CDDP), we present here a new paradigm of drug-induced self-assembly by harnessing phosphato-platinum complexation. Specifically, we show that a phosphato-platinum cross-linked micelle (PpY/Pt) can be generated by using a block copolymer methoxy-poly(ethylene glycol)-block-poly(l-phosphotyrosine) (mPEG-b-PpY). Coating of PpY/Pt with a R9-iRGD peptide by simple mixing affords a targeting micelle with near neutral-charged surface (iPpY/Pt). The micelles feature in well-controlled sizes below 50 nm and high stability under physiological conditions, and can withstand various environmental stresses. Importantly, the micelles demonstrate on-demand drug release profiles in response to pathological cues such as high ATP concentration and acidic pH. In vitro, the micelles are efficiently internalized and almost equally potent compared to CDDP. Moreover, iPpY/Pt induce greater cytotoxicity than PpY/Pt in a 3D tumor spheroid model likely due to its deeper tumor penetration. In vivo, the micelles exhibit prolonged circulation half-lives, enhanced tumor accumulation, excellent tumor growth inhibition in a xenograft HeLa model and an orthotropic mammary 4T1 model, and improved safety profiles evidenced by the reduced nephrotoxicity. Together, this work demonstrates for the first time that phosphato-platinum complexation can be exploited for effective delivery of CDDP, and suggests a paradigm shift of constructing nanosystems for other anticancer metallodrugs.

ATP-Responsive and Near-Infrared-Emissive Nanocarriers for Anticancer Drug Delivery and Real-Time Imaging.

Stimuli-responsive and imaging-guided drug delivery systems hold vast promise for enhancement of therapeutic efficacy. Here we report an adenosine-5'-triphosphate (ATP)-responsive and near-infrared (NIR)-emissive conjugated polymer-based nanocarrier for the controlled release of anticancer drugs and real-time imaging. We demonstrate that the conjugated polymeric nanocarriers functionalized with phenylboronic acid tags on surface as binding sites for ATP could be converted to the water-soluble conjugated polyelectrolytes in an ATP-rich environment, which promotes the disassembly of the drug carrier and subsequent release of the cargo. In vivo studies validate that this formulation exhibits promising capability for inhibition of tumor growth. We also evaluate the metabolism process by monitoring the fluorescence signal of the conjugated polymer through the in vivo NIR imaging.

ATP-responsive DNA-graphene hybrid nanoaggregates for anticancer drug delivery.

Stimuli-triggered drug delivery systems are primarily focused on the applications of the tumor microenvironmental or cellular physiological cues to enhance the release of drugs at the target site. In this study, we applied adenosine-5'-triphosphate (ATP), the primary "energy molecule", as a trigger for enhanced release of preloaded drugs responding to the intracellular ATP concentration that is significantly higher than the extracellular level. A new ATP-responsive anticancer drug delivery strategy utilizing DNA-graphene crosslinked hybrid nanoaggregates as carriers was developed for controlled release of doxorubicin (DOX), which consists of graphene oxide (GO), two single-stranded DNA (ssDNA, denoted as DNA1 and DNA2) and ATP aptamer. The single-stranded DNA1 and DNA2 together with the ATP aptamer serve as the linkers upon hybridization for controlled assembly of the DNA-GO nanoaggregates, which effectively inhibited the release of DOX from the GO nanosheets. In the presence of ATP, the responsive formation of the ATP/ATP aptamer complex causes the dissociation of the aggregates, which promoted the release of DOX in the environment with a high ATP concentration such as cytosol compared with that in the ATP-deficient extracellular fluid. This supports the development of a novel ATP-responsive platform for targeted on-demand delivery of anticancer drugs inside specific cells.