Mitochondria‐Targeted Cancer Therapy Using a Light‐Up Probe with Aggregation‐Induced‐Emission Characteristics

Subcellular organelle‐specific reagents for simultaneous tumor targeting, imaging, and treatment are of enormous interest in cancer therapy. Herein, we present a mitochondria‐targeting probe (AIE‐mito‐TPP) by conjugating a triphenylphosphine (TPP) with a fluorogen which can undergo aggregation‐induced emission (AIE). Owing to the more negative mitochondrial membrane potential of cancer cells than normal cells, the AIE‐mito‐TPP probe can selectively accumulate in cancer‐cell mitochondria and light up its fluorescence. More importantly, the probe exhibits selective cytotoxicity for studied cancer cells over normal cells. The high potency of AIE‐mito‐TPP correlates with its strong ability to aggregate in mitochondria, which can efficiently decrease the mitochondria membrane potential and increase the level of intracellular reactive oxygen species (ROS) in cancer cells. The mitochondrial light‐up probe provides a unique strategy for potential image‐guided therapy of cancer cells.     

Fluorescent Coumarin–Artemisinin Conjugates as Mitochondria‐Targeting Theranostic Probes for Enhanced Anticancer Activities

Mitochondria‐targeting theranostic probes that enable the simultaneously reporting of and triggering of mitochondrial dysfunctions in cancer cells are highly attractive for cancer diagnosis and therapy. Three fluorescent mitochondria‐targeting theranostic probes have been developed by linking a mitochondrial dye, coumarin‐3‐carboximide, with a widely used traditional Chinese medicine, artemisinin, to kill cancer cells. Fluorescence images showed that the designed coumarin–artemisinin conjugates localized mainly in mitochondria, leading to enhanced anticancer activities over artemisinin. High cytotoxicity against cancer cells correlated with the strong ability to accumulate in mitochondria, which could efficiently increase the intracellular reactive oxygen species level and induce cell apoptosis. This study highlights the potential of using mitochondria‐targeting fluorophores to selectively trigger and directly visualize subcellular drug delivery in living cells.     

Targeting Cancer with PCPA‐Drug Conjugates: LSD1 Inhibition‐Triggered Release of 4‐Hydroxytamoxifen

Targeting cancer with small molecule prodrugs should help overcome problems associated with conventional cancer‐targeting methods. Herein, we focused on lysine‐specific demethylase 1 (LSD1) to trigger the controlled release of anticancer drugs in cancer cells, where LSD1 is highly expressed. Conjugates of the LSD1 inhibitor trans‐2‐phenylcyclopropylamine (PCPA) were used as novel prodrugs to selectively release anticancer drugs by LSD1 inhibition. As PCPA‐drug conjugate (PDC) prototypes, we designed PCPA‐tamoxifen conjugates 1 a and 1 b , which released 4‐hydroxytamoxifen in the presence of LSD1 in vitro. Furthermore, 1 a and 1 b inhibited the growth of breast cancer cells by the simultaneous inhibition of LSD1 and the estrogen receptor without exhibiting cytotoxicity toward normal cells. These results demonstrate that PDCs provide a useful prodrug method that may facilitate the selective release of drugs in cancer cells.     

Synthesis and biological evaluations of mono‐ and bis‐ferrocene uracil derivatives

Mono‐ ( 3a – 3e and 4a – 4e ) and bis‐ferrocene ( 5a – 5e and 6a – 6e ) conjugated 5‐substituted uracil derivatives that are bridged by 1,2,3‐triazole linker were synthesized. The impact of ferrocene unit and spacer between ferrocene and triazole on radical scavenging potency was observed. Bis‐ferrocenyl uracil derivatives exhibited better antiproliferative activities than their mono‐ferrocenyl analogs. Bis‐ferrocenyl methyl‐ ( 5b ) and halogen‐substituted ( 5e , 6c , and 6d ) uracil derivatives showed pronounced and selective cytostatic activities on colon adenocarcinoma (CaCo‐2) and Burkitt lymphoma (Raji) cells, with higher potency and selectivity than the reference drug 5‐fluorouracil. Generation of reactive oxygen species (ROS) in CaCo‐2 and Raji cells when treated with compounds 5b , 5e , and 6d was observed. Bis‐ferrocenyl 5‐chlorouracil 6c induced significant disruption in mitochondrial membrane potential that is accompanied by activation of apoptosis in CaCo‐2, Raji, and acute lymphoblastic leukemia (CCRF‐CEM) cells, while 6d caused mitochondrial dysfunction and apoptosis induction in CaCo‐2 and Raji cells. Potent antiproliferative activity of 6c and 6d could be associated with mitochondrial membrane potential disruption accompanied by apoptosis induction. Our findings highlighted 6c and 6d with potent and selective antiproliferative activity on CaCo‐2, Raji, and CCRF‐CEM cells that may be associated with targeting cancer cell mitochondria, as a molecular target.     

A Drug‐Free Tumor Therapy Strategy: Cancer‐Cell‐Targeting Calcification

Herein, we propose a drug‐free approach to cancer therapy that involves cancer cell targeting calcification (CCTC). Several types of cancer cells, such as HeLa cells, characterized by folate receptor (FR) overexpression, can selectively adsorb folate (FA) molecules and then concentrate Ca²⁺ locally to induce specific cell calcification. The resultant calcium mineral encapsulates the cancer cells, inducing their death, and in vivo assessments confirm that CCTC treatment can efficiently inhibit tumor growth and metastasis without damaging normal cells compared with conventional chemotherapy. Accordingly, CCTC remarkably improve the survival rate of tumor mice. Notably, both FA and calcium ions are essential ingredients in human metabolism, which means that CCTC is a successful drug‐free method for tumor therapy. This achievement may further represent an alternative cancer therapy characterized by selective calcification‐based substitution of sclerosis for tumor disease.     

Sonodynamic Therapeutic Effects of Sonosensitizers with Different Intracellular Distribution Delivered by Hollow Nanocapsules Exhibiting Cytosol Specific Release

Sonodynamic therapy (SDT) is a novel promising noninvasive therapy involving utilization of low‐intensity ultrasound and sonosensitizer, which can generate reactive oxygen species (ROS) by sonication. In SDT, a high therapeutic effect is achieved by intracellular delivery and accumulation at the target sites of sonosensitizer followed by oxidative damage of produced ROS by sonication. Here, pH‐ and redox‐responsive hollow nanocapsules are prepared through the introduction of disulfide cross‐linkages to self‐assembled polymer vesicles formed from polyamidoamine dendron‐poly(l‐ lysine) for the efficient delivery of sonosensitizer. As sonosensitizer, doxorubicin (DOX), an anticancer drug accumulating into cell nucleus, is selected. Also, the conjugate of DOX and triphenylphosphonium (TPP‐DOX) is synthesized as sonosensitizer with mitochondrial targeting ability. DOX and TPP‐DOX are delivered to nucleus and mitochondria by nanocapsules. Furthermore, DOX‐ or TPP‐DOX‐loaded nanocapsules exhibit in vitro sonodynamic therapeutic effect to HeLa cells with sonication, which might be through oxidative damage to nucleus and mitochondria.     

Anticancer Aminoferrocene Derivatives Inducing Production of Mitochondrial Reactive Oxygen Species

Elevated levels of reactive oxygen species (ROS) and deficient mitochondria are two weak points of cancer cells. Their simultaneous targeting is a valid therapeutic strategy to design highly potent anticancer drugs. The remaining challenge is to limit the drug effects to cancer cells without affecting normal ones. We have previously developed three aminoferrocene (AF)-based derivatives, which are activated in the presence of elevated levels of ROS present in cancer cells with formation of electron-rich compounds able to generate ROS and reduce mitochondrial membrane potential (MMP). All of them exhibit important drawbacks including either low efficacy or high unspecific toxicity that prevents their application in vivo up to date. Herein we describe unusual AF-derivatives lacking these drawbacks. These compounds act via an alternative mechanism: they are chemically stable in the presence of ROS, generate mitochondrial ROS in cancer cells, but not normal cells and exhibit anticancer effect in vivo.     

Three‐Arm,Biotin‐Tagged Carbazole–Dicyanovinyl–Chlorambucil Conjugate: Simultaneous Tumor Targeting,Sensing, and Photoresponsive Anticancer Drug Delivery

The design, synthesis, and in vitro biological studies of a biotin–carbazole–dicyanovinyl–chlorambucil conjugate (Bio‐CBZ‐DCV‐CBL; 6 ) are reported. This conjugate ( 6 ) is a multifunctional single‐molecule appliance composed of a thiol‐sensor DCV functionality, a CBZ‐derived phototrigger as well as fluorescent reporter, and CBL as the anticancer drug, and Bio as the cancer‐targeting ligand. In conjugate 6 , the DCV bond undergoes a thiol–ene click reaction at pH<7 with intracellular thiols, thereby shutting down internal charge transfer between the donor CBZ and acceptor DCV units, resulting in a change of the fluorescence color from green to blue, and thereby, sensing the tumor microenvironment. Subsequent photoirradiation results in release of the anticancer drug CBL in a controlled manner.     

Mechanism‐Guided Design and Synthesis of a Mitochondria‐Targeting Artemisinin Analogue with Enhanced Anticancer Activity

Understanding the mechanism of action (MOA) of bioactive natural products will guide endeavor to improve their cellular activities. Artemisinin and its derivatives inhibit cancer cell proliferation, yet with much lower efficiencies than their roles in killing malaria parasites. To improve their efficacies on cancer cells, we studied the MOA of artemisinin using chemical proteomics and found that free heme could directly activate artemisinin. We then designed and synthesized a derivative, ART‐TPP, which is capable of targeting the drug to mitochondria where free heme is synthesized. Remarkably, ART‐TPP exerted more potent inhibition than its parent compound to cancer cells. A clickable probe ART‐TPP‐Alk was also employed to confirm that the attachment of the TPP group could label more mitochondrial proteins than that for the ART derivative without TPP (AP1). This work shows the importance of MOA study, which enables us to optimize the design of natural drug analogues to improve their biological activities.     

Four‐fold Channel‐Nicked Human Ferritin Nanocages for Active Drug Loading and pH‐Responsive Drug Release

Human ferritins are emerging platforms for non‐toxic protein‐based drug delivery, owing to their intrinsic or acquirable targeting abilities to cancer cells and hollow cage structures for drug loading. However, reliable strategies for high‐level drug encapsulation within ferritin cavities and prompt cellular drug release are still lacking. Ferritin nanocages were developed with partially opened hydrophobic channels, which provide stable routes for spontaneous and highly accumulated loading of Fe^II‐conjugated drugs as well as pH‐responsive rapid drug release at endoplasmic pH. Multiple cancer‐related compounds, such as doxorubicin, curcumin, and quercetin, were actively and heavily loaded onto the prepared nicked ferritin. Drugs on these minimally modified ferritins were effectively delivered inside cancer cells with high toxicity.     

GRP78‐Targeted HPMA Copolymer‐Photosensitizer Conjugate for Hyperthermia‐Induced Enhanced Uptake and Cytotoxicity in MCF‐7 Breast Cancer Cells

Photodynamic therapy (PDT) is a promising cancer treatment approach. However, the photosensitizers (PS) used for PDT are often limited by their poor solubility and selectivity for tumors. The goal of this study is to improve water solubility and delivery of the photosensitizer 2‐[1‐hexyloxyethyl]‐2‐divinyl pyropheophorbide‐a (HPPH) to breast cancer cells. An N‐(2‐hydroxypropyl)methacrylamide (HPMA) copolymer–HPPH photosensitizer conjugate is synthesized with heat shock receptor glucose‐regulated protein 78 (GRP78), targeting to GRP78 receptors of MCF‐7 cells, which are upregulated under mild hyperthermia. It is found that the uptake of the GRP78 targeted pep‐HPMA‐HPPH copolymer conjugate in MCF‐7 cells is improved through heat induction. Under mild hyperthermia the targeted copolymers are more effective compared to free HPPH. These results show potential for the utility of mild hyperthermia and copolymer delivery vehicles to enhance the efficacy of photodynamic therapy.     

Carboplatin‐Complexed and cRGD‐Conjugated Unimolecular Nanoparticles for Targeted Ovarian Cancer Therapy

Platinum‐based chemotherapy has been widely used to treat cancers including ovarian cancer; however, it suffers from dose‐limiting toxicity. Judiciously designed drug nanocarriers can enhance the anticancer efficacy of platinum‐based chemotherapy while reducing its systemic toxicity. Herein the authors report a stable and water‐soluble unimolecular nanoparticle constructed from a hydrophilic multi‐arm star block copolymer poly(amidoamine)‐b‐poly(aspartic acid)‐b‐poly(ethylene glycol) (PAMAM‐PAsp‐PEG) conjugated with both cRGD (cyclo(Arg‐Gly‐Asp‐D‐Phe‐Cys) peptide and cyanine5 (Cy5) fluorescent dye as a platinum‐based drug nanocarrier for targeted ovarian cancer therapy. Carboplatin is complexed to the poly(aspartic acid) inner shell via pH‐responsive ion–dipole interactions between carboplatin and the carboxylate groups of poly(aspartic acid). Based on flow cytometry and confocal laser scanning microscopy analyses, cRGD‐conjugated unimolecular nanoparticles exhibit much higher cellular uptake by ovarian cancer cells overexpressing α_vβ₃ integrin than nontargeted (i.e., cRGD‐lacking) ones. Carboplatin‐complexed cRGD‐conjugated nanoparticles also exhibit higher cytotoxicity than nontargeted nanoparticles as well as free carboplatin, while empty unimolecular nanoparticles show no cytotoxicity. These results indicate that stable unimolecular nanoparticles made of individual hydrophilic multi‐arm star block copolymer molecules conjugate with tumor‐targeting ligands and dyes (i.e., PAMAM‐PAsp‐PEG‐cRGD/Cy5) are promising nanocarriers for platinum‐based anticancer drugs for targeted cancer therapy.

     

A Dual‐Targeted Organic Photothermal Agent for Enhanced Photothermal Therapy

The development of highly effective anticancer drugs that cause minimal damage to the surrounding normal tissues is a challenging topic in cancer therapy. Herein, we demonstrate a dual‐targeted organic molecule that functions as a photothermal agent by actively targeting tumor tissue and mitochondria to selectively kill cancer cells. The synthesized photothermal agent exhibited high photothermal conversion efficiency, low cytotoxicity, and good biological compatibility. In vivo experiments showed an excellent tumor inhibitory effect of the dual‐targeted photothermal agent.     

A Small‐Molecule Drug Conjugate for the Treatment of Carbonic Anhydrase IX Expressing Tumors

Antibody–drug conjugates are a very promising class of new anticancer agents, but the use of small‐molecule ligands for the targeted delivery of cytotoxic drugs into solid tumors is less well established. Here, we describe the first small‐molecule drug conjugates for the treatment of carbonic anhydrase IX expressing solid tumors. Using ligand–dye conjugates we demonstrate that such molecules can preferentially accumulate inside antigen‐positive lesions, have fast targeting kinetics and good tumor‐penetrating properties, and are easily accessible by total synthesis. A disulfide‐linked drug conjugate with the maytansinoid DM1 as the cytotoxic payload and a derivative of acetazolamide as the targeting ligand exhibited a potent antitumor effect in SKRC52 renal cell carcinoma in vivo. It was furthermore superior to sunitinib and sorafenib, both small‐molecule standard‐of‐care drugs for the treatment of kidney cancer.     

Engineered Targeted Hyaluronic Acid–Glutathione‐Stabilized Gold Nanoclusters/Graphene Oxide–5‐Fluorouracil as a Smart Theranostic Platform for Stimulus‐Controlled Fluorescence Imaging‐Assisted Synergetic Chemo/Phototherapy

A theranostic platform with integrated diagnostic and therapeutic functions as well as specific targeted and controlled combination therapy to enhance treatment efficacy is of great importance for a wide range of biomedical applications. Here, we first attempted to develop biocompatible hyaluronic acid (HA)–glutathione (GSH) conjugate stabilized gold nanoclusters (GNCs) combined with graphene oxide (GO), accompanied by loading 5‐fluorouracil (5FU), as a novel theranostic platform (HG‐GNCs/GO‐5FU, HG refers to HA‐GSH). Multifunctional HG‐GNCs possessed excellent fluorescence, photosensitivity and specific targeting ability to the cancer cells while their fluorescence and singlet oxygen generation could be strongly inhibited by GO and then effectively restored by lysosomal hyaluronidase in tumor cells. The sustained and complete release of 5FU from HG‐GNCs/GO could also be stimulated successively by enzymatic degradation of HA and light‐induced heat effect of GO under laser irradiation so that turn‐on cell imaging‐assisted synergistic therapeutic strategies associated with triple enzyme/light‐controlled chemo/photothermal/photodynamic therapy could be achieved at the same time, reducing greatly the side effects of materials to normal cells. Our study presents a novel strategy to combine targeting and bioimaging with triple therapies to enhance the antitumor effect.     

DNAzyme‐Loaded Metal–Organic Frameworks (MOFs) for Self‐Sufficient Gene Therapy

DNAzymes have been recognized as potent therapeutic agents for gene therapy, while their inefficient intracellular delivery and insufficient cofactor supply precludes their practical biological applications. Metal–organic frameworks (MOFs) have emerged as promising drug carriers without in‐depth consideration of their disassembled ingredients. Herein, we report a self‐sufficient MOF‐based chlorin e6‐modified DNAzyme (Ce6‐DNAzyme) therapeutic nanosystem for combined gene therapy and photodynamic therapy (PDT). The ZIF‐8 nanoparticles (NPs) could efficiently deliver the therapeutic DNAzyme without degradation into cancer cells. The pH‐responsive ZIF‐8 NPs disassemble with the concomitant release of the guest DNAzyme payloads and the host Zn²⁺ ions that serve, respectively, as messenger RNA‐targeting agent and required DNAzyme cofactors for activating gene therapy. The auxiliary photosensitizer Ce6 could produce reactive oxygen species (ROS) and provide a fluorescence signal for the imaging‐guided gene therapy/PDT.     

Conjugated‐Polyelectrolyte‐Based Polyprodrug: Targeted and Image‐Guided Photodynamic and Chemotherapy with On‐Demand Drug Release upon Irradiation with a Single Light Source

Nanomaterials that combine diagnostic and therapeutic functions within a single nanoplatform are highly desirable for molecular medicine. Herein we report a novel theranostic platform based on a conjugated‐polyelectrolyte (CPE) polyprodrug that contains functionality for image, chemo‐ and photodynamic therapy (PDT), and on‐demand drug release upon irradiation with a single light source. Specifically, the PEGylated CPE serves as a photosensitizer and a carrier, and is covalently conjugated to doxorubicin through a linker that can be cleaved by reactive oxygen species (ROS). Under appropriate light irradiation, the CPE can generate ROS, not only for PDT, but also for on‐demand drug release and chemotherapy. This nanoplatform will offer on‐demand PDT and chemotherapy with drug release triggered by one light switch, which has great potential in cancer treatment.     

Cancer‐Targeted Selenium Nanoparticles Sensitize Cancer Cells to Continuous γ Radiation to Achieve Synergetic Chemo‐Radiotherapy

Cancer radiotherapy with ¹²⁵I seeds demonstrates higher long‐term efficacy and fewer side effects than traditional X‐ray radiotherapy owing to its low‐dose and continuous radiation but is still limited by radioresistance in clinical applications. Therefore, the design and synthesis of sensitizers that could enhance the sensitivity of cancer cells to ¹²⁵I seeds is of great importance for future radiotherapy. Selenium nanoparticles (SeNPs) have been found to exhibit high potential in cancer chemotherapy and as drug carriers. In this study, we found that, based on the Auger‐electron effect and Compton effect of Se atoms, cancer‐targeted SeNPs in combination with ¹²⁵I seeds achieve synergetic effects to inhibit cancer‐cell growth and colony formation through the induction of cell apoptosis and cell cycle arrest. Detailed studies on the action mechanisms reveal that the combined treatments effectively activate intracellular reactive oxygen species (ROS) overproduction to regulate p53‐mediated DNA damage apoptotic signaling pathways and mitogen‐activated protein kinase (MAPK) phosphorylation and to prevent the self‐repair of cancer cells simultaneously. Taken together, the combination of SeNPs with ¹²⁵I seeds could be further exploited as a safe and effective strategy for next‐generation cancer chemo‐radiotherapy in clinical applications.     

An N‐Acetyl Cysteine Ruthenium Tricarbonyl Conjugate Enables Simultaneous Release of CO and Ablation of Reactive Oxygen Species

We have designed and synthesised a [Ru(CO)₃Cl₂(NAC)] pro‐drug that features an N‐acetyl cysteine (NAC) ligand. This NAC carbon monoxide releasing molecule (CORM) conjugate is able to simultaneously release biologically active CO and to ablate the concurrent formation of reactive oxygen species (ROS). Complexes of the general formulae [Ru(CO)₃(L)₃]²⁺, including [Ru(CO)₃Cl(glycinate)] (CORM‐3), have been shown to produce ROS through a water–gas shift reaction, which contributes significantly, for example, to their antibacterial activity. In contrast, NAC‐CORM conjugates do not produce ROS or possess antibacterial activity. In addition, we demonstrate the synergistic effect of CO and NAC both for the inhibition of nitric oxide (formation) and in the expression of tumour‐necrosis factor (TNF)‐α. This work highlights the advantages of combining a CO‐releasing scaffold with the anti‐oxidant and anti‐inflammatory drug NAC in a unique pro‐drug.     

Novel Cyclometalated Fe(II) Complex with NCN Pincer and BODIPY‐Appended 4'‐Ethynyl‐2,2':6',2”‐terpyridine as Mitochondria‐Targeted Photodynamic Anticancer Agents

BODIPY (boron dipyrromethene) derivatives and iron complexes are two types of functional compounds that have found wide applications in the fields of biology and medicine. The new class of cyclometalated Fe(II) complex with NCN pincer and meso‐phenyl‐4'‐ethynyl‐2,2':6',2”‐terpyridine BODIPY ligands of formula [Fe(L)(tpy‐BODIPY)] , 1, in which HL:5‐methoxy‐1,3‐bis (1‐methyl‐1H‐benzo[d]imidazol‐2‐yl)benzene, tpy‐BODIPY: 8‐(4‐phenyl‐4'‐ethynyl‐2,2':6',2”‐terpyridine) BODIPY, has been synthesized and studied as mitochondria‐targeted photodynamic therapy (PDT). Complex 1 showed photocytotoxicity in HeLa cells at 500 nm with low dark toxicity. The phototoxicity of complex 1 on the nontumorigenic MRC‐5 cell line showed the same trend observed for HeLa cells, that is moderately photocytotoxic against the nontumorigenic MRC‐5 cell line (IC₅₀ = 36.21 μM). Moreover, complex 1 selectively localizes into mitochondria of the HeLa cells. The photophysical properties, cellular uptake, reactive oxygen species (ROS) generation, and cellular apoptosis of complex 1 have also been studied.Overall, the new Fe(II) complex with BODIPY moiety is significantly photocytotoxic in HeLa cells when irradiated with visible light of 500 nm giving as mitochondria targeting. Therefore, we present cyclometalated Fe(II) pincer complex induced mitochondria‐targeted PDT involving the BODIPY moiety that develops persuasively designed photoactivatable Fe(II) complexes.