Sulfatases are critical enzymes regulating the dynamic sulfation states of biomolecules and have profound implications in human physiology and pathology. Abnormal sulfatase activity is strongly connected to various diseases such as hormone-dependent cancers, infectious diseases like tuberculosis, lysosomal storage disorders, and bacterial virulence, making these enzymes emerge as crucial biomarkers and therapeutic targets. This review highlights the advances in small-molecule probes for selectively detecting sulfatase activity through various imaging modalities, such as fluorescence, photoacoustic, in-gel fluorogenic assays, and bioluminescence imaging. Additionally, the enzyme-activatable probes that exploit mechanisms via self-immolative linkers, intramolecular charge transfer (ICT), photoinduced electron transfer (PET), ratiometric sensing, followed by aggregation-induced emission (AIE), to achieve high specificity and sensitivity. Along similar lines, we explored substrate-based and affinity-based designs, covering turn-on, ratiometric, and dual-modal near-infrared fluorescence, chemiluminescence, and photoacoustic (NIRF/PA) probes that enable real-time, noninvasive, and deep-tissue imaging. A special focus is given to activity-based sensing of steroid sulfatases (STS) in hormone-related cancers and on profiling bacterial sulfatases for rapid mycobacterial strain differentiation. Furthermore, emerging strategies using nanoprobes and AIEgens demonstrate potential for image-guided surgery, inhalable diagnostics, and high-throughput screening of enzyme inhibitors. Together, these innovations establish a strong framework for sulfatase activity profiling and offer powerful capabilities for disease diagnostics, inhibitor screening, and therapeutic monitoring by enabling spatiotemporal visualization of sulfatase activity both in vitro and in vivo. This review may inspire the development of novel activatable sensors suited for practical biomedical applications by consolidating recent advances in sulfatase-targeted probe design and imaging strategies.

Photodynamic therapy (PDT) offers superior spatiotemporal control compared with conventional cytotoxic cancer therapies through localized light activation. However, most photosensitizers rely on passive tumor accumulation and lack intrinsic mechanisms for active cancer cell targeting. Here, we report Ir_Erl, the first cyclometalated iridium (III) photosensitizer covalently conjugated to the EGFR inhibitor erlotinib through C10 alkyl linker. Ir_Erl retains the bright phosphorescence of the [Ir(pq)2(bpy)] core, enabling label-free imaging and intracellular tracking, while efficiently generating singlet oxygen through a strictly light-dependent type II reactive oxygen species (ROS) generation pathway. Ir_Erl preferentially accumulates in cancer cells relative to non-cancerous controls, and its uptake is markedly reduced by EGFR pre-blockage with gefitinib, indicating an EGFR-dependent accumulation mechanism. Upon white-light irradiation, Ir_Erl exerted potent photocytotoxicity (IC50 13 µM) while maintaining minimal dark toxicity (> 100 µM). In contrast, erlotinib alone displays no detectable phototoxicity under identical conditions. Collectively, these results demonstrate that Ir_Erl integrates receptor-directed delivery with efficient light-triggered ROS generation, establishing a modular strategy for the development of tumor-selective, light-activatable metal-complex photosensitizer. First iridium (III) photosensitizer conjugated to erlotinib for EGFR-targeted PDT. EGFR dependent cellular uptake was confirmed by competition with gefitinib. Erlotinib-iridium metal complex conjugate shows outstanding light-gated cytotoxicity with negligible dark toxicity.

BACKGROUND:Influenza infections are significantly affected by the genetics of the viruses and the cells they infect. Our previous studies showed that various influenza A subtypes uniquely infect different cell lines, offering insights into viral infection mechanisms. Meanwhile, Korean red ginseng extract (RGE) is known for its anti-influenza properties, attributed to its rich composition of saponin and non-saponin components. PURPOSE:This study evaluates the antiviral effects of RGE and its non-saponin (GNSF) and saponin (GSF) fractions against H1N1 and H9N2 influenza A subtypes in diverse cell lines. STUDY DESIGN:Using various cell types and specialized assays, we explored the effect of pretreatment and continuous treatment with RGE and its fractions on viral infectivity and subsequent cellular responses. METHODS:We treated several cell lines with varying concentrations of RGE, GNSF, and GSF and measured the cytotoxic effect, viral infectivity, oxidative stress levels, immune responses, autophagy activity, and changes in cellular structure. RESULTS:Our findings demonstrate that RGE and its fractions significantly reduced H9N2 infection levels across multiple cell lines under pretreatment and continuous treatment conditions. However, continuous treatment elicited variable responses to H1N1, with increased infection levels in certain cell lines. Additionally, it elevated the production of reactive oxygen species and altered inflammatory responses, especially in A549 and NCI-H292 cells. GSF also modulated autophagy activity and MUC1 expression in response to H1N1. CONCLUSION:These findings highlight the potential of ginseng components as targeted influenza treatment, with cell line-specific responses that could guide treatment approaches.

The aggregation state of chemosensors significantly influences their photophysical properties, making it critical for their applications in sensing and imaging. In this study, we systematically investigated the disaggregation properties of 11 representative fluorophores, including widely used dyes such as BODIPY, coumarin, fluorescein, rhodamine B, and Nile Red, as well as AIEgens like TPE and TPA. Through a quantitative approach, we determined the disaggregation concentration (DC50) in various aqueous-organic co-solvent mixtures and correlated these values with solvent polarity parameters, such as dielectric constant and ET(30). Our analysis uncovered distinct DC50 patterns that act as unique fingerprints for each compound, reflecting their structural characteristics. Notably, structurally similar compounds, such as BODIPY derivatives and dansyl-based chemosensors, exhibited comparable trends. This study underscores the utility of solvent polarity versus DC50 scatter plots for evaluating aggregation-dependent chemosensors and offers valuable insights into the molecular determinants of self-aggregation, enabling more rational chemosensor design.

Cellular redox homeostasis is tightly regulated by the oxidation-reduction reactions of nicotinamide metabolites, including NAD(H) and NADP(H), which serve as essential cofactors in enzymatic processes related to energy metabolism. Monitoring intracellular NADH levels is therefore of significant interest. Most chemosensor designs to date rely on fluorescence turn-on mechanisms triggered by NADH oxidation, but these reaction-based sensors are inherently limited by NADH concentration and reaction kinetics. While NADH exhibits intrinsic fluorescence, its low quantum yield has led to the development of redox-sensitive substrates that emit fluorescence upon NADH oxidation. Here, we report an alternative fluorescence enhancement strategy based on an NADH-binding RNA aptamer. The interaction between NADH and a 49-base-pair RNA aptamer induces a 1.4-fold increase in fluorescence emission in vitro and an 1.8-fold increase in live-cell imaging. This fluorescence enhancement arises from aptamer-induced structural rigidity, analogous to the mechanism by which 4-(p-hydroxybenzylidene)-5-imidazolidinone (HBI) enhances fluorescence in green fluorescent protein. Using our aptamer-based assay, we established a live-cell fluorescence emission assay for real-time monitoring of cellular NADH dynamics.

Cholesterol sulfate (CS), one of the most abundant cholesterol derivatives, recently emerged as a key regulatory molecule in several physiological processes. Here, we demonstrate multiple mechanisms by which CS reduces intracellular cholesterol levels. CS promotes the proteasomal degradation of 3-hydroxy-3-methylglutaryl-CoA reductase reductase by enhancing insulin-induced gene-mediated ubiquitination, thereby inhibiting cholesterol synthesis. In addition, CS blocks low-density lipoprotein receptor endocytosis, reducing low-density lipoprotein cholesterol uptake. CS further suppresses the proteolytic activation of sterol regulatory element-binding protein 2, a master transcription factor governing cholesterol synthesis and uptake. Using in vitro and in vivo models, we show that CS lowers cholesterol by targeting both the cholesterol synthesis and uptake pathways, while also modulating an important feedback loop via sterol regulatory element-binding protein 2. These findings highlight the potential of CS as a modulator of cholesterol metabolism, offering new therapeutic insights into cholesterol-related disorders.

Traditional mass spectrometry (MS)-based proteomics aims to detect and measure protein expression on a global scale and elucidate the link between protein function and phenotypic characteristics. Although advances in MS technology have significantly broadened the scope of detectable proteomes, these methodologies primarily provide data on protein abundance and offer limited insights into their functional activities. Phenotypic traits emerge from the interplay between protein abundance and functional activity, making the accurate measurement of activity a critical but challenging task, owing to the complexity of biological systems. Furthermore, the biological function of a protein is strongly linked to its interaction with other molecules within the cellular environment. Chemical proteomics offers a complementary approach that uses a toolkit developed in chemical biology to map the molecular interactome and provide initial insights into the activities of specific target proteins. However, the value of these techniques lies not in isolation, but as part of a broader experimental workflow that includes follow-up biological investigations to validate the findings and elucidate their functional relevance. This tutorial review highlights the design principles of chemical tools and examines their applications in two key areas: (i) functional activity profiling of biomolecules and (ii) molecular proximity profiling for interactome characterization. We also discuss the importance of the experimental context in shaping data interpretation and ensuring the practical adoption of these methods by biologists. Although chemical proteomics is not a standalone solution, it represents a promising step toward next-generation omics technologies and advances our understanding of biological functions at the molecular level.

Proteolysis-targeting chimeras (PROTACs) degrade target proteins through the ubiquitin-proteasome system. To date, PROTACs are primarily used to treat various diseases; however, they have not been applied in regenerative therapy. Herein, this work introduces MDM2-targeting PROTACs customized for application in bone regeneration. An MDM2-PROTAC library is constructed by combining Nutlin-3 and CRBN ligands with various linker designs. Through a multistep validation process, this work develops MDM2-PROTACs (CL144 and CL174) that presented potent degradation efficiency and a robust inductive effect on the biomineralization. Next, this work performs whole-transcriptome analysis to dissect the biological effects of the CL144, and reveals the upregulation of osteogenic marker genes. Furthermore, CL144 effectively induced bone regeneration in bone graft and ovariectomy (OVX) models after local and systemic administration, respectively. In the OVX model, the combination treatment with CL144 and alendronate induced a synergistic effect. Overall, this study demonstrates the promising role of MDM2-PROTAC in promoting bone regeneration, marking the first step toward expanding the application of the PROTAC technology.

ABSTRACT Prostate cancer is an epithelial malignancy with a high incidence among elderly men. Photochemistry‐based dye photodrugs (known as photosensitizers) offer a promising clinical approach for treating tumors. These agents work by inducing immunogenic cell death (ICD), which activates antitumor immune response. This approach is favored owing to its minimal invasiveness, low toxicity, and high efficiency. However, the immunosuppressive microenvironment of characteristics of “cold” tumors significantly restricts the clinical efficacy of photodrugs. Developing an advanced nanocarrier system to deliver photodrugs and immune agonists for efficient drug delivery to tumor lesion sites and to reshape the immunosuppressive microenvironment is crucial in clinical practice. Therefore, in this study, we designed an integrin‐targeted, activatable nano photodrug co‐assembly with an immune agonist (RPST@IMQ) for enhancing photoimmunotherapy in prostate cancer via the reprogramming of tumor‐associated macrophages. The active‐targeted nanosystem enhanced the dosage of photodrug at the lesion site through systemic administration. High doses of glutathione at the tumor site cleaved the disulfide bonds of RPST@IMQ, releasing the photodrug and the immune agonist imiquimod (IMQ). Under photoirradiation, the photodrug generated significant doses of singlet oxygen to eliminate tumor cells, thereby inducing ICD to activate antitumor immune responses. Simultaneously, the released IMQ reprograms immunosuppressive M2‐type tumor‐associated macrophages (TAMs) in the tumor microenvironment into M1‐type TAMs with tumor‐killing capabilities, thereby converting “cold” tumors into “hot” tumors. This conversion enhances the therapeutic efficacy against primary and distant tumors in vivo. This study offers new insights into the development of innovative, smart, activatable nano photodrugs to enhance anticancer therapeutic outcomes.

[This corrects the article DOI: 10.1021/acscentsci.4c01822.].

Photodynamic therapy (PDT) offers minimally invasive and repeatable cancer treatment options. Despite advancements in photosensitizer (PS) design, the optical control of PS activation remains unexplored. Here, we present the first photoswitchable PS based on a BOAHY-BODIPY dyad system. Inspired by BODIPY multimer structures and BOAHY's photoisomerization properties, we designed mono-(4 series) and bis-BOAHY-BODIPY (5 series) conjugates. These dyads primarily generate reactive oxygen species via a type-I process under white light. Notably, the 4 series compounds demonstrated effective photocytotoxicity and photoswitching properties in vitro. Building on these, we iodinated the monoconjugates to develop the highly efficient photoswitching PS, 6b, which exhibited enhanced intersystem crossing and type-II reactive oxygen species generation due to a reduced singlet-triplet energy gap. As the first demonstration of photoswitchable PDT agents, this strategy introduces a new approach with significant potential for selective cancer treatment and clinical applications.

Intestinal epithelial cells (IECs) are replenished by intestinal stem cells (ISCs) residing in the intestinal crypts and isthmus. Under homeostatic conditions, Lgr5+ crypt base columnar cells (CBCs) are considered the primary ISCs[1][1],[2][2]. The supply of IECs, however, remains continuous even in the absence of Lgr5+ CBCs[3][3], implying the existence of alternative ISC populations. Over the past decade, two contrasting models have emerged to explain the identities of these compensatory cells. The distinct cell population model proposes that slow-cycling, quiescent cells near the +4 position, act as reserve stem cells (rISCs)[3][3]-[10][4]. The cellular plasticity model suggests a broad spectrum of IECs can dedifferentiate into intermediate fetal-like states, thereby acquiring stemness[11][5],[12][6]. It is unclear which of these models is correct because the signaling mechanisms governing crypt regenerative responses remain poorly understood. Here, we demonstrate that Hedgehog (Hh) signaling orchestrates intestinal regeneration following Lgr5+ CBC loss by uniquely driving the emergence of de novo Lgr5+ CBCs from non-Lgr5+ CBCs clones. This regenerative mechanism is fundamentally distinct from both classical self-renewal of existing Lgr5+ CBCs and fetal-like dedifferentiation in intestinal villi. By accelerating the cell cycle and driving clonal competition among stem cell pools, Hh signaling leaves rare dormant stem cells in the crypt and isthmus regions. These rare cells act as damage-resistant “seeds” that can promote damage recovery, thereby conferring damage resistance on the tissue as a whole. Collectively, our findings identify novel molecular and cellular mechanisms underlying ISC regeneration and suggest potential therapeutic strategies for various intestinal epithelial diseases. ### Competing Interest Statement The authors have declared no competing interest. National Research Foundation of Korea, RS-2023-00208193, NRF-2022M3A9F3094559, RS-2023-00212238 IITP (Institute of Information & Communications Technology Planning & Evaluation)-ICAN (ICT Challenge and Advanced Network of HRD) grant, IITP-2024-RS-2024-00438263 [1]: #ref-1 [2]: #ref-2 [3]: #ref-3 [4]: #ref-10 [5]: #ref-11 [6]: #ref-12

Controlling the succession of chemical processes with high specificity in complex systems is advantageous for widespread applications, from biomedical research to drug manufacturing. Despite synthetic advances in bioorthogonal and photochemical methodologies, there is a need for generic chemical approaches that can universally modulate photodynamic reactivity in organic photosensitizers. Herein we present a strategy to fine-tune the production of singlet oxygen in multiple photosensitive scaffolds under the activation of bioresponsive and bioorthogonal stimuli. We demonstrate that the photocatalytic activity of nitrobenzoselenadiazoles can be fully blocked by site-selective incorporation of electron-withdrawing carbamate moieties and restored on demand upon uncaging with a wide range of molecular triggers, including abiotic transition-metal catalysts. We also prove that this strategy can be expanded to most photosensitizers, including diverse structures and spectral properties. Finally, we show that such advanced control of singlet oxygen generation can be broadly applied to the photodynamic ablation of human cells as well as to regulate the release of singlet oxygen in the semi-synthesis of natural product drugs.

Rational and effective design of a universal near-infrared (NIR) light-absorbed platform employed to prepare diverse activatable NIR fluorogenic probes for in vivo imaging and the imaging-guided tumor resection remains less exploited but highly meaningful. Herein, mandelic acid with a core structure of 4-hydroxylbenzyl alcohol to link recognition unit, a fluorophore and a quencher was employed to prepare activatable probes. We exemplified ester as carboxylesterase (CE)-recognized unit, ferrocene as quencher and phenothiazinium as NIR fluorophore to afford fluorogenic probes termed NBS-Fe-CE and NBS-C-Fe-CE. These probes enabled the conversion toward CE with significant fluorescence increases and successfully discriminate CE activity in cells. NIR light enhances the tumor penetration and enable imaging-guided orthotopic tumor resection. This specific case demonstrated that this platform can be effectively used to construct diverse NIR probes for imaging analytes in biological systems.

A ferritinophagy-assisted ferroptosis strategy addresses the limitations in iron-based anti-cancer therapy by inhibiting GSH, inducing ferritin degradation, overloading labile iron ions, and enhancing ferroptosis.

Taste buds, the neuroepithelial organs responsible for the detection of gustatory stimuli in the oral cavity, arise from stem/progenitor cells among nearby basal keratinocytes. Using genetic lineage tracing, Lgr5 and Lgr6 were suggested as the specific markers for the stem/progenitor cells of taste buds, but recent evidence implied that taste buds may arise even in the absence of these markers. Thus, we wanted to verify the genetic lineage tracing of lingual Lgr5- and Lgr6-expressing cells. Unexpectedly, we found that antibody staining revealed more diverse Lgr5-expressing cells inside and outside the taste buds of circumvallate papillae than was previously suggested. We also found that, while tamoxifen-induced genetic recombination occurred only in cells expressing the Lgr5 reporter GFP, we did not see any increase in the number of recombined daughter cells induced by consecutive injections of tamoxifen. Similarly, we found that cells expressing Lgr6, another stem/progenitor cell marker candidate and an analog of Lgr5, also do not generate recombined clones. In contrast, Lgr5-expressing cells in fungiform papillae can transform into Lgr5-negative progeny. Together, our data indicate that lingual Lgr5- and Lgr6-expressing cells exhibit diversity in their capacity to transform into Lgr5- and Lgr6-negative cells, depending on their location. Our results complement previous findings that did not distinguish this diversity.

In this concept, we present a comprehensive study on the development and application of COX-2-specific fluorescent probes for cancer imaging and diagnosis. To target cancer cells and measuring cancer-related activities in specific organelles quickly and accurately are crucial factors for early diagnosis and research on cancer pathology and treatment. This concept explores a variety of probes based on indomethacin (IMC), celecoxib, rofecoxib as well as CoxFluor and each one demonstrates unique mechanisms and high selectivity towards COX-2 enzymes. These probes were designed to enhance fluorescence upon binding to COX-2 which enable precise visualization of tumor and inflamed tissues. The research emphasizes the importance of COX-2 as a biomarker in cancer diagnostics, particularly in identifying cancer stem cells and inflamed tissues. This concept highlights the potentiality of these probes in non-invasive imaging techniques which offering significant advancements in cancer diagnosis and monitoring. The in vivo and in vitro experiments, including applications in mouse models and human tissue samples, confirm the efficacy of these probes in providing detailed imaging for clinical and research applications.

The differential sensing approach uses fingerprint patterning to distinguish uncharacterized biological samples. Inspired by natural sensory systems, an array of cross-reactive sensors generates unique response fingerprint depending on the samples. Until today, this array system has been developed using various materials, including the library of surface-charged nanoparticles and chemosensors. Many differential array systems have demonstrated accurate identification of bacterial species, viral subtypes, and cancer cells, as well as distinguishing disease states in blood or urine. This capability is particularly important for distinguishing between normal and abnormal states when specific marker molecules have not yet been identified, providing a powerful diagnostic tool. In this concept, we summarized representative outcomes of differential sensing applications for biological sample discrimination.

Target protein degradation (TPD) is a promising strategy for catalytic downregulation of target proteins through various cellular proteolytic pathways. Despite numerous reports on novel TPD mechanisms, the discovery of target-specific ligands remains a major challenge. Unlike small-molecule ligands, aptamers offer significant advantages, owing to their SELEX-based systematic screening method. To fully utilize aptamers for TPD, we designed an aptamer and N-degron ensemble system (AptaGron) that circumvents the need for synthetic conjugations between aptamers and proteolysis-recruiting units. In our AptaGron system, a peptide nucleic acid containing an N-degron peptide and a sequence complementary to the aptamer was designed. Using this system, we successfully degraded three target proteins, tau, nucleolin, and eukaryotic initiation factor 4E (eIF4E), which lack specific small-molecule ligands. Our results highlight the potential of the AptaGron approach as a robust platform for targeted protein degradation.