Details of a Researcher

Thermo-responsive targeting of polymeric micelles by controlling the cellular uptake based on the change of their surface arginine density.

Yamada S., Sasaki E., Ohno H., Nagase K., Hanaoka K.

Communications Chemistry 8 ( 1 ) 325 2025.11

Research paper (scientific journal), Lead author, Accepted, ISSN 23993669

　View Summary

Stimulus-responsive nanocarriers are good candidates for targeted drug delivery. Herein, inspired by the existence of a clear threshold number of arginine residues in oligoarginines for cell-penetrating peptide (CPP) activity, we developed a strategy to control the CPP activity by changing the local arginine density for thermo-responsive targeting. We constructed polymeric micelles whose shell consists of a thermo-responsive polymer based on N-isopropylacrylamide, with a low density of arginine moieties (named Arg-TRM). At physiological temperature (37 °C), internalization of Arg-TRM into cells was small and comparable to that of micelle without arginine. In contrast, upon heating at 42 °C, the arginine density on the micellar surface was increased by thermo-responsive shrinkage of the shell, thereby switching on the CPP activity and enabling efficient cellular uptake. The response of Arg-TRM at 42 °C occurred within a few minutes and the intracellular uptake was rapidly enhanced from 5 min after the heating. This response was transient, thus enabling reversible control of the enhancement by heating. As proof-of-concept, we show that intravenously administered Arg-TRM was effectively accumulated in one ear of a normal mouse by local heating. These results indicate that Arg-TRM is a promising drug carrier for on-demand targeted drug delivery in response to mild external heating.
- Access to Document (DOI)
Heat-guided drug delivery via thermally induced crosslinking of polymeric micelles

Yamada S., Sasaki E., Ohno H., Hanaoka K.

Communications Chemistry 7 ( 1 ) 287 2024.12

Research paper (scientific journal), Lead author, Accepted, ISSN 23993669

　View Summary

Targeted drug delivery in response to external stimuli is therapeutically desirable, but long-term drug retention at the target site after stimulation is turned off remains a challenge. Herein, we present a targeted-delivery strategy via irreversible aggregation of drug carriers in response to mild external heating. We constructed two types of polymeric micelles, DBCO-TRM and Az-TRM, having a thermo-responsive polymer shell based on N-isopropylacrylamide (NIPAAm) and incorporating alkyne and azide moieties, respectively. Upon heating at 42 °C, the micelles aggregated through hydrophobic interaction between their dehydrated shells. Further, the azide moieties of Az-TRM become exposed on the surface due to the thermally shrinkage of the shells, thereby enabling crosslinking between the two types of micelles via azide-alkyne click chemistry to form irreversible aggregates. These aggregates were efficiently accumulated at tumor sites in mice by local heating after intravenous administration of a mixture of the micelles, and were well retained after cessation of heating due to their increased size. As proof of concept, we show that delivery of doxorubicin in this heat-guided drug delivery system dramatically improved the anti-tumor effect in a mouse model after a single treatment. Our results suggest that this platform could be an efficient tool for on-demand drug delivery.
- Access to Document (DOI)
Visualizing Newly Synthesized Proteins and Their Degradation Dynamics by Using Long-Wavelength-Emitting Fluorescent Dye–DBCO Conjugates

Sumitani S., Sasaki E., Ohno H., Yamada S., Takayama O., Wei F.Y., Kuchitsu Y., Taguchi T., Hanaoka K.

Bioconjugate Chemistry 37 ( 5 ) 922 - 929 2026.05

ISSN 10431802

　View Summary

Understanding the spatiotemporal dynamics of protein synthesis and degradation is important for establishing how cells maintain protein homeostasis. Conventional methods for detecting newly synthesized proteins include metabolic labeling with radioactive [<sup>35</sup>S]methionine (Met) or the incorporation of l-azidohomoalanine (AHA) or l-homopropargylglycine followed by fluorescent labeling via copper(I)-catalyzed click chemistry. However, these methods typically require cell fixation, making them unsuitable for live-cell imaging. Here, we describe a fluorescence imaging technique to monitor newly synthesized proteins in living cells by utilizing a strain-promoted azide-alkyne cycloaddition (SPAAC) reaction, in which l-AHA-containing proteins are labeled with fluorescent dyes conjugated to dibenzocyclooctyne (DBCO). We synthesized orange-emitting tetramethylrhodamine (TAMRA)-DBCO and far-red-emitting silicon rhodamine (SiR)-DBCO. TAMRA-DBCO enabled the visualization of newly synthesized proteins and their time-dependent degradation throughout the entire cell. SiR-DBCO was similarly effective, but was mainly distributed to the cytoplasm. The time-dependent decrease of TAMRA-DBCO fluorescence intensity in living cells was suppressed by lysosomal enzyme inhibitors and a proteasome inhibitor, suggesting that newly synthesized proteins are degraded via both pathways. Moreover, imaging of drug-induced senescent cells with TAMRA-DBCO suggested that senescent cells have a lower protein degradation ability than nonsenescent cells. These methods should be useful for investigating protein homeostasis in living cells.
- Access to Document (DOI)
Enzymes that generate and regulate intracellular persulfides and polysulfides: mechanistic insights and inhibitors

Hirabayashi K., Sasaki E., Ohno H., Takayama O., Yamada S., Hanaoka K.

Frontiers in Physiology 17 1764165 2026.02

Accepted

　View Summary

Reactive sulfur species (RSS), which include various persulfides and polysulfides, are generated by multiple enzymes in vivo and play critical roles in mammalian physiological processes such as redox signaling, metabolic regulation, radical scavenging and anti-inflammatory responses. Cystathionine β-synthase (CBS), cystathionine γ-lyase (CSE) and 3-mercaptopyruvate sulfurtransferase (3MST) are well known to mediate endogenous production of hydrogen sulfide (H<inf>2</inf>S), and, together with the mitochondrial isoform of cysteinyl-tRNA synthetase (CARS2), are proposed to be major sources of intracellular persulfides and polysulfides. In mitochondria, enzymes involved in the sulfide oxidation pathway, including sulfide:quinone oxidoreductase (SQOR), persulfide dioxygenase (ETHE1) and thiosulfate sulfurtransferase (TST), also contribute to maintaining and regulating intracellular persulfide levels. Selective inhibitors targeting these enzymes are expected to be powerful tools for elucidating the functions of RSS, as well as having therapeutic potential. In this review, we present a comprehensive overview of these enzymes, focusing on their reaction mechanisms and inhibitors.
- Access to Document (DOI)
Recent advances in near-infrared dye conjugates for near-infrared photoimmunotherapy (NIR-PIT): enhancing therapeutic efficacy and immune mechanisms

Fuse Y., Sasaki E., Takayama O., Yamada S., Hanaoka K.

Rsc Chemical Biology 2026

　View Summary

Near-infrared photoimmunotherapy (NIR-PIT) is an innovative cancer treatment modality that was approved in Japan in 2020 for the treatment of unresectable locally advanced or locally recurrent head and neck cancer. This therapy uses an antibody-dye conjugate (Ab-IR700), which consists of a monoclonal antibody targeting a specific cell-surface antigen and a phthalocyanine-based near-infrared dye, IR700, that functions as a photosensitizer. After selective accumulation in tumor tissue, Ab-IR700 is irradiated with 690 nm NIR light, which initiates a photochemical reaction that selectively damages the cell membrane of target cells, thereby inducing immunogenic cell death. Its high tumor selectivity and therapeutic efficacy establish NIR-PIT as a promising next-generation cancer therapy. However, its further application to deep-seated solid tumors remains challenging, and will require IR700 analogs and novel dye scaffolds that can be activated by longer-wavelength light to achieve greater tissue penetration and that offer greater photochemical activation efficiency. This review covers the activation mechanism of IR700, the mechanisms of cytotoxicity of NIR-PIT, emerging applications of NIR-PIT in oncology and infectious diseases, the range of dye delivery vehicles, and the development of new dyes for NIR-PIT.
- Access to Document (DOI)

display all >>

Page Top ▲

Reviews, Commentaries, etc. 【 Display / hide 】

Enzymes that generate and regulate intracellular persulfides and polysulfides: mechanistic insights and inhibitors

Hirabayashi K., Sasaki E., Ohno H., Takayama O., Yamada S., Hanaoka K.

Frontiers in Physiology 17 1764165 2026.02

Article, review, commentary, editorial, etc. (scientific journal)

　View Summary

Reactive sulfur species (RSS), which include various persulfides and polysulfides, are generated by multiple enzymes in vivo and play critical roles in mammalian physiological processes such as redox signaling, metabolic regulation, radical scavenging and anti-inflammatory responses. Cystathionine β-synthase (CBS), cystathionine γ-lyase (CSE) and 3-mercaptopyruvate sulfurtransferase (3MST) are well known to mediate endogenous production of hydrogen sulfide (H2S), and, together with the mitochondrial isoform of cysteinyl-tRNA synthetase (CARS2), are proposed to be major sources of intracellular persulfides and polysulfides. In mitochondria, enzymes involved in the sulfide oxidation pathway, including sulfide:quinone oxidoreductase (SQOR), persulfide dioxygenase (ETHE1) and thiosulfate sulfurtransferase (TST), also contribute to maintaining and regulating intracellular persulfide levels. Selective inhibitors targeting these enzymes are expected to be powerful tools for elucidating the functions of RSS, as well as having therapeutic potential. In this review, we present a comprehensive overview of these enzymes, focusing on their reaction mechanisms and inhibitors.
- Access to Document (DOI)
Recent advances in fluorogenic probes based on twisted intramolecular charge transfer (TICT) for live-cell imaging

Ohno H., Sumitani S., Sasaki E., Yamada S., Hanaoka K.

Chemical Communications 61 ( 69 ) 12871 - 12884 2025.08

Article, review, commentary, editorial, etc. (scientific journal), ISSN 13597345

　View Summary

Fluorescence imaging is a powerful technique for visualizing biological events in living samples, and new fluorescence-control mechanisms are still needed to extend the scope of biomolecule-targeting fluorogenic probes. Twisted intramolecular charge transfer (TICT) is a unique fluorescence quenching mechanism that depends upon a twisted conformation to promote intramolecular charge separation. Probes utilizing TICT can detect biological molecules/phenomena, such as viscosity, polarity and extended protein structures, that cannot readily be accessed by probes employing other fluorescence-control mechanisms, such as photoinduced electron transfer or spirocyclization. In this review, we summarize recent work on molecular design strategies for TICT-based fluorogenic probes, focusing on structural-modification approaches to control the ease of TICT state formation.
- Access to Document (DOI)
アルギニン密度の温度スイッチングによる細胞内取込制御

山田創太，花岡健二郎

ケミカルバイオロジー 18 ( 1 ) 10 - 12 2025.06

Article, review, commentary, editorial, etc. (scientific journal), Lead author
熱に応答した架橋形成によるナノ粒子の標的選択的送達

山田創太，花岡健二郎

JSMI Report 18 ( 1 ) 31 - 35 2025.02

Article, review, commentary, editorial, etc. (scientific journal), Lead author
Recent advances in Si-rhodamine-based fluorescent probes for live-cell imaging

Ohno H., Sasaki E., Yamada S., Hanaoka K.

Organic and Biomolecular Chemistry 22 ( 16 ) 3099 - 3108 2024.03

Article, review, commentary, editorial, etc. (scientific journal), ISSN 14770520

　View Summary

Fluorescence imaging is a powerful technique for visualizing biological events in living samples with high temporal and spatial resolution. Fluorescent probes emitting far-red to near infrared (NIR) fluorescence are particularly advantageous for in vivo imaging due to their high tissue permeability and low autofluorescence, as well as their suitability for multicolor imaging. Among the far-red to NIR fluorophores, Si-rhodamine is one of the most practical fluorophores for the development of tailor-made NIR fluorescent probes because of the relative ease of synthesis of various derivatives, the unique intramolecular spirocyclization behavior, and the relatively high water solubility and high photostability of the probes. This review summarizes these features of Si-rhodamines and presents recent advances in the synthesis and applications of far-red to NIR fluorescent probes based on Si-rhodamines, focusing on live-cell imaging applications such as fluorogenic probes, super-resolution imaging and dye-protein hybrid-based indicators.
- Access to Document (DOI)