About

John P. Toscano is a professor of chemistry in the School of Arts and Sciences at Johns Hopkins University. His main research interests are focused on the study of the fundamental chemistry and biology of small molecule bioactive signaling agents such as nitric oxide (NO), nitroxyl (HNO), and hydrogen sulfide (H2S). His research explores the distinct biological activities of these molecules, their reactivity, and their potential therapeutic applications, particularly in cardioprotection and heart failure treatment. Toscano's work involves developing methods to generate and study these reactive species, understanding their biochemical effects, and investigating their roles in physiological processes. He received his baccalaureate degree in chemistry from Princeton University and his doctoral degree in organic chemistry from Yale University. After completing a National Institutes of Health postdoctoral fellowship at Ohio State University, he joined Johns Hopkins University in 1995 as an assistant professor, later becoming a full professor in 2003. He has served as vice-chair and chair of the Department of Chemistry, where he has contributed to faculty recruitment, research infrastructure, and interdisciplinary initiatives. His research group has made significant contributions to understanding the chemistry and biology of hydropersulfides, HNO, and related species, including their formation, reactivity, and biological effects, with implications for cellular protection and redox signaling.

Research topics

• Chemistry
• Biochemistry
• Organic chemistry
• Medicine
• Combinatorial chemistry
• Biophysics
• Biology
• Pharmacology
• Internal medicine
• Cancer research
• Stereochemistry

Selected publications

• Hydropersulfides promote angiogenesis and preserve vascular function

  Redox Biology · 2026-05-01

  articleOpen access

  Hydropersulfides (RSSH) are increasingly recognized as key mediators in redox signalling in mammalian cells, although their physiological functions, especially in angiogenesis, remain unknown. Direct mechanistic investigation of RSSH has been challenging due to their instability and high reactivity. To address this, donors that release RSSH in a controlled manner have been developed, enabling investigation of its biological roles. Herein, we employed thiol- and enzyme-activated RSSH donors along with mutant mouse models deficient in the Cys-SSH-producing enzyme cysteinyl-tRNA synthetase/cysteine persulfide synthase (CARS2/CPERS), to examine their role in vascular angiogenesis. We demonstrate that RSSH promotes angiogenesis via the CARS2/CPERS signalling axis and activation of the Akt-eNOS and NO-cGMP pathway. We also demonstrate that nitric oxide (NO) signalling in resistance vessels requires an intact CARS2/CPER2 pathway. These findings suggest that RSSH plays an important role in regulating angiogenesis and vascular tone.

  PublisherDOI

• Esterase‐Responsive Mitochondria‐Targeted Hydropersulfide Donors Mitigate Doxorubicin Cardiotoxicity While Preserving Anticancer Activity

  Angewandte Chemie · 2025-12-12

  articleSenior author

  Abstract Therapeutic agents that protect the heart from doxorubicin (DOX) toxicity without reducing its anticancer efficacy remain a critical unmet need. We report esterase‐activated hydropersulfide (RSSH) donors, alkyl sulfenyl thiocarbonate ( AST‐2 ), and acetoxy perthiocarbamate ( APT‐1 ), together with their mitochondria‐targeted analogs, AST‐2‐TPP and APT‐1‐TPP , which bear a triphenylphosphonium (TPP⁺) moiety. These compounds release RSSH upon esterase activation with tunable half‐lives (20–125 min in PBS, pH 7.4). LC–MS/MS analysis revealed that APT‐1 elevates hydropersulfide levels in the cytosol of H9c2 cardiomyoblasts, whereas its mitochondrial analog, APT‐1‐TPP , increases levels in mitochondria. All donors attenuated DOX‐induced toxicity in H9c2 cells, but in cancer cell lines (HepG2, MDA‐MB‐468, MCF‐7), APT‐1 did not blunt DOX cytotoxicity and APT‐1‐TPP synergistically enhanced its activity. Mechanistic studies revealed that both APT‐1 and APT‐1‐TPP rescue DOX‐induced mitochondrial membrane depolarization and ATP depletion in H9c2 cells but not in HepG2 cells. Further characterization indicated that cancer cells exhibit higher basal sulfane sulfur levels and mitochondrial membrane potentials compared to H9c2 cells, suggesting that divergent redox environments may underlie these contrasting effects. Collectively, these findings demonstrate that redox heterogeneity between cardiac and cancer cells can be exploited to develop cardioprotective interventions that preserve or enhance DOX's anticancer efficacy.

  PublisherDOI

• Arylsulfonothioates: Thiol-Activated Donors of Hydropersulfides which are Excreted to Maintain Cellular Redox Homeostasis or Retained to Counter Oxidative Stress

  Journal of the American Chemical Society · 2025-02-18 · 9 citations

  articleSenior authorCorresponding

  Despite their biological significance, the study of hydropersulfides (RSSH) is often limited due to their inherent instability. Here, we introduce arylsulfonothioates as thiol-activated RSSH donors and provide insight into cellular reactive sulfur species homeostasis. These precursors persulfidate physiologically relevant thiols (RSH) to form the corresponding RSSH. Real-time monitoring of hydrogen sulfide (H2S) generation via membrane inlet mass spectrometry (MIMS) was employed to follow RSSH production, revealing that electron-donating aryl substituents marginally slow RSSH release rates, whereas electron-withdrawing substituents slightly accelerate release. Furthermore, arylsulfonothioates with strong electron-withdrawing substituents offer superior protection against doxorubicin (DOX)-induced cardiotoxicity. Experiments using H9c2 cardiomyocytes affirmed the cell-permeability of arylsulfonothioates and their ability to increase intracellular RSSH levels and protein persulfidation levels. Notably, we observe the excretion of RSSH into the extracellular medium. Further investigations revealed the involvement of the cystine/glutamate antiporter SLC7A11, as cotreatment with its inhibitor, sulfasalazine, significantly reduce extracellular RSSH release. H9c2 cells exhibit tolerance to arylsulfonothioate 1g with an electron-withdrawing 4-cyano group at 1 mM; however, inhibition of the cystine antiporter results in a minor decrease in cell viability. Under oxidative stress conditions induced by DOX or hydrogen peroxide (H2O2), cotreatment with 1g diminishes the excretion of RSSH and confers cytoprotection against DOX or H2O2-mediated toxicity. Our findings show adaptive cellular responses to RSSH levels, demonstrating excretion under elevated conditions to maintain redox homeostasis and intracellular retention as a protective response during oxidative stress.

  PublisherDOI

• Development of Azoreductase-Activated Precursors for Efficient Hydropersulfide Release via 1,6-Elimination

  ACS Chemical Biology · 2025-10-25 · 1 citations

  articleSenior authorCorresponding

  Hydropersulfides (RSSH) are increasingly recognized for their potent redox-modulating and cytoprotective properties, yet their therapeutic potential remains underexplored due to their chemical instability. Here, we report the design and optimization of azoreductase (AzoR)-responsive RSSH donors for programmed release. We explore azo-linked precursors that undergo AzoR-mediated reduction to form phenylamino intermediates, which are designed to trigger RSSH release via spontaneous 1,6-elimination. A series of precursors was synthesized to evaluate the structure–activity relationships governing elimination efficiency. Direct attachment of aliphatic or aromatic RSSH moieties at the benzylic position of the azobenzene core (azodisulfides, AzDS-1, AzDS-2, AzDS-3) resulted in stable intermediates that failed to eliminate RSSH under physiological conditions. Even an electron-rich substituent such as a methoxy group on the azobenzene core was not sufficient to drive the elimination. To improve leaving group ability, we synthesized an azoperthiocarbonate donor (AzPTC) that enabled AzoR-triggered RSSH release but also underwent undesired nonenzymatic hydrolysis. Finally, incorporation of a hydrolytically stable perthiocarbamate yielded the precursor azoperthiocarbamate (AzPTB) that released RSSH selectively upon AzoR activation via 1,6-elimination, decarboxylation, and an intramolecular cyclization cascade. AzPTB demonstrates high enzymatic turnover and excellent stability under neutral aqueous conditions. These results provide key structural parameters that govern RSSH release via 1,6-elimination and establish AzPTB as a robust platform for site-selective delivery. This work expands the chemical biology toolkit for probing RSSH signaling and supports future efforts in redox-based therapeutic development.

  PublisherDOI

• Esterase‐Responsive Mitochondria‐Targeted Hydropersulfide Donors Mitigate Doxorubicin Cardiotoxicity While Preserving Anticancer Activity

  Angewandte Chemie International Edition · 2025-12-12 · 1 citations

  articleSenior authorCorresponding

  Therapeutic agents that protect the heart from doxorubicin (DOX) toxicity without reducing its anticancer efficacy remain a critical unmet need. We report esterase-activated hydropersulfide (RSSH) donors, alkyl sulfenyl thiocarbonate (AST-2), and acetoxy perthiocarbamate (APT-1), together with their mitochondria-targeted analogs, AST-2-TPP and APT-1-TPP, which bear a triphenylphosphonium (TPP⁺) moiety. These compounds release RSSH upon esterase activation with tunable half-lives (20-125 min in PBS, pH 7.4). LC-MS/MS analysis revealed that APT-1 elevates hydropersulfide levels in the cytosol of H9c2 cardiomyoblasts, whereas its mitochondrial analog, APT-1-TPP, increases levels in mitochondria. All donors attenuated DOX-induced toxicity in H9c2 cells, but in cancer cell lines (HepG2, MDA-MB-468, MCF-7), APT-1 did not blunt DOX cytotoxicity and APT-1-TPP synergistically enhanced its activity. Mechanistic studies revealed that both APT-1 and APT-1-TPP rescue DOX-induced mitochondrial membrane depolarization and ATP depletion in H9c2 cells but not in HepG2 cells. Further characterization indicated that cancer cells exhibit higher basal sulfane sulfur levels and mitochondrial membrane potentials compared to H9c2 cells, suggesting that divergent redox environments may underlie these contrasting effects. Collectively, these findings demonstrate that redox heterogeneity between cardiac and cancer cells can be exploited to develop cardioprotective interventions that preserve or enhance DOX's anticancer efficacy.

  PublisherDOI

• Hydropersulfides (RSSH) attenuate doxorubicin-induced cardiotoxicity while boosting its anticancer action

  Redox Biology · 2023 · 40 citations

  Senior authorCorresponding
  • Pharmacology
  • Cancer research
  • Chemistry

  S), sulfane sulfur, and reducing equivalents compared to cardiac cells. Thus, RSSH may represent a new promising avenue to fend off DOX-induced cardiotoxicity while boosting its anticancer effects.

  PublisherDOI

• β-Galactosidase-activated nitroxyl (HNO) donors provide insights into redox cross-talk in senescent cells

  Chemical Communications · 2023-01-01 · 1 citations

  article

  S)-dependent manner. Hence, the newly developed tool provides insights into redox cross-talk and establishes the foundation for new interventions that modulate levels of these species to mitigate oxidative stress and inflammation.

  PublisherDOI

• Reactive Sulfur Species in Biology and Medicine

  Antioxidants · 2023-09-13 · 8 citations

  editorialOpen accessSenior authorCorresponding

  S) has emerged as a third small-molecule bioactive signaling agent, along with nitric oxide (NO) and carbon monoxide (CO) [...].

  PublisherOA PDFDOI

• The reaction of hydropersulfides (RSSH) with S-nitrosothiols (RS-NO) and the biological/physiological implications

  Free Radical Biology and Medicine · 2022-07-06 · 7 citations

  articleOpen accessCorresponding

  S-Nitrosothiol (RS-NO) generation/levels have been implicated as being important to numerous physiological and pathophysiological processes. As such, the mechanism(s) of their generation and degradation are important factors in determining their biological activity. Along with the effects on the activity of thiol proteins, RS-NOs have also been reported to be reservoirs or storage forms of nitric oxide (NO). That is, it is hypothesized that NO can be released from RS-NO at opportune times to, for example, regulate vascular tone. However, to date there are few established mechanisms that can account for facile NO release from RS-NO. Recent discovery of the biological formation and prevalence of hydropersulfides (RSSH) and their subsequent reaction with RS-NO species provides a possible route for NO release from RS-NO. Herein, it is found that RSSH is capable of reacting with RS-NO to liberate NO and that the analogous reaction using RSH is not nearly as proficient in generating NO. Moreover, computational results support the prevalence of this reaction over other possible competing processes. Finally, results of biological studies of NO-mediated vasorelaxation are consistent with the idea that RS-NO species can be degraded by RSSH to release NO.

  PublisherDOI

• Photochemical Release of Hydropersulfides

  The Journal of Organic Chemistry · 2022-09-09 · 11 citations

  articleSenior authorCorresponding

  Hydropersulfides (RSSH) have received significant interest in the field of redox biology because of their intriguing biochemical properties. However, because RSSH are inherently unstable, their study is challenging, and as a result, the details of their physiological roles remain ill-defined. Herein, we report strategies to release RSSH utilizing photoremovable protecting groups. RSSH protection with the well-established p-hydroxyphenacyl (pHP) photoprotecting group resulted in inefficient RSSH photorelease along with complex chemistry. Therefore, an alternative precursor was examined in which a self-immolative linker was inserted between the pHP group and RSSH, providing nearly quantitative RSSH release following photolysis at 365 nm. Inspired by these results, we also synthesized an analogous precursor derivatized with 7-diethylaminocoumarin (DEACM), a visible light-cleavable photoprotecting group. Photolysis of this precursor at 420 nm led to efficient RSSH release, and in vitro experiments demonstrated intracellular RSSH delivery in breast cancer MCF-7 cells.

  PublisherDOI

Recent grants

• Generation and Solution of Reactivity of Nitroxyl (HNO)

  NSF · $471k · 2012–2016

• Generation and Solution Reactivity of Nitroxyl (HNO)

  NSF · $531k · 2005–2009

• Generation and Solution Reactivity of Small Molecule Signaling Agents

  NSF · $450k · 2016–2019

• Generation and Solution Reactivity of Small Molecule Sulfur-based Signaling Agents

  NSF · $500k · 2019–2022

• Generation and Solution Reactivity of Nitroxyl (HNO)

  NSF · $446k · 2009–2012

Frequent coauthors

• Nazareno Paolocci

  Johns Hopkins Medicine

  129 shared

• Carlo G. Tocchetti

  Federico II University Hospital

  101 shared

• David A. Kass

  Johns Hopkins University

  99 shared

• Jeffrey P. Froehlich

  Johns Hopkins University

  98 shared

• Brian O’Rourke

  Johns Hopkins Medicine

  84 shared

• Gerald M. Wilson

  University of Maryland, Baltimore

  83 shared

• Sabine Huke

  University of Alabama at Birmingham

  82 shared

• David A. Wink

  National Cancer Institute

  82 shared