New guidelines and definitions for type I photodynamic therapy

Abstract

The advent of photochemical technologies has revolutionized biology and medicine, offering groundbreaking innovations in cancer treatment and beyond. Among these, photodynamic therapy (PDT) has emerged as a promising approach to cancer therapy, leveraging cytotoxic reactive oxygen species (ROS) to eliminate cancer cells. While traditional type II PDT relies on high oxygen levels and consumes substantial amounts of oxygen, type I PDT requires less oxygen and holds great potential in addressing the hypoxic microenvironments characteristic of solid tumors. Over the last six years, our research team has made pioneering contributions to this field, with a particular focus on type I photosensitizers (PSs) and their diverse applications, including oxygen-sparing PDT, mitochondrial respiration inhibitors, modulation of cellular self-protection pathways, targeted cancer cell destruction, regulation of cellular signaling pathways, immune activation via nanomedicines, and intracellular oxygen-independent artificial photoredox catalysis. Notably, in 2018, our research proposed a “partial oxygen-recyclable mechanism” mediated by O₂˙⁻, successfully revealing why the type I mechanism can be used for overcoming PDT hypoxia resistance. This revitalized interest in type I PDT and inspired numerous research groups worldwide to develop a plethora of new O₂˙⁻ photogenerators. However, inconsistencies in mechanistic interpretations, detection methodologies, and application strategies have arisen due to fragmented communication within the field of photoscience and ambiguity in some key definitions. Given our research team's significant contributions and expertise in the type I PDT domain, we believe it is imperative to present a comprehensive review to establish standardized definitions, mechanisms, molecular designs, detection techniques, and clinical applications of type I PDT in cancer diagnosis and treatment. Our goal is to provide a clear and authoritative resource for both specialists and non-specialists, fostering a deeper understanding of type I PDT and inspiring future innovations to advance more effective and clinically relevant therapies for cancer treatment.

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From left to right: Lizhou Yue, Yang Zhou, Xiaoqiang Chen, Xiaojun Peng, Mingle Li, Yingying Zhang, Jianhua Xiong

Mingle Li (PhD, Dalian Univ. of Tech., 2019) is a full professor at Shenzhen University, specializing in type I PDT and O2-independent photocatalytic biomolecular regulation in living settings. Jianhua Xiong (PhD, Wuhan Univ., 2024) is a postdoc under Prof Xiaojun Peng at Shenzhen University, specializing in the design and synthesis of organic small-molecule prodrugs and their antitumor mechanisms. Yingying Zhang (PhD, Nanjing Forestry Univ., 2024) is a postdoc under Prof Xiaojun Peng at Shenzhen University, specializing in photosensitive dyes for type I PDT and photocatalytic therapy. Lizhou Yue (PhD, Guangxi Univ., 2024) is a postdoc under Prof Xiaojun Peng at Shenzhen University, specializing in molecular probes, biomedical imaging, and type I photosensitizers. Yang Zhou (PhD, Kent State Univ., 2017) is a full Professor at Hainan University, specializing in organic functional NO donors for biomedical applications. Xiaoqiang Chen (PhD, Dalian Univ. of Tech., 2007) is a full professor at Shenzhen University, specializing in organic/supramolecular photochemistry, biological sensors, and mRNA delivery materials. Xiaojun Peng (PhD, Dalian Univ. of Tech., 1990) is an Academician of the Chinese Academy of Sciences, specializing in functional dyes for fluorescent bioimaging/labeling and digital printing/recording.

Le Yu

Le Yu (PhD, Korea Univ., 2024) is currently a lecturer at Yunnan University, with research interests in fluorescence probes, photoacoustic imaging, photodynamic therapy, and photothermal therapy.

From left to right: Changyu Yoon, Jong Seung Kim, Yunjie Xu, Yujin Kim

Changyu Yoon (BS, Hallym Univ., 2022) is pursuing his PhD degree under Prof Jong Seung Kim at Korea University, focusing on designing disease-selective small-molecule photosensitizers with enhanced photochemical reactivity for PDT. Yujin Kim (BS, Korea Univ., 2023) is pursuing her PhD degree under Prof Jong Seung Kim at Korea University, focusing on the development of drug delivery systems in cancer therapy. Yunjie Xu (PhD, Jilin Univ., 2018) is currently pursuing postdoctoral training under the supervision of Prof Jong Seung Kim at Korea University, focusing on the antitumor mechanism (e.g., pyroptosis, ferroptosis, and autophagy) of prodrugs and small-molecular photoactive agents. Jong Seung Kim (PhD, Texas Tech Univ., 1993) is an Academician of the Korea Academy of Sciences and a distinguished Professor at Korea University in Seoul, focusing on drug delivery systems, in vivo imaging of pathologies, and phototherapeutic chemistry. He has been a highly cited researcher since 2014 and the author of more than 700 papers with an h-index of 127.

Key learning points

(1) The historical perspective and definitions of type I photodynamic therapy.

(2) The photophysical and photochemical processes of type I photosensitizers.

(3) Detection techniques for type I ROS in chemistry and biology.

(4) A palette of type I photosensitizers in cancer treatment.

(5) Challenges and opportunities in advancing type I PS research.

1. Introduction

Innovations in phototheranostic technologies have profoundly impacted life sciences, enabling precise exploration and manipulation of complex biochemical processes.^1–5 Among these, photodynamic therapy (PDT) has emerged as a groundbreaking approach in tumor treatment, celebrated for its unparalleled temporal and spatial precision, minimal invasiveness, and high therapeutic selectivity.^6,7 PDT relies on three essential components—photosensitizers (PSs), light sources, and sufficient oxygen—for its efficacy.⁸ By leveraging the metabolic activity of tumor cells, PSs preferentially accumulate in tumor tissue, where they can be activated by specific laser irradiation to generate reactive oxygen species (ROS), which can specifically ablate malignant cells and neovascularization with remarkable precision.^9,10 Such a dual targeting mechanism—combining preferential PS accumulation with precise light delivery—establishes PDT as a uniquely selective therapeutic modality with minimized collateral damage.¹

PDT is broadly classified into type I and type II mechanisms based on the mode of ROS generation.^11,12 The well-established type II PDT involves PS excitation upon light irradiation, transitioning to a triplet excited state through intersystem crossing (ISC).¹³ The triplet PS transfers energy to molecular oxygen, generating singlet oxygen (¹O₂), a potent oxidizing agent capable of inducing oxidative damage and cell death.¹⁴ However, the reliance of type II PDT on molecular oxygen presents a critical limitation in hypoxic tumor microenvironments (pO₂ < 5 mmHg),¹⁵ a hallmark of solid tumors that significantly reduces its therapeutic efficacy. Current strategies to overcome hypoxia include hyperbaric O₂ therapy, nanotechnology-based O₂ delivery and in situ O₂ production, but still face several concerns, such as the risk of oxygen poisoning and tumor-specific accumulation and controlled release.^16–19

In contrast, type I PDT has gained considerable attention due to its reduced O₂ dependency and a unique “partial oxygen-recyclable” anti-hypoxia mechanism (elaborated in Section 3.2.1).²⁰ Since our seminal discovery in 2018 of thio-Nile blue (ENBS) as a novel generator of superoxide anions (O₂˙⁻),²¹ interest in type I PDT has surged, leading to substantial research advancements globally. However, the diversity in detection methods and ROS identification mechanisms has introduced challenges, often complicating the accurate understanding and optimization of type I PDT.

Recognizing this need, we leverage our expertise and recent breakthroughs in tumor phototherapy to provide a comprehensive guide addressing these challenges. In this review, we redefine type I PDT, systematically outlining molecular design principles, photochemical mechanisms, and methodologies for ROS detection. Beginning with an in-depth exploration of type I photochemistry, this review consolidates cutting-edge advancements and offers practical insights into molecular PS design for type I PDT. In particular, we introduce, for the first time, fundamental chemical principles regarding “New Guidelines and Definitions for type I PDT”, aiming to inspire the development of more effective, innovative therapeutic strategies.

2 Status quo of type I PDT

Since its identification in 1991, type I PDT has undergone rapid development, transitioning into a new era after more than three decades of progress.²² Initially focused on understanding the electron transfer-based type I photosensitized oxidation and its biological application (i.e., type I PDT photochemical mechanism and therapeutic efficacy), research has now advanced to the systematic design of innovative type I PSs. These include not only organic small molecular dyes^23,24 but also inorganic (hybrid) nanomaterials²⁵ for achieving superb therapeutic outcomes.

The development of type I PDT can be roughly partitioned into two stages: (1) the first stage is from approximately 1991 to 2012. During this period, researchers unveiled the fundamental photochemical mechanism of type I PDT (i.e., electron transfer process)²⁶ and identified some PSs enabling the generation of O₂˙⁻ (but not specific), including organic molecular dyes (e.g., methylene blue and cationic boron-dipyrromethene),²⁷ water-soluble bacteriochlorophyll derivatives (e.g., WTS09 and WTS11)^28,29 and metal oxide nanoparticles (e.g., TiO₂, ZnO, Al₂O₃, CeO₂, and Fe₂O₃ nanoparticles).³⁰ (2) The second stage is in the last decade. To address the limitations of traditional type I PDT including (1) limited tissue penetration depth of excitation light; (2) non-specific O₂˙⁻ generation hampers anti-hypoxia efficiency in O₂-deficient environments (e.g., tumor microenvironment); (3) the lack of targeted specificity of PSs, leading to oxidative damage in normal tissues, researchers have developed and modified numerous type I PSs. For instance, Lin et al. introduced upconversion-based TiO₂ nanomaterials for near-infrared (NIR) light excitation, enhancing tissue penetration depth;³¹ Li and co-workers developed the first exclusive NIR O₂˙⁻ photogenerator (ENBS-B)²¹ and later designed a binary photodynamic O₂-economizer (SORgenTAM)³² to mitigate tumor hypoxia by inhibiting intracellular O₂ consumption, thereby improving therapeutic efficacy; additionally, Li et al. further proposed a structure-inherent targeting (SIT) strategy using Förster resonance energy transfer (FRET) theory to improve the tumor-targeting ability of type I PSs.³³

Despite these advancements, significant challenges remain in fully understanding the relationship between the therapeutic efficacy of type I PDT and its underlying biological mechanism. Recent studies have classified O₂-independent photoredox catalysis within cells as type I PDT.^34,35 However, according to the definition of PDT, light, oxygen, and PSs are the three essential elements required.²⁶ Without oxygen, these reactions should be categorized as photocatalytic therapy rather than PDT, as they merely borrow from the mechanisms of type I reactions without aligning with PDT's strict definition. Originally, type I PDT was believed to primarily induce apoptosis to achieve therapeutic effects. Recent findings, however, suggest that pyroptosis may also serve as a “potential contributor” to cell death,^36,37 although the dominance of apoptosis versus pyroptosis is likely influenced by multiple factors, including PS localization, intracellular ROS flux, light dose, and cellular context (e.g., caspase expression levels).³⁸ This raises critical questions: Are there other undiscovered therapeutic mechanisms underlying type I PDT? How can existing approaches be integrated to develop a “one for all” strategy that addresses type I PDT limitations? These challenges are driving scientists to conduct more comprehensive and in-depth research, paving the way for future advancements in clinical treatment.

3. New definition of type I PDT

In the evolving landscape of PDT, inconsistencies in terminology and definitions regarding type I photosensitized oxidation reactions have hindered progress. Discrepancies in reaction mechanisms, detection techniques, and application fields, often stemming from insufficient communication among professionals in photoscience, have created unnecessary misunderstandings and confusion. To foster a unified and accurate understanding of type I mechanisms, it is essential to bridge these gaps and establish a coherent framework. This section introduces an updated and comprehensive definition of type I PSs, highlighting their mechanisms of action and detection methodologies. By tracing the historical development of type I PDT, we provide a detailed exploration of its photophysical and photochemical processes, such as the photoinduced electron transfer mechanism in organic molecular dyes and the photon-generated electron–hole pair mechanism in inorganic nanomaterials. In particular, to present a holistic perspective of this dynamic field, we summarize the chemistry and biology of type I related ROS, including O₂˙⁻, hydroxyl radicals (˙OH), and hydrogen peroxide (H₂O₂). Drawing upon our research group's unique insights, we delve into the current status and challenges of type I PDT, offering a clear and forward-looking analysis of this transformative therapy. They are detailed in the following sections:

3.1. Photophysical and photochemical processes

Since its initial conceptualization by von Tappeiner and A. Jodlbauer in 1907,^39,40 PDT has undergone extensive development, demonstrating significant potential in the treatment of solid tumors. The pivotal distinction between type I and type II PDT mechanisms was first tentatively proposed by Foote in 1991,²² significantly advancing the field (Fig. 1). Subsequent discoveries have further elucidated type I PDT. For example, in 2002, methylene blue was identified as capable of generating O₂˙⁻ at aqueous micelle interfaces,²⁷ and by 2015, metal oxides were also implicated in type I PDT mechanisms.³¹ Jean Cadet and colleagues made a landmark contribution in 2017 by formally proposing “ten tips for defining type I and type II photosensitized oxidation reactions”, providing a comprehensive framework for distinguishing these two pathways.⁴¹ This work has charted a forward-looking path of guidance for subsequent research into type I PDT properties of both organic molecules and inorganic compounds. Despite these advancements, however, the fundamental question of why type I PDT exhibits a lower oxygen dependence compared to type II PDT remains unresolved. In 2018, our research group made a pivotal discovery with sulfur (S)-substituted Nile blue analogs (ENBS), which uniquely function as selective generators of O₂˙⁻ for PDT.²¹ Within cancer cells, it was revealed that photogenerated O₂˙⁻ could engage in superoxide dismutase (SOD)-mediated disproportionation reactions to form H₂O₂, then participate in Haber–Weiss or Fenton reaction to produce highly toxic ˙OH radicals. This mechanism permits an O₂-recycling cascade. Hence, even in a severely O₂-poor microenvironment, photoinduced O₂˙⁻ can be sustained at a therapeutically effective level.²¹ This breakthrough drew significant attention and spurred numerous subsequent studies, advancing the field of type I PDT and expanding its applications.^20,37,42,43 Currently, type I PDT has transcended traditional boundaries, finding relevance in supramolecular systems,^44–46 bacterial cell membrane disruption^47–49 and even environmental pollutant degradation.⁵⁰

Fig. 1 A concise timeline of key milestones in type I PDT research.

In type I PDT, PSs play a pivotal role in driving the therapeutic process. So far, a diverse array of PSs have been developed, which can be broadly categorized into two categories: (1) organic molecular dyes and (2) inorganic (hybrid) nanoparticles. At the core of type I PDT lies a fundamental photochemical reaction in which electrons are transferred from PSs to oxygen or other substrates, generating radical cations or anions. Notably, such an electron transfer mechanism differs significantly between organic molecules and inorganic nanoparticles, reflecting their distinct photophysical and photochemical properties.

As depicted in Fig. 2a, upon photoirradiation, type I organic molecular PSs absorb photons and transition to a short-lived excited singlet state (i.e., ¹[PS*]). From this state, PSs can return to their ground state (i.e., ⁰[PS]) via fluorescence emission or a nonradiative decay process (e.g., vibrational relaxation). Alternatively, ¹[PS*] can undergo intersystem crossing (ISC), a nonradiative transition to a long-lived triplet state (i.e., ³[PS*]), which is either isoenergetic or slightly lower in energy. Conventionally, ³[PS*] is understood to directly transfer electrons to surrounding O₂ (and water), resulting in the formation of O₂˙⁻ and ˙OH radicals. These radicals can initiate cascaded reactions that disrupt cellular components, such as membranes and lipids, ultimately leading to cell death (Fig. 2b).⁵¹ This mechanism represents the most well-characterized and widely studied type I pathway. However, an often-overlooked alternative pathway involves electron transfer between ³[PS*] and biomolecules (Fig. 2c). Cells contain a variety of electron-rich biomolecules, e.g., amino acids, proteins, RNA, DNA, and NAD(P)H. Upon photoexcitation, these biomolecules can transfer electrons to the excited PSs, forming anionic PS species. These intermediates subsequently transfer one electron to O₂ to generate O₂˙⁻, highlighting the ability of ³[PS*] to engage oxygen indirectly.^52–54 This mechanism, in theory, is similar to photoredox catalysis, thus it has gradually evolved into a newly recognized type I mechanism of action termed “photocatalytic biomolecular conversion” (discussed further in Section 6).⁵⁵

Fig. 2 Type I PDT mechanism in organic molecules. (a) Jablonski energy diagram of photochemistry in PDT. (b) Schematic illustration showing how the ROS are generated in conventional type II and type I mechanisms of action. (c) Schematic illustration of photocatalytic biomolecular conversion as a new type I PDT mode of action.

It is worth noting that theoretically, singlet state PSs (i.e., ¹[PS*]) are also capable of initiating electron transfer pathways to finish type I photochemical reactions. However, due to the extremely short lifetime of ¹[PS*], this pathway is less significant and is not considered a dominant mechanism in type I PDT, often being overlooked in mechanistic studies.

Different from type I organic molecular PSs, inorganic (hybrid) nanomaterials displayed a “photogenerated electron holes” mechanism (as depicted in Fig. 3a).^25,56 Upon photoirradiation, the energy of the light sources must exceed the bandgap of the nanomaterials. This excitation promotes electrons from the valence band (VB) to the conduction band (CB), creating a charge-separated state characterized by an electron–hole pair. In this process, electrons in the CB can transfer to molecular oxygen, generating O₂˙⁻ for type I PDT. Simultaneously, holes in the VB react with water molecules, forming ˙OH for a photochemotherapy (PCT) process that is independent of O₂ and distinct from PDT. Thus, type I PDT in inorganic (hybrid) nanomaterials is often accompanied by PCT. A prominent example is modified titanium dioxide (TiO₂), an ideal type I inorganic (hybrid) PS due to its efficient photogenerated charge separation.⁵⁶ Upon illumination, photogenerated electrons produce O₂˙⁻, and photogenerated holes react with water to form ˙OH radicals, representing an O₂-independent PCT process. Similar mechanisms have been observed in diverse inorganic (hybrid) nanomaterials (Fig. 3b), including covalent organic frameworks (COFs),^57,58 metal–organic frameworks (MOFs),⁵⁹ porous organic polymers (POPs),⁶⁰ and carbon dots (CDs).^61,62

Fig. 3 (a) Schematic diagram of the typical excitation process in inorganic nanomaterials, illustrating the transition of electrons from the valence band (VB) to the conduction band (CB) upon photoirradiation, followed by type I PDT and PCT processes. (b) Examples of diverse inorganic and hybrid nanomaterials employed in type I PDT, such as covalent organic frameworks (COFs), metal–organic frameworks (MOFs), porous organic polymers (POPs), and carbon dots (CDs).

3.2. Type I ROS chemistry and biology

ROS refers to a group of chemically reactive molecules derived from the incomplete reduction of oxygen or water, mainly including ¹O₂, O₂˙⁻, ˙OH, and H₂O₂.⁶³ These molecules are integral to the growth and development of living organisms, playing a pivotal role in regulating various physiological functions.⁶⁴ However, excessive ROS production can disrupt cellular homeostasis, leading to oxidative stress. This condition is implicated in numerous diseases, including chronic inflammation,⁶⁵ cancer,⁶⁶ neurodegenerative diseases,⁶⁷etc. Over the last century, significant advances have been made in understanding the intricate redox chemistry of ROS, which has, in turn, accelerated the development of ROS-based therapeutics, especially PDT.⁶⁸ This section focuses on the photophysical and photochemical properties of O₂˙⁻, ˙OH, and H₂O₂, the key ROS species involved in type I PDT (Fig. 4).

Fig. 4 Biomolecule oxidation pathways mediated by ROS generated during type I PDT. The diagram illustrates key oxidative reactions induced by ROS, including ˙OH, O₂˙⁻, and associated cascade reactions.

3.2.1. Superoxide radicals. Superoxide radicals (O₂˙⁻), the primary ROS generated during type I PDT, are monovalent reduction products of molecular oxygen. Their behavior is influenced by environmental factors such as surrounding pH and concentration, with lifetimes typically under 0.05 s and diffusion distances of only ∼40 μm under physiological conditions.⁶⁹ In neutral aqueous solutions, the deprotonation of O₂˙⁻ is weak due to the relatively low redox potential of the O₂/O₂˙⁻ pair (i.e., −0.16 eV vs. normalized hydrogen electrode). However, under acidic conditions, O₂˙⁻ readily transforms to HO₂˙, a more potent oxidant with a higher redox potential.⁷⁰ This shift enhances its oxidative capacity. Nevertheless, O₂˙⁻ demonstrates limited reactivity, primarily oxidizing small, highly reductive molecules such as catecholamines, tetrahydroflavins, leukoflavins, and sulfites. It is incapable of directly reacting with larger biomolecules like nucleic acids, carbohydrates, or lipids.

Despite its short lifespan and limited diffusion, O₂˙⁻ is highly effective in initiating cascade biochemical reactions that lead to target cell lethality, such as in cancer therapy.²¹

1. Disproportionation reaction

O₂˙⁻ is catalyzed by intracellular SOD (i.e., superoxide dismutase) to form H₂O₂ and molecular oxygen via a rapid disproportionation reaction with a rate constant of 6.4 × 10⁹ M⁻¹ s⁻¹ (reaction (1)).⁷¹

2. Fenton and Haber–Weiss reactions

H₂O₂ produced in the disproportionation participates in the Fenton reaction, reacting with Fe²⁺ to generate ˙OH, a highly toxic species. Concurrently, remaining O₂˙⁻ can react with Fe³⁺ to regenerate oxygen (reaction (2)).⁷² O₂˙⁻ also modulates H₂O₂ reduction through the Haber–Weiss reaction, producing ˙OH and oxygen (reaction (3)).

These highly reactive ˙OH radicals cause irreversible oxidative damage to target cells, effectively eliminating them. Furthermore, the oxygen generated during these processes alleviates hypoxia-resistance in cancer cells, enhancing the therapeutic efficacy of type I PDT and endowing it with a “partial oxygen-recyclable” anti-hypoxia mechanism (as proposed by Li et al. in 2018).

Reactions:

3.2.2. Hydroxyl radicals. Hydroxyl radicals (˙OH) are among the most reactive and potent ROS, typically generated through the homolytic cleavage of molecular water. Their lifetime is extremely short, often measured in nanoseconds.⁷³ Recent studies have simulated the lifetime of ˙OH in aqueous solutions to be approximately 30 ps,⁷⁴ limiting their diffusion distance to just a few molecular diameters. Physiologically, ˙OH radicals are primarily formed via the Fenton reaction as well as metal (i.e., intracellular Fe²⁺ and Cu⁺) catalyzed Haber–Weiss reaction.⁷⁰ These reactions ensure a localized and targeted production of ˙OH within cells.

Significantly, ˙OH is the most lethal ROS produced during the type I PDT process due to its extraordinary oxidative properties. The oxidative potential of the ˙OH/H₂O pair is as high as +2.30 eV, enabling ˙OH to abstract electrons from almost all nearby molecules at extremely high reaction rates (reaction rate constants between 10⁹ and 10¹⁰ M⁻¹ s⁻¹).⁷⁵ For instance, ˙OH readily participates in hydrogen-abstraction reactions, producing water and substrate radicals (R˙).⁷⁶ Additionally, ˙OH can react with low-valence metal cations through an electron transfer process, resulting in the formation of high-valence metal cations and OH⁻.⁷⁵ Furthermore, ˙OH interacts with the pentyl group of unsaturated fatty acids, initiating lipid peroxidation. This process generates incorporated ROS, such as peroxyl radicals (ROO˙), which further propagate oxidative damage.⁷⁷ Due to its potent oxidative properties, ˙OH generated in type I PDT inflicts significant damage on a variety of intracellular components, including proteins, DNA, amino acids, lipids, and NAD(P)H, ultimately leading to the destruction of targeted cells.⁷² Given its reliable and robust oxidative capacity, enhancing ˙OH production in type I PDT has emerged as a prominent research hotspot in recent years.

3.2.3. Hydrogen peroxide. Hydrogen peroxide (H₂O₂), a product of the two-electron reduction of molecular oxygen, predominantly exists in a neutral state under physiological conditions due to its high pK_a value (∼11.8).²³ A notable characteristic of H₂O₂ is its reactivity towards the thiol group (–SH), enabling it to interact with various thiol-containing compounds (e.g., homocysteine and glutathione). The reaction rate constant for these redox processes varies significantly, ranging from 10² M⁻¹ s⁻¹ to 10⁷ M⁻¹ s⁻¹ depending on the pH of the thiol compounds.⁷⁸ Additionally, H₂O₂ can be efficiently reduced by diverse transition metals (e.g., cobalt and manganese) to produce ˙OH, a property extensively leveraged in applications involving ˙OH generation.

It is noteworthy that type I PDT does not generate H₂O₂ directly but relies on the dismutation of O₂˙⁻, as discussed in Section 2.2.1. Biologically, the cumulation of intracellular H₂O₂ also primarily arises from O₂˙⁻ dismutation catalyzed by SOD (reaction (1)).⁷⁹ In addition to SOD, various enzymes, including peroxidases, catalases (CAT), and peroxiredoxins, play critical roles in H₂O₂ metabolism. Ascorbate peroxidase, a specific type of peroxidase, catalyzes H₂O₂ decomposition through a ping-pong mechanism, producing water and ascorbate radicals. This mechanism represents one of the primary pathways for intracellular H₂O₂ scavenging.⁸⁰ In addition, CAT also plays an important role in the catalysis of H₂O₂. Of note, CAT-mediated H₂O₂ decomposition yields molecular oxygen, distinguishing it from the ascorbate peroxidase pathway.⁸¹ This oxygen production is particularly significant in mitigating tumor hypoxia, thereby having been widely used for enhancing the effectiveness of PDT.

4. Methods to identify type I PDT

Type I PDT has rapidly emerged as a powerful tool for cancer and anti-infection therapy. However, there remains a lack of clear guidelines for identifying type I PSs. Accurate characterization of ROS is crucial for distinguishing between type I and type II PDT modalities. This section outlines fundamental chemical principles and methodologies for precisely analyzing ROS during PDT processes, both in solution tests and in cellular experiments.

4.1. Detection methods for O₂˙⁻

The detection of O₂˙⁻, a key ROS in type I PDT, can be achieved through three primary approaches: (1) optical absorption in ultraviolet and visible (UV) and infrared (IR) regions (Section 4.1.1); (2) fluorescence assay (Section 4.1.2); and (3) spin-trapping electron spin resonance (ESR, Section 4.1.3).

4.1.1. Optical absorption (UV and IR) for O₂˙⁻ detection. The UV absorption of the O₂˙⁻ can be observed at 245 nm, with a molar absorption coefficient of 2350 M⁻¹ cm⁻¹.⁸² However, direct measurement of O₂˙⁻ absorbance in solution poses big challenges due to its short wavelength and relatively low molar absorption coefficient. Additionally, protonation of O₂˙⁻ (i.e., ˙OOH) shifts the absorption peak to 225 nm, with a further reduced molar absorption coefficient of 1400 M⁻¹ cm⁻¹,⁸² making UV measurement more difficult. In contrast, IR spectroscopy offers a more reliable approach for detecting O₂˙⁻. Characteristic absorption bands for O₂˙⁻ are observed in the ranges of 1050–1200 cm⁻¹ or 1005–1016 cm⁻¹,⁸³ providing a robust alternative for identifying O₂˙⁻ in various environments.

4.1.2. Fluorescence assay for O₂˙⁻ detection. Fluorescence-based detection methods are widely favored due to their ease of use, rapid processing, and cost efficiency. Various fluorescence probes have been developed for detecting O₂˙⁻ in solutions or in vitro. Among these, dihydrorhodamine 123 (DHR123) is a well-established probe for O₂˙⁻ detection in solution.^84,85 DHR123 undergoes selective oxidation by O₂˙⁻ to produce the fluorescent compound rhodamine 123. Since DHR123 has been reported to be able to detect H₂O₂ in the presence of peroxidase,^86,87 its specificity for O₂˙⁻ has been a subject of debate.⁸⁵ To address this, we conducted a responsiveness experiment and demonstrated that DHR123 only reacts selectively with O₂˙⁻. Without peroxidase, DHR123 also shows no responses to H₂O₂ (Fig. 5). This finding confirms DHR123 as a reliable and selective probe for sensing O₂˙⁻ in various solvents (e.g., water, methanol, and DMSO). However, due to the ubiquitous presence of peroxidases in intracellular environments, DHR123 is unsuitable for detecting O₂˙⁻ in cells. Other typical O₂˙⁻ probes include dihydroethidium (DHE)⁸⁸ and MitoSOX Red,⁸⁹ which are widely used membrane permeable fluorescence indicators. DHE reacts with O₂˙⁻ to form 2-hydroxyethidium, which can integrate into chromosomal DNA to emit red fluorescence in the nucleus.⁸⁸ MitoSOX Red, a cationic derivative of DHE, is specifically designed to detect mitochondrial O₂˙⁻.⁸⁹ Its detection mechanism mirrors that of DHE, making it ideal for visualizing O₂˙⁻ within mitochondria. However, both DHE and MitoSOX Red required the presence of DNA for fluorescence emission, limiting their applicability to cellular studies rather than solution-based tests. Recently, HKSOX-1, a trifluoromethyl sulfonate ester-functionalized 5-carboxy-2′,4′,5′,7′-tetrafluorofluorescein, was developed as a novel O₂˙⁻ probe.⁹⁰ This commercially available probe can specifically detect O₂˙⁻ in both solutions as well as intracellular environments, overcoming some of the limitations of traditional fluorescence probes.

Fig. 5 Fluorescence intensity of DHR123 for the detection of various ROS and ONOO⁻ in aqueous solution, confirming that DHR123 exhibits selective response to O₂˙⁻.

While multiple fluorescence probes have been widely used for O₂˙⁻ detection, their limitations must be carefully considered. For DHR123, although its specificity for O₂˙⁻ in solution is confirmed (Fig. 5), its reactivity with H₂O₂ in the presence of peroxidases restricts its utility in cellular environments where endogenous peroxidases are abundant. DHE and MitoSOX Red, while membrane-permeable and suitable for intracellular O₂˙⁻ detection, require integration into DNA for fluorescence emission. This DNA dependency limits their application in non-nuclear or extracellular contexts (e.g., mitochondrial matrix and cytoplasmic vesicles). Furthermore, DHE exhibits non-specific oxidation by other ROS (e.g., ONOO⁻, HOCl) and redox-active metal ions in cells, leading to potential false-positive signals. To mitigate this, parallel control experiments using O₂˙⁻ scavengers (e.g., superoxide dismutase, SOD) is essential for validation. Based on their ease of use in experimental setups and the reliability of the results, DHR123 and DHE are considered the preferred probes for detecting O₂˙⁻ in solution and within cells, respectively (Table 1).

Table 1 A comparison of probe sensitivity, false-positive risk, and application scope between DHR123 and DHE

Probes	Analyte sensitivity	False-positive risks	Preferred scope of application	Validation protocols
Note: ^+high; ^−low
DHR123	+ + + (O2˙^−)	− − − (solution test)	Solution test	(1) Mixing DHR123 with PSs in an aqueous solution, followed by light irradiation.
	− − − (other ROS)	+ + + (in cell test)		(2) Measuring the fluorescence spectra to confirm O2˙^− generation.
DHE	+ + + (O2˙^−)	+ + + (solution test)	In-cell test	(1) Adding 10 μM DHE to PS-treated cells and incubating for 30 mins at 37 °C, then subjecting the cells to light irradiation.
	+ + + (other ROS)	− − − (in cell test)		(2) Using confocal microscopy to observe the red fluorescence in the nucleus.

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4.1.3. ESR spin trapping for O₂˙⁻ detection. Electron spin resonance (ESR) spectroscopy is a powerful technique for detecting O₂˙⁻ in aqueous solution, particularly when used with spin-trapping reagents. One of the most commonly employed spin traps is 5,5-dimethyl-1-pyrroline N-oxide (DMPO), which reacts with O₂˙⁻ to form a DMPO–O₂˙⁻ adduct (˙OOH/DMPO).⁹¹ This adduct exhibits a characteristic multiple spectral pattern, enabling the differentiation of O₂˙⁻ from other free radicals. However, a limitation of the instability of the DMPO–O₂˙⁻ adduct, which typically has a short lifetime of usually 0.5–1.3 min, leads to potential inaccuracies in O₂˙⁻ detection. This transient species is prone to convert to DMPO–˙OH adducts (˙OH/DMPO).⁹¹ To address this issue, advanced spin-trapping reagents have been developed, such as 5-diethoxyphosphoryl-5-methyl-1-pyrroline N-oxide (DEPMPO) and 5-tert-butoxycarbonyl-5-methyl-1-pyrroline N-oxide (BMPO).⁹² These reagents exhibit significantly extended adduct lifetimes, with BMPO–O₂˙⁻ and DEPMPO–O₂˙⁻ adducts persisting for over 10 min and 23 min, respectively.⁹³ This enhanced stability greatly improves the accuracy and reliability of O₂˙⁻ identification using ESR spectroscopy.

4.2. Detection methods for ˙OH

Due to its extremely high reactivity, ˙OH is often regarded as the most toxic ROS in type I PDT. The primary detection methods for ˙OH include (1) fluorescence assay (Section 4.2.1) and (2) spin-trapping ESR (Section 4.2.2).

4.2.1. Fluorescence assay for ˙OH detection. Several reagents capable of being oxidized by ˙OH to produce fluorescein have been extensively studied. Among these, 3′-(p-hydroxyphenyl) fluorescein (HPF) and 3′-(p-aminophenyl) fluorescein (APF) are the most commonly used fluorescence probes for the ˙OH detection in PDT.⁹⁴ Significantly, both HPF and APF exhibit selective reactivity with ˙OH, leading to the release of fluorescein molecules that emit bright green fluorescence. Notably, these probes are not influenced by other ROS, such as O₂˙⁻, ¹O₂, and H₂O₂. Additionally, their excellent photostability makes them well-suited for detecting ˙OH, even in complex intracellular environments.

4.2.2. ESR spin trapping for ˙OH detection. Spin trapping is a conventional and reliable method for detecting ˙OH production. As mentioned in Section 4.1.3, DMPO is widely used as a spin trap for ˙OH detection.⁹¹ In solution, DMPO reacts with ˙OH to form a ˙OH/DMPO adduct, which exhibits a characteristic quartet in its spectral parameters, enabling easy identification. In addition to DMPO, advanced spin traps such as BMPO,⁹² 5-(2,2-dimethyl-1,3-propoxycyclophosphoryl)-5-methyl-1-pyrroline N-oxide (CPYPMPO)⁹⁵ and α-(4-pyridyl-1-oxide)-N-tert-butylnitrone (POBN)⁹⁶ have also been employed to enhance the sensitivity and accuracy of ˙OH detection.

4.3. Detection methods for H₂O₂

Among the ROS involved in type I PDT, H₂O₂ stands out as the only stable molecule. This stability allows for its separate detection after the decay of other ROS (i.e., O₂˙⁻ and ˙OH). To date, two primary methods have been widely employed for H₂O₂ detection: (1) direct measurement of optical absorption in the UV region (Sections 4.3.1) and (2) fluorescence assays (Section 4.3.2).

4.3.1. UV absorption for H₂O₂ detection. H₂O₂ exhibits a molar absorption coefficient of only 0.01 M⁻¹ cm⁻¹ at 360 nm, increasing gradually to 13 M⁻¹ cm⁻¹ at 260 nm.⁶³ However, these coefficients are relatively low, making it challenging to detect the small quantity of H₂O₂ in solution using UV absorption alone. As a result, direct UV absorption measurement is generally less practical for low-concentration H₂O₂ detection.

4.3.2. Fluorescence assay for H₂O₂ detection. A variety of fluorescence probes have been developed for H₂O₂ detection, as summarized in Table 2. Among these, p-hydroxyphenylacetic acid (HPA)⁹⁷ and N-acetyl-3,7-dihydroxyphenoxazine (Amplex Red)⁹⁸ are widely used under diverse conditions. Both HPA and Amplex Red are readily oxidized by H₂O₂ in the presence of horseradish peroxidase, producing fluorescent molecules. These fluorescence signals can be efficiently detected using a fluorescence spectrophotometer or confocal laser scanning microscopy, depending on the experimental setup.

Table 2 Preferred standard assays for O₂˙⁻, ˙OH, and H₂O₂ detection based on commercially available probes

Probes	Analyte	λ abs (nm)^a	λ em (nm)^a	Scope of application
a In aqueous solution.
DHR123	O2˙^−	509	529	Solution test
DHE	O2˙^−	518	606	In-cell test
MitoSOX	O2˙^−	510	580	In-cell test
HKSOX-1	O2˙^−	509	534	Solution and in-cell tests
HPF	˙OH	490	515	Solution and in-cell tests
APF	˙OH	490	515	Solution and in-cell tests
HPA	H2O2	320	420	Solution and in-cell tests
Amplex Red	H2O2	572	583	Solution and in-cell tests
DMPO	˙OH or O2˙^−	—	—	Solution test
BMPO	˙OH or O2˙^−	—	—	Solution test
DEPMPO	O2˙^−	—	—	Solution test
CPYPMPO	˙OH	—	—	Solution test

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4.4. Validation via ROS scavengers

The identification of dominant ROS in type I PDT necessitates the use of selective scavengers as critical analytical tools. A panel of well-characterized scavengers was shown here, each targeting specific ROS with distinct mechanisms. For example, sodium pyruvate mediates non-enzymatic decarboxylation, specifically neutralizing H₂O₂ without affecting other ROS species.⁹⁹ Superoxide dismutase (SOD), a cell-impermeable enzyme, can only catalyze the dismutation of extracellular O₂˙⁻, while Tiron (4,5-dihydroxy-1,3-benzenedisulfonic acid) can penetrate cell membranes to quench intracellular O₂˙⁻ and modulate electron transfer pathways.⁸⁸ D-mannitol acts as an efficient ˙OH scavenger through radical hydrogen abstraction.¹⁰⁰ Co-treatment of PS-loaded systems with these scavengers under illumination, followed by comparative cytotoxicity or ROS quantification assays, provides direct evidence for ROS specificity and their relative contributions to PDT (Table 3).³²

Table 3 Selective scavengers for O₂˙⁻, ˙OH, and H₂O₂

ROS species	Scavenger	Mechanism	Scope of application
O2˙^−	SOD	Catalyzes O2˙^− dismutation to H2O2 and O2	Solution test
O2˙^−	Tiron	Scavenging O2˙^−	Solution and in-cell tests
˙OH	D-Mannitol	Hydroxyl radical quenching	Solution and in-cell tests
H2O2	Sodium pyruvate	Non-enzymatic H2O2 decomposition	Solution and in-cell tests

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5. A palette of type I PSs

This section presents a fresh perspective on the design mechanism of type I PSs, particularly from the viewpoint of small molecular chemistry. Despite significant advancements, systematic and rational approaches to designing exclusive type I PSs remain rare and elusive. To address this, we summarize state-of-the-art breakthroughs from the last seven years, focusing on molecular design, photochemical mechanisms, and their implications for cancer imaging, diagnosis, and therapeutics. Additionally, we explore methods to enhance type I photochemistry, providing a foundation for improved therapeutic efficacy. Detailed discussions are organized in the following subsections.

5.1. Phenothiazine derivatives

Efficient therapy in PDT requires careful selection of the underlying mechanism. The dynamics of substances generated during PDT differ significantly between type I and type II mechanisms. For instance, type II PDT often fails in a hypoxia environment due to oxygen depletion, making mechanism selection a critical factor in successful treatment design. Type I PDT offers a promising alternative in such environments, but its efficacy relies heavily on the PS molecules. A practical molecule design tailored to type I mechanisms often sparks debate due to the challenges in achieving specificity and efficacy. Continuous validation and refinement are crucial to ensuring therapeutic success. This section highlights key studies, particularly on ENBS, offering valuable insights and directions for future research.

In 2018, we reported the first-generation specific O₂˙⁻ generator, ENBS, as a landmark discovery stemming from an in-depth exploration of Nile blue dye modifications with chalcogen elements (Fig. 6a).²¹ ENBS is characterized by its amphiphilic properties and versatile chemical reaction sites, enabling functionalization and optimization for diverse applications (Fig. 6b). Over the last seven years, extensive research has demonstrated its utility in targeted cancer therapy, cell signaling regulation, immune activation, phage engineering, and nanomedicine.^{32,33,48,49,101–110} For example, ENBS conjugated with targeting biomolecules (e.g., ascorbate analog,¹¹¹ antibodies,¹⁰² and biotin²¹) selectively accumulates in cancer cells, achieving specific ablation (Fig. 6a and c). Additionally, the molecular design optimizes intersystem crossing to favor type I PDT, as evidenced by imbalanced optical absorbance and fluorescence changes (Fig. 6d).³³ This study gives an impressive perception of the molecular design for O₂˙⁻ generators. When combined with immunotherapy, ENBS exhibits synergistic effects. For instance, overexpressed ROS from PDT can induce pyroptosis and stimulate immune responses, especially when paired with 1-MT to inhibit IDO (indoleamine 2,3-dioxygenase, an immune suppressor), resulting in enhanced ROS release and tandem immunotherapy effects (Fig. 6e).¹¹² Some other studies from our group also expand the family of ENBS, such as targeting droplets for ferroptosis induction,¹⁰⁴ activating iterative revolutions of cancer immunity for photoimmunotherapy,¹¹³ electrostatically attracting siRNA for synergistic gene therapy,¹⁰⁹ engineering photo-PROTAC augmenter for pyroptosis activation,¹⁰⁷ and self-reporting photodynamic nanobody for sustainable large-volume tumor eradication.¹¹⁰ These studies have established ENBS as a benchmark for type I PDT research, solidifying our group's position in the field both domestically and internationally.

Fig. 6 First-generation exclusive type I O₂˙⁻ photogenerator (ENBS) utilizing a chalcogen element regulation strategy to modify Nile blue molecule. (a) ENBS-based emerging applications, such as photoimmunotherapy, synergetic photochemotherapy, photo-controlled drug release, photosensitizable phage, and nanomedicine. (b) Cellular mechanisms underlying type I PDT. (c) Schematic illustration of a biotin-targeted type I O₂˙⁻ photogenerator. (d) Schematic illustration of a FRET-based type I O₂˙⁻ photogenerator. (e) Schematic illustration of photo-controlled immune drug 1-MT release for synergistic type I photoimmunotherapy. Figure created with Biorender.com.

The excellent properties of ENBS explored in our research have inspired other groups, further advancing molecular design and broadening application scope. For example, ENBS can be integrated into nanocarriers to enable multifunctional applications, such as drug delivery, PDT, and MR imaging (Fig. 7).¹¹⁴ Besides, PSs with endoplasmic reticulum targeting abilities play a key role in enhancing the efficacy of PDT. NBS-ER¹¹⁵ was designed to target the ER and generate O₂˙⁻ under hypoxic conditions (Fig. 8a). This ability effectively triggers ER stress and induces apoptosis, showing significant PDT efficacy and good biocompatibility in 4T1 tumor-bearing mice. Some approaches utilizing O₂˙⁻ indirectly for therapeutic purposes were also reported (e.g., BPT in Fig. 8b).¹¹⁶ Peroxynitrite (ONOO⁻), a cytotoxic agent with potent anticancer activity, can overcome drug resistance through the in situ reaction of O₂˙⁻ and NO˙. By regulating O₂˙⁻ production via light control, precise ONOO⁻ production is achieved, overcoming challenges like low selectivity and short biological half-life (Fig. 8b). Insights from antibacterial studies also contribute to understanding PDT's selectivity in cancer treatment. For example, replacing oxygen in Nile blue derivatives with chalcogen elements alters ROS generation and antibacterial effects (Fig. 8c).¹¹⁷ It shows a different pattern depending on which chalcogen element (i.e., S or Se) replaces the oxygen site in the basic skeleton of Nile blue. The O₂˙⁻ generated by NBS-N shows high effectiveness in targeting Gram-positive bacteria. In contrast, NBSe-N primarily produces ¹O₂, which damages cell membranes non-selectively due to its high reactivity. This highlights the importance of selectivity in type I PDT, especially for bacteria treatment, where precision in targeting is essential. Aggregation of PSs in aqueous media is a well-known challenge as it can result in quenching of ROS production and conversion of absorbed light energy into heat, reducing the overall efficacy of PDT. To address this issue, recent research has shown that assembling immune interferon gene stimulants into polymers containing NBS (PNBS/diABZI)⁴² can form nanoagonists (Fig. 8d). These nanoagonists facilitate H-aggregation and intersystem crossing, enhancing the production of O₂˙⁻ and ¹O₂ by 3-fold. Additionally, combining ENBS (M-TPO)³⁷ with mitochondrial targeting drugs enables interference with signal pathway crosstalk, providing precise spatiotemporal control over inflammatory responses (Fig. 8e). This approach holds promise for designing strategies to mediate pyroptosis in cancer cells expressing GSDME, which could be a powerful therapeutic avenue. Besides, the design of ROS-activatable ENBS (NTP)¹¹⁸ for anticancer drug delivery further expands potential applications of type I PSs in cancer therapy (Fig. 8f). These findings not only demonstrate the effectiveness of ENBS as a robust O₂˙⁻ generator but also offer valuable insights for scientists exploring new strategies in cancer biomedicine.

Fig. 7 ENBS molecule mediated self-assembled vesicles of amphiphilic block copolymers for activated NIR fluorescence imaging and tissue-specific type I PDT under cellular and in vivo conditions. Figure created with Biorender.com.

Fig. 8 Schematic illustration of chemical structures and mechanisms of ENBS analog type I PSs. (a) Type I PS (NBS-ER) designed for endoplasmic reticulum targeting research. (b) Proposed molecular mechanism leading to the generation of ONOO⁻ and the stable photoproduct BPT upon red light excitation of the molecular hybrid BPT-NO. (c) Schematic illustration of O₂˙⁻ generated by NBS-N, along with the selective bactericidal effect on Gram-positive bacteria and the healing of Gram-positive bacterial infections in murine wound models using NBS-N. (d) Development of a nanoagonist (PNBS/diABZI) through the combination of stimulator of interferon genes (STING) and ENBS, aimed at enhancing tumor response. (e) ENBS PS (M-TOP) with mitochondrial targeting for optimized immunotherapy drug delivery. (f) ROS-activatable ENBS PS (NTP) for targeted anticancer drug delivery. Figure created with Biorender.com.

5.2. Porphyrin/chlorin/phthalocyanine-based photosensitizers

Light-induced radical generation is a hallmark of a photo-derived therapeutic approach. Since the pioneering introduction of porphyrin/phthalocyanine derivatives as PSs, these compounds have found wide applications across various fields, often incorporated into composite materials.^119,120 These PSs serve as a foundation for the development of type I PS through the following strategic approaches. (1) Electron-donating strategy: when a PS is incorporated with an electron-rich polymer, its type I efficiency is enhanced. (2) Self-assembly strategy: the effectiveness of type I photosensitization increases when the PS self-assembles into electron-rich nanoparticles, particularly in aqueous environments. These strategies have been explored in several studies, illustrating their impact on the photoactivation processes of PSs. For example, modulation of type I and II photoactivation mechanisms of PSs has been investigated by assessing how the carrier microenvironment affects the photophysical properties of the PS, subsequently influencing its therapeutic efficacy.¹²¹ As illustrated in Fig. 9a, incorporating 5,10,15,20-tetrakis(meso-hydroxyphenyl)porphyrin (mTHPP) to electron-rich polymer, (2-(diisopropylamino)ethyl methacrylate) (PDPA) micelles, significantly increased the generation of O₂˙⁻ over ¹O₂, transitioning the photoactivation process of mTHPP from type II to type I mode. The use of PDPA micelles (PEO-b-PDPA) enhanced phototoxicity in multiple cancer cell lines compared to the electron-deficient poly(D,L-lactide) control. These data suggest that micelle carriers not only improve the bioavailability of PS but also modulate their photochemical properties, enhancing PDT efficacy. In another study, the self-assembly of a novel phthalocyanine molecule into nanodot (NanoPcA) demonstrated highly efficient O₂˙⁻ generation through the type I mechanism (Fig. 9b).¹²² This innovative approach underscores the potential of self-assembled nanoparticles for advancing the effectiveness of type I PDT.

Fig. 9 Chemical structures of porphyrin/phthalocyanine derived type I PSs. (a) Schematic showing porphyrin-derived PSs (mTHPP) incorporated into electron-rich micelles can trigger a transition of PDT mechanism of action from type II to type I mode. The ROS generation mechanism of mTHPP under hypoxia and normoxia is shown. (b) Schematic illustration of phthalocyanine-derived nanodots can enhance type I PDT efficiency; the chemical structure of PcA and control molecules are shown.

In addition, a variety of type I PS based on chlorin derivatives have been developed due to their high yield of exogenous O₂˙⁻ generation, excellent therapeutic efficacy, as well as a straightforward preparation method. In 2009, two novel observations were made regarding light-induced ROS generation in aqueous solutions using water-soluble derivatives of the photosynthetic pigment bacteriochlorophyll a (Bchl), WST09 and WST11 (Fig. 10a).²⁹ These studies revealed that WST11 exclusively generates O₂˙⁻ and ˙OH, with no detectable formation of ¹O₂ in aqueous solutions. Furthermore, WST11 forms a noncovalent complex with human serum albumin (HSA), which functions as a photocatalytic oxidoreductase, enabling approximately 15 cycles of electron transfer from the HSA protein to molecular oxygen in the solution. These findings challenge the conventional paradigm of porphyrin- and chlorin-based PDT, which is traditionally considered to be primarily mediated by the formation of ¹O₂, particularly in vascular-targeted photodynamic therapy (VTP). In another study, Yang et al. measured the PDT activities for dicyano-bacteriochlorins, including the parent compound (BC), dicyano derivative (NC)₂BC, and corresponding zinc (NC)₂BC-Zn and palladium chelate (NC)₂BC-Pd, against cancer cells (Fig. 10b).¹²³ Their results demonstrated that the PS with the shortest triplet lifetime, (NC)₂BC-Pd, exhibited the highest activity. This outcome suggests that O₂˙⁻ and ˙OH, well-known precursors of important pathophysiological processes, can be leveraged for effective tumor eradication, leading to new design paradigms for type I PS undergoing photoactivation.

Fig. 10 Schematic illustration of porphyrin and phthalocyanine derived PS for enhanced O₂˙⁻ generation yield. (a) Chemical structure of [Pd] bacteriopheophorbide derivate, WST09, WST11, and Redaporfin. (b) Chemical structure of phthalocyanine ((NC)2BC-Zn and (NC)2BC-pd). (c) Chemical structure of phthalocyanine (PcAF) and its assemblies NanoPcAF used for type I PDT. (d) Chemical structure of silicon phthalocyanine (PcM) and its assemblies NanoPcM designed for turn-on imaging and PDT-based immunotherapy. (e) Chemical structure of silicon phthalocyanine derivates (PcSZ) for enhanced O₂˙⁻ generation via the FRET effect.

While porphyrin and chlorin-based type I PSs have been extensively studied, the development of type I PSs using phthalocyanine has also been actively pursued. A novel design was introduced to shift the photophysical and photochemical properties of traditional phthalocyanine-based PSs from type II photoreaction to efficient type I photoreaction and vibrational relaxation-induced photothermal conversion for photothermal therapy (PTT) as well as multimodal imaging (Fig. 10c).¹²⁴ One such innovation, NanoPcAF, is a nanostructured PS based on the self-assembly of silicon(IV) phthalocyanine axially monosubstituted with perphenazine. NanoPcAF demonstrated substantial O₂˙⁻ generation through the type I mechanism and excellent photothermal performance, overcoming tumor hypoxia in PDT. As a result, NanoPcAF exhibited a synergistic effect combining type I photoreaction and photothermal action, effectively inhibiting tumor growth. Furthermore, there is an increasing demand in clinical applications for titanium phthalocyanine-based PSs with high-definition fluorescence imaging capabilities, alongside outstanding therapeutic effects. A noteworthy example is the combination of morpholine-modified silicon phthalocyanine (PcM) with serum albumin (SA) through supramolecular self-assembly technology. This strategy not only reduces background noise but also enhances tumor-targeting fluorescence imaging, addressing key clinical demands (Fig. 10d).¹²⁵ In the latest research, the group explored the use of silicon(IV) phthalocyanine analogs (PcS) as energy donors and successfully paired them with zinc(II) phthalocyanine derivatives (serving as energy acceptors, denoted as PcZ). Using fluorescence resonance energy transfer (FRET) technology, they constructed an efficient O₂˙⁻ generator, PcSZ.¹²⁶ This innovation not only enhances energy conversion efficiency but also improves the safety of the system. These findings offer valuable insights for the future development of type I PSs using phthalocyanine derivatives, with the potential to drive further advancements in related fields (Fig. 10e).

5.3. BODIPY dyes

With the promising therapeutic outcomes of PDT, the development of advanced PS continues to gain momentum. Among these, boron dipyrromethene (BODIPY) stands out as one of the most actively studied PSs due to its high triplet quantum yield and efficient ROS generation. In general, BODIPY sensitizers are classified as type II PSs, capable of producing ¹O₂ with ultrahigh quantum yield.^127,128 However, through strategic molecular engineering, its derivatives offer notable advantages, including low oxygen-dependent type I mechanisms for enhanced hypoxic cancer treatment. Like porphyrin/phthalocyanine, BODIPY serves as an excellent starting point for designing type I PSs, utilizing strategies such as: (1) promoting electron donating efficiency via the introduction of electron-rich polymer (Fig. 11a); (2) triplet state formation from an effective excited-state relaxation by introducing α,β-linked BODIPY dimer/trimer (Fig. 11b); and (3) converting type II to type I PSs via host–guest strategies (Fig. 11c).

Fig. 11 Schematic illustration of chemical structures of BODIPY derived type I PSs. (a) Scheme of electron-rich polymer nanoparticles (NPs). (a-i) Chemical structure of BODIPY derived type I PS IBAB and its assemblies PPIABNP. (a-ii) Chemical structure of BODIPY derived type I PS BDPVDA and its assemblies PBVNPs. (b) Scheme of BODIPY dimers and trimers. (b-i) Chemical structures of traditional and newly developed BODIPY dimers. (c) Scheme of host–guest complexation. (c-i) BODIPY derivates as guests, pllar[5]arenes as hosts, forming nanoparticle assemblies to shift PDT from type II to type I PDT. (c-ii) BODIPY-derivated PS (DA) capable of directly catalyzing the oxidation of water to release highly cytotoxic ˙OH.

To address the limitations of traditional BODIPY-based PSs under hypoxic conditions, PPIABNPs were developed.¹²⁹ This aza-BODIPY-based nanoplatform incorporates a heavy atom to promote ISC and is encapsulated by the electron-rich polymer mPEG-PPDA for improved electron-donating efficiency (Fig. 11a–i).¹²⁹ This approach boosted O₂˙⁻ photogeneration, effectively overcoming hypoxia-induced challenges in PDT. Upon irradiation, aza-BODIPY rapidly transitions to its singlet state and subsequently changes to a charge-separated state facilitated by the electron-rich environment (i.e., electron-rich polymer, mPEG-PPDA), promoting O₂˙⁻ photogeneration. Building on this, PBVNPs were developed by encapsulating the BODIPY-based conjugate in an electron-rich amphiphilic mPEG-PPDA polymer (Fig. 11a-ii).¹³⁰ These NPs exhibited excellent core–shell intermolecular electron transfer, efficiently converting O₂ into O₂˙⁻ under NIR laser irradiation, even under hypoxic conditions. Moreover, in vitro and in vivo studies showed that PBVNPs could release vascular-disrupting agents (VDAs) at the tumor site, disrupting tumor vascular vasculature and hindering metastasis pathways.

Yang's research group reported BODIPY-based type I PSs without heavy atoms to mitigate dark toxicity.¹³¹ They constructed a series of α, β-linked BODIPY dimers and trimers to tune electron interaction between BODIPY moieties, enhancing ISC efficiency through connections between the electron-deficient α site and the electron-rich β site (Fig. 11b-i). This strategy leveraged effective excited state relaxation, where the triplet state (T₁) was populated by internal conversion (IC) from the intermediate triplet (T₂) state. Using this strategy, trimer 2 demonstrated significantly enhanced O₂˙⁻ generation due to its low reduction potential and an ultralong T₁ state lifetime. This was achieved through intermolecular charge transfer to molecular oxygen (Fig. 11b-i). Notably, the generation of ¹O₂ was effectively suppressed. This suppression occurred because the energy gap between the T₁–S₀ states was reduced to a level smaller than the gap between ³O₂ and ¹O₂. Consequently, the T₁ energy states were insufficient to enable ¹O₂ formation via excitation energy transfer, thereby inhibiting type II ROS generation.

Building on these findings, the same research group introduced a series of strategies to develop type I exclusive PSs.^45,132 To overcome the competition between triplet–triplet energy transfer and type II process, they utilized a host–guest complexation approach to convert conventional type II PS into type I PSs. Electron-rich pillar [5] arenes served as the hosts, while iodide BODIPY-based electron deficient type II PSs acted as the guests (Fig. 11c-i).¹³² Upon irradiation, host–guest complexation promoted intermolecular electron transfer from the host to the guest, which was subsequently delivered to molecular oxygen via the type I mechanism, resulting in efficient O₂˙⁻ generation. Antitumor experiments validated this type I-exclusive photosensitization strategy, demonstrating its high PDT efficacy even under hypoxic conditions.

In a recently launched in-depth study, Yang et al. successfully designed and synthesized a BODIPY PS featuring fluorene as an electron-donating group. This PS was synergistically assembled with perylene diimide, an efficient electron acceptor, to construct a supramolecular photodynamic system based on quadruple hydrogen bonding (Fig. 11c-ii).⁴⁵ The resulting innovative PS (DA) exhibits remarkable intermolecular electron transfer and charge separation capabilities under light excitation, efficiently facilitating the generation of radical ion pairs. Notably, under strictly anaerobic conditions, DA demonstrated the ability to directly catalyze water oxidation, releasing highly cytotoxic ˙OH. Concurrently, it transferred captured electrons to pyruvate molecules, enabling a catalytic cycle mechanism tailored for tumor systems. This dual-action functionality highlights the potential of this advanced PS design for enhancing PDT efficacy in challenging hypoxic tumor environments.

5.4. Transition metal complexes

Transition metal-based PSs (TMPs) are increasingly gaining attention in PDT, especially for their efficacy in addressing hypoxic tumors.^133–136 The unique photophysical and chemical properties of TMPs make them promising candidates for enhancing the type I PDT pathway. Key advantages of TMPs in improving PDT outcomes include their supportive electronic configurations, water solubility, remarkable endocytosis efficiency, organelle targeting capabilities, high molar absorption coefficients, and deep tissue penetration due to absorbance in the NIR region.¹³⁵ These PSs also exhibit excellent photostability and efficiently generate ROS by interacting with biomolecules such as amino acids, peptides, and proteins.⁵² The ability of TMPs to activate the type I pathway and generate ROS stems from their partially filled d-orbitals, which facilitate electron transfer to nearby biomolecules. Moreover, the absorbance wavelengths of TMPs can be finely tuned by modifying the central metal ion and electronic environment for their chelating ligands. Through metal–ligand charge transfer (MLCT), the central metal in its reduced form can participate in multiple redox cycles, transferring electrons to oxygen and nearby molecules to generate O₂˙⁻, ˙OH, H₂O₂, and other ROS.^137,138 Beyond the type I pathway, the heavy atom effect inherent in TMPs extends the triplet state lifetime, enabling more efficient energy transfer to molecular oxygen for the generation of ¹O₂. This dual capability enhances PDT efficacy by combining type I and type II pathways synergistically. TMPs are predominantly hydrophilic, facilitating their excretion from normal cells and reducing dark toxicity.¹³⁹ Moreover, certain transition metals can coordinate with serum proteins, extending their circulation time and promoting effective accumulation at the tumor site.¹³⁹ While their relatively large size can increase the risk of accumulation in organs and potential long-term toxicity, the high molar absorption coefficient and selective tumor targeting of TMPs allow for reduced dosing, mitigating toxicity concerns. Recent advancements have highlighted several examples of TMPs, particularly those based on ruthenium (Ru), iridium (Ir), osmium (Os), platinum (Pt), and rhenium (Rh), demonstrating efficient tumor therapy through the type I PDT pathway.

The success of a PS in type I PDT depends on its ability to efficiently transfer electrons to molecular oxygen and neighboring biomolecules, facilitating ROS generation and cancer cell eradication. In this section, we explore recent breakthroughs in TMP-based PDT, emphasizing studies that characterize type I pathways using spectroscopic and validate their efficacy in both in vitro and in vivo tumor models.

5.4.1. Ruthenium-based PSs. Ruthenium (Ru) is one of the most widely studied transition metals for designing PSs in PDT¹³⁴ and other photocatalytic applications.¹⁴⁰ Ru complexes exhibit numerous favorable properties such as partially filled d orbitals, high photostability, and higher molar extinction coefficient; also, their variable oxidation states enable multiple redox reactions, making them highly versatile for PDT.¹⁴¹ The electronic structure of Ru complexes allows efficient energy harvesting across a broad range of visible to NIR wavelengths, facilitating ROS generation (Fig. 12a). Moreover, Ru complexes can bind selectively to biomolecules like enzymes, proteins, and DNA which synergically enhances ROS production. Unlike many other metal-based PSs, Ru complexes are remarkably stable in biological environments and exhibit relatively low dark toxicity, making them ideal candidates for biological studies and applications. Given these advantageous properties, numerous Ru complexes have been synthesized and optimized for PDT efficiency.¹³⁴ Among them, the Ru(II) complexes TLD1433 (Fig. 12b) have demonstrated exceptional efficacy and are currently undergoing phase-IB human clinical trials for the PDT treatment of bladder carcinoma.¹⁴¹ The outstanding performance of TLD1433 is attributed to its unique electronic structure and its ability to undergo multiple transitions, facilitating efficient O₂˙⁻ generation.

Fig. 12 Schematic illustration of representative examples of transition metal-based PSs studied for type I pathway of PDT. (a) Exited state configuration of the ruthenium complex TLD1433, currently in phase II clinical trials. (b) Molecular structure of TLD1433. (b-i) Systematic presentation of the preparation process for the titanium-modified Ru complex TiO₂@Ru@SiRNA. (b-ii) Chemical structures of Ru(II) complexes RuNMe and illustration of the photoinduced ferroptosis triggered by RuNMe upon irradiation. (c) Chemical structure of Os-4T. (d-i) Chemical structures of Ir(III)–COUPY. (d-ii) Chemical structures of Ir-pzpy and Ir-TAP. (d-iii) Molecular structure of Ir-OA.

Encouraged by the early success of Ru complexes in PDT, recent studies have focused on developing Ru-based PSs that specifically and effectively activate the type I PDT pathway. For example, a hypoxia-compatible nanocomposite, TiO₂@Ru@siRNA, was developed by coating TiO₂-nanoparticles with a Ru PS and loading them with hypoxia-inducible factor-1α (HIF-1α) siRNA (Fig. 12b-i).¹⁴⁰ This nanocomposite was shown to localize successfully in lysosomes and, under visible light irradiation (525 nm), generate both O₂˙⁻ and ¹O₂, while escaping the HIF-1α signaling pathway. TiO₂@Ru@siRNA demonstrated effective tumor growth inhibition by reducing hypoxia, increasing ROS generation via the type I pathway, and enhancing the cancer immune response in an immune-deficient patient-derived xenograft (PDX) model. Additionally, Ru complexes with finely tuned conjugated ligands have shown impressive results in type I PDT. For instance, a series of Ru(II) polypyridine complexes were designed to generate a high quantum yield of O₂˙⁻ upon visible light irradiation.¹⁴² Among these, the RuNMe derivative (Fig. 12b-ii) demonstrated the most effective inhibition of tumor cells due to its superior cell uptake compared to other derivatives. Recent advances highlight the potential of Ru complexes as exceptional PS candidates for exclusive type I PDT pathways. To maximize their effectiveness, key design considerations should include achieving high quantum yields for electron transfer to biomolecules rather than oxygen, strong targeting and localization abilities, and enhanced biocompatibility. These features are crucial for ensuring efficient and selective PDT via the type I pathway.

5.4.2. Osmium-based PSs. Osmium (Os) is another transition metal whose polypyridyl complexes have been extensively investigated for PDT over the last two decades.¹⁴³ Compared to Ru analogs, Os complexes offer several advantages, including greater inertness and stability under biological media, higher oxidation states, slower ligand exchange kinetics, and stronger spin–orbit coupling.¹⁴⁴ These properties make Os complexes particularly effective in the type I pathway of PDT, largely due to their unique electronic configuration. Os features a low-lying, empty d-orbital that efficiently accepts electrons from the excited state. While Ru complexes typically absorb light up to 700 nm, Os complexes exhibit absorption up to the NIR region (approximately 900 nm), which is advantageous for deeper tissue penetration. Additionally, Os complexes exhibit preferential DNA binding and targeting capabilities, which contribute to their potent antiproliferative activity.¹³⁸ Due to these favorable characteristics, numerous Os complexes have been synthesized, demonstrating impressive results in type I pathways. Their performance is attributed to their ability to absorb longer wavelengths, enhanced spin–orbital coupling, and faster ISC, facilitated by the heavy atom effect.¹⁴³

Building on the intrinsic strengths of Os, McFarland et al. reported a series of Os(II) complexes conjugated with varying numbers of thiophene rings for application in PDT against hypoxic tumors. These complexes are structurally analogous to the ruthenium-based TLD1433,¹⁴⁵ which is currently in clinical trials. The Os(II) complexes exhibited broad absorption in the visible to NIR region, extending up to 740 nm, with photocytotoxicity increasing alongside the number of thiophene rings. These complexes were found to have good biocompatibility and exhibited comparatively low cytotoxicity to normal cells. Notably, the Os-4T complex (Fig. 12c)¹⁴⁵ is the first Os-based PS to demonstrate photocytotoxicity in a 1% oxygen environment, achieving the highest phototherapeutic index reported to date. This pivotal study highlighted the significant potential of osmium complexes in the type I pathway of PDT. Recent research has further demonstrated that Os complexes offer several advantages over other PSs such as a broader absorption spectrum, stronger spin–orbital coupling, faster ISC, and prolonged lifetime of triplet excited state. These properties make them highly promising candidates for the development of PSs specifically tailored for type I PDT applications.

5.4.3. Iridium-based PSs. In recent years, iridium (Ir) complexes have emerged as highly promising PSs for PDT applications.¹⁴⁶ Their distinctive properties—such as high quantum yields, broad absorption spectra in the visible region, microsecond level triplet state lifetimes, and prolonged excited-state durations—make them an ideal platform for designing PSs tailored to the type I pathway of PDT. Ir complexes are particularly adept at generating multiple free radicals in high yields via interacting with amino acids, proteins, and other biomolecules. In addition to their photophysical advantages, Ir complexes have desirable biological properties, including low toxicity, high stability under physiological conditions, and selective targeting of cancer cells. Recent advancements have introduced innovative design strategies to further enhance PDT efficiency, especially via the type I pathway. One such example is the coumarin-based COUPY dye-modified Ir complex, Ir(III)–COUPY, which was specifically designed and synthesized to improve PDT outcomes.¹⁴⁷ This complex exhibits excellent red light absorption and far-red-light-emitting properties (Fig. 12d-i). Upon visible light irradiation, Ir(III)–COUPY effectively generates O₂˙⁻, enabling type I PDT against HeLa cells. Additionally, Ir(III)–COUPY demonstrates excellent solubility in biological media, robust cellular uptake, and low dark toxicity, making it a highly compatible agent for cancer PDT.

Further advancements include two Ir-based complexes, Ir-pzpy and Ir-TAP (Fig. 12d-ii).¹⁴⁸ The 1,4,5,8-tetraazaphenanthrene (TAP) ligand-based complex Ir-TAP showed effective binding with the neighboring biomolecules and exhibited stronger cytotoxicity under hypoxic conditions, despite generating a lower quantum yield of ¹O₂. These findings underscore the importance of optimizing and tuning chelating ligands to increase the production of other ROS, thereby enhancing the efficacy of type I PDT under hypoxic conditions. Another notable example is the iridium PS Ir-OA, which demonstrated accelerated ROS production within mitochondria (Fig. 12d-iii).¹⁴⁹ This complex also exhibited the unique ability to monitor mitochondrial microenvironments, including polarity, viscosity, and morphology, further expanding its functionality in PDT applications. Designing TMPS specifically for the type I pathway of PDT requires careful consideration of the chelating ligand structure and its interaction with the central metal.

Recent studies highlight that the efficiency of TMPS can be significantly enhanced through strategic modifications, such as optimization of the structure of chelating ligands by incorporating donor groups, selection of a central metal, and accumulation of chromophores to enhance absorption ranges. TMPs offer immense potential to generate diverse ROS species, enabling highly efficient and selective type I PDT. Despite concerns about toxicity, these can often be mitigated by employing lower doses, thanks to the high molar extinction coefficient of these complexes. With their ability to deliver superior photophysical properties, stability under physiological conditions, and tumor selectivity, Ir complexes present versatile opportunities for designing PSs tailored to specific cancer types and therapeutic needs.

5.5. AIE gens

The concept of aggregate-induced emission (AIE) was first proposed in 2001.¹⁵⁰ AIE-active fluorophores, known as AIEgens, exhibit minimal fluorescence in solution but emit intensely in the aggregated state, effectively avoiding the aggregation-caused quenching (ACQ) effects. This unique property has made AIEgens a promising tool for various applications, including PDT. In PDT, aggregation is one of the strategies proposed to enhance ISC, thereby increasing ROS generation. However, traditional PSs (porphyrins, phthalocyanine indocyanine, etc.) being hydrophobic, often aggregate in physiological environments, resulting in ACQ effects. This strong π–π interaction between aggregates reduces their ROS generation efficiency. In contrast, PSs with AIE characteristics exhibit weak fluorescence and limited ROS generation in solution due to energy dissipation through intramolecular motion. Upon aggregation, this non-radiative heat dissipation is significantly reduced due to restricted intramolecular motion (RIM). This promotes fluorescence and enhances the ISC process, leading to increased ROS generation.¹⁵¹ Additionally, the twisted conformation of AIE PSs reduces intermolecular π–π stacking, contributing to both their AIE properties and aggregation-induced generation of ROS (AIG-ROS) characteristics.

Given these advantages, significant efforts have been devoted to developing innovative AIE-based PSs with enhanced ROS generation capabilities and improved therapeutic outcomes. The ISC process, which initiates ROS generation, is critical for the efficiency of AIE PSs. To optimize photosensitization, it is essential to achieve a higher ISC rate, ensuring adequate triplet-state production. Various studies have proposed strategies to enhance the ISC process in AIE PSs, including spin–orbit coupling (SOC) enhancement, donor–acceptor (D–A) molecular engineering,¹⁵² and other approaches. Although substantial progress has been made in designing type I AIE PSs, challenges remain in preferentially generating type I free radical species over type II ROS. This section summarizes the current strategies for developing type I AIE PSs, categorized into three main approaches: (1) D–A engineering, (2) anion–π⁺ incorporation, and (3) acceptor-shielding strategy, as shown in Fig. 13.

Fig. 13 Schematic illustration of AIE-based PSs in solution and aggregated states and their ROS generation behaviors. Illustration of (a) D–A and (b) anion–π⁺ strategies and associated photochemical factors facilitating type I ROS generation. Chemical structures of AIE-based type I PSs employing (a-i to a-iii) D–A and (b-i to b-iii) anion–π⁺ strategies respectively. (c) Chemical structure of AIE-based type PS designed with acceptor–shielding strategy, coupled with a schematic illustration of type I PDT and immunotherapy mechanisms. Figure b-iii created with Biorender.com.

5.5.1. D–A engineering for type I AIE PSs. In the general mechanistic framework of type I PDT, the excited triplet state of a PS (³PS*) must first transfer one electron to its surrounding O₂ environment to form O₂˙⁻. D–A (donor–acceptor) scaffolds are particularly advantageous in this context, as they can serve as intermediates for efficient electron transfer (Fig. 13a). Several type I AIE PSs incorporating D–A frameworks have been reported, including TTTMN,¹⁵³TTS-2F,¹⁵⁴DTTVBI,¹⁵⁵ and TCF-R.¹⁵⁶ Among these, TTTMN exemplifies a sophisticated D–A design. In TTTMN, the triphenylamine (TPA) moiety serves as an efficient electron donor, thiophene serves as both an auxiliary donor and π-bridge and two cyano units act as electron acceptors (Fig. 13a-i). The unique properties of TTTMN are attributed to its highly twisted molecular structure and distinct D–A interactions. In the aggregated state, these features significantly enhance the production of O₂˙⁻ and ˙OH, alongside exhibiting NIR emission. Additionally, TTTMN's efficient ISC process is supported by multiple transition pathways between the lowest singlet excited state (S₁) and the isoenergetic triplet state (Tn), facilitated by a high SOC value. Furthermore, the incorporation of tetraphenylethylene (TPE) introduces abundant molecular rotors to TTTMN, which effectively suppresses fluorescence quenching in the aggregated state. This not only reinforces its AIE properties but also enhances its overall performance in type I PDT applications.

Numerous efforts have been directed toward developing efficient type I AIE PSs using D–A strategies, focusing on enhanced O₂˙⁻ production capabilities and broad biomedical applications (Fig. 14).^156–165 Notably, molecular frameworks incorporating D–A designs, such as TTS-2F, DTTVBI, and TCF-R, provide additional benefits such as absorption in the NIR region. PSs with absorption and emission improve tissue penetration while minimizing auto-fluorescence interference. Furthermore, their broad absorption profiles enable the use of white light sources for ROS generation, thereby simplifying the PDT process. The structural tunability of type I AIE PSs for preferred photochemical mechanisms allows for dual-modal therapies, such as combined type I PDT and PTT, within a single molecular framework. For instance, the AIE PS TTS-2F (Fig. 13a-ii) demonstrates the potential for synergistic type I PDT and PTT.¹⁵⁴ PTT induces localized heating, enhancing O₂ supply in tumor tissues through improved blood flow, which, in turn, boosts the efficacy of type I PDT. The improved PDT then reinforces PTT efficacy, creating a positive feedback loop. Given the competitiveness of energy dissipation processes, careful design of molecular scaffold is crucial to regulate energy dissipation equilibrium through controlled intramolecular rotations. This strategic approach provides a foundation for advancing multifunctional AIE type I PS for NIR-II fluorescence-guided type I PDT and synergistic PTT therapies.

Fig. 14 Schematic illustration of molecular structures of AIE-based type I PSs developed through D–A engineering or anion–π⁺ incorporation strategies.

5.5.2. Anion–π⁺ incorporation for type I AIE PSs. The type I PDT process involves transferring one electron from ³PS* to the surrounding substrate. Enhancing the electron-rich environment around the PS is a practical approach to improve electron capture and, consequently, PDT performance. Anion–π⁺ interaction is such a mechanism for increasing electron transfer.^166,167 This principle underpins the development of several type I PSs, such as TPPM, MeTPPM, and MeOTPPM.¹⁶⁸ In these systems, a TPA moiety (electron donor) is connected to a positively charged pyridinium moiety (as an electron acceptor) through a benzene π-bridge, forming a propeller-shaped AIEgen with an enhanced ICT effect (Fig. 13b-i).¹⁶⁸ Methoxy (–OMe) and methyl (–Me) substituents on the TPA moiety further modulate the electronic properties by suppressing IC and promoting ISC. The incorporation of hexafluorophosphate as a counter ion and methoxy substituents yields the smallest singlet–triplet state energy gap (ΔE_ST), resulting in superior O₂˙⁻ generation efficiency in the order of TPPM < MeTPPM < MeOTPPM, as confirmed by theoretical studies, solution-phase experiments, and in vitro evaluations. These findings demonstrate that electron-donating substituents enhance the ICT effect, favoring a type I process over type II, and thereby improving free radical ROS production. Similar advancements were achieved with the development of AIE PSs, including TNZPY, MTNZPy, TBZPy, and MTBZPy (Fig. 13b-ii).¹⁶⁹ Among these, MTNZPy exhibited the smallest ΔE_st (0.08 eV) with the ROS efficiency ranked as MTNZPy > TNZPY > MTBZPy > TBZPy. These results affirm that introducing strong electron-rich anionic–π⁺ units is a viable strategy for developing AIE-based type I PSs. Expanding on this concept, other AIE-based type I PSs (i.e., TBP-2) have been explored for diverse therapeutic applications (Fig. 13b-iii). Interestingly, TBP-2 has been extensively applied for inhibiting breast cancer metastasis,¹⁷⁰ combating multidrug-resistant bacteria,¹⁷¹ and targeted cancer therapy.¹⁷² Such innovations underscore the potential of anion–π⁺ systems in advancing AIE-based type I PDT, enabling multiple anion–π⁺-based type I PSs to be developed, as shown in Fig. 14.^173–177

5.5.3. Acceptor shielding strategy for type I AIE PSs. Constructing an effective type I PS for PDT based on AIE requires meeting several critical conditions: First, the PS itself must exhibit strong light absorption, characterized by a high molar extinction coefficient (ε) and absorption in the longer wavelength region, allowing efficient transformation of excitation energy to the T₁ state. Second, a reduced ΔE_ST is essential for facilitating adequate triplet state production. This is achieved by employing electron-rich donors and acceptors to induce a strong ICT effect. Third, the T₁ state should have a sufficiently longer lifespan to enable electron transitions from ¹PS* to ³PS*, leading to a subsequent generation of ˙PS⁻. Importantly, the PS* environment must be shielded to ensure rapid electron transitions while minimizing undesired side reactions during the photophysical process. The acceptor shielding strategy addresses these requirements by introducing alkyl chains to the acceptor moiety, effectively shielding the *PS during light exposure. This concept was implemented in the design of MAP18-C12, which meets these criteria effectively. An N-substituted pyrenyl group on the pyrrole ring provides strong molar absorptivity due to its large π-conjugated planar structure. Furthermore, the N,N′-dimethylaniline groups at the 2,5-positions of pyrrole act as electron donors, extending absorption/emission wavelengths. A twisted conformation between pyrrole and phenyl rings also minimizes π–π interactions in the aggregated state, enhancing AIE properties. Studies using chemical indicators (DCFA-DA and ABDA) and ESR confirm the generation of type I ROS species. Notably, the ˙OH radicals produced upon irradiation by MAP18-C12 NPs exhibit a dual therapeutic effect. These radicals not only target cancer cells but also play a pivotal role in modulating the tumor microenvironment. Specifically, the ˙OH radicals induce the transformation of protumoral M2 macrophages into M1 macrophages, thereby enhancing immunotherapy outcomes. This dual action leads to the synergistic display of type I PDT and immunotherapy activities, effectively demonstrated in cancer cells, 3D multicellular spheroids, and in vivo tumor models (Fig. 13c).¹⁷⁸ This study underscores the critical importance of incorporating shielding groups on acceptor moieties in the design of type I PSs based on AIE for therapeutic PDT, offering a robust strategy to enhance both the efficacy and versatility of such systems.

5.6. Other examples

This section highlights a few promising PSs selected based on their innovative approaches and significant interest in the field. These examples represent diverse strategies for achieving type I PDT, aiming to inspire further research and deepen understanding in this area.

A notable example is the use of semiconducting materials with an acceptor–donor–acceptor (A–D–A) configuration to develop a π-conjugated molecular system exhibiting both type I & II capabilities, as well as NIR-II emission properties. One such system, COi6-4Cl (Fig. 15a), incorporates electron-rich oxygen atoms in the donor core to enhance electron-donating ability, linked to dicyanovinylindanone as the acceptor moiety with two chloride functional groups.¹⁷⁹ This structure results in a strong intramolecular charge transfer effect and a low band gap (1.27 eV). Further, COi6-4Cl demonstrates the ability to produce both ¹O₂ and ˙OH radicals due to small ΔE_ST (0.27 eV) and improved k_ISC, confirmed by solution tests. Theoretical studies revealed that the bridged molecular structure suppresses non-radiative decay by reducing molecular vibrations, while bulky side alkyl chains minimize π–π stacking, avoiding ACQ effects. For enhanced biocompatibility, COi6-4Cl was encapsulated into polystyrene5000-b-poly(ethylene glycol)5000 (PS-b-PEG), forming spherical nanoparticles with significant colloidal stability (up to 50 days). These nanoparticles exhibited concentration-dependent cytotoxicity against cancer cells under both hypoxic and normoxic conditions. The NIR-II imaging capability of COi6-4Cl nanoparticles enables deep tissue penetration and accurate discrimination between normal and cancerous tissues in tumor-bearing mouse models. As compared to chlorin e6 (Ce6) as a clinical PDT standard (660 nm laser irradiation), COi6-4Cl NPs (808 nm laser irradiation) showed significant anti-tumor efficacy.

Fig. 15 Schematic illustration of chemical structures of representative semiconducting materials serving as type I PSs for PDT. (a) Chemical structure of the π-conjugated molecule COi6-4Cl. (b) Chemical structure of PTS, PTSe, and PTTe used as NIR-II type I PDT/PTT PSs.

In recent years, interest in NIR-II-based PDT/PTT agents has surged due to their advantages, such as deep tissue penetration (>1 cm) and higher maximum permissible exposure limits (1.0 W cm⁻² for 1064 nm and 0.33 W cm⁻² for 808 nm). These properties hold promise for treating large, deeply infiltrated, or surgically inoperable malignant tumors while sparing patients from invasive procedures or radiotherapy. However, developing type I-based NIR-II PSs remains a challenge. Key hurdles include improving the ISC rate constant to generate triplet excitons efficiently and enabling electron transfer from molecular oxygen to generate O₂˙⁻ regardless of reducing intermediates. Additionally, achieving low Gibbs free energy (ΔG < 0) is crucial for producing H₂O₂ and ˙OH through superoxide disproportionation, while type II PDT requires that the T₁ state energy exceeds the oxygen sensitization threshold (0.98 eV). In general, type II processes tend to proceed at a faster rate compared to type I processes. To address this, a D–A strategy was employed to design three narrow-bandgap semiconducting materials, namely, PTS, PTSe, and PTTe (Fig. 15b).¹⁸⁰ These materials consist of thienoisoindigo (as an electron donor) and thiophene, selenophene, or tellurophene (as an electron donor) respectively. This approach enabled the achievement of NIR-II absorption and enhanced K_ISC to generate the triplet excitons (heavy atom effect, Se/Te). Additionally, these structural adjustments facilitated the type I process over the type II process by tunning the Gibbs free energy value between PTS, PTSe, PTTe, and ³O₂ to values below zero. This optimization enhanced the efficiency of type I PDT. The biocompatible NPs of PTTe NPs prepared using the nanoprecipitation method were validated for NIR-II (1064 nm) type I PDT/PTT applications. These validations included in vitro and in vivo models under both normoxic and hypoxic conditions, showcasing the therapeutic potential of PTTeNPs. The results, depicted in Fig. 15, emphasize the advantage of combining type I PDT and PTT capabilities in a single system for effective cancer treatment.

Another paradigm of type I PSs stems from the design of cyanine- and squaraine-based dyes. For instance, Li et al. developed an anionic pentamethine cyanine PS (CST-Pco)¹⁸¹ through a counterion engineering strategy to address the limitations of ACQ and low ROS efficiency in traditional cyanine dyes (Fig. 16). By incorporating a triphenylphosphonium cation (Pco) with oligoethylene glycol chains as the counterion, the authors enhanced dye–counterion interactions and optimized amphiphilicity, effectively suppressing excessive aggregation while preserving molecular symmetry and photophysical properties. In the aggregated state, CST-Pco exhibited strong NIR-II fluorescence emission (quantum yield up to 3.8%) and efficient type I ROS (e.g., O₂˙⁻) generation. Dual excitation wavelength modulation (760 nm for ROS production vs. 808 nm for fluorescence imaging) enabled dynamic theranostic switching. Formulated nanoparticles demonstrated enhanced tumor-targeting capability, achieving >6 mm imaging depth in vivo with a tumor-to-background signal ratio 7.6-fold higher than indocyanine green (ICG). Three PDT treatments yielded a 93.5% tumor suppression rate without systemic toxicity.

Fig. 16 Schematic illustration of chemical structures of CST-Pco and calculated aggregation behavior of C5T-Pco under nanoconfinement conditions based on MD simulations.¹⁸¹

In parallel, to challenge the conventional view that H-aggregation quenches photosensitization, a squaraine (SQ)-based supramolecular photosensitizer (TPE-SQ7)¹⁸² with ordered H-aggregates was designed (Fig. 17). By integrating tetraphenylethylene (TPE) and sulfonate groups, the H-aggregates promoted ISC and narrowed the singlet–triplet energy gap (ΔE_ST), leading to enhanced type I ˙OH generation while maintaining a high photothermal conversion efficiency of 54.2%. TPE-SQ7 NPs demonstrated potent cytotoxicity under both normoxia and hypoxia (2% O₂), achieving a 94.4% tumor inhibition rate after three type I PDT/PTT treatments. Additionally, a broad-spectrum antibacterial efficacy was also seen in TPE-SQ7 NPs, exhibiting >99% antibacterial efficacy against Staphylococcus aureus and Escherichia coli. This study redefines the role of H-aggregates in photodynamic processes, offering a dual-functional platform for hypoxia-tolerant cancer therapy and antimicrobial applications.

Fig. 17 Schematic illustration of molecular engineering of SQ-based PSs.

Interestingly, Peng et al. recently introduced a new type I PDT modality by integrating PDT with photocatalysis – offering a promising solution to the persistent challenge of drug resistance in tumor therapy. In this system, a julolidine-modified crystal violet (CVJ) was developed as a type I PS.¹⁸³ This molecular modification effectively mitigated the issue of intramolecular rotation-induced non-radiative decay, a key limitation that hampers ROS generation in conventional CV. In CVJ, the enhanced torsional strain reduces nonradiative transitions in highly viscous environments, thereby boosting type I ROS production. Further, Langlois’ reagent (CF₃SO₂Na) was employed, which under photocatalytic activation produces ˙CF₃ and SO₂. The ˙CF₃ radicals exhibit superior oxidative potential compared to conventional ROS, while SO₂ serves to deplete intracellular GSH, effectively disrupting redox homeostasis and inhibiting PS efflux from cancer cells (Fig. 18). Incorporation of this multifunctional system into biocompatible NPs demonstrated potent anticancer activity in both in vitro and in vivo models of drug-resistant tumors. This pioneering study not only underscores the therapeutic potential of ˙CF₃-based oxidative stress in cancer therapy but also introduces photocatalysis-induced ferroptosis as an innovative and mechanistically distinct strategy for overcoming multidrug resistance.

Fig. 18 Schematic illustration of CF₃-CVJ NPs preparation and underlying photocatalysis-based ferroptosis mechanism for drug-resistant cancer treatment. (a) Chemical structures of crystal violet, julolidine-based crystal violet, and its photocatalytic process. (b) Preparation of CF₃-CVJ NPs by the reverse evaporation method. (c) Mechanism of ferroptosis-based cell death triggered by CF₃-CVJ NPs under irradiation. Adapted with permission from ref. 183. Copyright 2022 Elsevier B.V.

While organic small molecular-based PSs typically function as type II PDT agents, their supramolecular assemblies can transition to type I PSs with appropriate structural modifications. For instance, the self-assembly of phthalocyanines bearing electron-donating amine groups favors type I ROS generation over type II activity.¹²² The electron-donating amine groups in proximity to the triplet state enable rapid reductive quenching, forming radical anions, which then transfer electrons to molecular oxygen and nearby substrates. However, introducing electron-donating substituents into organic scaffolds can be challenging. Kida et al. addressed this by developing supramolecular PS by using a charge-separated state (CS) of monomeric PS FI-C2 and FI-C18 (Fig. 19).¹⁸⁴ Under the CS state, electron transfer is stabilized over the triplet state, effectively suppressing type II ROS generation and favoring type I ROS production. Fluorescein, a conventional type II PS in its monomeric state, demonstrated photosensitization switching to type I activity upon self-assembly. Electrochemical and spectroscopic studies confirmed the CS state under irradiation, and ROS indicators (DHR123 and ABDA) alongside ESR experiments validated type I ROS generation. In cancer cells, the NPs demonstrated light intensity dependent type I PDT activity. This investigation offers a novel strategy for developing type I PSs in monomeric organic systems and supramolecular assemblies, expanding the toolkit for photodynamic cancer therapies.

Fig. 19 Schematic illustration of the assembly process of PSs. (a) Chemical structures of FI-C2 and FI-C18 PSs. (b) Schematic illustration of FI-C2 mediated type II and (c) FI-C18 mediated type I photosensitization mechanisms, respectively. Please note that the radiative decay forms of the excited states are omitted for better clarity.

Despite the remarkable strides made in type I PDT research, its therapeutic efficacy still faces significant challenges. One major limitation lies in the reliance of most reported type I nanosensitizers on the passive targeting mechanisms, which often result in suboptimal drug accumulation within tumor tissues, falling short of therapeutic expectations. To address this, researchers have explored advanced delivery strategies, such as tumor-selective antibodies, ligand modifications, and tumor-associated factor activation. However, these promising active targeting techniques have yet to see widespread implementation in type I nanosensitizer systems. In the latest research, Song et al. synthesized a novel amphiphilic PS (PS-02)¹⁸⁵ that combines thermally activated delayed fluorescence (TADF) with enhanced type I photosensitization capabilities (Fig. 20a). This design leverages a piperazine unit (6-NS) as an electron donor, covalently attached to the PS to create a localized “electron-rich environment”. The electron-deficient ligand 6-NS acts as an “electron transfer cage”, effectively suppressing the unwanted electron transfer (ET) process. Upon precise targeting and binding to the carbonic anhydrase IX (CAIX), the inherent electron-rich property of CAIX “unlocks” the ET effect, dramatically boosting type I photosensitization. Remarkably, this system achieves a 10.7-fold increase in the production of O₂˙⁻ compared to unmodified PS NPs. Further advancing the field, the team also developed PS@BSA, a novel type I PS integrating a fluorescein derivative with TADF properties into bovine serum albumin (BSA) (Fig. 20b).¹⁸⁶ This ingenious system harnesses the fluorescein derivative as an “electron pump” and BSA as an “electron reservoir”, thanks to its electron-rich amino acid composition. The PS “electron pump” draws electrons from BSA and transfers them to ³O₂, while BSA continuously replenishes electrons, driving efficient O₂ generation. This synergistic mechanism not only accelerates type I PDT processes but also provides new perspectives and a robust theoretical foundation for designing next-generation type I PSs.

Fig. 20 Schematic illustration of fluorescein derivatives thermally activated delayed fluorescence (TADF) type I PS. (a) Fluorescein derivatives PS-02 for promoting type I ROS. (b) Fluorescein derivatives PS PS@BSA as an “electron pump” and BSA as an “electron reservoir” for promoting type I photosensitive process.

6. Is O₂-independent photoredox catalysis a type I PDT?

Recent developments in type I PDT have introduced exciting new paradigms, such as the “photocatalytic oxidation of NADH in cells”, which has inspired the development of a variety of novel agents by research groups worldwide.^{52–55,187–189} In this section, we aim to highlight these significant advancements and emerging subdisciplines, to use cross-disciplinary insights to further accelerate the evolution of type I PDT.

In conventional type I PDT, photosensitizers (PSs) absorb light, which triggers an electron transfer process. In this process, the excited-state PSs (i.e., [PS*]) typically donate an electron to oxygen, leading to the generation of O₂˙⁻. However, this electron transfer is generally irreversible, meaning that PSs used in type I PDT are non-recyclable, resulting in the loss of PSs over time and diminishing the therapeutic efficacy of PDT. Fortunately, within living cells, numerous electron-rich substrates—such as proteins, nucleic acids, and small molecules (e.g., NADH, Fig. 21)—are readily available to donate electrons to exited PSs, even under mild reaction conditions (e.g., physiological environments). This interaction allows type I PSs to function like photocatalysts in the electron transfer process, making them recyclable. This catalytic nature opens up new avenues for developing alternative mechanisms of action in cancer phototherapy.

Fig. 21 Schematic illustration of the principle of PSs for photoredox catalysis of NADH in cells.

The process of utilizing transition metal complexes for photocatalytic oxidation of NADH is increasingly becoming a research focus. For example, modification to Ir(III) complexes with quinoline has led to the development of photocatalysts (QAIC) that can monitor real-time dynamic changes in NADH and trigger PDT through NADH depletion (Fig. 22a).¹⁹⁰ Furthermore, metal complexes substituted with Ru(II), such as chloromethyl-modified Ru(II) complexes (RuNH₂), have also been employed in photocatalytic NADH conversion, targeting mitochondria (Fig. 22b).¹⁹¹ These innovative designs enable the photocatalytic conversion of biomolecules while simultaneously generating O₂˙⁻ and regenerating the PSs (acting as photocatalysts), suggesting that the photocatalytic redox imbalance of biomolecules also serves as a potential antitumor mechanism of action in type I PDT.

Fig. 22 Schematic illustration of the chemical structure of transition metal complexes: (a) Ir and (b) Ru.

In contrast to the above-mentioned paradigms, a particularly impressive mechanism of action, namely O₂-independent photoredox catalysis in cells, has emerged recently. In 2019, Sadler and his colleagues made a groundbreaking contribution by reporting an intracellular photoredox catalysis mechanism with an O₂-independent mechanism of action, which could effectively ablate cancer cells under hypoxic conditions.⁵² Under light excitation, the transition metal catalyst, [Ir(ttpy)(pq)Cl]PF₆ (i.e., complex 1, Fig. 22a), not only catalyzed the oxidation of the cellular coenzyme NADH to form NAD⁺ but also drove the reduction of cytochrome C (Cyt c, Fe³⁺) to Cyt c (Fe²⁺) in an O₂-independent manner. It is well-established that NADH and Cyt c (Fe³⁺) are critical components in the electron transport chain (ETC) in mitochondria. Therefore, the photocatalytic redox imbalance of NADH/NAD⁺ and Cyt c (Fe³⁺)/Cyt c (Fe²⁺) disrupt energy metabolism and cause an imbalance of redox homeostasis in cancer cells. Motivated by these findings, our group further proposed a conditionally activatable photoredox catalysis (ConAPC) concept in 2021, wherein the inherent photocatalytic properties could be selectively activated by enzymes in cells.¹⁸⁷ In our work, we showed that a selenium-substituted Nile blue analog (Se–NH₂), a metal-free agent, is an effective NIR triplet state photocatalyst that could reach photooxidation of NADH to NAD⁺ and further induce the reduction of Cyt c (Fe³⁺), as shown in Fig. 23. By introducing a nitroreductase-responsive group, Se–NH₂ was further expanded into Se–NO₂, which is capable of both enzyme-responsive and light-activated. This system enabled high spatiotemporal resolution for selective fluorescence labeling of hypoxic tumor cells and cascade regulation of electron flow in the mitochondrial ETC.

Fig. 23 Schematic illustration of O₂-independent photoredox catalysis process. (a) Plausible tandem photocatalytic conversion of cellular electron source NADH and hemoprotein Cyt c via an O₂-independent pathway. (b) Chemical structure of newly developed NIR photoredox catalyst Se–NH₂ and its advantages. The optimized molecular conformation between Se–NH₂ and NADH is also shown. (c) Conditional activatable photoredox catalysis (ConAPC) for spatial-temporal control of photocatalytic activities in living systems. Abbreviations: TOF, turnover frequency; TON, turnover number; SET, singlet electron transfer. Adapted with permission from ref. 187. Copyright 2021 American Chemical Society.

Inspired by these pioneering works, more and more researchers are focusing on exploring O₂-independent modalities within the domain of PDT, such as Yao et al., who proposed O₂-independent RNA degradation via excitation energy transfer between Se-substituted Nile blue PSs and RNA molecules (also defined as type III PDT).¹⁹² However, this raises a pivotal question: can O₂-independent photoredox catalysis truly be classified as a form of PDT? In recent years, several definitions and research findings have contributed to the growing body of literature in this area. Through extensive discussions with prominent photochemists worldwide, a consensus has gradually emerged: photoredox catalysis, although it shares certain similarities with PDT, should not be strictly classified as PDT. PDT inherently involves three key components—O₂, PS, and light. Without the participation of O₂ in the process, it is unreasonable to categorize any phototherapeutic modality as PDT. This perspective fundamentally contradicts the essence of PDT in terms of its definition. Instead, a more accurate term for this approach would be “photocatalytic therapy”, which better captures the unique characteristics and mechanistic details.

7. Conclusion

The remarkable advancements in PDT over the last decade, particularly in the domain of type I PDT, have ushered in significant breakthroughs for the treatment of malignant tumors. Innovations in developing novel PSs to overcome the challenges posed by hypoxic tumor microenvironments, as well as emerging strategies targeting cellular organelles to enhance therapeutic efficacy, have not only challenged longstanding assumptions in biology and medicine but have also provided fresh perspectives and methodologies for tumor treatment. Researchers globally have been tackling the issue of tumor hypoxia by developing various PSs for type I PDT. However, despite these promising advancements, clear guidelines for designing effective type I PSs remain elusive.

Drawing from our research group's extensive experience in the field, we aim to offer a coherent design framework for both current and future researchers working on type I PDT. This review aspires to serve as a comprehensive “guidebook”, systematically organizing and defining key aspects such as the definition, mechanisms, detection methods, and applications of type I PDT. The ultimate goal is to provide researchers with an accessible and practical resource that will further propel advancements in PDT and encourage the innovative design and application of type I PSs.

However, several urgent challenges remain that need to be addressed:

(1) Designing exclusive type I PSs from a molecular perspective remains a complex task. Despite significant research, the scientific community has yet to establish a clear and general structure–activity relationship for type I PSs, making it difficult to predict the photoreactions based solely on the molecular structure and modifications of specific scaffolds. This complexity arises from the need to understand the multidimensional interplay between factors such as molecular architecture (e.g., electron donor–acceptor strength), excited-state dynamics, and the surrounding microenvironment. However, emerging computational approaches—particularly time-dependent density functional theory (TD-DFT) and machine learning algorithms—show promise in deciphering these relationships, enabling predictive modeling of electron transfer pathways and facilitating the targeted design of type I-dominant PSs.

(2) Standardizing strategies for calculating the yield of O₂˙⁻ is crucial. Current detection methods, such as ESR or fluorescent probes, provide qualitative assessments of type I ROS generation but lack quantitative precision.

(3) Oxygen dependence continues to be an issue, despite the partial oxygen cycling capabilities of many type I PSs. While some PSs can reduce their oxygen dependency, oxygen is still a vital component in photosensitive reactions.

(4) Translating in vitro and in vivo findings into clinical applications is challenging. The absence of a clear dose–response relationship remains a major obstacle to the clinical adoption and widespread use of type I PDT.

(5) Light source optimization plays a pivotal role in enhancing PDT outcomes, given its centrality in biomedical optical technology. Developing a light source that allows precise spatiotemporal control while minimizing side effects is essential for improving PDT's therapeutic efficacy. One promising avenue could be constructing a light transmission system using functionalized LED beads. This could represent an integrated approach to optimize future research and applications. Additionally, creating small, flexible alternating current electroluminescent (ACEL) devices that combine PSs and light sources into therapeutic patches capable of simultaneously performing photodiagnosis and PDT could further boost the effectiveness of type I PDT.

(6) Accurately assessing the therapeutic effects of type I PDT and optimizing treatment plans is another key challenge. This requires a holistic evaluation of factors such as tumor type, stage, size, and PS characteristics, alongside the parameters of the light source. A personalized treatment plan must integrate these variables for maximum impact. Although type I PDT can be combined with other therapeutic modalities (e.g., radiotherapy and chemotherapy) to enhance efficacy, determining the most effective combination and sequence of treatments, as well as evaluating the synergistic effects, still demands further investigation.

(7) Establishing clear design principles for PSs compatible with clinical workflows is important. Several critical factors should be prioritized in type I PS development: (i) biocompatibility and rapid clearance: emphasizing PSs with reduced systemic toxicity and short half-lives to minimize photosensitivity side effects; (ii) tumor microenvironment responsiveness: engineering PSs that respond to specific tumor hallmarks—such as hypoxia, acidosis, or elevated enzyme levels. This can significantly enhance therapeutic selectivity and minimize off-target effects; (iii) scalable synthesis: prioritizing cost-effective, reproducible synthesis routes to facilitate large-scale production for clinical use; (iv) multimodal integration: designing PSs compatible with imaging modalities (e.g., fluorescence-guided surgery) to streamline clinical workflows.

In conclusion, type I PDT has significantly expanded the spectrum of available therapeutic options within the complex physiological context of cancer treatment. Ongoing research and the development of both current and future PSs hold great promise, offering the potential to deepen our understanding of biological systems. This deeper insight not only paves the way for a more thorough understanding of cancer treatment mechanisms but also drives the development of more efficient and targeted PSs, especially those capable of precise tumor targeting and self-degradation. As technological innovation continues to advance at a rapid pace, there is a strong belief that type I PDT is poised for a bright future in providing precise, safe, and effective treatments.

Data availability

No primary research results, software or code have been included and no new data were generated or analysed as part of this review.

Conflicts of interest

There are no conflicts to declare.

Acknowledgements

M. L. wishes to thank the support of the National Natural Science Foundation of China (Grant No. 22308220), Shenzhen University Third-Phase Project of Constructing High-Level University (Grant No. 000001032104), the Research Team Cultivation Program of Shenzhen University (Grant No. 2023QNT005), and the Guangdong Province Key Areas Special Project for Regular Colleges and Universities (Grant No. 2024ZDZX2018). X. C. is thankful for the support of the National Natural Science Foundation of China (Grant No. 22090011), Shenzhen University 2035 Program for Excellent Research (Grant No. 00000208), Guangdong Basic and Applied Basic Research Foundation (Grant No. 2023B1515120001), and the Shenzhen Science and Technology Program (Grant No. RCBS20231211090515015). X. P. wants to thank the support of the Shenzhen University 2035 Program for Excellent Research (Grant No. 00000225). J. S. K is thankful for the National Research Foundation of Korea (2018R1A3B1052702). This work was also supported by the National Research Foundation of Korea (Grant No. 2018R1A3B1052702, J. S. K.; RS-2023-00241100, Y. X.). The authors also gratefully thank the support from the National Natural Science Foundation of China (Grant No. 82203050, Y. X.).

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Footnote

† Equally contributed.

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