4.2.1 Noble metal nanoparticles and metal oxides

Noble metal (mainly Au and Ag) nanoparticles exhibit localized surface plasmon resonance properties. These enable rapid and efficient energy transfer from a metal surface to molecular O₂ and the formation of ¹O₂, which is a typical type-II PDT process.^106-108 The tunable light scattering and absorption of metal nanoparticles enable tuning of the excitation wavelength to the NIR region to improve the tissue-penetration depth of the laser, thereby improving its efficacy in mediating the PDT of tumors in deeper tissues.¹⁰⁹ Vankayala et al.¹⁰⁶ reported that gold nanorods (Au NRs) under NIR light excitation could effectively generate ¹O₂, thus promoting the death of B16F0 melanoma tumors in mice. By varying the excitation wavelength, it is possible to control whether Au NRs produce ¹O₂ or heat, under light irradiation, resulting in tumor cell death by PDT or PTT, respectively (Figure 2A).

Metal oxide nanoparticles, including TiO₂, ZnO, Fe₂O₃, and so on.^109-111 TiO₂ exhibits photocatalytic ability. When TiO₂ is irradiated with ultraviolet (UV) light, electrons are excited from the valence band to the conduction band, producing negative electrons (e⁻) while leaving positively charged holes (h⁺) in the valence band. These free electrons react with O₂ and water molecules in the environment to form O₂^•−, H₂O₂, and •OH, which in turn react with tumor cell components, leading to apoptosis or necrosis.¹¹² Wang et al.¹¹³ reported an injectable thermosensitive hydrogel (BT-CTS thermogel) containing magnesium for local tumor treatment and the promotion of wound healing. Magnesium thermal reduction was used to produce structurally defective nanosized black titania (B-TiO₂-x) in TiO₂ nanocrystals. The presence of O₂ vacancies and disorder-induced lattice strain in B-TiO₂-x enables the light absorption of B-TiO₂-x to effectively extend into the visible and NIR regions and produce ROS and heat upon excitation by light. In the presence of B-TiO₂-x, which is rich in O₂ vacancies, the hydrogel showed the dual antitumor effects of PTT and PDT under NIR laser (808 nm) irradiation.

4.2.2 Noble metal complexes

In comparison with conventional organic PSs, noble metal complexes offer numerous advantages in the context of PDT applications. The incorporation of nonessential metals enables the utilization of diverse analytical techniques, such as X-ray absorption spectroscopy and inductively coupled plasma-mass spectrometry, facilitating quantitative analysis, and localization of the PSs within cellular and tissue environments. If the metal complex exhibits luminescent characteristics, its precise cellular localization can be further determined through confocal microscopy.¹¹⁴ More importantly, noble metal center complexes have received increasing interest as PSs because of their intrinsic heavy-atom effect, which facilitates strong spin-orbital coupling and enhances the rate of intersystem crossing. This phenomenon will prolong the time of the triplet excited state (T₁) to interact with dioxygen, biomolecular, or other O₂ substrates.¹¹⁵ Therefore, metal complexes offer a wider range of excited state electronic configurations that can be utilized in both O₂-dependent and nondependent cytotoxic pathways.^116-118

The typical metal complexes include Pd(II) complexes,¹¹⁴ Ru(II) complexes,¹¹⁹ Ir(III) complexes,⁹⁸ and Au(III) complexes.¹²⁰ Mazor et al.¹²¹ developed WST11 for vascular-targeted PDT and determined that the inclusion of a Pd center in WST11 enhances stability, excited state reactivity, and intersystem crossing rate of the PS. Fong et al.¹²² developed a Ru(II)-based agent TLD1433 that is currently undergoing clinical trials for intravesical application in nonmuscle invasive bladder cancers. TLD1433 demonstrated the ability to effectively engage in both type I (electron transfer) and type II (energy transfer) photoreactions, thereby transforming its excited state reactivity from single-linear O₂ sensitization in well-oxygenated environments to photo-oxidation–reduction reactions under anoxic conditions and displaying ¹O₂ quantum yields. Cole et al.¹²³ designed several families of Ru(II) bis-heteroleptic complexes. This new class of light-responsive, hypoxia-active agents incorporating the α-oligothienyl group might involve excited state pathways distinct from the ¹O₂ and photosubstitution pathways, as well as a novel mode of electron-transfer reaction specific to oligothiophenes. They exhibited phototherapeutic indices up to >500,000 in normoxia and >5800 in 1% O₂ hypoxia. Recently, Mani et al.¹²⁴ reported a series of Os(II) polypyridine complexes as PSs for PDT. As a result of the pronounced π-extended structure of the ligand and the heavy-atom effect imparted by the Os center, these complexes demonstrated that heightened absorption in the NIR region increased ¹O₂ quantum yields and shifted the maximum wavelength in comparison with their ruthenium analogues.

4.2.3 Carbon nanomaterials

Typical carbon nanomaterials include graphene quantum dot (GQD),^{73, 125} C₆₀,¹²⁶ and so on. Among them, the absorption spectrum of GQD spans the UV region and the entire visible region, which can be used as a PS for PDT to produce ¹O₂ via a multistate sensitization process (Figure 2B).¹²⁵ In addition, C₆₀ has unique photophysical properties. Under UV or visible-light irradiation, C₆₀ molecules can be transformed into excited trilinear states, producing single-linear states or other forms of ROS.¹²⁷ Furthermore, C₆₀ is known as a “free radical sponge” and can scavenge ROS by electron transfer in the absence of light, which has the potential to be applied to mitigate the damage caused by the local accumulation of ROS in biological tissues.¹²⁸

4.2.4 Graphene-like materials

Some graphene-like materials (e.g., black phosphorus (BP)) can be used as PSs for anticancer.¹²⁹ BP exhibits a thickness-dependent bandgap ranging from 0.3 eV in the parent material to 2.0 eV in the monolayer, resulting in tunable and broad optical absorption in the UV-to-visible range.¹³⁰ Wang et al.¹³¹ in 2015 first demonstrated that BP nanosheets can efficiently produce ¹O₂ with a quantum yield of approximately 0.91, which is higher than that of most reported PSs. In addition, BP-based nanomedicines have a better photothermal conversion efficiency. Zhang et al.¹³² found that BP-based nanodrugs can be used in combination with PTT and PDT in cancer treatment through the study of polyethylene glycol (PEG)ylated hyaluronic acid (HA)-modified BP (HA-BP) (Figure 2C). In addition, the phototherapy of HA-BP elicited ICD to mediate antitumor immunity by secreting damage-associated molecular patterns (DAMPs), including surface-exposed calreticulin (CRT), adenosine triphosphate (ATP), and high-mobility group protein B1 (HMGB1), to promote prominent dendritic cell maturation and then induce the activation of specific effector T cells (such as CD4⁺/CD8⁺ T cells) to further eliminate tumor cells.

4.3.1 Tetrapyrrole compounds

Tetrapyrrole-type PSs, which mainly include compounds such as porphyrins, phthalocyanines, chlorophylls, and bacteriochlorophylls, are the most commonly used PSs for anticancer therapy. In contrast to PSs with other structures, tetrapyrrole-type PSs (except bacteriochlorophylls) are primarily type II PSs that produce type II ¹O₂ rather than type I ROS.³³ Tetrapyrrole-type PSs are unique to PDT because of their high ¹O₂ generation efficiency and excellent fluorescence performance.¹³³ However, the lack of O₂ in the TME can seriously affect the therapeutic efficacy of type II PSs. Recently, scientists prepared type I PSs by modifying typical type II PSs to improve the anticancer effects of such PSs.¹³⁴ For example, Sun et al.¹³⁵ prepared porphyrin with O-linked cationic side chains that exhibited enhanced ROS generation in both type I and type II PDT pathways, and their PDT efficacy under hypoxic conditions was significant (Figure 3A).

4.3.2 Other organic dyes

Organic dyes, such as BODIPY, anthocyanine dyes, and phenothiazine dyes, can also be used as PSs.¹³⁶ Organic dyes such as carbocyanine iodide dyes, which are absorbed in the NIR region, can effectively convert NIR light into fluorescence, ROS, and heat, thus contributing to tumor extinction.¹³⁷ Yuan et al.¹³⁸ used mitochondria-targeting amphiphilic copolymers to load BODIPY (PBDPI-TPP) and generated ROS upon laser irradiation, impairing the biological function of organelles and causing apoptosis (Figure 3B). Schneider et al.¹³⁹ also demonstrated the advantageous properties of BODIPY-based agents in cancer therapy. These agents exhibited high photocytotoxicity and low dark toxicity, with a ratio of dark to light toxicity (phototoxic index) exceeding 830,000 and still exceeding 360,000 under low O₂ conditions.

4.3.3 CPs

CPs are a class of organic macromolecules characterized by a substantial π-conjugated backbone, light­harvesting capability, and efficient energy transfer.^{140, 141} The electronic conjugation between each repeating unit in their backbone generates a semiconductive “molecular wire,” allowing for efficient migration of excitons along the conjugated backbones upon excitation, thereby enhancing energy transfer efficiency to low-energy acceptors. Furthermore, it has been reported that encapsulating PSs with CPs into nanoparticles or covalently incorporating them into the main chain of CPs may further enhance their photosensitization efficiency.^142-144 Therefore, CPs-based PSs have great therapeutic potential for the treatment of malignant tumors. Xing et al.¹⁴⁵ reported in 2009 the first work utilizing the light-harvesting CP to amplify ROS generation. They synthesized anionic water-soluble polythiophene (PTP) and cationic porphyrin (TPPN) to form electrostatic complexes and undergo efficient energy transfer from PTP to TPPN under white light (400–800 nm) irradiation. The polymer PTP could transfer its excited state energy to TPPN via energy transfer, leading to enhanced intersystem crossing in the complex, and the production of TPPN's long-lived triplet state which further sensitized the ground state O₂ molecule to produce ¹O₂. Zhou et al.¹⁴⁶ employed a donor-acceptor strategy to design the backbone structure and synthesized a CP named PTDBD, incorporating electron-rich thiophene with electron-deficient benzothiadiazole and diketopyrrolopyrrole. When exposed to an 808 nm laser, PTDBD exhibited efficient conversion of light into heat while generating. Caverzán et al.¹⁴⁷ constructed CP-based nanoparticles and demonstrated their ability to generate ROS upon irradiation. Through in vitro and in vivo studies, it was found that the system was capable of inducing direct cancer apoptosis-involved cell death.

4.3.4 AIE molecules

Most organic PSs, particularly clinically approved porphyrins, have an intrinsic defect that leads to severe intermolecular π–π stacking at high concentrations or in the aggregated state leading to diminished or even complete loss of fluorescence, which is referred to as “Aggregation-Caused Quenching” (ACQ).¹⁴⁸ In contrast to conventional PSs, which show quenched fluorescence in the aggregated state and reduced single-linear O₂ production, AIE molecules can be excited by light in the aggregated state and emit strong fluorescence while efficiently producing reactive O₂ species. This new compound overcomes the problem of traditional PSs being prone to fluorescence burst and failure when molecules are aggregated, and thus, it has significant advantages in the field of PDT applications.^{149, 150}

AIE is a photophysical phenomenon discovered by Luo et al.¹⁵¹ in 2001. Owing to the restriction of the intramolecular motion (RIM) of the dye in the aggregated state, the nonradiative leap was suppressed, resulting in a significant enhancement of the fluorescence. Yuan et al.¹⁵² then developed a kind of AIE NPs with excellent fluorescence imaging and ROS generation capabilities in 2014. Based on the mode of ROS production, AIE-PSs are mainly classified as type I or type II, and they can generate ROS through both type I and type II pathways.¹⁵³ For example, Wang et al.¹⁵⁴ recently reported AIEgen Pys-QM-TT, which is capable of producing both type I and type II ROS and simultaneous NIR fluorescence imaging for the effective suppression of bacterial infection and ablation of tumor tissue (Figure 3C). In Pys-QM-TT, the strong electron-donating triphenylamine unit, π-bridge thiophene, and electron-withdrawing pyridinium salt unit can enhance the D-π-A behavior, thereby improving the intramolecular charge transfer effect and extending the emission wavelength. Simultaneously, the strong D-π-A effect should reduce π E_S-T and promote intersystem crossing processes, thereby increasing ROS production. In addition, the negatively charged anion in the pyridine salt moiety provides an electron-rich environment for the excited PS, thus promoting electron transfer to generate type I ROS. In general, the pathway of ROS production by AIEgens is strongly influenced by their structure.

4.3.5 Natural photoactive compounds derived from plants

With the development of separation and extraction techniques, numerous natural photoactive compounds have been isolated.¹⁵⁵ In recent years, many photoactive compounds such as polyacetylene, thiophene, and anthraquinones have been identified from plant extracts.^{156, 157} Some of these compounds are characterized by three-bonded carbon–carbon molecules and thiophene compounds. Typically, aliphatic compounds conjugated to three or more alkyne bonds are considered phototoxic in nature.¹⁵⁸ Polyacetylene and thiophene compounds have been reported to be activated or excited, with the polyacetylene compound producing ¹O₂ while the thiophene provides high optical yields, resulting in type II PDT reaction yields.^{159, 160}

Polycyclic quinones PSs are a class of natural PSs derived from mycorrhizal plants, mainly including bamboo red mycorrhizal, curcumin, and chrysin.¹⁶¹ Notably, chrysin can efficiently produce single-linear O₂ upon laser irradiation, increase superoxide dismutase activity, and decrease cellular glutathione levels.^{162, 163} PDT mediated by these compounds can inhibit the proliferation of a variety of tumor cells, including skin cancer,¹⁶⁴ head and neck squamous cell carcinoma,¹⁶⁵ breast cancer, and so on.¹⁶⁶ Due to its hydrophobic nature, chrysin predominantly localizes within lysosomal membranes as well as the Golgi apparatus, endoplasmic reticulum, and mitochondria. Moreover, it exhibits inhibitory effects on tumor cell growth and angiogenesis and induces apoptosis, necrosis, and autophagy of tumor cells.^{167, 168}

6.2.1 Harnessing the optical properties of nanomedicines

In recent years, UCNPs have emerged as a highly promising tool for cancer therapy. UCNPs usually consist of a crystalline matrix doped with lanthanide, transition metal, or actinide ions, which exhibit the unique ability to emit higher-energy photons by absorbing two or more low-energy NIR photons and converting them energetically.¹⁹⁰ This property allows UCNPs to emit light in the UV, visible, or NIR regions, facilitating ultrasensitive immunoassays, high-contrast imaging, and deep tissue imaging with minimal optical background. To enhance their therapeutic potential, PSs can be conjugated to UCNPs using various methods such as surface modification, chemical bonding, or encapsulation.¹⁹¹ These approaches leverage the upconversion luminescence properties of UCNPs and combine them with those of PSs, thereby improving drug delivery efficiency and therapeutic efficacy. Furthermore, the upconversion luminescence properties of UCNPs can enable precise tumor imaging and localization, providing accurate guidance for treatment. This integration of UCNPs and PSs holds great promise in advancing cancer therapy by offering enhanced imaging capabilities and improved treatment precision.¹⁹²

For example, Cui et al.¹⁹³ reported the application of UCNPs in PDT by developing a multifunctional PDT PS. UCNPs were first coated with a porous silica layer that contained high absorbance photosensitive molecules to match the emission spectrum of UCNPs. Specific antibodies were then covalently attached to the silica-coated nanoparticles to recognize specific antigens expressed on the surface of targeted cells. When irradiated with infrared light, the UCNPs emitted light that was absorbed by the photosensitive molecules on their surface. The excited photosensitive molecules interacted with the surrounding ground state molecular O₂ to generate singlet O₂, which led to oxidative damage of the cancer cells. A multifunctional nanostructure consisting of UCNPs and the PS zinc(II) phthalocyanine (ZnPc) has also been developed for the conversion of NIR light into visible light (Figures 5A and B). The surface of UCNPs was coated with folic acid-modified amphiphilic chitosan to anchor ZnPc close to the UCNPs, facilitating resonance energy transfer from UCNPs to ZnPc. In vivo anti-S180 tumor effect showed that this nanostructure-based NIR light-triggered PDT had significant therapeutic effects.

Two-photon absorption (TPA) is a nonlinear optical effect that offers several advantages over single-photon absorption, including the ability to achieve localized photoactivation in deep tissues using higher optical densities and longer wavelengths.³³ TPA can be used in PDT to selectively irradiate tissues at different depths by selecting the laser wavelength, and it can also improve the efficiency of PS activation and enhance the therapeutic effect.¹⁹⁴ Moreover, TPA can realize high-resolution 3D microscopic imaging in deep tissues due to its ability to achieve local excitation with a large laser beam diameter.

Currently, two-photon excited nanomedicines have been investigated based on various materials, including quantum dots,¹⁹⁵ Au nanomaterials,¹⁹⁶ silicon nanomaterials, carbon nanomaterials, and polymer nanomaterials. For instance, Secret et al.¹⁹⁷ developed a novel PS carrier mannitol-modified porous silica (pSiNPs). pSiNPs could absorb two-photons and act as an energy transfer donor to activate the PS by two-photons at 800 nm, thereby improving the therapeutic effect of PDT. The porphyrin loaded pSiNP nanodrug had a TPA cross section six times higher than that of porphyrin, which showed significant inhibitory effects on MCF-7 cells, suggesting that two-photon-excited nanodrugs could enhance the therapeutic effect of PDT.

Au nanoparticles are also being widely used in two-photon excited nanomedicines due to their excellent light scattering and absorbing properties. Their extremely high cross section or TPA cross section under two-photon excitation allows for deeper tissue penetration and more precise control compared with conventional single-photon absorption.¹⁹⁸ Chen et al.¹⁹⁹ conducted a study focusing on the enhancement of PDT efficacy using a nanocomposite composed of NaYF4:Yb/Er UCNPs conjugated with Au NRs. To achieve plasma-enhanced PDT, methylene blue was encapsulated within silica shells. The UCPs functioned as photoconverters, converting NIR light to visible light to excite methylene blue and generate ROS. The presence of Au NRs contributed to enhance upconversion efficiency and ROS production through the localized surface plasmon resonance effect. The researchers investigated the optimization of methylene blue loading, ROS generation capacity, and effective distance for plasma-enhanced ROS generation by adjusting the thickness of the silica shell. The mechanism of plasma-enhanced PDT was further elucidated by enhancing the upconversion luminescence intensity through the plasma field, thereby improving the light-harvesting ability and absorption cross-section of the system (Figures 5C and D). Comparative studies involving different surface plasmon resonance bands of Au nanoparticles confirmed the improved ROS generation. In vitro and in vivo experiments demonstrated that the nanocomposite exhibited substantial ROS generation and efficient PDT treatment.

6.2.2 Conversion of X-rays into visible light using nanoscintillators

To harness the tissue-penetrating power of X-rays for PDT, researchers often use scintillators with wide energy bandgaps to absorb X-rays and convert them into UV or visible light, which then excite PSs to produce ROS and achieve therapeutic effects. In a recent study, Chen and colleagues proposed a novel nanomaterial for X-ray-based PDT.²⁰⁰ This material was composed of a silicate nanoscintillator containing zinc, manganese, and bengal red (Figure 6A). After incubating these nanoparticles with U87MG cells, significant cancer cell death was observed only under X-ray irradiation. The researchers then intravenously injected the nanoparticles into tumor-bearing BALB/c nude mice and irradiated them with X-rays, leading to a remarkable tumor suppression rate of 98.1% compared with the control group. Notably, this silicate nanoscintillator could accumulate in tumors and significantly inhibit tumor progression at low doses of X-ray irradiation with minimal effects on normal tissues (Figure 6B). These findings suggest the potential of X-ray-based PDT in cancer treatment.

6.2.3 High-speed charged particles activate PSs

In 1934, the Soviet physicist Pavel A. Cherenkov (1904–1990) made a remarkable discovery, observing that high-velocity charged particles traveling through a medium emitted a faint blue visible light. This phenomenon, known as Cherenkov radiation, can be attributed to various processes such as external light sources, Compton γ-ray scattering, or the photoelectric effect. Cherenkov radiation has proven to be an effective approach for realizing free-electron laser light sources. Inspired by this, researchers have explored the potential of utilizing the visible light emitted when high-velocity charged particles traverse a medium to activate PSs in the surrounding area. In a study, Tang's team developed an isoquinoline organic salt derivative, TIdBO, based on a tetraphenylethylene backbone as a PS (Figure 6C). Under continuous light, small molecules underwent a photocyclization reaction via electron transfer involving free radicals. However, in aggregated form, the photocyclization reaction was suppressed, resulting in a higher proportion of type I ROS products. This demonstrated the feasibility of using molecular aggregation to regulate the competition between the two processes and enhance type I ROS production. Notably, TIdBO not only showed good PDT performance during the interaction with HeLa cells but also achieved self-monitoring of the PDT process through the relationship between the increase in fluorescence intensity and the change in cell morphology as an indicator of apoptosis.²⁰¹

6.2.4 Bioluminescence light source to activate PSs

Conventional PDT uses external light to irradiate tissues, which face the problems of poor penetration and rapid attenuation. To overcome these problems, researchers are actively exploring internal light sources for self-luminescence, including chemiluminescence and bioluminescence. However, the intensity and area of self-luminescence are difficult to control, which limits its clinical application. Therefore, it is important to develop a highly efficient and spatiotemporally controllable self-luminous PDT system. Tang's team reported a PDT system driven by electrochemiluminescence (ECL). The luminescence generated by [Ru(bpy)₃]²⁺ and the coreactant tripropylamine pair was both an optical readout for ECL imaging and a light source for excitation of the PS chlorin e6 (Ce6). This system relied on the effective energy transfer from ECL emission to the PS, which excited the surrounding O₂ for PDT. The dynamic processes of gradual morphological changes, changes in cell-matrix adhesion, and increases in cell membrane permeability during ECL-PDT were monitored with good spatial and temporal resolution under ECL microscopy (Figure 6D). It was expected to assist tumor treatment and imaging to reach deeper tumor sites for subsequent treatment.²⁰²

6.3.1 Passive targeting of PSs to tumors

The traditional enhanced permeability and retention (EPR) effect allows substances to preferentially infiltrate and remain in tumor tissues compared with normal tissues.²⁰³ This effect arises from the rapid and disorderly proliferation of tumor vessels, leading to structurally incomplete vessels with large gaps, and the lack of lymphatic vessels further contributes to the EPR effect. This phenomenon facilitates the accumulation of nanoparticles in tumor tissues for passive targeted therapy.²⁰⁴ To optimize the passive targeting efficiency, nanoparticles’ physicochemical properties, including size, charge, and shape, should be optimized. Strategies to enhance the EPR effect also include using external physical stimuli (e.g., radiation or heat) to temporarily increase tumor tissue permeability or employing drug administration to disrupt the TME and enhance nanoparticle accumulation. For instance, Gao et al.²⁰⁵ developed polymeric micelles containing the chemotherapeutic agent doxorubicin (DOX) and PS ZnPc. These micelles were self-assembled from amphiphilic block copolymers of methoxy-PEG and poly(β-benzyl-l-aspartate), which were expected to accumulate in tumor tissue through the EPR effect. The photoactivity of ZnPc was initially inhibited within the micelles due to self-aggregation. After internalization by tumor cells, the acidic and reducing intracellular conditions would trigger the release of both DOX and ZnPc, resulting in dual therapeutic effects (Figure 7A). To investigate the biodistribution of the DOX–ZnPc micelles, nude mice with HepG2 tumors were treated with an intravenous dose of these micelles. Fluorescence imaging revealed minimal fluorescence in the first 8 h, indicating ZnPc remained within the micelles with strong self-quenching. Over the next 7 days, fluorescence gradually increased, demonstrating preferential accumulation of DOX–ZnPc micelles at tumor sites through the EPR effect and successful release of ZnPc at the target site. Moreover, Ikeda-Imafuku et al.²⁰⁶ devised a potent anticancer strategy in which DOX was conjugated to PEG through a thioketal linker sensitive to ROS. The obtained amphiphilic PEG–DOX conjugate was used as the nanoparticle carrier for hydrophobic PS pheophorbide A. This system harnessed the EPR effect to augment nanoparticle accumulation within tumor tissues. This treatment approach not only activated pheophorbide A to generate cytotoxic ROS, but also initiated an ROS cascade, inducing structural disruption of the nanoparticles and facilitating the accelerated liberation of DOX and pheophorbide A. As a result, this approach significantly amplified the therapeutic efficacy of the anticancer treatment.

6.3.2 Active targeting of PSs to tumors

Recent investigations have unveiled that nanoparticles can primarily infiltrate into tumors via receptor-mediated cytosolic transport. Therefore, tailoring nanoparticles for binding to specific receptors on target cell surfaces has shown considerable promise in enhancing targeting precision. Consequently, it may be more productive to pivot research emphasis from EPR-mediated passive targeting to the strategic design of actively targeted nanotherapeutics. These systems can elevate drug concentrations within target cells, facilitated by the interaction between surface-bound ligands and membrane receptors on the target cells. A variety of ligands, including antibodies, antibody fragments, or short peptides, have been integrated into nanocarriers with remarkable specificity and affinity. For example, Yan et al.²⁰⁷ developed an epidermal growth factor receptor (EGFR)-targeted hyperpH-sensitive nano-PS and systematically evaluated its active targeting ability and therapeutic efficacy after modulation of the tumor vasculature system and stromal barrier (Figure 7B). EGFR, one of the most common targets for cancer therapy, has been reported to be overexpressed on the surface of a variety of cancer cells. And the Fab' fragment of Erbitux, a clinically used antibody against EGFR, was coupled to the nano-PS as a targeting moiety. The study demonstrated that antibody-modified nano-PSs exhibited specific targeting ability and superior cytotoxicity in vitro compared with their nontargeted counterparts. However, in vivo tumor targeting and inhibition were compromised due to stromal cell accumulation and nonspecific isolation. To improve the in vivo behavior of antibody-modified nano-PSs, the TME was perturbed (Figure 7C). The synergistic enhancement of tumor accumulation and targeting ability of antibody-modified nano-PSs was achieved by sequential modulation of the TME using thalidomide and PDT pretreatment, further highlighting the superiority of the active targeting strategy.

Although this active targeting approach allows for targeted delivery of PSs, incidental uptake of these couplers by normal cells is still unavoidable because most cancer-associated receptors are not exclusively expressed in cancer cells. To circumvent these problems, dual receptor targeting strategies have been explored to facilitate targeted delivery of photosensitizing drugs. Since cancer cells often overexpress more than one surface receptor, this dual-receptor targeting strategy can increase the likelihood of ligand–receptor interactions, resulting in better uptake and more precise targeting of cancer cells compared with single-receptor targeting approaches. Ng's group has devised a novel strategy for activating such a PS specifically in the target cells via dual receptor-mediated bioorthogonal coupling. This system consists of two components, a biotin-modified tetrazine-substituted PS and a cyclic EGFR-targeting peptide linked with a bicycle[6.1.0]non-4-yne dienophile, which are preferentially internalized only by cancer cells expressing both biotin receptors and EGFR, followed by a rapid bio-orthogonal inverse electron-demand Diels-Alder reaction to form the corresponding cycloadduct, thereby restoring the PS's ability to fluoresce and generate ROS.²⁰⁸

Due to the good in vitro targeting behavior of actively targeted nanomedicines, their related studies are increasing year by year. However, in vivo, the complex TME often affects the targeting effect of actively targeted nanoparticles. It was reported that after intravenous injection of nanoparticles, only a very small amount of particles can be delivered to solid tumors by the EPR effect.²⁰⁹ In addition, after leaving the tumor vasculature, actively targeted nanoparticles must traverse a long interstitial pathway to reach target cells. During this process, several physiological barriers will limit their penetration from the tumor vasculature into deeper tumor tissues, such as spatial blockage of the extracellular matrix and nonspecific uptake by tumor-associated fibroblasts and tumor-associated macrophages. These factors severely limit the delivery efficiency and efficacy of actively targeted nanomedicines. Therefore, the therapeutic efficacy of active targeting is still controversial.

6.3.3 Localized delivery of PS

Localized administration of PSs holds significant promise for treating superficial tumors while minimizing systemic phototoxicity. Yet, the efficacious migration of PSs into targeted tumor tissues is challenging due to the stratum corneum barrier.^{162, 210} In this context, dissolving microneedles has emerged as a promising solution, exhibiting distinct advantages to surmount the stratum corneum through creating several mechanical channels across the skin. Particularly in the case of superficial tumors, dissolving microneedles offer direct nanomedicine delivery to superficial lesion sites, effectively circumventing the uneven dispersion and potential toxicity inherent to systemic drug administration. Furthermore, dissolving microneedles can uphold PS stability and effectiveness during storage and transport, thereby addressing pivotal requisites in PDT.¹⁶⁴ Liu et al.²¹¹ introduced a novel approach using PS-loaded dissolving microneedle patches armed with O₂ propellant [referred to as PS-loaded active dissolving microneedle patches (DMNs)] to actively transport PS to tumor sites (Figure 7D). Constructed from polyvinylpyrrolidone-solubilized polymers infused with Ce6 and sodium percarbonate particles, the PS-loaded active DMNs could achieve precise and efficient drug delivery to tumors. Upon skin insertion, the embedded sodium percarbonate particles instantaneously reacted with the adjacent dermal mesenchyme to generate gaseous O₂ bubbles, which could enhance PS penetration depth while mitigating tumor hypoxia during laser irradiation, thereby amplifying PDT effectiveness. Notably, in vivo assessments using tumor-bearing mouse models exhibited a significant inhibition of tumor growth and a 50% increase in survival through active delivery. This investigation offers an expedient and efficient avenue for PS delivery, underscoring the potential of dissolving microneedle drug delivery as a novel strategy for advancing PDT of tumors. In another research, Luo et al.²¹² ingeniously combined chemo-phototherapy with microneedle drug delivery, employing MIL-100(Fe) nanoparticles as a carrier to codeliver the hydrophobic PS ZnPc and the chemotherapeutic agent DOX. By encapsulating the nanoparticles into microneedles for direct tumor delivery without systemic circulation, the concentration of ZnPc at the tumor site was elevated, while the potential cardiac toxicity of DOX was mitigated. A series of in vitro and in vivo investigations confirmed the synergistic potential of chemo-phototherapy in curtailing tumor progression. Consequently, the nanoparticle integrated microneedle patch, as pioneered in these studies, holds considerable promise as a pivotal benchmark in the realm of PDT for superficial tumors.