**3.1 O2 self-enriched metal-based nanoplatform for PDT improving**

Due to requiring a high concentration of O2 for PDT, one of the obstacles in PDT of solid tumors is the lack of enough oxygen because of the hypoxic microenvironment existence. The hypoxic microenvironment is a marking of solid tumors (50~60% of malignant solid tumors are recognized by hypoxia), which creates through imbalance between the oxygen consumption caused by tumor cell reproduction and insufficient oxygen supply created by tumor vascular systems abnormally. In addition to creating tumor resistance to PDT, hypoxia causes tumorigenesis and tumor progression, which can result from the production of hypoxia-inducible factors (HIFs), DNA methylation, and also the production of unwanted metabolites such as hydrogen peroxide (H2O2), which promotes tumor cell metastasis and invasion. Strategies for hypoxia relieving and PDT efficacy improvement, including: (1) hyperbaric oxygen therapy; (2) direct transport to tumor sites by oxygen-carrying agents (e.g., hemoglobin and perfluorocarbon); (3) in situ production of oxygen by degradation of chemicals (such as C3N4 and CaO2), which is delivered by nanomaterials into tumor sites; (4) O2 generation by catalytic conversion of some common appearances of ROSs such as O2 •− and H2O2 into the tumor microenvironment. Pathophysiologic mechanisms (e.g., NADPH oxidase (NOX) enzymes overexpression), in tumors, lead to the production of elevated H2O2 that has a generation rate of 5 nmol per 105 cells h−1.

Catalysis of endogenous H2O2 and conversion to O2 are carried out by natural catalase and catalyze-like oxygen generators or artificial enzymes such as manganese dioxide (MnO2), copper oxide (CuO), nanozymes, and biocatalytic cascades. Due to high sensitivity to TME and fast decomposition in an acidic environment, MnO2 nanostructures were considered a TME-responsive drug carrier. MnO2 and its various forms (e.g., MnO2 nanosheets, MnO2 nanoshell, and MnO2 nanozymes) enhance

#### *Metal-Based Nanomaterials Photodynamic Action with a Focus on Au and Ag Nanomaterials DOI: http://dx.doi.org/10.5772/intechopen.109220*

PDT efficacy through interaction with tumoral H2O2, as a result, O2 generation and ameliorating tumor hypoxia. Also, MnO2 moderates intratumoral GSH, as follows after MnO2 interaction with GSH, Mn4+ converts to Mn2+, which can form hydroxyl radical(• OH), through Mn2+ interaction with H2O2 by Fenton-like catalytic reaction. Tumor overexpressed GSH as a scavenger of ROS, causing tumor cells to become resistant to PDT-induced oxidative stress, thus reducing the efficiency of photodynamic therapy with intratumoral GSH. Nanozymes have enzyme mimetic activity. Nobel metal nanomaterials (such as platinum (Pt), palladium (Pd) nanoparticles, rhodium (Rh)-based nanomaterials), and cerium oxide nanoparticles have served as nanozymes and generate oxygen to promote ROS production for PDT of tumor cells. Rh decomposes H2O2 into O2 through its catalase-like activity, and Rh alloys have a more catalytic effect. Cerium oxide (ceria, CeO2) nanoparticles have the dual performance of superoxide dismutase (SOD) and catalase-like activity and could generate oxygen via the O2 •−and H2O2 catalysis. The bifunctional activity of ceria is the result of electron shuttle between Ce3+ and Ce4+, which are mixed valence states, as follows Ce3+ decompose superoxide anion (O2 •−) and Ce4+ catalysis hydrogen peroxide (H2O2). Natural catalase's poor stability and nonporous properties of artificial enzymes reduce 1 O2 yield and their effect on the tumor environment hypoxia. Also, the amount of H2O2 produced by tumor cells is not large enough that artificial enzymes have a good catalytic function and generate a high amount of O2. For this reason, smart biocatalytic cascade systems designed such as well-assembled multicatalytic nanoreactors have been interested. These systems can supply a large amount of molecular oxygen as a substrate for PDT and also provide more effective communication between catalyst and substrate in these systems [27, 34–36].

#### **3.2 Gold-based nanomaterials**

Gold nanomaterials (AuNMs) have different biomedical applications such as PTT and PDT, due to their physicochemical properties (**Figure 2**).

AuNMs features are including ease and the possibility of achieving different sizes of synthesis, flexibility in surface modification, great nanocarriers for transporting a wide range of components such as small molecules, nucleic acids, proteins, and antibodies, controlled-release, site-specific triggering, low toxicity, high conductivity, and localized surface plasmon resonance (LSPR). LSPR is one of the optical properties of AuNMs, which refers to the total oscillations of the conduction band electrons of metal nanoparticles. The absorption band of LSPR in AuNMs is affected by shape and aspect ratio, so that with tuning the shape and aspect ratio due to the surface photon confinement effect, the absorption spectrum reaches to NIR range. AuNMs are one of the NIR-responsive materials that have an absorption spectrum with a longer wavelength in the range of NIR (750–1000 nm), where the depth of light penetration into the target tissue is greater in this range. Therefore, they overcome the organic PSs' limitations, whose absorption spectrum has a shorter wavelength (300–700 nm) and a lower penetration depth (less than 1 mm) and can create a novel generation of PSs [2, 3, 37–40]. Gold nanoparticles' (Au NPs) appropriate wavelength for PDT is between 800 and 900 nm [12].

Au NPs can exist in different shapes, such as spheres, rods, shells, stars, cages, and gold nanoflowers (**Figure 2**). Au NPs can be isotropic and anisotropic based on their shapes and characteristics. Just as the size alterations affect the plasmon absorption band, the geometry alterations of the nanoparticles also affect the SPR band, so according to Mie's theory, the particle's electric surface charge density changes with

the alteration of the particle diameter. Therefore, with the increase in size, a shift in the location of the SPR absorption band toward longer wavelengths is observed. In addition, a small change in the particle geometry causes significant changes in the characteristics of the SPR absorption band. These alterations are also observed in anisotropic AuNPs, which are even higher than the changes caused by the increase in size in isotropic AuNPs. Anisotropic AuNPs have multiple SPR absorption bands contrary to the isotropic morphology, which has only one SPR absorption band. Therefore, the wavelength of the SPR absorption band shifts from the visible region to the near-infrared region (NIR), by changing the shape of the gold nanoparticle. Among the anisotropic morphologies of AuNPs, we can mention gold nanoflowers (AuNFs), gold nanostars (GNSs), gold nanorods (AuNRs), gold nanoplates, etc. Due to their anisotropic shape and symmetric structure with branches, GNSs possess tunable localized surface plasmon resonance (LSPR) in the near-infrared (NIR) region of the electromagnetic spectrum, and they are appropriate nanoplatforms for theranostic applications [3, 37, 41, 42].

The GNS@ICG-Ab-CIK nanoplatform designed by Shujing Liang et al. in 2020 is a nano-theranostics system that has the potential for cancer theranostics. In this nanoplatform, GNS (gold nanostars) was combined with anti-HER2 (trastuzumab) and targeted. Anti-HER2 (trastuzumab) is a monoclonal antibody that acts against HER2-overexpressing human breast cancer cells. This monoclonal antibody prevents cell cycle progression and inhibits angiogenesis and antibody-dependent cytotoxicity. GNS combined with trastuzumab was loaded with Indocyanine green (ICG) through electrostatic adsorption, and a new nanoprobe was formed that has the ability of trimodel imaging (photoacoustic (PA), computed tomography (CT), and fluorescence imaging) and also creates effective synergistic PTT/PDT of HER-2-positive breast cancer. GNSs protect ICG from photo-thermal damage and improve 1 O2 production, as well as increase their stability in the bloodstream and reduce blood clearance. To improve targeting efficiency, cytokine-induced killer (CIK) cells, which have the

*Metal-Based Nanomaterials Photodynamic Action with a Focus on Au and Ag Nanomaterials DOI: http://dx.doi.org/10.5772/intechopen.109220*

tumor-homing ability, were used. These cells activate the immune system after intravenous administration, so the constructed nanoplatform acts against cancer through immunotherapy in addition to PDT and PTT [37, 43].

AuNFs possess branch structures and flower-like shapes along with absorbing light in the red deep region (600—900 nm). AuNFs have a high surface-to-volume ratio and tissue penetration compared with other forms of AuNPs such as spherical and rod-shaped and are used as an agent for imaging, nano-vehicle, and therapy [3, 44]. PDA-Ce6-GSH-AuNF is a multifunctional system capable of cancer synergistic therapy through PDT and PTT treatments, in which AuNFs with a diameter of about 80 nm and an absorption band in the vis-near infrared (Vis–NIR) range of 800–900 nm are used. AuNFs were synthesized using HAuCl4 salt at 0°C, through the template-free method, which has two steps. In the first step, HAuCl4 was reduced by ascorbic acid (AA) and Au seeds were formed. In the second step, through the seed-mediated approach, and using NH2OH·HCl as a reducing agent, gold ions (Au3+) were converted into gold nanoparticles. These nanoparticles are deposited on Au seeds (crystal nucleus) and branch structures of AuNFs are constructed. Then glutathione (GSH) molecules are anchored to the gold surface by their thiol group, and it is utilized as a linker for connecting chlorin e6 (Ce6) to AuNFs through the activated carboxyl group of GSH. Then Ce6-modified AuNFs were coated with polydopamine (PDA). PDAs are surface modification agents with high chemical and thermal stability and containing catecholamine functional groups with great adhesion properties, which create thin nanoscale layers on the surface of organic materials. Thin PDA layers increase cellular absorption of nanoparticles and their mucopenetration and also play a role in hydrophobic drug delivery, which is due to the hydrophilicity and zwitterionic properties they give to the material. PDA has an elevated absorption capability in the NIR region and high efficiency of energy conversion and creates red-shift adsorption, so as a photothermal agent, it can increase the PTT efficiency of the nanoplatforms. The nanoplatform was examined in vitro and in vivo, through a near-infrared (NIR) laser with a wavelength of 660 nm for PDT and 808 nm for PTT. Based on the results, the amount of Ce6 loaded in Au NFs was 14.0 wt.%, the singlet oxygen production efficiency by the designed nanoplatform was approximately 91.0% of free Ce6, and the photothermal conversion efficiency was 23.6% (7.0% more than free Au NFs). As regards that the antitumor efficiency was increased by combining PDT and PTT treatments, PDA-Ce6- GSH-AuNFs were considered as dual phototherapy agents [3].

Au NRs are one-dimensional anisotropic nanoparticles. Contrary to spherical gold nanoparticles, which have one LSPR band in the range of 520 nm, AuNRs have two LSPR bands; transverse LSPR (t-LSPR) with a wavelength in the range of 520 nm and longitudinal-LSPR (l-LSPR) with higher wavelengths in the range of the biological window (560–950 nm) and (1000–1350 nm). l-LSPR can be adjusted by changing the aspect ratio of AuNRs, and compared with the LSPR of spherical nanoparticles, AuNRs are more sensitive to environmental variations resulting from dielectrics. These features make AuNRs possess wide applications in biomedical, including LSPR-biosensing. AuNRs are one of the metal nanoparticles that are used in non-aggregation plasmonic colorimetric sensors as a mediator in the etching/ growth process. This process alters the shape, size, and environmental dielectrics of metal nanoparticles, which follows the plasmon band changes. Based on the studies on Au NRs, gold nanoshells (nanocages, nanorod-in-shell, and nanoparticle-inshell) and gold nanobipyramids (Au NBPs) (**Figure 2**), have been used as PS in PDT and can produce singlet oxygen (1 O2) [45–47]. Au NBPs are, like the Au NRs, one of the novel plasmonic nanoparticles, which have longitudinal dipolar plasmon

with a tunable wavelength from the visible region to the near-infrared. Au NBPs have higher shape and size uniformity and against Au NRs, which have curved or flat ends, Au NBPs contain two sharp end apexes, so Au NBPs could create higher regional electric field enhancements with more slender peak width. Au NBPs possess broad plasmonic usage in the areas of photocatalysis, sensing, and switching (such as plasmonic index-change-based sensing method, colorimetric and selective immunoassay, and electrochemical plasmonic switching), biomedical applications (PTT, PDT, real-time bioassays, as thermocycling-creating nanoreactors for rapid and quantitative real-time PCR, plasmon-enhanced fluorescence bioimaging agents, etc.), and plasmon-enhanced spectroscopies (PL), surface-enhanced Raman spectroscopy (SERS), etc.) [48].

Au NP creates antimicrobial effects through mechanisms, which include interaction with bacterial cell barriers, interaction with biomolecules such as enzyme activity inhibiting, DNA binding or interference in protein synthesis, bacteria-killing by photothermal effect, the redox imbalance, and increasing the effectiveness of antimicrobial photodynamic therapy (aPDT). According to studies, AuNPs do not cause the redox imbalance by making changes in the ROS level, but they affect the level of glutathione (GSH) [49]. According to reports, the mechanisms through which gold nanoparticles can increase the effect of aPDT are as follows: improving the relative distribution and production of ROS, and also, it can increase the amount of PS excitation and improving the efficiency of aPDT through conjugation with PS such as AuNPs-conjugated methylene blue (MB) against bacteria such as the mature methicillin-resistant *Staphylococcus aureus* (MRSA) biofilm [50–52]. By reducing photobleaching, AuNPs also increase the efficiency of photodynamic therapy. In a study in 2021, the effect of biosynthetic AuNPs-mediated mycelium of *Mucor plumbeus* on aPDT efficiency was investigated. Based on the observations of this study, biogenic AuNPs in combination with (Mb) methylene blue as PS can cause effective aPDT of both Gram-negative (*Escherichia coli*) and Gram-positive (*S. aureus*) bacteria. Biogenic AuNPs reduce the photobleaching speed of methylene blue by changing the kinetics of the photofading process [50]. In 2017, phototheranostic nanoagents (PTNAs) were designed, which possess biosynthesized AuNPs using *Syzygium cumini'*s fruit extract (ScAuNPs). In this study, the effects of ScAuNPs have been compared with chitosan-mediated AuNPs (ChAuNPs). According to the results, antioxidant efficacy, drug loading, fluorescence quantum yield (Q ), singlet oxygen quantum yield, and antimicrobial PDT of ScAuNPs and their conjugated probes/nanoprobes were better than those of ChAuNPs [53].

Other forms of AuNMs are nanoclusters (**Figure 3**). Generally, metal nanoclusters (Au, Ag, Cu, etc.) have a different structure from metal nanoparticles, and subsequently, their characteristics, including their optical characteristics, change, too. These nanomaterials have several to 100 atoms and their size is less than 2 nm, which is comparable with the Fermi wavelength of electrons, which is usually less than 1 nm because as the size decreases, the continuous energy band of metals changes to a discrete energy level, which is associated with molecular-like electronic transitions (i.e., HOMO – LUMO transitions) in valence states. Metal nanoclusters (MNCs) could be categorized as water-soluble MNCs stabilized by biomolecules (proteins, peptides, DNA, etc.), and oil-soluble MNCs with easy crystallization, such as Au25(PET)18 (PET is phenethyl mercaptan), that is, oil-soluble mono-metallic NCs nobel metal nanoclusters (MNCs) have capping agents such as protein, DNA, and small organic molecules, good biocompatibility, and the ability to easily modify the surface characteristics. Also, they are good fluorescent labels, due to large stock

*Metal-Based Nanomaterials Photodynamic Action with a Focus on Au and Ag Nanomaterials DOI: http://dx.doi.org/10.5772/intechopen.109220*

**Figure 3.** *Properties of AuNCs and AgNCs.*

shift, size-dependent excitation and emission wavelength, and high optical stability [54–57]. Gold nanoclusters (AuNCs) consist of less than 100 gold atoms and have a size smaller than 2 nm, which is comparable with the Fermi wavelength of electrons (< 1 nm). AuNCs do not have localized surface plasmon resonance like gold nanoparticles because the electrons of the valence layer of AuNCs cannot move freely. AuNCs do not absorb plasma resonance in the visible region. The maximum fluorescence of gold nanoclusters is in the visible to NIR region, which is the appropriate wavelength for tissue penetration. Also, high photostability and great fluorescence lifetime in vivo conditions are other characteristics of AuNCs. Therefore, they have biomedical applications such as biosensing, bio-imaging, and therapy. AuNCs are used as inorganic PS in PDT. In 2014, in a study, a nanoprobe was designed in which Doxorubicin (DOX) was loaded in a nanocomposite consisting of AuNCs encapsulated in Zeolitic imidazolate framework-8 (ZIF-8). Due to their small size, AuNCs have limitations such as low enhanced permeability and retention (EPR) effect, short duration in blood circulation, and low accumulation in tumor sites due to fast clearance. To solve these limitations in this study, AuNCs were encapsulated in the inner of ZIF-8 and DOX in the channel of ZIF-8. This nanocomposite is pH-responsive and bifunctional so that it uses synergistic PDT/chemotherapy to treat breast cancer. And due to being pH-responsive, it is not released in neutral environments and normal cells are not affected. Their release occurs in acidic tumor sites, so the performance of PDT and chemotherapy is enhanced [29, 38, 54].

#### **3.3 Silver-based nanomaterials**

Due to the innate antibacterial properties of silver ions, it has been used by humans for many years and recently has been noticed in cancer therapeutics. According to studies, mechanism of Ag-based agents for therapy is irreversible apoptosis along with contacting Ag<sup>+</sup> to a thiol-rich protein that is located on the cell, and bacterial membrane leads to decrease of these proteins. So superficial contact is a factor that regulates antimicrobial effects. Ag<sup>+</sup> is restricted by a lack of stability

in the physiological body environment and target ability for tumors. So synthesis of nanoplatforms for Ag+ accumulation in the diseased region and making synergistic therapeutic effects is vital [58]. Silver nanoparticles usually possess a size between 20 and 25 nm due to their high surface-to-volume ratio [12]. They have wide contact areas with viruses and bacteria, which subsequently improves the bactericidal performance. Therefore, the effects of silver nanoparticles on Gram-positive and Gram-negative bacteria depend on the size, dose, shape, and total surface area of the nanoparticle.

According to the studies, silver nanoparticles have high antibacterial effects on *Streptococcus mutans* in low concentrations, so their toxicity is reduced. Silver nanoparticles affect the morphology of *E. coli* and *S. aureus*, also they can alter the expression of some coating proteins such as (OmpA, OmpC, OmpF, OppA, and MetQ ) and heat shock proteins (IbpA and IbpB). Ions released from silver nanoparticles can also have a biocidal effect [59]. Silver nanoparticles enhance photodynamic performance against the wide-broad spectrum of microorganisms bacterial and fungal species and including antibiotic-resistant strains, which is a public health threat all over the world. They can be used as carriers for PSs or as hybrids, composites, and conjugates with PSs [12, 60]. Silver core-mesoporous silica shell nanoparticlescontaining HPIX (Hematoporphyrin IX dihydrochloride) is a hybrid PS that was synthesized in 2016 by Tevhide Ozkaya Ahmadov et al. [61] which exhibited significant photodynamic inactivation ability against MRSA (a multidrug-resistant strain of *S.aureus*) in such a way that its lethal efficiency increased up to six times [62]. As the size of silver nanoparticles decreases, their antimicrobial effect will be greater due to the increase of surface-to-volume, such that silver nanoclusters (AgNCs) with a size smaller than 2 nm exhibit stronger antimicrobial effects. Similar to AuNCs, AgNCs participate in photodynamic therapy. AgNCs have remarkable luminescence properties, biocompatibility, tunable size and photoluminescence, photostability, and low toxicity. Ag NCs have more antimicrobial capability than AuNCs, but their stability is lower than AuNCs, in a way that they don't preserve for a long time such as a few months while some AuNCs could preserve for more than a few months [63, 64].

The optical features of AgNCs are attributed to factors such as the quantum confinement effect, surface ligand effect, and the electronic transition between the highest occupied molecular orbital (HOMO) and the lowest unoccupied molecular orbital (LUMO) (**Figure 3**) [65]. Surface ligand affects the intramolecular and intermolecular control of nanoclusters, which are two kinds of continuous systems of nanoclusters control and both together adjust nanocluster systems. Intramolecular control points out the manipulation of metal–ligand compositions and bonding environment at the single-molecule level. Intermolecular control refers to changes in the aggregation pattern in amorphous or crystallographic forms. Ligand exchange and heteroatom alloying are methods for intramolecular control. Among the methods used for intermolecular control, we can mention cluster-based metal–organic framework, aggregation-induced emission, and intercluster metallophilic reaction. Based on the studies, manipulation of nanocluster crystallographic networks at the supramolecular level and the adjustment of intramolecular and intermolecular interactions make the nanoclusters become CIEE (crystallization-induced emission enhancement)-active or -inactive nanomaterials [62].

There are various methods for metal nanoclusters synthesis, among these approaches, the template-based method is suitable for the generation of metal nanoclusters, and they involve improving the stability, shape, and size-controlled and fluorescence tunability of metal nanoclusters. Templates or capping agents

#### *Metal-Based Nanomaterials Photodynamic Action with a Focus on Au and Ag Nanomaterials DOI: http://dx.doi.org/10.5772/intechopen.109220*

can be dendrimers, proteins, peptides, and DNAs [50]. Bovine serum albumin (BSA) and human serum albumin (HSA) are the common proteins used as capping agents in the synthesis of nanoclusters (NCs). BSA has various functional groups such as -OH, -NH2, -COOH, and -SH, which are involved in the synthesis of metal nanoclusters (MNCs) by creating steric stabilization. BSA has disulfide bonds that establish a covalent bond with the MNCs core through the sulfur. Also, the pH of the environment participates in the formation of nanomaterials and MNCs. For instance, BSA converts silver ions into silver nanoparticles at pH (6–8) and generates silver nanoclusters at pH higher than 11. Of course, the protein itself can also reduce the silver ions (Ag<sup>+</sup> ) without adding external reducing agents and turning them into silver nanoparticles and nanoclusters. BSA obtains various conformations, including -N (native), B (basic), A (aged), and U (unfolded), along with alterations of environment pH from neutral to alkaline that different nanomaterials are stable in some of these conformations [65].

AgNCs are also constructed with DNA templates in the way that this nanocluster has a high affinity to N3 cytosine. The number of single-stranded cytosines installed in the primary and secondary structures of DNA (such as hairpin loops, i-motifs, and G-quadruplexes) controls the size and shape of AgNCs. Another feature of DNA-AgNCs is their resistance to photobleaching, which makes them have applications in nanophotonics and biosensing. One of the advantages of using oligonucleotides for the synthesis of AgNCs is the possibility of their synthesis with different sequences. AgNCs can exert antimicrobial effect through DNA intercalation and inactivation of type ΙΙ topoisomerases. In 2016, Siamak Javani et al. investigated the antibacterial performance of fluorescent DNA-AgNCs, which has an inhibitory effect on the growth of Gram-positive and -negative bacteria in sub-micromolar concentrations. These DNA-AgNCs were synthesized with three types of oligonucleotide sequences named Seq1, Seq2, Seq3, and the absorption wavelengths of the resulting nanoclusters are 370 nm, 480 nm, and 560 nm, respectively, and their emission wavelengths are 475 nm, 572 nm, and 630 nm, respectively. The result of the antibacterial test for *E.coli* showed that Seq3 had more inhibitory effect than Seq2 and Seq1 showed the least inhibitory activity. Based on the results of this study, the factors that regulate the antibacterial performance of DNA-AgNCs are the amount of silver and factors such as sequence, structure, and Ag arrangement [66, 67].

## **4. PDT effects on immune system**

PDT can lead to cell death through apoptosis, necrosis, autophagy, or proptosis. The type of cell death induced by PDT depends on the location of the PS and the amount of photodamage. Each of these cell death leads to the release of molecules such as alarmins or damage-associated molecular patterns (DAMPs), cytokines, growth factors, and other immunomodulating agents. Alarmins as immunostimulators are released after photooxidative damage that triggers innate immunity, which leads to the activation of adaptive immunity. They are detected by pattern recognition receptors (PRRs) that are expressed on immune cells, and subsequently, activate immune cells after DAMPs are attached to the PRRs. Similar to the lesion from infection and other tissue damage, PDT- mediated damage of tumor and endothelial cells is caused by release of inflammatory factors including arachidonic acid-derived metabolites arising from membrane lipids peroxidation (such as thromboxanes, prostaglandin, and leukotriene), high amount of cytokines such as the interleukin

(IL)-6, IL-1β, IL-2, IL-1β (possess a prominent role in PDT efficiency), MIP2 (CXCL2), tumor necrosis factorα (TNFα), acute phase proteins, pro-aggregatory and vasoactive mediator releasing (result in platelets activation and clotting cascade initiation). These factors lead to the migration of innate immune cells toward the tumor site for tumor cell elimination. The PDT immune cells from PDT-mediated damage include neutrophils, macrophages, natural killers (NK), and dendritic cells (DCs). Previous studies demonstrated that systemic neutrophilia is incorporated in immune-PDT interaction of tumor cells through various factors resulting from PDT-mediated damage, which can be mentioned in some cases, including: (1) Complement activation upon PDT is caused to release tumor tissue anaphylatoxins (such as C3a and C5a), subsequently, vascular permeability increasing and neutrophil infiltration. (2) Adhesion molecules such as E-selectin and ICAM1 for neutrophil adherence on tumor tissue and micro vessels, also adhesion to the subendothelial matrix by the β2 integrin receptors (3) Acute-phase proteins (APPs), which facilitate neutrophils migrate, mature neutrophil progenitors more quickly, and help them leave the bone marrow [68].

Macrophages as effector cells participate in immune-PDT of cancer cells. The release of HSP70 from tumor cells damaged by PTD will activate macrophages. Hsp70 is bound to PRRs (which are expressed on macrophages such as the toll-like receptors (TLRs) and as a result of this binding, TLR2/4 of macrophages is activated, followed by TNFα releasing. TNFα as a cytolytic cytokine leads to indirect tumor cell elimination. C3 and MBLs (mannose-binding lectins) opsonization of tumor cells provides their phagocytose possibility by macrophages, due to complement receptors expressed by macrophages could recognize opsonized agents. NK cells have also been interested in cancer PDT. According to reports, major histocompatibility (MHC) class I-related molecule (MICA) and Natural Killer Group 2D (NKG2D) ligands from photodamage are recognized by NKG2D receptors on NK cells and lead to activation of these receptors, consequently enhancing PDT-induced immunity. Dendritic cells (DCs) as antigen-presenting cells communicate between innate and adaptive immune systems. PRRs of DCs detect DAMPs, released by PDT-damage, following it to activate/mature dendritic cells leading to augment MHC class II and co-stimulatory factors expression, which will be associated with the presenting of antigens to T lymphocytes and activation of type 1, 2, and 3 of adaptive immunity. In type 1 immunity cytokines such as IL-12 and IFN-γ are expressed by CD4+ T cells leading to the activation of the cytotoxic function of CD8+ T cells. In type 2 immunity, CD4+ T cells deviate to the Th2 phenotype, which is associated with cytokines expression such as IL-4 and antibody generation from B cells. According to studies demonstrated, CD8+ T cells have a pivotal role in immune-PDT, in a way defects in these cells reduce the efficacy of PDT. Type 3 immunity induced by PDT arises from the increasing of T helper (Th17) cell numbers in the tumor-draining lymph nodes (TDLNs). Th17 cells are a subset of CD4+ T cells that generate IL-17 cytokine that according to studies, this molecule has a significant role in neutrophils recruitment in TDLNs upon PDT and also neutrophils are effective in activated CD8+ T cells accumulation into tumor cells [68, 69].
