It is widely recognized in the scientific community that treating glioblastoma (GBM) presents significant challenges, where the therapeutic regimen known as the Stupp protocol only results in a survival of 14–16 months [1]. This substantial issue gives rise to numerous research opportunities in the search for alternative therapies for this condition [2]. GBM is the most aggressive and lethal form of primary brain tumor, with a median survival of less than 15 months despite current standard treatments involving surgery, radiotherapy (RT), and chemotherapy (CTX). Recent advances in molecular and genetic profiling have improved our understanding of GBM’s complex biology, highlighting the role of mutations in the isocitrate dehydrogenase 1 and 2 genes (IDH1/2), MGMT promoter methylation, and other markers that are central to GBM’s pathogenesis and resistance mechanisms [3]. However, effective treatments that can significantly extend patient survival remain limited. One promising avenue is photodynamic therapy (PDT), which is a minimally invasive medical treatment that leverages the interaction between a photosensitizing agent, light, and molecular oxygen (O₂) to produce reactive O₂ species (ROS) capable of selectively destroying targeted cells [4]. The therapeutic potential of PDT was first recognized over a century ago, but its clinical application began to gain significant momentum in the 1970s with the discovery of porphyrins as effective photosensitizers (PSs). Initially developed for the treatment of skin cancers, PDT has evolved over the decades to include a broader range of applications, from non-oncological conditions to various types of cancer, including GBM, one of the most aggressive brain tumors [5, 6]. Early generations of PSs, like hematoporphyrin (HP) derivatives (HpD), laid the foundation for PDT by demonstrating the ability to accumulate in malignant tissues. However, their limited tissue penetration and prolonged photosensitivity hindered widespread use. As research progressed, second-generation PSs such as 5-aminolevulinic acid (5-ALA) and its derivatives improved selectivity and efficiency, allowing deeper tissue targeting and fewer side effects. Today, third-generation PSs are being developed, often in conjunction with nanotechnology, to further enhance targeting capabilities, improve bioavailability, and minimize dark toxicity but in the best scenario, there remain in preclinical trials [7]. Nanotechnology offers significant advantages in the development of PSs for PDT, enabling enhanced precision, efficiency, and safety in treatment [8]. By encapsulating PSs within nanocarriers, their solubility, stability, and bioavailability can be greatly improved, overcoming limitations associated with traditional PSs from the first generations, such as poor water solubility and rapid degradation. Nanoparticles (NPs) also provide the opportunity for image processing [9], diagnosis [10], treatment [11], targeted delivery [12], reducing off-target effects [13], gene transferring [14], and minimizing damage to healthy tissues [15], particularly in sensitive areas like the brain. Cancer therapy targeting tactics can be classified into passive and active approaches, each possessing distinct advantages and limitations in the context of brain tumors. Passive targeting utilizes the enhanced permeability and retention (EPR) phenomenon, whereby NPs aggregate in tumor tissues owing to compromised vasculature and deficient lymphatic outflow. This method encounters obstacles in brain tumors because of the blood-brain barrier (BBB) and the diverse characteristics of tumor vasculature, which frequently restrict efficient NP delivery. Active targeting employs ligands or antibodies coupled to NPs to selectively attach to overexpressed receptors on tumor cells and endothelial cells of the BBB, thereby promoting receptor-mediated transcytosis [7]. Although active targeting provides enhanced selectivity and efficiency, it is frequently impeded by inadequate penetration into the tumor bulk and possible off-target effects. Integrating both tactics with modern delivery systems and BBB-disrupting approaches, such as targeted ultrasound (US) or receptor-mediated transcytosis, offers potential to overcome these constraints. These methodologies can enhance the accuracy and therapeutic efficacy of NP-based interventions for brain tumors, promoting the advancement of more effective treatments.

Furthermore, nanotechnology allows for multifunctional systems, integrating imaging agents, targeting ligands, and therapeutic molecules into a single platform, thus enabling theranostic applications. These advancements not only enhance the therapeutic efficacy of PDT but also allow for more precise control over treatment parameters, such as light activation and ROS generation, paving the way for more personalized and effective cancer therapies.

Over the decades, the clinical applications of PDT have expanded, with approvals for treating various cancers, including lung, esophageal, and skin cancers, as well as ongoing trials for its use in treating brain tumors like GBM. However, many of the PSs tested in clinical studies for glioma patients belong to the first and second generations. Some of these studies show moderate efficacy and increased patient survival, but many other studies have not been completed, or the results remain inconclusive.

The growing understanding of tumor biology and advances in molecular techniques have enabled researchers to fine-tune PDT protocols, making it a promising complementary approach in cancer therapy, especially for tumors resistant to conventional treatments. This review explores the potential of PDT in GBM treatment with the most recent developments, focusing on its mechanisms, challenges, and the evolving landscape of PSs across different generations. For the writing of this review, a comprehensive analysis of articles related to the topic was conducted through a systematic search of scientific articles, reviews, preclinical studies, and clinical trials. The main databases consulted were PubMed, Scopus, Web of Science, and Google Scholar. Most of the included articles were published within the last 5–10 years, with a focus on research detailing the combined use of PDT with other therapies. The keywords used in the search were: malignant gliomas, GBM, photoassisted therapy, PDT, synergism, and pro-oxidant therapies, in all possible combinations. Additionally, a special focus was placed on the combinatory option with other therapies that have been investigated in recent years, demonstrating the versatility of light-assisted technology in damaging tumor cells.

Tumors located in the central nervous system (CNS) affect both the brain and spinal cord, with GBM being among the most aggressive and incurable of these malignancies [16]. GBM are highly vascular tumors characterized histologically by necrosis areas, microvascular proliferation, high mitotic rates, invasiveness, pleomorphism, and nuclear atypia [17–19]. GBM is generally considered a spontaneous tumor. The only confirmed risk factor at present is exposure to high-dose ionizing radiation [20]. Certain hereditary syndromes, including neurofibromatosis, Li-Fraumeni syndrome, and von Hippel-Lindau syndrome, have been shown to correlate with a minor percentage of GBM patients [21, 22]. Its development was associated with dysregulation of the G1/S checkpoint in the cell cycle. Additionally, disruptions in tumor suppressor pathways, including p53 and retinoblastoma (Rb), as well as mutations in genes regulating Rb, play a crucial role in the pathogenesis of these tumors [23]. There is a correlation between human cytomegalovirus (HCMV) and GBM development, as HCMV-encoded proteins activate intracellular signaling pathways involved in mitogenesis, mutagenesis, apoptosis, inflammation, and angiogenesis [21].

Malignant gliomas have long been categorized into two categories based on variations in molecular markers, clinical presentations, and disease progression. Primary GBM, commonly referred to as de novo GBM, develops without any apparent precursor lesions [24]. These are more common, typically diagnosed in patients over 50 years of age, and are associated with a more aggressive clinical course. These tumors have a unique molecular profile characterized by specific genetic alterations, such as mutations in the TERT promoter gene, loss of chromosome 10, suppression of the PTEN gene, amplification of the EGFR gene, and the presence of the EGFRvIII mutation. These genetic alterations promote tumor growth and activate the Shc-Grb2-Ras and phosphatidylinositol 3-kinase (PI3K) signaling pathways, resulting in increased ability to form tumors, cell division, and resistance to cell death. It is important to note that primary GBM tumors do not have mutations in the genes IDH1/2 [25]. In contrast, secondary GBM develops from pre-existing low-grade or anaplastic astrocytomas and generally has a more favorable clinical course. These tumors are characterized by mutations in codon 132 of IDH1 or codon 172 of IDH2 (IDH1/2 mutant), which inhibit the activity of α-KG-dependent dioxygenases—key enzymes involved in hypoxia detection, histone deacetylation, and DNA methylation. Additionally, secondary GBM frequently exhibits mutations in the ATRX and TP53 genes [23, 25].

Since 2021, the classification of gliomas has undergone significant changes, with the WHO proposing a new system based on molecular alterations, including IDH1/2 mutation status and 1p/19q co-deletion status. This updated classification system categorizes diffuse gliomas into three distinct types: oligodendroglioma with IDH1/2 mutation and 1p/19q co-deletion, astrocytomas with IDH1/2 mutations but without co-deletion, along with TP53 mutations and ATRX loss; and GBM with wild-type IDH1/2. These new categories replace the previous distinction between primary and secondary GBM and also exclude gliomas with IDH mutations from the GBM group (Figure 1) [26].

The standard clinical treatment for patients with gliomas, regardless of their classification, typically involves an initial surgical resection followed by ionizing radiation therapy, with doses of approximately 60 Gy. However, this treatment often yields suboptimal results, as tumors frequently recur, and RT commonly causes toxic effects on neural tissue. Consequently, adjuvant CTX is employed, utilizing alkylating agents to enhance patient survival [23, 25].

Advancements in molecular technologies have also facilitated the identification of key molecular markers, such as the methylation of the MGMT gene promoter, which predicts the response to alkylating CTX agents like temozolomide (TMZ). These markers enable the personalization of treatments and the selection of more effective therapies for patients, thereby improving clinical outcomes. MGMT is a DNA repair enzyme that can restore O-6-methylguanine during genomic damage caused by alkylating agents used in CTX [25]. Consequently, patients with methylated MGMT gene promoters are less likely to effectively repair the damage caused by TMZ, making them more responsive to the treatment [17]. In addition, IDH-mutant gliomas may respond differently to conventional therapies. For instance, they may exhibit increased sensitivity to certain chemotherapeutic agents, and the mutation status can guide the selection of more effective treatment regimens. For this reason, the unique metabolic and molecular characteristics of IDH1/2-mutant gliomas present opportunities for targeted therapies [27]. Ongoing research is being conducted on drugs that are particularly engineered to block the IDH1/2 enzymes or to offset the effects of 2-HG [3]. These medications possess the capacity to offer novel therapeutic options for individuals impacted by these illnesses. The IDH mutation status is commonly used as a criterion for classifying patients in clinical trials, which affects the development and evaluation of new therapies. Evaluating the IDH mutation status in gliomas is crucial for determining its suitability for experimental therapies and for reviewing clinical trial outcomes.

In this regard, Tumor-treating fields (TTFields) therapy is recognized as an effective adjunctive treatment for newly diagnosed patients with GBM [28]. In these malignancies, TTFields are low-intensity, intermediate-frequency alternating electric fields with a frequency of around 200 kHz, and the electric field strength typically remains below 5 V/cm in most brain areas [29]. Recent long-term survival results from a prospective study on TTFields have shown significant enhancements in overall survival (OS) and progression-free survival (PFS) when combined with adjuvant TMZ, compared to TMZ alone, in newly diagnosed GBM, integrated into standard therapy, without negatively impacting quality of life [30]. Some retrospective investigations have begun the assessment of the correlation between TTFields and the molecular characteristics of glioma malignancies, including IDH mutational status. Nonetheless, the majority of these investigations remain in initial stages, with limited outcomes. A study conducted by Zhang et al. [31] revealed that OS exhibited no significant relation with age, gender, Karnofsky performance status (KPS) score, MGMT status, IDH status, EGFR status, 1p/19q deletion status, or TERT promoter status.

In contrast to conventional and adjuvant treatments, PDT is a non-invasive treatment that employs a PS, light, and endogenous molecular O₂. Although these components are individually non-toxic, when the PS is exposed to light, ROS are generated, including superoxide anion and/or singlet O₂ (¹O₂), which induce cytotoxicity and cell death. PDT was explored in the context of GBM management with some encouraging results. However, many of these clinical studies have not gathered sufficient information to provide reports on the success or failure of PDT with different PSs such as Photofrin and 5-ALA (NCT01966809, NCT00118222, NCT00002647). Recently, the PS 3-(1-Butyloxy)ethyl-3-deacetyl-bacteriopurpurin-18-n-butylimide methyl ester, commercially known as Photobac, has entered phase 1 clinical trials in the recruitment stage (NCT05363826) as an adjuvant to resection of GBM. Similarly, 5-ALA HCl is in phase 1 clinical trials and the recruitment stage (NCT05736406) for GBM treatment is followed by standard therapy. A completed clinical study (NCT03048240) yielded promising results using 5-ALA HCl in combination with standard treatment, demonstrating increased survival without adverse effects, toxicity, or mortality. The study reported a median OS of 23.4 months.

In the following sections, we will review and discuss the mechanistic aspects, main advantages, and disadvantages of this therapeutic approach applied to malignant gliomas, as well as potential combinations that have been explored with this photo-assisted therapy.

PDT is a medical technique that uses light sources to trigger the activation of PSs for the treatment of cancer and other disorders. This approach is non-surgical in most of the cases and minimally invasive. PDT has received regulatory approval for the treatment of various cancers in several countries, though its specific use for gliomas is still under clinical investigation. In the United States, the FDA has approved PDT for cancers such as non-melanoma skin cancer, esophageal cancer, and non-small cell lung cancer, using PSs like Photofrin. While PDT for gliomas has not yet received formal FDA approval, ongoing clinical trials are evaluating its potential for treating GBM (NCT05736406, NCT05363826). Similarly, the European Medicines Agency (EMA) has approved PDT for certain cancers with glioma-specific applications currently in research. Japan, a leader in PDT innovation, has approved its use for cancers such as lung and gastric cancers, and like the U.S. and Europe, is investigating PDT’s efficacy for malignant gliomas. With the advancement of clinical research worldwide, it is expected that PDT would soon be established as a crucial therapy choice for aggressive brain cancers such as GBM as an adjuvant therapy.

PDT offers a promising approach for treating malignant gliomas by using light-sensitive compounds to generate massive ROS that selectively target and destroy tumor cells [32]. This minimally invasive technique allows for precise control of treatment, reducing damage to surrounding healthy brain tissue, and shows potential in overcoming the challenges of conventional therapies, such as drug resistance and limited efficacy in highly aggressive tumors like gliomas.

A PDT protocol for glioma treatment involves selecting a PS, such as porphyrin-based compounds or prodrugs, that preferentially accumulates in glioma cells. After oral, systemic, or local administration, a waiting period allows the PS to clear from healthy brain tissue. The tumor o remaining tumor cells after surgery are thereafter illuminated with light of a specified wavelength, carefully selected light intensity, usually employing red or near-infrared radiation (NIR), to trigger the activation of the PS [33]. Real-time monitoring and imaging guide the treatment, and post-treatment care includes monitoring for side effects and assessing tumor response [34]. One of the benefits of PDT is its adaptability, as it can be administered in either a single or multiple sessions, dependent on the tumor’s characteristics.

An ideal PS must possess several essential attributes. It should be readily accessible from widely available precursors and demonstrate high quantum efficiency for ¹O₂ production (ΦΔ). The absorption range for ideal PS must be between 680 and 800 nm, accompanied by a high molar extinction coefficient (ε max). Another characteristic is that PS must efficiently accumulate in cancer cells while exhibiting little dark toxicity in other organs and tissues. Additionally, it should be easy to administer, soluble in body fluids, and readily eliminated from it.

PDT can eradicate tumor cells via three established mechanisms: (a) direct cellular destruction of tumor cells through different types of cell death, (b) vascular impairment involving blood vessel destruction, alterations in barrier function, constriction of arteriolar vessels, thrombus formation, and stasis of blood flow, and (c) inflammation coupled with the activation of the immune response (Figure 2). The anti-vascular function is interesting to discuss if it is truly a desired mechanism of PDT in this type of tumor. It will be discussed later in the review.

ROS and ¹O₂ generated in the final stages of PDT process are highly cytotoxic, triggering mainly apoptotic, necrotic, or autophagy-related cell death mechanisms in tumor cells through oxidative processes. The extent and type of cell death depend on factors such as the subcellular localization of the PS, the oxidative stress intensity, and the tumor microenvironment (TME). Protein and membrane damage play a key role in optimizing the cytotoxic efficiency of PDT. When PDT-induced oxidative stress is moderate, mitochondria-mediated apoptotic pathways are activated. ¹O₂ and ROS damage mitochondrial membranes, leading to cytochrome c release and activation of caspases, such as caspase-9 and caspase-3. This process is observed in GBM cells treated with PS that preferentially localize in mitochondria, such as Photofrin or porphyrin-based compounds [35]. At higher oxidative stress levels, excessive ROS production disrupts cellular homeostasis, leading to rapid ATP depletion and plasma membrane rupture. This form of cell death is common in PDT regimens with high fluence rates or in cells with defective apoptotic pathways [36, 37]. Studies in GBM models show that necrotic death is often associated with oxidative damage to proteins [38].

PDT can activate autophagy as either a pro-survival or pro-death pathway, contingent upon ROS levels. In GBM, sublethal oxidative stress stimulates autophagic flux, resulting in the destruction of impaired organelles and proteins. Excessive autophagy can lead to autophagic cell death, marked by the buildup of autophagosomes and the lysosomal destruction of vital cellular components. This has been shown in GBM cells subjected to Hypericin (HYP) PS [39].

Currently, studies on new mechanisms of cellular damage with different PS that had already reported inducing a type of cell death continue to be conducted, demonstrating the triggering of new mechanisms of cellular damage and new pathways of cell death [40–42]. In the case of ferroptosis, phospholipid alterations result from lipid peroxidation, a process initiated by the generation of free radicals and ¹O₂. Once initiated, this mechanism becomes autocatalytic, leading to the formation of hydroperoxides and other byproducts [43, 44]. This oxidative lipid damage is particularly relevant, as it leads to the accumulation of lipid peroxides beyond the capacity of cellular antioxidant systems, such as glutathione (GSH) peroxidase 4 (GPX4), to neutralize them. In ferroptosis, these oxidative processes drive the peroxidation of polyunsaturated fatty acids (PUFAs), a key step in the loss of membrane integrity. Irreversible damage occurs when a hydrogen atom is abstracted from an unsaturated fatty acid or lipid with hydrogen (LH), generating a lipid radical (L•), which subsequently combines with an O₂ molecule to form a peroxyl radical (LOO•). This radical can further react with another LH fatty acid, initiating a new oxidation cycle that generates additional lipid hydroperoxide (LOOH) and L• [45]. During the propagation phase, the LOO• initiates new oxidation chains, while lipid hydroperoxides decompose into other intermediate radicals. In light-induced reactions, alkoxides are generated through direct contact between the triplet-state PS, lipid double bonds, and lipid hydroperoxides, leading to chain fragmentation by cleavage. The accumulation of truncated lipid aldehydes and other oxidation products exacerbates lipid membrane destabilization, triggering membrane rupture and subsequent cell leakage, hallmarks of ferroptosis. Ultimately, this uncontrolled lipid peroxidation overwhelms cellular repair mechanisms, culminating in iron-dependent oxidative cell death, which is a defining feature of ferroptosis [2, 46]. This offers opportunities based on the research conducted by Xu et al. [47], who developed nanocomposites (NC) that can traverse the BBB and specifically target GBM by adhering to an albumin-binding receptor that is highly expressed in both the BBB and GBM. Upon laser irradiation, the ROS-generating PS HYP, encapsulated within the NC, interacts with the ferroptosis inducer erastin to elicit a synergistic anti-GBM effect through PDT and ferroptosis, therefore suppressing GBM proliferation via excessive ROS generation.

Over the years, different generations of PSs have been developed (Figure 3), each addressing the limitations of their predecessors. First-generation PSs were the pioneers in clinical PDT applications but were hindered by suboptimal pharmacokinetics and low ΦΔ. In response, second-generation PS was introduced, designed with improved chemical properties, such as higher selectivity and better tissue penetration. More recently, third-generation PSs have been developed, integrating features like targeted delivery and multifunctional capabilities to enhance both therapeutic efficacy and safety. In the following section, the different PSs explored in the context of malignant gliomas will be described.

While both normal and malignant cells can produce ROS, the concentration of some PS, particularly third-generation nano-based PS, is generally much higher in tumors due to the EPR effect. This process enables PS to persist in the afflicted tissue for an extended period, hence decreasing their presence in healthy cells and mitigating adverse effects [48–50]. Moreover, the coupling of PS with NPs and other ligands, including monoclonal antibodies (mAbs), enhances its distribution and precise targeting to malignant cells [12]. Due to the elevated or differential expression of antigens and receptors in GBM cells, PS engineered to identify these biomarkers can be selectively internalized through endocytosis. This facilitates the formation of ROS primarily in tumor cells upon light activation, leading to their demise while preserving adjacent healthy tissue. Additionally, the use of nanotechnology in the delivery of photodynamic drugs enhances the pharmacokinetics and biodistribution of PS, thereby increasing their therapeutic efficacy. Advanced strategies, such as the conjugation of PS with NP targeting specific tumor receptors, further improve selective accumulation in GBM cells, ensuring a more precise and effective treatment.

PDT has emerged as a promising adjunctive treatment for malignant gliomas, with several clinical trials conducted worldwide. The contributions of research groups from Japan, Germany, Scotland, Canada, and Austria stand out. The contributions of Stummer’s German group to PDT in malignant gliomas are significant, particularly through their pioneering work on the use of 5-ALA for FGS. Their landmark study demonstrated that intraoperative fluorescence could enhance the resection of HGGs by clearly distinguishing tumor tissue from healthy brain tissue, which is crucial for improving surgical outcomes [121]. Stummer and his team conducted a phase III clinical trial that highlighted the benefits of 5-ALA-induced fluorescence, showing that it significantly increased the extent of tumor removal compared to conventional techniques [122]. The study reported improved PFS rates among patients treated with fluorescence-guided resection (FGR), establishing a new standard of care for glioma surgeries. Additionally, their research has explored the combination of PDT with CTX, further enhancing the efficacy of treatment regimens for malignant gliomas [122, 123]. This group examined the potential of PpIX as a serum marker for HGGs to monitor recurrence [124]. Patients [HGG: n = 23 pediatric (pHGG); n = 5 recurrent (rHGG)] who underwent FGR received 5-ALA following the standard clinical procedure. The control group of eight healthy volunteers (HCTR) also received 5-ALA. Serum was collected before and repeatedly up to 72 h after drug administration. A significant accumulation of PpIX in HGG was observed following 5-ALA administration [analysis of variance (ANOVA): p = 0.005, post-hoc: HCTR vs. pHGG p = 0.029, HCTR vs. rHGG p = 0.006]. Baseline PpIX levels were similar between patient and control groups. Therefore, 5-ALA is required for PpIX induction, which is safe at the standard clinical dose. PpIX may be a novel target for liquid biopsy in gliomas, so larger clinical studies are required to fully characterize its potential.

Another study by Pepper et al. [125], in which this PS was used, was the one in phase I/II conducted at the University Hospital of Münster, which evaluated the combination of oral 5-ALA and RT in patients with recurrent GBM. Thirty patients over 18 years old with histologically confirmed recurrent supratentorial GBM and good functional status (KPS ≥ 60) were recruited. Following a 3 + 3 dose-escalation design, patients who underwent repeat resection received RT at a dose of 36 Gy and fractions of PDT, which included oral 5-ALA administration before irradiation sessions. The study aimed to determine the maximum tolerated dose (MTD) of repeated 5-ALA administration, gradually increasing number of PDT fractions from one to eight, while closely monitoring safety and toxicity. Two cohorts received neoadjuvant treatment fractions before surgery. Follow-ups were conducted two and five months after treatment. The primary endpoints included the MTD, while secondary endpoints were event-free survival, PFS, and OS. Additionally, 5-ALA metabolites and tissue effects in resected samples were analyzed.

Despite the lack of improved outcomes with various other treatments, the application of 5-ALA as a radiosensitizer shows significant promise owing to its preferential absorption by glioma tumor cells, hence preserving normal brain tissue. Furthermore, 5-ALA is simple to administer and is typically well tolerated, as evidenced in the context of surgical procedures and PDT. This experiment was established to address recurrent GBM, due to the significant demand for effective salvage treatment alternatives in this patient population and the absence of a definitive treatment standard. In the event of favorable outcomes, additional assessment may be contemplated as an adjunct to conventional first-line therapy. Nevertheless, data regarding subsequent drug applications are not currently accessible. In the realm of brain tumor surgery, single doses of up to 60 mg/kg have been documented as well tolerated, thereby warranting a dose-escalation experiment with increments of 20 mg/kg. The researchers assert that the results from this inaugural single-center dose-escalation study in humans may provide a crucial instrument for treating GBM patients and herald a new era of enhanced RT efficacy against this highly aggressive tumor type.

The Japanese group of Kaneko et al. [126] conducted clinical trials concentrating on PDT for malignant gliomas. Their strategy comprised giving PSs such as 5-ALA before tumor excision and delivering intraoperative PDT by light diffusers or balloon devices inserted in the resection space. These trials indicated moderate improvements in survival, with PDT suggesting potential for better outcomes when coupled with surgical resection [127]. Another study assessed a different PS, talaporfin sodium, in a multicenter clinical trial investigating the efficacy and safety of intraoperative PDT utilizing talaporfin sodium in patients with primary malignant brain tumors, specifically GBM. The trial had 22 patients, demonstrating a 12-month OS rate of 95.5% and a 6-month PFS rate of 91%. The treatment was well tolerated, with adverse effects similar to standard neurosurgical practices, and no significant complications directly associated with the PDT [128]. However, the studies underlined the need for revised methods and further studies to enhance efficacy and safety of GBM treatment.

The Innsbruck group, under Kostron’s leadership, performed clinical experiments indicating that PDT with HpD can improve survival rates in patients with malignant gliomas [129]. The research demonstrated enhanced PFS and OS relative to conventional therapy. Patients exhibited little occurrence of serious adverse effects, reinforcing the safety and usefulness of PDT as a supplementary treatment to traditional therapies. The findings suggest that PDT may serve as a viable approach for the treatment of malignant gliomas. Nonetheless, the outcomes of cavitary PDT with HpD are promising, yet require a comprehensive phase III investigation. Another study investigated 5,10,15,20-tetrakis(3-hydroxyphenyl)chlorin (mTHPC)-mediated photodynamic diagnosis (PDD) and PDT for malignant brain tumors, notably a phase-II trial including 22 patients, which demonstrated that FGR utilizing mTHPC improves tumor detection and excision [130]. The study indicates a sensitivity of 87.9% and specificity of 95.7% for tumor detection, proposing that the integration of intraoperative visualization with therapy presents a promising approach to enhance surgical outcomes in brain cancer treatment.

Eljamel, a prominent Scottish researcher, has made significant contributions to the field of GBM treatment, particularly in PDT. His work focuses on developing and evaluating PDT as an adjuvant treatment for this challenging disease. Eljamel has conducted multiple clinical studies demonstrating the effectiveness of PDT combined with PSs like Photofrin^® in treating GBM. His research has shown that this therapy can extend patient survival, particularly in cases of recurrent GBM. In a prospective study, he evaluated FGR using ALA and Photofrin^® alongside repeat PDT sessions in GBM treatment. Twenty-seven patients were recruited (13 in the study group and 14 in the control group), with the study group achieving a median survival of 52.8 weeks compared to 24.6 weeks in the control group. FGR with ALA and Photofrin^® and repeat PDT provided a valuable survival benefit without additional risk to patients with GBM [131]. Another study included 73 patients with a median age of 59 years, 30 of whom received PDT and 43 did not. Median survival for patients treated with PDT was significantly higher than for those receiving standard therapy alone (62.9 weeks versus 20.6 weeks). Patients under 65 years of age had longer survival compared to those over 65. Intraoperative RT alone did not yield a significant survival benefit. However, the median survival of patients receiving PDT combined with intraoperative RT was greater than those receiving PDT alone. PDT for GBM was a statistically significant therapeutic modality, and its effects were further enhanced by combining it with intraoperative RT [132].

In another study, twenty patients with malignant brain tumors were administered 20 mg of ALA per kg of body weight orally, three hours before anesthesia. Surgery was conducted using image-guided systems and ALA-induced fluorescence microsurgical techniques. During the procedure, fluorescence intensity was categorized as red, pink, or blue and further measured using a 405 nm pulsed laser and a compact spectrometer with a contact probe placed on the tissue. The extent of tumor invasion was evaluated intraoperatively with standard white light, blue light, and spectroscopic measurements. The study revealed that fluorescence imaging showed GBM to be wider than previously indicated by contrast-enhanced magnetic resonance imaging (MRI), although metastases (MET)-enhanced MRI displayed a similar size to fluorescence imaging. Solid tumors were effectively identified and measured intraoperatively through fluorescence and spectroscopy, allowing for safe resection. Additionally, infiltrative tumor tissue could be identified and removed in non-eloquent brain areas, maximizing surgical outcomes [133].

Dr. Muller and Dr. Wilson’s team [134, 135] at St. Michael’s Hospital in Toronto has investigated PDT for the treatment of malignant brain tumors, particularly GBM and other primary brain tumors such as malignant astrocytoma (MA), malignant mixed glioma, and ependymoma (EP). In their clinical trials, they utilize adjuvant photodynamic treatment following tumor excision, employing a porphyrin-based PS that is injected prior to surgery. During the process, a specific device with an inflating balloon is utilized to administer light, ensuring optimal light distribution within the cavity formed by tumor removal, hence aiding in the eradication of leftover tumor cells. These studies demonstrate encouraging outcomes, indicating that certain patients experience extended intervals without recurrence following the combined treatment of PDT and resection. Furthermore, it concludes that patients receiving higher light doses exhibit prolonged survival relative to those receiving lower doses, and that there is no associated risk for patients who undergo PDT in conjunction with postoperative RT [134, 135].

Currently, PDT is being investigated as a complementary option for the treatment of GBM. Although there are very few direct comparative studies between PDT and other treatment modalities such as CTX, RT, or surgery, research has been conducted to evaluate the efficacy of PDT in combination with these treatments. An autopsy study was conducted on GBM patients who underwent PDT and surgical resection. This study assessed the therapeutic tissue depth of PDT using sodium talaporfin and a semiconductor laser in GBM, based on autopsy findings. Three patients—one newly diagnosed and two with recurrent GBM, received intraoperative PDT along with adjuvant therapies. While no local recurrence was observed in pre-mortem imaging, autopsy findings revealed tumor recurrence within the therapeutic depth in one case. Histopathological analysis showed PDT effects extending 9–18 mm from the resection cavity, characterized by glial scarring and immune infiltration. However, viable tumor cells were detected beyond this range, highlighting the need for strategies to enhance PDT efficacy in GBM [136].

While these studies offer promising insights into the use of complementary therapies such as PDT for GBM treatment, it is essential to conduct direct comparative clinical trials to establish their relative efficacy compared to standard treatment modalities.

One of the main components of PDT is the use of O₂ from the surrounding environment, which reacts with the PS employed. Therefore, solid tumors like GBM, which exhibit low O₂ tension, become one of the primary limitations of PDT [32]. Low O₂ concentrations, below 1 μM, in the TME underscore the challenges and reduced efficacy of PDT, as O₂ is essential for the therapy’s optimal performance [137]. Given that the impact of low O₂ tension on certain tumors has been recognized, negatively influencing PDT, research has demonstrated that modulating light fluence can favorably alter tissue oxygenation, thereby improving the therapeutic response [36, 138].

The regulation of redox regulatory pathways in tumor cells has been acknowledged as an effective method for their elimination [139]. The production of ROS, specifically ¹O₂, near the tumor site is essential for tumor eradication, as it facilitates focused and concentrated action of ¹O₂. However, PS will convert O₂ into ROS with cytotoxic potential, which causes O₂ consumption and further aggravates tumor hypoxia, resulting in decreased PDT action (Figure 4). On the other hand, the rapid growth of the tumor, O₂ consumption exceeds its supply, potentially leading to chronic hypoxia as the distance from the tumor-associated vasculature increases [140]. Therefore, in addition to pre-existing tumor hypoxia, the efficacy of PDT will be further impaired by the O₂ consumption involved in ROS generation [141]. Due to insufficient O₂, hypoxia triggers a multitude of intricate intracellular signaling cascades, including the hypoxia-inducible factor (HIF) pathway. Additional pathways linked to hypoxia encompass PI3K/protein kinase B (AKT)/mammalian target of rapamycin (mTOR), mitogen-activated protein kinase (MAPK) [often referred to as the extracellular signal-regulated kinase (ERK) pathway], nuclear factor kappa-light-chain-enhancer of activated B-cells (NFκB), and nuclear factor erythroid 2-related factor 2 (Nrf2) [142]. HIF-1 expression is linked to a worse prognosis in cancer patients, as it positively modulates the expression of genes related to metabolism, metastasis, angiogenesis, and immunosuppression [143]. Conversely, HIF-1a is increased by Nrf2 in hypoxic conditions, hence enhancing GBM cell survival. Nrf2 also enhances the expression of vascular endothelial growth factor (VEGF). Nrf2 demonstrates antiapoptotic, antioxidant, anti-inflammatory, and proliferative characteristics through various regulatory mechanisms [144]. Furthermore, Nrf2 is degraded in the proteasome by the action of Kelch-like ECH-associated protein 1 (KEAP1) under normal conditions (Nrf2 is ubiquitinated by the Cul3-KEAP1 E3 ubiquitin ligase complex), however, in the presence of ROS, KEAP1 is inactivated, allowing Nrf2 to be stable, and bind to antioxidant response elements, activating their cytoprotective functions [145]. Furthermore, oxidative stress-mediated activation of the transcription factor Nrf2 induces the expression of protective antioxidant genes. Nrf2-mediated regulation of the pentose phosphate pathway (PPP) affects glucose metabolism and ROS homeostasis in cancer cells [146, 147]. Studies have shown that TMZ treatment induces Nfr2 activation, and consequently, an increase in GSH concentration in tumor cells [148]. Among the main ROS detoxification mechanisms in GBM are superoxide dismutase (SOD), catalase and GPX. Furthermore, understanding the status of these antioxidant enzymes in GBM may prove key to the development of personalized therapies targeting ROS induction [93, 149]. GSH and its related enzymes are crucial in GBM for safeguarding against free radicals, radiation, CTX, and PDT [74, 93, 150]. The importance of the GSH redox cycle in drug resistance has been analyzed and it was found that TMZ-resistant cells had lower ROS and higher antioxidant capacity, with elevated levels of GSH reductase (GSR). GSR silencing increased cell sensitivity to TMZ, while its overexpression generated resistance. Modulation of redox status by GSR influenced drug resistance, suggesting that GSR is a potential therapeutic target in GBM. Furthermore, high GSR expression was associated with shorter PFS in patients [150].

Common risks of PDT encompass cutaneous and ocular photosensitivity subsequent to the administration of the PS. Nonetheless, these effects are transient and can be alleviated by evading direct sunlight exposure. A significant risk of PDT for brain tumors is uncontrolled cerebral edema, although its precise occurrence remains undefined, as it varies according to the type of PS utilized, the delivery technique, and the intensity of light applied during treatment [121].

A study assessed the efficacy of PDT utilizing HpD as an adjunctive treatment for brain gliomas. The median survival was 76.5 months for patients with anaplastic astrocytomas and 14.3 months for those with GBM. Advanced age correlated with a reduced prognosis, although laser dosages over 230 J/cm² indicated enhanced survival results. No fatalities were directly associated with PDT; nevertheless, three patients experienced cerebral edema, which was addressed with standard therapies. The occurrence of cerebral edema in these patients was about 0.04% after PDT with HpD [151]. In another study utilizing Photofrin^® at a dosage of 2 mg/kg, all patients exhibited good tolerance to the medication. Cerebral edema was noted in 46% of cases; however, in the majority of situations (10 cases), it was minor and did not necessitate particular intervention [126].

Modifications in lighting conditions and PS dosages are critical factors currently under investigation for each type of PS; these parameters are essential for inducing tumor cell mortality while preserving healthy tissue, mitigating complications such as cerebral edema, and preventing the emergence of treatment-resistant cells [36]. Animal experimental studies examine configurations employing surgically implanted LED lights to reliably and controllably illuminate the tumor site, enabling a metronomic application of PDT through smaller, repeated doses to enhance tumor cell destruction while reducing damage to healthy tissues [135].

Ps nanomaterials are very effective in generating ROS when exposed to light. While ROS can be beneficial in killing cancer cells, uncontrolled ROS production can damage cellular components, including DNA. ROS can induce DNA strand breakage, base alterations, and cross-linking, potentially resulting in mutations and chromosomal abnormalities in severe cases [152]. For instance, aluminium-phthalocyanine chloride tetra sulfonate (AlPcS₄Cl), a subclass of phthalocyanine, has been widely utilized in PDT for several tumor types, with its subcellular location determining the efficacy of PDT and the mechanism of cell death. A recent study revealed that AlPcS₄Cl, preferentially located in the nuclei, produced DNA double-strand breaks by upregulating ataxia telangiectasia mutated (ATM), a DNA damage sensor inducing cell death [153]. Nonetheless, the possible genotoxic consequences of certain PS, especially those with a predilection for nuclear localization, are increasingly concerning and require assessment. Genotoxicity denotes the capacity of a drug to inflict damage on DNA, potentially resulting in mutations, cancer, and other detrimental health consequences [154]. In the absence of a selective strategy to target tumor cells, PS may directly interact with DNA from normal or stem cells, particularly if they concentrate within the nucleus. This contact may result in physical damage to the DNA structure or disrupt DNA replication and repair mechanisms [155]. For NPs to exhibit primary genotoxicity, they must penetrate the cytosol or nuclear membrane and directly interact with DNA and its related proteins. Multiple studies have thoroughly reported the size-dependent entry of NPs and their accumulation outside endosomes and within certain cellular compartments, ultimately reaching the nuclei [156]. Nonetheless, in vitro research failed to accurately represent in vivo behavior. The distribution of PS NPs inside the body and their potential accumulation in specific tissues, such as the liver, spleen, and kidneys, may affect their genotoxic effects. From this viewpoint, PS accumulation in tissues with considerable regenerative capacity (e.g., bone marrow) might present an increased risk. The biodistribution of NP-based PS is affected by their chemical composition, sizes, and formulation, potentially leading to accumulation in bone marrow due to their small size and prolonged circulation time, hence enhancing their propensity for extravasation into highly vascularized tissues [157]. Nevertheless, limited research addresses bone marrow exposure to PS and assesses its potential harm [158, 159]. The bone marrow microenvironment, rich in hematopoietic stem cells (HSCs) and stromal cells, can sequester PS. This is particularly concerning because HSCs are highly sensitive to oxidative stress induced by PS-generated ROS. Various cell types found in different organs and tissues may be affected by NP-based PS and could be vulnerable to genotoxicity and cytotoxicity, including epithelial cells [160] and germ cells [161]. In the particular case of brain tumors, NPs can cause oxidative stress and DNA damage in neural stem cells, impairing their ability to differentiate into neurons and glial cells, leading to neurotoxicity [162].

While PS nanomaterials are highly effective in generating ROS for targeted cancer therapy, their potential to cause genotoxic and cytotoxic damage to normal and stem cells cannot be overlooked. To ensure the safe and effective use of PS nanomaterials, further research is essential to assess their long-term genotoxic and cytotoxic effects, particularly in vulnerable tissues, and to develop strategies that enhance tumor selectivity while minimizing harm to normal and stem cells.

A new direction emerges as encouraging results are obtained by combining different types of therapies, whether alternative or conventional. One therapeutic combination of interest is the application of PDT with various PSs and their integration with other therapies, whether primary or adjuvant. In recent years, synergistic effects have been observed in therapeutic combinations involving PDT, offering promising prospects for improved treatment outcomes. In this context, the following provides a brief overview of the most commonly used PS in PDT, as well as key studies on therapeutic combinations for the treatment of GBM (Figure 5).

Nanotechnology-based delivery systems hold immense promise for revolutionizing cancer therapy, particularly for highly aggressive and treatment-resistant malignancies like GBM. The integration of nanocarriers with PDT has already demonstrated significant advancements, including enhanced tumor targeting, reduced systemic toxicity, and multifunctional capabilities for theranostic applications. However, the road to clinical translation is fraught with challenges. Current hurdles, such as overcoming the BBB, addressing tumor-associated hypoxia, ensuring NP clearance, and reducing manufacturing complexities, must be systematically addressed. The heterogeneity of tumor vasculature and the intact BBB in certain glioma subtypes limit effective delivery of nanocarriers. Tumor-associated hypoxia, a hallmark of GBM, further reduces the efficacy of ROS-based therapies. Furthermore, insufficient clearance data on NPs from the body raises safety concerns, especially regarding potential long-term toxicity. Other barriers include the complexity and cost of large-scale NP production, the risk of off-target effects, and variability in therapeutic efficacy across different patients.

Future research should focus on designing smarter, m-NPs capable of adapting to the TME while ensuring patient safety and treatment efficacy. Innovative strategies, such as incorporating O₂-releasing mechanisms, hypoxia-activated PSs, and BBB-penetrating delivery systems, can significantly enhance therapeutic outcomes. The integration of molecular diagnostics with advanced nanotechnology could pave the way for personalized, precision-based therapies, tailoring treatments to individual patient profiles.

As the field progresses, collaboration between multidisciplinary teams of chemists, biologists, clinicians, and engineers will be crucial to overcome current limitations and unlock the full potential of nanomaterial-based delivery systems. With continued innovation and rigorous validation, these technologies could transform the landscape of cancer therapy, offering hope for improved survival and quality of life for patients with devastating diseases like GBM.

A field to explore is the economic evaluation of its implementation in the clinic. Many PSs have been developed in recent years that deserve validation and exploration in clinical trials for approval by regulatory agencies as an alternative or complement to conventional treatments.