Light Dosimetry for Intraperitoneal Photodynamic Therapy in a Murine Xenograft Model of Human Epithelial Ovarian Carcinoma

Few studies have been published to date measuring spatially resolved fluence rates in complex tissue geometries. Here the light distributions of three different intraperitoneal light delivery geometries in a murine ovarian cancer model were investigated to assess their influence on the tumorcidal efficacy of photodynamic therapy (PDT). In vivo fluence rate measurements in the peritoneal cavities of mice, with the light intensity being mapped in three transverse planes, were performed using fiber-optic detectors. Three different source fiber designs and placements were tested for their ability to provide uniform irradiation of the peritoneal cavity. The biological response to a PDT protocol comprising three separate treatments administered at 72 h intervals, each consisting of a 0.25 mg kg intraperitoneal injection of benzopor-phyrin derivative-mono acid ring A followed 90 min later by delivery of 15 J of 690 nm light, was measured. The tissue response was evaluated by measuring the number of remaining visible lesions and the total residual tumor mass. Fluence rate measurements showed large variations in the fluence rate distribution for similar intended treatments. The most uniform and reproducible illumination was achieved using two 18 mm long cylindrical emitting optical fibers. The biological response was comparable to that produced when a flat-cleaved end optical fiber is used to illuminate the four quadrants of the abdomen sequentially. While a good reproducibility in tumor induction in this animal model exists, no correlation was found between the fluence rate distribution measured in one group of animals and the biological response in a separate group of similarly treated animals. Due to the large intra-animal variability in fluence rate distribution, representative fluence rate mapping in complex tissue geometries is of limited value when applied to an individual PDT treatment. Thus, surveillance of the fluence rate during individual treatments will be required for acceptable PDT dosimetry. To improve the versatility of this particular animal model for PDT research, a large number of extended sources are required to increase uniformity of the illumination in order to reduce unwanted cytotoxic side effects resulting from foci of high fluence rates. In this way, subsequent increase of the total energy delivered to the tumor may be possible.     

Photodynamic Efficacy of Cercosporin in 3D Tumor Cell Cultures

In the present work, we study the photodynamic action of cercosporin (cerco), a naturally occurring photosensitizer, on human cancer multicellular spheroids. U87 spheroids exhibit double the uptake of cerco than T47D and T98G spheroids as shown by flow cytometry on the single cell level. Moreover, cerco is efficiently internalized by cells throughout the spheroid as shown by confocal microscopy, for all three cell lines. Despite their higher cerco uptake, U87 spheroids show the least vulnerability to cerco-PDT, in contrast to the other two cell lines (T47D and T98G). While 300 μm diameter spheroids consistently shrink and become necrotic after cerco PDT, bigger spheroids (>500 μm) start to regrow following blue-light PDT and exhibit high viability. Cerco-PDT was found to be effective on bigger spheroids reaching 1mm in diameter especially under longer exposure to yellow light (~590 nm). In terms of metabolism, T47D and T98G undergo a complete bioenergetic collapse (respiration and glycolysis) as a result of cerco-PDT. U87 spheroids also experienced a respiratory collapse following cerco-PDT, but retained half their glycolytic activity.     

A Combination of Visudyne and a Lipid‐anchored Liposomal Formulation of Benzoporphyrin Derivative Enhances Photodynamic Therapy Efficacy in a 3D Model for Ovarian Cancer

A major objective in developing new treatment approaches for lethal tumors is to reduce toxicity to normal tissues while maintaining therapeutic efficacy. Photodynamic therapy (PDT) provides a mechanistically distinct approach to treat tumors without the systemic toxicity of chemotherapy drugs. PDT involves the light‐based activation of a small molecule, a photosensitizer (PS), to generate reactive molecular species (RMS) that are toxic to target tissue. Depending on the PS localization, various cellular and subcellular components can be targeted, causing selective photodamage. It has been shown that targeted lysosomal photodamage followed by, or simultaneous with, mitochondrial photodamage using two different PS results in a considerable enhancement in PDT efficacy. Here, two liposomal formulations of benzoporphyrin derivative (BPD): (1) Visudyne (clinically approved) and (2) an in‐house formulation entrapping a lipid conjugate of BPD are used in combination with direct PS localization to mitochondria, endoplasmic reticulum and lysosomes, enabling simultaneous photodamage to all three organelles using a single wavelength of light. Building on findings by our group, and others, this study demonstrates, for the first time in a 3D model for ovarian cancer, that BPD‐mediated photodestruction of lysosomes and mitochondria/ER significantly enhances PDT efficacy at lower light doses than treatment with either PS formulation alone.     

Noninvasive Optical Imaging of UV‐Induced Squamous Cell Carcinoma in Murine Skin: Studies of Early Tumor Development and Vitamin D Enhancement of Protoporphyrin IX Production

Better noninvasive techniques are needed to monitor protoporphyrin IX (PpIX) levels before and during photodynamic therapy (PDT) of squamous cell carcinoma (SCC) of the skin. Our aim was to evaluate (1) multispectral fluorescent imaging of ultraviolet light (UV)‐induced cancer and precancer in a mouse model of SCC and (2) multispectral imaging and probe‐based fluorescence detection as a tool to study vitamin D (VD) effects on aminolevulinic acid (ALA)‐induced PpIX synthesis. Dorsal skin of hairless mice was imaged weekly during a 24‐week UV carcinogenesis protocol. Hot spots of PpIX fluorescence were detectable by multispectral imaging beginning at 14 weeks of UV exposure. Many hot spots disappeared after cessation of UV at week 20, but others persisted or became visible after week 20, and corresponded to tumors that eventually became visible by eye. In SCC‐bearing mice pretreated with topical VD before ALA application, our optical techniques confirmed that VD preconditioning induces a tumor‐selective increase in PpIX levels. Fluorescence‐based optical imaging of PpIX is a promising tool for detecting early SCC lesions of the skin. Pretreatment with VD can increase the ability to detect early tumors, providing a potential new way to improve efficacy of ALA‐PDT.     

Photobiomodulation Enhances Cisplatin Cytotoxicity in a Culture Model with Oral Cell Lineages

Cisplatin plays a central role in cancer chemotherapy, but resistance to this drug remains a major obstacle in treatment. Drawbacks related to cisplatin failure may be associated with cell energy metabolism. This study investigated whether photobiomodulation (PBM) can potentiate the effects of cisplatin on keratinocytes (HaCat) and cancer cells (SCC25 and HN12). Cells were treated with laser (GaAlAs; 660 nm; 60 mW; 0.33 J; 2.14 W cm⁻²; 11.7 J cm⁻² and 6 s) and cisplatin (7.8 μg mL⁻¹) to evaluate cell viability, Ki-67, VEGF, TGF-β1, EGF expression and ROS production. Observations were validated in the SCC25 cell lineage, where the type of cell death (necrosis/apoptosis) and the amount of ATP were assessed. Cell lineages showed increased sensitivity to cisplatin associated with PBM (Cis-PBM). Ki-67 was augmented in all cell lineages treated with Cis-PBM when compared to cisplatin alone (Cis). Cis or Cis-PBM significantly decreased VEGF expression in cancer cells, while no changes were seen in the expression of TGF-β1 or EGF compared to control. ROS levels were similar in the Cis and Cis-PBM groups. Cells treated with Cis-PBM died by apoptosis, leading to greater consumption of ATP. These observations suggest that PBM may potentiate the effects of cisplatin, leading to increased drug cytotoxicity and enhanced cell death.     

Photodynamic Therapy with Tumor Cell Discrimination through RNA-Targeting Ability of Photosensitizer

Photodynamic therapy (PDT) represents an effective treatment to cure cancer. The targeting ability of the photosensitizer is of utmost importance. Photosensitizers that discriminate cancer cells can avoid the killing of normal cells and improve PDT efficacy. However, the design and synthesis of photosensitizers conjugated with a recognition unit of cancer cell markers is complex and may not effectively target cancer. Considering that the total RNA content in cancer cells is commonly higher than in normal cells, this study has developed the photosensitizer QICY with RNA-targeting abilities for the discrimination of cancer cells. QICY was specifically located in cancer cells rather than normal cells due to their stronger electrostatic interactions with RNA, thereby further improving the PDT effects on the cancer cells. After intravenous injection into mice bearing a xenograft tumor, QICY accumulated into the tumor location through the enhanced permeability and retention effect, automatically targeted cancer cells under the control of RNA, and inhibited tumor growth under 630 nm laser irradiation without obvious side effects. This intelligent photosensitizer with RNA-targeting ability not only simplifies the design and synthesis of cancer-cell-targeting photosensitizers but also paves the way for the further development of highly efficient PDTs.     

Photodynamic Stromal Depletion Enhances Therapeutic Nanoparticle Delivery in 3D Pancreatic Ductal Adenocarcinoma Tumor Models

Pancreatic ductal adenocarcinoma (PDAC) is among the most lethal of human malignancies. PDAC is characterized by dense fibrous stroma which obstructs drug delivery and plays complex tumor-promoting roles. Photodynamic therapy (PDT) is a light-based modality which has been demonstrated to be clinically feasible and effective for tumors of the pancreas. Here, we use in vitro heterocellular 3D co-culture models in conjunction with imaging, bulk rheology and microrheology to investigate photodegradation of non-cellular components of PDAC stroma (photodynamic stromal depletion, PSD). By measuring the rheology of extracellular matrix (ECM) before and after PDT we find that softening of ECM is concomitant with increased transport of nanoparticles (NPs). At the same time, as shown by us previously, photodestruction of stromal fibroblasts leads to enhanced tumor response to PDT. Here we specifically evaluate the capability of PSD to enhance RNA nanomedicine delivery, using a NP carrying an inhibitor of miR-21-5P, a PDAC oncomiR. We confirm improved delivery of this therapeutic NP after PSD by observation of increased expression of PDCD4, a protein target of miR-21-5P. Collectively, these results in 3D tumor models suggest that PSD could be developed to enhance delivery of other cancer therapeutics and improve tumor response to treatment.     

Photodynamic Mechanisms induced by a Combination of Hypericin and a Chlorin Based‐Photosensitizer in Head and Neck Squamous Cell Carcinoma Cells

The aim of this study was to elucidate photodynamic therapy (PDT) effects mediated by hypericin and a liposomal meso‐tetrahydroxyphenyl chlorin (mTHPC) derivative, with focus on their 1:1 mixture, on head and neck squamous cell carcinoma cell lines. Absorption, excitation and photobleaching were monitored using fluorescence spectrometry, showing the same spectral patterns for the mixture as measured for single photosensitizers. In the mixture mTHPC showed a prolonged photo‐stability. Singlet oxygen yield for light‐activated mTHPC was Φ_Δ = 0.66, for hypericin Φ_Δ = 0.25 and for the mixture Φ_Δ = ~0.4. A linear increase of singlet oxygen yield for mTHPC and the mixture was found, whereas hypericin achieved saturation after 35 min. Reactive oxygen species fluorescence was only visible after hypericin and mixture‐induced PDT. Cell viability was also more affected with these two treatment options under the selected conditions. Examination of death pathways showed that hypericin‐mediated cell death was apoptotic, with mTHPC necrotic and the 1:1 mixture showed features of both. Changes in gene expression after PDT indicated strong up‐regulation of selected heat‐shock proteins. The application of photosensitizer mixtures with the features of reduced dark toxicity and combined apoptotic and necrotic cell death may be beneficial in clinical PDT. This will be the focus of our future investigations.     

Photodynamic Therapy Mediated by 5‐aminolevulinic Acid Promotes the Upregulation and Modifies the Intracellular Expression of Surveillance Proteins in Oral Squamous Cell Carcinoma

Expression of proteins related to cell surveillance has been described in tumors presenting resistance to photodynamic therapy (PDT). The aim of this study was to verify whether there was upregulation of proteins related to resistance in oral squamous cell carcinoma (OSCC) after PDT. OSCC was chemically induced in rats and treated after one cycle of PDT mediated by 5‐aminolevulinic acid (5‐ALA‐PDT). Immunolabeling of p‐NFκB, Bcl‐2, survivin, iNOS, p‐Akt, p‐mTOR and cyclin D1 was performed after the treatment. There was increased expression of Bcl‐2 (P = 0.008), iNOS (P = 0.020), p‐Akt (P = 0.020) and p‐mTOR (P = 0.010) by surviving neoplastic cells after PDT when compared to the control. In conclusion, after one cycle of 5‐ALA‐mediated PDT, Bcl‐2, p‐Akt, p‐mTOR and iNOS were upregulated in neoplastic cells of OSCC, suggesting an activation of antiapoptosis and cell proliferation pathways. This fact must be considered in the establishment of PDT protocols for OSCC treatment, mainly those in which PDT will be combined with chemotherapy drugs targeted at the studied proteins.     

MiR-335在口腔鳞状细胞癌中的表达变化

目的探明miR-335在口腔鳞状细胞癌组织中的表达异常情况。方法采用qRT-PCR分别检测25例人口腔鳞状癌细胞及相应邻近正常黏膜组织中miR-335的表达情况,对miR-335在口腔鳞状细胞癌及相应邻近正常黏膜组织中的表达情况进行比较。结果 miR-335在口腔鳞状细胞癌中的表达显著低于其癌旁正常黏膜组织(P0.05)。结论 miR-335在舌鳞癌中呈低表达,有可能在口腔鳞状细胞癌的发生、发展起重要作用。     

Targeted and Oxygen-Enriched Nanoplatform for Enhanced Photodynamic Therapy: In Vitro 2D Cell and 3D Spheroid Model Evaluation

Hypoxic microenvironment and limited penetration of photosensitizers within solid tumors are two crucial factors that restrict photodynamic therapy (PDT) efficacy. Herein, a new fluorinated mixed micelle ( M60@PFC-Ce6 ) is developed as a tumor-penetrating and oxygen-enriching nanoplatform, which consists of chlorin e6 (Ce6) and perfluorocarbons (PFCs) co-loaded into fluorinated micelles to relieve hypoxia conditions as well as folate as targeting ligand that facilitates the selective biodistribution within tumor solids. The incorporation of fluorinated copolymers into mixed micelles exhibits not only a great increase in the oxygen-loading capacity, but also improves the stability of liquid PFCs emulsion within micelles without leakage. M60@PFC-Ce6 shows excellent oxygen delivery capability, good intracellular reactive oxygen species (ROS) generation, and superior phototoxicity in vitro for both 2D monolayer of cells and 3D multicellular spheroid model. These results indicate the enriched oxygen delivery and increased cellular uptake resulting from folate-targeted ability to enhance ROS production and PDT efficacy. The penetration study of M60@PFC-Ce6 into a 3D spheroid confirms that small micellar size and folate-conjugation are beneficial for micelles to penetrate and accumulate within spheroids. Thus, a new nanoplatform with enriched oxygen-carrying amounts, better drug penetration, and stable micellar properties that relieve tumor hypoxia and improve PDT efficacy is provided.     

Effects of Sodium Butyrate on Cell Death Induced by Photodynamic Therapy in U373-MG and D54-MG Astrocytoma Cell Lines

The damage induced by end products of photodynamic therapy (PDT) in astrocytoma tumors leads to cytotoxicity and cell death. Chromatin modifiers such as sodium butyrate (NaB) induce several genes involved in apoptosis, among others. The PDT improvement was evaluated by the measurement of its effectiveness in the treatment of U373-MG and D54-MG astrocytoma cell lines exposed to NaB. Cells exposed to 80 μg mL⁻¹ of δ-aminolevulinic acid (ALA) as precursor of endogenous photosensitizer (PS), protoporphyrin IX (PpIX), induced 16.67% and 28.9% of mortality in U373-MG and D54-MG, respectively. The mortality increased to 70.62% and 96.7%, respectively, when U373-MG and D54-MG cells were exposed for 24 h to 8 m m NaB prior to ALA-induction. In this condition, re-expression of some genes related to apoptosis in U373-MG, and differentiation in D54-MG were induced. PpIX accumulation was higher than ALA-induction and the acetylation of histone H4 induced by NaB was verified by immunocytochemistry in both cells. It can be concluded that modified chromatin and genes induced by NaB increment the cellular death induced by PDT in astrocytoma cells using PpIX as endogenous PS.     

Targeted Photodynamic Therapy of Human Head and Neck Squamous Cell Carcinoma with Anti-epidermal Growth Factor Receptor Antibody Cetuximab and Photosensitizer IR700DX in the Mouse Skin-fold Window Chamber Model

Targeted photodynamic therapy (PDT) in head/neck cancer patients with a conjugate of the anti-epidermal growth factor receptor (EGFR) antibody, Cetuximab and a phthalocyanine photosensitizer IR700DX is under way, but the exact mechanisms of action are still not fully understood. In this study, the EGFR-overexpressing human head/neck OSC-19-luc2-cGFP tumor with transfected GFP gene was used in a skin-fold window chamber model in BALB/c nude mice. The uptake and localization of the conjugate in the tumor and its surrounding normal tissues were studied by an intravital confocal laser scanning microscopy with image analyses. The tumor was also irradiated with 690 nm laser light 24 h after conjugate administration. The vascular and tumor responses were examined by morphological evaluation and immunohistochemistry (IHC). The amount of conjugate in the tumor peaked at 24–48 h after injection. Image analyses of colocalization correlation parameters demonstrated a high fraction of the conjugate IR700DX colocalized in the GFP-expressing tumor cells. PDT-treated tumors showed extensive necrotic/apoptotic destruction with little vascular damage, while IHC showed no HIF-1α expression and decreased EGFR and Ki67 expression with activated caspase-3 overexpression, indicating a direct killing of tumor cells through both necrotic and apoptotic cell death.     

Magnetic Resonance Imaging Evaluation of Photodynamic Therapy-Induced Hemorrhagic Necrosis in the Murine M1 Tumor Model

Abstract— Proton magnetic resonance imaging (MRI) and histological methods were used to evaluate photodynamic therapy (PDT)-induced hemorrhagic necrosis in the murine Ml tumor within 72 h of treatment of male DBA/2 mice. The effects of three photosensitizing drugs were investigated: Photofrin (n = 4), Zn (II) phthalocyanine (n = 7) and benzoporphyrin derivative monoacid ring A (n = 11). As noted in previous studies of PDT using MRI, MRI makes possible serial, noninvasive, in vivo observation of tissue response to PDT. Our serial study of MRI and histological data confirms that tumors responded in the same way to PDT treatment using the three photosensitizing drugs: vascular damage followed by hemorrhagic necrosis. Most importantly and unlike previous MRI studies of PDT, we used a very high field magnet that enhanced the effect of magnetic susceptibility on image signal when blood is processed by the body after PDT-induced hemorrhagic necrosis. This last finding demonstrates the utility of high field magnets and the importance of localized, serial experiments in future magnetic resonance studies of PDT.     

Mutagenicity of Photodynamic Therapy as Compared to UVC and Ionizing Radiation in Human and Murine Lymphoblast Cell Lines

Abstract— The mutagenicity of photodynamic therapy (PDT) using red light and either Photofrin® (porfimer sodium) (PF) or aluminum phthalocyanine (AIPc) as the photosensitizer was determined at the thymidine kinase (TK) locus in the human lymphoblastic cell lines, TK6 and WTK1, and was compared to the mutagenicity of UVC and X-radia-tion in these cells as well as the mutagenicity of PDT in murine L5178Y lymphoblastic cell lines. Photodynamic therapy was found not to be mutagenic in TK6 cells, which possess an active p53 gene and which are relatively deficient in recombination and repair of DNA double-strand breaks. In contrast, PDT with either sensitizer was significantly mutagenic in WTK1 cells, which harbor an inactivating mutation in the p53 gene and are relatively efficient in recombination and double-strand break repair as compared to TK6 cells. The induced mutant frequency in WTK1 cells with PF as the photosensitizer was similar to that induced by UVC radiation but lower than that induced by X-radiation at equitoxic faiences/ doses. The mutant frequency induced by PDT in WTK1 cells with either photosensitizer was much lower than that induced in murine lymphoblasts at equitoxic fluences. The TK6 and WTK1 cells did not differ in their sensitivity to the cytotoxic effects of PDT, but the level of PDT-induced apoptosis was greater in TK6 than in WTK1 cells. These results indicate that the mutagenicity of PDT varies in different types of cells and may be related to the repair capabilities as well as the p53 status of the cells.     

Meta-analysis of gene expression for development and validation of a diagnostic biomarker panel for Oral Squamous Cell Carcinoma

We use a newly developed feature extraction and classification method to analyze previously published gene expression data sets in Oral Squamous Cell Carcinoma and in healthy oral mucosa in order to find a gene set sufficient for diagnoses. The feature selection technology is based on the relative dichotomy power concept published by us earlier. The resulting biomarker panel has 100% sensitivity and 95% specificity, is enriched in genes associated with oncogenesis and invasive tumor growth, and, unlike marker panels devised in earlier studies, shows concordance with previously published marker genes.     

3D Culture Models of Malignant Mesothelioma Reveal a Powerful Interplay Between Photodynamic Therapy and Kinase Suppression Offering Hope to Reduce Tumor Recurrence

In this issue, Cramer et al. introduce 3D culture models of metastatic mesothelioma to investigate basic cancer biology and new combination therapies for combating this complex and lethal disease. The results suggest that erlotinib‐enhanced photodynamic therapy could further improve the efficacy of intraoperative light‐activation to mop up residual tumor deposits in the clinic following surgical removal of macroscopic mesothelioma metastases.     

Modeling Epidermal Growth Factor Inhibitor‐mediated Enhancement of Photodynamic Therapy Efficacy Using 3D Mesothelioma Cell Culture

We have demonstrated that lung‐sparing surgery with intraoperative photodynamic therapy (PDT) achieves remarkably extended survival for patients with malignant pleural mesothelioma (MPM). Nevertheless, most patients treated using this approach experience local recurrence, so it is essential to identify ways to enhance tumor response. We previously reported that PDT transiently activates EGFR/STAT3 in lung and ovarian cancer cells and inhibiting EGFR via erlotinib can increase PDT sensitivity. Additionally, we have seen higher EGFR expression associating with worse outcomes after Photofrin‐mediated PDT for MPM, and the extensive desmoplastic reaction associated with MPM influences tumor phenotype and therapeutic response. Since extracellular matrix (ECM) proteins accrued during stroma development can alter EGF signaling within tumors, we have characterized novel 3D models of MPM to determine their response to erlotinib combined with Photofrin‐PDT. Our MPM cell lines formed a range of acinar phenotypes when grown on ECM gels, recapitulating the locally invasive phenotype of MPM in pleura and endothoracic fascia. Using these models, we confirmed that EGFR inhibition increases PDT cytotoxicity. Together with emerging evidence that EGFR inhibition may improve survival of lung cancer patients through immunologic and direct cell killing mechanisms, these results suggest erlotinib‐enhanced PDT may significantly improve outcomes for MPM patients.     

Distribution and Excretion of Radiolabeled Temoporfin in a Murine Tumor Model

The biodistribution and excretion of temoporfin (tetra[ m -hydroxyphenyl]chlorin, m -THPC), a recently developed photosensitizer, was investigated in BALB/c mice. [¹⁴C]temoporfin was administered intravenously (0.73 μmol/kg) to tumor-free mice or to mice implanted with the Colo 26 colorectal carcinoma. Blood, tissue and fecal samples were collected for 35 days and 10 days postdose from tumor-free mice and tumor-bearing mice, respectively. Blood concentrations fell rapidly such that at later time points they were indistinguishable from background counts. Tumor concentrations rose to a peak of 0.34 μg temoporfin equivalents/mL at 2 days and then declined in parallel (log plot) with the blood concentrations. Tumor : tissue ratios at 2 days for skin, adipose tissue and skeletal muscle underlying the tumor were 1.5, 2.3 and 3.8, respectively. By 4 days the corresponding values were 1.6, 3.4 and 4.0. Nearly 40% of the administered radioactivity was excreted in the feces in the first 24 h and more than 80% had been excreted by 20 days. Less than 0.2 % of the dose was recovered from the urine. An elimination half-life of 10–12 days was calculated from the excretion data.     

Measurement of Tumor Vascular Damage in Mice with 99m Tc-MIBI Following Photodynamic Therapy

Abstract— The clinical perfusion agent ^99mTc-MIBI was used to monitor changes in tumor vascular perfusion (TVP) induced by Photofrin® (Pll)-mediated photodynamic therapy (PDT). BALB/c mice bearing an EMT-6 tumor on each hind thigh were given an intravenous injection of 1, 2 or 5 mg kg⁻¹ PII. Twenty-four hours later, one tumor was illuminated (600–650 run, 200 mW cm⁻² 400 J cm⁻²) while the other served as a control. At various time intervals after PDT (0, 2 and 24 h) mice received an intravenous injection of ^99mTc-hexakismethoxy(sobutyusonitri-le (MIBI) (0.18 MBq g⁻¹) and were sacrificed 2 min later. The light-treated and the untreated tumors were then dissected, the radioactivity was counted and the percentage of the injected dose per gram of tumor (%ID g⁻¹) was calculated as a measure of TVP. We observed that TVP is drug dose dependent, develops progressively with time post-PDT and is inversely related to PDT efficacy. Our data show that early tumor retention of ^99mMIBI is a simple method to assess TVP and vascular damage induced by PDT.