Photophysical and photobiological properties of a sulfonated chlorin photosensitiser TPCS(2a) for photochemical internalisation (PCI).

This study investigated the photophysical and photobiological properties of a new amphiphilic chlorin photosensitiser, disulfonated tetraphenylchlorin (TPCS(2a)), for photochemical internalisation (PCI). The absorption and fluorescence spectra of TPCS(2a) were examined in a range of solvents together with fluorescence lifetime measurements. The fluorescence lifetime of TPCS(2a) was found to be 8.5 ns in methanol, whereas non-exponential decays were observed in distilled water due to sensitiser dimerisation. The singlet oxygen quantum yield of TPCS(2a) was determined as 0.62 in deuterated methanol by direct observation of singlet oxygen phosphorescence. In a human oral squamous carcinoma (HN5) cell line, intracellular co-localisation of TPCS(2a) and Alexa488-labelled saporin, a macromolecular toxin, was observed corresponding predominantly to a lysosomal distribution. Intracellular fluorescence redistribution of TPCS(2a) and Alexa488-saporin was observed after 405 nm irradiation. Using two-photon confocal microscopy at 840 nm, and fluorescence lifetime imaging (FLIM), the lifetime was measured as 6 ns in HN5 cells. PCI using TPCS(2a) was shown to be very effective, and a synergistic increase in saporin toxicity was achieved in HN5 cells where viability was significantly reduced after light exposure compared to saporin (25 nM) treatment alone. The results demonstrate the favourable photophysical and photobiological properties of TPCS(2a) for PCI, which induces the relocalisation of a macromolecular anti-cancer toxin inside cells and significantly enhances cell death.

Clinical assessment of a low-cost, hand-held, smartphone-attached intraoral imaging probe for 5-aminolevulinic acid photodynamic therapy monitoring and guidance.

Significance: India has one of the highest rates of oral squamous cell carcinoma (OSCC) in the world, with an incidence of 15 per 100,000 and more than 70,000 deaths per year. The problem is exacerbated by a lack of medical infrastructure and routine screening, especially in rural areas. New technologies for oral cancer detection and timely treatment at the point of care are urgently needed. Aim: Our study aimed to use a hand-held smartphone-coupled intraoral imaging device, previously investigated for autofluorescence (auto-FL) diagnostics adapted here for treatment guidance and monitoring photodynamic therapy (PDT) using 5-aminolevulinic acid (ALA)-induced protoporphyrin IX (PpIX) fluorescence (FL). Approach: A total of 12 patients with 14 buccal mucosal lesions having moderately/well-differentiated micro-invasive OSCC lesions (<2 cm diameter and <5 mm depth) were systemically (in oral solution) administered three doses of 20 mg/kg ALA (total 60 mg/kg). Lesion site PpIX and auto-FL were imaged using the multichannel FL and polarized white-light oral cancer imaging probe before/after ALA administration and after light delivery (fractionated, total 100 J/cm2 of 635 nm red LED light). Results: The handheld device was conducive for access to lesion site images in the oral cavity. Segmentation of ratiometric images in which PpIX FL is mapped relative to auto-FL enabled improved demarcation of lesion boundaries relative to PpIX alone. A relative FL (R-value) threshold of 1.4 was found to segment lesion site PpIX production among the patients with mild to severe dysplasia malignancy. The segmented lesion size is well correlated with ultrasound findings. Lesions for which R-value was >1.65 at the time of treatment were associated with successful outcomes. Conclusion: These results indicate the utility of a low-cost, handheld intraoral imaging probe for image-guided PDT and treatment monitoring while also laying the groundwork for an integrated approach, combining cancer screening and treatment with the same hardware.

How mainstream medicine sees photodynamic therapy in the United Kingdom.

Although photodynamic therapy (PDT) has been used successfully to treat an assortment of different types of cancer, it has yet to reach the level of mainstream medicine on either side of the Atlantic. Unsubstantiated claims of PDT's efficacy in the past may be part of the reason for this. However, perhaps the main obstacle to PDT's endorsement by conventional medicine is the limited number of high-quality randomized controlled trials (RCTs) comparing it with relevant comparators for all meaningful outcomes, including effectiveness, safety, adverse events, quality of life, survival, and cost. Based on a Health Technology Assessment report on the current status of PDT and consultation with professional groups, specialist societies, and clinical study groups in the United Kingdom, this article explores the current clinical guidelines for use of PDT in cancer treatment and the dearth of supportive data from RCTs.

Photodynamic therapy for prostate cancer--an emerging approach for organ-confined disease.

Since the first report of the use of photodynamic therapy (PDT) for prostate cancer in 1990 it has been investigated as a primary and a salvage treatment, using either whole gland or focal approaches. Since 1990 advances in the transperineal approach to the prostate, coupled with photosensitizers which have a short drug-light interval and minimal skin phototoxicity, have resulted in major advances in the field. This review will look at the work done to date, and the ongoing studies which help to define the place of PDT as a useful treatment modality for organ-confined prostate cancer.

Photodynamic Therapy in Primary Breast Cancer.

Photodynamic therapy (PDT) is a technique for producing localized necrosis with light after prior administration of a photosensitizing agent. This study investigates the nature, safety, and efficacy of PDT for image-guided treatment of primary breast cancer. We performed a phase I/IIa dose escalation study in 12 female patients with a new diagnosis of invasive ductal breast cancer and scheduled to undergo mastectomy as a first treatment. The photosensitizer verteporfin (0.4 mg/kg) was administered intravenously followed by exposure to escalating light doses (20, 30, 40, 50 J; 3 patients per dose) delivered via a laser fiber positioned interstitially under ultrasound guidance. MRI (magnetic resonance imaging) scans were performed prior to and 4 days after PDT. Histological examination of the excised tissue was performed. PDT was well tolerated, with no adverse events. PDT effects were detected by MRI in 7 patients and histology in 8 patients, increasing in extent with the delivered light dose, with good correlation between the 2 modalities. Histologically, there were distinctive features of PDT necrosis, in contrast to spontaneous necrosis. Apoptosis was detected in adjacent normal tissue. Median follow-up of 50 months revealed no adverse effects and outcomes no worse than a comparable control population. This study confirms a potential role for PDT in the management of early breast cancer.

Clinical evaluation of smartphone-based fluorescence imaging for guidance and monitoring of ALA-PDT treatment of early oral cancer.

SIGNIFICANCE: India has one of the highest rates of oral cancer incidence in the world, accounting for 30% of reported cancers. In rural areas, a lack of adequate medical infrastructure contributes to unchecked disease progression and dismal mortality rates. Photodynamic therapy (PDT) has emerged as an effective modality with potential for treating early stage disease in resource-limited settings, while photosensitizer fluorescence can be leveraged for treatment guidance. AIM: Our aim was to assess the capability of a simple smartphone-based device for imaging 5-aminolevulinic acid (ALA)-induced protoporphyrin IX (PpIX) fluorescence for treatment guidance and monitoring as part of an ongoing clinical study evaluating low-cost technology for ALA-based PDT treatment of early oral cancer. APPROACH: A total of 29 subjects with <2 cm diameter moderately/well-differentiated microinvasive ( < 5 mm depth) oral squamous cell carcinoma lesions (33 lesions total, mean area ∼1.23 cm2) were administered 60 mg / kg ALA in oral solution and imaged before and after delivery of 100 J / cm2 total light dose to the lesion surface. Smartphone-based fluorescence and white light (WL) images were analyzed and compared with ultrasound (US) imaging of the same lesions. RESULTS: We present a comparative analysis of pre- and post-treatment fluorescence, WL, and US images of oral lesions. There was no significant difference in the distribution of lesion widths measured by fluorescence and US (mean widths of 14.5 and 15.3 mm, respectively) and linear regression shows good agreement (R2 = 0.91). In general, PpIX fluorescence images obtained prior to therapeutic light delivery are able to resolve lesion margins while dramatic photobleaching (∼42 % ) is visible post-treatment. Segmentation of the photobleached area confirms the boundaries of the irradiated zone. CONCLUSIONS: A simple smartphone-based approach for imaging oral lesions is shown to agree in most cases with US, suggesting that this approach may be a useful tool to aid in PDT treatment guidance and monitoring photobleaching as part of a low-cost platform for intraoral PDT.

Optimal conditions for successful ablation of high-grade dysplasia in Barrett's oesophagus using aminolaevulinic acid photodynamic therapy.

Photodynamic therapy (PDT) using 5-aminolaevulinic acid (ALA-PDT) is an attractive alternative to PDT with porfimer sodium for the treatment of high-grade dysplasia (HGD) in Barrett's oesophagus (BO) because of the shorter duration of light photosensitivity and low risk of oesophageal stricture formation. Published results, however, show marked variation in its efficacy, and optimum treatment parameters have not been defined. This study investigated how the dose of ALA and the colour of the illuminating light influenced the biological effect. Twenty-seven patients were enrolled into a randomised controlled trial of red versus green (635 nm or 512 nm) laser light activation for the eradication of HGD with ALA-PDT in Barrett's oesophagus. A further 21 patients were subsequently treated with the most effective regimen. Regular endoscopic follow-up with quadrantic biopsies every 2 cm was performed. The primary outcome measure was eradication of HGD. Patient's receiving ALA at 30 mg/kg relapsed to HGD more than those receiving 60 mg/kg (P = 0.03). Additionally, for those treated with ALA 60 mg/kg, red laser light was more effective than green laser light (P = 0.008). Kaplan-Meier analysis of the 21 patients who were subsequently treated with this optimal regimen demonstrated an eradication rate of 89% for HGD and a cancer-free proportion of 96% at 36 months' follow-up. Using an ALA dose of 60 mg/kg activated by 1,000 J/cm red laser light, we found that ALA-PDT was a highly effective treatment for high-grade dysplasia in Barrett's oesophagus.

Elastic scattering spectroscopy for early detection of breast cancer: partially supervised Bayesian image classification of scanned sentinel lymph nodes.

Sentinel lymph node biopsy is a standard diagnosis procedure to determine whether breast cancer has spread to the lymph glands in the armpit (the axillary nodes). The metastatic status of the sentinel node (the first node in the axillary chain that drains the affected breast) is the determining factor in surgery between conservative lumpectomy and more radical mastectomy including axillary node excision. The traditional assessment of the node requires sample preparation and pathologist interpretation. An automated elastic scattering spectroscopy (ESS) scanning device was constructed to take measurements from the entire cut surface of the excised sentinel node and to produce ESS images for cancer diagnosis. Here, we report on a partially supervised image classification scheme employing a Bayesian multivariate, finite mixture model with a Markov random field (MRF) spatial prior. A reduced dimensional space was applied to represent the scanning data of the node by a statistical image, in which normal, metastatic, and nonnodal-tissue pixels are identified. Our results show that our model enables rapid imaging of lymph nodes. It can be used to recognize nonnodal areas automatically at the same time as diagnosing sentinel node metastases with sensitivity and specificity of 85% and 94%, respectively. ESS images can help surgeons by providing a reliable and rapid intraoperative determination of sentinel nodal metastases in breast cancer.

Photodynamic therapy of malignant biliary strictures using meso-tetrahydroxyphenylchlorin.

OBJECTIVES: The palliation of patients with malignant bile duct obstruction using metal or plastic biliary stents may be limited by stent occlusion. The aim of this study was to determine the safety and efficacy of endoscopically delivered meso-tetrahydroxyphenyl chlorin photodynamic therapy in the treatment of irresectable malignant biliary strictures and recurrent stent occlusion. METHODS: Thirteen patients with malignant biliary obstruction owing to carcinoma of the biliary tract (n=9), pancreas (n=3) or stomach (n=1), were studied. All had been initially palliated with metal (n=10) or polyethylene (n=3) biliary stents, but presented with recurrent obstructive jaundice because of local tumour progression. Patients received meso-tetrahydroxyphenyl chlorin 0.15 mg/kg intravenously 72 h before endoluminal light activation with an endoscopically placed optical fibre, followed by polyethylene stent insertion. RESULTS: Before photodynamic therapy, patients had a median of three (range 0-5) stent occlusions in the preceding 11 (2-22) months, with a median patency of plastic stents placed inside metal bile duct stents for recurrent stent occlusion of 3.5 (0.5-13) months. After photodynamic treatment, tumour necrosis and/or metal stent recanalization was seen in all patients, with a median of 0 (0-3) stent occlusions during 7 (1-43) months follow-up. The median patency of plastic stents placed inside metal stents after photodynamic therapy was 5 (1-43) months. The median survival after diagnosis and photodynamic therapy administration was 21 (10-56) and 8 (1-43) months, respectively. Photodynamic therapy was generally well tolerated but two patients developed cholangitis within the first week, complicated in one by a fatal liver abscess and two developed haemobilia within 4 weeks of treatment, one of whom died with a gall bladder empyema. CONCLUSION: In patients with malignant biliary obstruction, endoscopically delivered meso-tetrahydroxyphenyl chlorin photodynamic therapy causes efficient tumour necrosis and recanalization of blocked metal stents, but there is a significant risk of complications.

The vascular response to photodynamic therapy with ATX-S10Na(II) in the normal rat colon.

The mechanism of tissue damage from photodynamic therapy (PDT) may be cellular, vascular or both, depending on the photosensitising agent and the treatment conditions. Well established photosensitisers like porfimer sodium have an optimum drug light interval of two days and may cause skin photosensitivity lasting several weeks. ATX-S10Na(II) is a new photosensitiser that remains largely in the vasculature after systemic administration and clears from the body within a few hours. The present study looks at the factors controlling the extent of PDT necrosis using ATX-S10Na(II) and correlates these with changes in the circulation after PDT. Normal Wistar rats were sensitised with ATX-S10Na(II), 2 mg/kg. At laparotomy, a laser fibre was positioned just touching the colonic mucosa and 50 J light at 670 nm delivered varying the drug light interval (0.5-24 h) and light delivery regime (100 mW continuous, 20 mW continuous or 100 mW in five fractions). Some animals were killed at three days to document the area of necrosis, others received fluorescein shortly prior to death (from a few minutes to three days after PDT) to outline the zone of PDT induced vascular shutdown. Maximum necrosis was seen with the shortest drug light interval (0.5 h), with no effect by 6 h. Fractionating the light or lowering the power did not increase the necrosis. The area of fluorescein exclusion increased over the first 2 h after PDT (in contrast to the re-perfusion seen with other photosensitisers) and correlated with the area of necrosis. PDT with ATX-S10Na(II) is most effective with a drug light interval of less than one hour. It induces irreversible vascular shutdown that extends after completion of light delivery and which is largely independent of the light delivery regime.

Photodynamic therapy for pancreatic and biliary tract carcinoma.

The prognosis of patients with pancreatic and biliary tract cancer treated with conventional therapies such as stent insertion or chemotherapy is often poor, and new approaches are urgently needed. Surgery is the only curative treatment but is appropriate in less than 20% of cases, and even then it is associated with a 5-yr survival of less than 30% in selected series. Photodynamic therapy represents a novel treatment for pancreaticobiliary malignancy. It is a way of producing localized tissue necrosis with light, most conveniently from a low-power, red laser, after prior administration of a photosensitizing agent, thereby initiating a non-thermal cytotoxic effect and tissue necrosis. This review outlines the mechanisms of action of photodynamic therapy including direct cell death, vascular injury, and immune system activation, and summarizes the results of preclinical and clinical studies of photodynamic therapy for pancreaticobiliary malignancy.

Spectroscopic sensing of cancer and cancer therapy: current status of translational research.

Various types of optical spectroscopy have been investigated as methods to effect a non-invasive, real-time in-situ assessment of tissue pathology. All of these methods have one basic principle in common: the optical spectrum of a tissue contains information about the biochemical composition and/or the structure of the tissue, and that information conveys diagnostic information. The biochemical information can be obtained by measuring absorption, fluorescence, or Raman scattering signals. Structural and morphological information may be obtained by techniques that assess the elastic-scattering properties of tissue. These basic approaches are useful for the detection of cancer as well as for other diagnostic applications such as hemoglobin saturation, intra-luminal detection of atherosclerosis, and simply the identification of different tissue types during procedures. Optical spectroscopic measurements can also be employed in the management of disease treatment. The site-specific pharmacokinetics of chemotherapy and photodynamic therapy agents can be used to customize dosage to the patient, and diagnostic spectroscopy can be used to monitor response to treatment. In recent years clinical studies have provided indications of potential efficacy, and some of these modalities are now entering a translational research stage, with an eye to approval and commercialization. A benefit of these methods is their inherent low cost and ease of implementation, generally mediated with small portable instruments, not requiring any specialized facilities, and eventually not requiring expert interpretation. This paper reviews briefly the most common methods of diagnostic optical spectroscopy, and reviews in greater depth recent clinical translational research invoking scattering spectroscopy as the enabling technology, which has been the experience of the authors.

Elastic scattering spectroscopy for intraoperative determination of sentinel lymph node status in the breast.

The ability to provide the best treatment for breast cancer depends on establishing whether or not the cancer has spread to the lymph nodes under the arm. Conventional assessment requires tissue removal, preparation, and expert microscopic interpretation. In this study, elastic scattering spectroscopy (ESS) is used to interrogate excised nodes with pulsed broadband illumination and collection of the backscattered light. Multiple spectra are taken from 139 excised nodes (53 containing cancer) in 68 patients, and spectral analysis is performed using a combination of principal component analysis and linear discriminant analysis to correlate the spectra with conventional histology. The data are divided into training and test sets. In test sets containing spectra from only normal nodes and nodes with complete replacement by cancer, ESS detects the spectra from cancerous nodes with 84% sensitivity and 91% specificity (per-spectrum analysis). In test sets that included normal nodes and nodes with partial as well as complete replacement by cancer, ESS detects the nodes with cancer with an average sensitivity of 75% and specificity of 89% (per-node analysis). These results are comparable to those from conventional touch imprint cytology and frozen section histology, but do not require an expert pathologist for interpretation. With automation of the technique, results could be made available almost instantaneously. ESS is a promising technique for the rapid, accurate, and straightforward detection of metastases in excised sentinel lymph nodes.

Photodynamic therapy for pancreatic carcinoma: experimental and clinical studies.

Pancreatic carcinoma is the sixth leading cause of cancer-related mortality in the United Kingdom, with an overall 5-year survival of less than 5%. Attempted curative surgery is possible in less than 20% of cases and is associated with a 5-year survival of just 10-20%. Palliative radio-chemotherapy improves symptoms of pancreatic cancer but rarely extends median survival beyond 12 months. There is a need to develop novel therapies that improve outcome. Photodynamic therapy, which is a way of producing localised non-thermal tissue necrosis with light, is currently under evaluation as a treatment for pancreatic cancer. This review will examine some of the mechanisms underlying photodynamic therapy, and the preclinical work, which has led to this treatment being piloted in human studies.

CT contrast predicts pancreatic cancer treatment response to verteporfin-based photodynamic therapy.

The goal of this study was to determine dominant factors affecting treatment response in pancreatic cancer photodynamic therapy (PDT), based on clinically available information in the VERTPAC-01 trial. This trial investigated the safety and efficacy of verteporfin PDT in 15 patients with locally advanced pancreatic adenocarcinoma. CT scans before and after contrast enhancement from the 15 patients in the VERTPAC-01 trial were used to determine venous-phase blood contrast enhancement and this was correlated with necrotic volume determined from post-treatment CT scans, along with estimation of optical absorption in the pancreas for use in light modeling of the PDT treatment. Energy threshold contours yielded estimates for necrotic volume based on this light modeling. Both contrast-derived venous blood content and necrotic volume from light modeling yielded strong correlations with observed necrotic volume (R² = 0.85 and 0.91, respectively). These correlations were much stronger than those obtained by correlating energy delivered versus necrotic volume in the VERTPAC-01 study and in retrospective analysis from a prior clinical study. This demonstrates that contrast CT can provide key surrogate dosimetry information to assess treatment response. It also implies that light attenuation is likely the dominant factor in the VERTPAC treatment response, as opposed to other factors such as drug distribution. This study is the first to show that contrast CT provides needed surrogate dosimetry information to predict treatment response in a manner which uses standard-of-care clinical images, rather than invasive dosimetry methods.

Photodynamic therapy for photochemists.

Photodynamic therapy (PDT) is an evolving technique for localized control of diseased tissue with light after prior administration of a photosensitizing agent and in the presence of oxygen. The biological effect is quite different from surgery, radiotherapy and chemotherapy. With no temperature change during treatment, connective tissues like collagen are largely unaffected, so maintaining the mechanical integrity of hollow organs. PDT is of particular value for pre-cancer and early cancers of the skin (not melanomas) and mouth as the cosmetic and functional results are so good. Another key indication is for small areas of cancer that are unsuitable for or have persisted or recurred after conventional management. It can be applied in areas already exposed to the maximum safe dose of radiotherapy. Outside cancer, in ophthalmology, it is established for age-related macular degeneration, and has considerable potential in arterial disease for preventing restenosis after balloon angioplasty and in the treatment of infectious diseases, where the responsible organisms are accessible to both the photosensitizer and light. New developments on the horizon include techniques for increasing the selectivity for cancers, such as coupling photosensitizers to antibodies, and for stimulating immunological responses, but many further pre-clinical and clinical studies are needed to establish PDT's role in routine clinical practice.

Improved in vivo delivery of m-THPC via pegylated liposomes for use in photodynamic therapy.

Pegylated liposomal nanocarriers have been developed with the aim of achieving improved uptake of the clinical PDT photosensitiser, m-THPC, into target tissues through increased circulation time and bioavailability. This study investigates the biodistribution and PDT efficacy of m-THPC in its standard formulation (Foscan®) compared to m-THPC incorporated in liposomes with different degrees of pegylation (FosPEG 2% and FosPEG 8%), following i.v. administration to normal and tumour bearing rats. The plasma pharmacokinetics were described using a three compartmental analysis and gave elimination half lives of 90 h, 99 h and 138 h for Foscan®, FosPEG 2% and 8% respectively. The accumulation of m-THPC in tumour and normal tissues, including skin, showed that maximal tumour to skin ratios were observed at ≤ 24 h with FosPEG 2% and 8%, whilst skin photosensitivity studies showed Foscan® induces more damage compared to the liposomes at drug-light intervals of 96 and 168 h. PDT treatment at 24h post-administration (0.05 mg kg⁻¹) showed higher tumour necrosis using pegylated liposomal formulations in comparison to Foscan®, which is attributed to the higher tumour uptake and blood plasma concentrations. Clinically, this improved selectivity has the potential to reduce not only normal tissue damage, but the drug dose required and cutaneous photosensitivity.

Photochemical internalisation of a macromolecular protein toxin using a cell penetrating peptide-photosensitiser conjugate.

Photochemical internalisation (PCI) is a site-specific technique for improving cellular delivery of macromolecular drugs. In this study, a cell penetrating peptide, containing the core HIV-1 Tat 48-57 sequence, conjugated with a porphyrin photosensitiser has been shown to be effective for PCI. Herein we report an investigation of the photophysical and photobiological properties of a water soluble bioconjugate of the cationic Tat peptide with a hydrophobic tetraphenylporphyrin derivative. The cellular uptake and localisation of the amphiphilic bioconjugate was examined in the HN5 human head and neck squamous cell carcinoma cell line. Efficient cellular uptake and localisation in endo/lysosomal vesicles was found using fluorescence detection, and light-induced, rupture of the vesicles resulting in a more diffuse intracellular fluorescence distribution was observed. Conjugation of the Tat sequence with a hydrophobic porphyrin thus enables cellular delivery of an amphiphilic photosensitiser which can then localise in endo/lysosomal membranes, as required for effective PCI treatment. PCI efficacy was tested in combination with a protein toxin, saporin, and a significant reduction in cell viability was measured versus saporin or photosensitiser treatment alone. This study demonstrates that the cell penetrating peptide-photosensitiser bioconjugation strategy is a promising and versatile approach for enhancing the therapeutic potential of bioactive agents through photochemical internalisation.

Light penetration in the human prostate: a whole prostate clinical study at 763 nm.

Photodynamic therapy (PDT) is being investigated as a treatment for localized prostate cancer. Photodynamic therapy uses a photosensitizing drug which is activated by a specific wavelength of light, in the presence of oxygen. The activated drug reacts with tissue oxygen to produce reactive oxygen species which are responsible for localized tissue necrosis. One of the determinants of the PDT effect is the penetration of light in the prostate. This study assesses the penetration depth of 763 nm light throughout the prostate. Eight men undergoing multiple hollow needle insertion for high dose rate brachytherapy were recruited. 763 nm light, produced by a diode laser, was delivered to the prostate using cylindrically diffusing optical fibers within the plastic needles. Light was detected at different distances from the source, using an isotropic detector within nearby needles. Penetration depth was calculated using the Boltzmann approximation to the diffusion equation. Delivery detector fiber separation was measured on computed tomography. The mean penetration depth was 0.57 cm, but there was within patient variation of a mean factor of 4.3. Further work is ongoing to assess the effect of such variability in light penetration, on the PDT effect.

Intracellular re-localisation by photochemical internalisation enhances the cytotoxic effect of gelonin--quantitative studies in normal rat liver.

Photochemical internalisation (PCI) is a delivery technology that employs a sub-lethal form of photodynamic therapy (PDT) in which a photosensitiser is activated by light to break down intracellular membranes and release macromolecules into the cytosol where they can be biologically active. Although PCI does enhance the PDT killing of transplanted tumours in mice after local injection of the cytotoxic agent, gelonin, the redistribution of gelonin from intracellular organelles into the cytosol has only previously been demonstrated in vitro. This study is designed to understand the factors controlling the efficacy of PCI in vivo and to document the mechanism of action. Using the photosensitiser AlS(2)Pc in studies on normal rat liver, we have demonstrated in vivo that gelonin is initially taken up into lysosomes, but can be released into the cytosol using PCI. Furthermore, PCI enhances the PDT effect after systemic administration of gelonin (volume of necrosis increased x2.5 when gelonin is given one hour before light), with the remarkably low dose of 5 microg/kg (10,000 times lower than the LD50); in the absence of light, there is no effect with 500 microg/kg. These results suggest that PCI may have a useful role to play in the site specific activation of cytotoxic agents like gelonin, given at a dose level that has no effect in the absence of light.