Contrast enhanced MRI and intravital fluorescence microscopy indicate improved tumor microcirculation in highly vascularized melanomas upon short-term anti-VEGFR treatment.

Anti-angiogenic therapy by blocking VEGF signalling combined with standard chemotherapy is a novel strategy for clinical cancer treatment. The mechanisms for enhanced antitumoral effects are still a matter of controversial debate. Tumor vessel "normalization" upon anti-angiogenic therapy leading to improved drug delivery has been proposed as possible mechanism. Therefore, aim of the study was to investigate tumor microvascular function upon anti-VEGFR treatment in highly vascularized melanomas. A detailed intravital-microscopic analysis of tumor microcirculation including the distribution pattern of vessel diameters and blood flow velocities was performed in melanomas grown in dorsal skinfold chambers of hamsters. Animals with highly vascularized established tumors were treated by a VEGFR tyrosin kinase inhibitor (SU5416) on 3 repetitive days. Tumor tissue oxygenation was measured by phosphorescence quenching technique. Overall tumor microcirculation of subcutaneous tumors was investigated by contrast enhanced MRI (CE-MRI). Vessel density was significantly decreased in treated animals. A significant shift in the distribution patterns towards increased vessel diameters and faster red blood cell velocities in remaining tumor vessels was observed upon anti-VEGF treatment, compensating reduced vascular density. Moreover, a trend towards elevated pO(2) levels in treated tumors was observed. Compared to controls, inflow kinetics of tumors quantified by CE-MRI as well as overall uptake of contrast agent in tumor tissue were significantly increased following short-term SU5416 treatment. In conclusion the results confirm temporarily improved tumor microvascular function in highly vascularized melanomas upon short term anti-VEGFR treatment leading to enhanced tumor blood supply and oxygenation potentially improving the efficacy of simultaneous chemo- or radiotherapy.

[Zenker's diverticulum treated by transoral diverticulostomy: technique and results]. / Die Technik der transoralen Divertikulostomie bei Zenker-Divertikeln - Langzeitresultate und Literaturvergleich.

AIM OF THE STUDY: The surgical technique of transoral diverticulostomy by a modified Endo-GIAtrade mark Stapler (Multifire Endo GIA, Tyco Healthcare) is described. Experiences of this procedure in 31 patients are analysed and compared with different endoscopic and conventional surgical therapies of Zenker's diverticula, which are reported in the literature. METHOD: From January 1996 to December 2005, 31 transoral diverticulostomies were performed. All patients were included porspectively into the study. The median follow-up time after diverticulostomy was 54 months. Manometry, pH-study of the esophagus, endoscopy and swallow radiography were performed before and after surgery. All patients completed the Gastrointestinal quality of live index (GQLI) and the Grosshadern dysphagia score (GHDS). RESULTS: Subjective comfort of the patients as measured by the Smiley Index, the GQLI and the GHDS was increased significantly (p < 0.001) after therapy. Manometry showed that the upper esophageal sphincter functioned normally before and after intervention. A gastrografin swallow excluded leakage at the stapler suture-line in all cases. A conversion to a conventional cricomyotomy with resection of the diverticulum had to be performed once due to a dissection of the esophagus that occurred during insertion of the spreader. In one patient a bleeding out of the suture line was successfully treated with a metal clip. A prothesis broke due to the insertion of the spreader. Two patients developed relapses during the follow-up period of 54 months. CONCLUSION: Compared to standard procedure the endoscopic minimal-invasive therapy proved to be safer. The operation time and the postoperative stay are shorter.

Cationic lipid complexed camptothecin (EndoTAG-2) improves antitumoral efficacy by tumor vascular targeting.

Neo-vascular targeting by cationic colloidal carriers enables to realize an innovative approach for tumor therapy. EndoTag-2 is a novel vascular targeting agent, comprising the mammalian topoisomerase I inhibitor camptothecin in its carboxylate form complexed to cationic lipid (cationic lipid complexed camptothecin). Here we studied tumor vascular targeting properties, antitumoral effects and mode of action of EndoTag-2. Tumor vascular targeting properties of fluorescently labelled EndoTag-2 were investigated by in vivo microscopy using A-MEL-3 tumors grown in the dorsal skinfold chamber preparation and by fluorescence histology of s.c. LLC-1 carcinomas. Therapeutic effects have been investigated in the s.c. LLC-1 carcinoma model and the L3.6pl human pancreatic cancer model implanted orthotopically in athymic nude mice. Antivascular effects have been studied by histological investigation of tumor microvessel density and non invasive investigation of tumor blood flow by dynamic contrast enhanced MRI imaging (DCE-MRI). EndoTag-2 selectively targeted tumor microvessels as confirmed by quantitative fluorescence microscopy. Compared to controls EndoTag-2 revealed remarkable antitumoral efficiency in s.c. LLC-1 carcinomas implanted in C57/Bl6 mice. Growth and metastasis of orthotopic L3.6pl human pancreatic tumors was significantly inhibited by EndoTag-2 treatment. Quantitative analysis of tumor microvessel density revealed significant reduction of microvessel density in lewis lung carcinomas up to 50%. DCE-MRI confirmed significant reduction of intratumoral vascular volume as well as tumor perfusion upon EndoTag-2 treatment. In conclusion this study shows that cationic lipid complexed camptothecin (EndoTag-2) is a markedly active antitumor agent based on an innovative vascular targeting approach.

[Squamous cell carcinoma of the nasal vestibule]. / Fallbericht: Therapieresistenter "Furunkel". Die Diagnose kam zu spät- Nase adé.

The case of a 31-year-old man (smoker) with a progressively enlarging swelling and reddening in the region of the nasal vestibule is discussed. Before he was seen by us, he had been repeatedly treated with antibiotics, but to no avail. A deep excisional biopsy was obtained, and the histopathological work-up revealed a squamous cell carcinoma. Thereupon, complete surgical ablation in healthy tissue was carried out. This case shows that in chronic inflammation of the nasal vestibule that fails to respond to antibiotic treatment differential diagnostic consideration must be given to the possibility of a malignancy.

Paclitaxel encapsulated in cationic lipid complexes (MBT-0206) impairs functional tumor vascular properties as detected by dynamic contrast enhanced magnetic resonance imaging.

Cationic lipid complexes have been shown to be bound and internalized selectively by angiogenic tumor endothelial cells after intravenous injection. Based on this phenomenon, the chemotherapeutic agent paclitaxel was encapsulated into these lipid complexes providing a vascular targeting agent (MBT-0206). As noninvasive imaging techniques are of critical importance for optimizing antivascular cancer treatment in the clinic, we have evaluated the antivascular effects of MBT-0206 in the A-MEL-3 solid tumor model using dynamic contrast enhanced magnetic resonance imaging (DCE-MRI). Twenty-four hours after three intravenous applications of MBT-0206, tumors of treated animals demonstrated a significant decrease of intratumoral blood volume and an increase of vascular permeability in comparison to size-matched control tumors. In contrast, animals treated with conventional paclitaxel given as Taxol at equal drug dose did not show any significant differences in vascular parameters acquired by DCE-MRI in comparison to controls. Immunohistological analysis confirmed a significant reduction of microvessel density in MBT-0206 treated tumors. Moreover, a significant increase of intratumoral microvascular occlusion following MBT-0206 treatment was observed compared to controls and paclitaxel treated animals. In conclusion, antivascular tumor therapy with MBT-0206 significantly impairs functional tumor microcirculation. DCE-MRI is a promising tool to quantify the antivascular effects of MBT-0206 during treatment.

Protamine enhances uptake of cationic liposomes in angiogenic microvessels.

INTRODUCTION: Cationic liposomes have been shown to target angiogenic endothelial cells of solid tumours. Supposing a charge-related mechanism might be responsible for liposome-endothelial interaction, we investigated the effect of intravenous pre-injection of the charged molecules protamine, a polycationic protein, and fucoidan, a polyanionic polysaccharide on the accumulation of cationic liposomes within the blood vessels of a solid tumour. MATERIALS AND METHODS: Experiments were performed using the amelanotic hamster melanoma A-Mel-3 growing in a dorsal skinfold chamber of hamsters. Accumulation of fluorescently-labelled cationic liposomes was quantified by intravital macroscopy and digital image analysis of tumour (t) and surrounding normal host tissue (n) over an observation period of 6 h. All animals received an i.v. injection of cationic liposomes. Animals of the control group were pre-treated with an i.v. injection of 0.9% saline, while animals of group 2 received positively charged protamine and animals of group 3 negatively charged fucoidan prior to liposome injection. RESULTS: In control animals i.v. injection of cationic liposomes revealed a preferential targeting of the tumour vessels, indicated by a maximal t/n ratio of 2.2 +/- 0.24 and a maximal fluorescence intensity (fmax) corresponding to the tumour of 66 +/- 12 [% standard]. While there were no significant differences of liposome accumulation within normal host tissue, accumulation of cationic liposomes within the tumour was significantly enhanced after the pre-administration of protamine (fmax: 117 +/- 12 [% standard]). The t/n ratio was significantly increased in protamine pre-treated animals (5.3 +/- 1.7) in comparison to control and fucoidan treated animals. In contrast, pre-injection of fucoidan resulted in reduced maximal fluorescence intensities in tumour (47 +/- 8 [% standard]) and normal surrounding host tissue. CONCLUSION: Pre-administration of protamine increases the accumulation of cationic liposomes in a solid tumour animal model causing an increased selectivity of cationic liposomes in targeting angiogenic microvessels.

Soluble complement receptor 1 preserves endothelial barrier function and microcirculation in postischemic pancreatitis in the rat.

Components of the activated complement cascade are considered to play a pivotal role in ischemia-reperfusion-induced organ injury. With the use of intravital epifluorescence microscopy, we investigated the effect of complement inhibition by the recombinant soluble complement receptor 1 (sCR1; TP10) on the effect of macromolecular microvascular permeability, functional capillary perfusion, and leukocyte endothelium interaction in postischemic pancreatitis. Anaesthetized Sprague-Dawley rats were subjected to 60 min of normothermic pancreatic ischemia induced by microclipping of the blood-supplying arteries of the organ. Rats who received sCR1 (15 mg/kg body wt iv; n = 7) during reperfusion showed a significant reduction of permeability (1.77 +/- 1.34 x 10(-8) cm/s; n = 7) of tetramethylrhodamine isothiocyanate-labeled albumin injected 90 min after the onset of reperfusion compared with vehicle-treated animals (6.95 +/- 1.56 x 10(-8) cm/s; n = 7). At 120 min after the onset of reperfusion, the length of red blood cell-perfused capillaries (functional capillary density) was significantly improved (from 279 +/- 15.7 to 330 +/- 3.7 cm(-1); n = 7) and the number of leukocytes adherent to postcapillary venules was significantly reduced (from 314 +/- 87 to 163 +/- 71 mm(-2); n = 7) by sCR1 compared with vehicle treatment. Complement inhibition by sCR1 effectively ameliorates pancreatic ischemia-reperfusion-induced microcirculatory disturbances and might be considered for treatment of postischemic pancreatitis.

[Clinical and epidemiological data of patients with malignomas of the head and neck]. / Klinische und epidemiologische Daten zu Malignomen des Kopf-Hals-Bereichs.

BACKGROUND: The present study describes clinical and epidemiological data of patients with malignomas of the head and neck documented in the Munich Cancer Register. PATIENTS AND METHODS: Data of head and neck cancer patients treated at four departments of head and neck surgery and one of oral-maxillo-facial surgery in the area of Munich from 1978 up to now are reported. RESULTS: Incidence and mortality as a function of age, sex, and tumor localization are described in comparison to clinical and epidemiological data as specified in tumor registers of the Saarland and the USA. Moreover, TNM stages, survival, recurrence, and metastasis rates are presented. CONCLUSION: Based on the documentation of the Munich Cancer Register our study is the first to present a detailed description of clinical and epidemiological data of patients suffering from head and neck malignomas.

[Antivascular strategies--a new concept for diagnosis and therapy of head and neck cancer. A review]. / Antivaskuläre Strategien--ein neues Konzept zur Diagnose und Therapie von Kopf-Hals-Tumoren--Eine Ubersicht.

BACKGROUND: The development of a blood supply is a crucial step in the progression and metastasis of head and neck cancer. While conventional therapeutic approaches, e. g. chemotherapy and radiation, are focusing on tumor cells, antivascular therapy is directed against the tumor supplying blood vessels. Antivascular treatment can be divided in anti-angiogenesis and vascular targeting. While antiangiogenic therapy prevents neovascularisation by inhibiting new blood vessel growth, the aim of vascular targeting is the destruction of already existing tumor vasculature. METHODS: This review will summarise current pathophysiological mechanisms underlying tumor angiogenesis and the concepts of antivascular therapy with respect to possible applications in head and neck cancer. RESULTS: With experimental antivascular strategies a retardation of tumor growth or tumor remission may be achieved, demonstrating proof of principle. At present, numerous drugs are preclinically and clinically evaluated. CONCLUSIONS: Antivascular strategies are a promising concept for treatment of patients with head and neck carcinomas. They will most likely exhibit their potential in combination with standard tumor therapies in the future.

Orthogonal polarisation spectral imaging as a new tool for the assessment of antivascular tumour treatment in vivo: a validation study.

Tumour angiogenesis plays a key role in tumour growth, formation of metastasis, detection and treatment of malignant tumours. Recent investigations provided increasing evidence that quantitative analysis of tumour angiogenesis is an indispensable prerequisite for developing novel treatment strategies such as anti-angiogenic and antivascular treatment options. Therefore, it was our aim to establish and validate a new and versatile imaging technique, that is orthogonal polarisation spectral imaging, allowing for non-invasive quantitative imaging of tumour angiogenesis in vivo. Experiments were performed in amelanotic melanoma A-MEL 3 implanted in a transparent dorsal skinfold chamber of the hamster. Starting at day 0 after tumour cell implantation, animals were treated daily with the anti-angiogenic compound SU5416 (25 mg kg x bw(-1)) or vehicle (control) only. Functional vessel density, diameter of microvessels and red blood cell velocity were visualised by both orthogonal polarisation spectral imaging and fluorescence microscopy and analysed using a digital image system. The morphological and functional properties of the tumour microvasculature could be clearly identified by orthogonal polarisation spectral imaging. Data for functional vessel density correlated excellently with data obtained by fluorescence microscopy (y=0.99x+0.48, r2=0.97, R(S)=0.98, precision: 8.22 cm(-1) and bias: -0.32 cm(-1)). Correlation parameters for diameter of microvessels and red blood cell velocity were similar (r2=0.97, R(S)=0.99 and r2=0.93, R(S)=0.94 for diameter of microvessels and red blood cell velocity, respectively). Treatment with SU5416 reduced tumour angiogenesis. At day 3 and 6 after tumour cell implantation, respectively, functional vessel density was 4.8+/-2.1 and 87.2+/-10.2 cm(-1) compared to values of control animals of 66.6+/-10.1 and 147.4+/-13.2 cm(-1), respectively. In addition to the inhibition of tumour angiogenesis, tumour growth and the development of metastasis was strongly reduced in SU5416 treated animals. This new approach enables non-invasive, repeated and quantitative assessment of tumour vascular network and the effects of antiangiogenic treatment on tumour vasculature in vivo. Thus, quantification of tumour angiogenesis can be used to more accurately classify and monitor tumour biologic characteristics, and to explore aggressiveness of tumours.

Inhibition of the alpha-nu integrins with a cyclic RGD peptide impairs angiogenesis, growth and metastasis of solid tumours in vivo.

Anti-angiogenetic cancer therapy is a potential new form for treatment of solid tumours. The alpha(v)-integrins (alpha(v)beta3, alpha(v)beta5) mediate the contact of activated endothelial cells to proteins of the extracellular matrix during tumour angiogenesis as a prerequisite for survival of endothelial cells. The aim of this study was to investigate the effects of application of a methylated cyclic RGD-peptide as an alpha(v)-integrin antagonist on angiogenesis, microcirculation, growth and metastasis formation of a solid tumour in vivo. Experiments were performed in the dorsal skinfold preparation of Syrian Golden hamsters bearing the amelanotic hamster melanoma A-Mel-3. Animals were injected intraperitoneally with a methylated cyclic RGD-peptide every 12 h, the control group received an inactive peptide. Microcirculatory parameters of tumour angiogenesis including functional vessel density, red blood cell velocity, vessel diameter and leucocyte-endothelium interaction were analysed using intravital microscopy. In an additional study the effects on growth and metastasis of subcutaneous A-Mel-3 were quantified. Functional vessel density was markedly reduced on day 3 in treated animals compared to controls (37.2 +/- 12.1 vs 105.2 +/- 11.2 cm(-1); mean +/- s.e.m.; P<0.05) and increased subsequently in both groups. Red blood cell velocity at day 3 was below values of controls (0.026 +/- 0.01 vs 0.12 +/- 0.03 mm x s(-1); P<0.05). No differences were observed in vessel diameters and leucocyte-endothelium interaction was almost absent in both groups. Furthermore, growth and metastasis of subcutaneous tumours after administration of the cyclic RGD-peptide was significantly delayed in comparison to controls (P<0.05). Inhibition of alpha(v)-integrins by a cyclic RGD-peptide resulted in significant reduction of functional vessel density, retardation of tumour growth and metastasis in vivo. Taken together, these results implicate RGD-peptides as agents which have anti-tumour and anti-metastatic activity in vivo.

Quantitative imaging of tumour blood flow by contrast-enhanced magnetic resonance imaging.

Tumour blood flow plays a key role in tumour growth, formation of metastasis, and detection and treatment of malignant tumours. Recent investigations provided increasing evidence that quantitative analysis of tumour blood flow is an indispensable prerequisite for developing novel treatment strategies and individualizing cancer therapy. Currently, however, methods for noninvasive, quantitative and high spatial resolution imaging of tumour blood flow are rare. We apply here a novel approach combining a recently established ultrafast MRI technique, that is T(1)-relaxation time mapping, with a tracer kinetic model. For validation of this approach, we compared the results obtained in vivo with data provided by iodoantipyrine autoradiography as a reference technique for the measurement of tumour blood flow at a high resolution in an experimental tumour model. The MRI protocol allowed quantitative mapping of tumour blood flow at spatial resolution of 250 x 250 microm(2). Correlation of data from the MRI method with the iodantipyrine autoradiography revealed Spearman's correlation coefficients of Rs = 0.851 (r = 0.775, P < 0.0001) and Rs = 0.821 (r = 0.72, P = 0.014) for local and global tumour blood flow, respectively. The presented approach enables noninvasive, repeated and quantitative assessment of microvascular perfusion at high spatial resolution encompassing the entire tumour. Knowledge about the specific vascular microenvironment of tumours will form the basis for selective antivascular cancer treatment in the future.

Tissue distribution and penetration of 5-ALA induced fluorescence in an amelanotic melanoma after topical application.

BACKGROUND: Photodynamic therapy (PDT) following topical application of 5-aminolevulinic acid (ALA) is increasingly employed for several types of malignancies. However, data with respect to tissue penetration and distribution of ALA-induced porphyrins after topical application are scarce. Therefore, it was our aim to study tissue distribution and the penetration potency of topically applied ALA. MATERIAL AND METHODS: We used Syrian golden hamsters implanted with the amelanotic melanoma A-Mel-3 growing in a transparent dorsal skinfold chamber. ALA was topically applied in aqueous solution at a concentration of 3% for 4 hours. The fluorescence pattern was quantified by fluorescence microscopy and digital image analysis from cryosections and given as percentage of a reference standard in medians (25%, 75% quartiles). RESULTS: Fluorescence intensities in tumors were 90.8% (56.2%, 115.2% of a reference standard, p < 0.01 vs. normal tissue) significantly exceeding normal surrounding host tissue yielding fluorescence intensities of 12.1% (9.1%, 16.1%). The tumor selectivity, that is the ratio of fluorescence intensities between tumor and normal tissue, was 7.3 (6.1, 9.1). For superficial tumors with a thickness of approximately 1 mm no fluorescence gradients after topical application of ALA could be observed. CONCLUSION: In superficial cancerous lesions the fluorescence distribution of ALA induced porphyrins is tumor selective without significant fluorescence gradients throughout the tumor. Thus, by optimising the treatment modalities for topical ALA-PDT an enhanced efficacy and selectivity will be reached.

Vascular permeability in a human tumour xenograft: molecular charge dependence.

Molecular charge is one of the main determinants of transvascular transport. There are, however, no data available on the effect of molecular charge on microvascular permeability of macromolecules in solid tumours. To this end, we measured tumour microvascular permeability to different proteins having similar size but different charge. Measurements were performed in the human colon adenocarcinoma LS174T transplanted in transparent dorsal skinfold chambers in severe combined immunodeficient (SCID) mice. Bovine serum albumin (BSA) and IgG were fluorescently labelled and were either cationized by conjugation with hexamethylenediamine or anionized by succinylation. The molecules were injected i.v. and the fluorescence in tumour tissue was quantified by intravital fluorescence microscopy. The fluorescence intensity and pharmacokinetic data were used to calculate the microvascular permeability. We found that tumour vascular permeability of cationized BSA (pI-range: 8.6-9.1) and IgG (pI: 8.6-9.3) was more than two-fold higher (4.25 and 4.65x10(-7) cm s(-1)) than that of the anionized BSA (pI approximately 2.0) and IgG (pI: 3.0-3.9; 1.11 and 1.93x10(-7) cm s(-1), respectively). Our results indicate that positively charged molecules extravasate faster in solid tumours compared to the similar-sized compounds with neutral or negative charges. However, the plasma clearance of cationic molecules was approximately 2x faster than that of anionic ones, indicating that the modification of proteins enhances drug delivery to normal organs as well. Therefore, caution should be exercised when such a strategy is used to improve drug and gene delivery to solid tumours.

Validation of MR thermometry technology: a small animal model for hyperthermic treatment of tumours.

BACKGROUND: Local hyperthermia has been shown to be an effective adjuvant therapy for cancer. However, progress in this treatment modality requires the non-invasive assessment of temperature distribution in the entire tumour to enable administration of an efficient thermal dose to all tumour areas. Magnetic resonance (MR) imaging offers a promising tool to quantify, non-invasively and three-dimensionally, temperature distribution within tumours. An animal model taking into account the complex interrelationship between pathophysiological changes within a tumour during hyperthermia and temperature-sensitive MR parameters is warranted for the development and validation of new MR thermometry technology. METHODS: An experimental set-up was implemented to allow simultaneous measurements of temperature, tumour blood flow and temperature-sensitive MR parameters under standardised conditions in vivo. Local hyperthermia was induced at 44 degrees C for 20 min under inhalation anaesthesia on seven Syrian Golden hamsters bearing an amelanotic melanoma. Fibreoptic probes were used for reference temperature measurements. Laser Doppler flowmetry served for on-line tumour blood flow determination, and MR thermometry was performed using longitudinal T1 relaxation time measurements. RESULTS: The experimental design enables multifunctional MR thermometry. T1 relaxation times of tumours were 1.44 s (1.36, 1.46) and 1.53 s (1. 48, 1.75) at 37 degrees C and during hyperthermia at 44 degrees C, respectively (median, 25% and 75% quartiles, respectively; P<0.05). At the end of 20 min of hyperthermic treatment at 44 degrees C, relative tumour blood flow was reduced to 40.5% (20.7, 43.3) compared to values before treatment (median, 25% and 75% quartiles, respectively; P<0.05). Imaging of T1 relaxation times revealed a heterogeneous distribution in temperature during hyperthermic treatment. CONCLUSION: This novel in vivo model allows standardised investigations for the development and validation of MR thermography methods.

Primary tumor size-dependent inhibition of angiogenesis at a secondary site: an intravital microscopic study in mice.

Some primary tumors are capable of suppressing the growth of their metastases by presumably generating antiangiogenic factors such as angiostatin. We hypothesized that the amount of inhibitor(s) released by a tumor increases with tumor growth. We tested this hypothesis by evaluating the relationship between the size of a primary tumor and its ability to inhibit angiogenesis at a secondary site. Furthermore, we characterized the effects of the primary tumor on physiological properties of newly formed vessels at the secondary site. Angiogenesis and physiological properties were measured using intravital microscopy of angiogenic vessels in the gels containing basic fibroblast growth factor placed into cranial windows of immunodeficient mice bearing human prostatic carcinoma (PC-3) in their flank. The PC-3 tumor inhibited angiogenesis in the gels, and surgical resection of tumor reversed this inhibition. The inhibition of angiogenesis 20 days after gel implantation (range, 0-83%) correlated positively (r = 0.625; P < 0.008) with the tumor size on the day of gel implantation (range, 19-980 mm3). The primary tumor also suppressed leukocyte-adhesion in angiogenic vessels, thus helping them evade the immune recognition. These results provide an additional rationale for combining antiangiogenic treatment with local therapies.

Pharmacokinetics and selectivity of aminolevulinic acid-induced porphyrin synthesis in patients with cervical intra-epithelial neoplasia.

Photodynamic therapy (PDT), due to its tumor selectivity, represents an alternative approach to diagnose and treat cervical intra-epithelial neoplasia (CIN) without altering normal surrounding tissue. Our aim was to investigate the pharmacokinetics and the selectivity of 5-aminolevulinic acid (5-ALA)-induced porphyrin fluorescence after topical administration, to obtain basic clinical data for future diagnostic fluorescence imaging and PDT protocols for CIN. Twenty-eight non-pregnant women with a cytological diagnosis of low-grade or high-grade squamous intra-epithelial lesions were included. An aqueous solution containing 3% 5-ALA was topically applied 1 to 6 hrs prior to conization using a cervical cap. After excision, porphyrin-induced fluorescence was quantified in dysplastic (n = 14) and normal epithelium (n = 28) by means of quantitative fluorescence microscopy. High values of porphyrin fluorescence were found in squamous epithelium between 150 and 450 min, with a maximum at 300 min following administration of 5-ALA. Ratios of porphyrin fluorescence of dysplastic vs. surrounding normal epithelium were 1.3 and 1.21 for CIN 1 (n = 3) and CIN 2 (n = 3), respectively. In CIN 3 patients (n = 8), this ratio was 2.35; the best selectivity of 5-ALA-induced porphyrin fluorescence in CIN 3 lesions (ratio 3) was observed with a topical administration time of between 150 and 250 min. Our results demonstrate that patients with CIN 3 show higher 5-ALA-induced fluorescence compared with normal epithelium. The optimal administration time of topically applied 5-ALA was between 3 and 4 hr. Our data suggest that topical ALA-PDT and photodynamic diagnosis might be suitable for detecting CIN.

Distribution and pharmacokinetics of Photofrin in human bile duct cancer.

Prognosis of patients with bile duct tumors is mostly poor due to late diagnosis and a lack of adequate curative and palliative treatment modalities. To evaluate the potential of photodynamic therapy (PDT) as a novel and alternative treatment approach, we have investigated the uptake and tumor-specific localization of the photosensitizer Photofrin in human biliary tract neoplasms. We have quantified the distribution and the pharmacokinetics of Photofrin in normal and tumor tissue biopsies of the human bile duct by quantitative fluorescence microscopy and digital image analysis of cryosections. Fluorescence intensities (expressed as a percentage of a standard) are 19.0 +/- 11.4% and 25.2 +/- 12.7% for tumors and 10.9 +/- 2.9% and 13.2 +/- 9.1% (mean +/- SD) for normal bile duct tissue at 24 h (n = 5) and 48 h (n = 8) after Photofrin administration (2 mg kg-1 i.v.), respectively, and decrease afterwards in normal bile duct tissue over the period of investigation (4-35 days). The ratios of fluorescence in tumor versus normal tissue are found to be 1.7 +/- 0.7 and 2.3 +/- 1.2 (mean +/- SD) at days one and two after Photofrin administration, respectively. Thus, Photofrin preferentially accumulates in bile duct neoplasms, reaching peak values during the first two days. These data suggest that laser irradiation should be performed within this period after Photofrin injection to achieve tumor selectivity of PDT for effective treatment of bile duct carcinoma.

Changes in subcutaneous interstitial fluid pressure, tissue oxygenation, and skin red cell flux during venous congestion plethysmography in men.

OBJECTIVE: Venous congestion plethysmography enables noninvasive assessment of microvascular filtration capacity (Kf) in limbs. However, increases in fluid filtration might alter the balance of Starling forces: for example, progressive increases in interstitial fluid pressure (Pi) would reduce net fluid flux, thus underestimating Kf. Furthermore, elevation of cuff pressure to values close to diastolic blood pressure, as used in the protocol, may be itself impair tissue perfusion with unknown effects on the microvascular parameters investigated. METHODS: Pi was measured in healthy volunteers (n = 14) with a modified "Wick in needle" technique during small (8 mm Hg) cumulative increases in venous pressure (0-95 mm Hg). Changes in the hemoglobin (Hb) concentration, oxygenated hemoglobin (HbO2) concentration and oxidized cytochrome aa3 concentration were assessed in the calf using noninvasive near-infrared spectroscopy. Skin red blood cell flux close to the strain gauge was evaluated by laser Doppler fluxmetry. RESULTS: Pi at control was -0.89 +/- 0.8 mm Hg and during elevation of venous pressure remained constant until a cuff pressure of 30 mm Hg was reached. It rose thereafter to 1.57 +/- 1.3 mm Hg (mean +/- SD). Skin red cell flux was significantly reduced when cuff pressure exceeded 30 mm Hg and following cuff deflation, evidence of reactive hyperemia was obtained. Hb concentration increased significantly as a result of venous pressure elevation. No change in either HbO2 or cytochrome aa3 concentration was observed as long as cuff pressure remained under diastolic blood pressure. CONCLUSIONS: The small increase in Pi together with an absence of impaired tissue oxygenation during the venous congestion plethysmography protocol described by Gamble et al. supports the contention that this protocol enables accurate assessment of filtration capacity.