5-aminolevulinic acid photodynamic therapy for the treatment of high-grade gliomas.

INTRODUCTION: Photodynamic therapy (PDT) is a two-step treatment involving the administration of a photosensitive agent followed by its activation at a specific light wavelength for targeting of tumor cells. MATERIALS/METHODS: A comprehensive review of the literature was performed to analyze the indications for PDT, mechanisms of action, use of different photosensitizers, the immunomodulatory effects of PDT, and both preclinical and clinical studies for use in high-grade gliomas (HGGs). RESULTS: PDT has been approved by the United States Food and Drug Administration (FDA) for the treatment of premalignant and malignant diseases, such as actinic keratoses, Barrett's esophagus, esophageal cancers, and endobronchial non-small cell lung cancers, as well as for the treatment of choroidal neovascularization. In neuro-oncology, clinical trials are currently underway to demonstrate PDT efficacy against a number of malignancies that include HGGs and other brain tumors. Both photosensitizers and photosensitizing precursors have been used for PDT. 5-aminolevulinic acid (5-ALA), an intermediate in the heme synthesis pathway, is a photosensitizing precursor with FDA approval for PDT of actinic keratosis and as an intraoperative imaging agent for fluorescence-guided visualization of malignant tissue during glioma surgery. New trials are underway to utilize 5-ALA as a therapeutic agent for PDT of the intraoperative resection cavity and interstitial PDT for inoperable HGGs. CONCLUSION: PDT remains a promising therapeutic approach that requires further study in HGGs. Use of 5-ALA PDT permits selective tumor targeting due to the intracellular metabolism of 5-ALA. The immunomodulatory effects of PDT further strengthen its use for treatment of HGGs and requires a better understanding. The combination of PDT with adjuvant therapies for HGGs will need to be studied in randomized, controlled studies.

[Optical diagnostic methods for early tumour diagnosis in the upper aerodigestive tract: Quo vadis?]. / Optische Diagnoseverfahren zur Tumorfrühdiagnostik im oberen Luft-Speise-Weg: Quo vadis?

BACKGROUND: Optical diagnostic methods may simplify and improve the early diagnosis of tumours of the upper aerodigestive tract; however, these have not yet found their way into clinical routine. OBJECTIVE: This article aims to define the problems that have prevented routine use of optical diagnostic methods so far, as well as listing and also explaining potential trendsetting approaches to overcome these difficulties. MATERIALS AND METHODS: The study is based on a combined analysis of publically accessible databases (PubMed MEDLINE, Thompson Reuters Web of Science, SPIE. Digital Library; full time period available; search strings: "oral cavity", "pharynx", "larnyx", "optical diagnosis", "optical biopsy", "optical coherence tomography", "confocal endomicroscopy", "fluorescence endoscopy", "narrow band imaging", "non-linear imaging", "fluorescence lifetime imaging"), as well as personal experiences. RESULTS: Both conceptual and methodical problems were determined, and possible solutions based on current developments are discussed. CONCLUSION: Optical diagnostic methods have the potential to revolutionise early diagnosis of upper aerodigestive tract malignancies, providing the different hurdles listed in this review can be overcome.

Diagnostic efficacy of backscattering intensity measurements in optical coherence tomography of cervical intraepithelial dysplasia.

BACKGROUND AND OBJECTIVES: The aim of this study was to determine the diagnostic efficacy of backscattering intensity measurements in optical coherence tomography in identifying different grades of cervical intraepithelial dysplasia. STUDY DESIGN/MATERIALS AND METHODS: OCT images were taken from 153 unsuspicious and suspicious areas of 30 fresh conisation and hysterectomy specimens, evaluated by two blinded investigators using a six-grade classification (normal, inflammation, CIN1, CIN2, CIN3, squamous carcinoma) and later compared to the corresponding histology. Differences between judgments based on either the histology or the OCT images were investigated employing Correspondence Analysis (CA). Further, we explored the extent as to which backscattering intensity profiles of OCT images contained the essential information required for a reliable and valid diagnosis, using Linear Discriminant Analysis (LDA). RESULTS: The CA of histology- and OCT-based judgments suggests that the diagnostic process may be characterized in terms of two stochastically independent underlying ("latent") variables, the first of them reflecting the definiteness with which CIN classes are identified, the second reflecting a bias towards diagnosing inflammation on the side of the OCT-based judgments. This finding is supported by the results of LDAs, where histology and OCT categorizations differ in particular with respect to the positions of inflammation and CIN1. Possibly, a second canonical variable has to be assumed accounting for the evaluation of carcinoma. CONCLUSIONS: The systematic differences between histology-based and OCT-based diagnoses suggest that the use of available information is influenced by perceptual and/or cognitive biases. Apart from this it seems that the profiles appear to provide a remarkably large amount of information determining the main course of the diagnostic process.

3D optical coherence tomography of cervical intraepithelial neoplasia--early experience and some pitfalls.

OBJECTIVES: To compare two different systems for optical coherence tomography for the diagnosis of cervical dysplasia and to assess potential benefits of three-dimensional imaging. MATERIALS AND METHODS: OCT images were taken from unsuspicious and suspicious areas of fresh conisation specimens using two different imaging systems, one with the capability to produce three-dimensional images. All OCT images were separately evaluated by two blinded investigators based on a 6-grade classification (normal, inflammation, CIN 1, CIN 2, CIN 3, squamous carcinoma) and later compared to the corresponding histology. Sensitivity and specificity of OCT in detecting cervical dysplasia were determined. RESULTS: OCT images using both OCT systems were taken from 46 sites in ten conisation specimens and later compared to the corresponding histology. CIN lesions were diagnosed correctly by the two-dimensional OCT system with a sensitivity and specificity of 91% and 78% accordingly. Using the three-dimensional system sensitivity and specificity were 82% and 86% accordingly. CONCLUSIONS: Both OCT systems used were highly sensitive in identifying cervical intraepithelial neoplasia. Despite technical problems experienced in the present series, we believe that three-dimensional imaging has the potential to further improve the accuracy of optical coherence tomography.

Optical coherence tomography for bladder cancer -- ready as a surrogate for optical biopsy? Results of a prospective mono-centre study.

INTRODUCTION: New modalities like Optical Coherence Tomography (OCT) allow non-invasive examination of the internal structure of biological tissue in vivo. The potential benefits and limitations of this new technology for the detection and evaluation of bladder cancer were examined in this study. MATERIALS AND METHODS: Between January 2007 and January 2008, 52 patients who underwent transurethral bladder biopsy or TUR-BT for surveillance or due to initial suspicion of urothelial carcinoma of the bladder were enrolled in this study. In total, 166 lesions were suspicious for malignancy according to standard white light cystoscopy. All suspicious lesions were scanned and interpreted during perioperative cystoscopy using OCT. Cold cup biopsies and/or TUR-B was performed for all these lesions. For this study we used an OCT-device (Niris, Imalux, Cleveland, US), that utilizes near-infrared light guided through a flexible fibre-based applicator, which is placed into the bladder via the working channel of the cystoscope. The technology provides high spatial resolution on the order of about 10-20 microm, and a visualization of tissue to a depth of about 2 mm across a lateral span of about 2 mm in width. The device used received market clearance from the FDA and CE approval in Germany. The diagnostic and surgical procedure was videotaped and analyzed afterwards for definitive matching of scanned and biopsied lesion. The primary aim of this study was to determine the level of correlation between OCT interpretation and final histological result. RESULTS: Of 166 scanned OCT images, 102 lesions (61.4%) matched to the same site where the biopsy/TUR-BT was taken according to videoanalysis. Only these video-verified lesions were used for further analysis. Of all analyzed lesions 88 were benign (inflammation, edema, hyperplasia etc.) and 14 were malignant (CIS, Ta, T1, T2) as shown by final histo?pathology. - All 14 malignant lesions were detected correctly by OCT. Furthermore all invasive tumors were staged correctly by OCT regarding tumor growth beyond the lamina propria. There were no false negative lesions detected by OCT. Sensitivity of OCT for detecting the presence of a malignant lesion was 100% and sensitivity for detection of tumor growth beyond the lamina propria was 100% as well. Specificity of OCT for presence of malignancy was 65%, due to the fact that a number of lesions were interpreted as false positive by OCT. CONCLUSION: As a minimally invasive technique, OCT proved to have extremely high sensitivity for detection of malignant lesions as well as estimation of whether a tumor has invaded beyond the lamina propria. However, specificity of OCT within the bladder was impaired (65%), possibly due to a learning curve and/or the relatively low spatial resolution and visualization depth of the OCT technology. Further studies and technical development are needed to establish an adequate surrogate for optical biopsy.

Near-infrared autofluorescence imaging to detect parathyroid glands in thyroid surgery.

Objective To identify and save parathyroid glands during thyroidectomy by displaying their autofluorescence. Methods Autofluorescence imaging was carried out during thyroidectomy with and without central lymph node dissection. After visual recognition by the surgeon, the parathyroid glands and the surrounding tissue were exposed to near-infrared light with a wavelength of 690-770 nm using a modified Karl Storz near infrared/indocyanine green endoscopic system. Parathyroid tissue was expected to show near infrared autofluorescence at 820 nm, captured in the blue channel of the camera. Results We investigated 41 parathyroid glands from 20 patients; 37 glands were identified correctly based on near-infrared autofluorescence. Neither lymph nodes nor thyroid revealed substantial autofluorescence and nor did adipose tissue. Conclusions Parathyroid tissue is characterised by showing autofluorescence in the near-infrared spectrum. This effect can be used to identify and preserve parathyroid glands during thyroidectomy.

[Fluorescence cystoscopy at bladder cancer: present trials]. / Die Fluoreszenzzystoskopie beim Harnblasenkarzinom : Aktueller Stellenwert.

Bladder cancer is a frequent disease and represents the second most common genitourinary neoplasm. Although many aspects of the management of not-muscle-infiltrating bladder cancer are now well established, significant challenges remain, which influence patient outcome. Early detection and treatment of recurrent disease is required to optimize bladder preservation, reduce patient morbidity and increase quality of life and survival. Fluorescence cystoscopy, often referred to as "photodynamic diagnosis" (PDD) with intravesical application of photosensitizing agents has been developed in order to enhance the early detection of bladder cancer. Since March 2005 the hexyl-ALA ester (Hexvix) has been approved for the diagnosis of bladder cancer in 27 EU/EEA countries through the European Mutual Recognition Procedure. There is growing evidence that PDD enhances the detection of bladder cancer, particularly of high-grade flat lesions. Furthermore, transurethral resection of bladder tumor under fluorescence guidance has been shown to reduce the risk of recurrent tumors. Nevertheless, a resulting relatively decreased number of recurrences have still to be verified in prospective randomized trials.

[Fluorescence cytology. Improvement of urinary cytology]. / Die Fluoreszenzzytologie. Eine Weiterentwicklung der Urinzytologie.

Urothelial cancer of the bladder is a frequent disease, and urinary cytology often is used as a routine diagnostic tool. But this technique has an impaired sensitivity in low-grade tumours, and as a subjective method it is highly dependent on the experience of the cytologist. Here we present the technique of fluorescence cytology as an improvement of conventional cytology. This method is potentially able to compensate for the disadvantages of urinary cytology as it is an automated process that uses the principles of 5-Ala-induced photodynamic diagnosis (PDD).

[Fluorescence diagnosis and photodynamic therapy in urology]. / Fluoreszenzdiagnostik und photodynamische therapie in der urologie.

Urology is a preferential domain of endoscopy and as such an important research field for photodynamic procedures. An important milestone in the long-lasting and successful history of "photodynamics" in urology is the European approval of hexaminolevulinate (HAL, Hexvix) for fluorescence cystoscopy. All clinical studies carried out so far have demonstrated a significant increase in sensitivity of fluorescence versus standard cystoscopy for the detection of bladder cancer, especially concerning carcinoma in situ. The majority of the randomised, two-armed studies additionally show significantly reduced rates of residual tumour and recurrences. Tumor-selective fluorescence can also be observed in the kidney and prostate. Intraoperative fluorescence detection might thus simplify the achievement of high rates of R0 resections. Apart from the diagnostic potential of "photodynamics", also some possible therapeutic indications will be mentioned, including photodynamic therapy of bladder cancer and prostate cancer. Whereas initial clinical experience has been obtained for photodynamic therapy of bladder cancer, clinical studies for other indications are currently being designed. By providing an overview over methods and procedures as well as hitherto the available clinical results, we hope to provide reader with a basis for obtaining his/her own judgement.

Localization and staging of cervical intraepithelial neoplasia using double ratio fluorescence imaging.

A phase zero evaluation of a new fluorescence imaging technique for diagnosing cervical intraepithelial neoplasia (CIN) was performed. The fluorescence imaging prototype performed quantitative imaging of Protoporphyrin induced by a topically applied aminolevulinic acid using double ratio (DR) fluorescence imaging technique developed by our group. A total of 38 patients were in the protocol, with 16 colposcopically selected for biopsy. Fluorescence images of these 16 patients were taken, 19 sites were biopsied, and the disease was staged histopathologically. DR fluorescence imaging of the cervix using our general purpose prototype appeared to be cumbersome but feasible. In four cases strongly localized fluorescent hotspots were observed at the location where the disease was colposcopically visible. In the other cases the fluorescence showed a more diffuse multifocal image. The value of the DR determined at the site of biopsy correlated in a statistically significant way with the histopathologically determined stage of the disease [Spearman rank correlation, r=0.881, p<0.001 (confidence interval 0.7044-0.9552)]. This suggests that noninvasive staging of CIN using this technique is feasible. We believe that the results of this study justify the development of a dedicated device that combines regular white light colposcopy with DR fluorescence imaging.

Intraoperative detection of malignant gliomas by 5-aminolevulinic acid-induced porphyrin fluorescence.

OBJECTIVE: Survival after surgery and radiotherapy for the treatment of malignant gliomas is linked to the completeness of tumor removal. Therefore, methods that permit intraoperative identification of residual tumor tissue may be of benefit. In a preliminary investigation, we have studied the value of fluorescent porphyrins that accumulate in malignant tissue after administration of a precursor (5-aminolevulinic acid) for labeling of malignant gliomas in nine patients. METHODS: Three hours before the induction of anesthesia, 10 mg 5-aminolevulinic acid/kg body weight was administered orally. Intraoperatively, red porphyrin fluorescence was observed with a 455-nm long-pass filter after excitation with violet-blue (375-440 nm) xenon light and was verified by analysis of fluorescence spectra. Fluorescing and nonfluorescing samples taken from the tumor perimeters were examined histologically or used to study the photobleaching of porphyrins by excitation light and white light from the operating microscope. Plasma and erythrocyte porphyrin levels were determined by fluorescence photometry. RESULTS: Normal brain tissue revealed no porphyrin fluorescence, whereas tumor tissue was distinguished by bright red fluorescence. For a total of 89 tissue biopsies, sensitivity was 85% and specificity was 100% for the detection of malignant tissue. For seven of nine patients, visible porphyrin fluorescence led to further resection of the tumor. Under operating light conditions, fluorescence decayed to 36% in 25 minutes for violet-blue light and in 87 minutes for white light. Plasma and erythrocyte porphyrin contents increased slightly, without exceeding normal levels. CONCLUSION: Our observations suggest that 5-aminolevulinic acid-induced porphyrin fluorescence may label malignant gliomas safely and accurately enough to enhance the completeness of tumor removal.

Pharmacokinetics of fluorescent polyporphyrin Photofrin II in normal rat tissue and rat bladder tumor.

The transient behavior of the molecular components responsible for fluorescence emission of the photosensitizing polyporphyrin Photofrin II has been studied quantitatively in the liver, small intestines, bladder and muscles of rats. Relative concentrations of the substance were determined fluorometrically in vivo using a Kr(+)-laser (wavelength = 406.7 nm) and a mercury arc lamp (wavelength = 405 or 550 nm) for fluorescence excitation of Photofrin II. Fluorescence was detected at the maxima of the emission bands, at 630 or 690 nm. The results of the experiments show that Photofrin II can be clearly detected by its fluorescence in all the organs investigated from 3 h up to at least 28 days after systemic application of the substance. Within this investigational period the fluorescing components of Photofrin II are released continuously from the organs. In all the tissues examined, an initial decrease with time constants between 2 and 42 h followed by a slow decay with time constants between about 300 and 600 h can be observed. In addition the pharmacokinetics of the fluorescent components of Photofrin II in chemically induced rat bladder tumors with different stages of malignancy were compared to healthy rat bladder tissue. In a time range of 2-10 days after intravenous injection Photofrin II shows a fluorescence 2-5 times brighter in rat bladder tumors than in healthy bladder tissue.

Photodynamic effects induced by aminolevulinic acid esters on human cervical carcinoma cells in culture.

Fluorescence diagnosis and photodynamic therapy using 5-aminolevulinic acid (ALA) provide new methods for the detection and treatment of cervical cancer and especially its precursors. However, these techniques are restricted by the rate of uptake of the hydrophilic ALA, its poor diffusion through the bilayer of biological membranes or both. In this study we evaluated the effect of some esterified ALA derivatives on the induction of the endogenous photosensitizer, protoporphyrin IX (PpIX), and the photodamage in cultured human cervical cells (C33-A and CaSki). The kinetics of PpIX accumulation showed that ALA esters, especially the ALA-hexylester (h-ALA), induced significantly faster PpIX formation than ALA at the same concentration (0.5 mM). The PpIX induction showed a dose-dependent characteristic. The highest PpIX values could be achieved by an up to 1.3-13-fold lower concentration of ALA esters than with ALA. Using the Annexin V assay, apoptosis was found to be induced rapidly after irradiation in both ALA- and ALA esters-treated cells. On measuring mitochondrial activity, the incubation with h-ALA induced a more pronounced photodamage. The results indicate that improved or at least comparable photodynamic effects can be achieved by using remarkably lower doses of ALA esters.

Fluorescence-guided resection of glioblastoma multiforme by using 5-aminolevulinic acid-induced porphyrins: a prospective study in 52 consecutive patients.

OBJECT: It has been established that 5-aminolevulinic acid (5-ALA) induces the accumulation of fluorescent porphyrins in glioblastoma multiforme (GBM), a phenomenon potentially exploitable to guide tumor resection. In this study the authors analyze the influence of fluorescence-guided resection on postoperative magnetic resonance (MR) imaging and survival in a series of patients who underwent surgery in the authors' department. METHODS: Fifty-two consecutive patients with GBM received oral doses of 5-ALA (20 mg/kg body weight) 3 hours before induction of anesthesia. Intraoperatively, tumor fluorescence was visualized using a modified operating microscope. Fluorescing tissue was removed whenever it was considered safely possible. Residual enhancement on early postoperative MR imaging was quantified and related to each patient's characteristics to determine which factors influenced resection. Survival was analyzed using the Kaplan-Meier method and multivariate analysis was performed in which the Karnofsky Performance Scale (KPS) score, residual fluorescence, patient age, and residual enhancement on MR images were considered. Intraoperatively, two fluorescence qualities were perceived: solid fluorescence generally reflected coalescent tumor, whereas vague fluorescence mostly corresponded to infiltrative tumor. Complete resection of contrast-enhancing tumor was accomplished in 33 patients (63%). Residual intraoperative tissue fluorescence left unresected for safety reasons predicted residual enhancement on MR images in 18 of the 19 remaining patients. Age, residual solid fluorescence, and absence of contrast enhancement in MR imaging were independent explanatory factors for survival, whereas the KPS score was significant only in univariate analysis. No perioperative deaths and one case of permanent morbidity were encountered. CONCLUSIONS: The observations in this study indicate the usefulness of 5-ALA-induced tumor fluorescence for guiding tumor resection. The completeness of resection, as determined intraoperatively from residual tissue fluorescence, was related to postoperative MR imaging findings and to survival in patients suffering from GBM.

Fluorescence imaging and spectroscopy of 5-aminolevulinic acid induced protoporphyrin IX for the detection of neoplastic lesions in the oral cavity.

BACKGROUND: Semiquantitative fluorescence measurements following topical application of 5-aminolevulinic acid (5-ALA) in 16 patients with neoplastic lesions of the oral cavity were performed. METHODS: Time course and type of porphyrin accumulation were analyzed in neoplastic and surrounding healthy tissue by measuring emission spectra of 5-ALA-induced protoporphyrin IX fluorescence. Fluorescence images in the red and green spectral range from the tumor tissue were recorded with a charge-coupled device camera. RESULTS: Protoporphyrin IX fluorescence was detected in the oral mucosa of all patients after local application of 5-ALA. Protoporphyrin IX in neoplastic tissue accumulated earlier in comparison with the surrounding normal tissue. The maximum fluorescence contrast of 10:1 between tumor and host tissue was generally seen 1-2 hours after application, allowing a demarcation of tumor tissue even with the naked eye. CONCLUSION: Labeling of mucosal lesions of the oral cavity with Protoporphyrin IX fluorescence induced by the local application of 5-ALA seems to be a promising diagnostic procedure for neoplastic lesions that are difficult to visualize under white light examination. It is the aim of further investigations to evaluate the relevance of this new diagnostic procedure as a noninvasive and sensitive method for patients with oral cancer.

Detection of squamous cell carcinoma of the oral cavity by imaging 5-aminolevulinic acid-induced protoporphyrin IX fluorescence.

OBJECTIVES: Early cancer detection is the best way to improve the prognosis of patients with oral cancer. Therefore this study presents quantitative fluorescence measurements and results in the visualization of cancerous oral mucosa with 5-aminolevulinic acid (5-ALA)-induced protoporphyrin IX (PPIX). METHODS: Time progression and type of porphyrin accumulation were analyzed in neoplastic and surrounding healthy tissue of 58 patients with a suspected cancer of the oral cavity by measuring emission spectra of 5-ALA-induced PPIX fluorescence. Fluorescence images in the red and green spectral range from the tumor tissue were recorded with a charge-coupled device camera. RESULTS: After topical application of 0.4% 5-ALA and incubation for 1 to 2.5 hours, all patients revealed higher intensities of red fluorescence in neoplastic tissue compared with the surrounding normal tissue. Maximum contrast was reached after 1.5 hours of incubation. In 13.8% (n = 8) of the patients, additional findings like dysplasia, carcinoma in situ, primary tumor, secondary carcinomas, and tumor branches were found by means of fluorescence marking in contrast to white light examination. An evaluation of the biopsy specimens resulted in a specificity of 60% and a sensitivity of 99%. CONCLUSIONS: As a fluorescent marker, PPIX could represent a possible new diagnostic tool to detect early malignant and secondary lesions in the oral cavity. In addition, 5-ALA-induced PPIX fluorescence is promising as a useful intraoperative tool for determining adequate surgical margins of resection. Further investigations aim to assess this diagnostic procedure as a sensitive and clinically reliable method for patients with oral cancer.

Clinical experience with the integral photodynamic therapy of bladder carcinoma.

Photodynamic therapy (PDT) was given to 20 patients who had recurring superficial tumours after unsuccessful application of other treatments. The phototherapeutic results were evaluated by check-ups at 3 month intervals (endoscopy, cytology, bladder mapping, renal ultrasonography) and computed tomography (CT) examination at 8-13 month intervals. In six patients treated with PDT no tumour recurrence was found over the whole observation period up to nearly 5 years. Four patients remained free of tumour (12 and 14 months) after repeated transurethral resection (TUR) and Nd:YAG laser therapy following PDT. Due to an initial application of insufficient irradiation, four patients required a second photodynamic treatment. In one of these cases a circumscribed dysplasia which appeared at the left ostium 26 months following PDT was treated successfully using the Nd:YAG laser following TUR. In six patients slight mucosal atypia persisted for a period of at least 2.5 years. One cystectomy had to be performed because of bladder shrinkage. The dissected bladder was free of tumour. According to these preliminary results, PDT with strict patient selection (worst case situation with recommended cystectomy) is justified in the case of recurrent superficial TIS bladder carcinoma.

Fluorescence staining of oral cancer using a topical application of 5-aminolevulinic acid: fluorescence microscopic studies.

INTRODUCTION: Topical application of 5-aminolevulinic acid (5-ALA) by means of a rinsing solution has been shown to be a promising new procedure in the diagnosis of oral malignancies. However, for assessing the reliability of this method regarding fluorescence-guided tumor resections and photodynamic therapy, further information on the distribution and penetration depth of 5-ALA-induced protoporphyrin IX (PPIX) in the tissue is needed. METHODS: 24 patients suffering from oral cancer were included in this investigation. Biopsies were taken immediately after fluorescence examination and either used as native sections for immediate fluorescence microscopic examination (n = 3) or shock frozen in liquid nitrogen and prepared as frozen sections (n = 46). Fluorescence imaging and digital image processing were utilized in order to determine the presence of PPIX in regions of various histologies as well as the penetration depth of PPIX into solid tumor. RESULTS: PPIX fluorescence in the tissue was limited to the epithelium. Both normal and dysplastic epithelium showed PPIX fluorescence. In the stroma, no PPIX fluorescence was found. In some cases (n = 3/4) invasive carcinomas did not show PPIX fluorescence, while the adjacent or overlying normal epithelium was strongly fluorescent. The penetration depth of PPIX after topical application of 5-ALA was found to be limited to less than 1 mm. CONCLUSION: PPIX fluorescence induced by topical application of 5-ALA can be very useful in the determination of superficial tumor margins. However, due to the limited penetration depth there is a risk of not accurately recognizing the infiltration depth of solid tumors. The aim of further investigations will be to assess the tissue distribution and depth of penetration of PPIX following systemic application of 5-ALA.

Pharmacokinetics of 5-aminolevulinic acid-induced protoporphyrin IX in skin and blood.

The fluorescence and photosensitivity of endogenously synthesized protoporphyrin IX (PPIX) is increasingly used for the diagnosis and treatment of malignant and certain non-malignant diseases. A selective accumulation of PPIX can be induced by application of 5-aminolevulinic acid (5-ALA), which is a precursor of PPIX in the cellular biosynthetic pathway of heme. The purpose of this study was to monitor the in vivo accumulation of PPIX in different locations of the skin after oral ingestion and to determine the pharmacokinetics of 5-ALA and PPIX in human blood plasma for various routes of application. At the same time we wanted to achieve an optimal treatment scheme but also study possible side-effects of 5-ALA administration. After oral application of 5-ALA in a concentration of 40 mg kg-1 body weight, the fluorescence intensities of PPIX in the skin showed maxima between 6.5 and 9.8 h depending on the location and decreased to values lower than 5% related to the maximum after a mean time of about 40 h. The measured absolute intensities of PPIX fluorescence varied strongly between different patients and different locations on one patient. In the plasma of blood samples, PPIX could be detected via its fluorescence for all studied routes of application with the exception of the ointment, where PPIX levels were below the detection limit of 1 microgram l-1. The highest mean concentration of 742 micrograms l-1 PPIX in the plasma was measured 6.7 h after oral application. For inhalation of 5-ALA, a mean maximum concentration of 12 micrograms l-1 could be detected 4.1 h after application, for intravesical instillation, the mean maximum concentration was found to be 1 microgram l-1 2.9 h after application. The kinetics of 5-ALA in the plasma peaked much earlier with a maximum concentration of 32 mg l-1 about 30 min. after oral administration. The 5-ALA levels did not exceed normal reference values after topical application. The results of our experiments suggest that for a systemic application of 5-ALA side-effects in sensitive patients cannot be excluded.