The role of salvage extended lymph node dissection in patients with rising PSA and PET/CT scan detected nodal recurrence of prostate cancer.

BACKGROUND: Non-prostatic bed recurrence of prostate cancer (PCa) is usually treated with androgen deprivation therapy (ADT). We analyzed the impact of salvage extended lymph node dissection (sLND) on cancer control in patients with rising PSA and lymph node (LN) metastases. METHODS: Between 2009 and 2016 we performed sLND in 87 patients with biochemical recurrence (BCR) and positive LNs on 18FEC and 68Ga-PSMA positron emission tomography/X-ray computer tomography (PET/CT) after primary treatment (PT) of PCa. Intra- and postoperative complications according to Clavien-Dindo were assessed and the rates of biochemical response (BR), BCR-free and clinical recurrence (CR)-free survival, as well as time to initiation of systemic treatment were evaluated. RESULTS: Mean age of patients and mean PSA at sLND was 66.7 years (46-80 years) and 2.63 ng ml-1 (1.27-3.75 ng ml-1), respectively. With 87.4% radical prostatectomy (RP) was the most common PT. In all, 57.9% of patients additionally underwent adjuvant/salvage radiation therapy (RT) and 18.4% received ADT before sLND. Complete BR (cBR) was diagnosed in 27.5% of patients and incomplete BR in 40.6%. In total, 62.2% of patients remained without ADT at follow-up. With a median follow-up of 21 months (1-75 months), the cancer-specific mortality rate was 3.7%. The 3-year BCR-free, systemic therapy-free and CR-free survival rates for patients with cBR were 69.3%, 77.0% and 75%, respectively. CONCLUSIONS: sLND can be performed without significant complications and achieves an immediate BR, thus allowing a significant postponement of systemic therapy in selected patients with BCR and nodal recurrence of PCa. Therefore, sLND following 68Ga-PSMA PET/CT should be considered as part of a multimodal diagnostic and treatment concept for selective patients.

[Video-based teaching in pathology. Experience gained in the last 3 years at the RWTH Aachen University]. / Videobasierte Lehre der Pathologie. Erfahrungen der letzten drei Jahre an der RWTH Aachen.

Modern computer technology provides students with easier access to learning materials. Basic knowledge of pathological findings in organs is essential in medical education. We have produced didactic videos for teaching pathology in a clinical context in addition to regular lectures at the university. Didactic material includes macroscopic and histological findings, as well as cartoons explaining pathophysiology and clinical links. Videos can be downloaded in mv4 format as podcasts to a local hard disk or to an iPhone or iPod via iTunes University and are designed to improve classical medical literature. Analysis over 3 years of server traffic and subjective impressions by the students revealed regular use and high acceptance by users. Didactic material in clinical pathology can be successfully integrated in videos to complement lectures and practical training. Modern teaching methods in pathology make the specialty more understandable and therefore more attractive for students.

The extracellular matrix protein ITIH5 is a novel prognostic marker in invasive node-negative breast cancer and its aberrant expression is caused by promoter hypermethylation.

Inter-alpha-trypsin inhibitors (ITIs) are protease inhibitors stabilizing the extracellular matrix. ITIs consist of one light (bikunin) and two heavy chains (ITIHs). We have recently characterized ITIH5, a novel member of the ITIH gene family, and showed that its messenger RNA is lost in a high proportion of breast tumours. In the present study, an ITIH5-specific polyclonal antibody was generated, validated with western blot and used for immunohistochemical analysis on a tissue microarray; ITIH5 was strongly expressed in epithelial cells of normal breast (n=11/15), while it was lost or strongly reduced in 42% (92/217) of invasive breast cancers. ITIH5 expression in invasive carcinomas was associated with positive expression of oestrogen receptor (P=0.008) and histological grade (P=0.024). Correlation of ITIH5 expression with clinical outcome revealed that patients with primary tumours retaining abundant ITIH5 expression had longer recurrence-free survival (RFS; P=0.037) and overall survival (OS; P=0.044), compared to those with reduced expression (mean RFS: 102 vs 78 months; mean OS: 120 vs 105 months). Methylation-specific PCR analysis frequently showed strong methylation of the ITIH5 promoter in primary breast tumours (41%, n=109) and breast cancer cell lines (n=6). Methylation was significantly associated with mRNA loss (P<0.001; n=39), and ITIH5 expression was induced after treatment of tumour cell lines with the demethylating agent 5-aza-2'-deoxycytidine. Moreover, ITIH5 promoter methylation was significantly associated with reduced OS (P=0.008). The cellular function of ITIH5 was evaluated by forced expression of a full-length ITIH5 complementary DNA in the breast cancer cell line MDA-MB-231, which does not endogenously express ITIH5. ITIH5-expressing clones showed a 40% reduced proliferation rate compared to mock-transfected cells. Overall, these data show that promoter methylation-mediated loss of ITIH5 expression is associated with unfavourable outcome in breast cancer patients, and thus ITIH5 could be used as a prognostic marker, although this marker is not multivariate independent due to its close association with ER expression. Our data indicate that ITIH5 is a candidate class II tumour suppressor gene and could be involved in tumour progression, invasion and metastasis, as its absence is associated with increased proliferation rates and a prognostic value indicating poor clinical outcome.

Value of multicolour fluorescence in situ hybridisation (UroVysion) in the differential diagnosis of flat urothelial lesions.

AIMS: During the past 10 years, multitarget fluorescence in situ hybridisation has been established as a valuable adjunct in the cytological diagnosis of precancerous and malignant lesions of the urinary tract. The aim of the present study was to define its value in detecting chromosomal imbalances in patients with various flat urothelial lesions in routine paraffin-embedded bladder biopsy samples. In addition, the HER2 gene amplification and HER2 expression pattern were examined, since alterations of the HER2 expression patterns have been demonstrated in invasive bladder cancer. METHODS: 29 samples of normal urothelium and 86 flat urothelial lesions (hyperplasia, reactive atypia, dysplasia and carcinoma in situ (CIS)) from 73 patients were analysed patients using tissue microarrays and centromeric probes for chromosomes 3, 7 and 17, and gene-specific probes for 9p21/P16 and HER2 (UroVysion, PathVysion). The expression of HER2 was studied by immunohistochemistry. RESULTS: Polysomy of at least one of the chromosomes was found in about half of the dysplastic cells, and in more than 90% of cells in CIS or cells in invasive bladder tumours. Polysomic cells were found in only 17% of urothelial hyperplasia, reactive atypia and normal urothelium of healthy patients, whereas about 30% of non-neoplastic lesions in patients with concomitant urothelial carcinoma showed polysomy of at least one chromosome. These alterations indicate a field effect and are associated with synchronous development of dysplastic lesions of a higher grade. Deletion of the P16 locus was most frequently observed in aneuploid lesions, whereas overexpression of HER2 was found in 10-20% of invasive urothelial carcinomas, and only occasionally in CIS (5%). An altered HER2 expression pattern was present in non-neoplastic lesions (25%). CONCLUSIONS: UroVysion fluorescence in situ hybridisation is a valuable tool for the detection of genetically unstable flat urothelial lesions, and can help to resolve difficult cases, particularly the differential diagnosis of reactive atypia and dysplasia.

Analyzing effects of photodynamic therapy with 5-aminolevulinic acid (ALA) induced protoporphyrin IX (PPIX) in urothelial cells using reverse phase protein arrays.

Photodynamic therapy (PDT) using 5-aminolevulinic acid (ALA)-induced protoporphyrin IX (PPIX) is clinically established approach for a number of defined applications. However, in order to optimize the therapeutic benefits of PDT, the specific mode of cell destruction should be better defined. Apoptosis is favored over necrosis for clinical practice as the latter causes more side-effects. In the present study, we analyse PDT-induced cell death and its correlation to various PDT parameters (different doses applied, time after PDT treatment) in vitro using reverse phase protein arrays. Human urothelial cell lines with varying degrees of differentiation (UROtsa, RT4, RT112, J82) were subjected to in vitro-PDT using increasing doses of irradiation. In addition, positive controls for apoptosis, necrosis and un-/specific cellular damage were included. Cells were harvested over a specified time course, lysed and arrayed onto nitrocellulose-covered glass slides. The arrays were analyzed for expression of apoptosis-related proteins by immunohistochemistry. Analysis of caspase-3 and -9 expression, the activation of HIF-1alpha, Bcl2, Cox2 and the phosphorylation of AKT reveals signal activation due to a PDT-stimulus in correlation with the positive controls. Data were analyzed by unsupervised hierarchical clustering and depicted as a heat map revealing cell-specific patterns of pathway stimulation. Higher differentiated phenotypes showed a more distinct signal response in general and a higher apoptotic response in detail. Lower differentiated cell lines lost pathway regulation capabilities according to their state of dedifferentiation. Reverse phase protein arrays are a promising technique for signal pathway profiling: they exceed the range of traditional western blots by sensitivity, high-throughput capability, minimal sample consumption and easy quantification of results obtained.

[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.

Can an endocytoscope system (ECS) predict histology in neoplastic lesions?

BACKGROUND AND STUDY AIMS: An endocytoscope system (ECS) has recently been developed with the possibility of super-high magnification of gastrointestinal mucosa, thus allowing in vivo imaging of living cells. The aim of the present study was to assess the potential of ECS in the prediction of histology in both normal gastrointestinal mucosa and neoplastic lesions. PATIENTS AND METHODS: In total, 76 patients (57 men, 19 women; age range 37-86 years) with neoplastic lesions in the esophagus, stomach, or colon were enrolled into the study and underwent esophagogastroduodenoscopy or colonoscopy. After staining with 1% methylene blue, the mucosa was examined with the ECS probe (x 450 and x 1100 magnification), and video sequences were recorded on video disk. Biopsies from the examined areas were taken for histology and served as the gold standard. The endocytoscope video sequences were evaluated by two blinded pathologists. Finally the results were compared with those resulting from the evaluation of an experienced endoscopist who was aware of the macroscopic endoscopic pictures and the endocytoscope image results. RESULTS: A total of 25 patients with esophageal lesions, 28 patients with colonic lesions, and 23 patients with gastric lesions were examined. The sensitivity and specificity for the evaluation of the blinded pathologists was 81% and 100%, respectively, in the esophagus, 56% and 89% in the stomach, and 79% and 90% in the colon. If an endoscopist evaluated the endocytoscopic pictures in combination with the macroscopic endoscopic images sensitivity and specificity increased significantly. CONCLUSIONS: First experiences with ECS show good sensitivity rates even by blinded assessment for esophageal and colonic lesions. Sensitivity for neoplastic lesions in the stomach is lower because of gastric mucous secretion. Combining the endoscopic and cytoscopic appearance of the lesion may further enhance the diagnostic value of the method.

Artificial intelligence and bladder cancer arrays.

Non-muscle invasive bladder cancer is a heterogenous disease whose management is dependent upon the risk of progression to muscle invasion. Although the recurrence rate is high, the majority of tumors are indolent and can be managed by endoscopic means alone. The prognosis of muscle invasion is poor and radical treatment is required if cure is to be obtained. Progression risk in non-invasive tumors is hard to determine at tumor diagnosis using current clinicopathological means. To improve the accuracy of progression prediction various biomarkers have been evaluated. To discover novel biomarkers several authors have used gene expression microarrays. Various statistical methods have been described to interpret array data, but to date no biomarkers have entered clinical practice. Here, we describe a new method of microarray analysis using neurofuzzy modeling (NFM), a form of artificial intelligence, and integrate it with artificial neural networks (ANN) to investigate non-muscle invasive bladder cancer array data (n=66 tumors). We develop a predictive panel of 11 genes, from 2800 expressed genes, that can significantly identify tumor progression (average Logrank p = 0.0288) in the analyzed cancers. In comparison, this panel appears superior to those genes chosen using traditional analyses (average Logrank p = 0.3455) and tumor grade (Logrank, p = 0.2475) in this non-muscle invasive cohort. We then analyze panel members in a new non-muscle invasive bladder cancer cohort (n=199) using immunohistochemistry with six commercially available antibodies. The combination of 6 genes (LIG3, TNFRSF6, KRT18, ICAM1, DSG2 and BRCA2) significantly stratifies tumor progression (Logrank p = 0.0096) in the new cohort. We discuss the benefits of the transparent NFM approach with respect to other reported methods.

[Urine cytology and urine markers. Significance for clinical practice]. / Urinzytologie und Urinmarker. Bedeutung für die Praxis.

Urothelial carcinoma of the bladder is a frequent disease that can be identified timely by screening patients at high risk. Due to the high rate of disease recurrence, frequent follow-up procedures are necessary. For this purpose, cystoscopy is the standard procedure, and supplementary non-invasive procedures such as cytology or tumor marker tests are used. These tests have different advantages and disadvantages in terms of their sensitivities and specificities. Thus, they provide additional information, but are not able to replace cystoscopy as the standard instrument in the diagnosis of bladder cancer.

[Urinary cytology. Possibilities, limitations and the future]. / Die Urinzytologie. Möglichkeiten, Grenzen und Zukunft.

Even now, 60 years after the fundamental studies on oncological urinary cytology carried out by Papanicolaou and Marschall and the subsequent integration of the test they devised into the diagnostic investigations applied in the diagnosis of urothelial carcinoma, urinary cytology still maintains its place in the diagnosis of primary and recurrent tumours of the urinary tract. Newer diagnostic techniques involving urine-bound tumour markers have not so far achieved such high levels of acceptance as their method. It is possible, certainly, that a combination of these newer methods with cytological testing, or with other innovative diagnostic methods, such as photodynamic techniques, could prove very promising in the future and might overcome the limitations of urinary cytology.

New 5-aminolevulinic acid esters--efficient protoporphyrin precursors for photodetection and photodynamic therapy.

Photodetection (PD) and photodynamic therapy (PDT) with 5-aminolevulinic acid (ALA)-induced protoporphyrin IX (PPIX) accumulation are approaches to detect and treat dysplasia and early cancer in the gastrointestinal tract and in the urinary bladder. Because ALA-induced PPIX production is limited, we synthesized ALA ester hydrochlorides 3-22 and tested them in two different in vitro models (gastrointestinal tract: HT29-CCD18; urinary bladder: J82-UROTSA). PPIX accumulation after incubation with 0.12 mmol/L for 3 h and PPIX accumulation as a function of different incubation times were measured using flow cytometry. 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assays were performed to check cellular dark toxicity. Phototoxicity after irradiation was tested. ALA nonafluorohexylester hydrochloride 11, ALA thiohexylester hydrochloride 13 and ALA dibenzyldiester dihydrochloride 19 induced appreciably increased PPIX levels and showed improved phototoxicity compared with the references ALA hydrochloride 1, ALA hexylester hydrochloride 3 and ALA benzylester hydrochloride 4. Thus, the new compounds 11, 13 and 19 are promising compounds for PD and PDT.

[Reducing the risk of superficial bladder cancer recurrence with 5-aminolevulinic acid-induced fluorescence diagnosis. Results of a 5-year study]. / Senkung des Rezidivrisikos oberflächlicher Harnblasenkarzinome mittels 5-Aminolävulinsäure-induzierter Fluoreszenzdiagnostik. Resultate einer 5-Jahres-Studie.

A prospective monocentre randomized parallel-group Phase III trial was performed to investigate whether primary transurethral resection (TUR) with 5-aminolevulinic acid induced Fluorescence diagnosis (FD) allows for a more thorough TUR of superficial Bladder Carcinoma compared to conventional white light (WL). Evaluation of residual tumor rate and recurrence free survival were defined as the two primary study endpoints. The residual tumor rate was 25.2% in the WL arm (n=103) vs. 4.5% in the (n=88) FD arm (p<0.0001). Median follow up of the patients in the WL arm was 42 months (range 25-61) compared to 43 (range 24-61) in the FD arm. Recurrence free survival in the fluorescence diagnosis group was 90.9%, 90.9% und 85 % after 12, 24 and 48 months compared with 78.6%, 69.9% und 60.7 %, respectively, in the white light group (p=0.0005). This superiority proved to be independent of risk group. The adjusted hazard ratio of fluorescence diagnosis versus white light transurethral resection was 0.29 (95% CI: [0.15; 0.56]). ALA induced FD is statistically significantly superior to conventional WL TUR with respect to both residual tumor rate and recurrence-free survival. The differences in RFS imply that FD offers a clinically relevant procedure to reduce the number of tumor recurrences.

Optimization of three- and four-color multiparameter DNA analysis in lymphoma specimens.

BACKGROUND: Simultaneous analysis of DNA and immunophenotype of lymphoma cells by flow cytometry allows the calculation of the proliferative activity and aneuploidy in even a small lymphoma population. Unfavorable DNA binding characteristics or spectral features of DNA dyes impair the accuracy of multiparameter DNA analysis and limit their clinical application. We describe here a reliable and reproducible application of both three- and four-color multiparameter DNA analysis. METHODS: After immunostaining of fresh samples of peripheral blood, bone marrow and single cell suspensions of lymph nodes from healthy and lymphoma patients, a methanol fixation for TO-PRO-3 and DRAQ5 staining was tested. RESULTS: The red-excitable TO-PRO-3 on a FACSCalibur is limited to two-color antigen staining including fluorescein-isothiocyanate and phycoerythrin-labeled monoclonal antibodies due to its broad excitation spectrum. Although DRAQ5 is only applicable to flow cytometers equipped with a single argon laser emitting 488-nm light, its emission spectrum can be easily separated from the FITC, PE, and PE/Texas-Red emissions. DRAQ5 showed almost identical stoichiometric DNA binding characteristics as propidium iodide. Coefficient of variation produced by DRAQ5 staining is in the range of 3.5 and is adequate for detecting aneuploid amd near-diploid cells. CONCLUSIONS: These advantageous features of DRAQ5 make it a reliable candidate for multiparameter clinical studies.

Frequent genetic alterations in flat urothelial hyperplasias and concomitant papillary bladder cancer as detected by CGH, LOH, and FISH analyses.

Flat urothelial hyperplasia, defined as markedly thickened urothelium without cytological atypia, is regarded in the new WHO classification as a urothelial lesion without malignant potential. Frequent deletions of chromosome 9 detected by fluorescence in situ hybridization (FISH) have been previously reported in flat urothelial hyperplasias found in patients with papillary bladder cancer. Using comparative genomic hybridization (CGH) and microsatellite analysis, these hyperplasias and concomitant papillary tumours of the same patients were screened for other genetic alterations to validate and extend the previous findings. Eleven flat hyperplasias detected by 5-ALA-induced fluorescence endoscopy and ten papillary urothelial carcinomas (pTaG1-G2) from ten patients were investigated. After microdissection, the DNA of the lesions was pre-amplified using whole genome amplification (I-PEP-PCR). Loss of heterozygosity (LOH) analyses were performed with five microsatellite markers at chromosomes 9p, 9q, and 17p. CGH was performed using standard protocols. In 6 of 11 hyperplasias and 7 of 10 papillary tumours, deletions at chromosome 9 were simultaneously shown by FISH, LOH, and CGH analyses. There was a good correlation between FISH, LOH, and CGH analyses, with identical results in 6 of 10 patients. In addition to deletions at chromosome 9, further genetic alterations were detected by CGH in 9 of 10 investigated hyperplasias, including changes frequently found in invasive papillary bladder cancer (loss of chromosomes 2q, 4, 8p, and 11p; gain of chromosome 17; and amplification at 11q12q13). There was considerable genetic heterogeneity between hyperplasias and papillary tumours, but a clonal relationship was suggested by LOH and/or CGH analyses in 5 of 10 cases. These data support the hypothesis that flat urothelial hyperplasias can display many genetic alterations commonly found in bladder cancer and could therefore be an early neoplastic lesion in the multistep development of invasive urothelial carcinoma.

Do patients profit from 5-aminolevulinic acid-induced fluorescence diagnosis in transurethral resection of bladder carcinoma?

OBJECTIVES: To evaluate in a prospective study the influence of fluorescence diagnosis (FD) controlled transurethral resection of bladder tumors on therapeutic consequences. The aim was to determine in how many patients FD led to a change in treatment strategy compared with conventional white light (WL) cystoscopy. METHODS: A total of 279 patients with suspected bladder tumors underwent transurethral resection using FD in addition to WL cystoscopy. The number of additional tumor-positive patients, staging change, number of multilocular tumors exclusively detected by FD, and resulting therapeutic consequences compared with the results after WL cystoscopy were investigated. In addition a biopsy-based evaluation was performed. RESULTS: Tumor or dysplasia II degrees (moderate dysplasia) was detected in 177 patients. In 168 patients, tumor was detected by WL cystoscopy, and in 9 (5.1%) of the patients, tumor was completely overlooked by WL cystoscopy and diagnosed exclusively by FD (n = 3 TaG1-G2, n = 2 carcinoma in situ, n = 1 greater than T1, and n = 3 dysplasia II degrees ). Multilocular tumor involvement was detected in 10 cases using FD, and a change in the stage by detection of coexisting dysplasia II degrees and carcinoma in situ occurred in 8 patients. In 27 patients (15.3%), additional information was obtained by exclusive detection of tumors by FD. This resulted in a change in the treatment strategy for 16 patients (9%). CONCLUSIONS: FD leads to an improvement in the diagnosis of bladder carcinoma. It allows the early selection of the best treatment option and thus has a potentially positive effect on the prognosis of the affected patients.

Tumor-associated fibroblasts (part II): Functional impact on tumor tissue.

The article focuses on the functional impact of tumor-associated fibroblasts (TAF) on its surrounding cells. It intends to cover the recent knowledge on TAF, the phenotype, and expression profile of which have been described in the first part of the review series (Kunz-Schughart and Knuechel, 2002). The present review is subdivided into two main chapters: (1) functional impact of TAF on tumor cells and (2) fibroblast-host cell interactions in tumor tissue. In the first paragraph of chapter (1) about the role of fibroblasts in tumor cell growth and differentiation it is revealed, how strongly cellular interaction is dependent on fibroblast and tumor cell type as well as the spatial ratio between the cells. The variation of cellular behavior depending on quantity of molecules holds also true for the group of ECM molecules, e.g. the balance between MMPs and TIMPs, which provide an interesting therapeutic target in tumor tissue. This is one of the topics addressed in the second paragraph which focuses on tumor cell dissemination. Chapter (2) addresses the relation of TAF to other intra- or peritumoral host cells. The hypoxia-related angiogenesis induction of fibroblasts via growth factor secretion (e.g. VEGF) is considered as important as the immune modulatory properties of fibroblasts on immune cells, such as monocytes/macrophages. These cellular properties can be tested under controlled conditions in three-dimensional heterologous cultures of human cells, providing the chance for systematic modification to assess therapeutic effects in an in vivo like environment.

Tumor-associated fibroblasts (part I): Active stromal participants in tumor development and progression?

Phenotypic and functional characteristics of tumor associated fibroblasts (TAF) in contrast to normal fibroblasts are reviewed in this first synopsis (part I). Terms as tumor stroma, desmo-plasia, TAF, myofibroblast, and fetal-type fibroblast are defined, and experimental systems to study heterologous cell interactions are presented. While we only start to gather information on the genotype of TAF, a broad range of data deals with the expression profile of these cells, covering e.g. ECM and ECM-modulating molecules, growth factors and cytokines. Summarizing the recent state of knowledge indicates that TAF provide sources for tumor diagnosis and therapy, that have to be further defined in an organ-specific approach in terms of the functional impact on the tumor cell and its environment (see part II).

Improving the use of 5-aminolevulinic acid (ALA)-induced protoporphyrin IX (PPIX) for the gastrointestinal tract by esterification--an in vitro study.

Possible approaches to improve the diagnostic and therapeutic effects of 5-aminolevulinic acid (ALA)-induced protoporphyrin IX (PPIX) are the esterification of ALA for enhanced uptake and the choice of wavelength for irradiation. The human colonic cell lines HT29 [G2] and CCD18 (fibroblasts) were incubated with 0.6 mM ALA, ALA-hexylester or -benzylester respectively, and for further assays with hypotaurine, in addition. PPIX-accumulation was analyzed by flow cytometry and fluorescence spectroscopy. PPIX formation kinetics were continuously recorded. Incubated cells were irradiated with an incoherent light source lambda = 400-700 nm or lambda = 590-700 nm, respectively. After PDT treatment, clonogenicity assays were performed to determine cell viability. Esterification leads to increased PPIX-accumulation, decreased time for production of detectable amounts of PPIX as well as increased response to PDT. Tumor specificity is always maintained or exceeds values for ALA alone. ALA enters the cells via beta transporter whereas esters by passive diffusion. Altering irradiation wavelengths showed the independence of wavelength rather than light dose. Results emphasize the role of heme metabolism for generating tumor specificity rather than the process of ALA-uptake, an important detail for future clinical application.