Mechanistic studies on δ-aminolevulinic acid uptake and efflux in a mammary adenocarcinoma cell line

δ-aminolevulinic acid (ALA) is the precursor in the biosynthesis of porphyrins. The knowledge of both the regulation of ALA entrance and efflux from the cells and the control of porphyrin biosynthesis is essential to improve ALA-mediated photodynamic therapy. In this work, we studied the regulation of ALA uptake and efflux by endogenously accumulated ALA and/or porphyrins in murine mammary adenocarcinoma cells. Under our set of conditions, the haem synthesis inhibitor succinyl acetone completely prevented porphobilinogen and porphyrin synthesis from ALA, and led to an increase in the intracellular ALA pool. However, neither intracellular ALA nor porphyrin pools regulate ALA uptake or efflux during the first 15 min of the process. Based on temperature dependence data, ALA but not γ-aminobutyric acid (GABA) efflux is mediated by a diffusion mechanism. Moreover, the addition of extracellular GABA not only did not influence the rate of ALA efflux but on the contrary it affected ALA uptake, showing the contribution of a saturable mechanism for the uptake, but not for the efflux of ALA from the cells. © 2003 Cancer Research UK.Fil:Correa García, S. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina.Fil:Casas, A. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina.Fil:Perotti, C. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina.Fil:Batlle, A. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina.Fil:Bermúdez Moretti, M. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina

δ-Aminolevulinic acid transport in murine mammary adenocarcinoma cells is mediated by beta transporters

δ-aminolevulinic acid, the precursor of porphyrin biosynthesis has been used to induce the endogenous synthesis of the photosensitiser protoporphyrin IX for photodynamic therapy in the treatment of various tumours. The aim of this work was to characterise the δ-aminolevulinic acid transport system in the murine mammary adenocarcinoma cell line LM3 using 14C-δ-aminolevulinic acid, to finally improve δ-aminolevulinic acid incorporation in mammalian cells. Our results showed that δ-aminolevulinic acid is incorporated into these cells by two different mechanisms, passive diffusion which is important at the beginning of the incubation, and active transport. Specificity assays suggested that the transporter involved in δ-aminolevulinic acid incorporation is a BETA transporter, probably GAT-2

Early neoplastic and metastatic mammary tumours of transgenic mice detected by 5-aminolevulinic acid-stimulated protoporphyrin IX accumulation

A photodynamic technique for human breast cancer detection founded upon the ability of tumour cells to rapidly accumulate the fluorescent product protoporphyrin IX (PpIX) has been applied to transgenic mouse models of mammary tumorigenesis. A major goal of this investigation was to determine whether mouse mammary tumours are reliable models of human disease in terms of PpIX accumulation, for future mechanistic and therapeutic studies. The haeme substrate 5-aminolevulinic acid (5-ALA) (200 mg kg−1) was administered to mouse strains that develop mammary tumours of various histological subtypes upon expression of the transgenic oncogenes HRAS, Polyoma Virus middle T antigen, or Simian Virus 40 large T antigen in the mammary gland. Early neoplastic lesions, primary tumours and metastases showed consistent and rapid PpIX accumulation compared to the normal surrounding tissues, as evidenced by red fluorescence (635 nm) when the tumours were directly illuminated with blue light (380–440 nm). Detection of mouse mammary tumours at the stage of ductal carcinoma in situ by red fluorescence emissions suggests that enhanced PpIX synthesis is a good marker for early tumorigenic processes in the mammary gland. We propose the mouse models provide an ideal experimental system for further investigation of the early diagnostic and therapeutic potential of 5-ALA-stimulated PpIX accumulation in human breast cancer patients

ALA and ALA hexyl ester induction of porphyrins after their systemic administration to tumour bearing mice

The use of synthetic lipophilic molecules derived from 5-aminolevulinic acid (ALA) is currently under investigation to enhance cellular ALA penetration. In this work we studied the effect of systemic administration to mice of the hexyl ester of ALA (He-ALA) on porphyrin tissue synthesis as compared to ALA. In most normal tissues as well as in tumour, He-ALA induced less porphyrin synthesis than ALA after its systemic administration either intravenous or intraperitoneal, although explant organ cultures exposed to either ALA or He-ALA revealed equally active esterases. The only tissue that accumulated higher porphyrin levels from He-ALA (seven times more than ALA) was the brain, and this correlated well with a rapid increase in ALA/He-ALA content in brain after administration of He-ALA. This may be ascribed to a differential permeability to lipophilic substances controlled by the blood–brain barrier, a feature which could be further exploited to treat brain tumours

Optimum porphyrin accumulation in epithelial skin tumours and psoriatic lesions after topical application of δ-aminolaevulinic acid

Photodynamic therapy with topically applied δ-aminolaevulinic acid is used to treat skin tumours by employing endogenously formed porphyrins as photosensitizers. This study examines the time course of porphyrin metabolite formation after topical application of δ-aminolaevulinic acid. Porphyrin biosynthesis in human skin tumours (basal cell carcinoma, squamous cell carcinoma), in psoriatic lesions, and in normal skin was investigated. Skin areas were treated with δ-aminolaevulinic acid, and levels of total porphyrins, porphyrin metabolites and proteins were measured in samples excised after 1, 2, 4, 6, 9, 12 and 24 h. There was an increase in porphyrin biosynthesis in all tissues with maximum porphyrin levels in tumours between 2 and 6 h and in psoriatic lesions 6 h after treatment. The pattern of porphyrins showed no significant difference between normal and neoplastic skin, protoporphyrin being the predominant metabolite. The results suggest that optimum irradiation time for superficial epithelial skin tumours may be as soon as 2 h after application of δ-aminolaevulinic acid, whereas for treatment of psoriatic lesions an application time of 6 h is more suitable. © 1999 Cancer Research Campaig

European temporal trends in the use of lymph node dissection in patients with renal cancer

BACKGROUND: The role of lymph node dissection (LND) in renal cell carcinoma (RCC) is still under debate. We aimed to assess the utilization rates of LND over time in Europe. METHODS: A multi-institutional database of 13,581 RCC patients who underwent radical nephrectomy (RN) or nephron sparing surgery (NSS) between 1988 and 2014 was created within an European consortium. We analysed temporal trends in the frequency of LND by using Joinpoint regression. Logistic regression models were used to identify predictors of LND. RESULTS: Overall, 5114 patients (42.7%) underwent LND. Lymph node invasion was recorded in 566 cases (11% of LND patients) which represents 4.7% of the whole study cohort. A gradual decline in the use of LND started in the 1990s. After 2008 LND decreased significantly by 21.5% per year (95%CI -33.3 to -7.5, p < 0.01) until 2011 and stabilized thereafter (Annual Percentage Change 4.9%, 95%CI -3.4 to 13.8, p = 0.2). At multivariable analyses, patient age (OR 0.98, p < 0.0001), type of surgery (RN vs. NSS: OR 5.46, p < 0.0001), surgical approach (open vs. minimally invasive: OR 1.75, p < 0.0001), T stage (T2 vs. T1: OR 1.57; T3-4 vs. T1: OR 1.44, p < 0.0001), clinical tumour size (OR 1.14, p < 0.0001), and year of surgery (OR 0.95, p < 0.0001) were associated with higher probability of LND at nephrectomy. CONCLUSIONS: A trend towards lower LND was observed over time for RCC patients who underwent RN or NSS. LND is more frequently performed in younger patients, locally advanced diseases and in case of open surgery