The influence of photodynamic therapy with 5-aminolevulinic acid on senescent skin cancer cells.

BACKGROUND: Senescent cells, which are resistant to apoptosis, accumulate with age and after ultraviolet (UV) radiation, chemotherapy and radiation therapy. Preventing or eliminating senescent cells may be crucial for protection against skin cancer development and improving tumour treatment. The aim of the present study was to investigate the potential of photodynamic therapy (PDT) with 5-aminolevulinic acid (ALA) to induce senescence in skin cancer cells and to eliminate senescent cells induced by chemotherapy (bleomycin) or UVA (315-400nm) exposure. METHODS: WM115 and A431 cells were incubated with 1mM ALA for 2 and 4h, respectively, before exposure to blue light (10mW/cm2, 0-80s, 0-0.8J/cm2). WM115 cells were treated once with 106J/cm2 (58.4mW/cm2, 30.25min) UVA 6days before ALA-PDT or with 0.24IU/ml bleomycin for 7days to induce senescence before ALA-PDT. Cell viability was monitored by the MTT colorimetric assay. Senescent cells were detected using senescence-associated-beta-galactosidase (SA-ß-Gal) staining and morphological changes (enlarged, flat cells). RESULTS: ALA-PDT caused a light dose dependent increase in senescence. ALA-PDT induced senescence very effectively only in WM115 cells but not in A431 cells, while similar cytotoxic effects were observed in both cell lines. After ALA-PDT with 0.4J/cm2 around 70% of survived WM115 cells were senescent, while only around 5% of A431 cells were senescent after ALA-PDT with 0.8J/cm2. CONCLUSION: ALA-PDT can induce premature senescence and kill senescent cells induced by ALA-PDT itself, UVA and chemotherapy (bleomycin). Light doses must be properly chosen to photoinactivate ALA-PDT-induced senescent cells.

Malignant melanomas on head/neck and foot: differences in time and latitudinal trends in Norway.

BACKGROUND: Cutaneous malignant melanoma (CMM) incidence continues to increase in many parts of the world. Solar ultraviolet (UV) radiation is the main environmental risk factor for CMM. Different body locations are subjected to different doses and exposure patterns of solar UV. Time and latitudinal trends of CMMs on shielded and exposed skin give valuable information about the aetiology of these cancers. In this study, we have compared the time and latitudinal trends of CMM incidence on skin areas which are chronically (head and neck) and rarely (foot) exposed to UV radiation, to gain more information about the relationship between sun doses, exposure patterns and melanomagenesis. METHODS: We have analysed epidemiological data from the Cancer Registry of Norway, for foot and head and neck CMM for two time periods: 1966-1986 and 1987-2007. RESULTS: Cutaneous malignant melanoma incidence rate on head and neck has increased with time, while incidence rates of foot CMM have remained almost constant with time in Norway. There is a large north-south gradient in incidence rates of CMM on head and neck in Norway, while there is almost no north-south gradient for CMM incidence on foot. CONCLUSIONS: Comparisons of time trends and latitudinal trends of the incidence rates of CMM on head/neck and on foot indicate that solar radiation plays a role in the induction of the former CMM but probably not for the latter.

Comparison of the time and latitude trends of melanoma incidence in anorectal region and perianal skin with those of cutaneous malignant melanoma in Norway.

BACKGROUND: Melanoma incidence is increasing in many parts of the world. The main environmental risk factor is exposure to solar radiation. However, melanomas may arise also on non-sun-exposed areas (uveal and mucosal melanomas) and little is known about a possible relationship between sun exposure and melanoma on such locations. OBJECTIVES: We have compared the time and latitude trends of melanoma incidence in the anorectal region and perianal skin (non-sun-exposed sites) with those of cutaneous malignant melanoma (CMM) (sun-exposed skin) to gain more information about the relationship between sun exposure and melanoma on such sites. METHODS: We analysed epidemiological data from the Cancer Registry of Norway for melanomas of the anorectal mucosa, perianal skin and overall CMM for the time period 1966-2007. RESULTS: We found that melanoma incidence on these shielded sites tends to decrease or remain constant over a period during which the CMM rates increase. This is true both in the North and in the South regions of Norway. Comparison of latitudinal trends of the incidence rates of CMM and melanoma on these shielded sites shows that there is no latitude gradient for melanoma of the anorectal mucosa and perianal skin, whereas there is a strong one for CMM. CONCLUSIONS: The time and latitudinal trends are likely to support the assumption that melanomas on these shielded sites are not generated by ultraviolet radiation. Possible causes and significances of these trends are discussed.

Topical aminolaevulinic acid- and aminolaevulinic acid methyl ester-based photodynamic therapy with red and violet light: influence of wavelength on pain and erythema.

BACKGROUND: Photodynamic therapy (PDT) is based on the combination of an exogenously administered precursor of photosensitizer [protoporphyrin IX (PpIX)] synthesis and exposure to light. Choosing the optimal wavelength is important. Red light penetrates deeper into tissue, while violet light is more efficient in activating PpIX but does not penetrate so deeply. OBJECTIVES: We studied PpIX formation and the PDT effect after application to human skin of creams containing aminolaevulinic acid (ALA) and aminolaevulinic acid methyl ester (MAL). The aim of the study was to investigate whether the wavelength of the light used has an influence on pain sensations during topical PDT with the different prodrugs. METHODS: ALA cream (10%) and MAL cream (10%) were topically applied on the skin of 10 healthy volunteers. After 24 h the application site was exposed to 8 mW cm(-2) violet laser or to 100 mW cm(-2) red laser light. The erythema index was monitored up to 24 h after light exposure. For the first time the pain during topical ALA- and MAL-PDT was assessed by measuring the time taken for pain to occur. Also, for the first time, the intensities of the light sources were calibrated so as to have the same relative quantum efficiency. Results The pain sensation during ALA-PDT with red light came 22 s sooner than during ALA-PDT with violet light, which is statistically significant (P < 0.05). Moreover, ALA-PDT with red light gave stronger and more persistent erythema than ALA-PDT with violet light. ALA induced about three times more PpIX than MAL. No statistically significant differences were found for erythema, or for the time for pain to occur, in the case of MAL-PDT with red vs. violet light. CONCLUSIONS: Topical ALA-PDT with violet light allows longer exposure times before pain is induced and gives less erythema as compared with topical ALA-PDT with red light.

Calcitriol treatment improves methyl aminolaevulinate-based photodynamic therapy in human squamous cell carcinoma A431 cells.

BACKGROUND: Photodynamic therapy (PDT) using methyl aminolaevulinate (MAL) provides a new, approved method for treatment of skin cancer and its precursors. However, MAL-based PDT is not very efficient for poorly differentiated skin carcinoma. Thus, novel strategies to enhance the PDT effect are needed. OBJECTIVES: In order to improve the efficacy of MAL-based PDT, we investigated the effect of adding calcitriol, a prodifferentiation hormone, to human squamous cell carcinoma A431 cells in vitro. METHODS: A short course (24 h) of calcitriol pretreatment was applied in A431 cells, and, subsequently, MAL-induced protoporphyrin IX (PpIX) was measured. RESULTS: Calcitriol pretreatment of the cells elevated their PpIX levels. Furthermore, the cell damage after exposure to blue light was significantly higher in calcitriol-treated cells. Increased photoinactivation correlated with higher levels of PpIX in the calcitriol-pretreated cells. CONCLUSIONS: Calcitriol enhances MAL-based PDT in A431 cells.

Topical application of 5-aminolaevulinic acid, methyl 5-aminolaevulinate and hexyl 5-aminolaevulinate on normal human skin.

BACKGROUND: 5-Aminolaevulinic acid (ALA) and its ester derivatives are used in photodynamic therapy. Despite extensive investigations, the differences in biodistribution and pharmacokinetics of protoporphyrin IX (PpIX) induced by ALA and its derivatives are still not well understood, notably for humans. OBJECTIVES: To study porphyrin accumulation after topical application of ALA and two of its ester derivatives in normal human skin. METHODS: Creams containing 0.2%, 2% and 20% (w/w) of ALA, methyl 5-aminolaevulinate (MAL) and hexyl 5-aminolaevulinate (HAL) were applied on normal human skin of six volunteers. The amount and distribution of porphyrins formed in the skin was investigated noninvasively by means of fluorescence spectroscopy. RESULTS: Fluorescence emission and excitation spectra exhibited similar spectral shapes for the all drugs, indicating that mainly PpIX was formed. Low concentrations (0.2% and 2%) of MAL induced considerably less PpIX in normal human skin than similar concentrations of ALA and HAL. A high concentration (20%) of ALA gave higher PpIX fluorescence in normal human skin than was found for MAL and HAL. CONCLUSIONS: The concentrations inducing half of the maximal PpIX fluorescence are around 2% for ALA, 8% for MAL and 1% for HAL.

Photosensitization with protoporphyrin IX inhibits attachment of cancer cells to a substratum.

Effects of photodynamic therapy (PDT) on adhesion of human adenocarcinoma cells of the line WiDr to a plastic substratum were investigated. Protoporphyrin IX induced by 5-aminolevulinic acid (ALA) was used as a photosensitizer. Light exposure inhibited attachment of suspended cells to a substratum. The adhesion was most strongly pronounced for light exposures around 200 mJ/cm(2) causing cell death. However, sub-lethal exposures (42 mJ/cm(2), 97% survival) inhibited cell adhesion as well. Sub-lethal ALA-PDT increased the intracellular space in dense colonies of WiDr cells. This was attributed to formation of lamellipodia between the cells and to increased numbers of focal contacts containing alpha(V)beta(3) integrin in some of the cells. The E-cadherin distribution was not changed by the treatment. Complex processes, including changes in cellular shape and reorganization of the cytoskeleton, are suggested to participate in the observed ALA-PDT effect on the cell adhesion.

Production of protoporphyrin IX from 5-aminolevulinic acid and two of its esters in cells in vitro and tissues in vivo.

5-Aminolevulinic acid (ALA) and two of its esters were studied in cells in vitro and in vivo on skin of healthy hairless mice. In vitro, both esters, which are more lipophilic than ALA, induced higher PpIX fluorescence at lower concentrations compared with ALA. In vivo, ALA induced PpIX fluorescence more efficiently than the esters. The difference between ALA and the esters may be related to structures in the stratum corneum or to rate of penetration through this skin layer. The stratum corneum may bind the esters temporarily, and slow down their penetration into the living cells where PpIX is formed.