Penetration kinetics of 8-methoxypsoralen after 8-methoxypsoralen bath procedure with and without UVA irradiation.

Administration of 8-methoxypsoralen (8-MOP) in a dilute bath water solution is an effective therapeutic alternative for systemic application of 8-MOP, avoiding systemic side effects such as nausea and cataractogenesis. The aim of our study was to determine the epicutaneous penetration of 8-MOP in a dilute bath water solution with and without additional UVA irradiation in human skin under in vitro conditions. To simulate the PUVA bath procedure, 8 skin samples were exposed to radioactively labeled 8-MOP in a water solution. After 20 min, the test solution was removed and the skin surface was dried. Immediately after the bath procedure, 4 of the skin samples were irradiated with 0.5 J/cm2 UVA. During a test period of 15 h, the 8-MOP penetration was observed. In both test groups (with and without UVA irradiation) 8-MOP permeated through all skin layers between 30 min and 1 h after application. Compared to the unirradiated skin samples, the UVA-irradiated skin samples showed a significantly slower increase and a lower maximum of 8-MOP permeation. Following our results, UVA irradiation of 8-MOP-exposed skin samples led to a significantly decreased permeation rate. This might be due to UVA-induced links between 8-MOP molecules and human DNA. In addition, we investigated the levels of radioactivity emitted by tritium-labeled 8-MOP in stratum corneum, epidermis and dermis up to 30 min after 8-MOP bath in two further test groups with and without additional UVA irradiation. The statistical analysis revealed no significant differences between these two test groups. Thus, the levels of radioactivity remained constant in the epidermis and dermis during the test period of 30 min. Since the levels of radioactivity were constant up to 30 min after UVA irradiation, a previously supposed marked loss of 8-MOP concentration might not be responsible for the rapid extinction of observed in vivo photosensitivity within 1 h after PUVA bath observed in vivo in human skin.     

An action spectrum for the elicitation of erythema in skin persistently sensitized by photobound 8-methoxypsoralen

Human skin can be rendered persistently photosensitive by topical application of aqueous 8-methoxypsoralen (8-MOP) and exposure to a suberythemogenic dose of more than 380 nm radiation. We report an action spectrum for the elicitation of phototoxic erythema induced by a second exposure of skin pretreated in this way. After correction for unsensitized skin erythema this action spectrum resembles the absorption spectrum of the 4',5'-monoadduct of 8-MOP to DNA. This suggests that the monoadduct is the chromophore for erythema elicited by the second irradiation, and supports the DNA crosslink as the crucial photoproduct causing phototoxic erythema due to 8-MOP in human skin.     

A study of the relationship between photosensitizing and therapeutic activity of 4,5',8-trimethylpsoralen, and its major metabolite 4,8-dimethyl, 5'-carboxypsoralen

The molecular basis for the clinically observed differences in the skin photosensitizing activity and therapeutic effectiveness of the topically applied and orally administered drug trimethylpsoralen (TMP) was investigated. TMP, when tested topically, is a very potent photosensitizing and therapeutically effective furocoumarin in the treatment of psoriasis. When administered orally, however, it is significantly less photosensitizing and therapeutically a less effective drug than the commonly used furocoumarin 8-methoxypsoralen. This decreased reactivity of oral TMP is attributable to its poor solubility and rapid in vivo metabolic transformation to several inactive (nonphotosensitizing) metabolites, one of which is referred to as 4,8-dimethyl,5'-carboxypsoralen (DMeCP). The supporting evidence has been obtained by: (a) isolation of the urinary metabolite DMeCP and subsequent comparison of its properties with the synthetically prepared DMeCP and its methyl ester; (b) examining the dark and photochemical interactions of TMP, DMeCP, and DMeCP methyl ester with DNA and determining their ability to form interstrand cross-links with DNA; and (c) studying the inhibition of DNA and RNA synthesis in Ehrlich ascites tumor cells and the killing of bacteria and T2 bacteriophages. The structure-activity relationship of TMP and DMeCP also has been examined in normal human subjects and in patients with psoriasis. The order of topical therapeutic effectiveness in terms of ability to clear psoriasis plaques appeared to be: TMP greater than 8-MOP greater than DMeCP methyl ester greater than DMeCP. The data also suggest the methyl ester of DMeCP to be an interesting nonphotosensitizing furocoumarin that photoconjugates to DNA better than 8-MOP and is therapeutically effective in psoriasis.     

Assessment of minimal phototoxic dose following 8-methoxypsoralen bath: maximal reaction on average after 5 days

An essential procedure before starting bath psoralen ultraviolet (UV) A (PUVA) photochemotherapy is the evaluation of the minimal phototoxic dose (MPD), which is traditionally assessed 3 days after irradiation. However, there are no controlled studies supporting the 72 h peak of bath-PUVA erythema. The aim of this study was therefore to determine the exact time course of the erythematous reaction in human skin following bath-PUVA. For this purpose, the skin of 10 volunteers was exposed to 0.5-3.0 J/cm2 UVA directly after a 20-min 8-methoxypsoralen bath (0.5 mg/L, 37 degrees C). At 24, 48, 72, 96, 120 and 144 h (1-6 days) after irradiation, the MPD and the erythema sum score (ESS) were determined in each subject. The results showed a maximal erythematous reaction on average 5 days after irradiation. The mean MPD gradually decreased from day 2 (> 3.0 J/cm2) to day 5 (mean +/- SD 1.15 +/- 0.63 J/cm2) and started to increase at day 6 (mean +/- SD 1.6 +/- 0.52 J/cm2). The mean +/- SD ESS correspondingly increased from day 2 (0 +/- 0) to day 5 (10.5 +/- 3. 7) with a decrease at day 6 (7.5 +/- 3.1) (difference between day 3 and beyond statistically significant at P < 0.05). As our study indicates a maximal erythematous reaction to the bath-PUVA up to 5 days after irradiation, the traditional MPD assessment at 3 days generates a risk of phototoxic side-effects within the phototherapy course by underestimating the phototoxic effect in some patients. These findings contribute towards a more defined understanding of the kinetics of the phototoxic reaction in bath-PUVA therapy.     

Penetration,permeation, and resorption of 8-methoxypsoralen

Summary The penetration, permeation, and resorption of radioactively labelled 8-Methoxypsoralen was investigated in human skin. Simultaneously, the effects of time and ointment carrier on the penetration kinetics were ascertained. The carriers tested were: vaseline, aqueous wool-wax alcohol ointment, aqueous hydrophilic ointment and polyethylene glycol ointment. The absolute concentrations of 8-Methoxypsoralen were estimated in the horny layer, epidermis and dermis. With the most advantageous carrier, aqueous wool-wax alcohol ointment, 4–6 × 10^–5 M and 10^–5 M were attained in the epidermis and dermis, respectively. Moreover, it was shown that the substance penetrates rapidly (10 min) into the epidermis and dermis and the high concentrations reached remained constant over a period of 16 h.Only with a formulation of aqueous wool-wax alcohols is any accumulation at all achieved in the deeper areas of the horny layer. A uniform decrease in drug concentration with increasing depth of the horny layer is found with the other 3 vehicles, whereby slight variations in concentrations pertain from carrier to carrier. 4 h after local application, 8-Methoxypsoralen can be detected in the urine. Regardless of the ointment base employed, 8-Methoxypsoralen is no longer detectable in the urine 40 h after application.In comparison to the oral therapy, the same magnitude of percutaneous resorption into the central compartment is to be derived from the data, if half the body surface is treated locally.

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Zusammenfassung Die Penetration, Permeation und Resorption von radioaktiv markiertem 8-Methoxypsoralen wurde an menschlicher Haut untersucht. Gleichzeitig wurde die Abhängigkeit der Penetration von der Applikationsdauer und der verwendeten Salbengrundlage (Vaseline, wasserhaltige Wollwachsalkoholsalbe, wasserhaltige hydrophile Salbe und Polyäthylenglykolsalbe des Deutschen Arzneibuches (DAB 7) ermittelt. Es konnten Absolutkonzentrationen von 8-Methoxypsoralen in Hornschicht, Epidermis und Corium angegeben werden. Sie waren bei Verwendung des günstigsten Vehikels (wasserhaltige Wollwachsalkoholsalbe DAB 7) in der Epidermis 4–5 10^–6 molar und im Corium 4–5 10^–7 molar. Dabei zeigte sich, daß die Substanz schnell (10 min) in die Epidermis und das Corium eindringt und die schon anfänglich hohen Konzentrationen bis 1000 min Einwirkungszeit etwa konstant bleiben.In den unteren Hornschichtlagen kommt es allein bei der wasserhaltigen Wollwachsalkoholsalbe zu einer gewissen Anreicherung der markierten Substanz, die anderen Vehikel bewirken eine unterschiedlich große, jedoch gleichmäßig zur Tiefe hin abfallende Substanzverteilung in der Hornschicht. Vier Stunden nach der lokalen Applikation ist 8-Methoxypsoralen im Urin nachweisbar, nur bei Verwendung von Polyäthylenglykol als Vehikel, dauert die Ausscheidung doppelt so lange. Unabhängig von der Salbengrundlage ist ab ca. 40 h nach der Applikation kein 8-Methoxypsoralen mehr im Harn nachweisbar. Anhand der gefundenen Ergebnisse werden Hinweise für eine mögliche Verbesserung der Therapie mit 8-Methoxypsoralen gegeben.

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This work was supported by the Deutsche Forschungsgemeinschaft.     

Modulation of cytokine production by 8-methoxypsoralen and UVA

Modulatory effects on cytokine expression of 8-MOP in conjunction with UVA have been investigated in different systems using different cell types, including keratinocytes, lymphocytes, monocytes and endothelial cells. The vast majority of data have exhibited reduced production of cytokines in 8-MOP/UVA-treated cells and skin, reflecting its simplistic cellular damage. However, 8-MOP/UVA at modest doses stimulate T lymphocytes to produce Thl cytokines such as interferon-gamma, suggesting that some activational events may occur in certain types of cells phototreated with 8-MOP. Both the inhibitory and augmentative effects of 8-MOP/UVA on cytokine production appear to participate in the mechanisms underlying the therapeutic efficacy of PUVA and extracorporeal photochemotherapy (photopheresis). In particular, photopheresis may exert beneficial effects on cutaneous T-cell lymphoma as a cytokine modifier.     

Measurement of 5-methoxypsoralen and 8-methoxypsoralen in saliva of PUVA patients as a noninvasive, clinically relevant alternative to monitoring in blood

Abstract Background: Monitoring of psoralen concentration and time-course in PUVA patients is vital for efficient PUVA therapy. Blood sampling is invasive and labour-intensive and thus unsuited for routine use and repeat measurements over the course of therapy. Objective: Psoralen pharmacokinetics in saliva were investigated and validated as a noninvasive, simple and biologically relevant alternative to measurements in blood. Methods: The time-course of psoralen concentration was measured in saliva and serum of volunteers and patients receiving PUVA or extracorporeal photopheresis therapy. The samples were analysed by high-performance liquid chromatography. Three commonly used oral psoralen preparations were tested: Psoraderm5 (5-methoxypsoralen; 5-MOP), Meladinine and Oxsoralen (both 8-methoxypsoralen; 8-MOP). Results: The pharmacokinetic parameter C_max in saliva averaged 10% (range 6–20%) of the serum values for 8-MOP, and ≤ 4% for 5-MOP. These concentrations correspond to the therapeutically relevant, non-albumin-bound fraction of psoralen in serum that is available to diffuse into the tissues. The parameter t_max in saliva and serum coincided, indicating that psoralens diffuse rapidly between the two compartments. Conclusion: Monitoring of psoralens in saliva is a valuable, noninvasive alternative to measurements in serum, suitable for routine use. A series of five or six saliva samples is sufficient to determine t_max in a patient beginning photochemotherapy. To determine C_max, three independent saliva measurements at t = t_max are recommended. Received: 19 January 1999 / Received after revision: 31 May 1999 / Accepted: 4 June 1999     

The characteristics of erythema induced by topical 5-aminolaevulinic acid photodynamic therapy

BACKGROUND: Topical photodynamic therapy (PDT) is increasingly used to treat superficial non-melanoma skin cancers. Knowledge of the characteristics of 5-aminolaevulinic acid (ALA)-induced phototoxicity will increase our understanding of PDT and may facilitate optimisation of treatment regimes. METHODS: We examined the characteristics of ALA-induced erythema in 10 healthy subjects and investigated the effect of light source and body site. RESULTS AND CONCLUSIONS: Maximal erythema occurred within 1-2 h of PDT and inter-individual variation in ALA-induced phototoxicity was seen. No detectable differences were seen in the phototoxicity on back or leg sites or between coherent and non-coherent light sources. These data provide further information to allow us to optimise topical PDT regimes.     

Comparison of the minimal phototoxic dose in topical 4,5', 8-trimethylpsoralen PUVA treatment of caucasian skin and of oral mucous membrane

The minimal phototoxic dose values for UVA radiation of psoralen-treated skin and of oral mucous membrane were studied in 16 healthy volunteers. A commercial 0.01% trioxalen ointment was used as the topical photosensitizer. In all 16 persons the radiation dose needed to induce erythema was greater for the buccal mucosa than for the skin, and the average buccal minimal phototoxic dose was 2.3-fold that of the cutaneous minimal phototoxic dose.     

Photoreactions of 3,3',4',5-tetrachlorosalicylanilide with proteins.

The photoreactions of 3,3',4',5-tetrachlorosalicylanilide anion (TCSA-) with two serum proteins were studied. TCSA- and anions of two similar compounds, N-ethyl-3,5-dichlorosalicylamide and salicylanilide, bind noncovalently to human serum albumin (HSA) without irradiation in aqueous pH 7.4 buffered solutions. TCSA- noncovalently bound to HSA yields three types of photoproducts when irradiated with ultraviolet light (lambda greater than 360 nm). A covalently bonded photoadduct between TCSA- and HSA is formed and histidines in HSA are chemically modified. In addition to these two types of photoproducts which involve HSA, two of the four TCSA- photoproducts which form when HSA is absent are also formed when the TCSA-/HSA complex is irradiated. The results presented indicate that not all proteins in the skin are capable of being the carrier protein in photoallergy of TCSA- and that cross reactivity to other halogenated salicylanilides can be explained by further photochemical reactions of TCSA- photoproducts.     

A comparative study of toxic erythema of pregnancy and herpes gestationis

We compared the clinical features, histopathology, immunopathology and immunogenetics of 30 patients with toxic erythema of pregnancy and 24 patients with herpes gestationis. Although we found some clinical and histopathological overlap we highlighted several important differences. In toxic erythema of pregnancy prominent striae were frequently present. Herpes gestationis was suggested by the occurrence of periumbilical lesions, acute exacerbations immediately after delivery, and persistence of the eruption for more than 3 weeks post-partum. In herpes gestationis, immunofluorescence studies were consistently positive, there was a high frequency of HLA-B8 and an association with autoimmune thyrotoxicosis. Toxic erythema of pregnancy did not share these immunological features. Therefore we feel that toxic erythema of pregnancy and herpes gestationis should continue to be classified as separate disorders.     

甲氧沙林对培养人表皮黑素细胞黑素合成的影响及信号转导的研究

目的：观察甲氧沙林(8-甲氧补骨脂素，8-MOP)对体外培养的人表皮黑素细胞黑素合成和酪氨酸酶活性的影响，并初步探讨8-MOP诱导表皮黑素细胞分化的信号转导途径。方法：采用4种浓度(10～500μmol／L）的8-MOP作用于体外培养的人表皮黑素细胞，观察不同浓度、不同时间8-MOP对黑素细胞的形态、增殖、酪氨酸酶活性、黑素含量的影响，并用放射免疫法测定8-MOP对细胞内环磷腺苷酸(cAMP)含量的影响。结果：100μmol／L 8-MOP作用黑素细胞120h能显著促进酪氨酸酶的活性，提高黑素细胞的黑素含量，8-MOP抑制黑素细胞增殖和提高细胞内cAMP的水平呈浓度依赖性。结论：8-MOP在体外能直接刺激表皮黑素细胞的黑素合成，上述变化可能通过cAMP依赖的蛋白激酶(PK)A信号通路发挥作用。     

Skin photosensitizing and Langerhans' cell depleting activity of topical (bath) PUVA therapy: comparison of trimethylpsoralen and 8-methoxypsoralen

The skin photosensitizing and Langerhans' cell (LC) depleting effects of a single bath PUVA exposure were studied in 22 healthy young volunteers. The photosensitizing effect of bathing for 15 min in a 0.2 mg/1 trioxsalen-water solution was about 30 times as great as a similar treatment in an equipotent methoxsalen solution. The skin erythema induced by methoxsalen bath PUVA peaked on day 2 and diminished thereafter, whereas the trioxsalen reaction showed a broad plateau on days 2-5 after the irradiation. A reduction in LC density to about 30-40% of the starting value was seen in both trioxsalen and methoxsalen bath PUVA treated skin sites on day 4 after irradiation, and low or diminishing LC counts prevailed until day 10-11. The amount of UVA needed to produce a similar degree of LC depletion was 15-30 times as great in the case of methoxsalen, compared with trioxsalen. A perceptible erythema reaction, however, was, not a prerequisite for a reduction in LC density.     

Extended extracorporeal photochemotherapy with extracorporeal administration of 8-methoxypsoralen in systemic sclerosis. An Austrian single-center study

BACKGROUND: Extracorporeal photochemotherapy (EXP) is an immunomodulating therapy that has been used in a limited number of patients with systemic sclerosis (SSC) with controversial results. The present study was performed to evaluate the efficacy and safety of extended EXP with extracorporeal application of liquid 8-methoxypsoralen (8-MOP) in the treatment of SSC. METHODS: Eleven women with progressive SSC of recent onset were treated for a period of 16-57 months. Skin changes, physical performance, extracutaneous manifestations, and quality of life were evaluated before initiation of EXP and at regular intervals thereafter. RESULTS: From the start to the last set of EXP, we observed an overall improvement and/or stabilization of skin changes and physical performance in 5 of 11 patients (45%). Extracutaneous manifestations deteriorated in 10 of 11 patients (91%) (P<0.05), and quality of life deteriorated in 9 of 11 patients (82%) from a mean score of 10 before, to 17 at the last set of EXP (P<0.05). No major side effects were noted. CONCLUSION: Extended EXP with extracorporeal administration of 8-MOP is a safe and well tolerated treatment modality. However, it provides only (minor) improvement of skin changes of a subset of SSC patients and does not beneficially influence extracutaneous manifestations and quality of life.     

Inactivation of T4 bacteriophage by near ultraviolet light in the presence of 8-methoxypsoralen.

The killing action of near ultraviolet light emitted from a fluorescent black-light lamp in the presence of 8-methoxypsoralen was investigated using bacteriophage T4D and its mutants. It was found that a far ultraviolet-sensitive mutant, T4v, was no more sensitive to the present photoreaction than the wild type. T4px and T4y were a little more sensitive than T4D. Contrary to the results of illumination with monochromatic light, exponential survival curves with pronounced shoulders were obtained. The reason for this finding is discussed on the basis of the mode of photoreaction.     

A comparative study of fabric protection against ultraviolet-induced erythema determined by spectrophotometric and human skin measurements.

Historically, a textile's ability to protect against ultraviolet radiation (UVR)-induced erythema has been based on its UVR transmission. However, due to the nonuniformity of the fabric structure of a textile and its resultant nonuniform transmission, the above prediction may not hold. The fabric protection factors (FPF) of 5 metal meshes, to simulate the weave pattern and yarn dimensions of typical fabrics, and 6 textiles with variable construction (woven and knitted), fibre type and dye were determined using a spectrophotometric assay and human skin testing. All 5 meshs and 5 of the 6 textiles allowed spectrophotometric prediction of their FPF compared with off-skin (2 mm) human testing. However, on-skin human testing FPF were generally significantly lower than both the off-skin and spectrophotometric estimates. Although evidence is presented that the nonuniform nature of a textile's structure does influence its FPF predictability, in practice, properly conducted spectrophotometric analysis may yield the most typical indication of the protectiveness of a fabric against UVR-induced erythema.