Photodynamic Therapy with 5% δ-Aminolevulinic Acid is Safe and Effective Treatment of Acne Vulgaris in Japanese Patients.

BACKGROUND AND AIMS: Photodynamic therapy with aminolevulinic acid (ALA-PDT) is effective therapy for acne vulgaris; however, relatively strong side effects limit its wide usage. We have previously demonstrated that ALA-induced protoporphyrin IX distribution with lower concentrations and shorter contact time of ALA resulted in focused damage in sebaceous glands in vivo. We have formulated a protocol for ALA-PDT using 5% ALA with 2 hours contact time. The objective of this study was to establish the effectiveness and side effect profile of the new protocol in humans. SUBJECTS AND METHODS: Eleven Japanese patients (Fitzpatrick's skin type III - IV, mean age 23.7±7.2) with facial acne received topical application of 5% ALA for 2 hours with subsequent illumination by a broadband light (600 - 1100 nm, 15J/cm(2), 60 mW/cm(2)). Subjects were evaluated prior to the procedure, 1 month, and 3 months after the treatment by a blinded dermatologist using the global acne grading system (GAGS). Side effects were monitored through the treatment period. RESULTS: The mean GAGS score decreased from 22.1±3.8 at baseline to 19.4 at 1 month, and to 16.3 at 3 months after PDT (P<0.05). Ten of eleven patients experienced local side effects, such as erythema, which were of minimal to mild severity. However, most side effects were of minimal to mild severity, and all of them resolved within several days without post inflammatory hyper pigmentation. CONCLUSION: Our protocol was effective for acne in Japanese and did not exhibit severe side effects.

Targeting of sebaceous glands by δ-aminolevulinic acid-based photodynamic therapy: An in vivo study.

BACKGROUND AND OBJECTIVES: Wide application of ALA-PDT for acne is limited due to relative strong side effects, such as pain and erythema. The objective of this study was to establish a protocol for ALA-PDT that would provide specific destruction of sebaceous glands at the lowest concentrations and shortest contact times of ALA. MATERIALS AND METHODS: The rhino (hr(rh) hr(rh) ) murine model, an experimental acne model, was used in this study. A freshly prepared hydrophilic ALA hydrochloride ointment (2.5%, 5%, and 20%) was applied to the backs of 16-week-old male rhino mice. The fluorescence intensity (FI) of ALA-induced protoporphyrin IX (ALA-PpIX) on the skin was measured by spectrofluorometry. Skin samples were taken at 1, 2, and 4 hours after ALA application to determine the tissue distribution of ALA-PpIX by fluorescence microscopy. Light irradiation was also performed with a broadband light source (600-1100 nm, 15 J/cm² , 60 mW/cm²) with subsequent histological examination 1 day after treatment. RESULTS: Prominent increases of ALA-PpIX were observed 1 hour after application of 5% and 20% ALA, while no increase was observed with 2.5% ALA until 2 hours. A direct correlation was found between ALA concentration and ALA-PpIX FI. While no fluorescence was detected 1 hour after application of 2.5% or 5% ALA, 20% ALA produced a strong fluorescence in the epidermis, utricle walls, and sebaceous glands. Histological evaluation showed no damage to skin treated with 2.5% ALA-PDT incubated for 1 hour. Damage was still focused within the sebaceous glands with longer incubation times. Increased ALA concentrations resulted in more prominent damage to the epidermis and sebaceous glands, with deeper damage to the dermis when longer incubation times were used. CONCLUSION: Focused damage of sebaceous glands can be achieved with ALA-PDT when low concentrations of ALA (2.5-5.0%) and short incubation times (to 2 hours) were used.

Comparative split-face study of 5-aminolevulinic acid photodynamic therapy with intense pulsed light for photorejuvenation of Asian skin.

Photodynamic therapy (PDT) with 5-aminolevulinic acid (ALA) (ALA-PDT) using intense pulsed light (IPL) as a light source (IPL-ALA-PDT) has been used for photorejuvenation, but it is unclear if this protocol can be applied to darker skin types. We performed this study to assess our IPL-ALA-PDT protocol for photorejuvenation in Asian skin. To determine an appropriate dose, ALA ointment (0-20%) was applied to the upper arm of five healthy volunteers and the fluorescence intensity (FI) was measured using a spectrofluorometer. Non-linear regression analysis of FI 2 h after ALA application with global fitting gave a typical sigmoid dose-response curve with R² = 0.9705 and saturation after 5% ALA. The entire faces of 16 Japanese women with photodamage were then treated with IPL (500-670 and 870-1400 nm, 23-30 J/cm²) 2 h after application of 5% ALA to one side of the face. Three treatments were delivered at 4-week intervals with follow-up visits. Comparative analysis of photorejuvenation showed noticeable improvements on both sides of the face, although the reduction in the photoaging score from baseline did not differ significantly between the two sides in all subjects. Despite this finding, 75% of the patients felt that the IPL-ALA-PDT-treated side of the face showed greater improvement than the IPL-treated side. However, all IPL-ALA-PDT-treated sides showed adverse effects such as erythema and pain. Therefore, we conclude that the IPL-ALA-PDT protocol requires optimization for photorejuvenation in Asians.

Prospects for the use of differentiation-modulating agents as adjuvant of photodynamic therapy for proliferative dermatoses.

Current interest in photodynamic therapy (PDT) in dermatology stems from its recognized success in dermatological oncology, straightforward approach, easy accessibility and low cost. PDT is a photochemistry-based modality in which a light-activated photosensitizer (PS) destroys tissue through oxygen-dependent and -independent mechanisms. Although PDT has been used in dermatology for several decades, its application has still not extended significantly into the routine management of neoplastic and proliferative dermatoses because of continuing issues with the selectivity of the PS for affected tissues. This review analyzes prospects for optimization of PDT for the management of dermatoses with defects in keratinocyte proliferation/differentiation, and discusses the use of differentiating agents that redirect metabolic utilization within cells and lead to high levels of PS accumulation.

Photodynamic therapy for cutaneous leishmaniasis: the effectiveness of topical phenothiaziniums in parasite eradication and Th1 immune response stimulation.

Photodynamic therapy (PDT) is emerging as a therapeutic modality in the clinical management of cutaneous leishmaniasis (CL). The efficacy of PDT against CL has been demonstrated previously with aminolevulinic acid, although the prolonged terms of therapy were less than ideal, and the search for new photosensitizers (PS) is ongoing. However, phenothiaziniums have demonstrated high parasiticidal effects in vitro. The subject of our investigation is the in vivo activity of two PS, 5-ethylamino-9-diethylaminobenzo[a]phenoselenazinium chloride (EtNBSe) and (3,7-Bis(N,N-dibutylamino) phenothiazinium bromide (PPA904). The results of our comparative analysis of the efficacy of these two phenothiazinium analogues demonstrated a high antiparasitic activity of EtNBSe in vitro, and the higher efficacy of PPA904 in a mouse model of CL. The kinetics of photodestruction are different in parasite and mammalian cells, and with both dyes, the macrophages are more susceptible to photodynamic effects than L. major parasites. As the number of parasites in the lesions undergoes a biphasic change, temporarily increasing on days 2-4 and decreasing on days 5-7, more than one treatment is required within an interval of 5 to 7 days. We have also shown that PPA904-PDT can provide an immunomodulating, dose-dependent efflux on IL-12p70 production. This mechanism could be responsible for promoting a more rapid healing in PPA904-PDT treated mice. Our initial data indicate that phenothiaziniums exhibit a high parasiticidal effect in vivo against CL; this finding may be of use in establishing curative PDT regimens for future clinical trials.

Parasiticidal effect of delta-aminolevulinic acid-based photodynamic therapy for cutaneous leishmaniasis is indirect and mediated through the killing of the host cells.

Several clinical reports have shown promising, but not optimal, results from photodynamic therapy with delta-aminolevulinic acid-derived protoporphyrin IX, termed ALA-PDT, as a treatment for cutaneous leishmaniasis (CL). Therefore, understanding the basis of the phototoxic response of Leishmania parasites to ALA-PDT may be critical for optimization. We report here both in vitro and in vivo mechanistic studies of ALA-PDT against CL. Following in vitro co-incubation of Leishmania major with 0.1 microM ALA, the PpIX concentration remained at the basal level, whereas after co-incubation with 0.1 microM exogenous PpIX, the PpIX level was 100-fold higher. No differences in ALA-derived PpIX levels were detected between Leishmania-infected and non-infected J774.2 cells, and PDT did not demonstrate any parasiticidal effects on amastigotes. In contrast, in vivo topical ALA-PDT, performed on a murine CL model, resulted in significant reductions of the parasite loads and vigorous tissue destruction. After ALA-PDT, a dramatically decreased percentage of macrophages and increased levels of interleukin-6 were observed in the infected skin. The clinical outcome observed with ALA-PDT is likely the result of unspecific tissue destruction accompanied by depopulation of macrophages rather than direct killing of parasites.

The role of photosensitizer molecular charge and structure on the efficacy of photodynamic therapy against Leishmania parasites.

Photodynamic therapy (PDT) is emerging as a potential therapeutic modality in the clinical management of cutaneous leishmaniasis (CL). In order to establish a rationale for effective PDT of CL, we investigated the impact of the molecular charge and structure of photosensitizers on the parasitic phototoxic response. Two photosensitizers from the benzophenoxazine family that bear an overall cationic charge and two anionic porphyrinoid molecules were evaluated. The photodynamic activity of the photosensitizers decreases in the following order: EtNBSe > EtNBS > BpD > PpIX. The studies suggest that compared to hydrophobic anionic photosensitizers, the hydrophilic cationic benzophenoxazine analogs provide high effectiveness of PDT possibly due to (1) their strong attraction to the negatively charged parasitic membrane, (2) their hydrophilicity, (3) their high singlet oxygen quantum yield, and (4) their efficacy in targeting intracellular organelles.

Targeting of sebocytes by aminolevulinic acid-dependent photosensitization.

Photodynamic therapy using 5-aminolevulinic acid-induced protoporphyrin IX has been developed as a very useful therapeutic modality. Recently, several authors have reported on the efficacy of this procedure for acne. This approach is based on the fact that 5-aminolevulinic acid-induced protoporphyrin IX has strong selectivity for sebaceous glands. We used the immortalized human sebaceous gland cell line SZ95 to investigate cellular mechanisms of photodynamic therapy using 5-aminolevulinic acid-induced protoporphyrin IX. Quantification of induced protoporphyrin IX production showed dependence on the applied 5-aminolevulinic acid dose. When SZ95 sebocytes were differentiated by arachidonic acid treatment, there was no difference between them and the control cells with respect to both the amount of 5-aminolevulinic acid-induced protoporphyrin IX and the phototoxic effects. We altered protoporphyrin IX formation rates by growing cells scattered as single cells in the culture dishes. Single cells produced significantly lower protoporphyrin IX levels than those grown with intercellular contacts. Intracellular localization of protoporphyrin IX was imaged using confocal laser scanning microscopy. The differentiation-specific lipid droplets were virtually excluded from protoporphyrin IX fluorescence. In addition to weak mitochondrial and strong membrane fluorescence, distinctive spots with strong fluorescence were observed. These did not colocalize with fluorescent probes for mitochondria, lysosomes or the Golgi apparati.

Photoinactivation of Mycobacteria in vitro and in a new murine model of localized Mycobacterium bovis BCG-induced granulomatous infection.

Treatment of tuberculosis is currently hindered by prolonged antibiotic regimens and the emergence of significant drug resistance. Alternatives and adjuncts to standard antimycobacterial agents are needed. We propose that a direct attack utilizing photosensitizers and light-based treatments may be effective in curtailing Mycobacterium tuberculosis in discrete anatomical sites in the most infectious phase of pulmonary tuberculosis. To demonstrate experimental proof of principle, we have applied established photodynamic therapy (PDT) technology to in vitro cultures and an in vivo mouse model using Mycobacterium bovis BCG. We report here in vitro and in vivo PDT efficacy studies and the use of a three-dimensional collagen gel as a delivery vehicle for BCG, subcutaneously inserted, to induce specifically localized granuloma-like lesions in mice. When a benzoporphyrin derivative was utilized as the photosensitive agent, exposure to light killed extracellular and intracellular BCG in significant numbers. Collagen scaffolds containing BCG inserted in situ in BALB/c mice for 3 months mimicked granulomatous lesions and demonstrated a marked cellular infiltration upon histological examination, with evidence of caseating necrosis and fibrous capsule formation. When 10(5) BCG were present in the in vivo-induced granulomas, a significant reduction in viable mycobacterial cells was demonstrated in PDT-treated granulomas compared to those of controls. We conclude that PDT has potential in the treatment of localized mycobacterial infections, such as pulmonary granulomas and cavities.

Novel stereoconvergent transformation of 1,2a-disubstituted 1,2,2a,8b-tetrahydro-3H-benzo[b]cyclobuta[d]pyran-3-ones to 1,3-disubstituted 1,2,4a,9b-tetrahydrodibenzofuran-4-ols and its application to the second-generation synthesis of (+/-)-linderol A.

1,2a-Disubstituted 1,2,2a,8b-tetrahydro-3H-benzo[b]cyclobuta[d]pyran-3-ones bearing an electron-withdrawing group at the 2a-position were treated with two equivalents of dimethylsulfoxonium methylide to give r-1,t-4a,t-9b-1,3-disubstituted 1,2,4a,9b-tetrahydrodibenzofuran-4-ols stereoconvergently regardless of the stereochemistry of the 1-position on the benzocyclobutapyran ring. This methodology was applied to the second-generation synthesis of (+/-)-linderol A, a melanin biosynthesis inhibitory natural product.

Burns caused by dilute hydrofluoric acid in the bleach.

Two male cleaning workers aged 62 (patient 1) and 28-(patient 2) presented with red, swollen, aching hands and fingers. At the first interview, the fingers of the right hand of both patients were swollen from the proximal interphalangeal joint to the tip of the finger. The fingers were red and intensely painful. The bleach that both patients used contained 9.5% hydrofluoric acid, and therefore we diagnosed their symptoms as those of burns caused by this agent. We subcutaneously injected 8.5% calcium gluconate into the affected fingers and dressed them with gauze soaked in cooled 0.025% benzethonium chloride. The patients did not use gloves at work, neither of them knew that the cleaning fluid contained dilute hydrofluoric acid, and they were unaware of the danger of this agent. They had not received proper education about the care and handling of poisons and deleterious substances such as hydrofluoric acid. The doctors who had examined the patients in the emergency ward overlooked the possibility of hydrofluoric acid burns, although they suspected chemical burns and confirmed the trade name of the cleaning agent. In addition, although the patients presented with intense pain, no white areas of coagulation and blistering, or surrounding erythemas, which are characteristic symptoms of such burns, were evident. The component of the cleaning agent was described on the container label in very small print. Had the words "poison" and "hydrogen fluoride" been printed in large characters, the examining doctors in the emergency ward would probably not have overlooked the presence of hydrofluoric acid, and the patients would perhaps have been more careful when using it. Hydrofluoric acid can be easily obtained by anyone through the Internet, although general consumers could not obtain industrial quantities. Therefore, the number of burn patients who are not familiar with hydrofluoric acid may increase in the future.