Adjuvants that Empower the Action of Photodynamic Therapy.

Compounds have been devised whose supportive actions make them important adjuvants in the priming of photosensitization to selectively target cancer cells. Here, we highlight the paper by Maytin and Hasan in this issue of Photochemistry & Photobiology, which describes adjuvants methotrexate, 5-fluorouracil, vitamin D and its analogs leading to improved photodynamic therapy outcome. These small molecule adjuvants act by different mechanisms to enhance the cytotoxicity in tumor cells and the therapeutic effect in cancers. These findings add to the list of strategies for enhancement of photodynamic therapy.

Biologics for the primary care physician: Review and treatment of psoriasis.

Psoriasis is a chronic inflammatory disease that affects approximately 7.5 million people in the United States. The disease results in significant suffering, morbidity, and economic impact. Psoriasis is a multifaceted disease with a strong genetic component. Genetic data has revealed the presence of particular risk alleles in patients with psoriasis. Triggers of the disease have been elucidated and include factors such as trauma, obesity, infection, stress, and medications. At its core, psoriasis is a result of a dysfunctional immune response with T-cells at the center of immunogenesis. Clinically, psoriasis is characterized by discrete, erythematous scaly plaques. These lesions are often found on extensor surfaces, especially the elbows and knees. Although extensor surfaces are the prototypical destination of lesions, psoriasis may affect any area of the skin including the scalp, intertriginous areas, nails, palms, and soles. Location of lesions are important in assessing the impact on quality of life for patients. Diagnosis of psoriasis can typically be made clinically based on characteristic history and physical examination findings. In rare cases, biopsy may be needed to rule out other papulosquamous disease. Histologic findings of psoriasis can be non-specific and include marked epidermal hyperplasia, dilated vessels within the dermal papilla, and elongated rete ridges. Importantly, psoriasis is a systemic disease and organ systems outside of the skin must be considered. Co-morbidities of psoriasis include psoriatic arthritis, type 2 diabetes mellitus, cardiovascular disease, psychiatric disease, inflammatory bowel disease, neoplasms, and ocular disease. Management of psoriasis depends on the severity of the disease. In mild to moderate cases, topical medications are the cornerstone of treatment. Topical corticosteroids are the most commonly used and have limited systemic effects due to the localized application of medication. In moderate to severe cases of psoriasis, topical medications are ineffective and not feasible. Phototherapy and non-biologic systemic medications have been useful treatments; however, phototherapy is time consuming and non-biologic systemics have only modest response rates. In the last decade, biologic medications have become an important component of care for treating moderate to severe psoriasis. These medications target various cytokines responsible for psoriasis manifestations such as tumor necrosis factor (TNF-α), interleukin-12, interleukin-23, and interleukin-17. In the past 15 years, numerous biologic medications have been granted FDA approval, with the majority approved in the past several years. Some of the commonly used biologics include etanercept, adalimumab, infliximab, ixekizumab, secukinumab, brodalumab, guselkumab, ustekinumab, and tildrakizumab. Given the wealth of new biologics, current treatment guidelines have rapidly become outdated. This review provides summarized information of landmark trials that led to the approval of these medications.

Prevalence of Phototherapy in the Age of Biologics.

BACKGROUND: The prevalence of narrow-band ultraviolet B (NB-UVB) use in Europe for moderate and severe psoriasis is unknown, because national registries for psoriasis do not monitor this treatment. OBJECTIVES: To quantify the use of phototherapy, biologics or conventional treatments in psoriasis, in a setting where European Medicines Agency (EMA) eligibility criteria for biologics were strictly applied, and phototherapy was included among first-line treatments. METHODS: We followed a cohort of 1,090 patients who were referred to the only centre entitled to prescribe biologics and phototherapy during a 5-year period. RESULTS: The cumulative number of treatment cycles was: 1,047 with NB-UVB phototherapy, 650 with systemic treatments and 239 with biologics; 754 patients received at least 1 course of NB-UVB phototherapy, 422 at least 1 course with a systemic treatment and 137 with a biologic; 595 patients were treated only with phototherapy. CONCLUSIONS: Regular use of NB-UVB as first-line treatment for moderate and severe psoriasis and adherence to the EMA eligibility criteria for biologics led to a relatively restricted use of biologics.

Pulsed CO2 Laser Ablation of Superficial Basal Cell of Limbs and Trunk: A Comparative Randomized Clinical Trial With Cryotherapy and Surgical Ablation.

BACKGROUND: Pulsed CO2 laser is a treatment of superficial basal cell carcinoma (sBCC) although robust clinical evidence has not been reported so far. OBJECTIVE: The authors investigated efficacy, safety, time to wound healing, cosmetic outcome, patient satisfaction, and cost-effectiveness ratio of pulsed CO2 laser in comparison to cryotherapy and surgery. MATERIALS AND METHODS: BCCs of the trunk and extremities were randomized to one of the treatments. After 90 days, efficacy and cosmetic outcome were assessed. Patients recorded the time to complete healing of the wound and scored their overall satisfaction. RESULTS: Two hundred forty patients were randomized. After 3 months, complete remission (CR) rate with pulsed CO2 laser was 78.8%. This was significantly lower than surgery, whereas the CR rate with cryotherapy was not significantly different. Cosmetic result was better with surgery. High satisfaction was reported by 65.0% of patients treated with CO2 ablation. Time of wound healing was significantly shorter with CO2 laser. CONCLUSION: In comparison to cryotherapy, pulsed CO2 laser showed no statistically significant difference in efficacy, cosmetic outcome, and patient satisfaction. Time to healing was shorter; the cost and cost-effectiveness ratio were similar. Surgery had the greatest efficacy rate. The main limitation of this study was the short duration of follow-up (3 months).

Bilateral areolar leiomyomas in a patient undergoing BRAF inhibition therapy for melanoma.

BRAF inhibition therapy, used to treat melanomas with BRAF mutations, is associated with both neoplastic and non-neoplastic cutaneous side effects including squamous cell carcinomas, warty dyskeratomas, verrucous keratoses, photosensitivity and widespread eruptions that present histopathologically as acantholytic dyskeratosis. We report a case of a patient undergoing BRAF inhibition therapy for disseminated melanoma with a V600E mutation who developed bilateral areolar leiomyomas, one of which was biopsied and the other of which resolved after discontinuation of vemurafenib therapy. To our knowledge, this is the first reported case of a mesenchymal neoplasm developing in association with BRAF inhibition therapy.

Vitamin D Combined with Aminolevulinate (ALA)-Mediated Photodynamic Therapy (PDT) for Human Psoriasis: A Proof-of-Principle Study.

We previously showed that select agents (methotrexate or Vitamin D), when administered as a preconditioning regimen, are capable of promoting cellular differentiation of epithelial cancer cells while simultaneously enhancing the efficacy of 5-aminolevulinic acid (ALA)-mediated photodynamic therapy (PDT). In solid tumors, pretreatment with Vitamin D simultaneously promotes cellular differentiation and leads to selective accumulation of target porphyrins (mainly protoporphyrin IX, PpIX) within diseased tissue. However, questions of whether or not the effects upon cellular differentiation are inexorably linked to PpIX accumulation, and whether these effects might occur in hyperproliferative noncancerous tissues, have remained unanswered. In this paper, we reasoned that psoriasis, a human skin disease in which abnormal cellular proliferation and differentiation plays a major role, could serve as a useful model to test the effects of pro-differentiating agents upon PpIX levels in a non-neoplastic setting. In particular, Vitamin D, a treatment for psoriasis that restores (increases) differentiation, might increase PpIX levels in psoriatic lesions and facilitate their responsiveness to ALA-PDT. This concept was tested in a pilot study of 7 patients with bilaterally-matched psoriatic plaques. A regimen in which calcipotriol 0.005% ointment was applied for 3 days prior to ALA-PDT with blue light, led to preferential increases in PpIX (~130%), and reductions in thickness, redness, scaling, and itching in the pretreated plaques. The results suggest that a larger clinical trial is warranted to confirm a role for combination treatments with Vitamin D and ALA-PDT for psoriasis.

Biomodulatory approaches to photodynamic therapy for solid tumors.

Photodynamic Therapy (PDT) uses a photosensitizing drug in combination with visible light to kill cancer cells. PDT has an advantage over surgery or ionizing radiation because PDT can eliminate tumors without causing fibrosis or scarring. Disadvantages include the dual need for drug and light, and a generally lower efficacy for PDT vs. surgery. This minireview describes basic principles of PDT, photosensitizers available, and aspects of tumor biology that may provide further opportunities for treatment optimization. An emerging biomodulatory approach, using methotrexate or Vitamin D in combination with aminolevulinate-based PDT, is described. Finally, current clinical uses of PDT for solid malignancies are reviewed.

Narrowband UVB phototherapy as a novel treatment for Netherton syndrome.

Netherton syndrome (NS) is a rare congenital ichthyosis that is characterized by impaired skin barrier function. Topical medications are cautiously used in NS since toxicity from systemic absorption is a major concern. Narrowband ultraviolet B phototherapy is an alternative therapeutic option that demonstrated its beneficial and practical use in a patient with NS.

Poorly growing hair in a child.

A healthy 5-year-old Caucasian girl presented to the pediatric dermatology clinic for poor hair growth. The patient's father described slow hair growth and finely textured hair since birth. The patient had her first haircut 1 month before presentation. The family denied bald areas on the scalp, significant hair shedding, and sores on the scalp. The child did not scratch at her scalp or pull her hair. She are a normal diet and had normal growth and development otherwise. She was not taking any medications or over-the-counter supplements. There was no family history of hair disorders. On physical examination, the patient had short, fine, reddish blond hair that was of different lengths (Figure 1). There were no papules, pustules, scale, or crust on her scalp. Lymphadenopathy was absent. She had normal eyelashes and eyebrows. There were no lesions on her oral mucosa. No tooth or nail abnormalities were present. The rest of the physical examination was normal. The hair pull test resulted in 4 easily pulled hairs (3 anagen and 1 telogen). A hair mount was performed (Figure 2). The hair mount analysis revealed anagen hairs with distorted bulbs and ruffled cuticles extending a short distance distally from the bulb, consistent with loose anagen hair (Figure 2). All of the anagen hairs on the pull test demonstrated the above findings. Based on the patient's clinical presentation and the findings seen on light microscopy of the hair mount preparation, the patient was diagnosed with loose anagen syndrome.

A critical reappraisal of off-label indications for topical photodynamic therapy with aminolevulinic acid and methylaminolevulinate.

Photodynamic therapy (PDT) with methylaminolevulinate (MAL) has demonstrated high efficacy, minimal side effects, and improved cosmetic outcome in the treatment of its EU-approved clinical indications, i.e. actinic keratoses (AKs), basal cell carcinoma, and Bowen's disease. In addition, PDT with MAL or 5-aminolevulinic acid (ALA) is approved in the US for the treatment of AK. However, besides anti-tumoral activity, PDT has also demonstrated various anti-inflammatory and anti-infectious effects as well as those on the metabolism of connective tissue, keratinization of normal keratinocytes and maturation processes of sebaceous glands and hair follicles. These findings have expanded the spectrum of possible applications of PDT that now encompasses infectious (viral, bacterial, fungal, protozoal) disorders, epidermal and dermal inflammatory diseases, tumors of lymphocytes, adnexal diseases and premature skin aging due to sun exposure. However, the findings obtained so far must be considered in most off-label indications only preliminary and more detailed studies are required to clarify the role of PDT beyond the treatment of non-melanoma skin cancer. In addition, possible advantages over standard treatments remain to be clarified.

Molecular mechanisms of photodynamic therapy.

Despite its more than 100-year history in experimental and clinical use, photodynamic therapy (PDT) is only starting to be appreciated for its full potential. PDT combines a photosensitizer (PS) and light in the presence of oxygen to treat cancer and other disorders. This manuscript reviews molecular mechanisms that have been evaluated over the past years for the effects of PDT at the cellular level as well as in therapeutic settings in vivo. The availability of multiple PS with different structures and functional properties makes PDT an extremely versatile and, conversely, a challenging approach to cancer therapy. The advancing understanding of molecular pathways helps to design improved regimens. As most cancers are being treated with combination therapies, PDT is being integrated into rationally designed combined regimens that exploit molecular responses to PDT for improved efficacy.

Tumor cell lines resistant to ALA-mediated photodynamic therapy and possible tools to target surviving cells.

We isolated and characterized cell lines resistant to aminolevulinic acid (ALA)-mediated photodynamic therapy (PDT) derived from a murine adenocarcinoma and studied cross resistance with other injuries. The most resistant clones were numbers 4 and 8, which exhibited 6.7- and 4.2-fold increase in resistance respectively. Several characteristics were altered in these clones. A 2-fold increase in cell volume, higher cell spreading, and a more fibroblastic, dendritic pattern, were the morphology features that led us to think they could have different adhesive, invasive or metastatic phenotypes. The amount of porphyrins synthesized per cell in the resistant clones was similar to the parental line but, when it was expressed per mg protein, there was a 2-fold decrease, with a higher proportion of hydrophilic porphyrins. These cells were not cross-resistant to photosensitization with Benzoporphyrin derivative and Merocyanine 540, but exhibited a slight resistance to exogenous protoporphyrin IX treatment. Both clones displayed higher protein content and increased number of mitochondria, together with a higher oxygen consumption. The distinctive features found in the resistant lines led as to think how to exploit the changes induced by PDT treatment to target surviving cells. Those hypoxic cells can be also a preferential target of bioreductive drugs and hypoxia-directed gene therapy, and would be sensitive to treatment with other photosensitizers.

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.

Plasma matrix metalloproteinase 9 as biomarker of prostate cancer progression in Dunning (Copenhagen) rats.

BACKGROUND: Matrix metalloproteinases (MMPs) play an important role in invasion and metastatic spread of cancer cells. The objective of this study was to assess MMPs in plasma of Dunning tumor rats as indicators of the progression of prostate cancer and follow-up parameters after treatment. METHODS: Prostate cancer was induced in male Copenhagen rats by either subcutaneous or orthotopic implantation of R3327-MatLyLu cells. During the development of the tumor, plasma MMP-2, and MMP-9 were measured by gelatin-substrate zymography and Western blot technique with densitometry in untreated animals, rats treated with laser-induced hyperthermia, or with the new synthetic MMP inhibitor RO 28-2653. RESULTS: Normal prostatic tissue of the Copenhagen rats predominantly expressed proMMP-2 but not proMMP-9 whereas MMP-9 was only found in cancerous tissue. Elevated plasma MMP-9 values were demonstrated in rats with subcutaneous or orthotopic tumors. Animals with tumors and treated with the MMP inhibitor RO 28-2653 had both a lower tumor volume and a lower plasma MMP-9 concentration compared with controls. CONCLUSIONS: The determination of plasma MMP-9 in Dunning tumor rats is a useful tool to monitor the progression of prostate cancer and to assess the efficacy of drugs like MMP inhibitors or other treatment protocols.

The new synthetic matrix metalloproteinase inhibitor (Roche 28-2653) reduces tumor growth and prolongs survival in a prostate cancer standard rat model.

The therapeutic efficacy of synthetic inhibitors of matrix-metalloproteinases (MMPs) in various cancers has been demonstrated. A novel inhibitor, Ro 28-2653, with high selectivity for MMP2, MMP9 and membrane type 1-MMP was evaluated in an orthotopic prostate cancer rat model. Efficacy was determined by recording tumor growth and survival endpoints. Prostate cancer was induced by inoculating R3327 Dunning tumor cells (MatLyLu) into the ventral lobe of the prostates of 148 Copenhagen rats. Daily oral treatment with Ro 28-2653 (10-300 mg/kg per day) was started on day 1 or on day 6 after tumor cell injection. Animals were sacrificed on day 20 for determination of tumor weights. For survival studies, rats received daily oral Ro 28-2653 (100 mg/kg per day) or vehicle for up to 30 days. Tumor induction was successful in 100% of the animals. Ro 28-2653 reproducibly reduced the tumor weights by up to 90% in a dose-dependent manner. In addition, an inhibitory effect in rats with established tumors (treatment start at day 6) was shown. A significantly prolonged survival of Ro 28-2653-treated rats was also demonstrated. Selective inhibition of MMP activity is a novel therapeutic approach, which bears promise for studies in patients with prostate cancer.

Detection of female lower genital tract dysplasia using orally administered 5-aminolevulinic acid induced protoporphyrin IX: a preliminary study.

OBJECTIVE: Previous studies have suggested that 5-aminolevulinic acid (ALA) may be used topically on the cervix to allow optical detection of cervical dysplasia, based on the fluorescence of protoporphyrin IX (PpIX) synthesized in situ from ALA. However, the uniformity of distribution of topically applied PpIX and the sensitivity and specificity of detection are not optimal. The current study was undertaken to demonstrate the feasibility of administering ALA by mouth (po) with the hypothesis that systemic administration might provide a more reliable diagnostic tool. METHODS: Oral ALA was administered to 14 patients with abnormal Pap smears in a dose- and time-intensity design. Institutional review board approval was obtained. A starting dose of 10 mg/kg of po ALA was administered and colposcopy was performed in 3 patients at 1 h, 3 patients at 2 h, 6 patients at 3 h, and 2 patients at 4 h. The study was written with the intent to increase the dose in 10 mg/kg increments if fluorescence was not detected; however, fluorescence was detected at the first dose level. Liver function tests were checked pre and post ALA and follow-up telephone calls were made regarding possible side effects. Both white and blue light colposcopy examinations were performed by two blinded clinicians and biopsies of all abnormal areas were performed. RESULTS: All patients tolerated po ALA well, with no systemic side effects. At the 10 mg/kg dose there was no reported nausea or photosensitivity. Optimal fluorescence was achieved at the 3-h time point, with quenching noted at the 4-h time point. Excellent absorption was documented with fluorescence of the lip demonstrated with Wood's lamp. In some cases fluorescence correlated with dysplasia on biopsy. CONCLUSION: 5-ALA given via the po route and at the dose and time period studied is well tolerated and affords fluorescence of the cervix. Future study is needed to demonstrate the successful identification of dysplastic lesions, with the ultimate goal of treating dysplasia of the lower genital tract with 5-ALA and light therapy.

Degree of substitution of chlorin e6 on charged poly-L-lysine chains affects their cellular uptake, localization and phototoxicity towards macrophages and cancer cells.

Macromolecular photosensitizer conjugates are under investigation as improved delivery vehicles for dyes used in photodynamic therapy. We have previously described the use of conjugates between photosensitizers such as chlorine6 (ce6) and poly-L-lysine (pL) chains which are versatile molecular species because the size of the chain can be varied, and the overall charge can be altered from cationic through neutral to anionic. We now report on a series of pL-ce6 conjugates in their cationic (native), neutral (acetylated) and anionic (succinylated) forms, where the number of ce6 molecules attached to each chain was varied (pL: ce6 ratios, 1:4, 1:8, 1:12, and 1:16). The fluorescence emissions were measured in both saline and a disaggregating solvent. We studied two cell lines (an epithelial ovarian cancer, OVCAR-5 and a mouse macrophage, J774) and measured cellular uptake, subcellular localization (by confocal fluorescence microscopy) and phototoxicity. The cellular uptake of the conjugates with four substitution ratios all delivered at 2 µM ce6 equivalent concentration showed a maximum at 12 ce6 per chain for both cationic and anionic conjugates, but the uptake of the neutral conjugate was proportional to the substitution ratio. The macrophages took up several times more ce6 than the ovarian cancer cells. Confocal fluorescence micrographs showed more cellular fluorescence with the lower substitution ratios, and more lysosomal localization with the cationic conjugates. The phototoxicity was much higher for the neutral conjugates. For the cationic and neutral conjugates the 12 ce6 per chain was the most effective at killing cells, while for the anionic conjugate it was the 16 ce6 per chain. The anionic conjugate was better at killing OVCAR-5 cells, while the cationic was better for J774 cells, and the neutral was approximately the same. These data will help to optimize the parameters to be used in preparing polymeric-photosensitizer conjugates for photodynamic therapy.