Actinic keratosis and surrounding skin exhibit changes in corneocyte surface topography and decreased levels of filaggrin degradation products.

Actinic keratosis (AK) is a frequent premalignant skin lesion mainly caused by chronic sun exposure. AK lesions are often surrounded by invisible, subclinical alterations, called field of cancerization (FoC). Definition of FoC is of importance for therapy management; however, the criteria and non-invasive tools to characterize FoC are lacking. Atomic force microscopy (AFM) proved to be a suitable tool for detection of changes in the corneocyte surface topography in inflammatory skin diseases, which share similar clinical features with AK such as hyper- and parakeratosis. Therefore, in this study we applied AFM to investigate AK and surrounding skin obtained by non-invasive collection of the stratum corneum (SC) with adhesive tapes. Furthermore, we determined degradation products of structural protein filaggrin (natural moisturizing factor, NMF), which previously showed association with the changes in corneocyte surface topography. Ten patients with multiple AK on the face were recruited from the outpatient clinic. SC samples were collected from the AK lesion, skin sites adjacent to the AK, 5 cm from the AK and retroauricular area. Corneocyte surface topography was determined by AFM, and NMF by liquid chromatography. The AK lesion showed alterations of the corneocyte surface topography characterized by an increased number of nanosize protrusions, which gradually decreased with the distance from the lesion. NMF levels show an inverse pattern. Atomic force microscopy showed to be a suitable tool to detect changes in the corneocyte surface topography on the AK lesion and surrounding skin in a non-invasive manner.

Low irradiance compared with conventional photodynamic therapy in the treatment of actinic keratoses.

BACKGROUND: Photodynamic therapy (PDT) is an effective therapy treating photodamaged areas with multiple actinic keratoses (AK). Still pain during therapy is one of the most challenging obstacles for patients. This retrospective study compares pain and efficacy intra-individual in patients using conventional PDT (c-PDT) compared to a low irradiance PDT protocol (li-PDT) with a reduced irradiance to 25% of c-PDT. METHODS: Thirty-one patients were enrolled into this retrospective analysis treated with li-PDT and c-PDT on comparable fields of actinic damage on the forehead or the cheek. Pain was scored by the patients using a VAS. Moreover, number and time to therapy interruptions were documented. For effectiveness number and grade of AK were counted before and 4 weeks after PDT. RESULTS: Maintaining a total light dose of 37 J/cm2 , a decrease in irradiation in li-PDT patients resulted in significant less pain (VAS score 2.8 vs 7.6) and fewer therapy interruptions compared to treatment with c- PDT (P < 0.0005). No significant difference in treatment outcome was found (P = 0.068). CONCLUSION: Our data shows that li-PDT can reduce pain with at least comparable clinical outcome compared to c-PDT. Therefore, it is an effective and well-tolerated treatment for patients with multiple AK.

A prospective, randomized, within-subject study of ALA-PDT for actinic keratoses using different irradiation regimes.

BACKGROUND: Photodynamic therapy (PDT) can be used to treat large fields of actinic keratoses (AKs) with high clearance rates. A notable downside is the amount of pain that accompany the treatment. This study aimed to optimize the illumination protocol during conventional PDT in order to reduce pain without compromising treatment effectiveness. METHODS: In this prospective, randomized study with a split-face design, patients with, symmetrically distributed AKs were included. All patients were treated using a ALA 78 mg/g gel. One side was illuminated with the Aktilite® CL-128 lamp and the other side with the RhodoLED® lamp in which the light intensity gradually increased to a maximum of 60%. Both sides received a total light dose of 37 J/cm2 . Pain during the treatment was measured using a visual analogue scale. The clinical effectiveness of the 2 treated sides was assessed after 12 weeks. RESULTS: Twenty-nine patients with 399 AKs were included. Illumination with the gradually increasing light intensity resulted in a decrease in the median visual analogue scale score by 1.1 points. Clearance rates were similar between the 2 lamps. CONCLUSION: Minimizing the light intensity during the illumination phase of PDT reduces pain, while still preserving a high clearance rate of AKs.

Spanish version of the Actinic Keratosis Quality of Life questionnaire.

BACKGROUND: Patient-reported outcome measures are very important outcomes. For specific diseases, health-related quality of life-instruments (HRQoL) are increasingly used to provide data on patients' overall perceptions of the course of a given disease. Actinic keratoses (AKs) are common keratotic lesions that occur on chronically sunlight-exposed skin. Only few studies regarding HRQoL in AKs have been made. OBJECTIVE: In order to be able to compare HRQoL among different countries and cultures, we aimed to translate and validate the Actinic Keratosis Quality of Life (AKQoL) questionnaire into Spanish and quantify the impairment caused by AKs in Spanish patients. METHODS: The AKQoL was translated. Then, 15 patients with AKs were interviewed to ensure cultural adaption before it was tested in one hundred patients with AK lesions at the Melanoma Unit of Hospital Clinic in Barcelona. RESULTS: Validation showed high interitem correlations, as well as a high correlation of each item and the total score. Internal consistency (Cronbach's coefficient alpha) was also high at 0.91 and an alpha value of 0.90 at retest. The test-retest correlation was 0.96, and the intraclass coefficient was 0.98. CONCLUSION: The presented data support the AKQoL Spanish version as a valid and reliable HRQoL questionnaire for the description of AK-related QoL and may provide a method for comparison of AK specific QoL between different cultures and countries.

Comparison of morphologic criteria for actinic keratosis and squamous cell carcinoma using in vivo multiphoton tomography.

The routine diagnostic procedure of actinic keratosis (AK) and invasive squamous cell carcinoma (SCC) is a histological examination after taking a biopsy. In the past decades, non-invasive optical methods for skin examination have been developed. Patients with clinical diagnosis of AK or SCC were examined. The morphological criteria were determined for healthy, AK and SCC skin and compared for statistically significant differences. In this study, the applicability of multiphoton tomography (MPT) as an in vivo diagnostic tool for AK and SCC was evaluated. Changes in the morphology of the keratinocytes such as broadened epidermis, large intercellular spaces, enlarged nucleus and a large variance in cell shape could easily be recognized. The cell nuclei of AK and SCC were significantly larger compared to healthy skin cells in all cell layers. The nucleus-cytoplasm ratio was also significantly higher for AK and SCC than for the healthy skin cells. It was even higher in SCC compared to spinous and basal cell layer of AK. The cell density in AK and SCC was significantly lower than in the basal and spinous cell layers of healthy skin. In SCC, the cell density was significantly lower than in AK. Concerning the intercellular spaces, significant differences were found for AK and healthy skin in spinous and basal cell layer and for SCC compared to AK and healthy skin. In this study, MPT proved to be a valuable non-invasive imaging method for in vivo detection and discrimination of AK and SCC from healthy skin.

Comparison of three light doses in the photodynamic treatment of actinic keratosis using mathematical modeling.

Photodynamic therapy (PDT) is an emerging treatment modality for various diseases, especially for cancer therapy. Although high efficacy is demonstrated for PDT using standardized protocols in nonhyperkeratotic actinic keratoses, alternative light doses expected to increase efficiency, to reduce adverse effects or to expand the use of PDT, are still being evaluated and refined. We propose a comparison of the three most common light doses in the treatment of actinic keratosis with 5-aminolevulinic acid PDT through mathematical modeling. The proposed model is based on an iterative procedure that involves determination of the local fluence rate, updating of the local optical properties, and estimation of the local damage induced by the therapy. This model was applied on a simplified skin sample model including an actinic keratosis lesion, with three different light doses (red light dose, 37 J∕cm2, 75 mW∕cm2, 500 s; blue light dose, 10 J∕cm2, 10 mW∕cm2, 1000 s; and daylight dose, 9000 s). Results analysis shows that the three studied light doses, although all efficient, lead to variable local damage. Defining reference damage enables the nonoptimal parameters for the current light doses to be refined and the treatment to be more suitable.

Dysregulation of epidermal growth factor receptor in actinic keratosis and squamous cell carcinoma.

The epidermal growth factor receptor (EGFR) is a receptor tyrosine kinase. Its correct function is required for normal skin development and homeostasis, while dysregulation of EGFR signalling results in cellular hyper-proliferation and defects in differentiation, leading to impaired wound healing, the development of psoriasis-like lesions, structural and functional defects of hair follicles and tumourigenesis. Actinic keratosis, which is also known as solar keratosis, develops in sun-exposed areas of the skin. These are often called 'premalignant lesions' and are said to represent early squamous cell carcinoma (SCC) in situ, although debate over their classification continues. Anti-EGFR therapies have been approved for the treatment of several malignancies and are undergoing trials for others [1], including advanced cutaneous squamous cell carcinoma (CSCC). However, a number of questions remain regarding the treatment of CSCC with anti-EGFR inhibitors. A lower number of CSCC tumours are EGFR positive in comparison to other types of tumours, such as head and neck SCC (HNSCC), and it has been suggested that patients should be selected on the basis of high tumour EGFR expression. However, there are reports of patients with tumours showing no EGFR-positive staining responding to anti-EGFR therapy. EGFR is an oncogenic driver in many tumours. Does it drive the transformation of actinic keratosis to a tumourigenic phenotype? Many such questions remain, and here, we discuss the role of EGFR in SCC and its functions during the different stages of skin cancer development.

Comparing cold-air analgesia, systemically administered analgesia and scalp nerve blocks for pain management during photodynamic therapy for actinic keratosis of the scalp presenting as field cancerization: a randomized controlled trial.

BACKGROUND: Photodynamic therapy (PDT) using methyl aminolaevulinate (MAL) is an effective treatment for extensive actinic keratosis (AK). However, pain is a major side-effect of this therapy. OBJECTIVES: To investigate whether scalp nerve blocks (group 1) provide adequate pain relief during MAL-PDT of the scalp and forehead in 32 men with baldness. METHODS: The patients received intravenous (IV) analgesia [piritramide 7.5 mg IV, plus oral metamizole (40 drops 30 min prior to PDT)] in combination with cold-air analgesia (group 2; IV analgesia) and cold-air analgesia alone (group 3). Maximum pain was evaluated by means of a visual analogue scale (VAS) during and up to 300 min after PDT. Pain during PDT was further analysed according to a pain perception scale. Furthermore, we measured haemodynamics and investigated stress hormone levels in blood samples at different time points. RESULTS: Maximum pain during PDT (primary end point) was significantly reduced in the treatment group receiving scalp nerve blocks (VAS 2.1 ± 1.3) compared with the treatment groups receiving IV analgesia (VAS 7.3 ± 1.1) and cold-air analgesia (VAS 8.4 ± 2.0; P < 0.05). No significant difference was found between groups 2 and 3 with regard to pain relief (P = 0.32). The increase in systolic blood pressure during the first 3 min of PDT was significantly lower for group 1 than for groups 2 and 3 (P < 0.001). No correlation between stress hormone levels and pain were found. CONCLUSIONS: Scalp nerve blocks provide an effective method for pain management during PDT for patients with extensive AK.

Swiss clinical practice guidelines on field cancerization of the skin.

Actinic keratosis (AK) affects millions of people worldwide, and its prevalence continues to increase. AK lesions are caused by chronic ultraviolet radiation exposure, and the presence of two or more AK lesions along with photodamage should raise the consideration of a diagnosis of field cancerization. Effective treatment of individual lesions as well as field cancerization is essential for good long-term outcomes. The Swiss Registry of Actinic Keratosis Treatment (REAKT) Working Group has developed clinical practice guidelines for the treatment of field cancerization in patients who present with AK. These guidelines are intended to serve as a resource for physicians as to the most appropriate treatment and management of AK and field cancerization based on current evidence and the combined practical experience of the authors. Treatment of AK and field cancerization should be driven by consideration of relevant patient, disease, and treatment factors, and appropriate treatment decisions will differ from patient to patient. Prevention measures and screening recommendations are discussed, and special considerations related to management of immunocompromised patients are provided.

Can video thermography improve differential diagnosis and therapy between basal cell carcinoma and actinic keratosis?

Various noninvasive techniques (dermoscopy, confocal microscopy, etc.) have been introduced to help the clinical diagnosis in nonmelanoma skin cancer. Among them, the high definition video thermographic technique (VTG) has recently been proposed. The aim of this study is to define the VTG patterns, respectively of actinic keratosis (AK) and basal cell carcinoma (BCC), and to compare these data with them of dermoscopy. The study included 36 patients with a total number of 135 lesions who underwent clinical, VTG, and dermoscopic examination. The VTG showed the presence of a hyperthermic pattern in all the cases of AK, while in the case of the BCC, the pattern was hypothermic. Dermoscopy also showed distinct pattern for AK and for BCC, but in 22% of them the data were not conclusive. Our study permits us to define two specific VTG patterns, BCC and AK respectively.

c-Rel downregulation affects cell cycle progression of human keratinocytes.

The c-Rel protein, a member of the NF-κB transcription factor family, exerts unique and distinctive functions in various cell types. Although c-Rel is expressed in human epidermis, its functions in keratinocytes are poorly understood. Our small interfering RNA-based approach of c-Rel silencing in HaCaT keratinocytes induced altered cell morphology toward a spindle-shaped appearance. In addition, c-Rel downregulation resulted in increased apoptosis and significantly reduced proliferation towing to G2/M cell cycle delay, concomitant aberrant mitotic spindle formation, and induction of phospho-aurora A(Thr288). The relevance of c-Rel in epithelial carcinogenesis was further supported by detection of c-Rel expression in squamous cell carcinomas of the skin. Our studies indicate that c-Rel is a key regulator of cell fate decisions in keratinocytes such as cell growth and death and may have a role in epidermal carcinogenesis.

Vitiligo: is it grace or curse?

Vitiligo is a common skin disease, affecting approximately 0.5% of the general population. It is characterized by milky white macules and patches, which are a psychological burden to many patients. Although this disease has been known for a long time, the etiology is still under debate. Since melanin is a unique light absorbing and ultraviolet filtering pigment, it is generally accepted that its main function resides in the protection of skin cells against the deleterious effect of ultraviolet rays (UVRs). The occurrence of skin cancer in long lasting vitiligo is rare despite multiple evidence of DNA damage. The aim of this study was the immunohistochemical detection of p53 and Mdm2 in depigmented and "normal" pigmented skin of vitiligo patients to demonstrate the possible role of these proteins in the protection of vitiligo patients against actinic damage and non-melanoma skin cancer. Using standard immunohistochemical techniques, we examined 34 patients with vitiligo and 30 age- and sex-matched patients with noduloulcerative basal cell carcinoma as a control group. Both patients and control subjects had outdoor occupations. Skin biopsies were obtained from each case (from depigmented and "normal" pigmented UVR-exposed skin) and control subjects (from perilesional healthy skin). Both p53 and Mdm2 were strongly expressed in depigmented as well as "normal" pigmented skin of vitiligo patients. This expression involved the epidermis, skin adnexa and blood vessels, with significant differences between cases and controls. Both proteins showed nuclear and nucleo-cytoplasmic pattern of expression. Intense p53 and Mdm2 expression was in favor of generalized vitiligo. These results suggested that the over-expression of p53 and Mdm2 proteins in both depigmented and "normal" pigmented skin of patients with vitiligo could contribute to the decreased occurrence of actinic damage and non-melanoma skin cancer in these patients.

PpIX fluorescence combined with auto-fluorescence is more accurate than PpIX fluorescence alone in fluorescence detection of non-melanoma skin cancer: an intra-patient direct comparison study.

BACKGROUND: Previous research on fluorescence detection of non-melanoma had mixed results. An accurate non-invasive method for the detection of skin cancer is valuable to dermatologists because of the high incidence of skin cancer among the aging population. One notable difference between the methods of fluorescence detection previously studied was the use of the auto-fluorescence of the skin. Currently, there has not been a direct comparison between both methods of fluorescence detection. OBJECTIVE: To compare the accuracy of PpIX fluorescence and auto-fluorescence normalized PpIX fluorescence detection systems for the localization non-melanoma skin cancers (NMSC). METHODS: We conducted an observer blinded direct comparison of both methods. Thirty patients, 14 females and16 males, mean age 62 (SD = 9 years), skin type I to III and being suspected of having one or more NMSC, visited an independent treatment centre for dermatology. The patients were investigated using a fluorescence detection system capable of both normalized and non-normalized PpIX fluorescence measurements. Liposomal encapsulated 5-aminolevulinic acid was used as a photosensitizer. For each area being investigated, the associated normalized and non-normalized fluorescence measurements were directly compared. The results of the analysis were confirmed by clinical investigation using a dermatoscope. Both methods were evaluated based on the number of true and false positives and the number of true and false negatives. Specificity and sensitivity were calculated. Statistical significance of the findings was determined using Pearson's Chi-squared test. RESULTS: The non-normalized method was found to have a sensitivity of 27 % and a specificity of 39 % and the normalized method has a sensitivity of 97% and a specificity of 100%. This difference is statistically significant (p < 0.05). CONCLUSION: Using auto-fluorescence in PpIX fluorescence detection of NMSC is more accurate that PpIX fluorescence detection alone.

Current management of actinic keratoses.

An actinic keratosis (AK) is a pre-malignant cutaneous lesion that frequently manifests in sun-exposed areas of the skin as a small, rough, scaly erythematous papule. They are one of the most common presenting complaints for dermatologists. AKs should be treated due to their potential to progress into a squamous cell carcinoma (SCC). There are numerous treatments available for managing AKs including those broadly categorized as destructive, topical field, and procedural field therapies. The topical field therapies include 5-fluorouracil, imiquimod, and diclofenac gel. Recently, imiquimod 3.75% (Zyclara TM) has been approved for the treatment AKs on the face and scalp. It is a reasonable alternative to imiquimod 5%, as the approved indication includes a larger surface area for treatment, shorter duration course, and the potential for less severe local skin reactions. There is no widely accepted algorithm for the treatment of AKs, as comparative data is unavailable between all approaches. Therapy choices are guided by efficacy, adverse effects, cosmetic results, and patient compliance.

Pharmacokinetics of imiquimod 3.75% cream applied daily for 3 weeks to actinic keratoses on the face and/or balding scalp.

Imiquimod 3.75% cream is a topical formulation of imiquimod intended for daily application to treat actinic keratoses of the entire face or balding scalp. The objective of the study was to characterize serum imiquimod and metabolite pharmacokinetics. Nineteen subjects with actinic keratoses applied two packets of imiquimod 3.75% cream (18.75 mg imiquimod total) once daily for 21 days to a treatment area approximately 200 cm(2) in size on the face and/or balding scalp. Blood samples were obtained prior to application of doses 1, 7, 14 and 21, and at selected timepoints after application of doses 1 and 21. After multiple dosing (day 21) serum imiquimod mean C (max) was 0.323 (standard deviation 0.159) ng/mL, mean AUC(0-24) 5.974 (3.088) ng h/mL, and mean T(1/2) 29.3 (17.0) h. Steady-state was achieved by day 14; multiple dose accumulation ratios were 2.8 based on imiquimod C (max) and 3.9 based on AUC. Serum concentrations of imiquimod metabolites were only sporadically quantifiable in three subjects. One subject discontinued from study for adverse events of body aches and fatigue that were attributed to study drug. Treatment-related adverse events occurred in 42.1% (8/19) of the subjects. Systemic imiquimod exposure, as reflected by serum drug concentration, was low after daily application of two packets of imiquimod 3.75% cream for 21 days. Steady state was achieved by day 14, and the observed half-life of approximately 29 h supports daily dosing of the product.

Innovative agents for actinic keratosis and nanocarriers enhancing skin penetration.

Actinic keratosis and cutaneous squamous cell carcinoma are of increasing importance with aging and increased ultraviolet light exposure in Western societies. Efficient and well-tolerated therapy is still a matter of concern. As with tumours of other organs, new target sites and innovative drugs selectively addressing them are widely looked for. Due to the relevance for DNA synthesis and thus cell proliferation, human DNA polymerase alpha should be such a target, the more so as the three-dimensional structure of the active site has been proposed based on the application of molecular modelling methods and molecular dynamics simulations. The modelled structure of the active site was used for docking nucleotide analogues in order to design selective inhibitors. Consequently, well-fitting thymidine and guanosine analogues were synthesized and tested in vitro for their influence on normal and transformed human keratinocytes. In fact, the combination of modelling studies and in vitro tests allowed us to design antiproliferative and cytotoxic agents which are new drug candidates for the therapy of skin tumours, given the agents are no relevant substrates of nucleotide transporters (MRP-4, MRP-5) expressed by skin cancer cells. Essential kinases for nucleoside activation were detected, too, corresponding with the observed effects of nucleoside analogues. Due to the rather high molecular weight and poor solubility, however, skin penetration should be poor and thus topical therapy may require carriers to improve the uptake. This becomes feasible by lipidic and non-lipidic nanoparticles which can enhance the uptake of lipophilic agents up to 13-fold.

Triterpenes promote keratinocyte differentiation in vitro, ex vivo and in vivo: a role for the transient receptor potential canonical (subtype) 6.

It has been shown recently that triterpenes inhibit cancer cell growth of various cell types in vitro. In this work, the effect of highly purified triterpenes (TE) with betulin as the major compound (>80% w/w) on cell proliferation, apoptosis, and differentiation of human keratinocytes was analyzed in vitro, ex vivo, and in vivo. In vitro, TE increased calcium influx into primary keratinocytes and upregulated various differentiation markers including keratin 10. TE also specifically increased the expression of the non-selective transient receptor potential canonical (subtype) 6 (TRPC6) in keratinocytes, and knocking down TRPC6 inhibited keratin 10 upregulation. Ex vivo, in human skin explants TE induced the expression of TRPC6 in the epidermis and increased DNA fragmentation of terminally differentiating keratinocytes. Topical treatment with TE of actinic keratoses, that represent in situ squamous cell carcinomas with disturbed epithelial differentiation, resulted in downgrading of aberrant Ki67 expression and upregulation of keratin 10 in vivo. Our data indicate that TE promotes keratinocyte differentiation in vitro and in vivo. This effect seems to be mediated at least in part by TRPC6.

Italian guidelines and therapeutic algorithm for actinic keratoses.

The prevalence of actinic keratosis (AK) continues to rise among white people throughout the world and it is necessary to increase the level of attention paid to it from a diagnostic and a preventive point of view. Today, AK must be considered an in situ squamous cell carcinoma and as such, must be managed using one of the available approved therapeutic alternatives. However, when multiple AKs develop on severely photodamaged skin, the treatment of the lesion together with that of the field of cancerization is part of an optimal strategy that aims not only to solve alterations clinically evident but also those in the surrounding skin field cancerization, that most likely hosts genetic alterations and is the site of initial gradual replacement of normal cells with tumoral cells. This paper reports the most recent evidences from a careful review of the literature's key articles of the treatment of AKs and suggests guidelines for the clinicians. The guidelines indicated by the authors have also been based on practical evaluations and their own clinical experience. The present conclusions may be modified by new findings in the field of oncologic research.