Mobile health technologies in an interventional hybrid study on actinic keratosis: Results from an early phase randomized controlled trial investigating the safety and efficacy of a cytosolic phospholipase A2 inhibitor gel in photodamaged skin.

Hybrid trials are a new trend in dermatological research that leverage mobile health technologies to decentralize a subset of clinical trial elements and thereby reduce the number of in-clinic visits. In a Phase I/IIa randomized controlled hybrid trial, the safety and efficacy of an anti-proliferative and anti-inflammatory drug inhibiting cytosolic phospholipase A2 (AVX001) was tested using 1%, 3% or vehicle gel in 60 patients with actinic keratosis (AK) and assessed in-clinic as well as remotely. Over the course of 12 weeks, patients were assessed in-clinic at baseline, end of treatment (EOT) and end of study (EOS), as well as 9 times remotely on a weekly to biweekly basis. Safety outcomes comprising local skin reactions (LSR; 0-5), adverse events (AE) and cosmesis, were graded in-clinic and remotely using patient-obtained smartphone photographs (PSPs) and questionnaires; efficacy was assessed in-clinic based on clinically visible clearance of AK target area of >50%. A total of 55 participants (91.7%) completed the treatment course. The average submission rate of PSPs was high (≥85%), of which 93% were of sufficient quality. No serious AE were reported and only two experienced temporary LSR >2 (scale 0-4) and cosmesis remained stable throughout the study. Based on the mild AE and LSR profile, daily application of AVX001 gel for 1 month appears safe, tolerable, and cosmetically acceptable for use in patients with AK. At EOT, AVX001 achieved a subtle treatment response with clearance of AK target area of >50% in 18% of patients. Remote and in-clinic assessments of LSRs were in high agreement, suggesting that the use of mobile health technologies in early-phase hybrid studies of AK does not compromise patient safety.

Accelerating patient recruitment using social media: Early adopter experience from a good clinical practice-monitored randomized controlled phase I/IIa clinical trial on actinic keratosis.

Background: Patient recruitment is a major cause of delays in randomized controlled trials (RCT). Online recruitment is evolving into an alternative to conventional in-clinic recruitment for RCT. The objective of this study was to test the effectiveness of online patient recruitment for an RCT on actinic keratosis (AK). Methods: In this proof-of-concept study, adults with AK were recruited for a Phase I/IIa RCT (NCT05164393) via social media using targeted advertising Interested users were directed to a landing page to learn about the study, respond to questionnaires, and upload self-obtained smartphone pictures of potential AK. Facebook Analytics was used to track the number of advertisement views, individual users exposed to the advertisement, and advertisement clicks. Following eligibility-review by remote dermatologists, candidates were contacted for an in-clinic visit. A review of pertinent RCTs on AK (2012-2022) was conducted to compare recruitment metrics. Results: The online campaign served 886,670 views, reached 309,000 users, and generated 27,814 clicks. A total of 556 users underwent eligibility review, leading to 140 pre-evaluated potential study subjects. The RCT's enrollment target of 60 patients (68.8 ± 7.1 years, 43.3 % female) was reached in 53 days after screening 90 participants in-clinic, corresponding to a screen failure rate of 33.3 %. The total cost of this online recruitment campaign was 14,285 USD i.e. 238 USD per randomized patient. Compared to the existing literature (44 RCTs), our online approach resulted in 9 times more time-efficient recruitment per site. Conclusion: Using targeted advertisements, 60 patients with AK were recruited for a single-center Phase I/IIa RCT in 53 days. Social media appears to be an efficient platform for online recruitment of patients with low-grade AK for RCT.

Assessing anatomical distribution of atopic dermatitis identifies a cluster of patients with late onset and low risk of asthma and allergy: An observational study.

Background and Aims: A better understanding of distinct subgroups in atopic dermatitis (AD) is warranted. The aim was to identify and determine characteristics of clusters based on anatomical location of AD. Methods: In this 8-week, observational, decentralized study, patients with AD completed a baseline questionnaire about anatomical location and severity of AD, and a principal component analysis (PCA) was applied to identify clusters. The Patient-Oriented Eczema Measure (POEM) was completed weekly and photographs of affected body areas were captured by the participants' own smartphones. From the weekly photographs, the AD severity was evaluated using the intensity part of the SCORing Atopic Dermatitis. Results: Fifty-five participants were recruited, of which 53 completed the baseline questionnaire with a mean POEM of 14.5 (SD: 5.6). The PCA analysis revealed three clusters, with AD predominantly on the shins, knees, and genitals (Cluster 1), with involvement of the upper body (Cluster 2), and with AD on the hands and feet (Cluster 3). Cluster 1 had a lower mean POEM score (11.12, SD: 5.3) compared with Clusters 2 (12.64, SD: 4.5) and 3 (15.98, SD: 4.7), respectively (p = 0.007). Further, Cluster 1 had the highest age of AD onset (mean 9.5 vs. 2.5 and 4.7 years, p = 0.02) and the lowest proportion of asthma/allergy (47% vs. 82% and 90%, p = 0.01). Conclusion: Three clusters of patients with AD based on affected body areas were identified. The cluster with involvement of legs and genitals was characterized by the oldest age of AD onset and the lowest prevalence of asthma/allergy.

The Copenhagen Actinic Keratosis Study (COAKS). A decentralised clinical trial to evaluate tolerability, safety and efficacy of daily field-directed topical treatment with cytosolic phospholipase A2 inhibitor, AVX001, in participants with actinic keratosis: protocol for a randomised controlled phase I/IIa trial.

INTRODUCTION: Actinic keratosis (AK) is the most common precancerous skin condition caused by long-term UV exposure. Given the high recurrence rate of 15%-53%, identifying safe and effective treatment options is warranted. AVX001, a cytosolic phospholipase A2α (cPLA2α) enzyme inhibitor, is a novel anti-inflammatory drug for field-directed, self-administered, topical therapy of AK. METHODS AND ANALYSIS: This study is a single-centre, randomised, vehicle-controlled, double-blind, parallel-group hybrid clinical trial in adults with multiple AK lesions Olsen grade 1 or 2. The hybrid design combines decentralised participant tasks and assessments with conventional in-clinic visits. Recruitment using targeted advertising on social media and eligibility prescreening are conducted via the Studies&Me online recruitment platform. Participants (n=60) are randomly assigned to 1 of 3 treatment arms: AVX001 gel 1%, AVX001 gel 3% or vehicle gel. The trial consists of a 4-week treatment period with daily field-directed topical application of the gel and an 8-week follow-up period. Participants attend in-clinic visits at baseline, week 4 and week 12. The remote participant trial tasks include questionnaires and upload of smartphone-obtained photos of the treated skin area using a study-specific web-based app. Both remote and in-clinic assessments of safety and efficacy will be performed. The primary objective is to evaluate the local tolerability of daily application of AVX001 gel (1% or 3%) compared with vehicle gel. Secondary objectives include safety, efficacy, dose-response efficacy relationship, treatment satisfaction and cosmetic outcome. Exploratory objectives include evaluations of tolerability and efficacy assessed by dermatologists using smartphone photos uploaded by participants, comparisons of in-clinic and remote assessments and assessment of AK-related skin changes by non-invasive optical imaging. ETHICS AND DISSEMINATION: Approved by the Ethics Committee of the Capital Region of Denmark (H-21018064) and the Danish Medicines Agency (2021032485). Results will be submitted for publication in peer-reviewed scientific journals. TRIAL REGISTRATION NUMBERS: 2021-000934-32; NCT05164393.

Online Patient Recruitment in Clinical Trials: Systematic Review and Meta-Analysis.

BACKGROUND: Recruitment for clinical trials continues to be a challenge, as patient recruitment is the single biggest cause of trial delays. Around 80% of trials fail to meet the initial enrollment target and timeline, and these delays can result in lost revenue of as much as US $8 million per day for drug developing companies. OBJECTIVE: This study aimed to conduct a systematic review and meta-analysis examining the effectiveness of online recruitment of participants for clinical trials compared with traditional in-clinic/offline recruitment methods. METHODS: Data on recruitment rates (the average number of patients enrolled in the study per month and per day of active recruitment) and conversion rates (the percentage of participants screened who proceed to enroll into the clinical trial), as well as study characteristics and patient demographics were collected from the included studies. Differences in online and offline recruitment rates and conversion rates were examined using random effects models. Further, a nonparametric paired Wilcoxon test was used for additional analysis on the cost-effectiveness of online patient recruitment. All data analyses were conducted in R language, and P<.05 was considered significant. RESULTS: In total, 3861 articles were screened for inclusion. Of these, 61 studies were included in the review, and 23 of these were further included in the meta-analysis. We found online recruitment to be significantly more effective with respect to the recruitment rate for active days of recruitment, where 100% (7/7) of the studies included had a better online recruitment rate compared with offline recruitment (incidence rate ratio [IRR] 4.17, P=.04). When examining the entire recruitment period in months we found that 52% (12/23) of the studies had a better online recruitment rate compared with the offline recruitment rate (IRR 1.11, P=.71). For cost-effectiveness, we found that online recruitment had a significantly lower cost per enrollee compared with offline recruitment (US $72 vs US $199, P=.04). Finally, we found that 69% (9/13) of studies had significantly better offline conversion rates compared with online conversion rates (risk ratio 0.8, P=.02). CONCLUSIONS: Targeting potential participants using online remedies is an effective approach for patient recruitment for clinical research. Online recruitment was both superior in regard to time efficiency and cost-effectiveness compared with offline recruitment. In contrast, offline recruitment outperformed online recruitment with respect to conversion rate.

Memory Buttons in Combination with Mobile Application-Induced Objective and Subjective Effects in Patients with Atopic Dermatitis.

BACKGROUND: Atopic dermatitis (AD) is a chronic skin condition where nonadherence often results in lack of disease control. OBJECTIVE: We wanted to determine whether the combination of an electronic memory button and a supportive application (app) would affect the Quality of Life and subjective and objective severity measures among AD patients over one month following the patient's normal schedules of treatment. METHODS: A randomized, investigator-blinded, prospective observational feasibility study for one month where patients diagnosed with AD were randomized based on POEM severity score and divided into 3 groups. The 3 groups were (1) the control group with two consultations, (2) in addition to group 1, patients also received electronic memory buttons to click every time they used their topical products, and (3) in addition to group 2, patients also received an app to track their treatment schedules. At both consultations, patients were evaluated using SCORAD, EASI, POEM, and DLQI. RESULTS: 96 patients were enrolled and randomized, of which 83 patients completed the study. EASI and SCORAD scores were lower in all groups at 2nd consultation (p < 0.05); however, these were highly significant for group 3 (p < 0.05); however, these were highly significant for group 3 (p < 0.05); however, these were highly significant for group 3 (p < 0.05); however, these were highly significant for group 3 (. CONCLUSION: A reduction in severity following objective assessments of the AD was observed for all groups and was highly significant for patients offered a memory button and the corresponding app. Furthermore, patients reported a significant subjective beneficial effect if they used the memory button and app. This indicates that digital solutions may have a benefit in clinical practice and may reduce nonadherence and increase the wellbeing of the patients.

Tumor-Preferential Induction of Immune Responses and Epidermal Cell Death in Actinic Keratoses by Ingenol Mebutate.

UNLABELLED: The rapid and strong clinical efficacy of the first-in-class, ingenol mebutate, against actinic keratosis (AK) has resulted in its recent approval. We conducted the first comprehensive analysis of the cellular and molecular mode of action of topical ingenol mebutate 0.05% gel in both AK and uninvolved skin of 26 patients in a phase I, single-center, open-label, within-patient comparison. As early as 1 day after application, ingenol mebutate induced profound epidermal cell death, along with a strong infiltrate of CD4+ and CD8+ T-cells, neutrophils, and macrophages. Endothelial ICAM-1 activation became evident after 2 days. The reaction pattern was significantly more pronounced in AK compared with uninvolved skin, suggesting a tumor-preferential mode of action. Extensive molecular analyses and transcriptomic profiling of mRNAs and microRNAs demonstrated alterations in gene clusters functionally associated with epidermal development, inflammation, innate immunity, and response to wounding. Ingenol mebutate reveals a unique mode of action linking directly to anti-tumoral effects. TRIAL REGISTRATION: ClinicalTrials.gov NCT01387711.

Repeated treatments with ingenol mebutate for prophylaxis of UV-induced squamous cell carcinoma in hairless mice.

BACKGROUND AND AIM: The incidence of squamous cell carcinomas (SCC) is increasing, and effective chemopreventative strategies are needed. We hypothesized that repeated treatments with ingenol mebutate (IngMeb) would postpone development of SCC in hairless mice, and that application of a corticosteroid would reduce IngMeb-induced local skin responses (LSRs) without affecting tumor postponement. METHODS: Hairless mice (n=150; 6 groups á 25 mice) were irradiated with solar simulated ultraviolet radiation (UVR) until SCC developed. During UV-irradiation and before tumor development, five single treatments (Tx) with IngMeb were given at four-week intervals (days 21, 49, 77, 105, 133). Clobetasol propionate (CP) was applied once daily for 5days prior to IngMeb, as well as 6h and 1day post treatment. Tumor formation was evaluated weekly for 52weeks. LSR (scale 0-24) were assessed at baseline, 1h, 6h, 1-, 2-, 3-, 4-, 5-, 6-, and 7days after each IngMeb treatment. RESULTS: IngMeb significantly delayed tumor development compared to UVR alone (UVR day 168 vs. UVR+IngMeb day 189; p=0.025). LSR included erythema, flaking, crusting, bleeding, vesiculation, and ulceration. The composite LSR-scores were of moderate intensity in non-UV irradiated skin (max LSR IngMeb Tx 1-5: 1.5-2.5) and more pronounced in photodamaged skin (max LSR Tx 5; IngMeb 1.5 vs. UVR+IngMeb 1.8; p<0.001). LSR intensity correlated with tumor development by means of greater composite LSR-score resulted in longer tumor-free survival (r(2)=0.257, p<0.001). Contrary to our hypothesis, concurrent CP increased LSR (max LSR Tx 1-5: UVR+CP+IngMeb 3.2-4.9 vs. UVR+IngMeb 1.3-2.2, p<0.001) and postponed tumor development compared to IngMeb alone (UVR+CP+IngMeb day 217 vs. UVR+IngMeb day 189, p<0.001). CONCLUSION: Repeated field-directed treatments with IngMeb delay development of UV-induced SCC in hairless mice, and increased IngMeb induced LSRs correlated with improved clinical outcomes. The findings highlight the potential of IngMeb as a prophylactic remedy for SCC in humans.

Unique microRNAs appear at different times during the course of a delayed-type hypersensitivity reaction in human skin.

Diphencyprone (DPCP) is a hapten that induces delayed-type hypersensitivity (DTH) reactions. MicroRNAs (miRNAs) are short non-coding RNAs that negatively regulate gene expression and have been implicated in various inflammatory skin diseases, but their role in DTH reactions is not well understood. We generated global miRNA expression profiles (using next-generation sequencing) of DPCP reactions in skin of seven healthy volunteers at 3, 14 and 120 days after challenge. Compared to placebo-treated sites, DPCP-challenged skin at 3 days (peak inflammation) had 127 miRNAs significantly deregulated. At 14 days (during resolution of inflammation), 43 miRNAs were deregulated and, at 120 days (when inflammation had completely resolved), six miRNAs were upregulated. While some miRNAs have been observed in psoriasis or atopic dermatitis, most of the deregulated miRNAs have not yet been studied in the context of skin biology or immunology. Across the three time points studied, many but not all miRNAs were uniquely expressed. As various miRNAs may influence T cell activation, this may indicate that the miRNAs exclusively expressed at different time points function to promote or resolve skin inflammation, and therefore, may inform on the paradoxical ability of DPCP to treat both autoimmune conditions (alopecia areata) and conditions of ineffective immunity (melanoma).

Ingenol Mebutate Signals via PKC/MEK/ERK in Keratinocytes and Induces Interleukin Decoy Receptors IL1R2 and IL13RA2.

Squamous cell carcinoma (SCC) is the second most common human skin cancer and the second leading cause of skin cancer-related death. Recently, a new compound, ingenol mebutate, was approved for treatment of actinic keratosis, a precursor of SCC. As the mechanism of action is poorly understood, we have further investigated the mechanism of ingenol mebutate-induced cell death. We elucidate direct effects of ingenol mebutate on primary keratinocytes, patient-derived SCC cells, and a SCC cell line. Transcriptional profiling followed by pathway analysis was performed on ingenol mebutate-treated primary keratinocytes and patient-derived SCC cells to find key mediators and identify the mechanism of action. Activation of the resulting pathways was confirmed in cells and human skin explants and supported by a phosphorylation screen of treated primary cells. The necessity of these pathways was demonstrated by inhibition of certain pathway components. Ingenol mebutate inhibited viability and proliferation of all keratinocyte-derived cells in a biphasic manner. Transcriptional profiling identified the involvement of PKC/MEK/ERK signaling in the mechanism of action and inhibition of this signaling pathway rescued ingenol mebutate-induced cell death after treatment with 100 nmol/L ingenol mebutate, the optimal concentration for the first peak of response. We found the interleukin decoy receptors IL1R2 and IL13RA2 induced by ingenol mebutate in a PKC/MEK/ERK-dependent manner. Furthermore, siRNA knockdown of IL1R2 and IL13RA2 partially rescued ingenol mebutate-treated cells. In conclusion, we have shown that ingenol mebutate-induced cell death is mediated through the PKCδ/MEK/ERK pathway, and we have functionally linked the downstream induction of IL1R2 and IL13RA2 expression to the reduced viability of ingenol mebutate-treated cells.

Laser capture microdissection followed by next-generation sequencing identifies disease-related microRNAs in psoriatic skin that reflect systemic microRNA changes in psoriasis.

Psoriasis is a systemic disease with cutaneous manifestations. MicroRNAs (miRNAs) are small non-coding RNA molecules that are differentially expressed in psoriatic skin; however, only few cell- and region-specific miRNAs have been identified in psoriatic lesions. We used laser capture microdissection (LCM) and next-generation sequencing (NGS) to study the specific miRNA expression profiles in the epidermis (Epi) and dermal inflammatory infiltrates (RD) of psoriatic skin (N = 6). We identified 24 deregulated miRNAs in the Epi and 37 deregulated miRNAs in the RD of psoriatic plaque compared with normal psoriatic skin (FCH > 2, FDR < 0.05). Interestingly, 9 of the 37 miRNAs in RD, including miR-193b and miR-223, were recently described as deregulated in circulating peripheral blood mononuclear cells (PBMCs) from patients with psoriasis. Using flow cytometry and qRT-PCR, we found that miR-193b and miR-223 were expressed in Th17 cells. In conclusion, we demonstrate that LCM combined with NGS provides a robust approach to explore the global miRNA expression in the epidermal and dermal compartments of psoriatic skin. Furthermore, our results indicate that the altered local miRNA changes seen in the RD are reflected in the circulating immune cells, suggesting that miRNAs may contribute to the pathogenesis of psoriasis.

MicroRNA-223 and miR-143 are important systemic biomarkers for disease activity in psoriasis.

BACKGROUND: Psoriasis is a systemic inflammatory skin disease. MicroRNAs (miRNAs) are a class of small non-coding RNA molecules that recently have been found in the blood to be relevant as disease biomarkers. OBJECTIVE: We aimed to explore miRNAs potential as blood biomarkers for psoriasis. METHODS: Using microarray and quantitative real-time PCR we measured the global miRNA expression in whole blood, plasma and peripheral blood mononuclear cells (PBMCs) from patients with psoriasis and healthy controls. RESULTS: We identified several deregulated miRNAs in the blood from patients with psoriasis including miR-223 and miR-143 which were found to be significantly upregulated in the PBMCs from patients with psoriasis compared with healthy controls (FCH=1.63, P<0.01; FCH=2.18, P<0.01, respectively). In addition, miR-223 and miR-143 significantly correlated with the PASIscore (r=0.46, P<0.05; r=0.55, P<0.02, respectively). Receiver-operating characteristic analysis (ROC) showed that miR-223 and -143 have the potential to distinguish between psoriasis and healthy controls (miR-223: area under the curve (AUC)=0.80, miR-143: AUC=0.75). Interestingly, after 3-5 weeks of treatment with methotrexate following a significant decrease in psoriasis severity, miR-223 and miR-143 were significantly downregulated in the PBMCs from patients with psoriasis. CONCLUSION: We suggest that changes in the miR-223 and miR-143 expressions in PBMCs from patients with psoriasis may serve as novel biomarkers for disease activity in psoriasis; however, further investigations are warranted to clarify their specific roles.

A proteomics approach to the identification of biomarkers for psoriasis utilising keratome biopsy.

The discovery of plasma biomarkers for psoriasis vulgaris may aid clinicians in disease grading and monitoring of treatment response. We have therefore developed a proteomics/mass spectrometry based workflow which enables biomarker discovery from psoriasis patient samples. We have utilised keratome skin biopsy, which results in reduced cellular complexity compared to punch biopsy. Furthermore, we applied short term ex vivo culture in order to enrich for a "secretome" sub-proteome reflective of the disease and enriched in potential biomarkers. Using these sample preparation techniques we performed a quantitative proteomics screen of four patients with psoriasis using stable isotope dimethyl labelling and identified over 50 proteins consistently altered in abundance in psoriasis lesional versus non-lesional skin. This includes several canonical psoriasis related proteins (e.g. S100A7 [Psoriasin] and FABP5 [Epidermal Fatty Acid Binding Protein]) and more than 30 novel alterations. From this disease localised dataset we further assessed several proteins as potential biomarkers in the plasma of patients with psoriasis versus healthy controls utilising selected reaction monitoring mass spectrometry (SRM-MS/MS). BIOLOGICAL SIGNIFICANCE: The significance of this study for proteome research in psoriasis and dermal disease is threefold. 1) A novel sample preparation method for isolation of dermal proteomes enriched in extracellular proteins is described, which may be of interest to other researchers in the field. 2) Novel psoriasis associated alterations in protein abundance are described at the disease site, bolstering knowledge in an area dominated by transcript level studies and 3) Profilin 1 is described as a candidate plasma biomarker of psoriasis, this is of value in itself and it proves that our workflow can yield results in terms of biomarker discovery.

Ingenol mebutate: induced cell death patterns in normal and cancer epithelial cells.

We investigated the proposed necrotic mechanism of ingenol mebutate, a natural compound with anti-cancer properties in human keratinocytes, the human squamous cell carcinoma cell line HSC-5, and HeLa cervix carcinoma cells. Topical application of a clinical dose of ingenol mebutate 0.05% (1.15 mM) gel to human reconstituted full-thickness skin equivalents strongly reduced epidermal, but not dermal viability. Ingenol mebutate showed cytotoxic potency between 200-300 M on normal and cancer cells. When keratinocytes were induced to differentiate, they became significantly less sensitive to ingenol mebutate and half-maximal induction of cell death required more than 300 M ingenol mebutate. Cytotoxic concentrations of ingenol mebutate caused rupture of the mitochondrial network within minutes paralleled by cytosolic calcium release in all cells. Subsequently, plasma membrane integrity was lost as seen by propidium uptake into the cells. This was in sharp contrast to lysis of cells with low concentrations of the detergent Triton X-100 that permeabilized the plasma membrane within minutes without affecting organelle morphology. Buffering of intracellular calcium and inhibition of the mitochondrial permeability transition pore reduced the cytotoxic effect of ingenol mebutate in cancer cells, but not in normal keratinocytes. However, these inhibitors could not prevent cell death subsequent to prolonged incubation. Our findings reveal that ingenol mebutate does not mediate cytotoxicity by a simple lytic, necrotic mechanism, but activates distinct processes involving multiple cell organelles in a cell-type and differentiation-dependent manner. These data improve our understanding of ingenol mebutate-target cell interactions and offer new insights relevant to the removal of aberrant cells in human skin.

Comparison of microRNA expression using different preservation methods of matched psoriatic skin samples.

MicroRNAs are non-coding RNA molecules modulating gene expression post-transcriptionally. Formalin-fixed, paraffin-embedding (FFPE) is a standard preservation method often used in clinical practices, but induces RNA degradation. Extracting high-quality RNA from human skin can be challenging as skin contains high levels of RNases. As microRNAs are 19-23 nucleotides long and lack a poly-A tail, they may be less prone to RNA degradation than mRNAs. We investigated whether microRNAs in psoriatic (FFPE) samples reliably reflect microRNA expression in samples less prone to RNA degradation such as fresh-frozen (FS) and Tissue-Tek-embedding (OCT). We found a strong correlation of the microRNA expression levels between all preservation methods of matched psoriatic skin samples (r(s) ranging from 0.91 to 0.95 (P < 0.001)). These observations were further confirmed with qRT-PCR. Our results demonstrate that microRNA detection in human skin is robust irrespective of preservation method; thus, microRNAs offer an appropriate and flexible approach in clinical practices and for diagnostic purposes in skin disorders.

Diagnostic microRNA profiling in cutaneous T-cell lymphoma (CTCL).

Cutaneous T-cell lymphomas (CTCLs) are the most frequent primary skin lymphomas. Nevertheless, diagnosis of early disease has proven difficult because of a clinical and histologic resemblance to benign inflammatory skin diseases. To address whether microRNA (miRNA) profiling can discriminate CTCL from benign inflammation, we studied miRNA expression levels in 198 patients with CTCL, peripheral T-cell lymphoma (PTL), and benign skin diseases (psoriasis and dermatitis). Using microarrays, we show that the most induced (miR-326, miR-663b, and miR-711) and repressed (miR-203 and miR-205) miRNAs distinguish CTCL from benign skin diseases with > 90% accuracy in a training set of 90 samples and a test set of 58 blinded samples. These miRNAs also distinguish malignant and benign lesions in an independent set of 50 patients with PTL and skin inflammation and in experimental human xenograft mouse models of psoriasis and CTCL. Quantitative (q)RT-PCR analysis of 103 patients with CTCL and benign skin disorders validates differential expression of 4 of the 5 miRNAs and confirms previous reports on miR-155 in CTCL. A qRT-PCR-based classifier consisting of miR-155, miR-203, and miR-205 distinguishes CTCL from benign disorders with high specificity and sensitivity, and with a classification accuracy of 95%, indicating that miRNAs have a high diagnostic potential in CTCL.

Halting angiogenesis by non-viral somatic gene therapy alleviates psoriasis and murine psoriasiform skin lesions.

Dysregulated angiogenesis is a hallmark of chronic inflammatory diseases, including psoriasis, a common skin disorder that affects approximately 2% of the population. Studying both human psoriasis in 2 complementary xenotransplantation models and psoriasis-like skin lesions in transgenic mice with epidermal expression of human TGF-ß1, we have demonstrated that antiangiogenic non-viral somatic gene therapy reduces the cutaneous microvasculature and alleviates chronic inflammatory skin disorders. Transient muscular expression of the recombinant disintegrin domain (RDD) of metargidin (also known as ADAM-15) by in vivo electroporation reduced cutaneous angiogenesis and vascularization in all 3 models. As demonstrated using red fluorescent protein-coupled RDD, the treatment resulted in muscular expression of the gene product and its deposition within the cutaneous hyperangiogenic connective tissue. High-resolution ultrasound revealed reduced cutaneous blood flow in vivo after electroporation with RDD but not with control plasmids. In addition, angiogenesis- and inflammation-related molecular markers, keratinocyte proliferation, epidermal thickness, and clinical disease scores were downregulated in all models. Thus, non-viral antiangiogenic gene therapy can alleviate psoriasis and may do so in other angiogenesis-related inflammatory skin disorders.

MicroRNAs and potential target interactions in psoriasis.

BACKGROUND: Psoriasis is a chronic inflammatory skin disease often seen in patients with a genetic susceptibility. MicroRNAs (miRNA) are endogenous, short RNA molecules that can bind to parts of mRNA target genes, thus inhibiting their translation and causing accelerated turnover or transcript degradation. MicroRNAs are important in the pathogenesis of human diseases such as immunological disorders, as they regulate a broad range of biological processes. OBJECTIVE: We investigated miRNA-mRNA interactions in involved (PP) and non-involved (PN) psoriatic skin compared with healthy skin (NN). METHODS: Biopsies were obtained from PP, PN and NN, the miRNA and mRNA expression was analyzed by microarray techniques and a subset of miRNAs and mRNAs were validated by q-RT-PCR. Novel target interactions in psoriasis were found using PubMed, miRBase and RNAhybrid. In addition, TIMP3 protein expression was studied in PP, PN and NN. Finally, the miR-221/2-TIMP3 target interaction was studied in primary human keratinocytes by endogenous overexpression of the miRNAs. RESULTS: We identified 42 upregulated miRNAs and 5 downregulated miRNAs in PP compared with NN, and only few deregulated miRNAs in PN compared with NN. Based on the miRNA and mRNA profiles miR-21, -205, -221 and -222 were found to have the following potential mRNA targets in psoriatic skin: PDCD4, TPM1, P57, C-KIT, RTN4, SHIP2, TIMP3, RECK and NFIB. The identified target mRNAs were likely to be involved in cellular growth, proliferation, apoptosis and degradation of the extracellular matrix. Finally we found that TIMP3 is downregulated in psoriatic skin. In vitro overexpression of miR-221 and miR-222 lead to degradation of TIMP3 resulting in decreased TIMP3 protein level. CONCLUSION: Our data indicate several novel important associations for miRNAs in psoriasis and in particular the miR-221/2-TIMP3 target interaction could among others play a role in the psoriasis pathogenesis.

Significance of the S100A4 protein in psoriasis.

The S100A4 protein is reported as a pivotal player in the tumor microenvironment with a metastasis-promoting function. Moreover, the upregulation of S100A4 is found in other non-malignant human disorders as cardiac and pulmonary systems and rheumatoid arthritis. In this study, we investigated the expression and significance of S100A4 in psoriasis. We found significant upregulation of S100A4 in the dermis of psoriatic skin compared with normal skin. This pattern of S100A4 expression differs considerably from that of other S100 proteins, S100A7 and S100A8/9, with predominant expression in the epidermis of psoriasis. Furthermore, we revealed a massive release of the biologically active forms of S100A4 from psoriatic skin. Interestingly, we found stabilization (increase) of p53 in the basal layer of epidermis in close proximity to cells expressing S100A4. To examine the possible implication of S100A4 in the pathogenesis of psoriasis, we analyzed the effect of S100A4 blocking antibodies in a human psoriasis xenograft SCID mouse model and observed a significant reduction of the epidermal thickness and impairment in cell proliferation and dermal vascularization. In conclusion, we showed strong upregulation and release of S100A4 in the upper dermis of psoriatic skin and found evidence indicating that S100A4 might actively contribute to the pathogenesis of psoriasis.