The Activation of PPARγ by (2Z,4E,6E)-2-methoxyocta-2,4,6-trienoic Acid Counteracts the Epithelial-Mesenchymal Transition Process in Skin Carcinogenesis.

Cutaneous squamous cell carcinoma (cSCC) is the most common UV-induced keratinocyte-derived cancer, and its progression is characterized by the epithelial-mesenchymal transition (EMT) process. We previously demonstrated that PPARγ activation by 2,4,6-octatrienoic acid (Octa) prevents cutaneous UV damage. We investigated the possible role of the PPARγ activators Octa and the new compound (2Z,4E,6E)-2-methoxyocta-2,4,6-trienoic acid (A02) in targeting keratinocyte-derived skin cancer. Like Octa, A02 exerted a protective effect against UVB-induced oxidative stress and DNA damage in NHKs. In the squamous cell carcinoma A431 cells, A02 inhibited cell proliferation and increased differentiation markers' expression. Moreover, Octa and even more A02 counteracted the TGF-ß1-dependent increase in mesenchymal markers, intracellular ROS, the activation of EMT-related signal transduction pathways, and cells' migratory capacity. Both compounds, especially A02, counterbalanced the TGF-ß1-induced cell membrane lipid remodeling and the release of bioactive lipids involved in EMT. In vivo experiments on a murine model useful to study cell proliferation in adult animals showed the reduction of areas characterized by active cell proliferation in response to A02 topical treatment. In conclusion, targeting PPARγ may be useful for the prevention and treatment of keratinocyte-derived skin cancer.

Efficacy and safety of oral ritlecitinib for the treatment of active nonsegmental vitiligo: A randomized phase 2b clinical trial.

BACKGROUND: Vitiligo is a chronic autoimmune disorder characterized by depigmented patches of the skin. OBJECTIVE: To evaluate the efficacy and safety of ritlecitinib, an oral JAK3 (Janus kinase)/TEC (tyrosine kinase expressed in hepatocelluar carcinoma) inhibitor, in patients with active nonsegmental vitiligo in a phase 2b trial (NCT03715829). METHODS: Patients were randomized to once-daily oral ritlecitinib ± 4-week loading dose (200/50 mg, 100/50 mg, 30 mg, or 10 mg) or placebo for 24 weeks (dose-ranging period). Patients subsequently received ritlecitinib 200/50 mg daily in a 24-week extension period. The primary efficacy endpoint was percent change from baseline in Facial-Vitiligo Area Scoring Index at week 24. RESULTS: A total of 364 patients were treated in the dose-ranging period. Significant differences from placebo in percent change from baseline in Facial-Vitiligo Area Scoring Index were observed for the ritlecitinib 50 mg groups with (-21.2 vs 2.1; P < .001) or without (-18.5 vs 2.1; P < .001) a loading dose and ritlecitinib 30 mg group (-14.6 vs 2.1; P = .01). Accelerated improvement was observed after treatment with ritlecitinib 200/50 mg in the extension period (n = 187). No dose-dependent trends in treatment-emergent or serious adverse events were observed across the 48-week treatment. LIMITATIONS: Patients with stable vitiligo only were excluded. CONCLUSIONS: Oral ritlecitinib was effective and well tolerated over 48 weeks in patients with active nonsegmental vitiligo.

Research update of adipose tissue-based therapies in regenerative dermatology.

Mesenchymal stromal/stem cells (MSCs) have a spontaneous propensity to support tissue homeostasis and regeneration. Among the several sources of MSCs, adipose-derived tissue stem cells (ADSCs) have received major interest due to the higher mesenchymal stem cells concentration, ease, and safety of access. However, since a significant part of the natural capacity of ADSCs to repair damaged tissue is ascribable to their secretory activity that combines mitogenic factors, cytokines, chemokines, lipids, and extracellular matrix components, several studies focused on cell-free strategies. Furthermore, adipose cell-free derivatives are becoming more attractive especially for non-volumizing purposes, such as most dermatological conditions. However, when keratinocytes, fibroblasts, melanocytes, adipocytes, and hair follicle cells might not be locally sourced, graft of materials containing concentrated ADSCs is preferred. The usage of extracellular elements of adipose tissue aims to promote a self-autonomous regenerative microenvironment in the receiving area restoring physiological homeostasis. Hence, ADSCs or their paracrine activity are currently being studied in several dermatological settings including wound healing, skin fibrosis, burn, and aging.The present work analyzing both preclinical and clinical experiences gives an overview of the efficacy of adipose tissue-derivatives like autologous fat, the stromal vascular fraction (SVF), purified ADSCs, secretome and extracellular matrix graft in the field of regenerative medicine for the skin.

Therapeutic potential of adipose tissue-derivatives in modern dermatology.

Stem cell-mediated therapies in combination with biomaterial and growth factor-based approaches in regenerative medicine are rapidly evolving with increasing application beyond the dermatologic field. Adipose-derived stem cells (ADSCs) are the more frequently used adult stem cells due to their abundance and easy access. In the case of volumetric defects, adipose tissue can take the shape of defects, restoring the volume and enhancing the regeneration of receiving tissue. When regenerative purposes prevail on volume restoration, the stromal vascular fraction (SVF) rich in staminal cells, purified mesenchymal stem cells (MSCs) or their cell-free derivatives grafting are favoured. The therapeutic efficacy of acellular approaches is explained by the fact that a significant part of the natural propensity of stem cells to repair damaged tissue is ascribable to their secretory activity that combines mitogenic factors, cytokines, chemokines and extracellular matrix components. Therefore, the secretome's ability to modulate multiple targets simultaneously demonstrated preclinical and clinical efficacy in reversing pathological mechanisms of complex conditions such atopic dermatitis (AD), vitiligo, psoriasis, acne and Lichen sclerosus (LS), non-resolving wounds and alopecia. This review analysing both in vivo and in vitro models gives an overview of the clinical relevance of adipose tissue-derivatives such as autologous fat graft, stromal vascular fraction, purified stem cells and secretome for skin disorders application. Finally, we highlighted the major disease-specific limitations and the future perspective in this field.

Profiling Cancer-Associated Fibroblasts in Melanoma.

Solid tumors are complex systems characterized by dynamic interactions between neoplastic cells, non-tumoral cells, and extracellular components. Among all the stromal cells that populate tumor microenvironment, fibroblasts are the most abundant elements and are critically involved in disease progression. Cancer-associated fibroblasts (CAFs) have pleiotropic functions in tumor growth and extracellular matrix remodeling implicated in local invasion and distant metastasis. CAFs additionally participate in the inflammatory response of the tumor site by releasing a variety of chemokines and cytokines. It is becoming clear that understanding the dynamic, mutual melanoma-fibroblast relationship would enable treatment options to be amplified. To better characterize melanoma-associated fibroblasts, here we analyzed low-passage primary CAFs derived from advanced-stage primary skin melanomas, focusing on the immuno-phenotype. Furthermore, we assessed the expression of several CAF markers and the production of growth factors. To deepen the study of CAF-melanoma cell crosstalk, we employed CAF-derived supernatants and trans-well co-culture systems to evaluate the influences of CAFs on (i) the motogenic ability of melanoma cells, (ii) the chemotherapy-induced cytotoxicity, and (iii) the release of mediators active in modulating tumor growth and spread.

Anti-Inflammatory and Pro-Differentiating Properties of the Aryl Hydrocarbon Receptor Ligands NPD-0614-13 and NPD-0614-24: Potential Therapeutic Benefits in Psoriasis.

The aryl hydrocarbon receptor (AhR), a ligand-activated transcription factor expressed in all skin cell types, plays a key role in physiological and pathological processes. Several studies have shown that this receptor is involved in the prevention of inflammatory skin diseases, e.g., psoriasis, atopic dermatitis, representing a potential therapeutic target. We tested the safety profile and the biological activity of NPD-0614-13 and NPD-0614-24, two new synthetic AhR ligands structurally related to the natural agonist FICZ, known to be effective in psoriasis. NPD-0614-13 and NPD-0614-24 did not alter per se the physiological functions of the different skin cell populations involved in the pathogenesis of inflammatory skin diseases. In human primary keratinocytes stimulated with tumor necrosis factor-α or lipopolysaccharide the compounds were able to counteract the altered proliferation and to dampen inflammatory signaling by reducing the activation of p38MAPK, c-Jun, NF-kBp65, and the release of cytokines. Furthermore, the molecules were tested for their beneficial effects in human epidermal and full-thickness reconstituted skin models of psoriasis. NPD-0614-13 and NPD-0614-24 recovered the psoriasis skin phenotype exerting pro-differentiating activity and reducing the expression of pro-inflammatory cytokines and antimicrobial peptides. These data provide a rationale for considering NPD-0614-13 and NPD-0614-24 in the management of psoriasis.

Simultaneous Targeting Tumor Cells and Cancer-Associated Fibroblasts with a Paclitaxel-Hyaluronan Bioconjugate: In Vitro Evaluation in Non-Melanoma Skin Cancer.

BACKGROUND: Cancer-associated fibroblasts (CAFs) facilitate many aspects of cancer development by providing a structural framework rich in bioactive compounds. There are emerging studies proposing a combination of conventional anti-cancer therapies directed against neoplastic cells to molecules targeting tumor microenvironments. METHODS: The study evaluated the pharmacological properties of the anti-tumor agent paclitaxel conjugated to hyaluronic acid (HA) regarding non-melanoma skin cancer (NMSC) and the surrounding fibroblasts. This molecule, named Oncofid-P20 (Onco-P20), preferentially targets cells expressing high levels of CD44, the natural ligand of HA. RESULTS: Consistent with paclitaxel's mechanism of action involving interference with the breakdown of microtubules during cell division, highly sensitive carcinoma cells rapidly underwent apoptotic cell death. Interestingly, less sensitive cells, such as dermal fibroblasts, resisted the Onco-P20 treatment and experienced a prolonged growth arrest characterized by morphological change and significant modification of the gene expression profile. Onco-P20-treated fibroblasts exhibited reduced growth factor production, downmodulation of the Wnt signaling pathway, and the acquisition of a marked pro-inflammatory profile. Independently of direct exposure to taxol, in the presence of Onco-P20-treated fibroblasts or in their conditioned medium, carcinoma cells had a reduced proliferation rate. Similar to NHF, fibroblasts isolated from skin cancer lesions or from adjacent tissue acquired anti-neoplastic activity under Onco-P20 treatment. CONCLUSION: Collectively, our data demonstrate that Onco-P20, exerting both a direct and an NHF-mediated indirect effect on carcinoma cells, is a candidate for an innovative therapy alternative to surgery for the treatment of NMSC.

Alterations of the pigmentation system in the aging process.

Human skin aging is a natural phenomenon that results from continuous exposure to intrinsic (time, genetic factors, hormones) as well as extrinsic factors (UV exposure, pollution, tobacco). In areas that are frequently exposed to the sun, photoaging blends with the process of intrinsic aging, resulting in an increased senescent cells number and consequently accelerating the aging process. The severity of photodamage depends on constitutional factors, including skin phototype (skin color, tanning capacity), intensity, and duration of sunlight/UV exposure. Aging affects nearly every aspect of cutaneous biology, including pigmentation. Clinically, the phenotype of age pigmented skin has a mottled, uneven color, primarily due to age spots, with or without hypopigmentation. Uneven pigmentation might be attributed to the hyperactivation of melanocytes, altered distribution of pigment, and turnover. In addition to direct damage to pigment-producing cells, photodamage alters the physiological crosstalk between keratinocytes, fibroblasts, endothelial cells, and melanocytes responsible for natural pigmentation homeostasis. Interestingly, age-independent diffuse expression of senescence-associated markers in the dermal and epidermal compartment is also associated with vitiligo, suggesting that premature senescence plays an important role in the pathology.

An update on Vitiligo pathogenesis.

Vitiligo, the most common depigmenting disorder of the skin, is undergoing a period of intense advances in both disease understanding and therapeutic possibilities leading the way to the beginning of a new era for the disorder. Its pathophysiology has gathered the attention of researchers for years, and many advances have been made in the clarification of the interaction between different factors that result in depigmented macule formation. The complex interplay between non-immunological and immunological factors in vitiligo is key for the development of the disease, and the participation of cells other than melanocytes, such as keratinocytes, fibroblasts, natural killer cells, and innate lymphoid cells, has been shown. Recent advances have also brought to the understanding of the complex part played by a specific subtype of T cells: T-resident memory cells. This review analyzes some of the most recent insights in vitiligo pathogenesis underlining the interactions between different cell types, which are the basis for the therapeutic approaches under development.

A Framework of Major Tumor-Promoting Signal Transduction Pathways Implicated in Melanoma-Fibroblast Dialogue.

The development of a modified stromal microenvironment in response to neoplastic onset is a common feature of many tumors including cutaneous melanoma. At all stages, melanoma cells are embedded in a complex tissue composed by extracellular matrix components and several different cell populations. Thus, melanomagenesis is not only driven by malignant melanocytes, but also by the altered communication between melanocytes and non-malignant cell populations, including fibroblasts, endothelial and immune cells. In particular, cancer-associated fibroblasts (CAFs), also referred as melanoma-associated fibroblasts (MAFs) in the case of melanoma, are the most abundant stromal cells and play a significant contextual role in melanoma initiation, progression and metastasis. As a result of dynamic intercellular molecular dialogue between tumor and the stroma, non-neoplastic cells gain specific phenotypes and functions that are pro-tumorigenic. Targeting MAFs is thus considered a promising avenue to improve melanoma therapy. Growing evidence demonstrates that aberrant regulation of oncogenic signaling is not restricted to transformed cells but also occurs in MAFs. However, in some cases, signaling pathways present opposite regulation in melanoma and surrounding area, suggesting that therapeutic strategies need to carefully consider the tumor-stroma equilibrium. In this novel review, we analyze four major signaling pathways implicated in melanomagenesis, TGF-ß, MAPK, Wnt/ß-catenin and Hyppo signaling, from the complementary point of view of tumor cells and the microenvironment.

Paired Basic Amino Acid-cleaving Enzyme 4 (PCSK6): An Emerging New Target Molecule in Human Melanoma.

Although recent therapeutic developments raise hope, melanoma remains a devastating disease with a need for new treatment targets. In other tumours prohormone convertases have been shown to be pro-tumourigenic as they are involved in processing preforms of matrix-metalloproteinases, growth factors and adhesion molecules. The aim of this study was to look for new treatment options for melanoma, by investigating the role of the prohormone convertase Paired basic Amino acid-Cleaving Enzyme 4 (PACE4/PCSK6) in melanoma cell lines and human melanoma tissue. PACE4-transfected A375 melanoma cells displayed significantly increased proliferation, MMP-2 production, gelatinase activity and migratory capacity in vitro compared with sham-transfected cells. In vivo, elevated PACE4 expression resulted in significantly increased tumour growth on immunodeficient mice. In the majority of 45 human primary melanomas and melanoma metastases ex vivo PACE4 immunoreactivity was detectable, while it was absent in in situ melanomas. These results indicate PACE4 as a regulator of melanoma cell aggressiveness.

The Role of Dermal Fibroblasts in Nevoid Basal Cell Carcinoma Syndrome Patients: An Overview.

Nevoid basal cell carcinoma syndrome (NBCCS), also named Gorlin syndrome, is a rare multisystem genetic disorder characterized by marked predisposition to basal cell carcinomas (BCCs), childhood medulloblastomas, maxillary keratocysts, celebral calcifications, in addition to various skeletal and soft tissue developmental abnormalities. Mutations in the tumor suppressor gene PATCHED1 (PTCH1) have been found to be associated in the majority of NBCCS cases. PATCH1 somatic mutations and loss of heterozygosity are also very frequent in sporadic BCCs. Unlike non-syndromic patients, NBCCS patients develop multiple BCCs in sun-protected skin area starting from early adulthood. Recent studies suggest that dermo/epidermal interaction could be implicated in BCC predisposition. According to this idea, NBCCS fibroblasts, sharing with keratinocytes the same PTCH1 germline mutation and consequent constitutive activation of the Hh pathway, display features of carcinoma-associated fibroblasts (CAF). This phenotypic traits include the overexpression of growth factors, specific microRNAs profile, modification of extracellular matrix and basement membrane composition, increased cytokines and pro-angiogenic factors secretion, and a complex alteration of the Wnt/-catenin pathway. Here, we review studies about the involvement of dermal fibroblasts in BCC predisposition of Gorlin syndrome patients. Further, we matched the emerged NBCCS fibroblast profile to those of CAF to compare the impact of cell autonomous "pre-activated state" due to PTCH1 mutations to those of skin tumor stroma.

Premature cell senescence in human skin: Dual face in chronic acquired pigmentary disorders.

Although senescence was originally described as an in vitro acquired cellular characteristic, it was recently recognized that senescence is physiologically and pathologically involved in aging and age-related diseases in vivo. The definition of cellular senescence has expanded to include the growth arrest caused by various cellular stresses, including DNA damage, inadequate mitochondria function, activated oncogene or tumor suppressor genes and oxidative stress. While senescence in normal aging involves various tissues over time and contributes to a decline in tissue function even with healthy aging, disease-induced premature senescence may be restricted to one or a few organs triggering a prolonged and more intense rate of accumulation of senescent cells than in normal aging. Organ-specific high senescence rate could lead to chronic diseases, especially in post-mitotic rich tissue. Recently, two opposite acquired pathological conditions related to skin pigmentation were described to be associated with premature senescence: vitiligo and melasma. In both cases, it was demonstrated that pathological dysfunctions are not restricted to melanocytes, the cell type responsible for melanin production and transport to surrounding keratinocytes. Similar to physiological melanogenesis, dermal and epidermal cells contribute directly and indirectly to deregulate skin pigmentation as a result of complex intercellular communication. Thus, despite senescence usually being reported as a uniform phenotype sharing the expression of characteristic markers, skin senescence involving mainly the dermal compartment and its paracrine function could be associated with the disappearance of melanocytes in vitiligo lesions and with the exacerbated activity of melanocytes in the hyperpigmentation spots of melasma. This suggests that the difference may arise in melanocyte intrinsic differences and/or in highly defined microenvironment peculiarities poorly explored at the current state of the art. A similar dualistic phenotype has been attributed to intratumoral stromal cells as cancer-associated fibroblasts presenting a senescent-like phenotype which influence the behavior of neoplastic cells in either a tumor-promoting or tumor-inhibiting manner. Here, we present a framework dissecting senescent-related molecular alterations shared by vitiligo and melasma patients and we also discuss disease-specific differences representing new challenges for treatment.

Validation of a physician global assessment tool for vitiligo extent: Results of an international vitiligo expert meeting.

Currently, vitiligo lacks a validated Physician Global Assessment (PGA) for disease extent. This PGA can be used to stratify and interpret the numeric scores obtained by the Vitiligo Extent Score (VES). We investigated the interrater reliability of a 5-point PGA scale during an international vitiligo workshop. Vitiligo experts from five different continents rated photographs of non-segmental vitiligo patients with varying degrees of extent with the PGA score. Good interrater agreements (intraclass correlation coefficient >0.6) were observed between the raters overall and within each continent. All hypotheses to evaluate construct validity were confirmed. Median VES values per category were for limited 1.10 [IQR: 0.21-1.67], moderate 3.17 [IQR: 1.75-6.21], extensive 9.58 [IQR: 6.21-13.03] and very extensive 42.67 [IQR: 21.20-42.67]. Defined categories for vitiligo extent can be valuable for inclusion criteria and may impact future reimbursement criteria.

Acquired disorders with hypopigmentation: A clinical approach to diagnosis and treatment.

Acquired hypopigmented skin changes are commonly encountered by dermatologists. Although hypopigmentation is often asymptomatic and benign, occasional serious and disabling conditions present with cutaneous hypopigmentation. A thorough history and physical examination, centered on disease distribution and morphologic findings, can aid in delineating the causes of acquired hypopigmented disorders. The second article in this 2-part continuing medical education series focuses on conditions with a hypopigmented phenotype. Early diagnosis and appropriate management of these disorders can improve a patient's quality of life, halt disease progression, and prevent irreversible disability.

Acquired disorders with depigmentation: A systematic approach to vitiliginoid conditions.

Acquired disorders with depigmentation are commonly encountered by dermatologists and present with a wide differential diagnosis. Vitiligo, the most common disorder of acquired depigmentation, is characterized by well-defined depigmented macules and patches. Other conditions, such as chemical leukoderma, can present with similar findings, and are often easily mistaken for vitiligo. Key clinical features can help differentiate between acquired disorders of depigmentation. The first article in this continuing medical education series focuses on conditions with a vitiligo-like phenotype. Early recognition and adequate treatment of these conditions is critical in providing appropriate prognostication and treatment.

Inhibition of Stearoyl-CoA desaturase 1 reverts BRAF and MEK inhibition-induced selection of cancer stem cells in BRAF-mutated melanoma.

BACKGROUND: Combination therapy with BRAF and MEK inhibitors significantly improves survival in BRAF mutated melanoma patients but is unable to prevent disease recurrence due to the emergence of drug resistance. Cancer stem cells (CSCs) have been involved in these long-term treatment failures. We previously reported in lung cancer that CSCs maintenance is due to altered lipid metabolism and dependent upon Stearoyl-CoA-desaturase (SCD1)-mediated upregulation of YAP and TAZ. On this ground, we investigated the role of SCD1 in melanoma CSCs. METHODS: SCD1 gene expression data of melanoma patients were downloaded from TCGA and correlated with disease progression by bioinformatics analysis and confirmed on patient's tissues by qRT-PCR and IHC analyses. The effects of combination of BRAF/MEKi and the SCD1 inhibitor MF-438 were monitored by spheroid-forming and proliferation assays on a panel of BRAF-mutated melanoma cell lines grown in 3D and 2D conditions, respectively. SCD1, YAP/TAZ and stemness markers were evaluated in melanoma cells and tissues by qRT-PCR, WB and Immunofluorescence. RESULTS: We first observed that SCD1 expression increases during melanoma progression. BRAF-mutated melanoma 3D cultures enriched for CSCs overexpressed SCD1 and were more resistant than 2D differentiated cultures to BRAF and MEK inhibitors. We next showed that exposure of BRAF-mutated melanoma cells to MAPK pathway inhibitors enhanced stemness features by upregulating the expression of YAP/TAZ and downstream genes but surprisingly not SCD1. However, SCD1 pharmacological inhibition was able to downregulate YAP/TAZ and to revert at the same time CSC enrichment and resistance to MAPK inhibitors. CONCLUSIONS: Our data underscore the role of SCD1 as prognostic marker in melanoma and promote the use of SCD1 inhibitors in combination with MAPK inhibitors for the control of drug resistance.

Adipose tissue-derived extracellular fraction characterization: biological and clinical considerations in regenerative medicine.

BACKGROUND: Adipose tissue-derived stem cells are considered to be a promising source in the field of cell therapy and regenerative medicine. In addition to direct cell replacement using adipose tissue or purified stem cells, intercellular molecule exchange by the adipose tissue complex, a vast array of bioactive secretory factors, demonstrated beneficial effects by reducing tissue damage and stimulation of endogenous repair. However, for therapeutic purposes, the use of secretome derivatives, such as full conditioned media or purified exosomes generated in vitro, may present considerable disadvantages for cell manufacturing, storage, product safety, and their potential as a ready-to-go therapeutic product. METHODS: In this study, the effect of a liquid fraction of lipoaspirates isolated intraoperatively from 28 healthy donors was evaluated for their protective effect against oxidative stress and senescence, proliferation, and migration in vitro on normal human melanocytes, keratinocytes, and fibroblasts. Immunoenzymatic quantification of several growth factors and important signal molecules was used to define the biological profile of physiological adipose tissue secretome. RESULTS: Adipose tissue extracellular fraction (AT-Ex), isolated from lipoaspirate, exhibited significant potential for skin repair. AT-Ex augmented dermal and epidermal cell proliferation in a dose-dependent manner without promoting cancer cell growth. Moreover, migration of dermal fibroblasts, an important phenomenon implicated in endogenous repair, was enhanced by AT-Ex treatment. AT-Ex has a positive impact on oxidative stress damage when cells are exposed to extrinsic hostile factors and prevent a fibroblast senescence phenotype including paracrine functions associated with skin aging. CONCLUSIONS: Collectively, our findings propose natural systems carrying the physiological balance of in-vivo produced secretome that could improve cutaneous wound healing and tissue repair. This approach, representing an innovative perspective and therapeutic strategy in regenerative medicine, could also be combined with autologous stem cell grafts to treat chronic nonhealing wounds, stable vitiligo, severe burns, and post-oncological scarring.