The causal relationship between immune cells mediating FIT3L, CCL4, OSM, and skin-derived deteriorated tumors.

OBJECTIVE: Observational studies have identified a dual effect of circulating inflammatory proteins and immune cells on cancer progression. However, the specific mechanisms of action have not been clarified in the exacerbation of cutaneous-origin tumors. Therefore, this study aims to investigate whether the causal relationship between circulating inflammatory factors and basal cell carcinoma (BCC), cutaneous malignant melanoma (SKCM), and cutaneous squamous cell carcinoma (cSCC) is regulated by immune cells. METHODS: This study employed the Two-Sample Mendelian Randomization (TSMR) approach to investigate the causal relationships between 91 circulating inflammatory factors and three prevalent types of skin cancer from a genetic perspective. Bayesian Weighted Mendelian Randomization (BWMR) was also used to validate correlation and reverse MR to assess inverse relationships. Subsequent sensitivity analyses were conducted to limit the impact of heterogeneity and pleiotropy. Finally, the two-step Mendelian Randomization (two-step MR) method was utilized to ascertain the mediating effects of specific immune cell traits in the causal pathways linking circulating inflammatory factors with BCC, SKCM, and cSCC. RESULTS: The Inverse Variance Weighted (IVW) method and the Bayesian Weighted Algorithm collectively identified nine inflammatory factors causally associated with BCC, SKCM, and cSCC. The results from Cochran's Q test, mendelian randomization pleiotropy residual sum and outlier (MR-PRESSO), and MR-Egger intercept were not statistically significant (p < 0.05). Additionally, the proportions mediated by CD4+ CD8dim T cell %leukocyte, CD4-CD8-Natural Killer T %T cell, and CD20 on IgD-CD38-B cell for FIt3L, CCL4, and OSM were 9.26%, 8.96%, and 10.16%, respectively. CONCLUSION: Immune cell levels potentially play a role in the modulation process between circulating inflammatory proteins and cutaneous-origin exacerbated tumors. This finding offers a new perspective for the in-depth exploration of cutaneous malignancies.

Bioluminescent aptamer-based microassay for detection of melanoma inhibitory activity protein (MIA).

Melanoma inhibitory activity protein (MIA) does obviously offer the potential to reveal clinical manifestations of melanoma. Despite a pressing need for effective diagnosis of this highly fatal disease, there are no clinically approved MIA detection ELISA kits available. A recommended MIA threshold has not yet been defined, mostly by reason of variability in immunoglobulins' affinity and stability, the difference in sample preparation and assay conditions. Here we present a pair of high-affinity DNA aptamers developed as an alternative recognition and binding element for MIA detection. Their stability and reproducible synthesis are expected to ensure this analysis under standard conditions. The devised aptamer-based solid-phase microassay of model standard and control human sera involves luciferase NLuc as a highly sensitive reporter. Bioluminescence dependence on MIA concentration ranges in a linear manner from 2.5 to 250 ng mL-1, providing a MIA detection limit of 1.67 ± 0.57 ng mL-1.

Ultrasensitive quantification of PD-L1+ extracellular vesicles in melanoma patient plasma using a parallelized high throughput droplet digital assay.

The expression of programmed death-ligand 1 (PD-L1) on extracellular vesicles (EVs) is an emerging biomarker for cancer, and has gained particular interest for its role mediating immunotherapy. However, precise quantification of PD-L1+ EVs in clinical samples remains challenging due to their sparse concentration and the enormity of the number of background EVs in human plasma, limiting applicability of conventional approaches. In this study, we develop a high-throughput droplet-based extracellular vesicle analysis (DEVA) assay for ultrasensitive quantification of EVs in plasma that are dual positive for both PD-L1 and tetraspanin (CD81) known to be expressed on EVs. We achieve a performance that significantly surpasses conventional approaches, demonstrating 360× enhancement in the limit of detection (LOD) and a 750× improvement in the limit of quantitation (LOQ) compared to conventional plate enzyme-linked immunoassay (ELISA). Underlying this performance is DEVA's high throughput analysis of individual EVs one at a time and the high specificity to targeted EVs versus background. We achieve a 0.006% false positive rate per droplet by leveraging avidity effects that arise from EVs having multiple copies of their target ligands on their surface. We use parallelized optofluidics to rapidly process 10 million droplets per minute, ∼100× greater than conventional approaches. A validation study on a cohort of 14 patients with melanoma confirms DEVA's ability to match conventional ELISA measurements with reduced plasma sample volume and without the need for prior EV purification. This proof-of-concept study demonstrates DEVA's potential for clinical utility to enhance prognosis as well as guide treatment for cancer.

Reconsidering S100B: Why it is time to abandon its use in detecting disease recurrence in stage III melanoma patients.

BACKGROUND: Especially in the era of successful systemic therapy, there is an urgent need to detect early disease recurrence in stage III melanoma patients. This study investigates if serum S100 calcium-binding protein B (S100B) can detect disease recurrence in stage III melanoma patients. METHODS: A retrospective cohort study was conducted at the University Medical Center Groningen (UMCG). Adult AJCC 8th stage III melanoma patients in whom serum S100B was measured as part of follow-up from January 2010 until April 2023 were included. The association between serum S100B and disease recurrence was evaluated using standard definitions for sensitivity and positive predictive value (PPV). RESULTS: Overall, 147 patients were included (mean age was 60.4 years, 53.1 % were female). Most patients were classified as stage IIIB (39, 26.5 %) and IIIC (73, 49.7 %). During median follow-up of 56 months, 69 (46.9 %) patients experienced disease recurrence. Seventeen out of 18 patients with elevated serum S100B (≥0.15 µg/L) experienced disease recurrence (PPV of 94.4 %). However, 52 out of 69 patients with disease recurrence had normal serum S100B (sensitivity of 24.6 %). Eight out of 17 (47.1 %) patients were asymptomatic (P = 0.608), twelve (70.6 %) patients had at least four distant metastases (P < 0.001). CONCLUSION: The clinical value of serum S100B to detect disease recurrence in stage III melanoma patients is negligible since only one out of four patients with disease recurrence have elevated serum S100B. Furthermore, half of stage III melanoma patients with elevated S100B experienced symptoms, and most patients already have multiple distant metastases.

Causal relationship between genetically determined plasma metabolites and skin cancer: a two-sample Mendelian randomization study.

We aimed to unveil the underlying pathogenic mechanisms of skin cancer in relation to metabolic factors and pathway mechanisms. This study utilized the TwoSample Mendelian randomization (MR) method to investigate the causal relationship between 1400 plasma metabolites and skin cancer. The primary method employed was the inverse variance weighting (IVW). Through IVW analysis, we found 105 plasma metabolites associated with Basal Cell Carcinoma (BCC), with the highest association observed for Prolylglycine levels (OR [95% CI]: 1.1902 [1.0274, 1.3788]). For Malignant Melanoma of Skin (MSS), 68 plasma metabolites were linked, with the highest causal relationship seen for 3-Hydroxybutyrate levels (OR [95% CI]: 1.0030 [1.0013, 1.0048]). Regarding actinic keratosis (AK), and the highest association observed for Hexadecadienoate (16:2n6) levels (OR [95% CI]: 1.3302 [1.0333, 1.7125]). Glycerol to palmitoylcarnitine (16: n6) levels (OR [95% CI]: 1.3302 [1.0333, 1.125]) were found to be significant for BCC and AK. Palmitoylcarnitine (C16) had the most positive causal effect for BCC (OR [95% CI]: 1.1777 [1.0493, 1.3218]), while 5-hydroxy-2-methylpyridine sulfate levels had the highest effect for AK (OR [95% CI]: 1.1788 [1.0295, 1.3498]). And 4-guanidinobutanoate levels had the largest positive causal effect (OR [95% CI]: 1.0857 [1.0417, 1.1317]) for BCC, and X-11880 levels for MSS (OR [95% CI]: 1.0013 [1.0000, 1.0025]). The study revealed a positive association between hereditary Glycerol to palmitoylcarnitine (C16) and 5-hydroxy-2-methylpyridine sulfate levels with the risk of developing BCC and AK. Additionally, 4-guanidinobutanoate levels and X 11880 levels were found to be positively associated with the risk of BCC and MMS.

Prognostic Value of the Neutrophil-to-lymphocyte Ratio, Platelet-to- lymphocyte Ratio and Monocyte-to-lymphocyte Ratio in Melanoma Patients: A Cohort Study.

The prognostic value of the neutrophil-lymphocyte ratio, platelet-lymphocyte ratio and monocyte-lymphocyte ratio in patients with melanoma has yielded controversial results in the literature. A retrospective single-centre cohort study was conducted from 1998 to 2020, including patients diagnosed with invasive melanoma. A total of 2,721 patients were included in the study. The median follow-up was 8.23 years (IQR 4.41-13.25). The median baseline neutrophil- lymphocyte ratio, platelet-lymphocyte ratio and monocyte-lymphocyte ratio values increased significantly (p < 0.001) with the increasing American Joint Committee on Cancer stage. The optimal cut-off values for neutrophil-lymphocyte ratio, platelet-lymphocyte ratio and monocyte-lymphocyte ratio were determined as 2.1, 184 and 0.2, respectively. In the multivariate analysis, high levels of neutrophil-lymphocyte ratio (≥ 2.1), platelet-lymphocyte ratio (≥ 184) and monocyte-lymphocyte ratio (≥ 0.2) were independently associated with significantly shorter melanoma-specific survival (neutrophil-lymphocyte ratio: HR 1.30, 95% CI 1.06-1.60, p = 0.013; platelet-lymphocyte ratio: HR 1.37, 95% CI 1.06-1.76, p = 0.014; monocyte- lymphocyte ratio: HR 1.29, 95% CI 1.05-1.58, p = 0.015) and overall survival (neutrophil-lymphocyte ratio: HR 1.39, 95% CI 1.19-1.64, p < 0.001; platelet- lymphocyte ratio: HR 1.44, 95% CI 1.19-1.74, p < 0.001; monocyte-lymphocyte ratio: HR 1.42, 95% CI 1.21-1.66, p < 0.001). High levels of neutrophil- lymphocyte ratio and monocyte-lymphocyte ratio were also associated with poor relapse-free survival, while platelet-lymphocyte ratio was not. In conclusion, baseline neutrophil-lymphocyte ratio, platelet-lymphocyte ratio and monocyte-lymphocyte ratio were identified as independent predictors for the prognosis of melanoma.

Identification of Plasma Lipid Alterations Associated with Melanoma Metastasis.

The aim of this study was to apply a state-of-the-art quantitative lipidomic profiling platform to uncover lipid alterations predictive of melanoma progression. Our study included 151 melanoma patients; of these, 83 were without metastasis and 68 with metastases. Plasma samples were analyzed using a targeted Lipidyzer™ platform, covering 13 lipid classes and over 1100 lipid species. Following quality control filters, 802 lipid species were included in the subsequent analyses. Total plasma lipid contents were significantly reduced in patients with metastasis. Specifically, levels of two out of the thirteen lipid classes (free fatty acids (FFAs) and lactosylceramides (LCERs)) were significantly decreased in patients with metastasis. Three lipids (CE(12:0), FFA(24:1), and TAG47:2-FA16:1) were identified as more effective predictors of melanoma metastasis than the well-known markers LDH and S100B. Furthermore, the predictive value substantially improved upon combining the lipid markers. We observed an increase in the cumulative levels of five lysophosphatidylcholines (LPC(16:0); LPC(18:0); LPC(18:1); LPC(18:2); LPC(20:4)), each individually associated with an elevated risk of lymph node metastasis but not cutaneous or distant metastasis. Additionally, seventeen lipid molecules were linked to patient survival, four of which (CE(12:0), CE(14:0), CE(15:0), SM(14:0)) overlapped with the lipid panel predicting metastasis. This study represents the first comprehensive investigation of the plasma lipidome of melanoma patients to date. Our findings suggest that plasma lipid profiles may serve as important biomarkers for predicting clinical outcomes of melanoma patients, including the presence of metastasis, and may also serve as indicators of patient survival.

Cutaneous Melanoma and 486 Human Blood Metabolites: A Mendelian Randomization Study.

BACKGROUND: Cutaneous melanoma (CM) has long been recognized as a lethal form of cancer. Despite persistent research endeavors, the precise underlying pathological mechanisms remain largely unclear, and the optimal treatment for this patient population remains undetermined. OBJECTIVES: This study aims to examine the causal associations between CM and 486 metabolites. METHODS: A two-sample Mendelian randomization (MR) analysis was conducted to ascertain the causal relationship between blood metabolites and CM. The causality analysis involved the inverse variance weighted (IVW) method, followed by the MR-Egger and weighted median (WM) methods. To increase the robustness of our findings, several sensitivity analyses, including the MR-Egger intercept, Cochran's Q test, and MR-pleiotropy residual sum and outlier (MR-PRESSO), were performed. The robustness of our results was further validated in independent outcome samples followed by a meta-analysis. Additionally, a metabolic pathway analysis was carried out. RESULTS: The two-sample MR analysis yielded a total of 27 metabolites as potential causal metabolites. After incorporating the outcomes of the sensitivity analyses, seven causal metabolites remained. Palmitoylcarnitine (OR 0.9903 95% CI 0.9848-0.9958, p = 0.0005) emerged as the sole metabolite with a significant causality after Bonferroni correction. Furthermore, the reverse MR analysis provided no evidence of reverse causality from CM to the identified metabolites. CONCLUSIONS: This study suggested a causal relationship between seven human blood metabolites and the development of CM, thereby offering novel insights into the underlying mechanisms involved. NO LEVEL ASSIGNED: This journal requires that authors assign a level of evidence to each submission to which Evidence-Based Medicine rankings are applicable. This excludes Review Articles, Book Reviews, and manuscripts that concern Basic Science, Animal Studies, Cadaver Studies, and Experimental Studies. For a full description of these Evidence-Based Medicine ratings, please refer to the Table of Contents or the online Instructions to Authors www.springer.com/00266 .

Soluble PD-L1 predicts tumor response and immune-related adverse events in patients with advanced melanoma treated with anti-PD-1 antibodies.

Immune checkpoint inhibitors (ICIs) bring prognostic benefits to patients with malignancies. However, there is a substantial number of patients whose lesions are not improved by ICIs. In addition, ICIs may cause immune-related adverse events (irAEs), which could lead to an unfavorable prognosis with fatal consequences. Therefore, we conducted a retrospective study to evaluate the utility of circulating sPD-L1 (soluble programmed cell death 1 ligand 1) as a biomarker in patients with advanced melanoma treated with anti-PD-1 (programmed cell death 1 protein) antibodies. Sera from 31 consecutive patients were prospectively collected before and after anti-PD-1 antibody treatment and the serum level of sPD-L1 was evaluated. We found that high sPD-L1 levels before treatment were associated with better prognosis, and this association was observed only in patients with a low tumor burden. We also found that sPD-L1 levels were elevated in patients who developed severe irAEs after treatment, and the patients with severe irAEs had significantly higher fluctuations in sPD-L1 (delta sPD-L1) than those without severe irAEs. Our study suggests that serum sPD-L1 level is a useful biomarker to predict tumor response and irAE development in patients with advanced melanoma treated with anti-PD-1 antibodies.

Nodular type but not vitamin D levels increases the risk of second primary cancers in melanoma patients: An observational study of 663 patients / El melanoma de tipo nodular pero no los niveles de vitamina D aumenta el riesgo de desarrollar una segunda neoplasia no cutánea en pacientes con un melanoma: estudio observacional de 663 pacientes

Background Vitamin D deficiency associates with the risk of developing many diseases, including cancer. At the molecular level, vitamin D appears to have an antineoplastic effect. However, the role of vitamin D deficiency in cancer pathogenesis remains unelucidated and numerous studies have resulted in discordant results. This study aimed to determine whether vitamin D deficiency during melanoma diagnosis increases the risk of developing non-cutaneous second primary cancers (SPC). Materials and methods A retrospective study on 663 patients diagnosed with melanoma between 1 January 2011 and 31 October 2022. The effect of each variable on the development of a subsequent non-cutaneous cancer was performed using Kaplan–Meier curves and differences were assessed by log-rank tests. Cox proportional hazard univariate and multivariate models were used to quantify the effect of each variable in the time to develop a non-cutaneous neoplasia. Results Out of 663 patients, 34 developed a non-cutaneous SPC. There was no statistically significant association between vitamin D levels and non-cutaneous SPC development (log-rank, p=0.761). Age>60 years, stage III/IV, and nodular melanoma subtype were significantly associated with the development of a SPC. After multivariate analysis, only age>60 years (HR 3.4; HR CI 95%: 1.5–7.6) and nodular melanoma subtype (HR 2.2; HR CI 95%: 1.0–4.8) were included in the final model. Conclusions Our results suggest that vitamin D deficiency is not associated with an increased risk of developing non-cutaneous SPC in melanoma patients. However, age over 60 years and nodular melanoma subtype increase the risk for non-cutaneous SPC development (AU)

Antecedentes El déficit de vitamina D se asocia con un mayor riesgo de padecer varias enfermedades, incluido el cáncer. Molecularmente, esta parece tener un efecto antineoplásico. Sin embargo, el papel que juega en la patogénesis del cáncer no está bien esclarecido y hay resultados dispares en los estudios publicados. El objetivo del presente fue determinar si unos niveles de vitamina D deficientes en el momento del diagnóstico del melanoma aumentaba el riesgo de desarrollar un cáncer no cutáneo (CNC). Material y método Se diseñó un estudio retrospectivo de 663 pacientes diagnosticados de melanoma entre el 1 de enero de 2011 y el 31 de octubre de 2022. El efecto de cada una de las variables seleccionadas en el desarrollo de un CNC durante el seguimiento tras el diagnóstico del melanoma se realizó mediante el estudio de supervivencia con el método de Kaplan-Meier y las diferencias se evaluaron con la prueba de los rangos logarítmicos. Se elaboraron modelos uni y multivariados de riesgos proporcionales de Cox para cuantificar el efecto de cada valor de las variables de estudio en el tiempo para desarrollar un CNC. Resultados De los 663 pacientes, 34 desarrollaron un CNC tras el melanoma. No hubo diferencias estadísticamente significativas entre los grupos definidos por los niveles de vitamina D (log-rank, p = 0,761). Sin embargo, una edad > 60, el estadio III/IV, y el tipo nodular se asociaron significativamente al desarrollo de un CNC. Tras el análisis multivariado, solo la edad > 60 (hazard ratio [HR] 3,4; intervalo de confianza [IC] 95% HR:1,5-7,6) y el subtipo nodular de melanoma (HR 2,2; IC 95% HR:1,0-4,8) se mantuvieron en el modelo predictivo final. Conclusiones Nuestros resultados sugieren que unos niveles de vitamina D deficientes en el diagnóstico de melanoma no se asocian a un mayor riesgo de desarrollar un CNC. Sin embargo, en una edad > 60 y el subtipo nodular sí que aumentan el riesgo de desarrollar un CNC (AU)

[Artículo traducido] El melanoma de tipo nodular pero no los niveles de vitamina D aumenta el riesgo de desarrollar una segunda neoplasia no cutánea en pacientes con un melanoma: estudio observacional de 663 pacientes / Nodular Type but Not Vitamin D Levels Increases the Risk of Second Primary Cancers in Melanoma Patients: An Observational Study of 663 Patients

Antecedentes El déficit de vitamina D se asocia con un mayor riesgo de padecer varias enfermedades, incluido el cáncer. Molecularmente, esta parece tener un efecto antineoplásico. Sin embargo, el papel que juega en la patogénesis del cáncer no está bien esclarecido y hay resultados dispares en los estudios publicados. El objetivo del presente fue determinar si unos niveles de vitamina D deficientes en el momento del diagnóstico del melanoma aumentaba el riesgo de desarrollar un cáncer no cutáneo (CNC). Material y método Se diseñó un estudio retrospectivo de 663 pacientes diagnosticados de melanoma entre el 1 de enero de 2011 y el 31 de octubre de 2022. El efecto de cada una de las variables seleccionadas en el desarrollo de un CNC durante el seguimiento tras el diagnóstico del melanoma se realizó mediante el estudio de supervivencia con el método de Kaplan-Meier y las diferencias se evaluaron con la prueba de los rangos logarítmicos. Se elaboraron modelos uni y multivariados de riesgos proporcionales de Cox para cuantificar el efecto de cada valor de las variables de estudio en el tiempo para desarrollar un CNC. Resultados De los 663 pacientes, 34 desarrollaron un CNC tras el melanoma. No hubo diferencias estadísticamente significativas entre los grupos definidos por los niveles de vitamina D (log-rank, p = 0,761). Sin embargo, una edad > 60, el estadio III/IV, y el tipo nodular se asociaron significativamente al desarrollo de un CNC. Tras el análisis multivariado, solo la edad > 60 (hazard ratio [HR] 3,4; intervalo de confianza [IC] 95% HR:1,5-7,6) y el subtipo nodular de melanoma (HR 2,2; IC 95% HR:1,0-4,8) se mantuvieron en el modelo predictivo final. Conclusiones Nuestros resultados sugieren que unos niveles de vitamina D deficientes en el diagnóstico de melanoma no se asocian a un mayor riesgo de desarrollar un CNC. Sin embargo, en una edad > 60 y el subtipo nodular sí que aumentan el riesgo de desarrollar un CNC (AU)

Background Vitamin D deficiency associates with the risk of developing many diseases, including cancer. At the molecular level, vitamin D appears to have an antineoplastic effect. However, the role of vitamin D deficiency in cancer pathogenesis remains unelucidated and numerous studies have resulted in discordant results. This study aimed to determine whether vitamin D deficiency during melanoma diagnosis increases the risk of developing non-cutaneous second primary cancers (SPC). Materials and methods A retrospective study on 663 patients diagnosed with melanoma between 1 January 2011 and 31 October 2022. The effect of each variable on the development of a subsequent non-cutaneous cancer was performed using Kaplan–Meier curves and differences were assessed by log-rank tests. Cox proportional hazard univariate and multivariate models were used to quantify the effect of each variable in the time to develop a non-cutaneous neoplasia. Results Out of 663 patients, 34 developed a non-cutaneous SPC. There was no statistically significant association between vitamin D levels and non-cutaneous SPC development (log-rank, p=0.761). Age>60 years, stage III/IV, and nodular melanoma subtype were significantly associated with the development of a SPC. After multivariate analysis, only age>60 years (HR 3.4; HR CI 95%: 1.5–7.6) and nodular melanoma subtype (HR 2.2; HR CI 95%: 1.0–4.8) were included in the final model. Conclusions Our results suggest that vitamin D deficiency is not associated with an increased risk of developing non-cutaneous SPC in melanoma patients. However, age over 60 years and nodular melanoma subtype increase the risk for non-cutaneous SPC development (AU)

Efecto de los niveles séricos de vitamina D en el pronóstico del melanoma cutáneo / Prognostic Value of Vitamin D Serum Levels in Cutaneous Melanoma

Introducción: La vitamina D tiene un rol fundamental en múltiples vías metabólicas, incluidas vías implicadas en la proliferación celular y la respuesta inmune. Sus niveles han mostrado una asociación con el riesgo de desarrollar el melanoma cutáneo y su pronóstico. El objetivo de este estudio fue evaluar si los niveles séricos de vitamina D influyen en el pronóstico del melanoma. Materiales y métodos: Estudio de cohorte retrospectivo, observacional, longitudinal y analítico en 286 pacientes con diagnóstico histológico de melanoma, en los que se midieron los niveles séricos de vitamina D en el momento del diagnóstico. Se analizó la relación entre los niveles de vitamina D y las características epidemiológicas, clínicas y patológicas de los pacientes, y el efecto de la vitamina D en la supervivencia global de los pacientes. Mediante un bucle iterativo se encontró el punto de corte de los niveles séricos de vitamina D de 9,25ng/mL para su relación con la supervivencia. Resultados: Un nivel bajo de vitamina D (<9,25ng/mL) se relacionó con la ulceración en el análisis histológico. Tras una mediana de seguimiento de 39,4 meses, 24 pacientes (8,4%) fallecieron. Unos niveles de vitamina D<9,25ng/mL se asociaron con una menor supervivencia global, tanto en el análisis a través de curvas de Kaplan-Meier, como tras la regresión de Cox multivariada. Conclusión: Los niveles<9,25ng/mL de vitamina D se asocian a la presencia de ulceración histológica en el melanoma y son un factor pronóstico independiente para la supervivencia global en estos pacientes (AU)

Introduction: Vitamin D plays a fundamental role in many metabolic pathways, including those involved in cell proliferation and the immune response. Serum levels of this vitamin have been linked to melanoma risk and prognosis. This study aimed to assess the prognostic value of vitamin D serum level in melanoma. Material and methods: Retrospective, observational, longitudinal, and analytical study of 286 patients with a histologic diagnosis of melanoma in whom serum levels of vitamin D were measured at the time of diagnosis. We analyzed associations between serum level and epidemiologic and clinical variables and pathology findings; we also analyzed the influence of vitamin D on overall survival. An iterative loop was used to identify a vitamin D serum level to test for its an association with survival. Results: A vitamin D level less than 9.25ng/mL was associated with a histologic finding of ulceration. After a median follow-up period of 39.4 months, 24 patients (8.4%) had died. The cutoff of 9.25ng/mL was associated with lower overall survival according to both the Kaplan-Meier curves and multivariate Cox regression analysis. Conclusion: Vitamin D levels less than 9.25ng/mL are associated with ulceration in melanoma and serve as an independent prognostic factor for overall survival in this disease (AU)

[Translated article] Prognostic Value of Vitamin D Serum Levels in Cutaneous Melanoma / Efecto de los niveles séricos de vitamina D en el pronóstico del melanoma cutáneo

Introduction: Vitamin D plays a fundamental role in many metabolic pathways, including those involved in cell proliferation and the immune response. Serum levels of this vitamin have been linked to melanoma risk and prognosis. This study aimed to assess the prognostic value of vitamin D serum level in melanoma. Material and methods: Retrospective, observational, longitudinal, and analytical study of 286 patients with a histologic diagnosis of melanoma in whom serum levels of vitamin D were measured at the time of diagnosis. We analyzed associations between serum level and epidemiologic and clinical variables and pathology findings; we also analyzed the influence of vitamin D on overall survival. An iterative loop was used to identify a vitamin D serum level to test for its an association with survival. Results: A vitamin D level less than 9.25ng/mL was associated with a histologic finding of ulceration. After a median follow-up period of 39.4 months, 24 patients (8.4%) had died. The cutoff of 9.25ng/mL was associated with lower overall survival according to both the Kaplan-Meier curves and multivariate Cox regression analysis. Conclusion: Vitamin D levels less than 9.25ng/mL are associated with ulceration in melanoma and serve as an independent prognostic factor for overall survival in this disease (AU)

Introducción: La vitamina D tiene un rol fundamental en múltiples vías metabólicas, incluidas vías implicadas en la proliferación celular y la respuesta inmune. Sus niveles han mostrado una asociación con el riesgo de desarrollar el melanoma cutáneo y su pronóstico. El objetivo de este estudio fue evaluar si los niveles séricos de vitamina D influyen en el pronóstico del melanoma. Materiales y métodos: Estudio de cohorte retrospectivo, observacional, longitudinal y analítico en 286 pacientes con diagnóstico histológico de melanoma, en los que se midieron los niveles séricos de vitamina D en el momento del diagnóstico. Se analizó la relación entre los niveles de vitamina D y las características epidemiológicas, clínicas y patológicas de los pacientes, y el efecto de la vitamina D en la supervivencia global de los pacientes. Mediante un bucle iterativo se encontró el punto de corte de los niveles séricos de vitamina D de 9,25ng/mL para su relación con la supervivencia. Resultados: Un nivel bajo de vitamina D (<9,25ng/mL) se relacionó con la ulceración en el análisis histológico. Tras una mediana de seguimiento de 39,4 meses, 24 pacientes (8,4%) fallecieron. Unos niveles de vitamina D<9,25ng/mL se asociaron con una menor supervivencia global, tanto en el análisis a través de curvas de Kaplan-Meier, como tras la regresión de Cox multivariada. Conclusión: Los niveles<9,25ng/mL de vitamina D se asocian a la presencia de ulceración histológica en el melanoma y son un factor pronóstico independiente para la supervivencia global en estos pacientes (AU)

sIL-2R- an Immuno-biomarker for Prediction of Metastases in Uveal Melanoma.

BACKGROUND/AIM: Uveal melanoma (UM) is the most common primary intraocular malignancy in adults. High serum levels of soluble IL-2 receptor (sIL-2R) have been reported in acute inflammations and metastatic cancers. This study evaluated the potential of high/increasing sIL-2R levels in predicting metastases. PATIENTS AND METHODS: The study included a total of 1,546 sera samples of subjects from three groups: 119 healthy controls (73 subjects), 566 UM 10 year (10y) disease-free (DF) (220 patients), 861 metastatic UM (268 patients). Patients were followed-up biannually with liver ultrasound and liver function tests for the presence of metastases (Mets). Blood samples to measure the levels of sIL-2R were obtained at the time of primary diagnosis, soon after initial treatment (enucleation, brachytherapy), every 6 months, 10 years from diagnosis, at Mets confirmation by CT, and after additional treatments. RESULTS: Significantly higher sIL-2R levels were detected in the Mets patients compared to healthy controls and 10y DF patients. Compared to the upper limit of the normal levels of sIL-2R, 1,000 U/ml, its levels in metastatic UM were 61%, 25% in 10y DF UM, and 6.25% in the controls. High levels of sIL-2R in metastatic patients, decreased significantly post treatments. Individual kinetics of markers, indicated similar trends of sIL-2R compared to osteopontin and S-Protein 100, predicting metastases, which were confirmed on liver imaging. CONCLUSION: Significantly higher sIL-2R levels were evident in all UM patients with Mets. Significant increases in sIL-2R levels on serial evaluations indicated and predicted UM Mets, enabling earlier treatment of Mets, to improve survival.

Therapeutic Response Evaluation in Advanced Melanoma Patients Incorporating Plasma cfDNA, LDH, VEGF, PD-L1, and IFN-γ Measurements.

BACKGROUND/AIM: Current treatment strategies for advanced melanoma require serial assessment of disease status in affected patients. In this study, we sought to examine the relationship between radiographic tumour burden and blood borne biomarkers including plasma cfDNA, serum LDH, plasma VEGF, PD-L1 and IFN-γ in advanced melanoma patients receiving immunotherapy. We hypothesized that a combination of these explanatory variables in a suitable regression analysis model may predict changes in tumour burden during patient treatment. MATERIALS AND METHODS: We extracted and quantified circulating cfDNA, LDH, VEGF, PD-L1, and IFN-γ from thirty patients with stage IV melanoma at baseline and at six months. All participating patients were evaluated with paired blood sample collection and CT scan assessments during treatment. RESULTS: Changes in radiographic tumour burden correlated with changes in levels of cfDNA (p≤0.001), LDH (p≤0.001), VEGF (p≤0.001), and PD-L1 (p<0.05) during treatment. Multiple regression analysis consisting of the follow-up to baseline assessment ratios of cfDNA, LDH, VEGF and PD-L1 explained changes in tumour burden (F (4, 23)=32.05, p<0.001); with an R2 of 0.8479 (Y=ß0+ß1*B+ß2*C+ß3*D+ß4*E). CONCLUSION: A quantitative measure of cfDNA, LDH, VEGF and PD-L1 may complement current methods of assessing tumour burden in advanced melanoma patients.

Detection of clinical progression through plasma ctDNA in metastatic melanoma patients: a comparison to radiological progression.

BACKGROUND: The validity of circulating tumour DNA (ctDNA) as an indicator of disease progression compared to medical imaging in patients with metastatic melanoma requires detailed evaluation. METHODS: Here, we carried out a retrospective ctDNA analysis of 108 plasma samples collected at the time of disease progression. We also analysed a validation cohort of 66 metastatic melanoma patients monitored prospectively after response to systemic therapy. RESULTS: ctDNA was detected in 62% of patients at the time of disease progression. For 67 patients that responded to treatment, the mean ctDNA level at progressive disease was significantly higher than at the time of response (P < 0.0001). However, only 30 of these 67 (45%) patients had a statistically significant increase in ctDNA by Poisson test. A validation cohort of 66 metastatic melanoma patients monitored prospectively indicated a 56% detection rate of ctDNA at progression, with only two cases showing increased ctDNA prior to radiological progression. Finally, a correlation between ctDNA levels and metabolic tumour burden was only observed in treatment naïve patients but not at the time of progression in a subgroup of patients failing BRAF inhibition (N = 15). CONCLUSIONS: These results highlight the low efficacy of ctDNA to detect disease progression in melanoma when compared mainly to standard positron emission tomography imaging.

Naturally Occurring Telomerase-Specific CD4 T-Cell Immunity in Melanoma.

CD4 T cells play a key role in anticancer immunity. In this study, we investigate the clinical relevance of circulating CD4 T helper type 1 (Th1) response against telomerase (anti-TERT Th1 response) in patients with melanoma. The spontaneous anti-TERT Th1 response was detected in 54.5% (85/156) of patients with melanoma before treatment. The prevalence of this systemic response was inversely related to Breslow thickness >1 mm and American Joint Committee on Cancer stage ≥II (P = 0.001 and 0.032, respectively). In contrast to patients treated with targeted therapies, the anti-TERT Th1 immunity was associated with an objective response after immune checkpoint inhibitors treatment. Hence, 86% (18/21) of responder patients exhibited pre-existing anti-TERT Th1 versus 35% (6/19) in nonresponders (P = 0.001). This response was also associated with increased progression-free survival and overall survival in patients with melanoma treated with immune checkpoint inhibitors (P = 0.0008 and 0.012, respectively). Collectively, the presence of circulating anti-TERT Th1 response is inversely related to melanoma evolution and appears to be a predictive factor of response to immunotherapy. Our results highlight the interest in telomerase-specific CD4 Th1 response as a promising blood-based biomarker of immune checkpoint inhibitors therapy in melanoma.

MCAM/MUC18/CD146 as a Multifaceted Warning Marker of Melanoma Progression in Liquid Biopsy.

Human malignant melanoma shows a high rate of mortality after metastasization, and its incidence is continuously rising worldwide. Several studies have suggested that MCAM/MUC18/CD146 plays an important role in the progression of this malignant disease. MCAM/MUC18/CD146 is a typical single-spanning transmembrane glycoprotein, existing as two membrane isoforms, long and short, and an additional soluble form, sCD146. We previously documented that molecular MCAM/MUC18/CD146 expression is strongly associated with disease progression. Recently, we showed that MCAM/MUC18/CD146 and ABCB5 can serve as melanoma-specific-targets in the selection of highly primitive circulating melanoma cells, and constitute putative proteins associated with disease spreading progression. Here, we analyzed CD146 molecular expression at onset or at disease recurrence in an enlarged melanoma case series. For some patients, we also performed the time courses of molecular monitoring. Moreover, we explored the role of soluble CD146 in different cohorts of melanoma patients at onset or disease progression, rather than in clinical remission, undergoing immune therapy or free from any clinical treatment. We showed that MCAM/MUC18/CD146 can be considered as: (1) a membrane antigen suitable for identification and enrichment in melanoma liquid biopsy; (2) a highly effective molecular "warning" marker for minimal residual disease monitoring; and (3) a soluble protein index of inflammation and putative response to therapeutic treatments.

Altered Glycosylation of Human Alpha-1-Acid Glycoprotein as a Biomarker for Malignant Melanoma.

A high-resolution HILIC-MS/MS method was developed to analyze anthranilic acid derivatives of N-glycans released from human serum alpha-1-acid glycoprotein (AGP). The method was applied to samples obtained from 18 patients suffering from high-risk malignant melanoma as well as 19 healthy individuals. It enabled the identification of 102 glycan isomers separating isomers that differ only in sialic acid linkage (α-2,3, α-2,6) or in fucose positions (core, antenna). Comparative assessment of the samples revealed that upregulation of certain fucosylated glycans and downregulation of their nonfucosylated counterparts occurred in cancer patients. An increased ratio of isomers with more α-2,6-linked sialic acids was also observed. Linear discriminant analysis (LDA) combining 10 variables with the highest discriminatory power was employed to categorize the samples based on their glycosylation pattern. The performance of the method was tested by cross-validation, resulting in an overall classification success rate of 96.7%. The approach presented here is significantly superior to serological marker S100B protein in terms of sensitivity and negative predictive power in the population studied. Therefore, it may effectively support the diagnosis of malignant melanoma as a biomarker.