Association of miR-146a-5p and miR-21-5p with Prognostic Features in Melanomas.

BACKGROUND: Cutaneous melanoma (CM) is one of the most lethal tumors among skin cancers and its incidence is rising worldwide. Recent data support the role of microRNAs (miRNAs) in melanoma carcinogenesis and their potential use as disease biomarkers. METHODS: We quantified the expression of miR-146a-5p and miR-21-5p in 170 formalin-fixed paraffin embedded (FFPE) samples of CM, namely 116 superficial spreading melanoma (SSM), 26 nodular melanoma (NM), and 28 lentigo maligna melanoma (LMM). We correlated miRNA expression with specific histopathologic features including Breslow thickness (BT), histological subtype, ulceration and regression status, and mitotic index. RESULTS: miR-146a-5p and miR-21-5p were significantly higher in NM compared to SSM and LMM. The positive correlation between miR-146a-5p and miR-21-5p expression and BT was confirmed for both miRNAs in SSM. Considering the ulceration status, we assessed that individual miR-21-5p expression was significantly higher in ulcerated CMs. The increased combined expression of the two miRNAs was strongly associated with ulceration (p = 0.0093) and higher mitotic rate (≥1/mm2) (p = 0.0005). We demonstrated that the combination of two-miRNA expression and prognostic features (BT and ulceration) can better differentiate cutaneous melanoma prognostic groups, considering overall survival and time-to-relapse clinical outcomes. Specifically, miRNA expression can further stratify prognostic groups among patients with BT ≥ 0.8 mm but without ulceration. Our findings provide further insights into the characterization of CM with specific prognostic features. The graphical abstract was created with BioRender.com.

CAR+ extracellular vesicles predict ICANS in patients with B cell lymphomas treated with CD19-directed CAR T cells.

BACKGROUND: Predicting Immune-effector Cell Associated Neurotoxicity Syndrome (ICANS) in patients infused with Chimeric Antigen Receptor T cells (CAR-T) is still a conundrum. This complication, thought to be consequent to CAR-T cell activation, arises a few days after infusion, when circulating CAR-T cells are scarce and specific CAR-T cell-derived biomarkers are lacking. METHODS: Human CD19.CAR-T cells were generated to gain insight into CAR+ extracellular vesicle (CAR+EV) release upon target engagement. A prospective cohort of 100 B-cell lymphoma patients infused with approved CD19.CAR-T cell products (axi-cel, brexu-cel and tisa-cel) was assessed for plasma CAR+EVs as potential biomarkers of in vivo CD19.CAR-T cell activation and predictors of ICANS. Human induced pluripotent stem cells (iPSCs)-derived neural cells were used as a model for CAR+EV-induced neurotoxicity. RESULTS: In vitro, exosome-like CAR+EVs were released by CD19.CAR-T cells upon target engagement. In vivo, CAR+EVs were detectable as early as 1 hour in the plasma of patients. A concentration > 132.8 CAR+EVs/µl at hour +1 or > 224.5 CAR+EVs/µl at day +1 predicted ICANS in advance of 4 days, with a sensitivity up to 96.55% and a specificity up to 80.36%, outperforming other potential ICANS predictors. Enolase 2 (ENO2+) nanoparticles were released by iPSCs-derived neural cells upon CAR+EVs exposure and were increased in the plasma of ICANS patients. CONCLUSIONS: These results convey that plasma CAR+EVs are an immediate signal of CD19.CAR-T cell activation, are suitable predictors of neurotoxicity, and may be involved in ICANS pathogenesis. TRIAL REGISTRATION: NCT04892433, NCT05807789.

Circulating microRNA biomarkers in melanoma and non-melanoma skin cancer.

INTRODUCTION: Skin cancer is the most common type of cancer and is classified in melanoma and non-melanoma cancers, which include basal cell, squamous cell, and Merkel cell carcinoma. Specific microRNAs are dysregulated in each skin cancer type. MicroRNAs act as oncogene or tumor suppressor gene regulators and are actively released from tumor cells in the circulation. Cell-free microRNAs serve many, and possibly yet unexplored, functional roles, but their presence and abundance in the blood has been investigated as disease biomarker. Indeed, specific microRNAs can be isolated and quantified in the blood, usually in serum or plasma fractions, where they are uncommonly stable. MicroRNA levels reflect underlying conditions and have been associated with skin cancer presence, stage, evolution, or therapy efficacy. AREAS COVERED: In this review, we summarize the state of the art on circulating microRNAs detectable in skin cancer patients including all the studies that performed microRNA identification and quantification in the circulation using appropriate sample size and statistics and providing detailed methodology, with a specific focus on diagnostic and prognostic biomarkers. EXPERT OPINION: Circulating microRNAs display a relevant biomarker potential. We expect the development of methodological guidelines and standardized protocols for circulating miRNA quantification in clinical settings.

Peripheral blood cellular profile at pre-lymphodepletion is associated with CD19-targeted CAR-T cell-associated neurotoxicity.

Background: Infusion of second generation autologous CD19-targeted chimeric antigen receptor (CAR) T cells in patients with R/R relapsed/refractory B-cell lymphoma (BCL) is affected by inflammatory complications, such as Immune Effector Cell-Associated Neurotoxicity Syndrome (ICANS). Current literature suggests that the immune profile prior to CAR-T infusion modifies the chance to develop ICANS. Methods: This is a monocenter prospective study on 53 patients receiving approved CAR T-cell products (29 axi-cel, 24 tisa-cel) for R/R-BCL. Clinical, biochemical, and hematological variables were analyzed at the time of pre-lymphodepletion (pre-LD). In a subset of 21 patients whose fresh peripheral blood sample was available, we performed cytofluorimetric analysis of leukocytes and extracellular vesicles (EVs). Moreover, we assessed a panel of soluble plasma biomarkers (IL-6/IL-10/GDF-15/IL-15/CXCL9/NfL) and microRNAs (miR-146a-5p, miR-21-5p, miR-126-3p, miR-150-5p) which are associated with senescence and inflammation. Results: Multivariate analysis at the pre-LD time-point in the entire cohort (n=53) showed that a lower percentage of CD3+CD8+ lymphocytes (38.6% vs 46.8%, OR=0.937 [95% CI: 0.882-0.996], p=0.035) and higher levels of serum C-reactive protein (CRP, 4.52 mg/dl vs 1.00 mg/dl, OR=7.133 [95% CI: 1.796-28], p=0.005) are associated with ICANS. In the pre-LD samples of 21 patients, a significant increase in the percentage of CD8+CD45RA+CD57+ senescent cells (median % value: 16.50% vs 9.10%, p=0.009) and monocytic-myeloid derived suppressor cells (M-MDSC, median % value: 4.4 vs 1.8, p=0.020) was found in ICANS patients. These latter also showed increased levels of EVs carrying CD14+ and CD45+ myeloid markers, of the myeloid chemokine CXCL-9, as well of the MDSC-secreted cytokine IL-10. Notably, the serum levels of circulating neurofilament light chain, a marker of neuroaxonal injury, were positively correlated with the levels of senescent CD8+ T cells, M-MDSC, IL-10 and CXCL-9. No variation in the levels of the selected miRNAs was observed between ICANS and no-ICANS patients. Discussion: Our data support the notion that pre-CAR-T systemic inflammation is associated with ICANS. Higher proportion of senescence CD8+ T cells and M-MDSC correlate with early signs of neuroaxonal injury at pre-LD time-point, suggesting that ICANS may be the final event of a process that begins before CAR-T infusion, consequence to patient clinical history.