Melanoma Metastasis Research Articles

Abstract PR001: The CTC RPL/RPS gene signature: Targeting translation and the cell cycle inversely affects CTC metabolism but not metastasis

Abstract Melanoma brain metastasis (MBM) is the most devastating consequence of melanoma, is increasing in frequency, and is linked to poor prognosis and dismal survival. By employing an unbiased, multipronged approach, we discovered a CTC gene signature for ribosomal protein large/small subunits (RPL/RPS) which associated with MBM onset and progression. Experimental strategies involved capturing, transcriptional profiling, and interrogating CTCs, either directly isolated from blood of melanoma patients at distinct stages of progression, from CTC-driven MBM in experimental animals, or by developing/applying the first MRI CTC- derived MBM xenograft model (MRI-MBM CDX) to discriminate MBM spatial and temporal growth, thus faithfully recreating its clinical presentation. Accordingly, we hypothesized that targeting CTC ribogenesis can prevent melanoma metastasis in general, MBM in particular. We treated CDX cohorts with FDA-approved protein translation inhibitor omacetaxine in the presence or absence of CDK4/CDK6 inhibitor palbociclib, and monitored metastatic development and cell proliferation. Necropsies and IVIS imaging showed decreased MBM/extracranial metastasis in drug-treated mice, and RNA-Seq on mouse-blood-derived CTCs revealed downregulation of four RPL/RPS genes of the CTC signature. Notably, by performing the first-ever functional metabolic characterization of circulating neoplastic cells/CTCs in real- time, coupled with confirmatory RT-qPCR, we found that omacetaxine and palbociclib inversely affected glycolytic metabolism, and demonstrated that dual targeting of cell translation and proliferation is critical to suppress plasticity in metastasis-competent CTCs. This study identifies the relevance of targeting the CTC RPL/RPS signature to suppress melanoma metastasis, and set forward notions for novel therapies to affect metastasis by the deregulation of translation controlling CTC states. Studies involving structural and mechanistic analyses of CTC dormant vs active ribosomes by cryogenic electron microscopy are in progress. Citation Format: Dario Marchetti. The CTC RPL/RPS gene signature: Targeting translation and the cell cycle inversely affects CTC metabolism but not metastasis [abstract]. In: Proceedings of the AACR Special Conference: Liquid Biopsy: From Discovery to Clinical Implementation; 2024 Nov 13-16; San Diego, CA. Philadelphia (PA): AACR; Clin Cancer Res 2024;30(21_Suppl):Abstract nr PR001.

Robot-assisted laparoscopic hepatectomy for liver metastasis from clitoral malignant melanoma: a case report

Abstract Introduction Malignant melanomas occur most commonly in the skin, mucous membranes, or choroid. Clitoral malignant melanomas are extremely rare. Stage IV malignant melanomas have a poor prognosis, and molecularly targeted agents or immune checkpoint inhibitors are recommended. However, surgical resection is reportedly a valid option for improving the prognosis of patients with oligometastases, defined as a small number of metastases that can be completely resected. In this report, we describe hepatic resection for a recurrent liver metastasis in a patient who had undergone removal of a clitoral malignant melanoma 9 years previously. Case presentation An 82 year-old woman presented with a black nodule on her clitoris. Total resection of the nodule resulted in a diagnosis of clitoral malignant melanoma (pT4bN0M0, pStage IIC; UICC 8th edition). A follow-up computed tomography scan 4 years later revealed a single 5 mm mass in the lower lobe of the right lung, prompting partial resection of the right lung. Pathological examination of the operative specimen revealed a pulmonary metastasis of malignant melanoma. The patient was treated with pembrolizumab monotherapy as adjuvant chemotherapy for 1 year. A follow-up computed tomography scan 9 years after surgical removal of the primary lesion revealed an 18 mm mass in segment II of the liver, prompting robot-assisted laparoscopic left lateral sectionectomy. The provisional diagnosis of metastatic malignant melanoma in the liver was confirmed by histopathological examination of the operative specimen. The patient was treated with pembrolizumab monotherapy as postoperative adjuvant chemotherapy for 1 year. No further recurrence was detected at the 1.5 year follow-up. Conclusion We performed hepatectomy for oligometastasis of clitoral malignant melanoma, an extremely rare entity. Surgery has the potential to prolong the prognosis of patients with oligometastasis.

CSIG-11. TUMOR-CELL INTRINSIC CALCIUM OSCILLATIONS DRIVE BRAIN METASTASIS

Abstract Calcium is a ubiquitous secondary messenger which regulates many cellular processes. Recently, we have shown that a subpopulation of glioblastoma cells can intrinsically generate calcium oscillations which drive proliferative signalling. Here, we set out to investigate whether such cancer cell autonomous oscillations are also present in secondary brain tumours. Employing longitudinal intravital two-photon microscopy in awake mice, we demonstrate that brain metastases of melanoma and lung cancer exhibit spontaneous calcium oscillations in vivo along the metastatic cascade. These oscillations are already observed during intravascular arrest and in in vitro monocultures, suggesting a tumour-cell intrinsic origin. We developed a fully automatic image analysis pipeline to accurately quantify calcium oscillations both in vivo and in vitro. Mathematical modelling of cytosolic and ER calcium stores can predict stable calcium oscillations with similar frequency and peak characteristics as those experimentally observed. We discovered that these oscillations are tightly regulated by an interplay of calcium-induced calcium release and store-operated calcium entry (SOCE). Similarly, SOCE-related ion channels are enriched in tissue from human melanoma brain metastases compared to healthy brain controls and calcium activity can also be observed ex vivo in patient-derived brain metastasis resections. Finally, by correlating calcium activity and EdU incorporation we discovered that the presence of calcium oscillations predicts their proliferative potential in vitro. Importantly, by intravital two-photon microscopy we show that calcium activity correlates with metastasis size in preclinical models, suggesting a potential new therapeutic angle from which to target brain metastases.

CNSC-27. INTRATUMORAL CANCER CELL NETWORKS IN BRAIN METASTASES

Abstract Recently, we have shown that primary brain tumors form highly organized, small-world scale-free networks, reminiscent of early networks in neurodevelopment and highly resistant to therapy. Such putative cancer-cell intrinsic neural mechanisms are becoming an increasing focus in the field of Cancer Neuroscience, but it has not yet been shown whether tumors of non-CNS origin also have the ability to form multicellular networks which sustain proliferative signaling and increase resistance to therapy. We observed synchronized calcium activity using longitudinal microscopy both in vivo and in vitro that were reminiscent of gap junction coupled networks. Indeed, dye transfer experiments verified functional coupling of brain metastases cells in vitro. Similarly, inhibition of gap junctions with different pharmacological agents significantly reduced calcium oscillations and tumour proliferation in vitro. Furthermore, gap junctions are significantly upregulated in brain-tropic sublines compared to parental cells. To understand the functional relevance of these gap-junction coupled tumor cell networks, we performed bulk RNA-Sequencing of two melanoma metastasis models in monoculture under gap junction inhibition and control: We observed reduced calcium communication and a concordant downregulation of neurodevelopmental and synaptic pathways within the tumor cells upon network disconnection, highlighting a potential recapitulation of neural-like features in metastases to the brain. To investigate the therapeutic potential of targeting these networks, we treated mice with brain metastases with two brain-penetrant gap junction blockers and observed significantly reduced tumor burden. Finally, we investigated the presence of heterotypic networks between brain metastases cells and the brain microenvironment with our SR101 dye transfer pipeline. Contrary to our findings in primary brain tumors, no gap junction connections to cells of the brain microenvironment in those models with reduced tumor growth upon network inhibition could be detected, supporting a cancer cell intrinsic gap junction mediated network and highlighting a selectivity of these homogenous tumor networks and potential therapeutic vulnerability.

Identification of genetic fingerprint of type I interferon therapy in visceral metastases of melanoma

Malignant melanoma is a difficult-to-treat skin cancer with increasing incidence worldwide. Although type-I interferon (IFN) is no longer part of guidelines, several melanoma patients are treated with type-I interferon (IFN) at some point of the disease, potentially affecting its genetic progression. We run genome-wide copy number variation (CNV) analysis on previously type-I IFN-treated (n = 17) and control (n = 11) visceral metastases of melanoma patients. Results were completed with data from the TCGA and MM500 databases. We identified metastasis- and brain metastasis-specific gene signatures mostly affected by CN gains. Some cases were genetically resistant to IFN showing characteristic gene alterations (e.g. ABCA4 or ZEB2 gain and alterations of DNA repair genes). Analysis of a previously identified type-I IFN resistance gene set indicates that only a proportion of these genes was exclusive for the IFN-treated metastases reflecting a possible selective genomic pressure of endogenous IFNs during progression. Our data suggest that previous type-I IFN treatment and/or endogenous IFN production by immune response affect genomic progression of melanoma which may have clinical relevance, potentially influence immune checkpoint regulation in the tumor microenvironment.

Homeostatic self-MHC-I recognition regulates anti-metastatic function of mature lung natural killer cells

Natural killer (NK) cells are important innate immune effector cells for controlling tumor growth and metastasis. Differentiated mature NK cells preferentially reside in the peripheral tissues and express higher levels of self-major histocompatibility complex class I (MHC-I)-recognizing inhibitory receptors. MHC-I recognition by NK cells are known to be important for their development and maturation processes, however, the role of homeostatic MHC-I recognition in maintaining effector functions of mature NK cells in the peripheral tissues needs to be elucidated. In this study, we utilized a pan anti-MHC-I blocking monoclonal antibody (anti-MHC-I) to examine the role of homeostatic MHC-I recognition in the response of pulmonary mature NK cells in an experimental lung metastasis model of B16F10 melanoma. Anti-MHC-I treatment showed significant inhibition of the lung metastasis of B16F10 melanoma in NK cell- and IFN-γ-dependent mechanisms. The blockade of homeostatic MHC-I recognition increased mature lung NK cell responsiveness, such as direct cytotoxicity and IFN-γ production, rather than the number of lung NK cells. Mechanistically, the gene expression of activating receptors including DNAX accessory molecule-1 (DNAM-1) was upregulated in NK cells treated with anti-MHC-I, and further the enhanced NK cell cytotoxicity against B16F10 cells was DNAM-1-dependent. Collectively, homeostatic self-MHC-I recognition regulates anti-metastatic function of mature lung NK cells by restraining the expression of activating receptors.

Exosomal lncRNA Mir100hg derived from cancer stem cells enhance glycolysis and promote metastasis of melanoma through miR-16-5p and miR-23a-3p

Exosomal lncRNA Mir100hg derived from cancer stem cells enhance glycolysis and promote metastasis of melanoma through miR-16-5p and miR-23a-3p

Machine learning and single-cell RNA sequencing reveal relationship between intratumor CD8+ T cells and uveal melanoma metastasis

PurposeUveal melanoma (UM) is adults’ most common primary intraocular malignant tumor. It has been observed that 40% of patients experience distant metastasis during subsequent treatment. While there exist multigene models developed using machine learning methods to assess metastasis and prognosis, the immune microenvironment’s specific mechanisms influencing the tumor microenvironment have not been clarified. Single-cell transcriptome sequencing can accurately identify different types of cells in a tissue for precise analysis. This study aims to develop a model with fewer genes to evaluate metastasis risk in UM patients and provide a theoretical basis for UM immunotherapy.MethodsRNA-seq data and clinical information from 79 μm patients from TCGA were used to construct prognostic models. Mechanisms were probed using two single-cell datasets derived from the GEO database. After screening for metastasis-related genes, enrichment analysis was performed using GO and KEGG. Prognostic genes were screened using log-rank test and one-way Cox regression, and prognostic models were established using LASSO regression analysis and multifactor Cox regression analysis. The TCGA-UVM dataset was used as internal validation and dataset GSE22138 as external validation data. A time-dependent subject work characteristic curve (time-ROC) was established to assess the predictive ability of the model. Subsequently, dimensionality reduction, clustering, pseudo-temporal analysis and cellular communication analysis were performed on GSE138665 and GSE139829 to explore the underlying mechanisms involved. Cellular experiments were also used to validate the relevant findings.ResultsBased on clinical characteristics and RNA-seq transcriptomic data from 79 samples in the TCGA-UVM cohort, 247 metastasis-related genes were identified. Survival models for three genes (SLC25A38, EDNRB, and LURAP1) were then constructed using lasso regression and multifactorial cox regression. Kaplan-Meier survival analysis showed that the high-risk group was associated with poorer overall survival (OS) and metastasis-free survival (MFS) in UM patients. Time-dependent ROC curves demonstrated high predictive performance in 6 m, 18 m, and 30 m prognostic models. Cell scratch assay showed that the 24 h and 48 h migration rates of cells with reduced expression of the three genes were significantly higher than those of the si-NC group. CD8 + T cells may play an important role in tumour metastasis as revealed by immune infiltration analysis. An increase in the percentage of cytotoxic CD8 + T cells in the metastatic high-risk group was found in the exploration of single-cell transcriptome data. The communication intensity of cytotoxic CD8 was significantly enhanced. It was also found that the CD8 + T cells in the two groups were in different states, although the number of CD8 + T cells in the high-risk group increased, they were mostly in the exhausted and undifferentiated state, while in the low-risk group, the CD8 + T cells were mostly in the functional state.ConclusionsWe developed a precise and stable 3-gene model to predict the metastatic risk and prognosis of patients. CD8 + T cells exhaustion in the tumor microenvironment play a crucial role in UM metastasis.

Stereotactic Radiosurgery Dose Reduction for Melanoma Brain Metastases Patients on Immunotherapy or Target Therapy: A Single-Center Experience.

Better local control but higher rates of adverse radiation events (ARE) have been reported when combining American Society for Radiation Oncology (ASTRO)-guideline-suggested dose (SD) stereotactic radiosurgery (SRS) with immunotherapy or targeted therapy for melanoma brain metastases. The objective of this study is to explore the efficacy and safety of lower prescription doses compared with ASTRO guidelines for single-fraction SRS for patients with melanoma metastases who are concurrently receiving immunotherapy or targeted therapy. We conducted a retrospective, single-center study on 194 patients who underwent SRS between 2009 and 2022. After propensity score matching, 71 patients with 292 metastases were included in the ASTRO-SD (20-24 Gy for <2 cm, 18 Gy for ≥2 to <3 cm) group and 33 patients with 292 metastases in the reduced dose (RD, <20 Gy for <2 cm, <18 Gy for ≥2 to <3 cm) group. The median diameter (5.4 vs 5.2 mm, P = .6), prescription volume (0.2 vs 0.2 cm3, P = .2), and radiographic follow-up (11 vs 12 months, P = .2) were similar in the 2 groups. The cumulative incidence of progressing metastases was significantly higher in the SD compared with the RD group (P = .018). Higher prescription volumes and ASTRO-suggested radiation doses were associated with local progression in multivariable analysis. Radiographic AREs were significantly more common in the SD compared with the RD group (8.6% vs 3.1%, P = .005). BRAF and other tyrosine kinase inhibitors' concurrent use, higher prescription volumes, and ASTRO-suggested radiation doses were associated with an increased risk of radiographic ARE. This study provides evidence that RD SRS could offer reduced toxicity rates, while maintaining high local control as compared with the current guideline-SDs for the treatment of melanoma brain metastases.

Metastatic Melanoma to the Small Bowel and Colon: A Systematic Review of the Global Experience and Institutional Cohort Analysis Detailing a Rare Clinical Entity.

Cutaneous melanoma is among the most common solid tumors to metastasize to the gastrointestinal (GI) tract. Literature summarizing the clinical experience and features of this unique pathology is lacking. A systematic review of the available literature reporting clinically salient features of melanoma metastases to the small and large intestines was conducted. Additionally, we surveyed our institutional experience of surgically treated melanoma metastasis to the small bowel and colon. A descriptive analysis was performed. Kaplan-Meier curves with log-rank tests were used to analyze time-to-event intervals. Univariable and multivariable Cox logistic regression models were generated to identify predictors of survival. Over 100 studies including 1153 patients were included. GI metastases predominantly affected males, were in the small bowel/jejunum, equally presented as solitary and multiple lesions, and were generally not the first site of distant metastatic disease. The median time from primary lesion diagnosis to GI metastasis was 48 months. Analysis of our institutional cohort suggested that survival in patients receiving complete GI-specific surgical resection and immune checkpoint inhibitors (ICIs) was prolonged compared to palliative resection and without ICI therapy. Positive prognostic factors for survival following GI metastasis included fewer GI metastatic lesions, complete resection, and longer duration between primary tumor diagnosis and GI metastasis. GI metastases are a sign of advanced metastatic melanoma. Clinical suspicion of metastatic involvement in patients with a history of melanoma who develop any abdominal symptoms or anemia should remain high. Receipt of complete surgical resection and ICIs may prolong survival in disseminated melanoma.

Selenocysteine tRNA methylation promotes oxidative stress resistance in melanoma metastasis.

Selenocysteine-containing proteins play a central role in redox homeostasis. Their translation is a highly regulated process and is dependent on two tRNASec isodecoders differing by a single 2'-O-ribose methylation called Um34. Here we characterized FTSJ1 as the Um34 methyltransferase and show that its activity is required for efficient selenocysteine insertion at the UGA stop codon during translation. Specifically, loss of Um34 leads to ribosomal stalling and decreased UGA recoding. FTSJ1-deficient cells are more sensitive to oxidative stress and show decreased metastatic colonization in xenograft models of melanoma metastasis. We found that FTSJ1 mediates efficient translation of selenoproteins essential for the cellular antioxidant response. Our findings uncover a role for tRNASec Um34 modification in oxidative stress resistance and highlight FTSJ1 as a potential therapeutic target specific for metastatic disease.

In vitro effects of l-kynurenine and quinolinic acid on adhesion, migration and apoptosis in B16 F10 melanoma cells

IntroductionThe inhibition of melanoma adhesion through adhesion molecules, such as integrins and E-cadherin, may represent a promising strategy for managing melanoma metastasis. Compounds, namely l-kynurenine (L-kyn) and quinolinic acid (Quin), previously displayed anti-cancer effects at half-maximal inhibitory concentration (IC50) against B16 F10 melanoma cells in vitro. However, the role of these compounds in B16 F10 melanoma cell adhesion, migration and apoptosis remain unknown. MethodsPost-exposure to the compounds, flow cytometry was used to analyse the expression of very late antigen-5 (VLA-5), E-cadherin and cleaved caspase-3 in B16 F10 melanoma and RAW 264.7 murine macrophage cells. An adhesion assay was used to quantify the adhesion of both cell lines to vitronectin. A scratch migration assay was used to measure the possible inhibition of cell migration in B16 F10 cells in response to L-kyn and Quin. ResultsIn both B16 F10 and RAW 264.7 cells, neither L-kyn nor Quin induced significant effects on VLA-5 expression or cell adhesion to vitronectin. In B16 F10 cells, both L-kyn and Quin elevated E-cadherin expression and displayed a trend of suppressed migration. However, only L-kyn elevated E-cadherin in RAW 264.7 cells. L-kyn induced apoptosis by elevating cleaved caspase-3 expression in both cell lines. ConclusionL-kyn and Quin demonstrated promising antimetastatic effects in their ability to elevate E-cadherin expression and induce apoptosis in B16 F10 melanoma cells. However, these effects did not occur in response to vitronectin or VLA-5 integrin alterations. Furthermore, it cannot be excluded that L-kyn also induced apoptosis in RAW 264.7 cells. As such, these effects should be confirmed in additional control cell lines and substantiated with in vivo models.

Metachronous Cutaneous Melanoma Metastases of the Left Anterior Paranasal Sinuses and the Right Nasopharynx That Were Treated Surgically: A Case Report With Literature Review

Metachronous Cutaneous Melanoma Metastases of the Left Anterior Paranasal Sinuses and the Right Nasopharynx That Were Treated Surgically: A Case Report With Literature Review

PP2A negatively regulates NK cell T-bet expression and anti-tumor effector function.

The transcription factor T-bet is essential for the anti-tumor effector function of NK cells, but the mechanism regulating its expression in NK cells remains unclear. In this study, we aimed to identify an NK cell intrinsic regulator that controls T-bet expression. Using T-bet-luciferase reporter assay screening, we identified a protein phosphatase inhibitor as a potential activator of T-bet expression. A series of PP2A-specific inhibitors (PP2Ai) or PP2A siRNA induced the expression of T-bet. In PP2Ai-treated mice, the expressions of T-bet and its downstream effector molecules, granzyme B and IFN-γ, were also upregulated in NK cells. Mechanistically, PP2Ai increased the phosphorylation of mTOR and ribosomal protein S6 in NK cells, and mTOR inhibitor canceled the effects of PP2Ai in NK cells. Importantly, NK cells isolated from PP2Ai-treated mice showed higher cytotoxicity and IFN-γ production; therefore, they increased the anti-tumor effector function of NK cells. Accordingly, PP2Ai treatment inhibited lung metastasis of B16 melanoma by NK cell- and mTOR-dependent mechanisms. These results suggest that PP2A negatively regulates NK cell T-bet expression and effector function by an mTOR-dependent mechanism.

De-Differentiated Metastatic Melanoma: A Diagnostic Challenge

Abstract Introduction/Objective Melanoma is one of the common malignant neoplasms with several histological appearances. Its diagnosis is usually based on histomorphologic findings and immunohistochemistry (IHC). De-differentiated melanoma (DM) is a special form of malignant melanoma with different histo-morphologic and immunohistochemical characteristics than conventional malignant melanomas. It has a worse prognosis and can impose a diagnostic challenge. Methods/Case Report A 62-year-old male previously had melanoma on his chest, which was surgically removed with negative margins and no involvement of nearby sentinel lymph nodes. The patient presented two months later with a rapidly enlarging mass in the right axillary region. Subsequent lymph node dissection revealed several nodes, the largest measuring 14 cm with extensive necrosis. Microscopic examination of the largest mass revealed clusters of irregularly shaped cells displaying marked nuclear abnormalities, prominent nucleoli, and heightened mitotic activity. Immunohistochemical analysis of this mass showed negative results for SOX-10, HMB45, Melan-A, S100 protein, CAM5.2, BRAF, and myogenin. Conversely, the remaining lymph nodes exhibited typical signs of melanoma metastasis, expressing relevant melanoma markers. Given the patient’s history of melanoma and the presence of metastatic melanoma in nearby lymph nodes, the histological and immunohistochemical findings support the diagnosis of dedifferentiated cutaneous metastatic melanoma. Results (if a Case Study enter NA) N/A Conclusion The distinctive appearance and lack of typical markers in this type of melanoma pose considerable difficulties in distinguishing it from other malignancies like poorly differentiated carcinomas and high-grade sarcomas. Furthermore, dedifferentiated melanoma tends to be highly aggressive, often resisting standard and immunological treatments. Therefore, comprehensive clinical and histomorphological assessments are crucial for accurately diagnosing these complex and challenging cases.

The Expression of miR-211-5p in Sentinel Lymph Node Metastases of Malignant Melanoma Is a Potential Marker for Poor Prognosis

Metastatic primary cutaneous melanoma is a frequently fatal disease despite recent therapeutic advances. Biomarkers to stratify patients’ prognosis are lacking. MicroRNAs (miRNAs) are small, non-coding RNAs. We aimed to determine the expression of miR-211-5p in primary tumors and metastases of malignant melanoma and its potential use as a prognostic biomarker. We performed in situ hybridization for miRNA-211-5p on 109 FFPE melanoma samples from 76 patients, including 31 paired primary tumor/metastasis samples. For validation, we performed in silico analyses of TCGA skin cutaneous melanoma (SKCM) cohort. High miR-211-5p expression was more frequent in primary tumors (70.8%) compared to metastases (39.3%). In metastases, it was associated with a significantly worse overall survival. Data from TCGA SKCM cohort confirmed that high miR-211-5p expression in melanoma metastases, but not primary tumors, is associated with worse overall survival. MiR-211-5p expression in metastases is associated with a shorter survival, emphasizing the potential of miR-211-5p as a risk predictor for a less favorable clinical outcome in metastatic disease. In situ hybridization could be implemented in a routine laboratory workflow and can be performed on diagnostic tissue.

Penfluridol inhibits melanoma growth and metastasis through enhancing von Hippel‒Lindau tumor suppressor-mediated cancerous inhibitor of protein phosphatase 2A (CIP2A) degradation.

Melanoma's high metastatic potential, especially to the brain, poses significant challenges to patient survival. The blood‒brain barrier (BBB) is a major obstacle to the effective treatment of melanoma brain metastases. We screened antipsychotic drugs capable of crossing the BBB and identified penfluridol (PF) as the most active candidate. PF reduced melanoma cell viability and induced apoptosis. In animal models, PF effectively inhibited melanoma growth and metastasis to the lung and brain. Using immunoprecipitation combined with high-resolution mass spectrometry, and other techniques such as drug affinity responsive target stability, we identified CIP2A as a direct binding protein of PF. CIP2A is highly expressed in melanoma and its metastases, and is linked to poor prognosis. PF can restore Protein Phosphatase 2A activity by promoting CIP2A degradation, thereby inhibiting several key oncogenic pathways, including AKT and c-Myc. Additionally, von Hippel‒Lindau (VHL) is the endogenous E3 ligase for CIP2A, and PF enhances the interaction between VHL and CIP2A, promoting the ubiquitin‒proteasome degradation of CIP2A, thereby inhibiting melanoma growth and metastasis. Overall, this study not only suggests PF's potential in treating melanoma and its brain metastases but also highlights CIP2A degradation as a therapeutic strategy for melanoma.

Repurposing of Antidiarrheal Loperamide for Treating Melanoma by Inducing Cell Apoptosis and Cell Metastasis Suppression In vitro and In vivo.

Melanoma is the most common skin tumor worldwide and still lacks effective therapeutic agents in clinical practice. Repurposing of existing drugs for clinical tumor treatment is an attractive and effective strategy. Loperamide is a commonly used anti-diarrheal drug with excellent safety profiles. However, the affection and mechanism of loperamide in melanoma remain unknown. Herein, the potential anti-melanoma effects and mechanism of loperamide were investigated in vitro and in vivo. In the present study, we demonstrated that loperamide possessed a strong inhibition in cell viability and proliferation in melanoma using MTT, colony formation and EUD incorporation assays. Meanwhile, xenograft tumor models were established to investigate the anti-melanoma activity of loperamide in vivo. Moreover, the effects of loperamide on apoptosis in melanoma cells and potential mechanisms were explored by Annexin V-FITC apoptosis detection, cell cycle, mitochondrial membrane potential assay, reactive oxygen species level detection, and apoptosis-correlation proteins analysis. Furthermore, loperamide-suppressed melanoma metastasis was studied by migration and invasion assays. What's more, immunohistochemical and immunofluorescence staining assays were applied to demonstrate the mechanism of loperamide against melanoma in vivo. Finally, we performed the analysis of routine blood and blood biochemical, as well as hematoxylin- eosin (H&E) staining, in order to investigate the safety properties of loperamide. Loperamide could observably inhibit melanoma cell proliferation in vitro and in vivo. Meanwhile, loperamide induced melanoma cell apoptosis by accumulation of the sub-G1 cells population, enhancement of reactive oxygen species level, depletion of mitochondrial membrane potential, and apoptosis-related protein activation in vitro. Of note, apoptosis-inducing effects were also observed in vivo. Subsequently, loperamide markedly restrained melanoma cell migration and invasion in vitro and in vivo. Ultimately, loperamide was witnessed to have an amicable safety profile. These findings suggested that repurposing of loperamide might have great potential as a novel and safe alternative strategy to cure melanoma via inhibiting proliferation, inducing apoptosis and cell cycle arrest, and suppressing migration and invasion.

S3035 A Rare Case of Extensive Melanoma Metastasis to the Ascending Colon and Gallbladder

S3035 A Rare Case of Extensive Melanoma Metastasis to the Ascending Colon and Gallbladder

TCTN1 Induces Fatty Acid Oxidation to Promote Melanoma Metastasis.

Metabolic reprogramming promotes and sustains multiple steps of melanoma metastasis. Identification of key regulators of metabolic reprogramming could lead to the development of treatments for preventing and treating metastatic melanoma. Here, we identified that the tectonic family member TCTN1 promotes melanoma metastasis by increasing fatty acid oxidation (FAO). In clinical melanoma samples, high expression of TCTN1 correlated with increased metastasis and shorter patient survival. Functionally, TCTN1 promoted melanoma invasion and migration in vitro and distant metastasis in vivo, and TCTN1 induced a mesenchymal-like phenotype switch. Mechanistically, TCTN1 acted as a protein scaffold to promote the binding of HADHA and HADHB, subunits of the mitochondrial trifunctional protein complex, thus leading to FAO activation. TCTN1-mediated FAO activated the p38/MAPK signaling pathway in melanoma cells, promoting tumor EMT and stemness. Molecular docking indicated that the prostaglandin F receptor agonist fluprostenol can block HADHA/HADHB binding, which was confirmed experimentally. Treatment with fluprostenol was able to inhibit TCTN1-induced melanoma invasion and metastasis. Taken together, these findings elucidate the mechanism of TCTN1-mediated promotion of melanoma metastasis and support the potential application of fluprostenol for targeted therapy of metastatic melanoma.