SIRT6 promotes a mesenchymal, invasive phenotype in melanoma

Abstract

Melanoma is a highly aggressive cutaneous malignancy with significant metastatic potential, posing ongoing challenges in elucidating the molecular mechanisms driving its progression. Epithelial–mesenchymal transition (EMT) is a key process that enhances cancer cell migration and invasion, facilitating metastatic dissemination. Sirtuin 6 (SIRT6), a NAD⁺-dependent enzyme, has been reported to either promote or suppress EMT in a context-dependent manner; however, its function in melanoma remains poorly defined. We previously showed that SIRT6 is overexpressed in melanoma tissues and cells, where it contributes to tumor growth via an autophagy-dependent mechanism. In this study, we further investigated the role of SIRT6 in EMT using melanoma cells with stable overexpression or shRNA-mediated knockdown. Overexpression of SIRT6 enhanced cell proliferation and clonogenic survival and was associated with increased expression of mesenchymal markers and EMT-associated transcription factors. Conversely, SIRT6 knockdown significantly reduced the expression of these genes. Functionally, SIRT6 depletion markedly impaired cell migration and invasion, as demonstrated by wound-healing and Matrigel invasion assays. Collectively, these findings identify SIRT6 as a promoter of an invasive, mesenchymal phenotype in melanoma. Our results support a potential oncogenic role for SIRT6 and highlight its inhibition as a promising therapeutic strategy to limit melanoma progression and metastasis.

Introduction

Although targeted therapy and immunotherapy have yielded significant clinical advances in recent years, melanoma continues to carry a high mortality burden, with an estimated 112,000 new cases and 8,510 deaths projected in 2026 [1]. Further, melanoma with distant metastases has a 5-year relative survival rate of only 32% [2], leading to a sustained need to find more effective treatments for this metastatic cancer. This underlines the understanding that melanoma is most dangerous not because of the primary tumor, but because of its early and aggressive metastatic spread. Epithelial-mesenchymal transition (EMT)-like phenotype switching is an important mechanism of tumor progression and metastasis, which includes a loss of epithelial-type cell characteristics (cell–cell junctions, apicobasal polarity of the cell, and cobblestone morphology) and an acquisition of more mesenchymal-type characteristics (increased cell-matrix adhesions and motility, and fibroblast-like cell morphology) [3,4]. Further, EMT results in increased cell migration and invasion, which allows cancer cells to invade the surrounding vasculature, leading to tumor dissemination and metastases [5,6]. Mechanistically, EMT can be recognized by the downregulation of epithelial markers such as E-cadherin and cytokeratin, and the upregulation of mesenchymal markers such as N-cadherin, Vimentin and β-catenin as well as transcriptional regulators of EMT such as Snail and Zeb1 [3,7]. Like other cancer types, the phenomenon of EMT has been reported in melanoma [4,8]. In fact, EMT-associated signaling molecules are considered potential therapeutic targets, and EMT-modulating agents can serve as therapeutics against melanoma metastasis [4,9].

Sirtuin 6 (SIRT6) is a nuclear sirtuin that possesses mono-ADP-ribosylase and NAD⁺- dependent deacetylase activities. Due to the essential roles SIRT6 has in cellular homeostasis, disruptions of this enzyme appear to have strong effects on melanoma development. We have previously shown that SIRT6 is significantly overexpressed in human melanoma tissues and cells, and lentiviral short hairpin RNA-mediated knockdown of SIRT6 results in a marked anti-proliferative response in melanoma cells [10]. Further, a recent multi-omics analysis demonstrated that SIRT6 knockdown resulted in global transcriptional reprogramming, including the suppression of genes involved in cell migration/invasion, cytoskeletal remodeling, and pro-metastatic inflammatory signaling [11]. Interestingly, SIRT6 has been shown to promote or inhibit EMT in other cancer types, depending on the cancer [12–15]. However, its precise role in EMT and metastasis of melanoma remain unexplored. In this study, we determined the effect of SIRT6 overexpression on melanoma cell growth, long-term survival in terms of colony formation ability, and EMT-associated mesenchymal markers. In addition, we also validated the role of SIRT6 in melanoma phenotype switching by determining the effects of shRNA-mediated SIRT6 knockdown on melanoma cell migration, invasion, and mesenchymal markers.

Materials and methods

Cell culture

SK-MEL-3 human melanoma cell lines were purchased from ATCC and maintained in McCoy’s 5 A medium (Corning #10-050-CV) supplemented with 15% FBS (Sigma-Aldrich #F2442). A375 cells were maintained in DMEM (Corning #10-013-CV) supplemented with 10% FBS (Sigma-Aldrich #F2442). A375 nonsense shRNA (shNS)- and SIRT6-targeting shRNA (shSIRT6)-containing cells were transduced using lentiviral shRNA plasmids from Sigma-Aldrich as published previously [10], and then plated at low density in 10 cm² dishes until discrete colonies formed. Colonies were picked and the best clone with low SIRT6 levels was chosen for further experiments.

Using Lipofectamine 2000 (ThermoFisher #1168019), SK-MEL-3 cells were transfected with either pcDNA 3.1 or pcDNA3.1-SIRT6-Flag (SIRT6 Flag was a gift from Eric Verdin (Addgene plasmid #13817; RRID:Addgene_13817)) [16], allowed to recover for 24 hours, and then grown in G418 to select for the transfected clones. Cells were plated at low density in 10 cm² dishes and once discrete colonies formed, they were picked and passaged into larger dishes as appropriate. Once enough cells were available, cells were collected and subjected to immunoblot and viability assays as described below to determine the best clone to use for further studies.

All cells were maintained at standard cell culture conditions (37 °C, 5% CO₂ in a humidified incubator) and routinely (upon thawing, or if a cell line is in culture for more than 3 months) authenticated using STR testing through the University of Wisconsin Translational Research Initiatives in Pathology (TRIP) lab core facility. Cell lines were also routinely (monthly and upon cryo-recovery) tested for the absence of mycoplasma using the MycoAlert Plus Mycoplasma Detection Kit (Lonza #LT07–710) using the manufacturer’s instructions and reading on a BioTek Synergy H1 Multimode Plate Reader (Agilent).

Protein isolation, quantification, and immunoblotting

Protein was isolated from individual cell pellets with 1X RIPA lysis buffer (Millipore #20–188) supplemented with phenylmethylsulfonyl fluoride (PMSF, Amresco #0754) and protease inhibitor cocktail (Thermo Scientific/Pierce #78444), and quantified using Pierce BCA Protein assay kit (Thermo Scientific/Pierce #23225) using a BioTek Synergy H1 plate reader as per the manufacturer’s instructions. Proteins were subjected to SDS-PAGE and immunoblotting as described previously [10].

RealTime-Glo MT cell viability assay

RealTime-Glo MT cell viability assay (Promega #G9713) was performed on the clones using the manufacturer’s protocol. Briefly, cells were counted and 1,500 cells were added per well in 5–6 replicates in a white 96-well plate with the appropriate amounts of media and RealTime-Glo reagents. Luminescence measurements were taken at the indicated times using a BioTek Synergy H1 Multimode Plate Reader (Agilent), and individual well reads were normalized back to the initial 1 h timepoint reading to account for well-to-well variation.

Clonogenic cell survival assay

Cells were trypsinized and counted, and 200 viable cells from each group were plated in separate wells of a 6-well plate. Cells were maintained under standard tissue culture conditions, changing media every 3 days for approximately 10 days, until the cells reached optimal density. Cells were then stained at room temp with 0.5% crystal violet solution in 1:1 methanol:water, rinsed 2 times with phosphate-buffered saline (pH 7.4), and air-dried before images were captured, followed by manual counting of colonies.

RNA isolation, cDNA synthesis and RT-qPCR

Total RNA was isolated from cells using the RNeasy Plus Mini Kit (Qiagen #74134), per manufacturer’s protocol, with all optional steps performed. cDNA was synthesized using M-MLV Reverse Transcriptase (Promega #M1701), Oligo dT primers (Promega #C1101) and dNTP Mix (Promega #U1511) in an Eppendorf Mastercycler Nexus X2. RT-qPCR was performed with PowerTrack SYBR Green Master Mix (Applied Biosystems, A46109) and ran on a BioRad CFX 96 Touch Real-Time PCR machine. CFX Maestro 1.1 (Bio-Rad, version 4.1.2433.1219) software was used for RT-qPCR analysis using default normalization parameters and ACTB as the endogenous control. A list of the primers used in this study is provided in Supplemental Table 1.

Wound-healing scratch assay

Cells (5 × 10⁵ cells) were seeded into 3 wells per group into 6-well plates and incubated at standard conditions. Once cells reached confluency (24–48 h later), a wound was made by scratching the confluent monolayer with a sterile 200 μL pipette tip. The loose cells in the scratch were gently rinsed off with media, fresh media was added, and the remaining cells were allowed to continue growing in standard conditions. Images of cells were acquired at 0 and 24 h post-scratch using EVOS XL Core microscope (ThermoFisher) at 20× magnification. Cell migration towards the mid-line of the scratch was assessed and measured using ImageJ.

Invasion assay

Cell invasion assay was performed using Matrigel Invasion Chambers (Corning #354480) per the manufacturer’s protocol. Briefly, shNS or shSIRT6 A375 cells (2.5 × 10⁴ cells/well in 0.5 mL of DMEM medium without FBS) were seeded in the inner chamber of Matrigel-coated transwell kept in a 24-well plate. Chemoattractant (media with 10% FBS) was added to the outer chamber of the plate, followed by incubation at 37 °C and 5% CO₂ for 24–48 h. For the assessment of invasion, remaining cells in the upper surface of the chamber were removed and cells on the lower surface of the inner chamber were then gently rinsed with PBS and were stained with crystal violet staining solution at room temperature for 30 min, followed by washing 3 times with PBS. The images of stained cells were captured using an EVOS XL Core microscope (ThermoFisher) at 20× magnification and cells were manually counted for quantification.

Statistical analysis

Statistical analyses were performed using GraphPad PRISM 10.4.2 software (GraphPad Software, Inc., La Jolla, CA). Statistical significance was determined via multiple t-tests using the Holm–Sidak method for studies with two experimental groups. Data are shown as mean ± SEM with at least 2 biological replicates, and at least 3 technical replicates unless otherwise specified. p values less than 0.05 were considered significant.

Results and discussion

SIRT6 overexpression (OE) in SK-MEL-3 melanoma cells leads to enhanced cell growth and survival

SIRT6 acts as a double-edged sword in cancer, capable of functioning as either a tumor suppressor or an oncogene in a context-dependent manner [17]. Prior research has been conducted to attempt to better define the exact role of SIRT6 in various cancer types. In non-melanoma skin cancer (particularly squamous cell carcinoma), SIRT6 is shown to function as an oncogene [18], however, in some other cancer types, such as liver and intestinal cancer, SIRT6 is shown to be a tumor suppressor [19]. Previously, we found evidence that SIRT6 has an oncogenic role in human melanoma [10,20]. To further determine the direct role of SIRT6 in melanoma, we generated stable SIRT6 overexpression (OE) in SK-MEL-3 cells. As shown in Figure 1A, we successfully achieved SIRT6 OE in SK-MEL-3 human melanoma cells and validated SIRT6 protein expression levels in several different clones. Based on immunoblot data, Clone 2 showed the highest SIRT6 protein expression and was used for further experiments.

Figure 1.

Effects of SIRT6 overexpression on the growth and long-term survival of melanoma cells. SK-MEL-3 melanoma cells were used for SIRT6 overexpression (OE) along with an appropriate empty vector control (pcDNA). (A) Cell lysates of three different clones were subjected to an immunoblot analysis to confirm SIRT6 overexpression. β-tubulin was used as a loading control. Clone 2 was used for further experiments. (B) Cell growth and viability were determined by RealTime Glo MT viability assay, and quantitative results are shown as relative cell growth with SIRT6 overexpression compared to pcDNA vector control. (C) Representative images of clonogenic survival assay stained with crystal violet. (D) Quantification of the clonogenic survival assay. The data are expressed as mean ± SEM of two biological and three technical replicates, with ****p ≤ 0.0001. pcDNA: Empty vector; SIRT6 OE: SIRT6 overexpression.

Subsequently, we assessed the effects of SIRT6 OE on the proliferative potential of the SK-MEL-3 cells. Our data demonstrated that SIRT6 OE significantly increased cell growth and viability in SK-MEL-3 cells as compared to the pcDNA empty-vector control cells (Figure 1B). Additionally, we evaluated the long-term clonogenic survival of the SIRT6 OE SK-MEL-3 cells using a colony formation assay. As seen in Figure 1C, SIRT6 OE leads to a significant increase in the colony formation abilities of the cells when compared to the pcDNA control cells (Figure 1C,D), suggesting SIRT6 OE in SK-MEL-3 cells resulted in a significant increase in long-term proliferation potential. Our findings provide evidence supporting SIRT6 as a potential oncogene in melanoma, which corroborates our earlier published study showing SIRT6 is overexpressed in human melanoma cells and tissues and may contribute to melanoma development and/or progression [10].

SIRT6 OE in SK-MEL-3 melanoma cells increases EMT-related genes

EMT, which is an important phenotypic switching process through which epithelial-like cancers become invasive and metastatic by losing epithelial cell characteristics such as tight cell-cell junctions, apicobasal polarity, and cobblestone morphology, while acquiring mesenchymal characteristics such as increased cell-matrix adhesions and cell motility [21–24]. Cellular migration and invasion enable melanoma cells to detach from the primary tumor, degrade and traverse the extracellular matrix (ECM), enter lymphatic or blood vessels, and colonize distant organs (e.g., lung, brain, liver). Interestingly, pigmented melanocytes are typically derivative of a group of migratory embryonic cells referred to as the neural crest [25]. Studies suggest that EMT, a developmental program, is recapitulated during melanoma progression/metastasis. In melanoma (which is not strictly epithelial), EMT-like transcriptional reprogramming, often termed phenotype switching, plays a similar role in promoting invasion and metastasis. In melanoma, migration and invasion are tightly linked to EMT-like phenotype switching [26]. Caramel et al. reported that transcriptional regulators Twist1 and Zeb1 function in favor of melanocyte dedifferentiation and neoplastic transformation in response to NRAS/BRAF activation, which was further associated with enhanced phospho-ERK levels in late-stage melanoma [27]. The authors also posit that EMT may be considered an independent factor of poor prognosis in melanoma patients, and results in E-cadherin loss, enhanced migration, and invasion [27].

Interestingly, SIRT6 has been shown to either promote or inhibit EMT, depending on cellular context [28,29]. Thus, here we investigated the role of SIRT6 in modulating EMT in melanocytic cells. Using the shNS and SIRT6 OE SK-MEL-3 cells, we performed RT-qPCR analysis on key mesenchymal markers and transcriptional regulators of EMT. We showed that SIRT6 overexpression significantly increased expression of the mesenchymal markers N-cadherin, vimentin, and β-catenin as compared to the pcDNA empty vector control cells (Figure 2A). These molecules are well-established mediators of cytoskeletal reorganization, cell-cell adhesion remodeling, and enhanced cellular plasticity, all of which are critical features of the EMT program. Additionally, we found significant increases in the EMT-transcriptional regulators Zeb1, Snail, and Twist (Figure 2B) in SIRT6 OE SK-MEL-3 melanoma cells compared to control cells. These transcription factors function as master regulators of EMT by orchestrating widespread transcriptional reprogramming and phenotypic switching. This supports the idea that SIRT6 may be involved in the EMT-regulatory circuit, thereby promoting a sustained and coordinated mesenchymal transition.

Figure 2.

Effect of SIRT6 overexpression (OE) on key mesenchymal markers in human melanoma cells. Relative mRNA expression levels of (A) mesenchymal markers and (B) EMT transcriptional regulators in pcDNA control and SIRT6 OE SK-MEL-3 melanoma cells. Data shown as mean ± SEM for two biological and three technical replicates, with *p ≤ 0.05, **p ≤ 0.01, ***p ≤ 0.001, ****p ≤ 0.0001. pcDNA: Empty vector; SIRT6 OE: SIRT6 overexpression.

SIRT6 knockdown in A375 melanoma cells inhibits EMT-related genes

In a previous study, we showed that SIRT6 is upregulated in A375 melanoma cells, and SIRT6 knockdown (KD) via shRNA-mediated RNA interference resulted in a marked antiproliferative response (growth inhibition, cell cycle alteration, senescence, and autophagy dysregulation) [10]. In a follow-up study, we found that CRISPR/Cas9-mediated knockout of SIRT6 in A375 melanoma cells resulted in a significant decrease in cell viability, clonogenic survival, and induced G1-phase cell cycle arrest [20]. Interestingly, in this study, RT² Profiler PCR array analysis of SIRT6 KO cells resulted in modulation of several cancer pathways, including EMT [20].

To further validate the role of SIRT6 in EMT in melanoma, we performed RT-qPCR analysis for EMT-related genes after SIRT6 KD in A375 melanoma cells. As expected, SIRT6 KD inhibited expression of key mesenchymal markers N-cadherin, vimentin, and β-catenin (Figure 3A). This was the opposite of what we found with the SIRT6 OE cells above. Additionally, we found significant decreases in the EMT-transcriptional regulators Zeb1, Snail, and Twist in shSIRT6 A375 melanoma cells as compared to the shNS cells (Figure 3B). These findings strongly support a role for SIRT6 as a modulator of EMT at the transcriptional level. Given its function as a NAD⁺-dependent deacetylase and chromatin regulator, SIRT6 may influence EMT through epigenetic mechanisms, such as histone deacetylation at promoters or enhancers of EMT-related genes, ultimately facilitating transcriptional activation of mesenchymal programs. This raises the possibility that SIRT6 contributes to melanoma progression not only by altering gene expression profiles but also by promoting a more plastic and adaptable cellular state conducive to invasion and metastasis. Interestingly, SIRT6 has been shown to promote EMT-like phenotype switching through a multi-layered mechanism involving, but not limited to, deacetylation of Beclin-1 and FOXO3a in hepatocellular carcinoma cells [12], an increase in the HIF-1α expression in papillary thyroid cancer [14], direct deacetylation of Snail in non-small cell lung cancer [15], and Wnt/β-catenin signaling in prostate cancer [30].

Figure 3.

Effect of SIRT6 knockdown on key mesenchymal markers in human melanoma cells. Relative mRNA expression levels of (A) mesenchymal markers and (B) EMT transcriptional regulators in SIRT6-knockdown (shSIRT6) or non-sense control (shNS) A375 melanoma cells. Data shown as mean ± SEM for two biological and three technical replicates, with **p ≤ 0.01, ***p ≤ 0.001, ****p ≤ 0.0001.

SIRT6 KD in A375 melanoma cells inhibits migration and invasion

To gain a further understanding of the involvement of SIRT6 in melanoma metastasis, we evaluated its contribution to key cellular behaviors associated with metastatic progression, namely migration and invasion [31]. We performed a wound healing scratch assay to study cell migration and a Matrigel invasion assay to study invasion in the shNS and shSIRT6 A375 melanoma cells. The wound healing scratch assay showed marked inhibition of cell migration following SIRT6 KD in the melanoma cells (Figure 4A). Interestingly, 24 h after performing the scratch, the wound was almost 80% closed in shNS A375 control cells, whereas SIRT6 KD (shSIRT6) cells showed significantly less migration (Figure 4B), indicating a marked reduction in migratory potential. These findings strongly suggest that SIRT6 plays a critical role in facilitating melanoma cell motility.

Figure 4.

Effect of shRNA-mediated SIRT6 knockdown on cell migration and invasion ability of human melanoma cells. (A) Scratch assay was performed on the confluent monolayer of SIRT6-knockdown cells (shSIRT6) or non-sense control (shNS) A375 cells. Cell migration towards the mid-line (blue lines) of the scratch was assessed at 0 and 24 h post-scratch. Representative images captured using an EVOS microscope are shown at 20× magnification with a 20 μm scale bar. (B) The percent scratch area was calculated using ImageJ software. Quantitative results are shown in a bar graph with the percent scratch area. The data are expressed as mean ± SEM of two biological and three technical replicates, with statistical significance as determined as *p ≤ 0.05. (C) Representative images of Matrigel-coated transwell invasion assay showing shNS and shSIRT6 A375 cells. For the assessment of invasion potential, cells on the lower surface (invaded cells) were stained with crystal violet staining solution and photographed using an EVOS microscope at 20× with a 20 μm scale bar. (D) Three random fields were selected for manual cell counting. Quantitative results are shown in a bar graph with the number of invaded cells. The data are expressed as mean ± SEM of two biological and three technical replicates with statistical significance as determined by *p ≤ 0.05, **p ≤ 0.01.

Consistent with these observations, the Matrigel invasion assay demonstrated a substantial decrease in invasive capacity upon SIRT6 knockdown (Figure 4C). Quantitative analysis revealed a significant reduction in the number of cells traversing the Matrigel matrix in shSIRT6 cells compared to controls (Figure 4D), underscoring the importance of SIRT6 in promoting extracellular matrix degradation and invasion. Together, these results establish that SIRT6 not only supports melanoma cell movement but also enhances their ability to invade through physiologically relevant barriers, two essential steps in the metastatic cascade.

Importantly, these functional findings align with our molecular data showing that SIRT6 regulates EMT-associated gene expression. The observed attenuation of migration and invasion following SIRT6 depletion is consistent with a reversal or suppression of the mesenchymal phenotype, further supporting the notion that SIRT6 acts as a key modulator of EMT in melanoma. Given that EMT-like events play in melanoma phenotype switching that is inherently linked with cell migration, invasion and metastatic potential [4], our data provide mechanistic insight into how SIRT6 may contribute to melanoma progression from primary to metastatic stages.

Our findings are also consistent with previous reports implicating SIRT6 in promoting migration and invasion in other cancer types, including hepatocellular carcinoma [12] and ovarian cancer [32], where SIRT6 has been shown to regulate pathways associated with cytoskeletal remodeling, matrix interaction, and EMT-like transitions. SIRT6 is highly expressed in cutaneous squamous cell carcinoma (CSCC) cells, and silencing SIRT6 inhibited the proliferation, migration, and invasion abilities of CSCC cells [33]. Further, in a recent publication, SIRT6 has been shown to promote intrahepatic cholangiocarcinoma (ICC), where SIRT6 knockdown inhibited cell viability, proliferation, and colony formation, while its overexpression promoted survival and proliferation of ICC cells [34]. This cross-cancer consistency suggests that SIRT6 may function as a broader regulator of metastatic behavior, although its role remains context-dependent.

Notably, the ability of SIRT6 to influence migration and invasion may have greater clinical relevance than its effects on proliferation alone. While proliferation drives primary tumor growth, metastatic dissemination is the primary cause of melanoma-related mortality. Therefore, the observed suppression of migratory and invasive capacities upon SIRT6 knockdown highlights its potential importance in determining metastatic potential and, by extension, patient outcomes. From a clinical perspective, these findings suggest that targeting SIRT6 could limit metastatic spread by reducing melanoma cell invasiveness, suppress tumor progression by maintaining cells in a less aggressive state, and enhance therapeutic response, as EMT is often linked to resistance to cancer therapies. Importantly, inhibition of SIRT6 does not merely affect proliferation but specifically targets metastatic competency of melanoma cells, which is the primary cause of mortality in melanoma patients. Thus, SIRT6-directed therapies could be particularly valuable in advanced or metastatic disease settings, where controlling dissemination is critical. Furthermore, because EMT contributes to resistance against BRAF/MEK inhibitors and immune checkpoint blockade, combining SIRT6 inhibition with standard therapies may improve treatment efficacy by preventing or reversing resistance mechanisms. Thus, targeting SIRT6 may offer a promising therapeutic approach to restrict melanoma dissemination, overcome resistance, and improve clinical outcomes, particularly when integrated into combination treatment strategies.

Conclusion

Overall, our study supports a model in which SIRT6 promotes a pro-metastatic phenotype in melanoma by enhancing both EMT-associated molecular programs and the functional properties required for tumor cell dissemination. We found that SIRT6 overexpression increases melanoma cell proliferation, long-term survival, and upregulates mesenchymal markers and EMT transcriptional regulators. Further, SIRT6 downregulation showed a decrease in mesenchymal markers and EMT transcriptional regulators, as well as melanoma cell migration and invasion, suggesting that SIRT6 promotes EMT-like phenotype switching, leading to melanoma progression. A proposed diagram of the association between SIRT6 and this phenotypic switching is presented in Figure 5. However, there are some limitations in this study. The experimental framework primarily relies on in vitro melanoma cell models. For SIRT6 overexpression, we used SK-MEL-3 melanoma cells that are NRAS-mutant, non-BRAF–mutant, and for SIRT6 knockdown, we used A375 melanoma cells that harbor the BRAF^V600E mutation. Therefore, these results should be further validated across additional melanoma cell lines and in vivo melanoma models. However, due to the importance of EMT in cancer progression and the relationship we found between SIRT6 modulation and EMT, our findings provide strong mechanistic and translational justification for continued development of SIRT6-targeted therapeutic strategies in melanoma management. Pharmacologic targeting of SIRT6 alone or in combination with existing targeted or immune therapies may limit melanoma progression by simultaneously restricting proliferation and metastatic potential. Our study identifies a critical role for SIRT6 in promoting a mesenchymal, invasive phenotype in melanoma cells and suggests that targeting SIRT6 may therefore represent a promising therapeutic strategy aimed not only at limiting tumor growth but also at reducing metastatic spread.

Figure 5.

Schematic representation of the association of SIRT6 with EMT-like phenotype switching in melanoma progression. Melanoma progression exhibits higher expression of SIRT6 in human melanoma. Upregulation of mesenchymal markers and EMT transcriptional regulators with SIRT6 overexpression and downregulation with SIRT6 knockdown suggests that SIRT6 promotes EMT-like phenotype switching, contributing to an increase in melanoma cell migration and invasion abilities.

Supplementary Material

Supplemental data for this article can be accessed online at https://doi.org/10.1080/29944376.2026.2700162.

Data availability statement

The data that support the findings of this study are available upon request from the corresponding author.