Unité Mixte de Recherche Intitut National de la Santé et de la Recherche Médicale (INSERM) 1037, Centre National de la Recherche Scientifique (CNRS) 5071, Université Toulouse III - Paul Sabatier, Centre de Recherches en Cancérologie de Toulouse (CRCT), Toulouse, France.
Équipe labellisée Fondation Association (ARC), Toulouse, France.
Front Immunol. 2024 Jul 29;15:1421432. doi: 10.3389/fimmu.2024.1421432. eCollection 2024.
Advanced cutaneous melanoma is a skin cancer characterized by a poor prognosis and high metastatic potential. During metastatic spread, melanoma cells often undergo dedifferentiation toward an invasive phenotype, resulting in reduced expression of microphthalmia-associated transcription factor (MITF)-dependent melanoma antigens and facilitating immune escape. Tumor Necrosis Factor (TNF) is known to be a key factor in melanoma dedifferentiation. Interestingly, accumulating evidence suggests that TNF may play a role in melanoma progression and resistance to immunotherapies. Additionally, TNF has been identified as a potent regulator of sphingolipid metabolism, which could contribute to melanoma aggressiveness and the process of melanoma dedifferentiation.
We conducted RNA sequencing and mass spectrometry analyses to investigate TNF-induced dedifferentiation in two melanoma cell lines. experiments were performed to manipulate sphingolipid metabolism using genetic or pharmacologic alterations in combination with TNF treatment, aiming to elucidate the potential involvement of this metabolism in TNF-induced dedifferentiation. Lastly, to evaluate the clinical significance of our findings, we performed unsupervised analysis of plasma sphingolipid levels in 48 patients receiving treatment with immune checkpoint inhibitors, either alone or in combination with anti-TNF therapy.
Herein, we demonstrate that TNF-induced melanoma cell dedifferentiation is associated with a global modulation of sphingolipid metabolism. Specifically, TNF decreases the expression and activity of acid ceramidase (AC), encoded by the gene, while increasing the expression of glucosylceramide synthase (GCS), encoded by the gene. Remarkably, knockdown of AC alone via RNA interference is enough to induce melanoma cell dedifferentiation. Furthermore, treatment with Eliglustat, a GCS inhibitor, inhibits TNF-induced melanoma cell dedifferentiation. Lastly, analysis of plasma samples from patients treated with immune checkpoint inhibitors, with or without anti-TNF therapy, revealed significant predictive sphingolipids. Notably, the top 8 predictive sphingolipids, including glycosphingolipids, were associated with a poor response to immunotherapy.
Our study highlights that ceramide metabolism alterations are causally involved in TNF-induced melanoma cell dedifferentiation and suggests that the evolution of specific ceramide metabolites in plasma may be considered as predictive biomarkers of resistance to immunotherapy.
晚期皮肤黑色素瘤是一种预后不良、转移潜能高的皮肤癌。在转移扩散过程中，黑色素瘤细胞通常向侵袭表型去分化，导致小眼畸形相关转录因子（MITF）依赖性黑色素瘤抗原表达减少，并促进免疫逃逸。肿瘤坏死因子（TNF）被认为是黑色素瘤去分化的关键因素。有趣的是，越来越多的证据表明，TNF 可能在黑色素瘤的进展和对免疫疗法的耐药性中发挥作用。此外，TNF 已被确定为鞘脂代谢的有效调节剂，这可能有助于黑色素瘤的侵袭性和黑色素瘤去分化的过程。
我们通过 RNA 测序和质谱分析研究了两种黑色素瘤细胞系中 TNF 诱导的去分化。通过遗传或药理学改变操纵鞘脂代谢，并结合 TNF 处理，进行实验，旨在阐明这种代谢在 TNF 诱导的去分化中的潜在作用。最后，为了评估我们研究结果的临床意义，我们对 48 名接受免疫检查点抑制剂治疗的患者的血浆鞘脂水平进行了无监督分析，这些患者单独或联合使用抗 TNF 治疗。
本研究表明，TNF 诱导的黑色素瘤细胞去分化与鞘脂代谢的全局调节有关。具体来说，TNF 降低了基因编码的酸性神经酰胺酶（AC）的表达和活性，同时增加了基因编码的葡萄糖神经酰胺合酶（GCS）的表达。值得注意的是，通过 RNA 干扰单独敲低 AC 足以诱导黑色素瘤细胞去分化。此外，用 GCS 抑制剂 Eliglustat 治疗可抑制 TNF 诱导的黑色素瘤细胞去分化。最后，对接受免疫检查点抑制剂治疗的患者的血浆样本进行分析，无论是否联合抗 TNF 治疗，都揭示了具有显著预测作用的鞘脂。值得注意的是，前 8 个具有预测作用的鞘脂，包括糖脂，与免疫治疗反应不良相关。
我们的研究强调了神经酰胺代谢的改变在 TNF 诱导的黑色素瘤细胞去分化中起着因果关系，并表明血浆中特定神经酰胺代谢物的演变可以被认为是对免疫治疗耐药的预测生物标志物。
