Study of the role of glucosylceramide in cutaneous melanoma

Cutaneous melanoma (CM) is the most severe skin cancer whom the incidence is keeping growing. Despite of the remarkable improvement of CM management thanks to the development of targeted therapies and immune checkpoint inhibitors, the prognostic of CM patients remains still bad. Some
alterations in sphingolipid (SL) metabolism have been described in CM by our research team and others. These alterations could be involved in the tumoral development and the drug resistance, and hinge on a decrease of ceramide production, considered as an anti-oncometabolite and on an increase of sphingosine-1-phosphate production, considered as an oncométabolite. Glucosylceramidase (GCase), encoded by GBA gene, is a lysosomal enzyme, which hydrolyze glucosylceramide (GlcCer) into ceramide, the last step of SL catabolism. The analysis of exome from MC patients has revealed missense mutations in GBA gene. GCase deficiency results in Gaucher disease (GD), a lysosomal storage disease, in which patients display an organomegaly with or without a neurological involvement. The clinical presentation would result from the accumulation of undegraded GlcCer and of GlcSph corresponding with its deacylated form. Moreover, the patient with GD have an increase of risk for developing a cancer, including CM. The involvement of these lipids is evoked. On the other hand, mutations in GBA gene at homozygous or heterozygous state are a risk factor for developing Parkinson disease (PD), a neurodegenerative disorder, in which, alpha-synuclein aggregates are found in neurons. The accumulation of GlcCer and GlcSph, due to GCase deficiency, would promote alpha-synuclein aggregation. Interestingly, patients with PD have also a greater risk for developing CM. Thus, alphasynuclein aggregates, secondary to GCase deficiency, could be involved in CM development.

Mechanisms underlying between GD and CM remains elusive. Here, our aim is to determine the role of GCase in CM development. For that, human melanoma cell lines (A375) have been used, in which the expression and the catalytic activity of GCase have been reduced by either RNA silencing (shRNA against GBA) or by pharmacological inhibition (Conduritol B Epoxide), and A375 cell lines in which GCase wild type and mutated have been overexpressed by lentiviral transduction. In addition, A375 cell line have been treated by GlcSph in order to study its involvement in cancer development. Cancer cell properties such as cell proliferation, cell cycle and response to drugs have been tested as well as a SL analysis. Alpha-synuclein expression has been also assessed in these different cellular models.

We have shown that the inhibition of expression and/or activity of GCase do not modify cancer cell properties. However, cells treated by GlcSph exhibit a temporary and reversible decrease of cell growth and some of them have become giant multinucleated cells but their function is unknown. As
for assessment of alpha-synuclein expression, technical problems have us hampered to conclude.

Our preliminary results show a potential effect of GlcSph on development of CM and some indications about GlcSph metabolism.

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Thesis defended on 14 December 2021

Find the full text of the thesis on the Paul Sabatier University website