Organismal roles for the PI3Kα and ß isoforms: their specificity, redundancy or cooperation is context-dependent.

PI3Ks are important lipid kinases that produce phosphoinositides phosphorylated in position 3 of the inositol ring. There are three classes of PI3Ks: class I PI3Ks produce PIP3 at plasma membrane level. Although D. melanogaster and C. elegans have only one form of class I PI3K, vertebrates have four class I PI3Ks called isoforms despite being encoded by four different genes. Hence, duplication of these genes coincides with the acquisition of coordinated multi-organ development. Of the class I PI3Ks, PI3Kα and PI3Kß, encoded by PIK3CA and PIK3CB, are ubiquitously expressed. They present similar putative protein domains and share PI(4,5)P2 lipid substrate specificity. Fifteen years after publication of their first isoform-selective pharmacological inhibitors and genetically engineered mouse models (GEMMs) that mimic their complete and specific pharmacological inhibition, we review the knowledge gathered in relation to the redundant and selective roles of PI3Kα and PI3Kß. Recent data suggest that, further to their redundancy, they cooperate for the integration of organ-specific and context-specific signal cues, to orchestrate organ development, physiology, and disease. This knowledge reinforces the importance of isoform-selective inhibitors in clinical settings.

Lymphatic Vasculature Requires Estrogen Receptor-α Signaling to Protect From Lymphedema.

OBJECTIVE: Estrogens exert beneficial effect on the blood vascular system. However, their role on the lymphatic system has been poorly investigated. We studied the protective effect of the 17ß estradiol-the most potent endogenous estrogen-in lymphedema-a lymphatic dysfunction, which results in a massive fluid and fat accumulation in the limb. APPROACH AND RESULTS: Screening of DNA motifs able to mobilize ERs (estrogen receptors) and quantitative real-time polymerase chain reaction analysis revealed that estradiol promotes transcriptional activation of lymphangiogenesis-related gene expression including VEGF (vascular endothelial growth factor)-D, VEGFR (VEGF receptor)-3, lyve-1, and HASs (hyaluronan synthases). Using an original model of secondary lymphedema, we observed a protective effect of estradiol on lymphedema by reducing dermal backflow-a representative feature of the pathology. Blocking ERα by tamoxifen-the selective estrogen modulator-led to a remodeling of the lymphatic network associated with a strong lymphatic leakage. Moreover, the protection of lymphedema by estradiol treatment was abrogated by the endothelial deletion of the receptor ERα in Tie2-Cre; ERαlox/lox mice, which exhibit dilated lymphatic vessels. This remodeling correlated with a decrease in lymphangiogenic gene expression. In vitro, blocking ERα by tamoxifen in lymphatic endothelial cells decreased cell-cell junctions, inhibited migration and sprouting, and resulted in an inhibition of Erk but not of Akt phosphorylation. CONCLUSIONS: Estradiol protection from developing lymphedema is mediated by an activation of its receptor ERα and is antagonized by tamoxifen. These findings reveal a new facet of the estrogen influence in the management of the lymphatic system and provide more evidence that secondary lymphedema is worsened by hormone therapy.

Human genetic disorders of sphingolipid biosynthesis.

Monogenic defects of sphingolipid biosynthesis have been recently identified in human patients. These enzyme deficiencies affect the synthesis of sphingolipid precursors, ceramides or complex glycosphingolipids. They are transmitted as autosomal recessive or dominant traits, and their resulting phenotypes often replicate the abnormalities seen in murine models deficient for the corresponding enzymes. In quite good agreement with the known critical roles of sphingolipids in cells from the nervous system and the epidermis, these genetic defects clinically manifest as neurological disorders, including paraplegia, epilepsy or peripheral neuropathies, or present with ichthyosis. The present review summarizes the genetic alterations, biochemical changes and clinical symptoms of this new group of inherited metabolic disorders. Hypotheses regarding the molecular pathophysiology and potential treatments of these diseases are also discussed.

New insights into the organ-specific adverse effects of fumonisin B1: comparison between lung and liver.

Fumonisin B1 (FB1) is a well-known inhibitor of de novo sphingolipid biosynthesis, due to its ability to inhibit ceramide synthases (CerS) activity. In mammals, this toxin triggers broad clinical symptoms with multi-organ dysfunction such as hepatotoxicity or pulmonary edema. The molecular mechanism of CerS inhibition by FB1 remains unknown. Due to the existence of six mammalian CerS isoforms with a tissue-specific expression pattern, we postulated that the organ-specific adverse effects of FB1 might be due to different CerS isoforms. The sphingolipid contents of lung and liver were compared in normal and FB1-exposed piglets (gavage with 1.5 mg FB1/kg body weight daily for 9 days). The effect of the toxin on each CerS was deduced from the analysis of its effects on individual ceramide (Cer) and sphingomyelin (SM) species. As expected, the total Cer content decreased by half in the lungs of FB1-exposed piglets, while in contrast, total Cer increased 3.5-fold in the livers of FB1-exposed animals. Our data also indicated that FB1 is more prone to bind to CerS4 and CerS2 to deplete lung and to enrich liver in d18:1/C20:0 and d18:1/C22:0 ceramides. It also interact with CerS1 to enrich liver in d18:1/C18:0 ceramides. Cer levels were counterbalanced by those of SM. In conclusion, these results demonstrate that the specificity of the effects of FB1 on tissues and organs is due to the effects of the toxin on CerS4, CerS2, and CerS1.

Ceramide synthase 4 and de novo production of ceramides with specific N-acyl chain lengths are involved in glucolipotoxicity-induced apoptosis of INS-1 ß-cells.

Pancreatic ß-cell apoptosis induced by palmitate requires high glucose concentrations. Ceramides have been suggested to be important mediators of glucolipotoxicity-induced ß-cell apoptosis. In INS-1 ß-cells, 0.4 mM palmitate with 5 mM glucose increased the levels of dihydrosphingosine and dihydroceramides, two lipid intermediates in the de novo biosynthesis of ceramides, without inducing apoptosis. Increasing glucose concentrations to 30 mM amplified palmitate-induced accumulation of dihydrosphingosine and the formation of (dihydro)ceramides. Of note, glucolipotoxicity specifically induced the formation of C(18:0), C(22:0) and C(24:1) (dihydro)ceramide molecular species, which was associated with the up-regulation of CerS4 (ceramide synthase 4) levels. Fumonisin-B1, a ceramide synthase inhibitor, partially blocked apoptosis induced by glucolipotoxicity. In contrast, apoptosis was potentiated in the presence of D,L-threo-1-phenyl-2-palmitoylamino-3-morpholinopropan-1-ol, an inhibitor of glucosylceramide synthase. Moreover, overexpression of CerS4 amplified ceramide production and apoptosis induced by palmitate with 30 mM glucose, whereas down-regulation of CerS4 by siRNA (short interfering RNA) reduced apoptosis. CerS4 also potentiates ceramide accumulation and apoptosis induced by another saturated fatty acid: stearate. Collectively, our results suggest that glucolipotoxicity induces ß-cell apoptosis through a dual mechanism involving de novo ceramide biosynthesis and the formation of ceramides with specific N-acyl chain lengths rather than an overall increase in ceramide content.

Potential Role of Sphingolipidoses-Associated Lysosphingolipids in Cancer.

Sphingolipids play a key structural role in cellular membranes and/or act as signaling molecules. Inherited defects of their catabolism lead to lysosomal storage diseases called sphingolipidoses. Although progress has been made toward a better understanding of their pathophysiology, several issues still remain unsolved. In particular, whether lysosphingolipids, the deacylated form of sphingolipids, both of which accumulate in these diseases, are simple biomarkers or play an instrumental role is unclear. In the meanwhile, evidence has been provided for a high risk of developing malignancies in patients affected with Gaucher disease, the most common sphingolipidosis. This article aims at analyzing the potential involvement of lysosphingolipids in cancer. Knowledge about lysosphingolipids in the context of lysosomal storage diseases is summarized. Available data on the nature and prevalence of cancers in patients affected with sphingolipidoses are also reviewed. Then, studies investigating the biological effects of lysosphingolipids toward pro or antitumor pathways are discussed. Finally, original findings exploring the role of glucosylsphingosine in the development of melanoma are presented. While this lysosphingolipid may behave like a protumorigenic agent, further investigations in appropriate models are needed to elucidate the role of these peculiar lipids, not only in sphingolipidoses but also in malignant diseases in general.

FAN stimulates TNF(alpha)-induced gene expression, leukocyte recruitment, and humoral response.

Factor associated with neutral sphingomyelinase activation (FAN) is an adaptor protein that constitutively binds to TNF-R1. Microarray analysis was performed in fibroblasts derived from wild-type or FAN knockout mouse embryos to evaluate the role of FAN in TNF-induced gene expression. Approximately 70% of TNF-induced genes exhibited lower expression levels in FAN-deficient than in wild-type fibroblasts. Of particular interest, TNF-induced expression of cytokines/chemokines, such as IL-6 and CXCL-2, was impaired in FAN-deficient cells. This was confirmed by real time RT-PCR and ELISA. Upon i.p. TNF or thioglycollate injection, neutrophil recruitment into the peritoneal cavity was reduced by more than 50% in FAN-deficient mice. Nevertheless, FAN-deficient animals did not exhibit an increased susceptibility to different microorganisms including bacteria and parasites, indicating that FAN is not essential for pathogen clearance. Specific Ab response to BSA was substantially impaired in FAN-deficient mice and this was associated with a reduced content of leukocytes in the spleen of BSA-challenged FAN-deficient mice as compared with their wild-type counterparts. Altogether, our results indicate the involvement of FAN in TNF-induced gene expression and leukocyte recruitment, contributing to the establishment of the specific immune response.

Apolipoprotein E-deficient mice develop an anti-Chlamydophila pneumoniae T helper 2 response and resist vascular infection.

BACKGROUND: Hypercholesterolemia could inhibit the immune response against various pathogens. No information is available about its impact on the immune response toward Chlamydophila pneumoniae. METHODS: Apolipoprotein E (apoE)-deficient and wild-type mice fed a normal diet were infected with a single intranasal inoculation of viable C. pneumoniae. RESULTS: Whereas interferon gamma concentrations (T helper 1 response) were similar in the lungs and spleen of apoE-deficient and wild-type mice, increased concentrations of interleukin 10, interleukin 6, and interleukin 4 (T helper 2 response) were found in the lungs of apoE-deficient mice. The spleen B lymphocyte percentage and interleukin 4 levels and serum specific antibody titers were higher in apoE-deficient mice. C. pneumoniae infection was facilitated neither in the lungs nor in the aorta of these mice. On the contrary, the number of apoE-deficient mice with detectable levels of bacterial DNA in the aorta was clearly decreased. When low-density lipoprotein receptor-deficient mice fed a normal diet were similarly infected, no difference in the interleukin 4 concentration and infection level was observed in the lungs and no protection was found in the aorta. CONCLUSIONS: Mild hypercholesterolemia in mice does not facilitate C. pneumoniae persistence in the vascular wall. ApoE deficiency, rather than mild hypercholesterolemia, probably favors the development of an unusual anti-C. pneumoniae T helper 2 response and protects against vascular infection.

Phosphoproteomics Identifies PI3K Inhibitor-selective Adaptive Responses in Pancreatic Cancer Cell Therapy and Resistance.

The PI3K pathway is highly active in human cancers. The four class I isoforms of PI3K are activated by distinct mechanisms leading to a common downstream signaling. Their downstream redundancy is thought to be responsible for treatment failures of PI3K inhibitors. We challenged this concept, by mapping the differential phosphoproteome evolution in response to PI3K inhibitors with different isoform-selectivity patterns in pancreatic cancer, a disease currently without effective therapy. In this cancer, the PI3K signal was shown to control cell proliferation. We compared the effects of LY294002 that inhibit with equal potency all class I isoenzymes and downstream mTOR with the action of inhibitors with higher isoform selectivity toward PI3Kα, PI3Kß, or PI3Kγ (namely, A66, TGX-221 and AS-252424). A bioinformatics global pathway analysis of phosphoproteomics data allowed us to identify common and specific signals activated by PI3K inhibitors supported by the biological data. AS-252424 was the most effective treatment and induced apoptotic pathway activation as well as the highest changes in global phosphorylation-regulated cell signal. However, AS-252424 treatment induced reactivation of Akt, therefore decreasing the treatment outcome on cell survival. Reversely, AS-252424 and A66 combination treatment prevented p-Akt reactivation and led to synergistic action in cell lines and patient organoids. The combination of clinically approved α-selective BYL-719 with γ-selective IPI-549 was more efficient than single-molecule treatment on xenograft growth. Mapping unique adaptive signaling responses to isoform-selective PI3K inhibition will help to design better combinative treatments that prevent the induction of selective compensatory signals.

Pancreatic cancer intrinsic PI3Kα activity accelerates metastasis and rewires macrophage component.

Pancreatic ductal adenocarcinoma (PDAC) patients frequently suffer from undetected micro-metastatic disease. This clinical situation would greatly benefit from additional investigation. Therefore, we set out to identify key signalling events that drive metastatic evolution from the pancreas. We searched for a gene signature that discriminate localised PDAC from confirmed metastatic PDAC and devised a preclinical protocol using circulating cell-free DNA (cfDNA) as an early biomarker of micro-metastatic disease to validate the identification of key signalling events. An unbiased approach identified, amongst actionable markers of disease progression, the PI3K pathway and a distinctive PI3Kα activation signature as predictive of PDAC aggressiveness and prognosis. Pharmacological or tumour-restricted genetic PI3Kα-selective inhibition prevented macro-metastatic evolution by hindering tumoural cell migratory behaviour independently of genetic alterations. We found that PI3Kα inhibition altered the quantity and the species composition of the produced lipid second messenger PIP3 , with a selective decrease of C36:2 PI-3,4,5-P3 . Tumoural PI3Kα inactivation prevented the accumulation of pro-tumoural CD206-positive macrophages in the tumour-adjacent tissue. Tumour cell-intrinsic PI3Kα promotes pro-metastatic features that could be pharmacologically targeted to delay macro-metastatic evolution.

Neutral Sphingomyelinase 2 Heightens Anti-Melanoma Immune Responses and Anti-PD-1 Therapy Efficacy.

Dysregulation of lipid metabolism affects the behavior of cancer cells, but how this happens is not completely understood. Neutral sphingomyelinase 2 (nSMase2), encoded by SMPD3, catalyzes the breakdown of sphingomyelin to produce the anti-oncometabolite ceramide. We found that this enzyme was often downregulated in human metastatic melanoma, likely contributing to immune escape. Overexpression of nSMase2 in mouse melanoma reduced tumor growth in syngeneic wild-type but not CD8-deficient mice. In wild-type mice, nSMase2-overexpressing tumors showed accumulation of both ceramide and CD8+ tumor-infiltrating lymphocytes, and this was associated with increased level of transcripts encoding IFNγ and CXCL9. Overexpressing the catalytically inactive nSMase2 failed to alter tumor growth, indicating that the deleterious effect nSMase2 has on melanoma growth depends on its enzymatic activity. In vitro, small extracellular vesicles from melanoma cells overexpressing wild-type nSMase2 augmented the expression of IL12, CXCL9, and CCL19 by bone marrow-derived dendritic cells, suggesting that melanoma nSMase2 triggers T helper 1 (Th1) polarization in the earliest stages of the immune response. Most importantly, overexpression of wild-type nSMase2 increased anti-PD-1 efficacy in murine models of melanoma and breast cancer, and this was associated with an enhanced Th1 response. Therefore, increasing SMPD3 expression in melanoma may serve as an original therapeutic strategy to potentiate Th1 polarization and CD8+ T-cell-dependent immune responses and overcome resistance to anti-PD-1.

Palmitoyl protein thioesterase 1 modulates tumor necrosis factor alpha-induced apoptosis.

Induction of apoptosis by TNF has recently been shown to implicate proteases from lysosomal origin, the cathepsins. Here, we investigated the role in apoptosis of palmitoyl protein thioesterase 1 (PPT1), another lysosomal enzyme that depalmitoylates proteins. We show that transformed fibroblasts derived from patients with the infantile form of neuronal ceroid lipofuscinosis (INCL), a neurodegenerative disease due to deficient activity of PPT1, are partially resistant to TNF-induced cell death (57-75% cell viability vs. 15-30% for control fibroblasts). TNF-initiated proteolytic cleavage of caspase-8, Bid and caspase-3, as well as cytochrome c release was strongly attenuated in INCL fibroblasts as compared to control cells. Noteworthy, activation of p42/p44 mitogen-activated protein kinase and of transcription factor NF-kappaB by TNF, and induction of cell death by staurosporine or chemotherapeutic drugs in INCL cells were unaffected by PPT1 deficiency. Resistance to TNF-induced apoptosis was also observed in embryonic fibroblasts derived from Ppt1/Cln1-deficient mice but not from mice with a targeted deletion of Cln3 or Cln5. Finally, reconstitution of PPT1 activity in mutant cells was accompanied by resensitization to TNF-induced caspase activation and toxicity. These observations emphasize for the first time the role of PPT1 and, likely, protein depalmitoylation in the regulation of TNF-induced apoptosis.

Are Glucosylceramide-Related Sphingolipids Involved in the Increased Risk for Cancer in Gaucher Disease Patients? Review and Hypotheses.

The roles of ceramide and its catabolites, i.e., sphingosine and sphingosine 1-phosphate, in the development of malignancies and the response to anticancer regimens have been extensively described. Moreover, an abundant literature points to the effects of glucosylceramide synthase, the mammalian enzyme that converts ceramide to ß-glucosylceramide, in protecting tumor cells from chemotherapy. Much less is known about the contribution of ß-glucosylceramide and its breakdown products in cancer progression. In this chapter, we first review published and personal clinical observations that report on the increased risk of developing cancers in patients affected with Gaucher disease, an inborn disorder characterized by defective lysosomal degradation of ß-glucosylceramide. The previously described mechanistic links between lysosomal ß-glucosylceramidase, ß-glucosylceramide and/or ß-glucosylphingosine, and various hallmarks of cancer are reviewed. We further show that melanoma tumor growth is facilitated in a Gaucher disease mouse model. Finally, the potential roles of the ß-glucosylceramidase protein and its lipidic substrates and/or downstream products are discussed.

Experimental pancreatic cancer develops in soft pancreas: novel leads for an individualized diagnosis by ultrafast elasticity imaging.

Rapid, easy and early pancreatic cancer diagnosis and therapeutic follow up continue to necessitate an increasing attention towards the development of effective treatment strategies for this lethal disease. The non invasive quantitative assessment of pancreatic heterogeneity is limited. Here, we report the development of a preclinical imaging protocol using ultrasonography and shear wave technology in an experimental in situ pancreatic cancer model to measure the evolution of pancreatic rigidity. Methods: Intrapancreatic tumors were genetically induced by mutated Kras and p53 in KPC mice. We evaluated the feasiblity of a live imaging protocol by assessing pancreas evolution with Aixplorer technology accross 36 weeks. Lethality induced by in situ pancreatic cancer was heterogeneous in time. Results: The developed method successfully detected tumor mass from 26 weeks onwards at minimal 0.029 cm3 size. Elastography measurements using shear wave methodology had a wide detection range from 4.7kPa to 166.1kPa. Protumorigenic mutations induced a significant decrease of the rigidity of pancreatic tissue before tumors developed in correlation with the detection of senescent marker p16-positive cells. An intratumoral increased rigidity was quantified and found surprisingly heterogeneous. Tumors also presented a huge inter-individual heterogeneity in their rigidity parameters; tumors with low and high rigidity at detection evolve very heterogeneously in their rigidity parameters, as well as in their volume. Increase in rigidity in tumors detected by ultrafast elasticity imaging coincided with detection of tumors by echography and with the detection of the inflammatory protumoral systemic condition by non invasive follow-up and of collagen fibers by post-processing tumoral IHC analysis. Conclusion: Our promising results indicate the potential of the shear wave elastography to support individualization of diagnosis in this most aggressive disease.

Paclitaxel induces lymphatic endothelial cells autophagy to promote metastasis.

Cytotoxic therapy for breast cancer inhibits the growth of primary tumors, but promotes metastasis to the sentinel lymph nodes through the lymphatic system. However, the effect of first-line chemotherapy on the lymphatic endothelium has been poorly investigated. In this study, we determined that paclitaxel, the anti-cancer drug approved for the treatment of metastatic or locally advanced breast cancer, induces lymphatic endothelial cell (LEC) autophagy to increase metastases. While paclitaxel treatment was largely efficacious in inhibiting LEC adhesion, it had no effect on cell survival. Paclitaxel inhibited LEC migration and branch point formation by inducing an autophagy mechanism independent of Akt phosphorylation. In vivo, paclitaxel mediated a higher permeability of lymphatic endothelium to tumor cells and this effect was reversed by chloroquine, an autophagy-lysosome inhibitor. Despite a strong effect on reducing tumor size, paclitaxel significantly increased metastasis to the sentinel lymph nodes. This effect was restricted to a lymphatic dissemination, as chemotherapy did not affect the blood endothelium. Taken together, our findings suggest that the lymphatic system resists to chemotherapy through an autophagy mechanism to promote malignant progression and metastatic lesions. This study paves the way for new combinative therapies aimed at reducing the number of metastases.

Pharmacodynamic monitoring of the conversion from mycophenolate mofetil to enteric-coated mycophenolate sodium in stable kidney-allograft recipients.

BACKGROUND: The formulations of mycophenolic acid, i.e., mycophenolate mofetil (MMF) and enteric-coated mycophenolate sodium (EC-MPS), seem to have different pharmacokinetic profiles. The aim of this study was to compare the effects MMF and EC-MPS on T-cell proliferation, T-cell activation, T-cell function, and lymphocyte subsets. CLINICAL STUDY AND METHODS: Ten stable kidney-transplant patients on standard maintenance therapy of tacrolimus and MMF (1 g/d), with or without steroids, were converted from MMF to EC-MPS at equivalent dose (720 mg/d). Tacrolimus and steroid doses remained unchanged before, and at 1, 2, 3, and 6 months (M) after conversion. Intra T-lymphocyte cytokines IL-2 and TNF-alpha, lymphocyte-activation surface markers (CD25 and CD71), T-cell proliferation (PCNA+ PI(high)), total lymphocyte count, as well as lymphocytes subsets (CD2, CD3, CD4, CD8, CD19, NK cells) were measured by flow cytometry before conversion and at M1, M2, M3, and M6. RESULTS: We found no significant differences of MMF versus EC-MPS on lymphocyte function. T-cell proliferation and T-cell activation (CD25 and CD71 expression), but not cytokine expression (TNF-alpha and IL-2), showed a trend to increase after conversion from MMF to EC-MPS. Total lymphocyte, CD2, CD3, CD4, CD8, and NK cells counts were not significantly modified. CONCLUSION: This study revealed a trend to a lower immunosuppression with EC-MPS as compared to MMF in stable renal transplant patients.

Signal-Targeted Therapies and Resistance Mechanisms in Pancreatic Cancer: Future Developments Reside in Proteomics.

For patients with metastatic pancreatic cancer that are not eligible for surgery, signal-targeted therapies have so far failed to significantly improve survival. These therapeutic options have been tested in phase II/III clinical trials mostly in combination with the reference treatment gemcitabine. Innovative therapies aim to annihilate oncogenic dependency, or to normalize the tumoural stroma to allow immune cells to function and/or re-vascularisation to occur. Large scale transcriptomic and genomic analysis revealed that pancreatic cancers display great heterogeneity but failed to clearly delineate specific oncogene dependency, besides oncogenic Kras. Beyond these approaches, proteomics appears to be an appropriate approach to classify signal dependency and to identify specific alterations at the targetable level. However, due to difficulties in sampling, proteomic data for this pathology are scarce. In this review, we will discuss the current state of clinical trials for targeted therapies against pancreatic cancer. We will then highlight the most recent proteomic data for pancreatic tumours and their metastasis, which could help to identify major oncogenic signalling dependencies, as well as provide future leads to explain why pancreatic tumours are intrinsically resistant to signal-targeted therapies. We will finally discuss how studies on phosphatidylinositol-3-kinase (PI3K) signalling, as the paradigmatic pro-tumoural signal downstream of oncogenic Kras in pancreatic cancer, would benefit from exploratory proteomics to increase the efficiency of targeted therapies.

Glucosylceramidases and malignancies in mammals.

Sphingolipids represent a major class of lipids that are essential constituents of eukaryotic cells. They are predominantly located in plasma membrane microdomains, and play an important structural role in regulating membrane fluidity. They are also bioactive effectors involved in diverse key cellular functions such as apoptosis and proliferation. The implication of some sphingolipids in cancer is well established whereas that of some others is still a matter of intense investigation. Glucosylceramide is the backbone of more than 300 structurally different glycosphingolipids including gangliosides and sulfatides, and is essential for mammalian development. Therefore, glucosylceramidases (also named GBA1, GBA2 and GBA3 ß-glucosidases), the enzymes that hydrolyse ß-glucosylceramide, play important functions. GBA1 is a lysosomal hydrolase whose deficiency causes Gaucher disease, the most prevalent inherited lysosomal storage disorder. GBA2 is a ubiquitous non-lysosomal glucosylceramidase whose mutations have been associated with some forms of hereditary spastic paraplegia. GBA3 is a cytosolic ß-glucosidase, mostly present in the kidney, liver, spleen, intestine and lymphocytes of mammals, the function of which is still unclear. Whereas glucosylceramide synthase is implicated in multidrug resistance, the role of glucosylceramide breakdown in cancer is not yet fully appreciated. Defective GBA1 enzyme activity in humans, i.e., Gaucher disease, is associated with an increased risk of multiple myeloma and other malignancies. Putative molecular links between Gaucher disease and cancer, which might implicate the malignant cell and/or its microenvironment, are reviewed. The functions of GBA2 and GBA3 in cancer progression are also discussed.

Downregulation of sphingosine kinase-1 induces protective tumor immunity by promoting M1 macrophage response in melanoma.

The infiltration of melanoma tumors by macrophages is often correlated with poor prognosis. However, the molecular signals that regulate the dialogue between malignant cells and the inflammatory microenvironment remain poorly understood. We previously reported an increased expression of sphingosine kinase-1 (SK1), which produces the bioactive lipid sphingosine 1-phosphate (S1P), in melanoma. The present study aimed at defining the role of tumor SK1 in the recruitment and differentiation of macrophages in melanoma. Herein, we show that downregulation of SK1 in melanoma cells causes a reduction in the percentage of CD206highMHCIIlow M2 macrophages in favor of an increased proportion of CD206lowMHCIIhigh M1 macrophages into the tumor. This macrophage differentiation orchestrates T lymphocyte recruitment as well as tumor rejection through the expression of Th1 cytokines and chemokines. In vitro experiments indicated that macrophage migration is triggered by the binding of tumor S1P to S1PR1 receptors present on macrophages whereas macrophage differentiation is stimulated by SK1-induced secretion of TGF-ß1. Finally, RNA-seq analysis of human melanoma tumors revealed a positive correlation between SK1 and TGF-ß1 expression. Altogether, our findings demonstrate that melanoma SK1 plays a key role in the recruitment and phenotypic shift of the tumor macrophages that promote melanoma growth.