EP4746876A1

EP4746876A1 - Udp-glucose ceramide glucosyltransferase synthase (ugcg) inhibitors in the treatment of cancer

Info

Publication number: EP4746876A1
Application number: EP24748042.9A
Authority: EP
Inventors: Jaroslav Belotserkovsky; Jonas DEHAIRS; Vincent DE LAAT; Jakub IDKOWIAK; Johannes SWINNEN; Ali Talebi; Frank VANDERHOYDONC
Original assignee: KU Leuven
Current assignee: KU Leuven
Priority date: 2023-07-20
Filing date: 2024-07-22
Publication date: 2026-05-27
Also published as: WO2025017212A1; JP2026526781A

Abstract

The invention relates to chemotherapy agents such as gemcitabine and and inhibitors of UDP-glucose ceramide glucosyltransferase synthase (UGCG) such as miglustat, lucerastat, sinbaglustat, eliglustat, ibiglustat, for use in the treatment of cancers, such as pancreatic cancers.

Description

UDP-GLUCOSE CERAMIDE GLUCOSYLTRANSFERASE SYNTHASE (UGCG) INHIBITORS IN THE TREATMENT OF CANCER

Field of the invention

The invention relates to cancer treatment, more particular pancreatic cancer treatment.

The invention further relates to the combined use of

Background of the invention

Based on the clinical staging procedures for pancreatic cancer, a tumour is classified as resectable, borderline resectable, locally advanced, or metastatic disease. A treatment decision is made based on these findings, but also considers the general and nutritional status of the patient.

For pancreatic cancer that has not spread beyond the pancreas, surgical excision remains the primary treatment option. It is also the only effective treatment, although the 5-year survival rate is about 20%. Approximately 60%-70% of pancreatic cancers arise in the head of the pancreas, 20%-25% in the body and tail, and the remaining 10%-20% diffusely involve the pancreas3. Pancreatic cancer is a term for a collection of diseases that mark distinct subtypes of tumours that originate in the pancreas. The classification of pancreatic cancer subtypes is based on histopathological characteristics and cellular origin. The two main subtypes are exocrine and endocrine tumours. Endocrine tumours account for a minority (5%) of pancreatic malignancies. Endocrine tumours, also termed pancreatic neuroendocrine tumours (PNETs), develop from the endocrine cells that produce hormones. PNETs can be further classified into a number of subtypes depending on the type of hormone they generate, including insulinomas, gastrinomas, and glucagonomas. Exocrine tumours are by far (95%) the most common type of pancreatic cancer. They develop in the exocrine tissue that produces digestive enzymes, and may arise from ductal epithelium, acinar cells, or connective tissue. Adenocarcinoma is the most common type of exocrine pancreatic cancer, and accounts for about 80% of all pancreatic cancers. The term pancreatic cancer is often used to refer to pancreatic ductal adenocarcinoma (PDAC). Moreover, only about 20% of patients with pancreatic ductal adenocarcinoma (PDAC) have a tumour that is regarded as resectable at the time of diagnosis. For resection, pancreaticoduodenectomy (Whipple procedure) is the most commonly used surgical procedure. It involves the removal of the pancreatic head, duodenum, gallbladder and bile duct. However, depending on the tumour location, surgery might entail distal pancreatectomy (removing the pancreatic tail and body) or total pancreatectomy. Given the poor results of surgery alone, patients receive adjuvant (after surgery) chemotherapy to improve survival. The use of FOLFIRINOX, gemcitabine/capecitabine, or gemcitabine monotherapy is considered as standard of care for this setting, depending on patient fitness. Although adjuvant chemotherapy leads to increased survival, only a minority of patients with resectable pancreatic cancer achieve long-term survival.

In 30%-40% of patients, the tumour is confined to the pancreatic region but resection is not possible due to vascular invasion. These tumours are classified as borderline- resectable disease, and neoadjuvant (administered before surgery) chemotherapy (gemcitabine, gemcitabine plus nab-paclitaxel, gemcitabine and cisplatin, or FOLFIRINOX (a triplet chemotherapy regimen combining 5-fluorouracil (5-FU; a DNA replication inhibitor), irinotecan (a topoisomerase I inhibitor) and oxaliplatin (a platinum based antineoplastic agent) with folinic acid (a vitamin B derivative that may enhance the effect of 5-FU)) has been shown to improve overall survival, although unexpectedly without increasing resection rate [Springfield et al. (2023) Nat Rev Clin Oncol 20, 318-337]. Additional radiation therapy has been assessed in multiple trials and shows promise but the practice currently suffers from lack of definitive data [Hall et al. (2021) JCO 39, 3773-3777],

When there are no metastases and the tumour is not considered resectable or borderline resectable, the tumour is considered as locally advanced. The optimal treatment of this group of patients is still highly debatable. The standard of care is gemcitabine, but in some studies chemoradiation has been shown to slightly improve outcome for this subgroup of patients. However, all randomized trials combining radiotherapy and chemotherapy in patients with locally advanced cancer have failed to meet their primary end points. Regardless of treatment strategy, the average overall survival for these patients remains low.

Protein-bound paclitaxel, also known as nanoparticle albumin-bound paclitaxel (nab- paclitaxel), is an injectable formulation of paclitaxel used to treat breast cancer, lung cancer and pancreatic cancer, among others. Paclitaxel kills cancer cells by preventing the normal breakdown of microtubules during cell division. Nab-paclitaxel is designated as first-line treatment, in combination with gemcitabine, for metastatic adenocarcinoma of the pancreas. A combination of nab-paclitaxel with capecitabine (a 5-FU prodrug) has been evaluated in a phase II clinical trial in patients with metastatic pancreatic adenocarcinoma [Scheithauer et al. (2016) J Gastrointest Oncol 7, 234-238], The majority of patients with pancreatic cancer progress to metastatic disease and about 50% of the patients are initially diagnosed with distant metastatic disease (stage IV PDAC), for which the 5-year survival rate is merely 3%. These patients often require palliative and supportive care to relieve biliary and/or duodenal obstruction, malnutrition, and pain. For oncologic treatment, gemcitabine monotherapy was established as the standard of care in 1997 [Burris et al. (1997) J Clin Oncol. 15, 2403-2413]. The demonstration of the efficacy of FOLFIRINOX resulted in significant advances in the treatment of metastatic disease. Despite an increase in toxicity, the FOLFIRINOX regimen has proven to be more effective than gemcitabine alone [Conroy et al. (2011) /V. Eng. J. Med. 364, 1817-1825]. However, after decades of failed randomized trials attempting to add a drug to gemcitabine alone, the combination of gemcitabine plus nab-paclitaxel (hereinafter gem-nab- paclitaxel) was found to be superior to gemcitabine alone in metastatic patients [Von Hoff et al. (2014) N. Eng. J. Med. 370, 479-480]. Yet, there is no data for a direct comparison between FOLFIRINOX and gem-nab-paclitaxeL An indirect comparison of the two regimens indicated that FOLFIRINOX has slightly higher activity but also higher toxicity. Since 2016, NALIRIFOX -a regimen based on FOLFIRINOX where irinotecan is replaced with nanoliposomal irinotecan (nal-IRI) with the aim to maintain anti-tumour efficacy while minimizing drug-related toxicities was considered as second-line therapy option for fit patients [Wang Gilliam et al. (2016) Lancet 387, 545-557]. Very recently, a phase III trial found that NALIRIFOX modestly but significantly improved overall survival and progression-free survival compared to gem-nab-paclitaxel in the first-line setting, with a manageable safety-profile. However, there is no phase III data on a direct comparison between NALIRIFOX and FOLFIRINOX.

In conclusion, there are multiple treatment options for patients with pancreatic cancer and the choice is mainly influenced by disease stage and the overall health (performance status) of the patient. In practice, also local factors such as reimbursement and infrastructure can affect the treatment choice. Of note, targeted therapies have been developed as a result of genomic profiling, but only for very small subsets of PDAC patients. Although these treatments can improve outcome and/or quality of life, demonstrated by the increase in 5-year survival rate for pancreatic cancer patients from 6% to 12% over the past decade, treatments are rarely curative.

Strikingly, the survival rate of patients with advanced PDAC treated with gemcitabine is 4% after two years, and is only increased to 9% when treated with the standard of care combination of nab-paclitaxel with gemcitabine. A recent phase III study confirms this poor response to gemcitabine-based treatment [Wainberg et al. (2023) ICO 41, LBA661]. Even in patients with resectable pancreatic cancer, long-term response to gemcitabine is low. For these reasons, there is high interest in the biological mechanisms that govern resistance to gemcitabine.

Gemcitabine, or 2',2'-difluoro-2'-deoxycytidine (dFdC), is a cytidine nucleoside analog. By interfering with DNA synthesis, gemcitabine blocks the cell cycle, inhibits cellular proliferation and induces apoptosis. First, nucleoside transporters allow for gemcitabine influx through the cell membrane. Next, gemcitabine undergoes complex intracellular conversion to the cytotoxic nucleotides gemcitabine diphosphate (dFdCDP) and triphosphate (dFdCTP). Finally, gemcitabine affects DNA synthesis in multiple ways: by direct inhibition of DNA polymerase, by depleting deoxyribonucleotide pools through direct inhibition of ribonucleoside reductase, and by incorporation into DNA where it induces DNA strand termination. Gemcitabine also affects many signalling pathways, although the precise mechanisms are often unclear.

Given that gemcitabine depends on and targets multiple proteins, the up- or downregulation of these targets might support gemcitabine resistance. Multiple resistance mechanisms have been described. These include reduced gemcitabine uptake by hampered nucleoside transporter expression, and reduced conversion of gemcitabine to dFdCDP and dFdCTP. It was recently found that tumour-associated macrophages (TAMs) exchange pyrimidine molecules with PDAC cells, which inhibit gemcitabine uptake and metabolism in PDAC cells [Halbrook et al (2019) Cell Metab 29, 1390- 1399)]. Also other elements of the TME support chemoresistance, such as hypoxia and stroma. Moreover, genome-scale CRISPR screens have identified multiple drug resistance-related genes and pathways. Metabolic processes such as glycolysis and fatty acid synthesis have also been reported to support resistance to gemcitabine. Unfortunately, these efforts have thus far not been translated to an improved outcome for patients with PDAC.

Gemcitabine is used in the treatment of various carcinomas, e.g. as a first-line treatment alone for pancreatic cancer; in combination with cisplatin for advanced or metastatic bladder cancer or advanced or metastatic non-small cell lung cancer; as a second-line treatment in combination with carboplatin for ovarian cancer; in combination with paclitaxel for breast cancer that is metastatic or cannot be surgically removed; as well as off-label to treat cholangiocarcinoma and other biliary tract cancers. UDP-glucose ceramide glucosyltransferase (UGCG) (alternative names: glucosylceramide synthase (GCS), ceramide glucosyltransferase, UDP-glucose:N- acylsphingosine D-glucosyltransferase, EC 2.4.1.80), is an enzyme that catalyzes the attachment of glucose to ceramide lipid species to produce the resulting core structure g lycol ipid/g lycosph ing olip id molecule glucosylceramide (GlcCer, glucocerebroside). One variant (UGCG, 1-BP DUP, 142A ) has been previously reported with unknown significance. Other enzymes can modify GlcCer towards more complex sugar chains and thus produce a wide variety of glycolipids. These include galactolipids, sulfolipids, gangliosides, cerebrosides, globosides and others [Daniotti et al. (2016) Front Oncol. 5, 300]

The role of UGCG in regulating ceramide levels has been explored, since this molecule may induce apoptotic cell death. The role of UGCG in maintaining cholesterol/glycolipid 'rafts', cell-surface membrane domains of specialized permeability and functionality that appear to be involved in a variety of signal transduction events, has also been investigated. Glucosylceramide and structurally related glycolipids accumulate in the lysosomes of patients with genetically inherited diseases, which result from a mutation in one of the essential glycolipid-degrading, catabolizing enzymes, e.g. Gaucher disease, GM1 or GM2 gangliosidoses, and Fabry disease. Secondary lysosomal glycosphingolipid accumulation also occurs in other lysosomal storage diseases, such as Niemann-Pick type C disease. UGCG inhibitors may be applied to reduce the rate of glycolipid synthesis in diseased cells so that there is less glycosphingolipid present to be accumulated, a treatment approach termed substrate reduction therapy.

Lysosomal acid glucosylceramidase (GBA) is a lysosomal enzyme involved in the catabolism of glucosylceramide, i.e. its breakdown to ceramide. In human, homozygous or compound heterozygous mutations of the glucocerebrosidase gene (GBA), were initially discovered in patients suffering from Gaucher Disease (GD), a lysosomal-storage disease characterised by accumulation of glucosylceramide substrate due to reduced p-glucocerebrosidase enzyme activity

Similar to the lysosomal acid glucosylceramidase (GBA) (which catabolizes glucosylceramide (GlcCer) in the lysosome or late endosome), also the non-lysosomal glucosylceramidase (GBA2) (alternative names: non-lysosomal p-glucocerebrosidase 2, non-lysosomal p-glucocerebrosidase, non-lysosomal glucosylceramidase 2, beta- glucocerebrosidase 2, glucosylceramidase beta 2, bile acid beta-glucosidase, EC 3.2.1.45, UniProtKB-Q9HCG7) catalyzes the catabolism of GlcCer to ceramide. However, GBA2 is a non-lysosomal enzyme found mostly in the endoplasmic reticulum and on the cell membrane. Deletion of the GBA2 gene in Gaucher disease mice and Niemann-Pick type C mice markedly rescues their clinical phenotype, despite further increasing the already highly elevated levels of GlcCer.

Glycolipids are widely expressed on the outer surface of eukaryotic plasma membranes and play various roles, especially in cell recognition. As such, glycolipids such as gangliosides partake in cell proliferation and differentiation. Moreover, it has been reported that gangliosides are overexpressed in cancer cells, where they can contribute to tumour growth and immune evasion. Also in pancreatic cancer, glycolipids have been implicated in cancer cell growth and invasion [Sasaki et al. (2019) Sci Rep 9, 19369],

Multiple therapies have been successfully developed that recognize these complex glycolipids as targets. Such therapies include monoclonal antibodies and, more recently, CAR-T cells, and they have demonstrated clinical benefit or even received regulatory approval.

Other therapies have focused on the inhibition of glycolipid metabolism. As UGCG catalyses the first step in glycolipid synthesis, it is an interesting target for drug development. Multiple UGCG inhibitors have been developed and are in clinical use for Gaucher's disease, a genetic disorder characterized by glucocerebroside accumulation. Pharmacological UGCG inhibition has been suggested as an anti-cancer therapy. Specifically, it has been proposed that UGCG plays a role in therapy resistance [Yandi et al. (2013) Cancer Chemother Pharmacol. 7, 13-20].

SUMMARY OF THE INVENTION

The present invention demonstrates that pharmacological modulation of glycolipid metabolism, especially pharmacological inhibition of glucosylceramide synthase (UGCG) and/or inhibition of non-lysosomal glucosylceramidase (GBA2), may overcome resistance to gemcitabine. The present invention relates to a therapeutic strategy to enhance the response of tumours to gemcitabine by targeting lipid metabolism. Pharmacological inhibition of UGCG, which catalyses the rate-limiting step in glucosylceramide synthesis, enhances the response of cancer models to gemcitabine. In cell line models of pancreatic, breast, lung and ovarian cancer, the combination of UGCG inhibitors (such as eliglustat, miglustat, ibiglustat, or sinbaglustat) and gemcitabine acted synergistically to cause combinatorial lethality. The invention demonstrates the potential therapeutic use of a combinatorial approach with chemotherapy agents such as especially gemcitabine in the treatment of cancer. The availability of clinical grade inhibitors with a good safety profile may greatly enhance the translational potential of this approach.

A first aspect of the invention of the invention relates to chemotherapy agents or pharmaceutically acceptable salts thereof, and inhibitors of UDP-glucose ceramide glucosyltransferase synthase (UGCG) or pharmaceutically acceptable salts thereof for use in the treatment of cancer.

Typically the chemotherapy agent is a cytidine nucleoside analogue such as gemcitabine.

Embodiments of a chemotherapy agent are an alkylating agent, an antimetabolite, an antitumour antibiotic, a microtubule inhibitor, a topoisomerase inhibitor, or combination thereof.

The chemotherapy agent and UGCG inhibitor are typically used in the treatment of pancreatic cancer, more particularly pancreatic ductal adenocarcinoma (PDAC).

Typical UGCG inhibitors are eliglustat, miglustat, ibiglustat, lucerastat, sinbaglustat, AL01211 and nizubaglustat; or eliglustat, miglustat, ibiglustat, lucerastat or AL01211 A typical the UGCG inhibitor is sinbaglustat.

More typically a UGCG inhibitor does not cross the blood brain barrier, such as eliglustat or miglustat.

Specific combinations of a chemotherapy agent and a UGCG are one of gemcitabine, gemcitabine + paclitaxel and gemcitabine + nab-paclitaxel (abraxane), combined with one of eliglustat, miglustat, lucerastat, ibiglustat, AL01211, nizubaglustat and sinbaglustat.

More specific combinations are:

-gemcitabine an eliglustat,

-gemcitabine and miglustat,

-gemcitabine and ibliglustat,

-gemcitabine and lucerastat,

-gemcitabine and sinbaglustat.

A second aspect of the invention is a modulator of glycolipid metabolism, or a pharmaceutically acceptable salt thereof for use in a method of treating cancer; said method comprising administering to a subject in need thereof a pharmaceutical composition comprising an effective amount of said modulator of glycolipid metabolism, or a pharmaceutically acceptable salt thereof; said method further comprising administering to said subject an effective amount of one or more chemotherapy agent(s); or a pharmaceutically acceptable salt thereof. Embodiments as outlined for the above first aspect are equally applicable for this second aspect.

A third aspect of the invention relates to methods of treating cancer, comprising administering to a subject in need thereof an effective amount of a modulator of glycolipid metabolism, or a pharmaceutically acceptable salt thereof and administering to said subject an effective amount of one or more chemotherapy agent.

Typically the cancer is a carcinoma or an adenocarcinoma, or a metastatic cancer. Examples of a cancer treated by the methods of the invention are pancreatic cancers such as pancreatic ductal adenocarcinoma (PDAC).

Examples of chemotherapy agents used in these methods are an antimetabolite drug, a platinum compound, a topoisomerase inhibitor, a microtubule inhibitor, an alkylating agent, an anti-tumour antibiotic.

Herein typical examples of an antimetabolite drug are

-a cytidine nucleoside analogue,

-a nucleoside analogue antimetabolite drugs such as 5-fluorouracil (5-FU) or precursors thereof such as capecitabine, alone or in combination with folate analogue antimetabolites such as methotrexate, pemetrexed, or folinic acid leucovorin,

- a purine analogue antimetabolite such as 6-mercaptopurine (6-MP) and 6- thioguanine (6-TG).

More specifically the cytidine nucleoside analogue is selected from the group consisting of azacytidine, decitabine, gemcitabine, cytarabine, clofarabine, fludarabine, and vidarabine and gemcitabine.

Alternatively cytidine nucleosides analogues are selected from the group consisting of gemcitabine, a combination of gemcitabine with paclitaxel, or a combination of gemcitabine with nab-paclitaxel.

Specific examples of a cytidine nucleoside are azacytidine, decitabine, gemcitabine, cytarabine, clofarabine, fludarabine, and vidarabine and gemcitabine.

In the methods of the invention the modulator of glycolipid metabolism is typically - a dual inhibitor of glucosylceramide synthase (UGCG inhibitor) and of non-lysosomal glucosylceramidase (GBA2) such as miglustat, lucerastat, nizubaglustat, or sinbaglustat or a pharmaceutically acceptable salt thereof or

- a selective UGCG inhibitor such as eliglustat, ibiglustat, or AL01211 or a pharmaceutically acceptable salt thereof.

Typical examples of glycolipid metabolism modulators are miglustat, lucerastat, nizubaglustat, sinbaglustat, eliglustat, ibiglustat, or AL01211 or a pharmaceutically acceptable salt thereof.

Alternatively, typical examples of glycolipid metabolism are an iminosugar selected from the group consisting of miglustat, lucerastat, nizubaglustat, and sinbaglusta; or a pharmaceutically acceptable salt thereof.

Specific examples of glycolipid metabolism modulators are eliglustat, ibiglustat, or AL01211, or a pharmaceutically acceptable salt thereof.

More specific examples are eliglustat or AL01211, or a pharmaceutically acceptable salt thereof.

A specific example of a modulator is the iminosugar sinbaglustat, or a pharmaceutically acceptable salt thereof, for example administered in a total dose of about 10 to 150 mg per day.

Specific combinations of glycolipid metabolism modulators and chemotherapy agents are sinbaglustat and gemcitabine or eliglustat.

Certain embodiments of the invention are :

1. A composition comprising a chemotherapy agent; and a modulator of glycolipid metabolism which is an inhibitor of UDP-glucose ceramide glucosyltransferase synthase (UGCG), for use in the treatment of cancer.

2. The composition for use according to embodiment 1, wherein the chemotherapy agent is a cytidine nucleoside analogue.

3. The composition for use according to embodiment 1, wherein the chemotherapy agent is an alkylating agent, an antimetabolite, an antitumour antibiotic, a microtubule inhibitor, or a topoisomerase inhibitor; or any combination thereof.

4. The composition for use according to any one of embodiments 1 to 3, wherein the cancer is pancreatic cancer.

5. The composition for use according to any one of embodiments 1 to 4, wherein the cancer is pancreatic ductal adenocarcinoma (PDAC). 6. The composition for use according to embodiment 2, wherein the cytidine nucleoside analogue is gemcitabine.

7. The composition for use according to embodiment 1, wherein the modulator of glycolipid metabolism is eliglustat, miglustat, ibiglustat, lucerastat, sinbaglustat, or AL01211, or nizubaglustat (in a sub-embodiment: eliglustat, miglustat, ibiglustat, lucerastat or AL01211; in another sub-embodiment: sinbaglustat).

8. The composition for use according to any one of embodiments 1 to 5, wherein the inhibitor of UDP-glucose ceramide glucosyltransferase synthase does not cross the blood brain barrier.

9. The composition for use according to embodiment 8, wherein the inhibitor of UDP-glucose ceramide glucosyltransferase synthase (UGCG) is eliglustat.

DESCRIPTION OF THE FIGURES

Figure 1. UGCG expression is associated with poor survival in patients with PDAC

(a) Bulk hexosylceramide abundance in PDAC clinical samples. Paired two-sided Student's t-test. (b) Overall survival of patients with PDAC with either high (n=45) or low (n=44) gene UGCG expression. Log-rank (Mantel-Cox) test, (c) Disease-free survival of patients with PDAC with either high (n=33) or low (n=36) gene UGCG expression. Log-rank (Mantel-Cox) test, (d) Progression-free survival of patients with PDAC with either high (n=89) or low (n=88) gene UGCG expression. Log-rank (Mantel-Cox) test, (e) Disease-specific survival of patients with PDAC with either high (n=86) or low (n=85) gene UGCG expression. Log-rank (Mantel-Cox) test, (f) UGCG gene expression in tumour samples from patients with PDAC per disease stage. Oneway ANOVA with Sldak's multiple comparisons test.

Figure 2. Hexosylceramide synthesis is associated with resistance to gemcitabine

(a) Volcano plot of the lipidomic analysis of PANC-1 cells upon resistance to gemcitabine (n = 4). Two-tailed Student's t-tests with Benjamini-Hochber's multiple comparisons test, (b) Hexosylceramide abundance in PANC-1 cells upon resistance to gemcitabine (n = 4). Unpaired two-sided Student's t-test.

Figure 3. Pharmacological inhibition of glucosylceramide synthesis sensitizes cancer cells to gemcitabine in vitro (a-e) Confluence after 200h of PANC-1 (a), KPC Y 2838c3 (b), A2780 (c), A549 (d) and MCF7 (e) cells treated with either the UGCG inhibitors E I igl ustat, Miglustat or Ibiglustat or in combination with gemcitabine. One-way ANOVA with Sidak's multiple comparisons test.

Figure 4. Glucosylceramide partially rescues combined effect of gemcitabine and eliglustat

Confluence after 200h KPCY 2838c3 cells treated with gemcitabine, eliglustat, and supplemented with glucosylceramide. One-way ANOVA with Sidak's multiple comparisons test.

Figure 5. Preclinical evidence for a therapeutic strategy that combines gemcitabine treatment with a pharmacological UGCG inhibitor

(a) Volcano plot of the lipidomic analysis of PANC-1 xenograft tumours in NMRInu/nu mice upon resistance to gemcitabine (n = 12). (b) Hexosylceramide abundance of PANC-1 xenograft tumours in NMRInu/nu mice after 1 week of treatment with vehicle (n = 5) or gemcitabine (n = 5), or upon reaching a tumour volume of 2000 mm3 under vehicle (n=3) or gemcitabine (n = 12) treatment. One-way ANOVA with Sidak's multiple comparisons test, (c) Tumour volume of subcutaneous KPCY tumours of mice treated with either vehicle (n = 7), Eliglustat (n = 9), gemcitabine (n = 6) or eliglustat and gemcitabine (n = 6). One-way ANOVA with Sidak's multiple comparisons test. Error bars represent s.e.m. from mean of biologically independent samples.

Figure 6. Sinbaglustat sensitizes cancer cells to gemcitabine in vitro

(a-b) Confluence after 200h of KPC 2838c3 cells treated with either the UGCG inhibitor Sinbaglustat or in combination with gemcitabine. One-way ANOVA with Sidak's multiple comparisons test.

Figure 7. Lucerastat sensitizes cancer cells to gemcitabine in vitro

(a-b) Confluence after 200h of KPC 2838c3 cells treated with either the UGCG inhibitor Lucerastat or in combination with gemcitabine. One-way ANOVA with Sidak's multiple comparisons test.

DETAILED DESCRIPTION OF THE INVENTION

1) A first embodiment relates to a composition comprising

• one or more chemotherapy agent(s) (or alternatively named, chemotherapeutic agent(s)), notably comprising a cytidine nucleoside analogue (especially gemcitabine); and

• a modulator of glycolipid metabolism which is an inhibitor of UDP-glucose ceramide glucosyltransferase synthase (UGCG); for use in the treatment of cancer.

2) The composition for use according to embodiment 1), wherein the chemotherapy agent is a cytidine nucleoside analogue.

3) The composition for use according to embodiment 1), wherein the chemotherapy agent is an alkylating agent, an antimetabolite, an antitumour antibiotic, a microtubule inhibitor, or a topoisomerase inhibitor; or any combination thereof.

4) The composition for use according to any one of embodiments 1) to 3), wherein the cancer is pancreatic cancer.

5) The composition for use according to any one of embodiments 1) to 4), wherein the cancer is pancreatic ductal adenocarcinoma (PDAC).

6) The composition for use according to embodiment 2), wherein the cytidine nucleoside analogue is gemcitabine.

7) The composition for use according to embodiment 1), wherein the modulator of glycolipid metabolism is eliglustat, miglustat, ibiglustat, lucerastat, sinbaglustat, or AL01211, or nizubaglustat (in a sub-embodiment: eliglustat, miglustat, ibiglustat, lucerastat or AL01211; in another sub-embodiment: sinbaglustat).

8) The composition for use according to any one of embodiments 1) to 5), wherein the inhibitor of UDP-glucose ceramide glucosyltransferase synthase does not cross the blood brain barrier

9) The composition for use according to embodiment 8), wherein the inhibitor of UDP- glucose ceramide glucosyltransferase synthase (UGCG) is eliglustat.

The term "chemotherapy" refers to the treatment of cancer with one or more cytotoxic anti-neoplastic agents ("chemotherapy agents"). The term especially refers to conventional cytotoxic chemotherapeutic agents which act by killing cells that divide rapidly, one of the main properties of most cancer cells. Chemotherapy may use one drug at a time (single-agent chemotherapy) or several drugs at once (combination chemotherapy or polychemotherapy).

The term "chemotherapy agent" as used herein refers to an active anti-neoplastic agent inducing apoptosis or necrotic cell death. When used in combination with the, UGCG inhibitor, the term especially refers to one or more conventional cytotoxic chemotherapy agents such as: a) antimetabolite drugs including especially cytidine nucleoside analogues; as well as other nucleoside analogue antimetabolite drugs such as 5-fluorouracil (5-FU) or precursors thereof such as capecitabine, alone or in combination with folate analogue antimetabolites such as methotrexate, pemetrexed, or especially folinic acid/leucovorin; and purine analogue antimetabolites such as 6-mercaptopurine (6-MP) and 6-thioguanine (6-TG) [especially antimetabolite drugs refer to cytidine nucleoside analogues (in particular gemcitabine)]; and/or b) platinum compounds; and/or c) topoisomerase inhibitors; and/or d) microtubule inhibitors; and/or e) alkylating agents; and/or f) anti-tumour antibiotics.

When used in combination with the UGCG inhibitors, preferred chemotherapy agents are a. the above-mentioned antimetabolite drugs, notably said cytidine nucleoside analogues (including especially azacytidine, decitabine, gemcitabine, cytarabine, clofarabine, fludarabine, and vidarabine; in particular gemcitabine); alone or in combination with a microtubule inhibitor or a platinum compound; [in particular gemcitabine alone or in combination with paclitaxel (including nab-paclitaxel), or in combination with cisplatin]; or b. a combination of at least two (preferably a combination of a total of two, three or four) cytotoxic chemotherapy agents comprising i. at least one cytidine nucleoside analogue which is 5-fluorouracil (5-FU) or a prodrug/precursor thereof (such as capecitabine), alone or (preferably) in combination with folinic acid/leucovorin; and

II. a platinum drug (especially oxaliplatin); and/or ill. a microtubule inhibitor (especially paclitaxel (including nab-paclitaxel); and/or iv. a topoisomerase inhibitor (especially irinotecan). A non-limiting list of cytidine nucleoside analogues includes

Azacytidine (5-Azacytidine or 5-Aza-CR). Azacytidine incorporates into DNA and RIMA, leading to inhibition of DNA methyltransferases and reactivation of tumour suppressor genes.

Decitabine (5-Aza-2'-deoxycytidine or 5-Aza-CdR). Decitabine is incorporated into DNA, leading to DNA hypomethylation and reactivation of silenced genes.

Gemcitabine (2',2'-difluorodeoxycytidine or dFdC). Gemcitabine undergoes intracellular phosphorylation to its active triphosphate form, which inhibits DNA synthesis by incorporation into DNA and causes chain termination.

Cytarabine (1-p-D-arabinofuranosylcytosine or Ara-C). Cytarabine is phosphorylated intracellularly to its active triphosphate form, which inhibits DNA polymerase and causes chain termination.

Clofarabine is metabolized intracellularly to its active triphosphate form, which inhibits DNA polymerase and ribonucleotide reductase, leading to DNA synthesis inhibition.

Fludarabine (9-p-D-arabinofuranosyl-2-fluoroadenine or F-ara-A) is phosphorylated intracellularly to its active triphosphate form, which inhibits DNA synthesis and causes chain termination.

Vidarabine (adenine arabinoside or ara-A). It inhibits viral DNA polymerase and incorporates into viral DNA, leading to chain termination.

In a preferred embodiment the cytidine nucleoside analogue is gemcitabine.

Typically a cytidine nucleoside analogue may be used alone or in combination with one or more further cancer chemotherapeutic agents such as nab-paclitaxel (abraxane), carboplatin or cisplatin. In particular, gemcitabine may be used in combination with nab-paclitaxel.

Other chemotherapeutic agents for use in the treatment of cancer, in particular pancreas cancer, in combination with a glucosylceramide synthase (UGCG) inhibitors are:

Alkylating agents, which include: nitrogen mustards such as cyclophosphamide, ifosfamide, and mechlorethamine (mustine); alkyl sulfonates such as busulfan; ethylenimines such as thiotepa;

Triazines such as temozolomide; and Nitrosoureas such as carmustine (BCNU), lomustine (CCNU), and semustine (MECCNU).

Platinum compounds, which include: cisplatin, carboplatin, oxaliplatin, satraplatin, lobaplatin, nedaplatin, picoplatin, phenanthriplatin, and triplatin tetranitrate.

Antimetabolites which include nucleoside analogue antimetabolites such as gemcitabine, 5-fluorouracil (5- FU), cytarabine (Ara-C), azacitidine, decitabine and clofarabine; folate analogue antimetabolites such as methotrexate and pemetrexed; and purine analogue antimetabolites such as 6-mercaptopurine (6-MP) and 6- thioguanine (6-TG).

Antitumour antibiotics which include anthracyclines such as doxorubicin, daunorubicin, idarubicin and epirubicin; bleomycins such as bleomycin A2, bleomycin B2; mitomycins such as mitomycin C; and actinomycin-D

Microtubule inhibitors which include vinca alkaloids such as vinblastine, vincristine, vinorelbine and vindesine; and taxanes such as especially paclitaxel (including nab-paclitaxel), and docetaxel.

Topoisomerase inhibitors which include topoisomerase I inhibitors such as irinotecan (including nanoliposomal irinotecan) or topotecan; and topoisomerase II inhibitors such as etoposide, or teniposide.

A non-limiting list of modulators of glycolipid metabolism which are glucosylceramide synthase (UGCG) inhibitors includes:

Eliglustat (Genz-112638, commonly used as the tartrate salt), a potent selective UGCG inhibitor: Miglustat, also known as N-butyldeoxynojirimycin (NB-DNJ), is an iminosugar derivative and a potent dual inhibitor of UGCG and GBA2;

Miglustat ((2R,3R,4R,5S)-piperidine-l-pentyl-2-(hydroxymethyl)-3,4,5-triol, N-butyl deoxynojirimyin, NB-DNJ) is an iminosugar of the natural D-glucose configuration:

Miglustat is a UGCG inhibitor indicated in the EU and the US for the treatment of Gaucher type 1 disease (GD-1), and in the EU also for the treatment of Niemann- Pick disease Type C (NPC). Miglustat was shown to penetrate into the brain in the rat inhibit, in addition to UGCG, the non-lysosomal glucosylceramidase (GBA2)

Ibiglustat (alternatively named venglustat, GZ/SAR402671), a potent UGCG inhibitor, works by reducing glucosylceramide levels;

Lucerastat [(2R,3S,4R.,5S)-l-butyl-2-(hydroxymethyl)piperidine-3,4,5-triol, N- butyldeoxygalactonojirimycin: WO1994/026714, W02018/220131] is an iminosugar of galactose configuration. Lucerastat is an inhibitor of UGCG and of GBA2 and acts by reducing the production of globotriaosylceramide (Gb3);and is in development for Fabry disease. AL01211, ([E]-N-((lR,2R)-l-(3-chloro-4-cyclopropoxyphenyl)-l-hydroxy-3-

(pyrrolid in-l-y I) propan-2-yl)-2-(6-chloronaphthalen-2-yl)-2-(hydroxyimino) acetamide) is a non-brain penetrant UGCG inhibitor:

Nizubaglustat ((2S,3R,4R,5S)-l-{5-[(2-fluoro-[l,l'-biphenyl]-4yl)methoxy-

]pentyl}-2-(hydroxymethyl) piperidine-3,4,5-triol) is an iminosugar having the L-ido Nizubaglustat is a dual inhibitor of UGCG and non-lysosomal glucosyl ceramidase (GBA2)

Sinbaglustat, [also named (2S,3R,4R,5S)-piperidine-l-pentyl-2-(hydroxymethyl)- 3,4,5-triol N-pentyl-deoxyidonojirimycin, ACT-519276 or OGT2378: W02002/055498] is an iminosugar having the L-ido configuration:

Sinbaglustat is an inhibitor of UGCG. W02002/055498 discloses sinbaglustat for use in the treatment of glycosphingolipid storage diseases, diseases associated with glycosphingolipid accumulation, cancers in which glycosphingolipid synthesis is abnormal, infectious diseases caused by organisms which use cell surface glycosphingolipid as receptors, infectious diseases in which synthesis of glucosylceramide is essential or important, diseases in which excessive glycosphingolipid synthesis occurs, neuronal disorders and neuronal injury. In addition to its function as UGCG inhibitor, sinbaglustat is also reported as a potent inhibitor of GBA2. Sinbaglustat does not inhibit to a significant extend the enzyme lysosomal acid glucosylceramidase (GBA). Sinbaglustat furthermore is not expected to inhibit to a pharmacologically relevant extend off-target enzymes such as intestinal glycosidases. Thus, sinbaglustat can be classified as an iminosugar which is a selective dual GBA2 I UGCG inhibitor. Sinbaglustat has been studied in a phase I clinical single-ascending dose (SAD) and multiple-ascending dose (MAD) study). In the MAD study, the effect of sinbaglustat on the biomarker GlcCerin blood plasma was determined by LC-MS/MS. During the treatment period, the total dose of 600 mg per day (dose of 300 mg b.i.d.) reduced the amount of GlcCer in blood plasma with no significant difference to the higher 1000 mg b.i.d dose; the lower dose of a total of 60 mg per day (30 mg b.i.d.) had overall no, and the medium dose of a total of 200 mg per day (100 mg b.i.d.) overall had partial effect, reducing the amount of GlcCer in blood plasma, but not to the extend observed with 300 mg b.i.d..

PDMP (D-threo-l-phenyl-2-decanoylamino-3-morpholino-l-propanol) is a synthetic

UGCG inhibitor; and

L-PDMP (L-threo-l-phenyl-2-decanoylamino-3-morpholino-l-propanol). L-PDMP is an enantiomer of PDMP and has similar UGCG inhibitory activity. UDP-glucose ceramide glucosyltransferase (UGCG) (alternative names: glucosylceramide synthase (GCS), ceramide glucosyltransferase, UDP-glucose:N- acylsphingosine D-glucosyltransferase, EC 2.4.1.80), is an enzyme that catalyses the attachment of glucose to ceramide lipid species to produce the resulting core structure glycolipid/glycosphingolipid molecule glucosylceramide (GlcCer, glucocerebroside). One variant (UGCG, 1-BP DUP, 142A ) has been previously reported with unknown significance. Other enzymes can modify GlcCer towards more complex sugar chains and thus produce a wide variety of glycolipids. These include galactolipids, sulfolipids, gangliosides, cerebrosides, globosides and others [Daniotti et al. (2016) Front Oncol. 5, 300]

In specific embodiments the inhibitor of glucosylceramide synthase (UGCG inhibitor) is an imino sugar such as miglustat, lucerastat, nizubaglustat, or sinbaglustat, or a non-iminosugar UGCG inhibitor such as eliglustat, ibiglustat, or AL01211; in a subembodiment such UGCG inhibitor is eliglustat, miglustat, ibiglustat, lucerastat, or AL01211; in another sub-embodiment such UGCG inhibitor is sinbaglustat.

Iminosugars such as miglustat, lucerastat, nizubaglustat, or sinbaglustat are modulators of glycolipid metabolism which act as dual inhibitors of glucosylceramide synthase (UGCG inhibitor) and of non-lysosomal glucosylceramidase (GBA2) [wherein it is understood that, at low concentrations/low dosage, such iminosugars may act as selective inhibitors of non-lysosomal glucosylceramidase (GBA2)]. Notably, for cancers other than brain cancers a UGCG inhibitor that does not effectively cross the blood brain barrier may be advantageous; for example eliglustat is reported to be immediately transported back out of the CNS by the multidrug transporter Pgp-1; and AL01211 is reported to have minimal brain penetration.

As an alternative to UGCG inhibitors, Ceramide synthase inhibitors or Glycosphingolipid metabolism inhibitors can be used.

Examples of further UGCG inhibitors or such alternatives to UGCG inhibitors are Fumonisin Bl, D-threo-l-phenyl-2-decanoylamino-3-morpholino-l-propanol (PDMP), Genz-123346, N-(5'-adamantane-l'-yl-methoxy)-pentyl]-deoxynojirimycin (AMP-DNM), N-(n-nonyl)-deoxynojirimycin (NN-DNJ), D-threo-l-phenyl-2- decanoylamino-3-pyrrolidino-l-propanol (PPPP) and DL-threo-l-phenyl-2- palmitoylamino-3-morpholino-l-propanol (PPMP). Chemotherapy may be given with a curative intent or it may aim to prolong life or to palliate symptoms.

• Induction chemotherapy is the first line treatment of cancer with a chemotherapeutic drug. This type of chemotherapy is used for curative intent.

• Consolidation chemotherapy is the given after remission in order to prolong the overall disease-free time and improve overall survival. The drug that is administered is the same as the drug that achieved remission.

• Intensification chemotherapy is identical to consolidation chemotherapy but a different drug than the induction chemotherapy is used.

• Combination chemotherapy involves treating a patient with a number of different drugs simultaneously. The drugs differ in their mechanism and side effects. Combination chemotherapy may minimize the chances of resistance developing to any one agent. Also, the drugs may often be used at lower doses, potentially reducing toxicity.

• Neoadjuvant chemotherapy is given prior to a local treatment such as surgery, and is designed to shrink the primary tumour. It is also given to cancers with a high risk of micrometastatic disease.

• Maintenance chemotherapy is a repeated low-dose treatment to prolong remission.

• Salvage chemotherapy or palliative chemotherapy is given without curative intent, but simply to decrease tumour load and increase life expectancy. For these regimens, a better toxicity profile is generally expected.

10) Another aspect of the invention relates to a method of treating cancer; said method comprising administering to a subject in need thereof a pharmaceutical composition comprising an effective amount of a modulator of glycolipid metabolism, or a pharmaceutically acceptable salt thereof; said method further comprising administering to said subject an effective amount of one or more chemotherapy agent(s) [or alternatively named, chemotherapeutic agent(s)], notably comprising a cytidine nucleoside analogue (especially gemcitabine); or a pharmaceutically acceptable salt thereof.

For avoidance of doubt, it is understood that said modulator of glycolipid metabolism and said one or more chemotherapy agent(s) are administered simultaneously or separately, such that they are pharmaceutically active at the same time in said subject [e.g. for at least 6 hours, especially at least 12 hours, in particular 24 hours of a day; wherein it is understood that such modulators of glycolipid metabolism and certain chemotherapy agent(s) may be administered for example enterally (especially orally) in a once or twice daily dosage regimen, while other chemotherapy agent(s) may be administered parenterally, such as especially intravenously, for example once a week].

"Simultaneously", when referring to an administration type, means in the present application that the administration type concerned consists in the administration of two or more active ingredients and/or treatments at approximately the same time; wherein it is understood that a simultaneous administration will lead to exposure of the subject to the two or more active ingredients and/or treatments at the same time. When administered simultaneously, said two or more active ingredients may be administered in a fixed dose combination, or in an equivalent non-fixed dose combination (e.g. by using two or more different pharmaceutical compositions to be administered by the same route of administration at approximately the same time), or by a non-fixed dose combination using two or more different routes of administration; wherein said administration leads to essentially simultaneous exposure of the subject to the two or more active ingredients and/or treatments. "Fixed dose combination", when referring to an administration type, means in the present application that the administration type concerned consists in the administration of one single pharmaceutical composition comprising the two or more active ingredients.

"Separately", when referring to an administration type, means in the present application that the administration type concerned consists in the administration of two or more active ingredients and/or treatments at different points in time; wherein it is understood that a separate administration will lead to a treatment phase (e.g. at least 1 hour, notably at least 6 hours, especially at least 12 hours) where the subject is exposed to the two or more active ingredients and/or treatments at the same time; but a separate administration may also lead to a treatment phase where for a certain period of time (e.g. at least 12 hours, especially at least one day) the subject is exposed to only one of the two or more active ingredients and/or treatments. Separate administration especially refers to situations wherein at least one of the active ingredients and/or treatments is given with a periodicity substantially different from daily (wherein "daily" especially refers to once or twice daily) administration (e.g. wherein one active ingredient and/or treatment is given e.g. once or twice a day, and another is given e.g. once a week or at even longer distances). For example, when used in combination with gemcitabine, the present UGCG inhibitors would possibly be used "separately".

The production of the pharmaceutical compositions can be effected in a manner which will be familiar to any person skilled in the art (see for example Remington, The Science and Practice of Pharmacy, 21st Edition (2005), Part 5, "Pharmaceutical Manufacturing" [published by Lippincott Williams & Wilkins]) by bringing the crystalline forms of the present invention, optionally in combination with other therapeutically valuable substances, into a galenical administration form together with suitable, non-toxic, inert, pharmaceutically acceptable solid or liquid carrier materials and, if desired, usual pharmaceutical adjuvants.

Whenever a certain dosage refers to a unit dose of a certain amount in mg, it is understood that such unit dose refers to such amount in mg of the respective active ingredient in free base form. In case the active ingredient is administered e.g. in form of a pharmaceutically acceptable salt, it is understood that the respective amount of active pharmaceutical ingredient (e.g. said pharmaceutically acceptable salt) in a pharmaceutical composition will be adapted accordingly.

The term "treat" or "treatment” or "treating" used with reference to a disease means either that said disease is cured in the patient or animal; or that, although the animal or patient remains affected by the disease, part or all of the symptoms of said disease are either reduced or eliminated.

The terms "subject", and likewise, "patient" refers to mammals, especially humans. Notably, the term "subject" refers to a human patient.

The term "cancer" refers to all sorts of cancers such as especially carcinomas; adenocarcinomas, squamous cell carcinomas, adenosquamous carinomas, and colloid carcinoma; leukemias; sarcomas; lymphomas; myelomas; metastatic cancers; pancreatic cancers; bladder cancers; lung cancers; ovarian cancers; breast cancers; gastro-intestinal cancers; brain tumours; neuroblastomas; prostate cancers; endometrial cancers; skin cancers; head and neck cancers; neuroendocrine tumours; cervical cancers; oral tumours; nasopharyngeal tumours; thoracic cancers; and virally induced tumours.

Particular examples are cancers which may respond to gemcitabine, alone or in combination; such as pancreatic cancer (including notably pancreatic ductal adenocarcinoma (PDAC) and metastatic pancreatic cancer); bladder cancer (including notably advanced or metastatic bladder cancer); lung cancer (including notably advanced or metastatic non-small cell lung cancer); ovarian cancer; breast cancer (including notably breast cancer that is metastatic or cannot be surgically removed); and biliary tract cancers (including notably cholangiocarcinoma); further particular examples are osteosarcoma; neuroblastoma; Ewing sarcoma; gliomas; and retinoblastoma.

One of the key factors limiting success of therapeutic approaches for the treatment of advanced cancers is the often-limited response elicited by therapeutic modalities, be it chemotherapeutics, targeted approaches of immunotherapies. Combinatorial treatments, those aiming at inducing synthetic lethality, have evolved as promising and safe approaches to improve the success of cancer therapies^{71 72}. Typically, synthetic lethality approaches are based on genetic screens exploiting the plethora of genetic mutations found in tumours. While the limits of such sequence-based cancer genetic target discovery are being reached, previous focus was on non-genetic targets -such as lipids-, that more directly reflect adaptive changes in tumours. Lipids are a very attractive and poorly explored class of molecules in this sense. Cancer progression is accompanied by dramatic changes in lipid profiles, some of which directly contribute to therapy resistance and have shown potential in novel combinatorial approaches. Through the application of advanced lipidomics approaches, including spatial lipidomics by mass spectrometry imaging (MSI), a class of glycosphingolipids (GSL) was identified as being highly overexpressed in pancreatic cancer. GSL form a complex class of membrane lipids that are composed of a ceramide backbone that is embedded in the lipid core of the plasma membrane and a hydrophilic saccharide headgroup that faces the extracellular space. Because of their unique physicochemical properties, GSL play important roles in membrane organization and the formation of subdomain structures called membrane microdomains. These micro-domains help organize and stabilize membrane protein complexes and hence play an active role in cellular signalling including the activation of several oncogenic pathways, this way orchestrating the cancer cells' state and fate [Daniotti et al. cited above; Jain et al. (2023) Cancer discovery 13, 454-473].

The first and rate-limiting step in GSL synthesis is the glycosylation of ceramide by the enzyme UDP- glucose ceramide glucosyltransferase (L/GCG), also known as glucosylceramide synthase. The resulting glucosyl-ceramide basic unit can then be further extended by other enzymes catalyzing the stepwise addition of further monosaccharides as well as other modifying groups, leading to the generation of several hundreds of structurally distinct glycosphingolipids including cerebrosides, sulfatides, globosides, and gangliosides. Several of these enzymes are known to be affected in lipid storage diseases including Gaucher disease and Tay-Sachs. GSL are also known to be dysregulated in several tumour types. In clinical cohorts UGCG expression is increased in PDAC compared to healthy pancreas (Fig. la). To probe the clinical indication of increased hexosylceramide abundance, an association between clinical outcome and UGCG expression assessed.

Clinical data from The Cancer Genome Atlas (TCGA) indicates that UGCG expression correlates with poor survival outcomes in patients with PDAC (Fig. lb-e), indicating that UGCG expression contributes to pancreatic cancer progression. However, UGCG expression does not correlate with clinical tumour stage (Fig. If), indicating that hexosylceramide synthesis might not affect pancreatic cancer pathophysiology directly, but could rather affect response to therapy.

Hexosylceramide synthesis is associated with resistance to gemcitabine

As a synthetic pyrimidine nucleoside analogue, gemcitabine has been used for over 20 years to treat a variety of cancer types including pancreatic, lung, breast and ovarian cancers as first or second line therapy when other treatments have failed or are not effective. To this day, gemcitabine remains a cornerstone of various treatment options for patients with PDAC.

Interestingly, lipidomic profiling revealed increased hexosylceramide abundance in gemcitabine resistant PDAC cells (Fig. 2a, b). Together, these findings indicate a functional role for hexosylceramide in resistance to gemcitabine and indicates that UGCG inhibition might overcome gemcitabine resistance.

Clinical-grade UGCG inhibitors (such as Eliglustat, Miglustat, Ibiglustat, Lucerastat and AL01211) are available and are standard of care treatment for certain patients with Gaucher disease. Moreover, these UGCG inhibitors have shown good efficacy and long-term safety. To investigate whether UGCG contributes to chemoresistance, PDAC cell lines PANC-1 and KPCY 2838C3 were co-treated with gemcitabine and clinical- grade, pharmacological UGCG inhibitors (eliglustat, miglustat and ibiglustat, lucerastat and sinbaglustat).

Moreover, as gemcitabine can also be used to treat patients with lung cancer, breast cancer and ovarian cancer, cell lines A549, MCF7 and A2780 were also included. In all models tested, it was found that UGCG inhibition sensitized cancer cells to gemcitabine (Fig. 3). Supplementing cell culture medium with glucosylceramide rescued this combinatorial lethal effect partially (Figure 4), indicating that a decreased glucosylceramide synthesis does not affect the response to gemcitabine directly but rather indirectly, potentially through more complex GSL such as galactolipids, sulfolipids, cerebrosides, gangliosides or globosides.

To assess the role of hexosylceramide synthesis in resistance to gemcitabine in a preclinical model, the lipidome of xenograft PANC-1 tumours that progressed under gemcitabine treatment was analysed. Corroborating in vitro findings (Fig. 2b, c), an increased hexosylceramide abundance was found in gemcitabine resistant tumours (Fig. 5a, b). The effect of the combination of Eliglustat with gemcitabine on tumour growth is assessed in a preclinical subcutaneous KPCY 2838c3 PDAC model (Fig. 5c). Also a synergistic effect of the combination of gemcitabine and sinbaglustat on KPCY 2838c3 cells (fig 6a, b) and the combination of gemcitabine and lucerastat on KPCY 2838c3 (fig 7a, b) was observed.

Glycolipids have a functional role in many biological processes, particularly in the neurological system, as shown by the creation of genetically modified mice with abnormalities in certain biochemical steps of ganglioside synthesis. Mainly expressed on membranes, glycolipids modulate cell signalling processes and cell-to-cell and cell- to-matrix interactions. As such, they are linked to a variety of physiological processes, including growth, differentiation, migration, and cell death. Cancer cells have abnormally glycosylated lipids on their surface, and there is growing evidence that these molecules play a functional role in a number of cancer-related pathways. For this reason, glycolipids are attractive immunotherapeutic targets for the treatment of cancer. This led to the creation of numerous therapies with glycolipids as immunological targets [Daniotti et al. (2013) Front. Oncol. 3, 306]. On the other hand, targeting the synthesis of glycolipids has also been proposed as a therapeutic strategy for cancer, especially the targeting of UGCG (glucosylceramide synthase) to overcome anti-cancer drug resistance [Li et al. (2021) Biomed. Pharmacother. 139, 111565],

The present invention shows that UGCG is overexpressed in PDAC compared to healthy pancreatic tissue. This finding is in accordance with UGCG overexpression in multiple other tumour types. Moreover, the present invention demonstrates that UGCG inhibition sensitizes cancer cells to gemcitabine. Eliglustat and Miglustat are UGCG (UDP-glucose ceramide glucosyltransferase) inhibitors that were developed and approved to treat Gaucher disease, a disease that results from glucocerebroside buildup.

11) A further embodiment, thus, relates to the method according to embodiment 10), wherein said administration of a modulator of glycolipid metabolism, or of said pharmaceutically acceptable salt thereof, leads to a sensitization of the cancer cells to said one or more chemotherapy agent(s) [especially one or more chemotherapy agent(s) comprising gemcitabine]; wherein it is understood that said sensitization may be measured by comparing the effect of said one or more chemotherapy agent(s) on such cancer cell when used in combination with said modulator of glycolipid metabolism, with the effect of said one or more chemotherapy agent(s) alone.

12) Another embodiment relates to the method according to embodiments 10) or 11), wherein said cancer is a carcinoma or adenocarcinoma.

13) Another embodiment relates to the method according to embodiments 10) or 11), wherein said cancer is a pancreatic cancer (including notably pancreatic ductal adenocarcinoma (PDAC) and metastatic pancreatic cancer); a bladder cancer (including notably advanced or metastatic bladder cancer); a lung cancer (including notably advanced or metastatic non-small cell lung cancer); an ovarian cancer; a breast cancer (including notably breast cancer that is metastatic or cannot be surgically removed); or a biliary tract cancer (including notably cholangiocarcinoma); or said cancer is an osteosarcoma; a neuroblastoma; a Ewing sarcoma; a glioma; or a retinoblastoma.

14) Another embodiment relates to the method according to embodiments 10) or 11), wherein said cancer is pancreatic cancer, especially pancreatic ductal adenocarcinoma (PDAC) or metastatic pancreatic cancer.

15) Another embodiment relates to the method according to any one of embodiments 10) to 14), wherein said cancer is resistant to said one or more chemotherapy agent(s) [especially one or more chemotherapy agent(s) comprising gemcitabine]. 16) Another embodiment relates to the method according to any one of embodiments 10) to 15), wherein said one or more chemotherapy agent(s) is/are independently selected from

• an antimetabolite drug, including especially cytidine nucleoside analogues; as well as other nucleoside analogue antimetabolite drugs such as 5-fluorouracil (5-FU) or precursors thereof such as capecitabine, alone or in combination with folate analogue antimetabolites such as methotrexate, pemetrexed, or especially folinic acid/leucovorin; and purine analogue antimetabolites such as 6-mercaptopurine (6-MP) and 6-thioguanine (6-TG) [especially antimetabolite drugs refer to cytidine nucleoside analogues (in particular gemcitabine)]; and/or

• a platinum compound; and/or

• a topoisomerase inhibitor; and/or

• a microtubule inhibitor; and/or

• an alkylating agent; and/or

• an anti-tumour antibiotic.

17) Another embodiment relates to the method according to any one of embodiments 10) to 15), wherein said one or more chemotherapy agent(s) is/are

• an antimetabolite drug [notably a cytidine nucleoside analogue (including especially azacytidine, decitabine, gemcitabine, cytarabine, clofarabine, fludarabine, and vidarabine; in particular gemcitabine)]; or

• a combination of an antimetabolite drug [notably a cytidine nucleoside analogue (including especially azacytidine, decitabine, gemcitabine, cytarabine, clofarabine, fludarabine, and vidarabine; in particular gemcitabine)] with a microtubule inhibitor;

[in particular a combination of gemcitabine with paclitaxel (including nab- paclitaxel)]; or

• a combination of an antimetabolite drug [notably a cytidine nucleoside analogue (including especially azacytidine, decitabine, gemcitabine, cytarabine, clofarabine, fludarabine, and vidarabine; in particular gemcitabine)] with a platinum compound;

[in particular a combination of gemcitabine with cisplatin]; or

• a combination of at least two (preferably three or four) cytotoxic chemotherapy agents comprising at least one cytidine nucleoside analogue which is 5-fluorouracil (5-FU) or a prodrug/precursor thereof (such as capecitabine), alone or (preferably) in combination with a vitamin B derivative such as folinic acid/leucovorin; and a platinum drug (especially oxaliplatin); and/or a microtubule inhibitor (especially paclitaxel (including nab-paclitaxel); and/or a topoisomerase inhibitor (especially irinotecan).

[in particular (i) a combination of 5-fluorouracil (5-FU) or capecitabine with folinic acid/leucovorin, and oxaliplatin and irinotecan; or (ii) a combination of 5- fluorouracil (5-FU) or capecitabine (each alone or in combination with folinic acid/leucovorin), and nab-paclitaxel].

18) Another embodiment relates to the method according to any one of embodiments 10) to 15), wherein said one or more chemotherapy agent(s) is/are

[in particular a combination of gemcitabine with cisplatin].

19) Another embodiment relates to the method according to any one of embodiments 10) to 15), wherein one or more chemotherapy agent(s) is/are

• gemcitabine; or

• a combination of gemcitabine with paclitaxel (including nab-paclitaxel).

20) Another embodiment relates to the method according to any one of embodiments 10) to 19), wherein said modulator of glycolipid metabolism is • a dual inhibitor of glucosylceramide synthase (UGCG inhibitor) and of non- lysosomal glucosylceramidase (GBA2) such as miglustat, lucerastat, nizubaglustat, or sinbaglustat; or a pharmaceutically acceptable salt thereof; or

• a selective UGCG inhibitor such as eliglustat, ibiglustat, or AL01211; or a pharmaceutically acceptable salt thereof.

21) Another embodiment relates to the method according to any one of embodiments 10) to 19), wherein said modulator of glycolipid metabolism is miglustat, lucerastat, nizubaglustat, sinbaglustat, eliglustat, ibiglustat, or AL01211; or a pharmaceutically acceptable salt thereof.

22) Another embodiment relates to the method according to any one of embodiments 10) to 19), wherein said modulator of glycolipid metabolism is an iminosugar selected from miglustat, lucerastat, nizubaglustat, and sinbaglustat; or a pharmaceutically acceptable salt thereof.

23) Another embodiment relates to the method according to any one of embodiments 10) to 19), wherein said modulator of glycolipid metabolism is eliglustat, ibiglustat, or AL01211; or a pharmaceutically acceptable salt thereof.

24) Another embodiment relates to the method according to any one of embodiments 10) to 19), wherein said modulator of glycolipid metabolism is eliglustat or AL01211; or a pharmaceutically acceptable salt thereof.

25) Another embodiment relates to the method according to any one of embodiments 10) to 19), wherein said modulator of glycolipid metabolism is an iminosugar which is sinbaglustat; or a pharmaceutically acceptable salt thereof.

26) Another embodiment relates to the method according to embodiments 22) or 25), wherein said iminosugar is administered at a dosage regimen that results in a predominant blockade of non-lysosomal glucosylceramidase (GBA2).

The term "dosage regimen resulting in a predominant blockade of non-lysosomal glucosylceramidase (GBA2)" may notably be defined as dosage regimen leading to an exposure of said iminosugar suitable for a predominant blockade of GBA2 (e.g. in blood plasma, or in the brain), wherein especially, on average over 24 hours, UGCG is not inhibited to an extent that results in pharmacologically efficacious inhibition of UGCG. Especially said dosage regimen leads to an exposure of said iminosugar providing for pharmacologically efficacious inhibition of GBA2, wherein the exposure is such that on average over 24 hours UGCG is not inhibited to an extent that results in pharmacologically efficacious inhibition of UGCG.

It is understood that the exposure I pharmacological effect of said iminosugar resulting in its predominant blockade of non-lysosomal glucosylceramidase (GBA2) can be determined by routine measurement of appropriate biomarkers in plasma or in the CSF (such as preferably glycosphingolipids (GSLs) : glucosylceramide (GlcCer) and/or ganglioside GM3, as well as ganglioside GM2; or, less preferred, other glycosphingolipids such as lactosylceramide (LacCer), ganglioside GM1, glycosylsphingosine (GlcSph), and/or any other GSL with a 0-glucose moiety proximal to ceramide). In another aspect of the invention, alternatively, the exposure of said iminosugar on average over 24 hours can be determined in plasma or in the CSF and can preferably be expressed as AUC?4h.

All glycosphingolipids (GSLs) contain a head group which is a carbohydrate moiety, and an amino-alcohol sphingoid base moiety, most commonly (2S,3R,4E)-2- aminooctadec-4-ene-l,3-diol in mammals. Glycosphingolipids are named using common/trivial nomenclature system as IUPAC names would be too cumbersome and too precise due to length and structural heterogeneity, respectively. Common names can be found in a variety of databases (e.g. http://www.lipidmaps.org/, Fahy et aL, J Lipid Res. 2009 (50, Suppl) : S9-S14; http://www.hmdb.ca/). The common lipid name typically includes information on the head group (e.g. Glc for glucose in GlcCer, or Lac for lactose in LacCer), and the sphingoid base containing lipid moiety (e.g. Cer for ceramide in GlcCer). A ceramide indicates a sphingoid base in which the amino group is acylated with a fatty acid. In addition to the head group there is structural hetereogenity in the sphingoid base moiety I ceramide part of the molecule. Typically, the length and the type of the sphingosine, as well as the length and the type of the fatty acid may be indicated to identify a particular member of GSLs [e.g. GlcCer(dl8: l/18:0) indicates a glucosyl ceramide containing a glucosyl head group, a (2S,3R,4E)-2-aminooctadec-4-ene-l,3-diol sphingoid base (chain length 18 including 1 double bond : (18: 1), which is derivatized on the amino group with stearic acid (18:0)]. Where a selective technique, such as LC-MS/MS is used to quantify a sphingolipid, information on the ceramide substructure can be reasonably inferred and is then given. For sphingolipids in which the amino group is not derivatised (i.e. acetylated with a fatty acid), Sph is used and implies a (2S,3R,4E)-2-aminooctadec- 4-ene-l,3-diol sphingoid base unless otherwise stated (e.g. GlcSph). The term glycosphingolipid (GSL) is used to indicate any sphingolipid in which the head group contains a carbohydrate moiety, in the context of the present invention the term GSL especially refers to a GSL with a beta-glucose moiety proximal to the sphingoid base moiety I ceramide. Important GSLs in the context of the present invention are GlcCer, ganglioside GM3, and ganglioside GM2. GlcCer, GM2, and GM3 can be measured routinely by bioanalytical assays, using for example LC-MS/MS.

If not explicitly stated otherwise, any reference to a certain GSL (such as any reference to GlcCer, to ganglioside GM3, or to ganglioside GM2) refers to sum of different ceramide isoforms of such GSL.

Notably, the term "predominant blockade of GBA2" may be defined as pharmacological action of said iminosugar leading to the following pharmacological effects:

• an increase of levels of GlcCer by at least 25% (notably by at least 50%, especially by at least 100%) when compared to baseline level; and levels of ganglioside GM3 which are within 50 % (notably within 25 %, especially within 10 %) of baseline level; and/or levels of ganglioside GM2 which are within 50 % (notably within 25 %, especially within 10 %) of baseline level.

It is further understood that at least one measurement to determine the predominant blockade of GBA2 (treatment measurement) is to be performed, wherein said treatment measurement(s) is/are preferably performed at least 24 hours after a baseline measurement (in case a baseline measurement is performed), but not more than 36 hours (notably not more than 24 hours, especially not more than 12 hours) after the last administration of said iminosugar. Preferably, in case the corresponding study is a multiple dose study wherein the iminosugar is given for example in two separate unit dosages per day, said treatment measurements is/are performed trough, or, less preferred, in case the corresponding study is a single dose study said treatment measurements is/are performed between 6 hours and not more than 12 hours after the last administration of said iminosugar.

In another aspect of the invention, the term "predominant blockade of GBA2" may be defined as pharmacologically efficacious inhibition of GBA2, wherein the exposure is such that on average over 24 hours UGCG is not inhibited to an extent that results in pharmacologically efficacious inhibition of UGCG. UGCG may be considered as not being inhibited, on average over 24 hours, to an extent that results in pharmacologically efficacious inhibition of UGCG in case the exposure of a given iminosugar (as may be measured in plasma or in the CSF) on average over 24 hours (while resulting in pharmacologically efficacious inhibition of GBA2) does not exceed the IC50 of said iminosugar on UGCG (wherein for example the IC50 of sinbaglustat on UGCG has been determined at about 630 nM).

Unless used regarding temperatures, the term "about" placed before a numerical value "X" refers in the current application to an interval extending from X minus 10% of X to X plus 10% of X, and preferably to an interval extending from X minus 5% of X to X plus 5% of X. In the particular case of temperatures, the term "about" placed before a temperature "Y" refers in the current application to an interval extending from the temperature Y minus 10 °C to Y plus 10 °C, preferably to an interval extending from Y minus 5 °C to Y plus 5 °C, notably to an interval extending from Y minus 3 °C to Y plus 3 °C. Room temperature means a temperature of about 25 °C. When in the current application the term n equivalent(s) is used wherein n is a number, it is meant and within the scope of the current application that n is referring to about the number n, preferably n is referring to the exact number n.

Whenever the word "between" or "to" is used to describe a numerical range, it is to be understood that the end points of the indicated range are explicitly included in the range. For example: if a temperature range is described to be between 40°C and 80°C (or 40°C to 80°C), this means that the end points 40°C and 80°C are included in the range; or if a variable is defined as being an integer between 1 and 4 (or from 1 to 4), this means that the variable is the integer 1, 2, 3, or 4.

27) Another embodiment relates to the method according to any one of embodiments 10) to 26), wherein said modulator of glycolipid metabolism is an iminosugar which is especially sinbaglustat, or a pharmaceutically acceptable salt thereof, wherein said cancer is treated by a predominant blockade of non-lysosomal glucosylceramidase (GBA2); wherein especially sinbaglustat is administered in a total dose of about 10 to 150 mg per day, about 20 to 100 mg per day, or about 10 to 60 mg per day (in a sub-embodiment about 40 to 60 mg, or in another sub-embodiment about 10 to 30 mg per day); wherein said total dose is administered in one unit dosage, or in two or three (especially two) separate unit dosages.

28) Another embodiment relates to the method according to embodiments 22) or 25), wherein said iminosugar is administered at a dosage wherein said dosage results in pharmacologically effective dual inhibition of UDP-glucose ceramide glucosyltransferase (UGCG) and non-lysosomal glucosylceramidase (GBA2).

29) Another embodiment relates to the method according to any one of embodiments 10) to 25), wherein said modulator of glycolipid metabolism is an iminosugar which is especially sinbaglustat, or a pharmaceutically acceptable salt thereof, wherein said cancer is treated by a dual blockade of UDP-glucose ceramide glucosyltransferase (UGCG) and non-lysosomal glucosylceramidase (GBA2); wherein especially sinbaglustat is administered in a total dose of 300 mg per day or more (in particular a total dose of about 300 mg to 1000 mg per day; a total dose of about 500 mg per day, or a total dose of about 600 mg per day); wherein said total dose is administered in one unit dosage, or in two or three (especially two) separate unit dosages.

30) Another embodiment relates to the method according to any one of embodiments 10) to 29), wherein said treatment of cancer results in the slowing of the rate of progression of said cancer.

31) Another embodiment relates to the method according to any one of embodiments 10) to 30), wherein the cancer treated is a metastatic cancer.

For avoidance of any doubt, any method of treatment of a certain disease or disorder, said method comprising a modulator of glycolipid metabolism, or a pharmaceutically acceptable salt thereof, in combination with one or more chemotherapy agent(s), or a pharmaceutically acceptable salt thereof, as set out in any one of embodiments 8) to31) herein, also discloses

• said method comprising administering said one or more chemotherapy agent(s), or a pharmaceutically acceptable salt thereof, in combination with said modulator of glycolipid metabolism, or a pharmaceutically acceptable salt thereof, as set out in any one of embodiments 10) to 31);

• said modulator of glycolipid metabolism, or a pharmaceutically acceptable salt thereof, for use in the method of treatment of said disease or disorder as set out in any one of embodiments 10) to 31) herein;

• said one or more chemotherapy agent(s), or a pharmaceutically acceptable salt thereof, for use in the method of treatment of said disease or disorder as set out in any one of embodiments 10) to 31) herein;

• the use of said modulator of glycolipid metabolism, or of a pharmaceutically acceptable salt thereof, in the preparation of a medicament for the treatment of said disease or disorder as set out in any one of embodiments 10) to 31) herein; wherein said medicament is to be administered in combination with said one or more chemotherapy agent(s), or a pharmaceutically acceptable salt thereof; • the use of said one or more chemotherapy agent(s), or of a pharmaceutically acceptable salt thereof, in the preparation of a medicament for the treatment of said disease or disorder as set out in any one of embodiments 10) to 31) herein; wherein said medicament is to be administered in combination with said modulator of glycolipid metabolism, or a pharmaceutically acceptable salt thereof;

• the use of a pharmaceutical composition comprising said one or more chemotherapy agent(s), or a pharmaceutically acceptable salt thereof, and said modulator of glycolipid metabolism, or a pharmaceutically acceptable salt thereof; in the preparation of a medicament for the treatment of said disease or disorder as set out in any one of embodiments 10) to 31) herein;

• a medicament for the treatment of said disease or disorder as set out in any one of embodiments 10) to 31) herein, said medicament comprising said modulator of glycolipid metabolism, or a pharmaceutically acceptable salt thereof; and said one or more chemotherapy agent(s), or a pharmaceutically acceptable salt thereof; and the like.

Likewise, if a modulator of glycolipid metabolism, or a pharmaceutically acceptable salt thereof, in combination with a one or more chemotherapy agent(s), or a pharmaceutically acceptable salt thereof, is described as useful for the treatment of a certain disease or disorder as set out herein, said modulator of glycolipid metabolism, or a pharmaceutically acceptable salt thereof, is likewise suitable:

• for use in the preparation of a medicament for the treatment of said disease or disorder as set out herein;

• for use in a method to treat of said disease or disorder, said method comprising administering said modulator of glycolipid metabolism, or a pharmaceutically acceptable salt thereof, as set out herein;

• as a medicament for the treatment of said disease or disorder as set out herein, said medicament comprising said modulator of glycolipid metabolism, or a pharmaceutically acceptable salt thereof; and the like.

The invention further relates to a kit comprising

• a pharmaceutical composition, said composition comprising a pharmaceutically acceptable carrier material, and a modulator of glycolipid metabolism wherein said modulator of glycolipid metabolism is as defined in any one of embodiments 1) to 31); • and instructions how to use said pharmaceutical composition for the prevention or the treatment of a cancer, in combination with one or more chemotherapy agent(s) according to any one of the methods of embodiments 10 to 31).

Particular embodiments of the invention are described in the following examples, which serve to illustrate the invention in more detail without limiting its scope in any way.

EXAMPLES

Cell culture

PANC-1, A549 and MCF7 cells were obtained from ATCC. KPCY 2838c3 cells were obtained from Kerafast. A2780 cells were obtained from Sigma-Aldrich. Cell lines were propagated in DMEM High Glucose (Sigma), supplemented with 10% FBS (Life Technologies) and 4 mM glutamine (ThermoFisher). Cell cultures were periodically tested for mycoplasma contamination using the MycoAlert Mycoplasma Detection Kit (Lonza) and confirmed to be mycoplasma free.

For compound treatments, media was supplemented with 30 nM Gemcitabine (MedChem Express), 20 pM Eliglustat (MedChem Express), 100 pM Miglustat (MedChem Express) 20 pM Ibiglustat (MedChem Express), 200 pM sinbaglustat (MedChem Express), and 400 pM lucerastat (MedChem Express), . Cells were cultured at 37°C in a humidified incubator (ICOmed 150, Memmert) with 5% CO2.

Cell growth

Cells were seeded in 96-well plates at a density of 4 x 10³ cells per well and treated with the indicated compounds on the second day. Proliferation (confluency) was assessed with an IncuCyte ZOOM system (Sartorius) based on phase contrast images.

Bulk lipidomics

For lipid extraction an amount of cells or tissue containing 10 pg of DNA was homogenized in 700 pL of water with a handheld sonicator and was mixed with 800 pl HCI(1M) :CH3OH 1 :8 (v/v), 900 pl CHCI3, 200 pg/ml of the antioxidant 2,6-di-tert- butyl-4-methylphenol (BHT; Sigma Aldrich) and 3 pl of SPLASH® LIPIDOMIX® Mass Spec Standard (#330707, Avanti Polar Lipids). After vortexing and centrifugation, the lower organic fraction was collected and evaporated using a Savant Speedvac spdlllv (Thermo Fisher Scientific) at room temperature and the remaining lipid pellet was stored at - 20°C under argon. Just before mass spectrometry analysis, lipid pellets were reconstituted in 100% ethanol. Lipid species were analyzed by liquid chromatography electrospray ionization tandem mass spectrometry (LC-ESI/MS/MS) on a Nexera X2 UHPLC system (Shimadzu) coupled with hybrid triple quadrupole/linear ion trap mass spectrometer (6500+ QTRAP system; AB SCIEX). Chromatographic separation was performed on a XBridge amide column (150 mm x 4.6 mm, 3.5 pm; Waters) maintained at 35°C using mobile phase A (1 mM ammonium acetate in water-acetonitrile 5:95 (v/v)) and mobile phase B (1 mM ammonium acetate in water-acetonitrile 50:50 (v/v)) in the following gradient: (0-6 min: 0% B 6% B; 6-10 min: 6% B 25% B; 10-11 min: 25% B 98% B; 11-13 min: 98% B 100% B; 13-19 min: 100% B; 19-24 min: 0% B) at a flow rate of 0.7 mL/min which was increased to 1.5 mL/min from 13 minutes onwards. SM, CE, CER, DCER, HCER, LCER were measured in positive ion mode with a precursor scan of 184.1, 369.4, 264.4, 266.4, 264.4 and 264.4 respectively. TAG, DAG and MAG were measured in positive ion mode with a neutral loss scan for one of the fatty acyl moieties. PC, LPC, PE, LPE, PG, PI and PS were measured in negative ion mode by fatty acyl fragment ions. Lipid quantification was performed by scheduled multiple reactions monitoring (MRM), the transitions being based on the neutral losses or the typical product ions as described above. The instrument parameters were as follows: Curtain Gas = 35 psi; Collision Gas = 8 a.u. (medium); lonSpray Voltage = 5500 V and -4,500 V; Temperature = 550°C; Ion Source Gas 1 = 50 psi; Ion Source Gas 2 = 60 psi; Declustering Potential = 60 V and -80 V; Entrance Potential = 10 V and -10 V; Collision Cell Exit Potential = 15 V and -15 V. The following fatty acyl moieties were taken into account for the lipidomic analysis: 14:0, 14: 1, 16:0, 16:1, 16:2, 18:0, 18:1, 18:2, 18:3, 20:0, 20: 1, 20:2, 20:3, 20:4, 20:5, 22:0, 22:1, 22:2, 22:4, 22:5 and 22:6 except for TGs which considered : 16:0, 16: 1, 18:0, 18: 1, 18:2, 18:3, 20:3, 20:4, 20:5, 22:2, 22:3, 22:4, 22:5, 22:6. Peak integration was performed with the MultiQuantTM software version 3.0.3. Lipid species signals were corrected for isotopic contributions (calculated with Python Molmass 2019.1.1) and were quantified based on internal standard signals and adheres to the guidelines of the Lipidomics Standards Initiative (LSI) (level 2 type quantification as defined by the LSI). Unpaired T-test p-values and FDR corrected p- values (using the Benjamini/Hochberg procedure) were calculated in Python StatsModels version 0.10.1. Patient Survival and Expression Data

The survival and expression data of patients with pancreatic cancer were from PDAC databases (TCGA-PAAD and QCMG databases) and were obtained through cBioPortal⁹². Gene expression from healthy pancreas was obtained from the GTEX database through UCSC Xena Resource. Protein expression of MAPK14 (pPT180/Y182) was used to probe p38 phosphorylation status.

Animal Experiments

To probe the effect of UGCG inhibition on gemcitabine response, 3 x 10⁵ KPCY 2838c3 cells dissolved in PBS were injected subcutaneously into 6-8 week-old female NMRInu/nu mice (Janvier). Tumours were allowed to grow up a diameter of 5 mm before treatment started. Tumour bearing mice were either treated with vehicle, gemcitabine (UZ Leuven Hospitals, 100 mg/kg, twice weekly by intraperitoneal injection) or eliglustat (100 mg/kg, daily oral delivery through drinking water, dissolved in 5% gum Arabic and 5% PEG). Tumour size was measured with a digital caliper (Fowler Sylvac) and mice were euthanized upon reaching humane endpoints (tumour size exceeding 1000 mm³ or ulcerating tumours), after which tumours were collected and snap-frozen for further analysis.

Statistical analysis

The results were analyzed using GraphPad Prism v.9.1.2. Statistical data analysis was performed on n > 3 biologically independent replicates. Details on statistical tests and post-tests are presented in the figure legends. P-values of <0.05 were considered to be statistically significant.

Claims

1. A chemotherapy agent or pharmaceutically acceptable salt thereof, and an inhibitor of UDP-glucose ceramide glucosyltransferase synthase (UGCG) or pharmaceutically acceptable salt thereof for use in the treatment of cancer.

2. The chemotherapy agent and UGCG inhibitor for use in the treatment of cancer according to claim 1, wherein the chemotherapy agent is a cytidine nucleoside analogue.

3. The chemotherapy agent and UGCG inhibitor for use in the treatment of cancer according to claim 2, wherein the cytidine nucleoside analogue is gemcitabine.

4. The chemotherapy agent and UGCG inhibitor for use in the treatment of cancer according to any one of claim 1 to 3, wherein the chemotherapy compound is an alkylating agent, an antimetabolite, an antitumour antibiotic, a microtubule inhibitor, a topoisomerase inhibitor, or combination thereof.

5. The chemotherapy agent and UGCG inhibitor for use in the treatment of cancer according to any one of claims 1 to 4, wherein the cancer is pancreatic cancer.

6. The chemotherapy agent and UGCG inhibitor for use in the treatment of cancer according to any one of claims 1 to 5, wherein the cancer is pancreatic ductal adenocarcinoma (PDAC).

7. The chemotherapy agent and UGCG inhibitor for use in the treatment of cancer according to any one of claims 1 to 6, wherein the UGCG inhibitor is selected form the group consisting of eliglustat, miglustat, ibiglustat, lucerastat, sinbaglustat, AL01211 and nizubaglustat.

8. The chemotherapy agent and UGCG inhibitor for use in the treatment of cancer according to any one of claims 1 to 7, wherein the UGCG inhibitor is selected form the group consisting of eliglustat, miglustat, ibiglustat, lucerastat or AL01211.

9. The chemotherapy agent and UGCG inhibitor for use in the treatment of cancer according to any one of claims 1 to 8, wherein the UGCG inhibitor is sinbaglustat.

10. The chemotherapy agent and inhibitor for use in the treatment of cancer according to any one of claims 1 to 9, wherein the UGCG inhibitor does not cross the blood brain barrier.

11. The chemotherapy agent and UGCG inhibitor for use according to any one of claims 1 to 10, wherein the UGCG inhibitor is eliglustat or miglustat.

12. The chemotherapy agent and UGCG inhibitor for use according to any one of claims 1 to 11 :

-wherein the chemotherapy agent is selected from the group consisting of gemcitabine, or gemcitabine + paclitaxel and gemcitabine + nab-paclitaxel (abraxane), and

-wherein the UGCG inhibitor is selected from the group consisting of eliglustat, miglustat, lucerastat, ibiglustat, AL01211, nizubaglustat and sinbaglustat.

13. The chemotherapy agent and inhibitor for use according to any one of claims 1 to 12, wherein the chemotherapy agent is gemcitabine and the inhibitor is eliglustat, or wherein the chemotherapy agent is gemcitabine and the inhibitor is miglustat, or wherein the chemotherapy agent is gemcitabine and the inhibitor is iblig lustat, or wherein the chemotherapy agent is gemcitabine and the inhibitor is lucerastat, or wherein the chemotherapy agent is gemcitabine and the inhibitor is sinbaglustat.

14. A modulator of glycolipid metabolism, or a pharmaceutically acceptable salt thereof for use in a method of treating cancer; said method comprising administering to a subject in need thereof a pharmaceutical composition comprising an effective amount of said modulator of glycolipid metabolism, or a pharmaceutically acceptable salt thereof; said method further comprising administering to said subject an effective amount of one or more chemotherapy agent(s); or a pharmaceutically acceptable salt thereof.

15. A method of treating cancer, the method comprising administering to a subject in need thereof an effective amount of a modulator of glycolipid metabolism, or a pharmaceutically acceptable salt thereof; and administering to said subject an effective amount of one or more chemotherapy agent.

16. The method according to claim 15, wherein the cancer is a carcinoma or an adenocarcinoma.

17. The method according to claim 15 or 16 wherein the cancer is pancreatic cancer.

18. The method according to any one of claims 15 to 17 wherein the cancer is pancreatic ductal adenocarcinoma (PDAC).

19. The method according to any one of claims 15 to 18, wherein the cancer treated is a metastatic cancer.

20. The method according to any one of claims 15 to 19, where the chemotherapy agent is one or more independently selected from the group consisting of an antimetabolite drug, a platinum compound; and/or a topoisomerase inhibitor; and/or a microtubule inhibitor; and/or an alkylating agent; and/or an anti-tumour antibiotic.

21. The method according to any one of claims 15 to 20, wherein chemotherapy agent is an antimetabolite drug selected from the group consisting of:

-a cytidine nucleoside analogue,

22. The method according to any one of claim 15 to 21, wherein the chemotherapy agent is a cytidine nucleoside analogue selected from the group consisting of azacytidine, decitabine, gemcitabine, cytarabine, clofarabine, fludarabine, and vidarabine and gemcitabine.

23. The method according to any one of claim 15 to 21, wherein the chemotherapy agent is a cytidine nucleoside selected from the group consisting of gemcitabine, a combination of gemcitabine with paclitaxel, or a combination of gemcitabine with nab-paclitaxel.

24. The method according to any one of claim 15 to 21, wherein the chemotherapy agent is a cytidine nucleoside selected from the group consisting of azacytidine, decitabine, gemcitabine, cytarabine, clofarabine, fludarabine, and vidarabine and gemcitabine.

25. The method according to any one of claim 15 to 24, wherein said modulator of glycolipid metabolism is:

- a dual inhibitor of glucosylceramide synthase (UGCG inhibitor) and of non- lysosomal glucosylceramidase (GBA2) such as miglustat, lucerastat, nizubaglustat, or sinbaglustat or a pharmaceutically acceptable salt thereof or

26. The method according to any one of claim 15 to 25, wherein said modulator of glycolipid metabolism is selected from the group consisting of miglustat, lucerastat, nizubaglustat, sinbaglustat, eliglustat, ibiglustat, or AL01211 or a pharmaceutically acceptable salt thereof.

27. The method according to any one of claim 15 to 25, wherein said modulator of glycolipid metabolism is an iminosugar selected from the group consisting of miglustat, lucerastat, nizubaglustat, and sinbaglusta; or a pharmaceutically acceptable salt thereof.

28. The method according to any one of claim 15 to 25, wherein said modulator of glycolipid metabolism is eliglustat, ibiglustat, or AL01211, or a pharmaceutically acceptable salt thereof.

29. The method according to any one of claim 15 to 25, wherein said modulator of glycolipid metabolism is eliglustat or AL01211, or a pharmaceutically acceptable salt thereof.

30. The method according to any one of claim 15 to 25, wherein said modulator of glycolipid metabolism is an iminosugar which is sinbaglustat, or a pharmaceutically acceptable salt thereof.

31. The method according to any one of claim 15 to 25, wherein said modulator of glycolipid metabolism is an iminosugar which is especially sinbaglustat, or a pharmaceutically acceptable salt thereof, wherein especially sinbaglustat is administered in a total dose of about 10 to 150 mg per day.

32. The method according to any one of claims 15 to 31, wherein the modulator of glycolipid metabolism is sinbaglustat or eliglustat, and wherein the chemotherapy agent is gemcitabine.