L-asparaginase anti-tumor activity in pancreatic cancer is dependent on its glutaminase activity and resistance is mediated by glutamine synthetase.

l-Asparaginase is a cornerstone of acute lymphoblastic leukemia (ALL) therapy since lymphoblasts lack asparagine synthetase (ASNS) and rely on extracellular asparagine availability for survival. Resistance mechanisms are associated with increased ASNS expression in ALL. However, the association between ASNS and l-Asparaginase efficacy in solid tumors remains unclear, thus limiting clinical development. Interestingly, l-Asparaginase also has a glutaminase co-activity that is crucial in pancreatic cancer where KRAS mutations activate glutamine metabolism. By developing l-Asparaginase-resistant pancreatic cancer cells and using OMICS approaches, we identified glutamine synthetase (GS) as a marker of resistance to l-Asparaginase. GS is the only enzyme able to synthesize glutamine, and its expression also correlates with l-Asparaginase efficacy in 27 human cell lines from 11 cancer indications. Finally, we further demonstrated that GS inhibition prevents cancer cell adaptation to l-Asparaginase-induced glutamine starvation. These findings could pave the way to the development of promising drug combinations to overcome l-Asparaginase resistance.

An avian embryo patient-derived xenograft model for preclinical studies of human breast cancers.

Lack of preclinical patient-derived xenograft cancer models in which to conduct large-scale molecular studies seriously impairs the development of effective personalized therapies. We report here an in vivo concept consisting of implanting human tumor cells in targeted tissues of an avian embryo, delivering therapeutics, evaluating their efficacy by measuring tumors using light sheet confocal microscopy, and conducting large-scale RNA-seq analysis to characterize therapeutic-induced changes in gene expression. The model was established to recapitulate triple-negative breast cancer (TNBC) and validated using TNBC standards of care and an investigational therapeutic agent.

Methionine tumor starvation by erythrocyte-encapsulated methionine gamma-lyase activity controlled with per os vitamin B6.

Erymet is a new therapy resulting from the encapsulation of a methionine gamma-lyase (MGL; EC number 4.4.1.11) in red blood cells (RBC). The aim of this study was to evaluate erymet potential efficacy in methionine (Met)-dependent cancers. We produced a highly purified MGL using a cGMP process, determined the pharmacokinetics/pharmacodynamics (PK/PD) properties of erymet in mice, and assessed its efficacy on tumor growth prevention. Cytotoxicity of purified MGL was tested in six cancer cell lines. CD1 mice were injected with single erymet product supplemented or not with vitamin B6 vitamer pyridoxine (PN; a precursor of PLP cofactor). NMRI nude mice were xenografted in the flank with U-87 MG-luc2 glioblastoma cells for tumor growth study following five intravenous (IV) injections of erymet with daily PN oral administration. Endpoints included efficacy and event-free survival (EFS). Finally, a repeated dose toxicity study of erymet combined with PN cofactor was conducted in CD1 mice. Recombinant MGL was cytotoxic on 4/6 cell lines tested. MGL half-life was increased from <24 h to 9-12 days when encapsulated in RBC. Conversion of PN into PLP by RBC was demonstrated. Combined erymet + PN treatment led to a sustained Met depletion in plasma for several days with a 85% reduction of tumor volume after 45 days following cells implantation, and a significant EFS prolongation for treated mice. Repeated injections in mice exhibited a very good tolerability with only minor impact on clinical state (piloerection, lean aspect) and a slight decrease in hemoglobin and triglyceride concentrations. This study demonstrated that encapsulation of methioninase inside erythrocyte greatly enhanced pharmacokinetics properties of the enzyme and is efficacy against tumor growth. The perspective on these results is the clinical evaluation of the erymet product in patients with Met starvation-sensitive tumors.

Pancreatic tumor sensitivity to plasma L-asparagine starvation.

OBJECTIVES: In this study, our aim was to test whether asparagine synthetase (ASNS) deficiency in pancreatic malignant cells can lead to sensitivity to asparagine starvation. We also investigated, in tumor-bearing mice, the efficacy of L-asparaginase entrapped in red blood cells (RBCs), a safe formulation, to induce asparagine depletion. METHODS: First, ASNS expression was evaluated by immunohistochemistry in sporadic pancreatic ductal adenocarcinoma. Then, 4 pancreatic carcinoma cell lines were examined by Western blot, immunocytochemistry, and cytotoxicity assay to L-asparaginase and in asparagine-free or reduced-asparagine media. Finally, mice bearing the most in vitro sensitive cell line received RBC-entrapped L-asparaginase to investigate the anticancer efficacy of serum asparagine depletion in vivo. RESULTS: Approximately 52% of pancreatic adenocarcinomas expressed no or low ASNS. The highest in vitro cytotoxicity to L-asparaginase or to reduced asparagine medium was observed with SW1990 line when ASNS expression was the lowest. In vivo sensitivity was confirmed for this cell line. CONCLUSIONS: Plasma asparagine depletion by RBC-entrapped L-asparaginase in selected patients having no low or no ASNS may be a promising therapeutic approach for pancreatic cancer.