Adjuvant multimodal metabolic therapy of neuroblastoma

Neuroblastoma is a pediatric tumor of the sympathetic nervous system. Pediatric patients with high-risk neuroblastoma still have poor outcomes, despite receiving multi-modal intensive therapy. Thus, there is an urgent need for new NB therapies. Targeting cellular metabolism is emerging as a promising strategy in cancer treatment. In particular, neuroblastoma tumors, like many other solid tumors, reduce their respiration and strongly depend on glucose for energy production, which renders them highly susceptible to glucose deprivation. Recent preclinical data of researchers at the Department of Pediatrics of the Paracelsus Medical University in Salzburg indicate that targeting this metabolic phenotype by a ketogenic diet, which has a high fat and low protein/carbohydrate content, slows neuroblastoma tumor growth and enhances the effectiveness of a classical chemotherapy. However, ketogenic diet alone is not very efficient at reducing blood glucose levels. Thus, some studies have used caloric restriction in combination with ketogenic diet to further reduce plasma glucose, but this therapeutic approach is not applicable in patients with low body mass index. As an alternative, we propose that combining ketogenic diet and specific metabolic inhibitors can be used as an adjuvant therapy in neuroblastoma. This approach should target not only glucose dependent bulk cancer cells, but also respiration-competent cancer stem cells. Several studies have demonstrated that the antidiabetic drug metformin, which inhibits the mitochondrial energy metabolism and antibiotics, which inhibit mitochondrial protein synthesis, can block tumor cell proliferation. Our first aim is to determine the effect of multi-level metabolic targeting on neuroblastoma proliferation in a murine xenograft model. Classical low-dose chemotherapy will be combined with KD and drugs targeting cancer metabolism, such as metformin (to inhibit gluconeogenesis and oxidative phosphorylation) and/or antibiotics like doxycycline (to reduce mitochondrial protein synthesis) and the effect on the proliferation of NB xenografts will be determined. Our second aim is to demonstrate that the proposed multi-modal therapy is also effective in an immunocompetent genetic neuroblastoma model. We will compare different treatment groups in terms of markers of proliferation and cell death. Because dietary intervention and application of drugs interacting with body/cellular metabolism will influence cellular and extracellular metabolites, our final aim is to determine the effect of the multi-modal therapy on the plasma and tumor metabolomes. Despite the fact that numerous approaches to target the tumor metabolism have been reported, none has tried to target different pathways of the tumor metabolism. The proposed preclinical studies targeting neuroblastoma metabolism will provide a strong foundation for multi-level metabolic targeting of other types of cancer as well as the initiation of clinical studies. The combination therapy aims at increasing the survival of the patients and at reducing therapy induced long-term toxicity by the possibility to reduce the duration of the therapy.

Neuroblastoma is a tumor of the sympathetic nervous system that occurs primarily in early childhood. Despite intensive multimodal therapies, high-risk patients have a poor prognosis. Nearly half of these patients have multiple copies of a gene called MYCN. This gene is one of the best markers of poor prognosis in neuroblastoma patients. Unfortunately however, there are still no therapeutic approaches that target MYCN. Like most solid tumors, neuroblastomas are characterized by a massive dependence on glucose to meet energy demands and a decrease in respiration (i.e., oxidative metabolism in mitochondria). MYCN is able to increase both glucose and oxidative metabolism in neuroblastoma. Therefore, targeting the metabolic vulnerabilities induced by MYCN may represent an innovative and unexplored approach to treat high-risk neuroblastoma. Metformin, a common anti-diabetic drug, as well as certain antibiotics are to inhibit cell respiration. Pre-clinical studies revealed an anti-tumor effect of a low carbohydrate, high fat ketogenic diet, especially when combined with conventional chemotherapy. Thus, the main aim of the present study was to evaluate if metformin or antibiotics might enhance the effect of a ketogenic diet and low dose chemotherapy in high-risk neuroblastoma mouse models. Furthermore, we aimed to elucidate mechanisms mediating the anti-tumor effect of the multimodal therapies. First, we showed that metformin reduces respiration in neuroblastoma cells and, like certain antibiotics, reduced tumor cell growth in cell cultures in a dose-dependent manner. In vivo we observed that the antibiotic tigecycline, at a dose that was therapeutically relevant, induced substantial liver toxicity. On the other hand, metformin was well tolerated, notably in combination with the ketogenic diet and chemotherapy. Moreover, the multi-modal therapy with the ketogenic diet, metformin and low dose chemotherapy decreased the tumor growth and increased the survival of mice with neuroblastoma. The combination therapy increased the expressions of genes involved in the catabolism of fatty acids and suppressed an enzyme involved in fatty acid synthesis. This metabolic perturbation might lead to redox stress in the neuroblastoma cells and ultimately, slow tumor growth. Thus, the multi-modal therapy opens up new therapeutic approaches able to reduce the dose of standard chemotherapy, but with an enhanced anti-tumor effect. Furthermore, ketogenic diet seems to increase the quality of life of a range of cancer patients. The use of a ketogenic diet and metformin needs to be tested as adjuvant therapy in cancer patients in future clinical studies. The potential introduction in the standard medical practice might help to reduce the time and intensity of incriminating therapy, severe side effects as well as the hospitalization of the cancer patients.