Cara Valvona

There are over 120 types of brain tumor and approximately 45% of primary brain tumors are gliomas, of which glioblastoma multiforme (GBM) is the most common and aggressive with a median survival rate of 14 months. Despite progress in our knowledge, current therapies are unable to effectively combat primary brain tumors and patient survival remains poor. Tumor metabolism is important to consider in therapeutic approaches and is the focus of numerous research investigations. Lactate dehydrogenase A (LDHA) is a cytosolic enzyme, predominantly involved in anaerobic and aerobic glycolysis (the Warburg effect); however, it has multiple additional functions in non-neoplastic and neoplastic tissues, which are not commonly known or discussed. This review summarizes what is currently known about the function of LDHA and identifies areas that would benefit from further exploration. The current knowledge of the role of LDHA in the brain and its potential as a therapeutic target for brain tumors will also be highlighted. The Warburg effect appears to be universal in tumors, including primary brain tumors, and LDHA (because of its involvement with this process) has been identified as a potential therapeutic target. Currently, there are, however, no suitable LDHA inhibitors available for tumor therapies in the clinic.

Medulloblastoma is the most common malignant paediatric brain tumour and current therapies often leave patients with severe neurological disabilities. Four major molecular groups of medulloblastoma have been identified (Wnt, Shh, Group 3 and Group 4), which include additional, recently defined subgroups with different prognosis and genetic characteristics. Lactate dehydrogenase A (LDHA) is a key enzyme in the aerobic glycolysis pathway, an abnormal metabolic pathway commonly observed in cancers, associated with tumour progression and metastasis. Studies indicate MBs have a glycolytic phenotype; however, LDHA has not yet been explored as a therapeutic target for medulloblastoma. LDHA expression was examined in medulloblastoma subgroups and cell lines. The effects of LDHA inhibition by oxamate or LDHA siRNA on medulloblastoma cell line metabolism, migration and proliferation were examined. LDHA was significantly overexpressed in Group 3 and Wnt MBs compared to non-neoplastic cerebellum. Furthermore, we found that oxamate significantly attenuated glycolysis, proliferation and motility in medulloblastoma cell lines, but LDHA siRNA did not. We established that aerobic glycolysis is a potential therapeutic target for medulloblastoma, but broader LDH inhibition (LDHA, B, and C) may be more appropriate than LDHA inhibition alone.

INTRODUCTION: Medulloblastomas (MBs) are the most common solid malignant childhood brain tumour. Integrated genomic data has identified four distinct medulloblastoma sub-groups (Wnt, Shh, Group 3 and Group 4), which have different characteristic genetic abnormalities, and result in different clinical outcomes. Group 3 has the poorest prognosis, is the most frequently metastatic and characteristically over-expresses c-Myc. Lactate dehydrogenase A, known for its key role in aerobic glycolysis, is a downstream target of c-Myc and HIF1α. Previous studies using magnetic resonance spectroscopy and 18fluorodeoxyglucose positron emission tomography have shown that medulloblastomas have a glycolytic metabolic phenotype. We hypothesised LDHA inhibition would result in a decrease in lactate concentrations and a change from a glycolytic to an oxidative phosphorylation metabolic phenotype, leading to decreased medulloblastoma viability, proliferation and motility. METHODS: We used oxamate, a structural analogue of pyruvate, which competes with pyruvate to inhibit LDHA activity and LDHA siRNA to investigate the therapeutic potential of targeting LDHA in MB. RESULTS: We found that LDHA expression was significantly elevated in Group 3 and LDHB was significantly elevated in SHH medulloblastomas compared to non-neoplastic cerebellar tissue. Furthermore we showed that oxamate significantly inhibited LDHA activity, lactate production, aerobic glycolysis, proliferation and motility in medulloblastoma cell lines under normoxic and hypoxic conditions (1% O2) and also upregulated oxidative phosphorylation under normoxic conditions. Additionally, at low concentrations, oxamate inhibited LDHA activity and lactate production in the non-neoplastic paediatric astrocyte cell line CC2565 but it had no significant effects on the amount of viable, metabolically active cells or their motility under normoxic or hypoxic conditions. Despite showing 80% LDHA knockdown, our LDHA siRNA transfections did not have any significant downstream effects on MB cell line metabolism, growth or motility. However we found that LDHA siRNA also inhibited LDHC expression but not LDHB expression. CONCLUSIONS: It is possible that oxamate inhibits multiple LDH family members as the active site for LDH isoenzymes, comprised of LDHA and LDHB subunits are identical. The results of further investigations will be critical as LDHA may prove to be an inadequate target for MB and a broader LDH family inhibitor or lactate inhibitor may be more appropriate. However these studies, combined with extensive research into the literature, support the concept and provide proof of principle that targeting aerobic glycolysis and lactate production in medulloblastoma is worthwhile therapeutic avenue worth pursuing further.

INTRODUCTION: Medulloblastomas (MBs) are the most common type of solid, malignant childhood brain tumour. Recently, integrated genomics has identified four distinct sub-groups of which display different demographics, and clinical outcomes. Approximately 30% of MB patients present with metastatic spread at the time of diagnosis. Group 3 are considered the most metastatic, carry the worst prognosis and characteristically overexpress c-Myc. C-Myc, is involved in the initiation of several genes responsible for tumour growth and metabolism. Lactate dehydrogenase A (LDHA), a downstream target of c-Myc is often de-regulated in metastatic and invasive tumours. We have begun to investigate the role of LDHA and lactate on MB metabolism. METHOD: LDHA protein was examined in three medulloblastoma cell lines using Immunocytochemistry (ICC) and western blot analysis (WB). To investigate the role of LDHA in MB cell lines, LDHA was inhibited either by shRNA or with sodium oxamate. Metabolic assays and analysis include lactate and pyruvate assays. RESULTS: The three MB cell lines demonstrated varying levels of LDHA protein however ICC analysis showed consistency in localisation of LDHA in cytoplasmic and nuclear areas. Successful knockdown of LDHA protein using shRNA was confirmed with WB analysis. Knockdown of LDHA resulted in decreased lactate levels. LDHA inhibition by sodium oxamate also caused a decrease in lactate levels in a time and concentration dependent manner. Interestingly, pyruvate levels also decreased when LDHA was inhibited. CONCLUSION: Targeting medulloblastoma metabolism, such as LDHA, warrants further investigation to determine if this approach will affect not only metabolism but also cancer cell spread.