Aurora A kinase inhibition destabilizes PAX3-FOXO1 and MYCN and synergizes with Navitoclax to induce Rhabdomyosarcoma cell death

The clinically aggressive alveolar rhabdomyosarcoma subtype is characterized by expression of the oncogenic fusion protein PAX3-FOXO1, which is critical for tumorigenesis and cell survival. Here we studied the mechanism of cell death induced by loss of PAX3-FOXO1 expression and identified a novel pharmacological combination therapy that interferes with PAX3-FOXO1 biology at different levels. Depletion of PAX3-FOXO1 in fusion positive (FP)-RMS cells induced intrinsic apoptosis in a NOXA-dependent manner. This was pharmacologically mimicked by the BH3 mimetic navitoclax, identified as a top compound in a screen of 208 targeted compounds. In a parallel approach, and to identify drugs that alter the stability of PAX3-FOXO1 protein, the same drug library was screened and fusion protein levels were directly measured as a read-out. This revealed that inhibition of Aurora kinase A most efficiently negatively affected PAX3-FOXO1 protein levels. Interestingly, this occurred through a novel specific phosphorylation event in and binding to the fusion protein. Aurora kinase A inhibition also destabilized MYCN, which is both a functionally important oncogene and transcriptional target of PAX3-FOXO1. Combined treatment with an Aurora kinase A inhibitor and navitoclax in FP-RMS cell lines and patient-derived xenografts synergistically induced cell death and significantly slowed tumor growth. These studies identify a novel functional interaction of Aurora kinase A with both PAX3-FOXO1 and its effector MYCN, and reveal new opportunities for targeted combination treatment of FP-RMS

Repeated intermittent oral amphetamine administration results in locomotor tolerance not sensitization

Background: The phenomenon of locomotor sensitization to injected amphetamine is well-characterised. The increased locomotor activity found acutely is enhanced with repeated intermittent treatment. This effect arises due to hypersensitization of the dopaminergic system and is linked to drug addiction. A clinical population exposed to chronic repeated intermittent amphetamine treatment, such as is found for attention deficit hyperactivity disorder (ADHD), may be expected to be more at risk of addiction following this treatment. However, evidence suggests the opposite may be true. This suggests the route of administration may determine the direction of effects. Aims and methods: We aimed to establish how an oral amphetamine treatment regimen, similar to that used in ADHD, impacts on locomotor activity, specifically whether tolerance or sensitization would arise. Healthy hooded Lister rats were given amphetamine (2 mg/kg, 5 mg/kg and 10 mg/kg) or a vehicle solution once daily for 4 weeks with a 5 day on, 2 day off schedule. Locomotor activity was measured on the first day of treatment to establish the acute effects and on the final day of treatment to examine the chronic effects. Results: As expected, acute doses of amphetamine increased locomotor activity, although this only reached statistical significance for the 5 mg/kg and 10 mg/kg doses. By contrast, after chronic treatment, animals administered these doses showed reduced activity indicating drug tolerance rather than sensitization had occurred. Conclusion: We suggest that the route of administration used in ADHD, which results in more stable and longer duration drug levels in the blood, results in tolerance rather than sensitization and that this effect could explain the reduced likelihood of substance addiction in those treated with psychostimulants for ADHD