University of Minnesota Minneapolis, Minnesota, United States
Purpose: Medulloblastoma (MB) is the most common malignant pediatric brain tumor and has the potential for leptomeningeal spread, often responsible for the preponderance of morbidity and mortality in MB patients. Panobinostat is a non-selective histone deacetylase (HDAC) inhibitor that has been shown to be effective against MBs in vitro. It has been widely believed that panobinostat has limited blood-brain barrier (BBB) penetration. However, the statements regarding the central nervous system (CNS) penetration of panobinostat have been divergent. The objective of the current study is to evaluate the key pharmacokinetic (PK) parameters and the mechanisms influencing the CNS distribution of panobinostat in mouse models, that are often used in MB preclinical efficacy trials. Methods: An in vitro stability study was conducted in PBS, mouse plasma, brain homogenate, spinal cord homogenate, and human plasma at 37°C, room temperature, and 4°C, with agitation for 24 hours. The pH of PBS and all mouse matrices was adjusted to 7.4. Pharmacokinetic studies were conducted using FVB wild-type (WT) and P-glycoprotein-knockout (PKO, Mdr1a/b–/–) transporter-deficient mice. Panobinostat was administered intravenously at 10 mg/kg and an LC-MS/MS method was developed to quantify the concentrations of panobinostat at selected time points in the plasma, brain and spinal cord. PK parameters, including systemic clearances, volumes of distribution, half-lives, and areas-under-the-concentration time curve (AUC) were calculated using WinNonlin with non-compartmental analysis. Brain and spinal cord regional distribution of panobinostat was examined in WT mice. The brain was divided into six regions based on the anatomical structure of the mouse brain, including cortex, thalamus and hypothalamus, cerebellum, midbrain, pons and medulla. The spinal cord was separated into the cervical, thoracic and lumbosacral regions. Concentrations in each region was determined separately. Results: Panobinostat was stable in PBS and human plasma, however, instable in the various mouse matrices in vitro. The decomposition rate of panobinostat was temperature-dependent, with the fastest rate observed at 37°C and the lowest rate at 4°C. Therefore, all samples from animal studies were snap-frozen using dry ice and kept at -80°C until analysis. In WT mice, the CNS penetration was initially limited; however, it increased with time and the brain-to-plasma (Kp, brain) and spinal cord-to-plasma (Kp, spinal cord) distributional partition coefficients (AUCinf ratios) were 1.37 and 0.76, respectively. The Kp, brain and Kp, spinal cord of panobinostat in BBB transporter-deficient mice (PKO) were 2.56- and 2.8-fold higher than those in WT mice, respectively. This suggests that P-glycoprotein is limiting the CNS penetration of panobinostat. The spatial distribution of panobinostat in the mouse brain following systemic administration was heterogeneous. The rank order of panobinostat brain-to-plasma ratio (concentration ratios) at 4 hour after administration was thalamus and hypothalamus > pons > cerebellum > whole brain > midbrain > cortex > medulla. Concentrations in different spinal cord regions were not significantly different from each other. Conclusion: Panobinostat has shown different in vitro stability in human and mouse plasma, which could be translated to in vivo PK in the two species. The instability in mouse plasma might help explain the failure of panobinostat to generate in vivo efficacy following systemic administration even though exhibiting robust efficacy in vitro. Our data suggest that P-glycoprotein localized in the BBB can limit the CNS delivery of panobinostat and influence the delivery to tumors in the brain following systemic intravenous administration. The pons and cerebellum had levels greater than the whole brain average, indicating that treating the MBs would not be limited by the heterogeneous drug distribution. As such, studies investigating targeted administration routes that can bypass the BBB, such as intrathecal injection of novel formulations, are ongoing.
Acknowledgements: The studies are founded by the Eunice Kennedy Shriver National Institute of Child Health and Human Development (RO1 - HD099543) and National Institutes of Health (NIH)-National cancer institute (NCI) (U19 - CA264362).
