Gerald Grant - US grants

The blood-brain barrier (BBB), a dynamic and selective interface between the blood and brain parenchyma, consists of endothelial cells ensheathed by glia that line brain microvessels. Preliminary data suggest that perivascular astrocytes and glucocorticoids (i.e. dexamethasone) modulate the induction and maintenance of BBB function, however, mechanisms of action for glia and dexamethasone (DEX) on the BBB are largely unknown. The notion that glial influences are essential for the induction and-maintenance of the i3BB, together with the fact that intraluminal flow causes further differentiation of EC, has prompted the development of sophisticated in vitro models of the BBB, such as the dynamic in vitro BBB model (DIV-BBB) (41,42,43,44). Recent advances in molecular biotechnology have produced tools capable of analyzing patterns of changes in gene expression (i.e. cDNA array hybridization) on a comprehensive scale at distinct time points in a biological process. Applying these tools to the study of BBB induction will provide insight into the critical determinants of endothelial differentiation and BBB repair mechanisms. We hypothesize that 1) endothelial cell changes in gene expression can be identified and characterized at distinct time points which correlate with phenotypic changes; and that 2) DEX may affect non-brain endothelial pathways of gene expression-phenotypic changes similar to those observed during normal BBB induction by glia. A systematic study of this complex biological process has not been performed and would provide valuable insight into strategies for the treatment of human disease and CNS drug delivery.

DESCRIPTION (provided by applicant): The current treatment of malignant diffuse gliomas with combined radiation and chemotherapy is limited and often results in high rates of persistent and recurrent disease with poor survival. Inadequate delivery across the blood-brain barrier (BBB) has been identified as a significant factor contributing to the failure of systemic chemotherapy for malignant brain tumors. The properties of the BBB that shield the brain from deleterious agents are thus the same that prevent drugs from treating disease. The primary goal of this work is to manipulate the molecular structure of the BBB to enhance drug delivery into a brain tumor. It was recently shown that targeted suppression of claudin 5, an endothelial cell specific tight junction protein on the BBB, following injection of siRNA targeting claudin 5, caused both a transient and size-selective increase in paracellular permeability of the BBB (Campbell et al, J Gene Med, 2008). Many preclinical studies testing molecular therapeutics and chemotherapeutics rely on xenograft tumor models to predict tumor response and survival. Preliminary work in the PI's laboratory demonstrated a preferential tumor microvessel vulnerability to the siRNA targeting claudin 5 compared to adjacent normal microvessels. The working hypothesis is that this novel strategy of targeting claudin 5 in vivo will preferentially target brain tumor microvessels and result in a transient tumor selective opening of the BBB, also referred to as the blood-tumor barrier. To better characterize the molecular and functional changes with this approach, we propose to test the following: 1) characterize the temporal course of modulation of BBB permeability;2) determine the molecular pore size threshold in the tumor microvessels and extent of various sized tracers into the perivascular space using intravital cranial window microscopy;and 3) test this approach to augment the delivery of known antitumor agents. Using this novel molecular approach, these important studies will lay the foundation for future translational studies to better deliver chemotherapeutics into malignant brain tumors and the surrounding microenvironment. PHS 398/2590 (Rev. 09/04, Reissued 4/2006) Page Continuation Format Pag1e PUBLIC HEALTH RELEVANCE: Virtually all malignant gliomas in children recur even after the most aggressive combination therapy, usually at the original site of presentation. Poor drug delivery is one of the important features which is responsible for treatment failure and recurrence. The studies outlined here in mice will set the stage for future treatment strategies and test a novel molecular strategy to selectively open the tumor barrier to enhance the delivery of chemotherapy agents into the brain and avoid systemic toxicity.

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. Terminal study with these animals could be performed under our protocol entitled "Mapping the Rodent Brain with MR Microscopy.", A125-04-04 approved 04/22/04. This protocol would have to be modified for survival studies. However, as I understand, animals leaving CCIF cannot be returned. Additionally, being nude mice they are immuno-compromised. For starters, they could be brought over here and imaged the same day then sacrificed. The goal of this study is to perform in vivo MR permeability studies to assess the correlation of BBB permeability with tumor size and rate of tumor growth in a mouse xenograft brain tumor model. Congenitally athymic nude mice will be injected with an intracranial human glioblastoma multiforme xenograft. We have demonstrated that the increase in blood-tumor barrier permeability correlates with intracranial tumor size and that there is marked heterogeneity in permeability within each xenograft tumor as well as between xenograft tumors. The tumor core in the xenograft mouse model is characterized by the highest tumor permeability which inversely correlates with the distance from the tumor. The pilot study will image these mice using MRI (7T) between days 10-15 post injection of the xenograft using an MR contrast agent (ProHance) injected via a tail vein under anesthesia. Following the MRI, standard quantitative permeability measurements will be performed using 14C labeled AIB (ARC 145B-aminoisobutyric acid) in the CCIF which will validate the MR permeability data. For these pilot studies, we will use the xenograft tumor that is rapidly growing (GBM 270) and with a high blood-tumor barrier permeability. It remains unknown how the blood-tumor barrier permeability changes with increased tumor size. In addition, the ability to image the modulation of the blood-brain and blood-tumor barrier permeability has tremendous translational appeal for predicting tumor-specific drug delivery and for validating molecular vascular targeting strategies to treat brain tumors.

The objective of this proposal is to disseminate mouthguard sensors to a community of researchers studying the short- and long-term effects of head impacts on brain health. Traumatic brain injury (TBI) has long been known to be a leading cause of death and disability among children and young adults. However, it has only recently been argued that concussion, a form of ?mild? TBI, increases the risk for Alzheimer's and other neurodegenerative diseases. Understanding the mechanisms of concussion is crucial for injury diagnosis and prevention. However, past research efforts have been unable to identify a clear link between head impact acceleration dose and neurological response indicative of injury. This is mainly due to the lack of a large, high quality concussion dataset. The difficulty in measuring rare injury scenarios has hampered individual investigators from making substantial progress. Thus the required dataset can only be gathered through large- scale, multi-institution dissemination of a validated head impact measurement sensor. We propose to widely disseminate our rigorously validated Stanford mouthguard (MiG2.0), and subsequently develop a platform to share collected data among many investigators. The MiG2.0 is a unique design that rigidly couples to the skull through upper dentition. This project will disseminate the MiG2.0 to a large community of concussion researchers that study high injury rate populations such as contact sports players. At the end of the project, we will have a large concussion database with accurate head impact measurements that will further our understanding of concussion mechanisms. Our long-term goal is that the knowledge gained from this dissemination effort will improve concussion management and long-term brain health by enabling (1) real-time field diagnosis, and (2) design improvements in protective equipment for athletics, the military, transportation and other high-risk activities.