(442b) PAMAM Dendrimers for Brain Delivery of Therapeutics for the Treatment of Cerebral Palsy: Chemistry, In Vivo Efficacy and Imaging

Maternal intrauterine inflammation resulting in microglial activation has been implicated in the development of periventricular leukomalacia and cerebral palsy. N-acetyl cysteine (NAC) is a drug that is currently being explored for the treatment of neuroinflammation in neonatal and perinatal applications. However, plasma binding of NAC significantly reduces the bioavailability requiring very high doses (100-300 mg/kg in animal models). Our translational research effort is focused on the preparation, in vitro, and in vivo evaluation of PAMAM dendrimer-based nanodevices for the delivery of therapeutics in general, and NAC in particular.

Nanodevice preparation and characterization: We have developed neutral PAMAM dendrimer-based nanodevices where a disulfide linker is used to link the drug to the dendrimer. The conjugates were characterized, by 1H NMR, MALDI-TOF, and HPLC. Each molecule of the dendrimer had approximately 16 molecules of NAC, and the conjugate exhibited high water solubility. The presence of the disulfide linker enables tailored intracellular release of the drug in a manner sensitive to the glutathione levels (low in blood circulation, high inside the cells).

In vivo imaging: FITC-labeled dendrimers were administered in neuroinflammation-associated pregnant rabbit models of cerebral palsy, through subdural and intravenous injections. The histological studies were correlated with microPET studies on 64Cu-labeled dendrimers. Our results are the first to show that dendrimers are able to cross the BBB in a neuroinflammation model. Our studies suggested that dendrimers localize in the activated microglial cells and astrocytes in the corpus callosum, hippocampus, corona radiata, internal capsula, external capsula and around the lateral ventricles in the endotoxin kits. In normal animals, which do not have appreciable activated microglia or astrocyte proliferation in these regions, no appreciable uptake was found. This paves the way for potentially useful drug delivery approaches using dendrimers for the treatment of neuroinflammation, even without specific targeting moieties. When this is combined with the ability of dendrimers to selectively localize in activated microglial cells, significant improvements in vivo performance can be achieved.

In vivo efficacy: The nanodevices were evaluated extensively in a rabbit model of cerebral palsy. The biodistribution and efficacy of intravenously administered dendrimer-drug conjugates are compared with those of the free drug using a combination of tools. The efficacy is evaluated using a combination of neurobehavioral analysis, assessment of brain tissue level inflammatory cytokine analysis. Our studies show that dendrimer-drug conjugates are 10-100 times more efficacious that free drug, suggesting that these conjugates (~ 18000 Da) are able to cross the blood brain barrier and deliver the drug significantly better than free drug. Detailed mechanistic and efficacy studies will be presented.