Quantifying the Lymphatic Transport of Model Therapeutics from the Brain in Rats

In recent years, the drainage of fluids, immune cells, antigens, fluorescent tracers, and other solutes from the brain has been demonstrated to occur along lymphatic outflow pathways to the deep cervical lymph nodes in the neck. To the best of our knowledge, no studies have evaluated the lymphatic transport of therapeutics from the brain. The objective of this study was to determine the lymphatic transport of model therapeutics of different molecular weights and lipophilicity from the brain using cervical lymph cannulation and ligation models in rats. To do this, anesthetized Sprague-Dawley rats were cannulated at the carotid artery and cannulated, ligated, or left intact at the cervical lymph duct. Rats were administered C-14-ibuprofen (206.29 g/mol, logP 3.84), H-3-halofantrine HCl (536.89 g/mol, logP 8.06), or H-3-albumin (similar to 65,000 g/mol) via direct injection into the brain striatum at a rate of 0.5 mu L/min over 16 min. Plasma or cervical lymph samples were collected for up to 6-8 h following dosing, and brain and lymph nodes were collected at 6 or 8 h. Samples were subsequently analyzed for radioactivity levels via scintillation counting. For C-14-ibuprofen, plasma concentrations over time (plasma AUC(0-6h)) were >2 fold higher in lymph-ligated rats than in lymph-intact rats, suggesting that ibuprofen is cleared from the brain primarily via nonlymphatic routes (e.g., across the blood-brain barrier) but that this clearance is influenced by changes in lymphatic flow. For H-3-halofantrine, >73% of the dose was retained at the brain dosing site in lymph-intact and lymph-ligated groups, and plasma AUC(0-8h) values were low in both groups (<0.3% dose.h/mL), consistent with the high retention in the brain. It was therefore not possible to determine whether halofantrine undergoes lymphatic transport from the brain within the duration of the study. For H-3-albumin, plasma AUC(0-8h) values were not significantly different between lymph-intact, lymph-ligated, and lymph-cannulated rats. However, >4% of the dose was recovered in cervical lymph over 8 h. Lymph/plasma concentration ratios of H-3-albumin were also very high (up to 53:1). Together, these results indicate that H-3-albumin is transported from the brain not only via lymphatic routes but also via the blood. Similar to other tissues, the lymphatics may thus play a significant role in the transport of macromolecules, including therapeutic proteins, from the brain but are unlikely to be a major transport pathway from the brain for small molecule drugs that are not lipophilic. Our rat cervical lymph cannulation model can be used to quantify the lymphatic drainage of different molecules and factors from the brain.