Brain drug delivery and meningeal permeability

The BBB: a challenge in drug development

The blood-brain barrier protects the brain from harmful pathogens, but it also blocks most drugs, particularly biologics, from reaching their target. Innovative strategies like nanoparticles or anti-TfR conjugation offer ways to bypass this barrier. Regardless of the approach, it’s crucial to quantitatively assess whether new drug candidates can cross the BBB. Neurotar’s in vivo trans-BBB pharmacokinetic (PK) assay is designed to provide this critical information.

Why two-photon microscopy for brain drug delivery?

In vitro assays, while useful, can’t replicate the complex environment of the living brain. To truly understand a compound’s ability to cross the BBB, it must be studied within a living organism. Conventional preclinical imaging falls short in resolution, but in vivo two-photon imaging excels.

Here’s why two-photon microscopy is the best choice for trans-BBB PK studies:

• High-resolution imaging down to the organelle level
• Conducted in the brains of living mice
• Reliable quantification of trans-BBB transport dynamics
• Low phototoxicity enables longitudinal studies, allowing multiple assessments in the same brain region over time (days to months)
• Using the mouse as its own control enhances statistical power
• Possibility of using humanized mouse models to increase translational relevance.

Sample results from a trans-BBB PK study at Neurotar: The vasculature of the somatosensory cortex was imaged using two-photon microscopy at the specified time points following the injection of fluorescently labeled lead compounds. Quantification of parenchymal and vascular fluorescence shows that Lead 1 effectively crosses the BBB.

What drug delivery and permeability studies can Neurotar perform for you?

Visualizing large molecules crossing the blood-brain barrier in wild-type mice

We specialize in visualizing how large molecules cross the blood-brain barrier in wild-type mice. Please note that this assay utilizes fluorescent labeling, so the test compound’s molecular weight must exceed 10 kD.

Combining pharmacokinetics with target engagement in transgenic mouse models

Our capabilities extend to combining pharmacokinetics studies with target engagement in transgenic mouse models featuring labeled brain deposits (such as amyloid plaques, tau tangles, or synuclein deposits). Additionally, we can measure efficacy by assessing whether your lead compound reduces amyloid plaque burden. Check out our Abeta plaque dynamics page for more details. What’s more, we also combine pharmacokinetics studies with localization analysis in mice with genetically or virally labeled cell populations.

Sample results from a combined BBB PK and Abeta plaque targeting study at Neurotar: The somatosensory cortex of an APPSL mouse (an Alzheimer’s model) was imaged using two-photon microscopy at specified time points following the injection of a fluorescently labeled antibody. Data at 24 hours demonstrate that the lead antibody crosses the BBB, while images at 72 and 144 hours show targeted binding to Abeta plaques.

Studying trans-BBB PK in the hippocampus

We utilize two-photon imaging via glass microperiscopes to study trans-BBB pharmacokinetics in the hippocampus. This assay was developed with expertise transferred from Micael Goard’s lab at the University of California, Santa Barbara. Follow this link to the relevant publication.

Meningeal permeability and vasodilation studies

We investigate dural permeability and vasodilation associated with local neuropeptide release, which is key in migraine development. By measuring FITC-dextran accumulation in dural tissue in PACAP-treated vs. vehicle-treated controls, we can clearly differentiate between dural and cortical tissues, focusing on the meninges — a hotbed of migraine pain. Additional readouts include the diameter of middle meningeal artery (MMA) branches and blood flow rate.

How can Neurotar facilitate your drug delivery assessment?

Neurotar is the world’s leading commercial provider of in vivo two-photon brain imaging in mice. With over a decade of experience, we’ve gained valuable insights from observing multiple drug candidates attempting to cross the blood-brain barrier. We’re eager to share this expertise with you.

Here’s how we can assist:

• Initial consultation: We’ll begin by discussing your research questions. Together, we’ll select the most suitable compound labeling ratio and concentration, and design a robust in vivo imaging study.
• Mouse line and model selection: We can help you choose the appropriate mouse lines or transgenic models and assist in sourcing licenses and mice from trusted commercial providers. Additionally, our team can advise on using viral vectors to highlight specific cell populations for localization studies.
• Optimizing drug administration: We’ll guide you in selecting the best drug administration routes — whether oral (gavage), i.p., s.c., i.v., i.t. (intrathecal), or Cisterna Magna injection.
• Imaging and analysis: After injecting a fluorescently labeled compound into the brain, we’ll repeatedly image the same cortical region, analyze the images, and quantify the redistribution through the BBB into the cortical parenchyma. Alternatively, we can focus on dural permeability, specifically examining the meninges and quantifying FITC-dextran accumulation.
• Tissue harvesting: If needed, we can harvest and preserve tissues at the end of the study for further analysis, either by your in-house team or a dedicated in vitro CRO partner.
• Reporting and support: Upon completing the study, we’ll present the results, interpret the data, and recommend follow-up steps. We’ll address all your questions and finalize the report based on your feedback.
• Publication assistance: If you wish to publish the study results, we’re here to help with publications, poster abstracts, investor presentations, and other communications.

Other two-photon brain imaging services offered by Neurotar

	Neurodeg. Disease (e.g. AD, PD)	Stroke and TBI	Neuropathic Pain and Migraine	Neuropsychiatry (e.g. Schizophrenia)	Epilepsy
Blood-brain barrier integrity	✓	✓	✓
Trans-BBB pharmacokinetics	✓	✓	✓
Dendritic spine turnover	✓	✓	✓	✓	✓
Microglial dynamics or response to injury	✓	✓	✓	✓	✓
Calcium signaling	✓	✓	✓	✓	✓
Abeta Plaque or Tau Tangle dynamics	✓	✓	✓
Mitochindria dysfunction	✓	✓	✓	✓	✓
Ischemic Stroke model		✓
Regeneration of peripheral neurons	✓	✓	✓

References

Neurotar’s publications

• Hoehlig K, Johnson K, Pryazhnikov E, Maasch C, Clemens-Smith A, Purschke W, Vauléon S, Buchner K, Jarosch F, Khiroug L, Vater A, Klussmann S. (2015) A novel CGRP-neutralizing Spiegelmer attenuates neurogenic plasma protein extravasation. Br J Pharmacol. DOI: 10.1111/bph.13110.
• Pradier L, Blanchard-Brégeon V, Bohme A, Debeir T, Menager J, Benoit P, Barneoud P, Taupin V, Bertrand P, Dugay P, Cameron B, Shi Y, Naimi S, Duchesne M, Gagnaire M, Weeden T, Travaline T, Reczek D, Khiroug L, Slaoui M, Brunel P, Fukuyama H, Ravetch J, Canton T, Cohen C. (2018) SAR228810: an antibody for protofibrillar amyloid β peptide designed to reduce the risk of amyloid-related imaging abnormalities (ARIA). Alzheimers Res Ther. 2018 10(1):117. DOI: 10.1186/s13195-018-0447-y.

Other relevant publications

• Kadry H, Noorani B, Cucullo L. (2020) A blood–brain barrier overview on structure, function, impairment, and biomarkers of integrity. Fluids Barriers CNS. Nov 18;17(1):69. DOI: 10.1186/s12987-020-00230-3.
• Pardridge WM. (2020) Blood-Brain Barrier and Delivery of Protein and Gene Therapeutics to Brain. Front Aging Neurosci. Jan 10;11:373. DOI: 10.3389/fnagi.2019.00373.
• Bhowmik A, Khan R, and Ghosh MK. (2015) Blood Brain Barrier: A Challenge for Effectual Therapy of Brain Tumors. Biomed Res Int. 2015: 320941. Mar 19. DOI: 10.1155/2015/320941.
• Arvanitis CD, Ferraro GB, Jain RK. (2020) The blood–brain barrier and blood–tumour barrier in brain tumours and metastases. Nature Reviews Cancer. 20:26–41. DOI: https://doi.org/10.1038/s41568-019-0205-x.
• Pandit R, Chen L, Götz J. (2020) The blood-brain barrier: Physiology and strategies for drug delivery. Adv Drug Deliv Rev. 165-166:1-14. DOI: 10.1016/j.addr.2019.11.009.
• Ferraris C, Cavalli R, Panciani PP, Battaglia L. (2020) Overcoming the Blood-Brain Barrier: Successes and Challenges in Developing Nanoparticle-Mediated Drug Delivery Systems for the Treatment of Brain Tumours. Int J Nanomedicine. Apr 30;15:2999-3022. DOI: 10.2147/IJN.S231479.
• Parvez S, Kaushik M, Ali M, Alam MM, Ali J, Tabassum H, Kaushik P. (2022) Dodging blood brain barrier with “nano” warriors: Novel strategy against ischemic stroke. Theranostics. Jan 1;12(2):689-719. DOI: 10.7150/thno.64806.
• Abrahao A, Meng Y, Llinas M, Huang Y, …, Zinman L, (2019) First-in-human trial of blood-brain barrier opening in amyotrophic lateral sclerosis using MR-guided focused ultrasound. Nat Commun. Sep 26;10(1):4373. DOI: 10.1038/s41467-019-12426-9.
• Levy D, Moskowitz MA. Meningeal Mechanisms and the Migraine Connection. Annu Rev Neurosci. 2023 Jul 10;46:39-58. DOI: 10.1146/annurev-neuro-080422-105509.