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Summary

Convection-enhanced delivery (CED) is a method enabling effective delivery of therapeutics into the brain by direct perfusion of large tissue volumes. The procedure requires the use of catheters and an optimized injection procedure. This protocol describes a methodology for CED of an antibody into a mouse brain.

Abstract

Convection-enhanced delivery (CED) is a neurosurgical technique enabling effective perfusion of large brain volumes using a catheter system. Such an approach provides a safe delivery method by-passing the blood brain barrier (BBB), thus allowing treatment with therapeutics with poor BBB-permeability or those for which systemic exposure is not desired, e.g., due to toxicity. CED requires optimization of the catheter design, injection protocol, and properties of the infusate. With this protocol we describe how to perform CED of a solution containing up to 20 µg of an antibody into the caudate putamen of mice. It describes preparation of step catheters, testing them in vitro and performing the CED in mice using a ramping injection program. The protocol can be readily adjusted for other infusion volumes and can be used for injecting various tracers or pharmacologically active or inactive substances, including chemotherapeutics, cytokines, viral particles, and liposomes.

Introduction

The blood brain barrier (BBB) forms a semipermeable border separating the central nervous system (CNS) from the blood circulation. Reaching the CNS with therapeutics is however necessary in context of various diseases, like brain tumors, Alzheimer’s disease (AD) or Parkinson’s disease (PD) among others1. This becomes important in the development of new therapies, especially if the tested drug exhibits poor BBB permeability or its systemic exposure can lead to dangerous toxicity1,2. Some of the clinically used antibodies display both of these features. A solution to this prob....

Protocol

All methods described here have been approved by the Swiss Cantonal Veterinary Office under license number ZH246/15.

1. Preparation of the Step Catheters

  1. Preparation of a fused silica tubing for the step of the catheter
    1. Cut the fused silica capillary with inner diameter of 0.1 mm and wall thickness of 0.0325 mm tubing to a length of 30 mm.
    2. Examine the tubing for cracks and heat polish the ends using a microforge to ensure the tubing openings .......

Representative Results

This protocol enables preparation of step catheters (Figure 1) for use in the CED procedure in a laboratory environment. In order to control the catheters for leakage, reflux along the needle tract and clogging, we recommend performing injections of a dye, e.g., trypan blue solution, into an agarose block. Figure 3 depicts a cloud of trypan blue forming after injection of 1 µL at 0.5 µL/minute using a CED catheter (Figure 3A

Discussion

Convection-enhanced delivery, or pressure-mediated drug infusion into the brain, was first proposed in the early 19903. This approach promises perfusion of large brain volumes behind the blood brain barrier in a controlled manner2. However, so far, only a few clinical trials have been performed using this approach, partially because CED in a clinical setup has shown to be technically demanding24,25. Recent developme.......

Acknowledgements

This work was supported by grants of the University of Zurich (FK-15-057), the Novartis Foundation for medical-biological Research (16C231) and Swiss Cancer Research (KFS-3852-02-2016, KFS-4146-02-2017) to Johannes vom Berg and BRIDGE Proof of Concept (20B1-1_177300) to Linda Schellhammer.

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Materials

NameCompanyCatalog NumberComments
10 μL syringeHamilton7635-01
27 G blunt end needleHamilton7762-01
AgarosePromegaV3121
AtipamezolJanssen
Bone waxBraun1029754
BuprenorphineIndivior Schweiz AG
CarprofenPfizer AG
Dental drill bits, steel, size ISO 009Hager & Meisinger1RF009
Ethanol 100%Reuss-Chemie AG179-VL03K-/1
FentanylHelvepharm AG
FITC-Dextran, 2000 kDaSigma AldrichFD2000S
FlumazenilLabatec Pharma AG
FormaldehydeSigma AldrichF8775-500ML
High viscosity cyanoacrylate glueMigros
Iodine solutionMundipharma
MedetomidinOrion Pharma AG
MicroforgeNarishigeMF-900
MidazolamRoche Pharma AG
Ophthalmic ointmentBausch + LombVitamin A Blache
PBSThermoFischer Scientific10010023
Polyclonal goat anti-rat IgG (H+L) antibody coupled with Alexa Fluor 647Jackson Immuno
ScalpelsBraunBB518
Silica tubing internal diameter 0.1 mm, wall thickness of 0.0325 mmPostnovaZ-FSS-100165
Stereotactic frame for miceStoelting51615
Stereotactic robotNeurostarDrill and Injection Robot
SuccroseSigma AldrichS0389-500G
Topical tissue adhesiveZoetisGLUture
Trypan blueThermoFischer Scientific15250061
WaterBichsel1000004

References

  1. Scherrmann, J. M. Drug delivery via the blood-brain barrier. Vascular Pharmacology. 38 (6), 349-354 (2002).
  2. Barua, N. U., Gill, S. S. Convection-enhanced drug delivery: prospects for glioblastoma treatment.

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