1. What is “CED”?

by The Infusion Physics Study Group*

5-14-09

Getting large molecules into the brain:

The brain is naturally protected from harmful agents by the blood-brain barrier (BBB), a

cellular barrier that selectively controls the movement of molecules between the circulating

blood and the neuronal tissue. It allows the movement of substances essential to metabolic

function but restricts the passage of large molecules (proteins and microorganisms). This

capacity to block the entrance of large molecules has impeded progress in developing

effective drugs for many years.

A way around the BBB is by direct intracerebral infusion, an invasive method. A cannula (or

hollow needle) is inserted into the brain through an opening created in the skull (burr hole)

until its tip reaches the vicinity of where the treatment is needed. Then, a solution containing

the drug is infused through the cannula.

Distributing the substance in the brain:

Once the solution is in the brain, it needs to be distributed throughout the intended target.

There are two mechanisms for dispersion: diffusion and convection.

Diffusion is the process in which the drug molecules move from regions of high concentration,

as prevails immediately at the cannula opening, to regions of lower concentration. Through

spontaneous motion diffusion is very slow at temperatures tolerable for living tissue. As

represented in Figure 1, it produces a concentration profile that drops off very rapidly and

results in limited spreading of the drug.

As an alternative to diffusion, NIH researchers proposed to use the pressure built-up in the

cannula to force the solution containing the drug through the free spaces that exist in the

surrounding tissue1. In this method, the solution spreads further with a more homogenous

concentration of the drug, as shown in Figure 2. This method was named “Convection

Enhanced Delivery” (CED).

Experience with pressure-driven infusion:

A typical method to study the dynamics of infusion is to inject a colored solution into a

medium that has some of the characteristics of brain tissue. Agarose gel is widely used for this

purpose. As it is transparent, the distribution of a colored solution can be observed with the

naked eye.2

A detailed description of the experimental methods we used can be found in a

companion report, “Factors Affecting Drug Distribution Through Infusion.”

Figure 3A shows an example of ideal convection enhanced delivery in a gel model: a

spherical distribution of the colored solution. As the infusion proceeds, the sphere grows.

However, instead of spreading evenly in all directions, the infusate frequently moves up along

the outer surface of the cannula. This type of flow, illustrated in Figure 3B, is called

backflow.

Both phenomena (spherical distribution or backflow) also occur in vivo. They can be

visualized in “real time” by infusing a solution containing a contrast agent, such as

gadolinium, while performing high resolution Magnetic Resonance Imaging (MRI) 3 A

description of the experimental methods we used can be found in the companion report

mentioned above.

Figure 4A shows a coronal MRI view of a brain region with a predominantly spherical

distribution of infusate in the putamen nucleus, a gray matter area of the brain often targeted

to treat Parkinson’s disease. Figure 4B shows a different case in which there is intense

backflow. As the flow moves up along the outside of the cannula, it may reach the corpus

callosum. This is highly undesirable as the corpus callosum will divert (leak) the infusate

away from the putamen.

The success of convection enhanced delivery by infusion depends on identifying a method

that gives:

maximum coverage of the target brain region,

in a reproducible manner,

without causing leakage.

Flow through the cannula:

The flow through the cannula and the flow from the cannula into the medium are described by

two laws of classical fluid dynamics. Poiseuille’s Law describes the flow through the

cannula, with the equation:

P = Q * L/KA

where P is the pressure difference applied over a cannula

L is the length of the cannula

A is the cross-sectional area of the cannula

Q is the flow rate

and K is called the hydraulic conductivity, a property of the

solution

Flow in the surrounding medium:

The flow of the infused solution in the medium, away from the tip of the cannula, is described

by D’Arcy’s Law, which says that the pressure that must prevail at the tip of the cannula is

proportional to the rate at which fluid is transported away from the tip, Q. The slope of

proportionality is related to the permeability of the surrounding material.

Figure 5 shows the pressure required to infuse a solution into water and into 0.6% agarose gel

for different flow rates. The infused solution and cannula length is the same in both cases, yet

the slope is steeper in the case of gel.

It appears that both the resistance of the cannula (and its design) and the resistance of the

surrounding medium (its permeability) are at work.

The pressure observed at a flow rate of 1 µL/min is shown for different gels, and also for dead

and living tissue in Table 1. As expected, denser gels – which have fewer pores and

consequently lower permeability – provide a higher resistance to flow, requiring higher

pressure.

Magnetic Resonance Imaging (MRI) measures distribution in tissue:

MRI can be used to evaluate drug infusion by measuring the distribution of the drug relative

to the desired target brain structure. For Parkinson’s disease, the target brain structure is

usually the putamen. The drug is made visible in MRI by adding a contrast agent to the

solution such as Gadolinium that appears bright in the image.

MRI gives three parameters necessary to quantify the infusion:

Vd: The distribution volume

The pressure at the tip of the cannula pushes the injected solution (volume Vi ) into the free

spaces in the medium. Because these spaces make up only a part of the total volume of the

medium, the infusate will occupy a larger volume than Vi of the surrounding medium. The

volume occupied is called the distribution volume, Vd.

Vput: Distribution volume within the putamen

The distribution volume of drug within the putamen is calculated by measuring the region of

Gadolinium enhancement after infusion within the boundary of the putamen. Ideally, Vput will

be as close as possible to Vd. If Vput is smaller than Vd some infusate presumably escaped.

%put: Portion of putamen covered by infusate

The third parameter calculated from the MRI is the percentage of the putamen that is covered

by the infusate. It is calculated by dividing the volume of infusate in the putamen (Vput) by the

total volume of the putamen. %put needs to be high enough for biologial effectiveness.

A successful infusion is shown in Figure 6. The concentration of the infusate around the

cannula tip follows an approximately spherical distribution (red is the highest, violet is the

lowest). The parameters of this infusion, Vi = 95 µL, Vd = 569 µL, and consequently Vd/Vi =

5.9, suggest that the interstitial volume is about 15% of the tissue volume. 4

Near the catheter,

the concentration is higher. This likely indicates that the pressure of the infusion has

expanded the extracellular space locally. The ratio, Vput/Vd, is 0.47, suggesting that almost

half of the infusate ended up within the desired region. We achieved coverage of 49% of the

total putamen volume. At the outer edges of the infusion, the concentration drops to low

levels, probably due to the effect of diffusion.

Does infusion work in humans as expected?

In Parkinson’s Disease, infusions are intended to deliver drugs, such as growth factors, to the

main areas affected by the disease. As we mentioned before, the putamen is a preferred target.

Interspecies differences in the sizes of the brain regions can be significant and require

different methods of infusion for optimal delivery. Figure 7 shows the volume of the

putamen for rats, non human and human primates7

The large size of the human putamen relative to animal models makes it harder to achieve

uniform coverage of the entire putamen in humans than in animal models. The inadequacy of

infusion methods in human applications was illustrated in an analysis of clinical trials testing

the effects of Glial Derived Neurotrophic Factor (GDNF). 5 Replicating the infusion

conditions in animal models suggested that the growth factor delivery in a key clinical trial

may have reached less than 10% of a human putamen.6

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Next – a look at details of infusion methods.

*The Infusion Physics Study Group:

Research Contributor

Senior investigators and in

vivo/ex vivo experiments Dr. Krystof Bankiewicz, University of

California at San Francisco

Dr. Marina E. Emborg, University of

Wisconsin, Madison

Fluid physics Dr. Raghu Ragavan, Therataxis

Dr. Martin Brady, Therataxis

Simulations/engineering Chris Ross, Engineering Resources

Group, Inc.

MRI physics Dr. Andrew Alexander, University of

Wisconsin, Madison

Dr. Tracy McKnight, University of

California at San Francisco

Technical contributors

(in alphabetical order) Janine Beyer, University of California at

San Francisco

John Bringas, University of California at

San Francisco

Dr. Kevin Brunner, University of

Wisconsin, Madison

Michael Dobbert, University of

Wisconsin, Madison

Ronald Fisher, University of Wisconsin,

Madison

Valerie Joers, University of Wisconsin,

Madison

Philip Pivirotto, University of California

at San Francisco

James J. Raschke, University of

Wisconsin, Madison

Dr. Dali Yin, University of California at

San Francisco

Elizabeth Zakszewski, University of

Wisconsin, Madison

Project management Ken Kubota, Kinetics Foundation

Tom Dunlap, Kinetics Foundation

References:

1. A good summary of early work can be found in “Convection-enhanced delivery of

macromolecules in the brain”, R. Hunt Bobo, Douglas W. Laske, Aytac Akbasak, Paul F.

Morrison, Robert L. Dedrick, and Edward H. Oldfield, Proc. Natl. Acad. Sci. USA, Vol. 91,

pp. 2076-2080, 1994

More recent work is summarized, with application to brain tumors, in “Convection-enhanced

delivery of nanocarriers for the treatment of brain tumors”, Emilie Allard, Catherine Passirani,

Jean-Pierre Benoit, Biomaterials, Vol. 30, pp. 2302-2318, 2009

2. A thorough study using gels can be found in “A realistic brain tissue phantom for

intraparenchymal infusion studies”, Zhi-Jian Chen, George T. Gillies, William C. Broaddus,

Sujit S. Prabhu, Helen Fillmore, Ryan M. Mitchell, Frank D. Corwin and Panos P. Fatouros,

J. Neurosurg, 101: 314-322, 2004

3. For an early review of this technique, please see “Real-time Imaging and Quantification of

Brain Delivery of Liposomes”, Michal T. Krauze, John Forsayeth, John W. Park, and Krystof

S. Bankiewicz, Pharmaceutical Research, pp. 1-12, 2006

4. This is of the same magnitude as observed using a different experimental technique. See

“Diffusion in Brain Extracellular Space”, Eva Sykova and Charles Nicholson, Physiol. , Rev

88, pp. 277-1340, 2007

5. An overview of all these trials can be found in “Crossroads in GDNF Therapy for

Parkinson’s Disease”, Todd B. Sherer, PhD, Brian K. Fiske, PhD, Clive N. Svendsen, PhD,

Anthony E. Lang, MD, FRCP, and J. William Langston, MD, Movement Disorders, Vol 21,

No. 2, pp. 136-141, 2006

A good summary of the different clinical trials from the point of view of infusion can be

found in “Convective delivery of glial cell line-derived neurotrophic factor in the human

putamen”, Paul E. Morrison, PhD, Russell R. Lonser, MD, and Edward H. Oldfield, MD, J.

Neurosurg., Vol. 107, pp. 74-83, 2007

6. “Point source concentration of GDNF may explain failure of phase II clinical trial”,

Michael F. Salvatore, Yi Ai, Brent Fischer, Amanda M. Zhang, Richard C. Grondin, Zhiming

Zhang, Greg A. Gerhardt, Don M. Gash, Experimental Neurology, 2006

7. “The Human Brain in Figures and Tables, A Quantitative Book,” Samuil M. Blinkov and

Il’ya I. Gezer, Basic Books, Inc, Publishers Plenum Press, Table 135, 1968.

Cannula

1-2 mm

Concentration

Distance from cannula0

Figure 1 Sketch of the concentration distribution of a drug spread by diffusion

from the point where it is introduced.

Figure 2 Sketch of the concentration distribution of a drug spread by flow from a pressurized

cannula. Note that the concentration remains constant from the point of

introduction out to the periphery of the distribution.

Cannula

Concentration

Figure 3a Ideal infusion into gel. The colored infusate is distributed spherically.

Time

Figure 3b Backflow: the infusate moves up along the outer surface of the cannula.

Figure 4a In vivo coronal MRI; “good infusion”.

Figure 4b In vivo coronal MRI; with backflow.

Figure 5 Pressure vs. Flow Rate for infusion into water and into 0.6% agarose gel. The

pressure is proportional to the flow rate. The infusate requires more pressure to

move into a medium with lower permeability.

Table 1 Infusing into tissues with lower permeability requires higher pressure to maintain

the same flow rate. For example, infusing into tissue requires higher pressure than

infusing into gel or water.

The flow rate in all cases is held constant at 1uL/min and the pressure is measured

in mmHg.

60.0Ex vivo liver

23.0Ex vivo brain

12.0In vivo brain

5.4Gel, 2.0

4.0Gel, 1.0

0.4Gel, 0.6

0.25Gel, 0.2

0.20Water

PressureMedium

Mode of

Distribution

Threshold

(mmol/l)

Vi

(µl)

Vd

(µl)

Vd/Vi Vd(put)

(µl)

Vd(put)/

Vd

Coverage

(%)

Diffusion +

Convection

0.05 95 569 6.0 269 0.47 49%

Convection 0.20 95 223 2.4 143 0.64 26%

Figure 6 Illustration of a successful in-vivo infusion into the putamen. The image on the left

displays the effects of both convection and diffusion with the concentration of infused

gadolinium in color over a co-registered T1-map. The image on the right depicts the

approximate area covered by convection alone. The scale at the far left shows the

concentration in mmol/liter, and as a percentage of the infused concentration.

Figure 7 Typical putamen volumes of subjects in Parkinson’s studies.