Convection-enhanced delivery of M13 bacteriophage to the brain

Laboratory investigation

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Object

Recent studies indicate that M13 bacteriophage, a very large nanoparticle, binds to β-amyloid and α-synuclein proteins, leading to plaque disaggregation in models of Alzheimer and Parkinson disease. To determine the feasibility, safety, and characteristics of convection-enhanced delivery (CED) of M13 bacteriophage to the brain, the authors perfused primate brains with bacteriophage.

Methods

Four nonhuman primates underwent CED of M13 bacteriophage (900 nm) to thalamic gray matter (4 infusions) and frontal white matter (3 infusions). Bacteriophage was coinfused with Gd-DTPA (1 mM), and serial MRI studies were performed during infusion. Animals were monitored for neurological deficits and were killed 3 days after infusion. Tissues were analyzed for bacteriophage distribution.

Results

Real-time T1-weighted MRI studies of coinfused Gd-DTPA during infusion demonstrated a discrete region of perfusion in both thalamic gray and frontal white matter. An MRI-volumetric analysis revealed that the mean volume of distribution (Vd) to volume of infusion (Vi) ratio of M13 bacteriophage was 2.3 ± 0.2 in gray matter and 1.9 ± 0.3 in white matter. The mean values are expressed ± SD. Immunohistochemical analysis demonstrated mean Vd:Vi ratios of 2.9 ± 0.2 in gray matter and 2.1 ± 0.3 in white matter. The Gd-DTPA accurately tracked M13 bacteriophage distribution (the mean difference between imaging and actual bacteriophage Vd was insignificant [p > 0.05], and was –2.2% ± 9.9% in thalamic gray matter and 9.1% ± 9.5% in frontal white matter). Immunohistochemical analysis revealed evidence of additional spread from the initial delivery site in white matter (mean Vd:Vi, 16.1 ± 9.1). All animals remained neurologically intact after infusion during the observation period, and histological studies revealed no evidence of toxicity.

Conclusions

The CED method can be used successfully and safely to distribute M13 bacteriophage in the brain. Furthermore, additional white matter spread after infusion cessation enhances distribution of this large nanoparticle. Real-time MRI studies of coinfused Gd-DTPA (1 mM) can be used for accurate tracking of distribution during infusion of M13 bacteriophage.

Abbreviations used in this paper:AAV = adenoma-associated virus; AD = Alzheimer disease; CED = convection-enhanced delivery; PBS = phosphate-buffered saline; PD = Parkinson disease; Vd = volume of distribution; Vi = volume of infusion.

Article Information

Address correspondence to: Russell R. Lonser, M.D., Surgical Neurology Branch, National Institute of Neurological Disorders and Stroke, National Institutes of Health, 10 Center Drive, Building 10, Room 3D20, Bethesda, Maryland 20892-1414. email: lonserr@ninds.nih.gov.

Please include this information when citing this paper: published online May 18, 2012; DOI: 10.3171/.2012.4.JNS111528.

© AANS, except where prohibited by US copyright law.

Headings

Figures

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    A: Schematic illustration showing the shape and exterior coat proteins of M13 bacteriophage. The M13 bacteriophage's coat is composed of approximately 3000 copies of major and several copies of minor proteins. This coat surrounds a single-stranded, closed circular DNA. B: Size comparison of M13 bacteriophage (900 nm) relative to albumin (8 nm), AAV (25 nm), and adenovirus (90 to 100 nm).

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    Immunohistochemical and MRI studies of thalamic infusion. Left: Representative 40-μm brain section showing immunostaining for M13 bacteriophage. This section shows a distinct dark-staining region in the thalamic gray matter without evidence of white matter tracking. The Vd is calculated by adding areas of the stained region in serial sections. Right: Coronal T1-weighted MRI study obtained with contrast agent showing CED to thalamic gray matter of bacteriophage M13 and Gd-DTPA. The distinct hyperintense region shows Gd-DTPA distribution (Vd) and corresponds to the M13 staining seen on immunohistochemical studies.

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    Magnetic resonance imaging and immunohistochemical studies of the frontal white matter infusion showing M13 bacteriophage distribution and axonal transport. A: Coronal T1-weighted MRI study obtained with contrast agent showing CED to frontal white matter of bacteriophage M13 and Gd-DTPA. The distinct hyperintense region shows Gd-DTPA distribution (Vd) and corresponds only to the dark-staining region seen on immunohistochemical studies. B: Representative 40-μm section showing immunohistochemical staining for bacteriophage M13 in frontal white matter. This section shows a distinct darker-staining region representing the initial infusion, with a lighter-stained region extending through the corpus callosum and to the cortex, representing axonal transport. C: Immunohistochemical staining for bacteriophage M13 at higher magnification (× 10) of inset in panel B, further showing lighter-stained M13 bacteriophage extension emanating from the initial infusion site. D: Immunohistochemical staining for bacteriophage M13 at high magnification (× 40) of inset in panel C, showing fibers (arrows). This is histological evidence for axonal transport.

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    Graph showing a linear relationship between MRI-confirmed Vd and Vi. There was a linear relationship (R2 = 0.98) between Vi and Vd.

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