Therapy of glioblastoma, a particularly difficult-to-reach cancer type, is highly limited by the blood brain barrier (BBB) and will benefit greatly from effective drug and contrast agent delivery. Here, we are expanding on the functionality of M13 bacteriophage as an engineerable nanoprobe for targeting glioma cancer sites. M13 bacteriophage, a naturally monodisperse multifunctional nanostructure, consists of thousands of distinct protein subunits organized in a high aspect ratio, filamentous viral capsid; 900nm in length and 6nm in diameter. All M13 coat proteins are amenable to mutation, and can be tuned for the binding and nucleation of inorganics and nanoparticles, chelation of metal ions, and expression of targeting ligands or even enzymes. To harness these capabilities for medical imaging and therapy applications, we (a) develop a chlorotoxin (CTX) motif on the M13 capsid to cross the BBB, and (b) tailor the assembly of M13 into smaller “inho” phages. CTX display allows phage crossing of the BBB as well as uptake by glioma cells. Second window near infrared (NIR-II) through-skull mouse imaging of CTX-phage, complexed with single walled carbon nanotubes (SWNT) or small molecule NIR-II dyes, reveals localization of the phage to patient derived, orthotopic glioma xenografts, and makes possible phage based shuttling of drug loads to the tumor cells and simultaneous, non-invasive NIR-II tracking of the therapy progression. Such NIR-II optical imaging in the 900nm to 1700nm range allows high depth of tissue penetration and low tissue autofluorescence using quick, inexpensive machinery. To further build on the delivery potential of the M13 phage based nanoprobes, we have constructed a phagemid assembly system to alter the length of M13. Filamentous phages of short lengths, sizes ranging from 20nm to 900nm, are made possible by constructing our own set of small viral ssDNA that are packaged by M13 capsid proteins. With the ability to control the aspect ratio of these rigid, rod-like “inho”-phages we can improve on M13 based disease detection by optimizing for <i>in vivo </i>phage trafficking and tumor extravasation, focusing on inho-phage sizes 20nm to 200nm. Overall, targeted and miniaturized M13 phage is considered here as a complete nano-theranostic that could augment the therapeutic efficacy of drugs shuttled to the site of glioma while also providing clinicians with the information needed to easily track and debulk early-stage, deeply embedded tumor masses.

BBB-penetrating delivery to glioma using engineered, size-controlled nano phage

2019 Fall National Meeting

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