M13 Phage Assembly System for Optimal Blood Trafficking, Tumor Penetration, and Passage Across the Blood Brain Barrier

Technology

This technology is a novel method of delivering therapeutics and imaging molecules to the brain using the filamentous bacteriophage M13. M13 phage are an attractive candidate for a molecular courier due to their small diameter (~5-6nm), rod-like shape, and the ability to modify coat proteins to attach functional molecules. These inventors made two major improvements to M13 phage to increase BBB penetration and GBM targeting. Firstly, they decreased the length of the phage. M13 phage length is directly proportional to the size of the packaged genome, so these inventors devised a two-plasmid system to decrease the size of the packaged ssDNA genome. One plasmid contains all of the coding information for the protein coat and packaging components, and the second contains only ssDNA of tunable length containing a packaging signal. By changing the size of the plasmid with the packaging signal, the length of the M13 phage can be finely tuned between 880nm for a wild type phage down to only 50nm for phage with a packaged genome of 285 DNA base pairs. Secondly, the inventors included a chlorotoxin (CTX) motif on the M13 phage. CTX has a high positive charge that aids in BBB penetration and has additionally been shown to target very specifically to the electronegative GBM extracellular matrix. The combined M13-CTX delivery system results in highly efficient BBB penetration and GBM targeting while additionally facilitating addition of therapeutic or imaging molecules to the M13 coat proteins. As a proof of principle, the inventors attached single walled carbon nanotubes (SWNT) to the M13-CTX phage and injected the composition into an in vivo mouse model of GBM. The SWNT-M13-CTX phage demonstrated efficient tumor targeting and facilitated sub-millimeter resolution near infrared imaging.

Problem Addressed

Glioblastoma (GBM) is the most common cancer of the brain. GBM is very fast growing and is considered one of the most lethal cancers, with a 5-year survival rate of only 5%. One of the major challenges in treating GBM is finding drugs that can cross the blood brain barrier (BBB), which is composed of very tight junctions in the blood vessel walls of the brain. These tight junctions are so restrictive that they prevent most small molecule drugs from reaching the brain, and 98% of all small molecule drugs are incapable of penetrating this barrier. These inventors have developed a novel strategy to use M13 bacteriophage as a courier to deliver therapeutics and imaging molecules across the BBB.

Advantages

• Improved M13 phage blood brain barrier penetration
• Specific targeting of M13 to GBM tumors
• Therapeutic or imaging molecule delivery through conjugation to M13-CTX phage
• Rod-like shape improves extravasation, immune cell escape, and internalization by cancer cells in comparison to traditional nanoparticles
• Sub-millimeter resolution imaging with near infrared imaging of SWNT-M13-CTX phage