Designing Chemical Strategies to Promote Therapeutic Access to Restricted Sites in CYTO

Jennifer L Rowe, Purdue University

Abstract

Therapeutically restricted sites present a formidable barrier in medicine. Herein, chemical strategies to overcome two restricted sites, HIV reservoirs and intracellular bacteria, will be discussed. First, cellular and anatomical HIV reservoirs, such as those in the brain, limit HIV eradication using currently known therapeutic regimes. HIV therapies are unable to localize in the brain, in part, due to high expression of efflux transporters, such as P-glycoprotein (P-gp), at the BBB, because many of these therapies are P-gp substrates. In an effort to overcome therapeutically restricted HIV sanctuaries, a dimerized combination HIV therapy was designed to act two-fold. First, the dimer acts as a P-gp inhibitor allowing therapeutic access to restricted sites. Second, the dimer acts as a prodrug, which once in the reducing environment of the cell, may release monomeric HIV therapies. The dual conjugate, Abacavir-S2-Darunavir, was shown to potently inhibit P-gp across two separate cell lines, was able to regenerate the component monomers in a reducing environment and contained modest anti-HIV activity. Further, mammalian cells create sanctuary sites for bacteria to grow and proliferate, because many common antibiotic therapies are unable to cross the mammalian cell membrane. Therefore, these pathogens are able to proliferate without therapeutic constraint. Here, a chemical strategy was developed to deliver a dual antibiotic therapy inside mammalian cells in an effort to clear these intracellular pathogens. First, a new synthetic strategy was developed for facile synthesis of dual conjugates, composed of an aminoglycoside and a cell penetrating peptide (CPP) linked with a reversible disulfide tether, using kanamycin and the known CPP Arg8 as a model system. Next, this synthetic methodology was expanded for use with the aminoglycoside tobramycin and the known broad-spectrum antibiotic and cell penetrating peptide, P14LRR, once again linked via the reversible disulfide tether (TobP14). Two distinct isomers of TobP14 were synthesized, isolated, and fully characterized by 2D NMR. The TobP14 isomers were shown to be an effective antibiotic across various Gram positive and negative pathogens such as MRSA, S. epidermidis, P. aeruginosa, and A. baumannii. Further, the isomers effectively released the monomeric therapies (tobramycin and P14-SH) in a reducing environment and were nontoxic to mammalian cells up to 16 µM. Finally, the dual conjugate isomers significantly reduce two different strains of intracellular A. baumannii within macrophages.

Degree

Ph.D.

Advisors

Chmielewski, Purdue University.

Subject Area

Energy|Design|Cellular biology|Oncology|Organic chemistry|Pharmaceutical sciences|Plastics|Polymer chemistry

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