n-azidoacetylmannosamine and Breast-Neoplasms

Metabolic sugar labeling followed by the use of reagent-free click chemistry is an established technique for in vitro cell targeting. However, selective metabolic labeling of the target tissues in vivo remains a challenge to overcome, which has prohibited the use of this technique for targeted in vivo applications. Herein, we report the use of targeted ultrasound pulses to induce the release of tetraacetyl N-azidoacetylmannosamine (Ac4 ManAz) from microbubbles (MBs) and its metabolic expression in the cancer area. Ac4 ManAz-loaded MBs showed great stability under physiological conditions, but rapidly collapsed in the presence of tumor-localized ultrasound pulses. The released Ac4 ManAz from MBs was able to label 4T1 tumor cells with azido groups and significantly improved the tumor accumulation of dibenzocyclooctyne (DBCO)-Cy5 by subsequent click chemistry. We demonstrated for the first time that Ac4 ManAz-loaded MBs coupled with the use of targeted ultrasound could be a simple but powerful tool for in vivo cancer-selective labeling and targeted cancer therapies.

Topics: Animals; Azides; Breast; Breast Neoplasms; Carbocyanines; Cell Line, Tumor; Click Chemistry; Drug Delivery Systems; Female; Fluorescent Dyes; Hexosamines; Mice, Inbred BALB C; Microbubbles; Optical Imaging; Ultrasonography, Mammary

Metabolic oligosaccharide engineering (MOE) is a maturing technology capable of modifying cell surface sugars in living cells and animals through the biosynthetic installation of non-natural monosaccharides into the glycocalyx. A particularly robust area of investigation involves the incorporation of azide functional groups onto the cell surface, which can then be further derivatized using "click chemistry." While considerable effort has gone into optimizing the reagents used for the azide ligation reactions, less optimization of the monosaccharide analogs used in the preceding metabolic incorporation steps has been done. This study fills this void by reporting novel butanoylated ManNAc analogs that are used by cells with greater efficiency and less cytotoxicity than the current "gold standard," which are peracetylated compounds such as Ac₄ ManNAz. In particular, tributanoylated, N-acetyl, N-azido, and N-levulinoyl ManNAc analogs with the high flux 1,3,4-O-hydroxyl pattern of butanoylation were compared with their counterparts having the pro-apoptotic 3,4,6-O-butanoylation pattern. The results reveal that the ketone-bearing N-levulinoyl analog 3,4,6-O-Bu₃ ManNLev is highly apoptotic, and thus is a promising anti-cancer drug candidate. By contrast, the azide-bearing analog 1,3,4-O-Bu₃ ManNAz effectively labeled cellular sialoglycans at concentrations ∼3- to 5-fold lower (e.g., at 12.5-25 µM) than Ac₄ ManNAz (50-150 µM) and exhibited no indications of apoptosis even at concentrations up to 400 µM. In summary, this work extends emerging structure activity relationships that predict the effects of short chain fatty acid modified monosaccharides on mammalian cells and also provides a tangible advance in efforts to make MOE a practical technology for the medical and biotechnology communities.

Topics: Acylation; Adenocarcinoma; Animals; Antineoplastic Agents; Apoptosis; Azides; Breast Neoplasms; Butyric Acid; Cell Cycle; Cell Line, Tumor; CHO Cells; Click Chemistry; Cricetinae; Cricetulus; Drug Design; Glycocalyx; Glycoconjugates; Hexosamines; Humans; Jurkat Cells; Ketones; Molecular Structure; N-Acetylneuraminic Acid; Pancreatic Neoplasms; Structure-Activity Relationship