Enzyme Prodrug Therapy with Photocrosslinkable Anti-EGFR Affibodies Conjugated to Upconverting Nanoparticles

• Published in: ACS nano Vol. 16; no. 10; pp. 15873 - 15883
• Main Authors: Roy, Shambojit, Curry, Shane D., Bagot, Conrad Corbella, Mueller, Evan N., Mansouri, Abdulrahman M., Park, Wounjhang, Cha, Jennifer N., Goodwin, Andrew P.
• Format: Journal Article
• Language: English
• Published: 21-09-2022
• ISSN: 1936-0851; 1936-086X
• DOI: 10.1021/acsnano.2c02558

In this work, we demonstrate that a photocrosslinkable conjugate of upconverting nanoparticles and cytosine deaminase can catalyze prodrug conversion specifically at tumor sites
in vivo
. Noncovalent association of proteins and peptides with cellular surfaces leads to receptor-mediated endocytosis and catabolic degradation. Recently, we showed that covalent attachment of proteins such as affibodies to cell receptors yields extended expression on cell surfaces with preservation of protein function. To adapt this technology for
in vivo
, conjugates were prepared of upconverting nanoparticles and fusion proteins of affibody and cytosine deaminase enzyme (UC-ACD). The affibody allows covalent photocrosslinking to Epidermal Growth Factor Receptors (EGFR) overexpressed on Caco-2 human colorectal cancer cells under near infrared (NIR) light. Once bound, the cytosine deaminase portion of the fusion protein converts the prodrug 5-fluorocytosine (5-FC) to the anticancer drug 5-Flurouracil (5-FU). NIR covalent photoconjugation of UC-ACD to Caco-2 cells showed four-fold higher retention than cells that were not irradiated
in vitro
. Next, athymic mice expressing Caco-2 tumors showed five-fold greater UC-ACD accumulation in the tumors than either conjugates without the CD enzyme or UC-ACDs in the absence of NIR excitation. With oral administration of 5-FC prodrug, tumors with photoconjugated UC-ACD yielded two-fold slower tumor growth than control groups, and median survival increased from 28 days to 35 days. These experiments demonstrate that enzyme-decorated nanoparticles can remain viable after a single covalent photoconjugation
in vivo
, which can in turn localize prodrug conversion to tumor sites for multiple weeks.