Therapeutic proteins exhibit highly specific functions, few side effects, and low immune responses, which are typically difficult to achieve with small-molecule drugs. Hence, protein-based drugs continue to dominate the market, as shown from the list of top-selling drugs from 2019, where 4 out of 5 were protein therapeutics. Despite its distinctive advantages, the innate structural instability of proteins has been a drawback, which has led to studies that involve the conjugation of synthetic polymers to enhance the half-life of the drugs in the bloodstream. Key factors to consider in the conjugation step are site-specificity, yield, and ease of purification as they directly affect protein activity, product homogeneity, and upscale-applicability. The most commonly used polymer-conjugation method, direct chemical modification, lacks site-specificity that often leads to reduced protein activity, and the bulk of the polymer often results in low conjugation yields and difficulties in purification. Therefore, new strategies are in demand.<br/>In this work, protein prenylation and in situ controlled radical polymerization are exploited to overcome the limitations in conventional protein-polymer conjugation techniques. A tumor-targeting antibody mimetic DAPRin (Designed Ankyrin Repeat Protein) is site-specifically functionalized with an ATRP (Atom Transfer Radical Polymerization) initiator moiety via protein prenylation. In situ polymerization is then employed from the DARPin-macroinitiator with a variety of customized-monomers to yield DARPin-polymer constructs capable of tumor-targeted imaging and drug delivery. We demonstrate that utilization of the described methods results in DARPin-polymer constructs with high therapeutic efficacies due to the highly retained protein activity, enhanced pharmacokinetics, and high drug-to-antibody ratios (DAR).

Site-specific DARPin-polymer conjugates functionalized via in situ polymerization for tumor-targeted imaging and drug delivery

ACS Spring Meeting 2021

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