Synthesis of a Series of Folate-Terminated Dendrimer‑b‑PNIPAM Diblock Copolymers: Soft Nanoelements That Self-Assemble into Thermo- and pH-Responsive Spherical Nanocompounds

Dendrimers are large polymeric structures with nanosize dimensions and unique physicochemical properties. The major advantage of dendrimers compared with linear polymers is their spherical-shaped structure. During synthesis, the size and shape of the dendrimer can be customized and controlled, so the finished macromolecule will have a specific “architecture” and terminal groups. These characteristics will determine its suitability for drug delivery, diagnostic imaging, and as a genetic material carrier. Dendrimer-based multifunctional nanodevices desirable for a wide range of applications can be synthesized via multifunctionalization through site-specific modifications. In nanomedicine, for example, co-attaching dendrimers to oncolytic drugs, stimuli-responsive segments and target groups have enabled the creation of multifunctional nanocarriers. As one of the most significant classes of targeting agents, folates are commonly used in the active targeting of cancer cells. Moreover, folated-targeted therapy usually depend on overexpression of the folate receptor alpha protein (Frα) on the epithelial cancer cell surfaces.

Besides increasing the binding affinity with Frα, dendrimer-folate conjugates also improve anti-cancer activity by preventing drug resistance and internalization over free drugs. Generating dendritic polymer conjugate by combining dendrimers with polymers has proved to be an effective approach for increasing the complexity, size and functionality of dendritic nanocarriers. To this end, a number of dendritic polymer conjugates, such as hyperbranched and polymers, have been reported. Compared with dendrimers alone or other block copolymers, dendritic polymer conjugates exhibit unique structure-property correlations.

Poly(N-isopropylacrylamide) (PNIPAM) polymer is widely used in composing block copolymers owing to its remarkable stimuli-responsiveness property, biocompatibility and low critical solution temperature (LCST). However, at a temperature exceeding the transition temperature, PNIPAM chains collapse and become hydrophobic because of the dominating intramolecular hydrogen bonds. Additionally, an increase in the temperature or addition of additives that reduce LCST to about room temperature could induce microphase segregation of PNIPAM-containing block copolymers.

Although dendritic conjugates with PNIPAM have been extensively studied, only a few examples of dendrimer-PNIPAM conjugates have been reported. Moreover, a systematic investigation of the structure-activity relationship on various dendrimer-b-PNIPAM copolymers is lacking. Herein, Dr. Diego Bertuzzi, Dr. Carolyne Braga and Professor Catia Ornelas from University of Campinas – Unicamp in Brazil together with Dr. Melody Morris and Professor Bradley Olsen from Massachusetts Institute of Technology developed a new nanostructured folate-terminated dendrimer-b-PNIPAM diblock carrier as an innovative anti-cancer drug. The work is currently published in the journal, Macromolecules.

The research team used a divergent approach to synthesize the bifunctional dendrimers comprising one azide group and 5,15 and 44 folate termini. Through the grafting-to approach facilitated by the alkyne-azide cycloaddition between the PNIPAM and the dendrimers, three dendrimer-b-PNIPAM diblock copolymers containing folates (G1-11k, G2-30k, and G3-95k) were prepared. Furthermore, the authors fabricated poly(Nisopropylacrylamide) homopolymers via reversible addition-fragmentation chain-transfer polymerization and end functionalization with a cyclooctyne derivative. Finally, the dendrimers and associated intermediaries were characterized using different techniques.

The authors showed that high purity conjugates and all associated intermediaries were obtained with reasonable reaction yields. Investigation of the solution self-assembly revealed the resulting pH- and thermo-responsive core-shell spherical micelles exhibited significantly low polydispersities. The nanosized micelles within 70 – 160 nm size range displayed negative zeta potential. Additionally, within the nanoperiodic concept, the behavior of the obtained dendrimer-b-PNIPAM copolymers was akin to that of SI-type nanoelements that self-assemble into soft: soft spherical nanocompounds. Furthermore, the molecular weight of these copolymers and the hydrodynamic diameter of the micelles showed a linear relationship.

In summary, the synthesis and self-assembly of smart folate-terminated dendrimer-bPNIPAM diblock copolymers for targeting Frα-positive cancer cells were reported. Interestingly, while the folate terminal groups render the micelles pH-responsive, the PNIPAM block rendered them thermo-responsive. In a joint statement to Advances in Engineering, the authors explained the presented dendrimer-b-PNIPAM diblock copolymers containing folate ligands have extraordinary characteristics, making them promising candidates for anti-cancer targeted drug delivery.