skip to main content

Title: Poly(β‐amino ester)‐ co ‐poly(caprolactone) Terpolymers as Nonviral Vectors for mRNA Delivery In Vitro and In Vivo

The production of new proteins with messenger RNA (mRNA) has gained a broad interest due to its potential for addressing a wide range of diseases. Here, the design and characterization of novel ionizable poly(β‐amino ester)‐co‐poly(caprolactone) terpolymers, synthesized via the combination of the ring opening polymerization and the Michael step‐growth polymerization, are reported. The versatility of this method is demonstrated by varying the number of caprolactone units attached to each poly(β‐amino ester) (PBAE) terpolymer. The ability of the novel poly‐caprolactone (PCL)‐based PBAE materials to deliver mRNA is shown to depend on the physiochemical characteristics of the material, such as lipophilicity, as well as the formulation method used to complex the polymer with the oligonucleotide. This latter variable represents a previously unstudied aspect of PBAE library screens that can play an important role in identifying true top candidates for nucleic acid delivery. The most stable terpolymer is injected intravenously (IV) in mice and shows a transfection efficacy several times higher than the polyethylenimine (PEI) which is focused in the spleen, opening the possibility to use these biodegradable carriers in the intravenous delivery of antigen‐encoding mRNA for cancer immunotherapy and vaccination.

more » « less
Author(s) / Creator(s):
 ;  ;  ;  
Publisher / Repository:
Wiley Blackwell (John Wiley & Sons)
Date Published:
Journal Name:
Advanced Healthcare Materials
Medium: X
Sponsoring Org:
National Science Foundation
More Like this
  1. Abstract

    Bioreducible polymeric mRNA carriers are an emerging family of vectors for gene delivery and vaccine development. A few bioreducible systems have been generated through aqueous‐phase ring‐opening polymerization of lipoic acid derivatives, however this methodology limits hydrophobic group incorporation and functionality into resulting polymers. Herein, a poly(active ester)disulfide polymer is synthesized that can undergo facile aminolysis with amine‐containing substrates under stoichiometric control and mild reaction conditions to yield a library of multifunctional polydisulfide polymers. Functionalized polydisulfide polymer species form stable mRNA‐polymer nanoparticles for intracellular delivery of mRNAsin vitro. Alkyl‐functionalized polydisulfide‐RNA nanoparticles demonstrate rapid cellular uptake and excellent biodegradability when delivering EGFP and OVA mRNAs to cellsin vitro. This streamlined polydisulfide synthesis provides a new facile methodology for accessing multifunctional bioreducible polymers as biomaterials for RNA delivery and other applications.

    more » « less
  2. null (Ed.)
    The synthesis and degradation mechanisms of a class of pH-sensitive, rapidly degrading cationic poly(α-aminoester)s are described. These reactive, cationic polymers are stable at low pH in water, but undergo a fast and selective degradation at higher pH to liberate neutral diketopiperazines. Related materials incorporating oligo(α-amino ester)s have been shown to be effective gene delivery agents, as the charge-altering degradative behavior facilitates the delivery and release of mRNA and other nucleic acids in vitro and in vivo . Herein, we report detailed studies of the structural and environmental factors that lead to these rapid and selective degradation processes in aqueous buffers. At neutral pH, poly(α-aminoester)s derived from N -hydroxyethylglycine degrade selectively by a mechanism involving sequential 1,5- and 1,6-O→N acyl shifts to generate bis( N -hydroxyethyl) diketopiperazine. A family of structurally related cationic poly(aminoester)s was generated to study the structural influences on the degradation mechanism, product distribution, and pH dependence of the rate of degradation. The kinetics and mechanism of the pH-induced degradations were investigated by 1 H NMR, model reactions, and kinetic simulations. These results indicate that polyesters bearing α-ammonium groups and appropriately positioned N -hydroxyethyl substituents are readily cleaved (by intramolecular attack) or hydrolyzed, representing dynamic “dual function” materials that are initially polycationic and transform with changing environment to neutral products. 
    more » « less
  3. Abstract

    Ring‐opening polymerization (ROP) of lactones or cyclic (di)esters is a powerful method to produce well‐defined, high‐molecular‐weight (bio)degradable aliphatic polyesters. While the ROP of lactones of various ring sizes has been extensively studied, the ROP of the simplest eight‐membered lactone, 7‐heptanolactone (7‐HL), has not been reported using metal‐based catalysts. Accordingly, this contribution reports the ROP of 7‐HL via metal‐catalyzed coordinative‐insertion polymerization to the corresponding high‐molecular‐weight polyester, poly(7‐hydroxyheptanoate) (P7HHp). The resulting P7HHp is a semi‐crystalline material, with aTmof 68 °C, which is ~10 °C higher than poly(ε‐caprolactone) derived from the seven‐membered lactone. Mechanical testing showed that P7HHp is a hard and tough plastic, with elongation at break >670%. P7HHp‐based polyesters with higherTmvalues have been achieved through stereoselective copolymerization of 7‐HL with an eight‐membered cyclic diester, racemic dimethyl diolide (rac‐8DLMe), known to lead to highTmpoly(3‐hydroxyburtyrate) (P3HB). Notably, catalyst's strong kinetic preference for polymerizingrac‐8DLMeover 7‐HL in the 1/1 comonomer mixture rendered the formation of di‐block copolymer P3HB‐b‐P7HHp, showing two crystalline domains withTm1 ~ 65 °C andTm2 ~ 160 °C. Semi‐crystalline random copolymers withTmup to 164 °C have also been obtained by adjusting copolymerization conditions. Mechanical testing showed that P3HB‐b‐P7HHp can synergistically combine the high modulus of isotactic P3HB with the high ductility of P7HHp.

    more » « less
  4. Abstract

    Hydrolytically degradable poly(β‐thioether ester ketal) thermosets are synthesized via radical‐mediated thiol‐ene photopolymerization using three novel dialkene acyclic ketal monomers and a mercaptopropionate based tetrafunctional thiol. For all thermoset compositions investigated, degradation behavior is highly tunable based on the structure of the incorporated ketal and pH. Complete degradation of the thermosets is observed upon exposure to acidic and neutral pH, and under high humidity conditions. Polymer networks composed of cross‐link junctions based on acyclic dimethyl ketals degrade the quickest, whereas networks containing acyclic cyclohexyl ketals undergo hydrolytic degradation on a longer timescale. Thermomechanical analysis reveals low glass transition temperatures and moduli typical of thioether‐based thermosets.

    more » « less
  5. Abstract

    SARS‐CoV‐2 has led to a worldwide pandemic, catastrophically impacting public health and the global economy. Herein, a new class of lipid‐modified polymer poly (β‐amino esters) (L‐PBAEs) is developed via enzyme‐catalyzed esterification and further formulation of the L‐PBAEs with poly(d,l‐lactide‐coglycolide)‐b‐poly(ethylene glycol) (PLGA‐PEG) leads to self‐assembly into a “particle‐in‐particle” (PNP) nanostructure for gene delivery. Out of 24 PNP candidates, the top‐performing PNP/C12‐PBAE nanoparticles efficiently deliver both DNA and mRNA in vitro and in vivo, presenting enhanced transfection efficacy, sustained gene release behavior, and excellent stability for at least 12 months of storage at −20 °C after lyophilization without loss of transfection efficacy. Encapsulated with spike encoded plasmid DNA and mRNA, the lipid‐modified polymeric PNP COVID‐19 vaccines successfully elicit spike‐specific antibodies and Th1‐biased T cell immune responses in immunized mice even after 12 months of lyophilized storage at −20 °C. This newly developed lipid‐polymer hybrid PNP nanoparticle system demonstrates a new strategy for both plasmid DNA and mRNA delivery with the capability of long‐term lyophilized storage.

    more » « less