Benjamin M. Wohl

Macromolecular prodrugs of ribavirin combat side effects and toxicity with no loss of activity of the drug

Chemi-enzymatic synthesis of ribavirin acrylate and subsequent RAFT co-polymerization with acrylic acid afforded a formulation of a broad spectrum antiviral drug which avoids accumulation in erythrocytes, the origin of the main side effect of ribavirin. In cultured macrophages the macromolecular prodrugs exhibited decreased toxicity while maintaining the anti-inflammatory action of ribavirin.

Highly active macromolecular prodrugs inhibit expression of the hepatitis C virus genome in the host cells.

Efficacious, potent, and at the same time nontoxic macromolecular prodrugs of ribavirin are designed taking advantage over prodrug activation by the intracellular milieu. Activity of these prodrugs is illustrated in the cells hosting hepatitis C virus replication and also in the cells implicated in the inflammatory response to the viral infection.

Macromolecular Prodrugs for Controlled Delivery of Ribavirin

Ribavirin (RBV)-containing polymers are synthesized based on poly(N-vinylpyrrolidone) and poly(acrylic acid), two polymers with extensive characterization in biomedicine. The copolymers are shown to exhibit a minor to negligible degree of association with erythrocytes, thus effectively eliminating the origin of the main side effects of RBV. The therapeutic benefit of macromolecular RBV prodrugs is illustrated by matched efficacy in suppressing production of nitric oxide by stimulated cultured macrophages as compared to pristine RBV with no associated cytotoxicity, which is in stark contrast to an RBV-based treatment which results in a significant decrease in cell viability. These results contribute to the development of antiviral polymer therapeutics and delivery of RBV in particular.

Macromolecular Prodrugs of Ribavirin: Concerted Efforts of the Carrier and the Drug

Polymers in tune. Automated parallel polymer synthesis is developed to obtain libraries of macromolecular prodrugs of ribavirin, a broad-spectrum antiviral agent. As many as 10 identified lead polymer conjugates exhibit therapeutic efficacy matching that of the pristine drug and at the same time suppressed the origin of the main side effect of ribavirin.

Narrow Therapeutic Window of Ribavirin as an Inhibitor of Nitric Oxide Synthesis is Broadened by Macromolecular Prodrugs

Ribavirin (RBV), a broad-spectrum antiviral agent, is a standard medication against hepatitis C virus (HCV). However, despite the decades of clinical success, the mechanism of action of this drug against HCV remains a subject of debate. Furthermore, the appeal of this therapeutic agent is considerably lessened by unfavorable pharmacokinetics. This interdisciplinary study contributes to the understanding of intracellular effects exerted by RBV and presents a successful design of macromolecular prodrugs of RBV to achieve a safer treatment. Specifically, we demonstrate that RBV exhibits a pronounced anti-inflammatory activity in cultured macrophages as is evidenced by a 2-fold decrease in the levels of produced nitric oxide achieved using a clinically relevant concentration of this drug. However, this effect was characterized by a rather narrow therapeutic window with experimental values of EC50 and IC50 being 7 and 19 μM, respectively. Macromolecular prodrugs were obtained using an acrylate derivative of RBV, RAFT polymerization technique, and N-vinyl pyrrolidone as a partner monomer. The synthesized polymers were characterized with uniform molecular weights, relatively narrow polydispersities, and gradually increasing content of RBV. The resulting polymer therapeutics were effective in delivering their payload to the cultured macrophages and afforded a significantly wider therapeutic window, as much as >1000 μM (18-fold in relative values). Taken together, this work contributes significantly to the development of safer methods for delivery of RBV, as well as understanding the mechanism of action and origins of the side effects of this broad-spectrum antiviral agent.

Macromolecular (pro)drugs in antiviral research

Macromolecular (pro)drugs are a sub-discipline of medicinal and polymer chemistries aiming to optimize the delivery of drugs to their site of action. In recent decades, this field of science has undergone a tremendous development, with the soundest achievements registered in the delivery of anticancer drugs. Surprisingly, the development of these tools for applications in antiviral treatment lags significantly behind – despite the fact that the first in vivo successes of polymers in fighting viruses were reported half a century ago. Furthermore, the unique scope and utility of polymers in antiviral research is that macromolecules themselves exhibit highly potent activity against diverse viruses. Herein, in an attempt to revive the research interest in this field, we aim to provide an overview of successes (and failures) of polymers as antiviral agents and macromolecular prodrugs. Specifically, we discuss inhibition of the entry of the virus into mammalian cells by polymers, give an overview of the synthetic schemes applied for the conjugation of drugs to carrier polymers, and also present guidance with regard to potential reporter systems which can be used for the characterization of novel drug delivery systems in virus-free cell cultures.

Macromolecular (pro)drugs with concurrent direct activity against the hepatitis C virus and inflammation.

Macromolecular prodrugs (MPs) are a powerful tool to alleviate side-effects and improve the efficacy of the broad-spectrum antiviral agent ribavirin. In this work, we sought an understanding of what makes an optimal formulation within the macromolecular parameter space--nature of the polymer carrier, average molar mass, drug loading, or a good combination thereof. A panel of MPs based on biocompatible synthetic vinylic and (meth)acrylic polymers was tested in an anti-inflammatory assay with relevance to alleviating inflammation in the liver during hepatitis C infection. Pristine polymer carriers proved to have a pronounced anti-inflammatory activity, a notion which may prove significant in developing MPs for antiviral and anticancer treatments. With conjugated ribavirin, MPs revealed enhanced activity but also higher toxicity. Therapeutic windows and therapeutic indices were determined and discussed to reveal the most potent formulation and those with optimized safety. Polymers were also tested as inhibitors of replication of the hepatitis C viral RNA using a subgenomic viral replicon system. For the first time, negatively charged polymers are revealed to have an intracellular activity against hepatitis C virus replication. Concerted activity of the polymer and ribavirin afforded MPs which significantly increased the therapeutic index of ribavirin-based treatment. Taken together, the systematic investigation of the macromolecular space identified lead candidates with high efficacy and concurrent direct activity against the hepatitis C virus and inflammation.

Polyanionic Macromolecular Prodrugs of Ribavirin: Antiviral Agents with a Broad Spectrum of Activity.

Macromolecular prodrugs are developed as multiarmed agents against diverse viral pathogens. Lead candidates inhibit infectivity and replication of HIV, Ebola, influenza, measles, RSV, etc-thus being broad-spectrum antiviral agents.

Poly(vinyl alcohol) Physical Hydrogel Nanoparticles, Not Polymer Solutions, Exert Inhibition of Nitric Oxide Synthesis in Cultured Macrophages

Hydrogel nanoparticles (HNP) are an emerging tool of biomedicine with unique materials characteristics, scope, and utility. These hydrated, soft colloidal carriers can penetrate through voids with dimensions narrower than the size of the particle, provide stabilization for fragile biological cargo and allow diffusion and exchange of solutes with external phase. However, techniques to assemble HNP are few; solitary examples exist of biocompatible polymers being formulated into HNP; and knowledge on the biomedical properties of HNP remains rather cursory. In this work, we investigate assembly of HNP based on a polymer with decades of prominence in the biomedical field, poly(vinyl alcohol), PVA. We develop a novel method for production of PVA HNP through nanoprecipitation-based assembly of polymer nanoparticles and subsequent physical hydrogelation of the polymer. Polymer nanoparticles and HNP were visualized using scanning electron microscopy and fluorescence imaging, and characterized using dynamic light scattering and zeta potential measurements. Interaction of PVA HNP with mammalian cells was investigated using flow cytometry, viability screening, and measurements of nitric oxide production by cultured macrophages. The latter analyses revealed that PVA administered as a polymer solution or in the form of HNP resulted in no measurable increase in production of the inflammation marker. Unexpectedly, PVA HNP exerted a pronounced inhibition of NO synthesis by stimulated macrophages, that is, had an anti-inflammatory activity. This effect was accomplished with a negligible change in the cell viability and was not observed when PVA was administered as a polymer solution. To the best of our knowledge, this is the first observation of inhibition of NO synthesis in macrophages by administered nanoparticles and specifically hydrogel nanoparticles. Taken together, our results present PVA HNP as promising colloidal hydrogel nanocarriers for biomedical applications, specifically drug delivery and assembly of intracellular biosensors.