Enhancing pharmaceutical formulations to improve efficacy and delivery of drug molecules

Thesis (Ph. D. in Biological Chemistry)--Massachusetts Institute of Technology, Dept. of Chemistry, 2013.Cataloged from PDF version of thesis.Includes bibliographical references.Major impediments to the full utility of current and potential drugs include issues of resistance and delivery. To address these challenges, in this thesis two directions of research were pursued: (1) the use of multivalent polymeric inhibitors to overcome drug resistance in human and avian influenza and (2) low-viscosity, high-concentration protein suspensions for therapeutic antibody, in particular monoclonal antibody (MAb), delivery. (1) Influenza resistance to small molecule neuraminidase (NA) inhibitors is spreading. Little emphasis, however, has been placed on alternative formulations of inhibitors. We investigated the design of multivalent antivirals, wherein small molecule ligands of viral proteins are conjugated via a linker to a linear polymeric backbone. Unexpectedly, we found that a poly-L-glutamine bearing pendant zanamivir (ZA) groups is at least as potent as those containing both ZA and sialic acid (SA). By examining the structure-activity relationship of such monofunctional conjugates, we show that the most potent one has 10% ZA attached to a neutral, high molecular weight backbone through a short alkyl linker. Importantly, we also demonstrate that such a polymer conjugate entirely compensates for weakened binding in and has 2,000-fold enhanced anti-viral potency against, ZA-resistant strains. We further evaluated this optimized inhibitor in vivo and observed that it is an effective therapeutic of established infection in ferrets and reduces viral titers up to 190-fold when used as a combined prophylactic/therapeutic in mice. Additionally, we see no evidence that the conjugate stimulates an immune response in mice upon repeat administration. (2) Typically, high doses of MAb therapeutics are required for clinical effect. Ideally, these MAbs would be delivered by subcutaneous injection of a small liquid volume. Such highly concentrated MAb solutions, however, are far more viscous than the 50 centipose (cP) permitted by the FDA. We evaluated approaches to reduce formulation viscosity by forming protein suspensions. Aqueous suspensions induced by poly(ethylene glycol), precipitating salts, or ethanol actually increased viscosity. However, non-aqueous suspensions of amorphous antibody powders in organic solvents that have s 1 hydrogen atom available for hydrogen-bonding, exhibited up to a 38-fold decrease in viscosity.by Alisha K. Weight.Ph.D.in Biological Chemistr

Zanamivir Conjugated to Poly-L-Glutamine is Much More Active Against Influenza Viruses in Mice and Ferrets Than the Drug Itself

Purpose: Previously, polymer-attached zanamivir had been found to inhibit influenza A viruses in vitro far better than did small-molecule zanamivir (1) itself. The aim of this study was to identify in vitro—using the plaque reduction assay—a highly potent 1-polymer conjugate, and subsequently test its antiviral efficacy in vivo. Methods: By examining the structure-activity relationship of 1-polymer conjugates in the plaque assay, we have determined that the most potent inhibitor against several representative influenza virus strains has a neutral high-molecular-weight backbone and a short alkyl linker. We have examined this optimal polymeric inhibitor for efficacy and immunogenicity in the mouse and ferret models of infection. Results: 1 attached to poly-L-glutamine is an effective therapeutic for established influenza infection in ferrets, reducing viral titers up to 30-fold for 6 days. There is also up to a 190-fold reduction in viral load when the drug is used as a combined prophylactic/therapeutic in mice. Additionally, we see no evidence that the drug conjugate stimulates an immune response in mice upon repeat administration. Conclusions: 1 attached to a neutral high-molecular-weight backbone through a short alkyl linker drastically reduced both in vitro and in vivo titers compared to those observed with 1 itself. Thus, further development of this polymeric zanamivir for the mitigation of influenza infection seems warranted.National Institutes of Health (U.S.) (Grant U01-AI074443