Lewis acid-promoted cycloaddition reactions of aziridines and cyclopropanes

I. Lewis Acid-Promoted Carbon-Carbon Bond Cleavage of Aziridines: Divergent Cycloaddition Pathways of the Derived Ylides The formation of azomethine ylides from aziridines under Lewis acidic conditions and the productive reactivity of these species are described. Efficient carbon-carbon bond cleavage of aziridines through treatment with stoichiometric or catalytic amounts of Zn(II) salts has been shown to occur under relatively mild conditions. Depending on the constitution of the dipolarophile, the resulting azomethine ylides undergo cycloadditions with electron-rich olefins in either a [3+2] or [4+2] manner producing pyrrolidine or tetrahydroquinoline derivatives. R CO2R' N CO2R' ZnX2 R N O OR' R'O O Ar ZnXn N Ar R CO2R' CO2R' [3+2] [4+2] H2C EDG Y N R Y CO2R' CO2R' Z Z n n EDG R = Aryl or H X = Cl or OTf II. Heterocycle Synthesis Via [3+2] Reactions of Cyclopropanes A review of the literature concerning the use of cyclopropanes in the one-step synthesis of five-membered heterocycles is presented. Methodology for the convenient, stereoselective preparation of substituted ?-lactones, ?-lactams, tetrahydrofurans, pyrrolidines, and various other heterocycles from simple cyclopropane precursors is described. Lewis acid-promotion/catalysis, as well as thermal methods for the ring cleavage of cyclopropanes are discussed. Where possible, proposed models are given for the stereochemical outcome of these annulation reactions. III. Enantiospecific and Diastereoselective Tin-Catalyzed Cycloaddition Reactions of Aldehydes and Donor-Acceptor Cyclopropanes: The Synthesis of 2,5- Disubstituted Tetrahydrofurans The diastereoselective synthesis of a variety of cis-2,5-disubstituted tetrahydrofurans via Sn(II)- or Sn(IV)-catalyzed cycloaddition reactions of donor-acceptor (D-A) cyclopropanes with aldehydes is described. In some cases, the use of enantioenriched cyclopropane has been shown to afford optically active tetrahydrofuran derivatives with excellent stereochemical transfer. Evidence is presented supporting an SN2 reaction in which the aldehyde participates as a nucleophilic species. R CO2Me CO2Me R'CHO Sn(OTf)2 or SnCl4 (cat.) R O R' CO2Me MeO2C + R = Aryl, heteroaryl, alkenyl, alkyl R' = Aryl, heteroaryl, alkenyl, alkynyl, alkyl IV. Application of the Tin-Catalyzed Cyclopropane/Aldehyde Cycloaddition Reaction: Progress Towards the 2-Oxabicyclo[4.3.0]nonane Core of the Cladiellin Diterpenes Efforts towards the construction of the hydroisobenzofuran core common to the cladiellin diterpenes are discussed. The use of a model system in the catalytic cyclopropane/aldehyde cycloaddition manifold has been explored. The stereochemical results of this successful reaction are discussed in the context of possible extension to a more sophisticated system. Synthetic efforts herein have disclosed a potential straightforward route for the preparation electron-poor alkenylidene cyclopropanes via an unprecedented 3- exo-dig cyclization pathway

Analytical and data strategy for continuous downstream manufacturing

As advances emerge in developing continuous biomanufacturing processes, there is an increased need to deploy PAT tools to characterize, monitor, and control key quality attributes and a criticality to have a data infrastructure to support the immense amount of information being generated. While the desire for these tools exists in traditional batch processing, in a continuous operation, these become a requirement to ensure consistent product quality and enable proactive approaches in maintaining performance. The ultimate goal is to deploy PAT tools to reliably provide real-time information on product and process impurities throughout the entire operation. However, in its current state, there is a reliance on a mixture of inline, at-line, and offline technologies. By identifying the time criticality of CQAs, efforts can be focused on where to prioritize real-time measurements or instead, quicker or more automated testing for a subset of analytics. This work describes the application of this approach in the development of small-scale, compact in-line UV instruments to measure real-time protein concentration and in the integration of an automated sampling system with at-line and offline instrumentation for in-process impurity characterization. Introduction of these PAT tools add to the complexity of the data infrastructure as it introduces requirements for platforms capable of supporting spectral data, chemometric model deployment, spectral instrument management, and time-alignment of discrete data. With the vast amount of information produced in a continuous environment, interface and analysis tools need to be developed so that any end-user can digest data into a format that easily allows them to gain insight into an ongoing batch. This work will highlight the data architecture of the continuous platform, with a focus on software tools selected for aggregation and real-time data visualization. The capabilities of these software packages were demonstrated through a proof-of-concept study using single-pass tangential flow filtration (SPTFF) as a model unit operation, which allowed integration of continuous, spectral, and discrete data. These tools allowed scientists to go from viewing real-time data across multiple, equipment-specific software to one consolidated interface, which in turn reduced time spent in compiling data for analysis and reporting. In addition, advanced capabilities of deploying model predictive control in SPTFF were demonstrated to show the application of a closed loop process control in continuous manufacturing

Nanofabricated particles for engineered drug therapies: A preliminary biodistribution study of PRINT™ nanoparticles

A novel method for the fabrication of polymeric particles on the order of tens of nanometers to several microns is described. This imprint lithographic technique called PRINT™ (Particle Replication In Non-wetting Templates), takes advantage of the unique properties of elastomeric molds comprised of a low surface energy perfluoropolyether network, allowing the production of monodisperse, shape-specific nanoparticles from an extensive array of organic precursors. This engineered nature of particle production has a number of advantages over the construction of traditional nanoparticles such as liposomes, dendrimers, and colloidal precipitates. The gentle “top down” approach of PRINT enables the simultaneous and independent control over particle size and shape, composition, and surface functionality, and permits the loading of delicate cargos such as small organic therapeutics and biological macromolecules. Thus, this single tool serves as a comprehensive platform for the rational design and investigation of new nanocarriers in medicine, having applications ranging from therapeutics to advanced diagnostics. Preliminary in vitro and in vivo studies were conducted, demonstrating the future utility of PRINT particles as delivery vectors in nanomedicine. Monodisperse 200 nm poly(ethylene glycol)-based (PEG) particles were fabricated using PRINT methodology and characterized via scanning electron microscopy and dynamic light scattering. Incubation with HeLa cells showed very little cytotoxicity, even at high concentrations. The biodistribution and pharmacokinetics of [125I]-labeled particles were studied in healthy mice following bolus tail vein administration. The particles were distributed mainly to the liver and the spleen with an apparent distribution t1/2 of approximately 17 min followed by slow redistribution with a t1/2 of 3.3 h. The volume of distribution for the central and peripheral compartments was found to be approximately 3 mL and 5 mL, respectively

Rendering Protein-Based Particles Transiently Insoluble for Therapeutic Applications

Herein we report the fabrication of protein (bovine serum albumin, BSA) particles which were rendered transiently insoluble using a novel, reductively labile disulfide-based cross-linker. After being cross-linked, the protein particles retain their integrity in aqueous solution and dissolve preferentially under a reducing environment. Our data demonstrates that cleavage of the cross-linker leaves no chemical residue on the reactive amino group. Delivery of a self-replicating RNA was achieved via the transiently insoluble PRINT protein particles. These protein particles can provide new opportunities for drug and gene delivery

Reductively Responsive siRNA-Conjugated Hydrogel Nanoparticles for Gene Silencing

A critical need still remains for effective delivery of RNA interference (RNAi) therapeutics to target tissues and cells. Self-assembled lipid- and polymer-based systems have been most extensively explored for transfection with small interfering RNA (siRNA) in liver and cancer therapies. Safety and compatibility of materials implemented in delivery systems must be ensured to maximize therapeutic indices. Hydrogel nanoparticles of defined dimensions and compositions, prepared via a particle molding process that is a unique off-shoot of soft lithography known as PRINT (Particle Replication in Non-wetting Templates), were explored in these studies as delivery vectors. Initially, siRNA was encapsulated in particles through electrostatic association and physical entrapment. Dose-dependent gene silencing was elicited by PEGylated hydrogels at low siRNA doses without cytotoxicity. To prevent disassociation of cargo from particles after systemic administration or during post-fabrication processing for surface functionalization, a polymerizable siRNA pro-drug conjugate with a degradable, disulfide linkage was prepared. Triggered release of siRNA from the prodrug hydrogels was observed under a reducing environment while cargo retention and integrity were maintained under physiological conditions. Gene silencing efficiency and cytocompatibility were optimized by screening the amine content of the particles. When appropriate control siRNA cargos were loaded into hydrogels, gene knockdown was only encountered for hydrogels containing releasable, target-specific siRNAs, accompanied by minimal cell death. Further investigation into shape, size, and surface decoration of siRNA-conjugated hydrogels should enable efficacious targeted in vivo RNAi therapies

The Pursuit of a Scalable Nanofabrication Platform for Use in Material and Life Science Applications

In this Account, we describe the use of perfluoropolyether (PFPE)-based materials that are able to accurately mold and replicate micro- and nanosized features using traditional techniques such as embossing as well as new techniques that we developed to exploit the exceptional surface characteristics of fluorinated substrates. Because of the unique partial wetting and nonwetting characteristics of PFPEs, we were able to go beyond the usual molding and imprint lithography approaches and have created a technique called PRINT (Particle [or Pattern] Replication In Nonwetting Templates)

Reductively Responsive siRNA-Conjugated Hydrogel Nanoparticles for Gene Silencing

A critical need still remains for effective delivery of RNA interference (RNAi) therapeutics to target tissues and cells. Self-assembled lipid- and polymer-based systems have been most extensively explored for transfection with small interfering RNA (siRNA) in liver and cancer therapies. Safety and compatibility of materials implemented in delivery systems must be ensured to maximize therapeutic indices. Hydrogel nanoparticles of defined dimensions and compositions, prepared via a particle molding process that is a unique off-shoot of soft lithography known as particle replication in nonwetting templates (PRINT), were explored in these studies as delivery vectors. Initially, siRNA was encapsulated in particles through electrostatic association and physical entrapment. Dose-dependent gene silencing was elicited by PEGylated hydrogels at low siRNA doses without cytotoxicity. To prevent disassociation of cargo from particles after systemic administration or during postfabrication processing for surface functionalization, a polymerizable siRNA pro-drug conjugate with a degradable, disulfide linkage was prepared. Triggered release of siRNA from the pro-drug hydrogels was observed under a reducing environment while cargo retention and integrity were maintained under physiological conditions. Gene silencing efficiency and cytocompatibility were optimized by screening the amine content of the particles. When appropriate control siRNA cargos were loaded into hydrogels, gene knockdown was only encountered for hydrogels containing releasable, target-specific siRNAs, accompanied by minimal cell death. Further investigation into shape, size, and surface decoration of siRNA-conjugated hydrogels should enable efficacious targeted in vivo RNAi therapies