Photoinducible Oncometabolite Detection.

Dysregulated metabolism can fuel cancer by altering the production of bioenergetic building blocks and directly stimulating oncogenic gene-expression programs. However, relatively few optical methods for the direct study of metabolites in cells exist. To address this need and facilitate new approaches to cancer treatment and diagnosis, herein we report an optimized chemical approach to detect the oncometabolite fumarate. Our strategy employs diaryl tetrazoles as cell-permeable photoinducible precursors to nitrileimines. Uncaging these species in cells and cell extracts enables them to undergo 1,3-dipolar cycloadditions with endogenous dipolarophile metabolites such as fumarate to form pyrazoline cycloadducts that can be readily detected by their intrinsic fluorescence. The ability to photolytically uncage diaryl tetrazoles provides greatly improved sensitivity relative to previous methods, and enables the facile detection of dysregulated fumarate metabolism through biochemical activity assays, intracellular imaging, and flow cytometry. Our studies showcase an intersection of bioorthogonal chemistry and metabolite reactivity that can be applied for biological profiling, imaging, and diagnostics.

Photo-responsive gel droplet as a nano- or pico-litre container comprising a supramolecular hydrogel.

Photo-responsive gel droplets having nano- or pico-L volume that showed photo-induced gel-sol transition were successfully developed, for which the inter-droplet mass transport and the subsequent enzymatic reactions in the interior of the gel droplets were photo-triggered.

Photopolymerization and shrinkage kinetics of in situ crosslinkable N-vinyl-pyrrolidone/poly(epsilon-caprolactone fumarate) networks.

Biodegradable, injectable and in situ photocrosslinkable macromers based on fumaric acid and polycaprolactone (PCLF) were prepared and characterized by FTIR, 1HNMR, and 13CNMR spectroscopy. The multifunctional macromers dissolved in N-vinyl pyrollidone (NVP) were photopolymerized by visible light irradiation in the presence of camphorquinone as photoinitiator. The photocrosslinking reaction was monitored by measuring shrinkage strain and shrinkage strain rate. The degree of photopolymerization reaction i.e. degree of conversion (DC%) was traced using FTIR spectroscopy. A three level factorial design was developed to study the effects of initiator concentration, NVP concentration, and molecular weight of PCLF upon photocrosslinking characteristics including degree of conversion and shrinkage strain. Results revealed that although neat PCLF was photopolymerized, but it was putty like after 220 seconds of irradiation and showed a very low degree of conversion (29%). Adding about 20% NVP caused a dramatic increase in its degree of conversion (63.33%). Increasing NVP up to 50% resulted in a decrease in DC% because of lower reactivity of NVP and leaving more unreacted NVP monomers. Sol fraction studies supported these results indicating that at higher NVP concentration, most of NVP and PCLF have not undergone the crosslinking reaction, leading to 55% decrease in DC%. Shrinkage strain measurement also confirmed the FTIR results.

Effect of macromer molecular weight on in vitro ophthalmic drug release from photo-crosslinked matrices.

The objective of this study was to investigate the effect of poly(propylene fumarate) (PPF) molecular weight on the release kinetics of two ophthalmic model drugs, acetazolamide (AZ) and timolol maleate (TM), from matrices prepared by photo-induced copolymerization of PPF with N-vinyl pyrrolidone (NVP). PPF macromers of different number average molecular weight (M(n)) and polydispersity index (PI) were used in the experiments. Photo-crosslinked matrices were loaded with 5wt.% AZ or TM. The amounts of released drug and NVP were determined using high-performance liquid chromatography (HPLC). The release kinetics of both drugs was influenced by the molecular weight of the constituent PPF macromer. An increased M(n) led to an increased burst release and an accelerated drug release. Dependent on the PPF M(n), the initial AZ loading was released within periods ranging from 35 days (M(n) = 3670, PI = 1.9) to 220 days (M(n) = 2050, PI=1.5). TM-loaded matrices revealed similar release kinetics dependent on the PPF M(n). The amount of released NVP from photo-crosslinked matrices during the course of a release experiment was independent of the PPF M(n) for both drugs. Matrix swelling and erosion were determined gravimetrically. The network structures of non-loaded matrices were further characterized by determining their crosslinking densities and the relative double bond conversions of fumaric acid (FAA) and NVP. Independent of PPF M(n), PPF and NVP similarly participated in the formation of the PPF/polyNVP copolymer network. The observed differences in drug release might therefore be explained by differences in the microstructural organization of the copolymer networks. Overall, the results demonstrate that drug release kinetics from photo-crosslinked PPF/polyNVP matrices is strongly dependent on the M(n) of the PPF macromer.

Poly(propylene fumarate) bone tissue engineering scaffold fabrication using stereolithography: effects of resin formulations and laser parameters.

Stereolithography using photo-cross-linkable polymeric biomaterials is an effective technique for fabricating highly complex three-dimensional (3D) scaffolds with controlled microstructures for tissue engineering applications. In this study, we have optimized the UV curable polymer solution composition and laser parameters for the stereolithography machine. Poly(propylene fumarate) (PPF) was used as the biomaterial, diethyl fumarate (DEF) was used as the solvent, and bisacrylphosphrine oxide (BAPO) was used as the photoinitiator. Three different weight ratios of PPF/DEF and BAPO contents were characterized by measuring the viscosities and thermal properties of the un-cross-linked solutions and the mechanical properties of the formed scaffolds. After optimizing the resin composition by satisfying both the viscosity limitation and the mechanical requirement, laser parameters such as critical exposure (Ec) and penetration depth (Dp) were determined from the working curve and the relationship between laser speed and energy by measuring the thickness of predesigned windows fabricated in stereolithography with different ranges of Ec and Dp. Three-dimensional scaffolds with various pore sizes, pore shapes, and porosities were designed in computer-aided design (CAD) software and were fabricated in stereolithography. The fabricated scaffolds were characterized by measuring external dimensions, porosities, mean pore sizes, and compressive moduli and were compared to the CAD models. Feature accuracy in the xy-plane was achieved and overcuring of the resin in z-axis was minimized. The stereolithographically fabricated scaffolds with controlled microstructures can be useful in diverse tissue engineering applications.