Analytical techniques and bioactivity assays to compare the structure and function of filgrastim (granulocyte-colony stimulating factor) therapeutics from different manufacturers

The FDA has approved more than 100 protein and peptide drugs with hundreds more in the pipeline (Lanthier et al. in Nat Rev Drug Discov 7(9):733–737, 2008). Many of these originator biologic products are now coming off patent and are being manufactured by alternate methods than the innovator as follow-on drugs. Because changes to the production method often lead to subtle differences (e.g., degradation products, different posttranslational modifications or impurities) in the therapeutic (Schiestl et al. in Nat Biotechnol 29(4):310–312, 2011), there is a critical need to define techniques to test and insure the quality of these drugs. In addition, the emergence of protein therapeutics manufactured by unapproved methodologies presents an ongoing and growing regulatory challenge. In this work, high-resolution mass spectrometry was used to determine the presence or absence of posttranslational modifications for one FDA-approved and three foreign-sourced, unapproved filgrastim products. Circular dichroism (CD) was used to compare the secondary structure and probe the temperature stability of these products. Native 2D ¹H,¹⁵N-heteronuclear singular quantum coherence (HSQC) NMR test was applied to these samples to compare the higher-order structure of the four products. Finally, a cell proliferation assay was performed on the filgrastims to compare their bioactivity, and stressed filgrastim was tested in the bioassay to better understand the effects of changes in protein structure on activity. The results showed that orthogonal approaches are capable of characterizing the physiochemical properties of this protein drug and assessing the impact of structural changes on filgrastim purity and potency.     

Why is Firefly Oxyluciferin a Notoriously Labile Substance?

The chemistry of firefly bioluminescence is important for numerous applications in biochemistry and analytical chemistry. The emitter of this bioluminescent system, firefly oxyluciferin, is difficult to handle. The cause of its lability was clarified while its synthesis was reinvestigated. A side product was identified and characterized by NMR spectroscopy and X‐ray crystallography. The reason for the lability of oxyluciferin is now ascribed to autodimerization of the coexisting enol and keto forms in a Mannich‐type reaction.     

Statistical disordering of chalcogen atoms in the cluster fragments of K4[Re6(µ3-S)8-y (µ3-Te) y ,(CN)6]

An X-ray diffraction analysis was performed for three crystals K₄[Re₆(Μ_3-S)_8-y(Μ_3-Te)_y(CN)₆], which are members of a continuous series of solid solutions obtained by high-temperature reactions using different substrates. The crystals are cubic, space group Ia3. Statistical disordering of chalcogen atoms in the [Re₆(Μ_3-S)_8-y(Μ₃-Te)_y(CN)₆] cluster fragments is examined using the data for the three crystals. The results of this analysis show that the extent of the statistical disordering of Te and S affects the crystal structure dimensions.     

Small-angle approximation for an ion beam in an external electromagnetic field

A small-angle approximation method is formulated for a curvilinear ion beam propagating in an electromagnetic field. The effects of particle velocity scatter and multiple elastic scattering on the beam path in a magnetic field, the electric field of a cylindrical capacitor, and mutually orthogonal electric and magnetic fields are considered. An analytical model for beam power compression is developed.     

Method for measuring integrated sensitivity of solar cells and multielement photoconverters using an X-Y scanner

We describe a method for automated measurement of the integrated sensitivity of solar cells (SCs) and multielement photoconverters (MPCs) using an experimental apparatus including a Pentium III personal computer (PC), an HP-34401A digital multimeter (DM), a stabilized radiation source (SRS), a controllable focusing system, an X-Y positioning device based on CD-RW optical disk storage devices. The method provides high accuracy in measuring the size of photosensitive areas of the solar cells and multielement photoconverters and inhomogeneities in their active regions, which makes it possible to correct the production process in the development stage and during fabrication of test prototypes for the solar cells and multielement photoconverters. The radiation power from the stabilized radiation source was ≤1 W; the ranges of the scanning steps along the X, Y coordinates were 10–100 μm, the range of the transverse cross sectional diameters of the focused radiation beam was 10–100 μm, the measurable photocurrents were 10⁻⁹ A to 2 A; scanning rate along the X, Y coordinates, ≤100 mm/sec; relative mean-square error (RMSE) for measurement of the integrated sensitivity of the solar cells, 0.2 ≤ γS _int ≤ 0.9% in the ranges of measurable photocurrents 1 mA ≤ I_ph ≤ 750 mA and areas 0.1 ≤ A ≤ 25 cm² for number of measurements equal to ≤ 2· 10⁵; instability of the radiation power (luminosity) ≤ 0.08% for 1 h or ≤ 0.4% for 8 h continuous operation; stabilized power range for the stabilized radiation source, 10⁻²–10² W. The software was written in Delphi 7.0. __________ Translated from Zhurnal Prikladnoi Spektroskopii, Vol. 73, No. 5, pp. 670–675, September–October, 2006.