Facile Fabrication and Instant Application of Miniaturized Antibody-Decorated Affinity Columns for Higher-Order Structure and Functional Characterization of TRIM21 Epitope Peptides

2013 ◽  
Vol 85 (21) ◽  
pp. 10479-10487 ◽  
Author(s):  
M. Al-Majdoub ◽  
K. F. M. Opuni ◽  
C. Koy ◽  
M. O. Glocker
Keyword(s):  
Functional Characterization ◽  
Higher Order ◽  
Order Structure ◽  
Facile Fabrication

Physicochemical, functional, and pharmacologic comparability between the proposed biosimilar rituximab GP2013 and originator rituximab as the foundation for biosimilarity.

2013 ◽  
Vol 31 (15_suppl) ◽  
pp. 3075-3075 ◽  
Author(s):  
Antonio da Silva ◽  
Ulrich Kronthaler ◽  
Ines Meyer ◽  
Cornelius Fritsch ◽  
Timo Schneiderer ◽  
...  
Keyword(s):  
Amino Acid ◽  
B Cell ◽  
Functional Characterization ◽  
Higher Order ◽  
Order Structure ◽  
Pharmacological Characterization ◽  
Originator Product ◽  
Acceptance Range ◽  
Size Heterogeneity

3075 Background: Development of a biosimilar involves extensive characterization of the originator product and a target-directed iterative development process ensuring comparability to the originator with similar clinical efficacy, safety and quality. Here we report the physicochemical, functional and pre-clinical pharmacological characterization of a proposed rituximab biosimilar (GP2013). Methods: A variety of physicochemical methods were used to analyze primary and higher order structure, post-translational modifications and size heterogeneity. Functional characterization included a series of bioassays (in vitro target binding, ADCC, CDC and apoptosis) and SPR-based Fc receptor binding assays. Comparative PK and PD were assessed in cynomolgus monkeys, the pharmacologically most relevant species. Results: GP2013 has the same primary amino acid sequence and higher order structure as the originator rituximab and both were comparable with regard to charge variants, specific amino acid modifications, glycan pattern and size heterogeneity (low- and high-molecular weight variants & particles). Functionally GP2013 could not be distinguished from originator rituximab preclinically. In primates, PK analysis confirmed bioequivalence between GP2013 and originator rituximab with nearly identical AUC values and 90% CIs entirely within the standard acceptance range of 0.8-1.25. Bioequivalence of PD response was also shown, with 95% CIs of areas under the effect-time curves (AUEC) ratios for relative change from baseline in B-cell populations within the 0.8-1.25 acceptance range. Conclusions: Using a broad panel of analytical methods it was shown that GP2013 is highly similar to originator rituximab at the physicochemical level. In addition, the preclinical comparability exercise confirmed that GP2013 and originator rituximab are pharmacologically similar with regard to FcR and CD20 binding, ADCC, CDC and apoptosis potency, PK exposure and B-cell depletion. As such, we anticipate that the ongoing clinical trials will help provide confirmatory evidence of similar efficacy and safety to the originator product.

scholarly journals Chemometric outlier classification of 2D-NMR spectra to enable higher order structure characterization of protein therapeutics

2020 ◽  
Vol 199 ◽  
pp. 103973 ◽  
Author(s):  
David A. Sheen ◽  
Vincent K. Shen ◽  
Robert G. Brinson ◽  
Luke W. Arbogast ◽  
John P. Marino ◽  
...  
Keyword(s):  
Nmr Spectra ◽  
2D Nmr ◽  
Higher Order ◽  
Protein Therapeutics ◽  
Structure Characterization ◽  
Order Structure ◽  
2D Nmr Spectra

Characterization of Protein Higher Order Structure Using Vibrational Circular Dichroism Spectroscopy

2013 ◽  
Vol 14 (2) ◽  
pp. 199-208 ◽  
Author(s):  
Radhika P. Nagarkar ◽  
Brian M. Murphy ◽  
Xiaotong Yu ◽  
Mark Cornell Manning ◽  
Wasfi A. Al-Azzam
Keyword(s):  
Circular Dichroism ◽  
Higher Order ◽  
Circular Dichroism Spectroscopy ◽  
Vibrational Circular Dichroism ◽  
Order Structure ◽  
Circular Dichroïsm

Physicochemical and functional characterization of trastuzumab-dkst, a trastuzumab biosimilar

2021 ◽  
Author(s):  
Parag Goyal ◽  
Jyoti Iyer ◽  
Laxmi Adhikary ◽  
Bhavesh Vats ◽  
Pradeep Kabadi ◽  
...  
Keyword(s):  
Functional Characterization ◽  
Binding Activity ◽  
Order Structure ◽  
Repeat Dose ◽  
Antibody Dependent Cellular Cytotoxicity ◽  
Charge Heterogeneity ◽  
Tertiary Structures ◽  
Cynomolgus Monkeys ◽  
Comparative Similarity

Aims: Preclinical comparative similarity studies of trastuzumab-dkst, a Herceptin® biosimilar, are reported. Materials & methods: Primary sequence and higher-order structure and pharmacological mechanisms of action were compared using multiple techniques. Pharmacokinetics and repeat-dose toxicity were assessed in cynomolgus monkeys. Results: Primary structures were identical; secondary and tertiary structures were highly similar. Non-significant differences were observed for charge heterogeneity. Twelve of 13 glycan species were highly similar, with slightly higher total mannose levels in trastuzumab-dkst. FcγR and FcRn binding activity was highly similar. Each drug equally inhibited HER2+ cell proliferation, demonstrating equivalent relative potency in mediating HER2+ cell cytolysis by antibody-dependent cellular cytotoxicity. Pharmacokinetic and toxicological profiles in cynomolgus monkeys were similar. Conclusion: Trastuzumab-dkst, US-licensed trastuzumab and EU-approved trastuzumab demonstrate high structural and functional similarity.

Characterization of nucleic acid higher order structure by gas-phase H/D exchange in a quadrupole-FT-ICR mass spectrometer

Biopolymers ◽  
2009 ◽  
Vol 91 (4) ◽  
pp. 256-264 ◽  
Author(s):  
Jingjie Mo ◽  
Gabrielle C. Todd ◽  
Kristina Håkansson
Keyword(s):  
Nucleic Acid ◽  
Gas Phase ◽  
Mass Spectrometer ◽  
Higher Order ◽  
Order Structure ◽  
Ft Icr

scholarly journals CENP-N promotes the compaction of centromeric chromatin

2021 ◽  
Author(s):  
Keda Zhou ◽  
Magdalena Gebala ◽  
Dustin C Woods ◽  
Kousik Sundararajan ◽  
Garrett Edwards ◽  
...  
Keyword(s):  
Chromatin Structure ◽  
Higher Order ◽  
Histone Variant ◽  
Order Structure ◽  
Interacting Protein ◽  
Biophysical Characterization ◽  
Centromeric Chromatin

The histone variant CENP-A is the epigenetic determinant for the centromere, where it is interspersed with canonical H3 to form a specialized chromatin structure that nucleates the kinetochore. The arrangement of nucleosomes at the centromere into higher order structure is unknown. Here we demonstrate that the CENP-A interacting protein CENP-N promotes the stacking of CENP-A containing mono-nucleosomes and nucleosomal arrays through a previously undefined interaction between the α6 helix of CENP-N with the DNA of a neighboring nucleosome. We describe the cryoEM structures and biophysical characterization of such CENP-N mediated nucleosome stacks and nucleosomal arrays and demonstrate that this interaction is responsible for the formation of densely packed chromatin at the centromere in the cell. Our results provide first evidence that CENP-A, together with CENP-N, promotes specific chromatin higher order structure at the centromere.

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