Sticky Samples?

I recently had a customer write in regarding a challenging analytical size exclusion chromatography method. Their sample is a small protein that is bound to an antibody. Employing a traditional aSEC column, their sample does not want to elute, even when extra NaCl is added to the mobile phase! They were curious to learn if there are any aSEC columns on the market which are designed to prevent more challenging proteins from interacting with them.

The answer is, Yes, Sepax Zenix-C SEC columns!

Similar to how an Antibody Drug Conjugate?(ADC) would have a small molecule cytotoxic payload attached to the antibody, in this case, it is a small protein attached to the antibody. As we know with classical ADCs, they tend to become very hydrophobic and exhibit weird behaviors on classical aSEC.

A few years ago, Sepax Technologies, Inc. launched our "C" surface chemistry for customers who specifically were experiencing this issue. We worked directly with customers to develop an aSEC column that could be run under normal phosphate conditions without needing to include modifiers for these weird samples (you can certainly still add modifiers if you need to though!)

Tanabe Research Labs actually published a nice paper that beautifully displays the performance of the Sepax Zenix-C with their FynomAbs sample (Fynomers (7-kDa globular proteins) conj. with antibody (150kD)).

Analytical SEC HPLC of a bispecific antibody with Sepax Zenix-C SEC-300

A bispecific antibody that targets IL-6 receptor and IL-17A for the potential therapy of patients with autoimmune and inflammatory diseases

Company Name:?Tanabe Research Labs

Separation Mode:?SEC

Product:?LC Column, Sepax, Zenix-C SEC-300, 3 um, 300 A, 7.8 x 300 mm

Part Number:?233300-7830

Supelco/Sigma Aldrich Part Number:?Z777092

Mobile Phase:?PBS

Flow Rate:?1mL/min

Instrument:?HPLC, UV 280

Injection:?10ug

Top Chromatogram Sample:?MT-6194, a bispecific antibody targeting both IL-6R and IL-17A that was developed with the FynomAb technology

Bottom Chromatogram Sample:?tocilizumab

Abstract:

"Despite the success of current biological therapeutics for rheumatoid arthritis, these therapies, targeting individual cytokines or pathways, produce beneficial responses in only about half of patients. Therefore, better therapeutics are needed. IL-6 and IL-17A are proinflammatory cytokines in many autoimmune and inflammatory diseases, and several therapeutics have been developed to specifically inhibit them. However, targeting both of these cytokines with a bispecific therapeutic agent could account for their nonoverlapping proinflammatory functions and for the fact that IL-6 and IL-17A act in a positive feedback loop. Here, we present the development of MT-6194, a bispecific antibody targeting both IL-6R and IL-17A that was developed with the FynomAb technology. We also present data from mouse inflammatory disease experiments, indicating that simultaneous inhibition of both IL-6 and IL-17A yields enhanced efficacy compared with inhibition of each cytokine alone."

Full Title:?A bispecific antibody that targets IL-6 receptor and IL-17A for the potential therapy of patients with autoimmune and inflammatory diseases

Citation:?Lyman, Michael, et al. "A bispecific antibody that targets IL-6 receptor and IL-17A for the potential therapy of patients with autoimmune and inflammatory diseases." Journal of Biological Chemistry 293.24 (2018): 9326-9334.

Link to Paper:?https://doi.org/10.1074/jbc.M117.818559

Thank you to the authors and supporting scientists!

Robyn Richardson Currently Employed @ Neurocrine Biosciences

Richard Woods Currently Employed @ 恒瑞医药

Dragan Grabulovski

Michael Lyman Currently Employed @ Novasenta

Vincent Lieuw Currently Employed @ Tanabe Research Laboratories U.S.A, Inc.

Anjuli Timmer Currently Employed @ Aardvark Therapeutics

Christine Stewart Currently Employed Tanabe Research Laboratories U.S.A, Inc.

Steve W. Granger Currently Employed @ Salimetrics, LLC

Michela Silacci Melkko Currently Employed @ The Janssen Pharmaceutical Companies of Johnson & Johnson

Roland Newman ?Currently Retired