Contributed Commentary By Kunhua Chen
March 13, 2018 | Monoclonal antibodies, the modern version of the “magic bullets” that Paul Ehrlich first envisioned 100 years ago, have become invaluable components of diagnostic development and drug discovery. Take, for example, the anti-CEA monoclonal antibody: when labeled with a short-lived radioactive 99mTc, it can be used for cancer diagnostics (arcitumomab); when labeled with the toxin SN-38, it can be used for cancer treatment (labetuzumab-SN-38).
Applications like these are advancing more rapidly than ever, and the industry needs to keep pace. But the methods we have relied on for the past century to develop and produce antibodies — hybridomas, phage, and yeast displays — have been plagued by low affinity, low specificity, lack of repeatability, and long production times.
We need to ensure that the next generation of magic monoclonal antibody bullets can hit their targets without these problems. They must be delivered quickly, and at a price that does not drain the precious resources of researchers and clinical developers. Two recent advances in antibody development, the use of plasma cells and adoption of rabbits, promise to give scientists better results from their antibody development efforts.
Plasma Cells
One promising approach to improved development of monoclonal antibodies involves the use of single plasma cells. Eliminating low-quality antibodies from the start with single-cell technology reduces the guesswork associated with antibody effectiveness.
These terminally differentiated plasma cells contain fully matured antibody clones. Therefore, they are far more likely to generate high-affinity, high-specificity antibodies than methods that rely on selecting from their precursor cells. Plasma cells represent a small fraction of the standard B lymphocytes; by starting with them instead of the broader universe of B cells, scientists can dramatically reduce the amount of time required for downstream screening. In addition, plasma cells have much more antibody mRNA than B cells, making them richer sources of material for these important development pipelines.
The use of plasma cells addresses many of limitations associated with conventional antibody development techniques, such as hybridoma and phage display technologies. These methods make it very difficult to identify the best antibodies from a vast number of low-affinity or non-specific clones.
In addition, plasma cell-sourced antibody production improves on earlier single-cell techniques that use individual B cells or memory cells loaded onto microtiter plates. The former, which depends on cells sticking to an antigen-coated plastic plate, is prone to the inclusion of immature B cells with abundant cell surface IgG or IgM as well as the loss of mature plasma cells due to the growth bias during incubation. The latter relies on a library of memory cell-surface markers that are not available in common organisms used for antibody production.
A final benefit of working with single plasma cells is turnaround time. Unlike traditional processes for monoclonal antibody development, which typically take as long as four months to complete, a plasma cell workflow can take as little as two weeks. That involves plasma cell isolation, cloning, expression, screening, and selection.
Rabbit Model
Though most antibody development centers on the traditional mouse model, efforts to improve results have begun exploring different animal models. As it turns out, rabbits are especially useful. They are well known for mounting a strong immune response against foreign antigens, leading to useful polyclonal antibodies. Indeed, rabbit-derived polyclonal antibodies were among the first-established and most widely-used research tools in immunology. Rabbit antibodies also have a higher affinity and specificity to many antigens than rodent antibodies, making them strong choices for diagnostic development.
The rabbit immune system has unique properties that result in the recognition of more antigen epitopes. Even subtle changes in epitopes, such as post-translational modifications, are recognized. Rabbits are also superior in their ability to develop antibodies against small moieties.
In an immunoassay, higher affinity means that you can use less antibody without a corresponding loss of signal strength. This ensures the highest sensitivity and specificity.
A Question of Reliability
Diagnostic tests are not reliable if different batches of the “same” antibody give unpredictable results. Indeed, reproducibility has been a serious problem with commercial antibodies.
A 2008 study connected to the Human Protein Atlas (DOI 10.1074/mcp.R800013-MCP200) found that fewer than half of 20,000 commercially available antibodies exclusively recognized their specified targets, and a 2015 story in Nature estimated that the problem costs the U.S. biomedical research community $350 million per year and leads to a sea of inaccurate biological findings. More than 100 scientists banded together in response to the report and called for drastic changes in the production of antibodies to address this challenge.
I believe that it is now possible to overcome these problems by rapidly generating a more diverse array of antibodies directly from affinity matured plasma cells and using rabbit as our model. The associated savings of time and money will also facilitate important advances within the diagnostics industry.
Kunhua Chen is CEO of ExonBio, a recombinant antibody service company, providing rabbit monoclonal antibody development with Single Plasma Cell Interrogation (SPIN) Technology. Prior he was a research fellow at the Salk Institute for Biological Studies. He received his PhD in cellular and molecular biology from Rutgers University. Kunhua can be reached at kchen@exonbio.com.