Antidote tested for

biologic therapy side effects

Published September 01 2017


Schematic of the antidote mechanism: The role of FcRn recycling for circulating antibodies (top): Proteins in circulation are taken up by fluid phase pinocytosis by endothelial cells, where the majority of antibodies bind FcRn in the mildly acidic early endosome. Then proteins bound to FcRn are trafficked to a recycling endosome and returned to the cell surface, where they dissociate and return to circulation at the near-neutral pH of the serum. After antidote (bottom): The antidote blocks FcRn binding, and the therapeutic antibody follows the default fluid phase endocytic path to the lysosome for degradation.
Figures courtesy of alyse portnoff

About 120 years ago, Paul Ehrlich proposed a revolutionary idea, the side-chain theory, to explain immune response in cells. We now know the process he described is the binding of antigens to receptors on the surface of immune cells. Ehrlich compared this process to a key fitting in a lock and had the foresight to coin the term “magic bullet” in reference to the precision and lethality of immune cells reacting to specific targets. Fast forward to 1975, when George Köhler and Cesar Milstein developed a method to produce monoclonal antibodies and custom-made “magic bullets” were born as therapeutic antibodies.

Adapting monoclonal antibodies to be used as therapeutic agents involved solving problems with large-scale production and immunogenicity, fine-tuning their specificity and harnessing their molecular and cellular mechanisms as therapeutic tools. Now, monoclonal antibodies are an important part of the physicians’ arsenal to treat cancer, organ transplant rejection, and autoimmune and infectious diseases.

However, new issues have emerged, such as the inherent risks of treating a patient with therapeutic antibodies that remain in circulation for extended periods, exercising long-lasting effects. Alyse Portnoff, a postdoctoral fellow in the Antibody Discovery and Protein Engineering group at MedImmune, explained that “because every patient responds to a medicine differently,” she and her colleagues needed to create “safety mechanism(s) for their (therapeutic) proteins, such that they could be removed from a patient in the event of an adverse reaction — much like a safety net for patients undergoing treatment.” Portnoff and her colleagues provide proof of principle for this concept in a Journal of Biological Chemistry paper.

 
Antidotes that block FcRn binding profoundly impact antibody pharmacokinetics in huFcRn transgenic mice: Antibody serum concentration curves for the therapeutic antibody containing a nonnatural amino acid (purple) and for the antibody after antidote conjugation (orange) in huFcRn transgenic mice.

For this study, the researchers took advantage of the mechanism responsible for the naturally long half-lives of antibodies in circulation, on average about 21 days. This mechanism involves the pH-dependent interaction of the crystallizable fragment of an IgG antibody, Fc region, with the neonatal Fc receptor FcRn. The binding of an antibody to FcRn allows the former to escape lysosomal degradation, return to circulation and continue its function.

Borrowing the tRNA for the amber codon, UAG, from the anaerobic archeabacter Methanosarcina mazei and its pyrrolsyl-tRNA synthetase, the researchers designed and produced variants of a therapeutic antibody. These variants were modified with a nonnatural amino acid, which was strategically positioned in different regions of the antibody–FcRn interface. Binding of these mutant antibodies to FcRn and in vivo half-life and clearance rate appeared roughly unaltered despite having the nnAA substitution. However, addition of a small-molecule antidote covalently modified the nnAA by click chemistry, negatively affecting antibody affinity for FcRn and significantly increasing in vivo clearance in a mouse model.

The authors acknowledge the need to improve in vivo click chemistry reaction efficiency and antidote size and structure, but they point out that this preliminary study represents an important first step toward developing safety switches for therapeutic antibodies, potentially broadening the population of patients that could benefit from these novel therapies. A good example would be patients suffering from autoimmune diseases such as rheumatoid arthritis, lupus or psoriasis, Portnoff said. These are chronic diseases often treated with immunosuppressive therapies involving antibody therapeutic treatments for the rest of the patients’ lives, she said. “While a patient may respond well to his or her medicine, immunosuppressive therapies can sometimes prevent the body from healing unrelated infections,” she said. “Our research into turning off binding to the FcRn receptor may create a way to temporarily suppress treatment, enabling the patient to fight the infection and then return to his or her regular course of immunosuppressive treatment.”

 

Mariana Figuera-Losada Mariana Figuera-Losada is a research consultant at Montefiore Medical Center.