Doctor of Philosophy, Massachusetts Institute of Technology (2011)
Bachelor of Science, Johns Hopkins University (2006)
Chris Garcia, Postdoctoral Faculty Sponsor
Interleukin-2 (IL-2) regulates lymphocyte function by signaling through heterodimerization of the IL-2Rβ and γc receptor subunits. IL-2 is of considerable therapeutic interest, but harnessing its actions in a controllable manner remains a challenge. Previously, we have engineered an IL-2 "superkine" with enhanced affinity for IL-2Rβ. Here, we describe next-generation IL-2 variants that function as "receptor signaling clamps." They retained high affinity for IL-2Rβ, inhibiting binding of endogenous IL-2, but their interaction with γc was weakened, attenuating IL-2Rβ-γc heterodimerization. These IL-2 analogs acted as partial agonists and differentially affected lymphocytes poised at distinct activation thresholds. Moreover, one variant, H9-RETR, antagonized IL-2 and IL-15 better than blocking antibodies against IL-2Rα or IL-2Rβ. Furthermore, this mutein prolonged survival in a model of graft-versus-host disease and blocked spontaneous proliferation of smoldering adult T cell leukemia (ATL) T cells. This receptor-clamping approach might be a general mechanism-based strategy for engineering cytokine partial agonists for therapeutic immunomodulation.
View details for DOI 10.1016/j.immuni.2015.04.018
View details for Web of Science ID 000354827400010
Interleukin-2 (IL-2) is a pleiotropic cytokine that regulates immune cell homeostasis and has been used to treat a range of disorders including cancer and autoimmune disease. IL-2 signals via interleukin-2 receptor-β (IL-2Rβ):IL-2Rγ heterodimers on cells expressing high (regulatory T cells, Treg) or low (effector cells) amounts of IL-2Rα (CD25). When complexed with IL-2, certain anti-cytokine antibodies preferentially stimulate expansion of Treg (JES6-1) or effector (S4B6) cells, offering a strategy for targeted disease therapy. We found that JES6-1 sterically blocked the IL-2:IL-2Rβ and IL-2:IL-2Rγ interactions, but also allosterically lowered the IL-2:IL-2Rα affinity through a "triggered exchange" mechanism favoring IL-2Rα(hi) Treg cells, creating a positive feedback loop for IL-2Rα(hi) cell activation. Conversely, S4B6 sterically blocked the IL-2:IL-2Rα interaction, while also conformationally stabilizing the IL-2:IL-2Rβ interaction, thus stimulating all IL-2-responsive immune cells, particularly IL-2Rβ(hi) effector cells. These insights provide a molecular blueprint for engineering selectively potentiating therapeutic antibodies.
View details for DOI 10.1016/j.immuni.2015.04.015
View details for Web of Science ID 000354827400009
Cytokines exert a vast array of immunoregulatory actions critical to human biology and disease. However, the desired immunotherapeutic effects of native cytokines are often mitigated by toxicity or lack of efficacy, either of which results from cytokine receptor pleiotropy and/or undesired activation of off-target cells. As our understanding of the structural principles of cytokine-receptor interactions has advanced, mechanism-based manipulation of cytokine signaling through protein engineering has become an increasingly feasible and powerful approach. Modified cytokines, both agonists and antagonists, have been engineered with narrowed target cell specificities, and they have also yielded important mechanistic insights into cytokine biology and signaling. Here we review the theory and practice of cytokine engineering and rationalize the mechanisms of several engineered cytokines in the context of structure. We discuss specific examples of how structure-based cytokine engineering has opened new opportunities for cytokines as drugs, with a focus on the immunotherapeutic cytokines interferon, interleukin-2, and interleukin-4.
View details for DOI 10.1146/annurev-immunol-032713-120211
View details for PubMedID 25493332
Cytokines play crucial roles in regulating immune homeostasis. Two important characteristics of most cytokines are pleiotropy, defined as the ability of one cytokine to exhibit diverse functionalities, and redundancy, defined as the ability of multiple cytokines to exert overlapping activities. Identifying the determinants for unique cellular responses to cytokines in the face of shared receptor usage, pleiotropy, and redundancy will be essential in order to harness the potential of cytokines as therapeutics. Here, we discuss the biophysical (ligand-receptor geometry and affinity) and cellular (receptor trafficking and intracellular abundance of signaling molecules) parameters that contribute to the specificity of cytokine bioactivities. Whereas the role of extracellular ternary complex geometry in cytokine-induced signaling is still not completely elucidated, cytokine-receptor affinity is known to impact signaling through modulation of the stability and kinetics of ternary complex formation. Receptor trafficking also plays an important and likely underappreciated role in the diversification of cytokine bioactivities but it has been challenging to experimentally probe trafficking effects. We also review recent efforts to quantify levels of intracellular signaling components, as second messenger abundance can affect cytokine-induced bioactivities both quantitatively and qualitatively. We conclude by discussing the application of protein engineering to develop therapeutically relevant cytokines with reduced pleiotropy and redirected biological functionalities.
View details for DOI 10.1016/B978-0-12-800100-4.00001-5
View details for Web of Science ID 000331017400001
View details for PubMedID 24388212