Analysis of trafficking dynamics and cellular response in the interleukin-2 ligand/receptor system

Abstract

Interleukin-2 (IL-2) has been widely studied as a protein therapeutic due primarily to its central role in potentiating cell-based immunity. The effects of IL-2 on its cellular targets are mediated solely through interaction with one or more forms of the IL-2 receptor (IL- 2R). The overall goal of this thesis is to quantitatively characterize IL-2/IL-2R interactions toward the scientific end of advancing the understanding of the effects of molecular perturbations on cell function, and the technological end of providing insight into the design of improved IL-2 therapies. An interleukin-2 (IL-2) variant displays binding affinity to the heterotrimeric IL-2 receptor similar to that of wild-type (WT) IL-2, and was previously found to exhibit increased bioactivity in a T cell proliferation assay. Dedicated trafficking studies show that endocytic trafficking of this 20 I variant might be responsible for this increased potency, as a significantly increased fraction of internalized 201 is sorted to recycling instead of to lysosomal degradation. Denaturation experiments indicate that wild-type IL-2 and the 201 analog have similar stabilities at neutral pH; however, the analog displays decreased stability of the native and intermediate states under endosomal sorting conditions when compared to wild-type IL-2. This behavior correlates with differential pH-sensitivities of receptor binding affinity measured for IL-2 and the 201 variant. improved sorting leads to an extended half-life of intact 20 I, providing enhanced mitogenesis as compared to IL-2. We further conclude that this IL-2 variant minimizes undesired stimulation of NK cells. We describe a mathematical model that relates ligand-receptor trafficking properties to T cell proliferation in response to interleukin-2 (IL-2). The steady-state sorting behavior of the 201 variant is predictable from the model, as are criteria for designing IL-2 variants with further improvements in bioactivity. The concept of altering trafficking dynamics may offer a generalizable approach to generating improvements in the pharmacological efficacy of therapeutic cytokines.