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691 MDK-271: A dual function molecule consisting of empirically-designed peptidyl agonists of IL-2/15Rβγc and IL-7Rαγc, unrelated to IL-2, IL-15, or IL-7, incorporated into a bispecific Fc fusion protein
  1. William Dower,
  2. Steven Cwirla,
  3. Alice Bakker,
  4. Praechompoo Pongtornpipat,
  5. Blake Williams,
  6. Prarthana Joshi,
  7. Sandra Wang,
  8. Michael Needels and
  9. Ronald Barrett
  1. Medikine, Menlo Park, CA, USA


Background Activation of IL-2/15Rβγc or IL-7R on immune cells using modified versions of IL-2 or IL-7 is under investigation as monotherapy, or in combination with checkpoint inhibitors, engineered T or NK cells, or neo-antigen vaccines. We previously described small synthetic peptides, unrelated to IL-2, IL-15, or IL-7, that selectively activate either IL-2/15Rβγc or IL-7R. IL-2/15Rβγc and IL-7R activation exhibit complementary effects on immune cells that when combined may offer benefits over each independent mechanism. We now report the creation of an Fc-fusion protein that incorporates both IL-2/15Rβγc and IL-7R agonist peptides, and characterize its properties in cell lines and human (PBMC) lymphocyte subpopulations.

Methods Peptide agonists of IL-2/15Rβγc (MDK1169) and IL-7R (MDK1319) were separately fused to each chain of obligate heterodimeric (asymmetric) Fc molecules. The Fc-fusion was purified by protein-A and size exclusion chromatography, and characterized by LC-MS. Receptor-mediated signaling, proliferation, and cell-surface receptor expression in cell lines, PBMCs, mixed and isolated lymphocyte subpopulations were determined by flow cytometry and ELISA to evaluate effects of IL-2, IL-2v, IL-7, MDK-202 or MDK-701 (Fc-fusions of MDK1169 and MDK1319, respectively), and combinations (mixtures) of these molecules, in comparison with the dual agonist MDK-271.

Results LC-MS analysis indicates MDK-271 is a heterodimeric molecule containing one copy each of MDK1169 (IL-2/15Rβγc-biased agonist) and MDK1319 (IL-7R agonist) fused to individual Fc-chains. Cell-based assay of MDK-271 demonstrates potent, fully efficacious phosphorylation of STAT5 in TF-1 cells expressing Rγc, and engineered to express either IL-2/15Rβ or IL-7Rα. PBMCs exposed ex vivo to MDK-271 exhibit additive, complementary, and synergistic effects among various lymphocyte subpopulations: CD4+Tn, Teff, Treg, Tmem; C8+Tn, Teff, Tmem; and NK cells, in this analysis. The mono-specific agonists MDK-202 and MDK-701 produce proliferative effects and signaling patterns in responsive cell lines and lymphocyte subsets similar to those induced by IL-2v (an IL-2/15Rβγc-biased mutant of IL-2) and IL-7, respectively. Combining both activities in MDK-271 induces response profiles that differ in some T-cell subsets from those of mono-specific agonists of the two receptors. Animal studies designed to understand the effects of these differences are underway.

Conclusions IL-2/15Rβγc and IL-7R are both currently undergoing extensive scrutiny as potential immuno-oncology therapeutic targets. The biology of these cytokines is both overlapping and complementary in stimulating and supporting T-cell populations; and some recent evidence suggests possible superiority of the combination. Based on in vitro properties, the Fc-peptide fusion reported here, exhibiting both IL-2/15Rβγc-biased agonist and Il-7Rαγc agonist activities, could be valuable in anti-tumor therapeutic applications.

Ethics Approval The use of human PBMC in this study was authorized under Minimal Risk Research Related Activities at Stanford Blood Center (SQL 79075)

This is an open access article distributed in accordance with the Creative Commons Attribution 4.0 Unported (CC BY 4.0) license, which permits others to copy, redistribute, remix, transform and build upon this work for any purpose, provided the original work is properly cited, a link to the licence is given, and indication of whether changes were made. See:

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