Development of an attenuated interleukin-2 fusion protein that can be activated by tumour-expressed proteases

Immunology. 2011 Jun;133(2):206-20. doi: 10.1111/j.1365-2567.2011.03428.x. Epub 2011 Mar 23.


The ability to alter the cytokine microenvironment has the potential to shape immune responses in many physiological settings, including the immunotherapy of tumours. We set out to develop a general approach in which cytokines could be functionally attenuated until activated. We report the development and initial characterization of fusion proteins in which human or mouse interleukin-2 (IL-2), a potent growth factor for immune cells, is joined to a specific IL-2 inhibitory binding component separated by a protease site. The rationale is that upon cleavage by a protease the cytokine is free to dissociate from the inhibitory component and becomes biologically more available. We describe the successful development of two attenuation strategies using specific binding: the first uses the mouse IL-2 receptor alpha chain as the inhibitory binding component whereas the second employs a human antibody fragment (scFv) reactive with human IL-2. We demonstrated that the fusion proteins containing a prostate-specific antigen or a matrix metalloproteinase (MMP) protease cleavage site are markedly attenuated in the intact fusion protein but had enhanced bioactivity of IL-2 in vitro when cleaved. Further, we showed that a fusion protein composed of the IL-2/IL-2 receptor alpha chain with an MMP cleavage site reduced tumour growth in vivo in a peritoneal mouse tumour model. This general strategy should be applicable to other proteases and immune modulators allowing site-specific activation of immunomodulators while reducing unwanted side-effects.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Animals
  • Antineoplastic Agents / pharmacology
  • Cell Proliferation / drug effects
  • Disease Models, Animal
  • Enzyme-Linked Immunosorbent Assay
  • Humans
  • Immunoblotting
  • Interleukin-2 / genetics*
  • Interleukin-2 / metabolism*
  • Interleukin-2 Receptor alpha Subunit / genetics
  • Interleukin-2 Receptor alpha Subunit / metabolism
  • Male
  • Mice
  • Mice, Inbred C57BL
  • Mice, Transgenic
  • Neoplasms / physiopathology
  • Prostate-Specific Antigen / metabolism
  • Prostate-Specific Antigen / pharmacology
  • Recombinant Proteins / biosynthesis
  • Recombinant Proteins / metabolism*
  • Recombinant Proteins / pharmacology


  • Antineoplastic Agents
  • Interleukin-2
  • Interleukin-2 Receptor alpha Subunit
  • Recombinant Proteins
  • Prostate-Specific Antigen