Stanford and Zurich researchers have engineered a variant of IL-2 that shows a better antitumor response in mice than wild-type IL-2.1 The new IL-2 superkine variant, dubbed super-2, has been licensed by Teva Pharmaceutical Industries Ltd., which is planning additional preclinical studies of the compound.

IL-2 is an immunostimulatory cytokine that binds and activates a wide range of immune cells. Novartis AG markets Proleukin aldesleukin IL-2 to treat metastatic melanoma and renal cancer.

In patients with metastatic melanoma or metastatic renal cell carcinoma, high-dose IL-2 delivery, either alone or in combination with tumor vaccines, has led to therapeutic responses in about 13%-20% of cases and to long-term survival in about 10% of cases.2 However, components of IL-2's biology have limited its use.

One of the three parts of the cytokine's receptor complex-IL-2 receptor α-chain (IL2-RA; CD25)-is often expressed at very low levels or not at all on key immune cells such as cytotoxic T cells and NK cells. Those cells are rather insensitive to IL-2 (see "IL-2 regulation of T cells"). As a result, an IL-2-based immunotherapy has to be given at high doses in order to achieve significant stimulation of cytotoxic T cells, which play a central role in the host antitumor and antiviral response.

However, delivering high levels of systemic IL-2 can trigger vascular leak syndrome, leading to pulmonary edema, liver cell damage and renal failure.2 This process is thought to be CD25 dependent.2

To address those limitations, researchers from the Stanford University, Stanford University, University Hospital Zurich and the University of Zurich set out to eliminate the functional requirement of IL-2 for CD25. The group started by using in vitro evolution to produce IL-2 mutant libraries and screened for IL-2 variants-superkines-that bound the receptor complex and activated IL-2 signaling independently of CD25, thus generating a T cell response even when CD25 was absent.

In vitro, the top three variants bound IL-2 receptor b-chain (IL2-RB; CD122), another component of the receptor complex, much more strongly than wild-type IL-2. Structural studies showed the superkines' increased affinity for CD122 resulted from mutations in the core of the protein.

Next, the researchers set out to confirm that the increased affinity for CD122 was sufficient to activate IL-2 signaling in T cells independent of CD25.

Indeed, one of the superkines triggered proliferation in both CD25-deficient T cells and NK cells as well as in CD25-expressing T cells and NK cells.

The team then compared the effects of the superkine with those of both wild-type IL-2 and an IL-2-anti-IL-2 mAb complex. In prior mouse studies, the mAb complex had a longer half-life2 and generated more potent antitumor responses with less CD25-dependent pulmonary edema3-5 than wild-type IL-2.

In healthy mice, the superkine induced more than three times the number of cytotoxic T cells and led to less pulmonary edema than wild-type IL-2 (p<0.01).

In mice with melanoma, lung cancer or colon cancer, the superkine significantly decreased tumor growth after 18 days and induced less edema compared with wild-type IL-2 (p<0.05). The superkine and mAb complex produced similar results, and a detailed comparison of the two molecules awaits further studies.

Results were published in Nature.

More super

The next step for the superkine is longer-term studies in a variety of animal models.

"The work gives a good indication of increased antitumor effects, but more work is needed to discern effects on primary T cell responses versus simply bystander memory T cell expansion," said William Murphy, professor of dermatology and internal medicine at the University of California, Davis. "In addition, long-term efficacy, possible toxicities and immunogenicity against the superkine will have to be evaluated. Their study follows the treatment for only 18 days to provide a rather isolated snapshot of immunomodulation."

He said mouse studies of cancer immunotherapies need to run much longer to detect any signs of an impaired secondary antitumor response, long-term toxicities with repeated administration and potential tumor or immune rebound effects.

The Nature findings "can be validated by testing in nonhuman primates," said Jeffrey Bluestone, executive vice chancellor and provost and professor of medicine, pathology, microbiology and immunology at the University of California, San Francisco. "The structure of mouse IL-2 is very different from human IL-2. However, nonhuman primate IL-2 is very similar to human IL-2. Therefore, providing evidence that their human IL-2 superkine has an effect in monkeys would lend support to their strategy."

Regardless of animal type, Peter Rhode, VP of R&D at Altor BioScience Corp., said it will be important to better characterize the half-life of the superkines. Proleukin has a short half-life and is dosed every eight hours for five days per cycle in an in-patient hospital setting. Whether the IL-2 superkine can alter this dosing regimen needs to be probed, he said.

Bluestone said that even if the IL-2 superkine has a similar or shorter half-life than wild-type IL-2, it may not matter if the superkine is used in conjunction with a tumor vaccine. "Delivering the superkine as a short pulse with tumor vaccine inoculation may be sufficient to provide a strong antigen-specific immune response that could help initiate responses to tumors and not require continual treatment," he said.

This would depend on "whether IL-2 is used in vitro or in vivo to stimulate antigen-specific T cells," said Onur Boyman, professor of the Swiss National Science Foundation at the University of Zurich, senior consultant physician with the Allergy Unit of University Hospital Zurich and co-corresponding author on the Nature paper. "Moreover, it also depends whether IL-2 signaling is required for the survival or function of antigen-specific T cells in vivo." Those outcomes would have to be shown.

Altor's ALT-801, a fusion protein of IL-2 and a soluble T cell receptor (TCR) that specifically recognizes tumor cells that overexpress p53 antigen, is in Phase Ib/II testing to treat metastatic melanoma and metastatic urothelial cancer.

Stanford has filed for a patent covering super-2. The IP is licensed to Cephalon Inc., which was acquired by Teva in 2011. Teva has the IL-2 superkine in preclinical development.

Baas, T. SciBX 5(16); doi:10.1038/scibx.2012.405
Published online April 19, 2012


1.   Levin, A.M. et al. Nature; published online March 25, 2012; doi:10.1038/nature10975
Contact: K. Christopher Garcia, Stanford University School of Medicine, Stanford, Calif.
Contact: Onur Boyman, University of Zurich, Zurich, Switzerland

2.   Boyman, O. & Sprent, J. Nat. Rev. Immunol. 12, 180-190 (2012)

3.   Boyman, O. et al. Science 311, 1924-1927 (2006)

4.   Krieg, C. et al. Proc. Natl. Acad. Sci. USA 107, 11906-11911 (2010)

5.   Létourneau, S. et al. Proc. Natl. Acad. Sci. USA 107, 2171-2176 (2010)


      Altor BioScience Corp., Miramar, Fla.

      Novartis AG (NYSE:NVS; SIX:NOVN), Basel, Switzerland

      Stanford University, Stanford, Calif.

      Stanford University School of Medicine, Stanford, Calif.

      Teva Pharmaceutical Industries Ltd. (NASDAQ:TEVA),
Petah Tikva, Israel

      University of California, Davis, Calif.

      University of California, San Francisco, Calif.

      University Hospital Zurich, Zurich, Switzerland

      University of Zurich, Zurich, Switzerland