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 CBER Research Projects

Project Title

Interleukins, Receptors, and Targeting in Tumor Biology and AIDS

Principal Investigator

Raj K. Puri

Laboratory

Laboratory of Molecular Tumor Biology; Division of Cellular and Gene Therapy; Office of Therapeutics Research and Review

Project Summary

Interleukin-4 (IL-4), its mutants and soluble receptors are being tested in the clinic by systemic injection or by gene transfer (using viral or plasmid vectors) for the treatment of rheumatoid arthritis, bronchial asthma, and cancer. It is expected that IL-13, its mutants and soluble receptors will also be tested in the clinic in the near future. Studies are underway to characterize the structure, function, signal transduction and targeting of receptors (R) for IL-4 and IL-13 to immune and cancer cells.

  1. Previous reconstitution studies have demonstrated that IL-13R alpha1 chain, but not alpha2 chain, is a novel component of the IL-4R system. Thus, IL-4R and IL-13R share two chains (IL4R alpha and IL-13R alpha1) with each other that may help explain similar biological activities of IL-4 and IL-13 on many different cell types.

  2. To determine the subunit structure of IL-13 receptor RT-PCR and immunofluorescence studies were performed in primary brain tumor cells. These studies established that IL-13R alpha2 chain is over expressed on glioma cells, which may serve as a marker, or a target for receptor directed cancer therapy. Later studies demonstrated that IL-13R alpha2 chain is internalized after binding to the ligand and that a dileucine motif in the transmembrane domain of IL-13R alpha2 chain is responsible for this internalization.

  3. To create a mutant of IL-13 that is cancer IL-13R selective, we created a novel IL-13 mutant by site directed mutagenesis. This molecule termed IL-13R112D in which arginine (R) at position 112 was substituted to aspartic acid (D). This mutant was a 10x better agonist than wild type (wt) IL-13. We then generated another mutation in the alpha helix A of the IL-13 molecule. In this molecule E was changed to K at aa 13. This molecule turned out to be a powerful antagonist. IL-13E13K bound to IL-13R with 4 fold higher affinity than wtIL-13 and it neutralized the biological activity of wtIL-13 in EBV B cells, monocytic cell line and cancer cells. In addition, IL13E13K inhibited IL-13 induced STAT6 activation in cancer cells. Furthermore, IL-13E13K neutralized the cytotoxic activity of a fusion protein composed of wtIL-13 and a mutated form of Pseudomonas exotoxin (IL-13-PE38QQR). Based on these results, we concluded that IL-13E13K interacts with much stronger affinity than wtIL-13 and that Glu13 plays an important role in the interaction with its receptors. IL13E13K may be a useful agent for conditions where neutralization of IL-13 would be desirable e.g., inflammatory diseases such as asthma, allergic diseases and perhaps in cancer in which IL-13 acts as an autocrine growth factor.

  4. The IL-4 and IL-13R directed targeting of a Pseudomonas exotoxin, Diphtheria toxin, or alternatively receptor directed gene transfer is also being investigated. The receptors for these two interleukins are expressed in abundance on many human tumor cell lines and offer an attractive target for toxin therapy or gene therapy. In vivo experiments in immunodeficient mice with human head and neck tumors have demonstrated complete responses in a dose-dependent and route of administration-dependent manner in response to IL-4 toxin administration. Similarly, IL-13-toxin has shown remarkable antitumor activity in animal models of prostate cancer and head and neck cancer particularly when tumor cells are forced to express high levels of IL-13R alpah2 chain.

  5. Our previous studies on IL-4R targeting resulted in a Phase I clinical trial for malignant glioma. This trial is currently ongoing at many major medical Centers in the USA and in Germany. IL-13-toxin is also in clinical trial for brain tumor therapy at many clinical sites. Abstracts of these studies will be presented at the Congress of Neurological Surgeons in San Diego and Society of Neuro-Oncology in Washington, DC this fall. These clinical studies will help elucidate the safety and efficacy of these and other recombinant chimeric toxins being tested in clinic under various INDs.

Publications

  • Cancer Res 2000 Mar 1;60(5):1168-72
    Interleukin-13 receptor alpha chain: a novel tumor-associated transmembrane protein in primary explants of human malignant gliomas.
    Joshi BH, Plautz GE, Puri RK
    Pub Med

  • J Biol Chem 2000 May 5;275(19):14375-14380
    Conversion of Interleukin-13 into a High Affinity Agonist by a Single Amino Acid Substitution.
    Oshima Y, Joshi BH, Puri RK
    Pub Med

  • Blood 2000 Jun 1;95(11):3506-3513
    Interleukin-13 fusion cytotoxin as a potent targeted agent for AIDS-Kaposi's sarcoma xenograft.
    Husain SR, Puri RK
    Pub Med

  • Cancer Res 2000 Jun 1;60(11):2981-7
    Structure, function, and targeting of interleukin 4 receptors on human head and neck cancer cells.
    Kawakami K, Leland P, Puri RK
    Pub Med

  • Clin Cancer Res 2000 Jun;6(6):2157-65
    Intratumoral administration of recombinant circularly permuted interleukin-4-Pseudomonas exotoxin in patients with high-grade glioma.
    Rand RW, Kreitman RJ, Patronas N, Varricchio F, Pastan I, Puri RK
    Pub Med

  • Mol Med 6: (3) 165-178 MAR 2000
    Human breast carcinoma cells express type II IL-4 receptors and are sensitive to antitumor activity of a chimeric IL-4-Pseudomonas exotoxin fusion protein in vitro and in vivo.
    Leland P, Taguchi J, Husain SR, Kreitman RJ, Pastan I, Puri RK
    Pub Med

  • Hum Gene Ther 2000 Sep 1;11(13):1829-1835
    Sensitization of Cancer Cells to Interleukin 13-Pseudomonas Exotoxin-Induced Cell Death by Gene Transfer of Interleukin 13 Receptor alpha Chain.
    Kawakami K, Joshi BH, Puri RK
    Pub Med

  • J Biol Chem 2001 May 4;276(18):15185-91
    Characterization of a powerful high affinity antagonist that inhibits biological activities of human interleukin-13.
    Oshima Y, Puri RK
    Pub Med

  • Blood 2001 May 1;97(9):2673-2679
    The interleukin-13 receptor alpha2 chain: an essential component for binding and internalization but not for interleukin-13-induced signal transduction through the STAT6 pathway.
    Kawakami K, Taguchi J, Murata T, Puri RK
    Pub Med

  • J Biol Chem 2001 July 6;276(27):25114-25120
    Identification of distinct roles for a di-leucine motif and a tyrosine internalization motif in the interleukin-13 binding component IL-13 receptor {alpha}2 chain.
    Kawakami K, Takeshita F, Puri RK
    Pub Med

  • Cancer Res 2001 Aug 15;61(16):6194-200
    Interleukin-13 Receptor-targeted Cancer Therapy in an Immunodeficient Animal Model of Human Head and Neck Cancer.
    Kawakami K, Kawakami M, Joshi BH, Puri RK
    Pub Med

  • FASEB J 2001 Jun;15(8):1469-71
    A novel interleukin 13 (IL-13) antagonist that blocks the biological activity of human IL-13 in immune and nonimmune cells.
    Oshima Y, Puri RK
    Pub Med

  • Int J Cancer 2001 Apr 15;92(2):168-175
    Interleukin-13 receptor as a unique target for anti-glioblastoma therapy.
    Husain SR, Joshi BH, Puri RK
    Pub Med

Last Updated: 4/1/2002

 

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Date created: September 25, 2003
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