title: Heterogeneity in interleukin-13 receptor expression and subunit structure in squamous cell carcinoma of head and neck: differential sensitivity to chimeric fusion proteins comprised of interleukin-13 and a mutated form of Pseudomonas exotoxin.
abstract: Squamous cell carcinoma of the head and neck (SCCHN) is characterized by a high proliferation index and marked propensity for local invasion resulting in poor prognosis for these patients. To develop tumor-targeted novel therapeutic agents, here we demonstrate that SCCHN cell lines express receptors for an immune regulatory cytokine, interleukin (IL) 13. By reverse transcription-PCR (RT-PCR), we found that 16 SCCHN cell lines express equally strong RT-PCR positive bands for mRNA of IL-13Ralpha1 and IL-4Ralpha chains. However, only three cell lines, HN12, YCUM911, and KCCT873, expressed a strong band for transcripts for IL-13Ralpha2 chain and five cell lines, YCUL891, KCCTC871, KCCL871, KCCTCM901, and RPMI 2650 expressed faint bands. Transcripts for IL-2Rgamma(c) chain were absent in all of the cell lines tested. Indirect immunofluorescence analysis for four different receptor chains confirmed RT-PCR results and showed pronounced expression of IL-13Ralpha2 protein in three high IL-13R expressing cell lines. All of the cell lines were equally positive for IL-13Ralpha1 and IL-4Ralpha chains. Receptor-binding studies demonstrated that IL-13Ralpha2-positive cell lines expressed a high density of IL-13 receptors. Using two chimeric proteins composed of IL-13 and mutated forms of Pseudomonas exotoxin (IL-13-PE38 or IL-13-PE38QQR), we found that these two fusion toxins were highly and equally cytotoxic to IL-13Ralpha2-positive SCCHN, whereas IL-13Ralpha2-negative cell lines showed low or no sensitivity to IL-13 toxins. To additionally substantiate the critical role of the IL-13Ralpha2 chain in IL-13R-mediated cytotoxicity, two head and neck tumor cell lines (YCUMS861 and KB), devoid of the transcripts of this chain, were transfected with IL-13Ralpha2 cDNA and then tested for cytotoxicity. Transient transfection of the IL-13Ralpha2 chain highly sensitized these cells to IL-13 toxin as compared with mock-transfected control cells. Thus, our results indicate that IL-13Ralpha2 is present in 50% SCCHN tumor cell lines; of these, 19% are high expresser for this chain and respond to IL-13 cytotoxin. Thus, IL-13 cytotoxin may be a useful agent for high IL-13R-expressing SCCHN.
authors.affiliation: Laboratory of Molecular Tumor Biology, Division of Cellular and Gene Therapies, Center for Biologics Evaluation and Research, Food and Drug Administration, Bethesda, MD 20892, USA.
authors.forename: Bharat H
authors.initials: BH
authors.lastname: Joshi
authors.forename: Koji
authors.initials: K
authors.lastname: Kawakami
authors.forename: Pamela
authors.initials: P
authors.lastname: Leland
authors.forename: Raj K
authors.initials: RK
authors.lastname: Puri
mesh_terms: D036002:ADP Ribose Transferases / Q000627:therapeutic use*; D001427:Bacterial Toxins / Q000627:therapeutic use*; D002294:Carcinoma, Squamous Cell / Q000188:drug therapy* / Q000235:genetics / Q000378:metabolism; D002455:Cell Division / Q000187:drug effects / Q000502:physiology; D003114:Colony-Forming Units Assay; D017931:DNA Primers / Q000737:chemistry; D019008:Drug Resistance, Neoplasm; D005098:Exotoxins / Q000627:therapeutic use*; D005455:Fluorescent Antibody Technique; D006258:Head and Neck Neoplasms / Q000188:drug therapy* / Q000235:genetics / Q000378:metabolism; D006801:Humans; D018793:Interleukin-13 / Q000627:therapeutic use*; D053661:Interleukin-13 Receptor alpha1 Subunit; D012333:RNA, Messenger / Q000378:metabolism; D012334:RNA, Neoplasm / Q000235:genetics / Q000378:metabolism; D018123:Receptors, Interleukin / Q000235:genetics / Q000378:metabolism*; D053660:Receptors, Interleukin-13; D011993:Recombinant Fusion Proteins / Q000627:therapeutic use*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014407:Tumor Cells, Cultured / Q000187:drug effects / Q000378:metabolism; D037521:Virulence Factors / Q000627:therapeutic use*; D000097668:Pseudomonas aeruginosa Exotoxin A
chemical_list: D036002:ADP Ribose Transferases; D001427:Bacterial Toxins; D017931:DNA Primers; D005098:Exotoxins; D018793:Interleukin-13; D053661:Interleukin-13 Receptor alpha1 Subunit; D012333:RNA, Messenger; D012334:RNA, Neoplasm; D018123:Receptors, Interleukin; D053660:Receptors, Interleukin-13; D011993:Recombinant Fusion Proteins; D037521:Virulence Factors; D000097668:Pseudomonas aeruginosa Exotoxin A; C508181:IL13RA1 protein, human