Increased cytotoxicity of interleukin 2-pseudomonas exotoxin (IL2-PE) chimeric proteins containing a targeting signal for lysosomal membranes.
IL2-PE40 is a chimeric protein composed of human interleukin 2 (IL2) genetically fused to the amino terminus of a truncated form of pseudomonas exotoxin lacking its cell recognition domain (PE40). IL2-PE40 is extremely cytotoxic to IL2 receptor positive cells. This chimeric protein was found to be an effective and selective immunosuppressive agent for IL2 receptor targeted therapy in many models of disorders of the immune response where activated T-cells play a crucial role. In an attempt to produce an improved IL2-PE40 chimeric protein, we constructed new IL2-PE derivatives. This was done by inserting defined DNA sequences within the chimeric gene encoding IL2-PE40. Inserted sequences represent motifs of other proteins known to be targeted and/or sorted to specific compartments inside or outside the cell. One of the proteins, IL2-PE40(LAP+DUP), containing a targeted signal for lysosomal membrane, was 2-3-fold more active than IL2-PE40. The insertion of the LAP sequence also increased the cytotoxicity of another IL2-PE derivative, IL2-PE664Glu. Our results suggest that a selective targeting of IL2-PE chimeric proteins to lysosomes may enable the proteins to reach the cytosol more efficiently, thus improving its specific cytotoxicity. The LAP (lysosomal alkaline phosphatase) sequence may be used as a common motif for increasing the cytotoxicity of other chimeric proteins to be used for targeted immunotherapy.
Fishman
A
A
Department of Cellular Biochemistry, Hebrew University-Hadassah Medical School, Jerusalem, Israel.
Bar-Kana
Y
Y
Steinberger
I
I
Lorberboum-Galski
H
H
D036002:ADP Ribose Transferases; D000246:Adenosine Diphosphate Ribose / Q000378:metabolism; D001427:Bacterial Toxins; D001483:Base Sequence; D016923:Cell Death; D002478:Cells, Cultured; D004247:DNA / Q000096:biosynthesis; D005098:Exotoxins / Q000235:genetics / Q000378:metabolism / Q000494:pharmacology*; D006801:Humans; D007376:Interleukin-2 / Q000235:genetics / Q000378:metabolism / Q000494:pharmacology*; D007425:Intracellular Membranes / Q000187:drug effects; D008213:Lymphocyte Activation; D008247:Lysosomes / Q000187:drug effects* / Q000648:ultrastructure; D008969:Molecular Sequence Data; D009841:Oligonucleotides / Q000737:chemistry; D010835:Phytohemagglutinins / Q000494:pharmacology; D021382:Protein Sorting Signals; D011993:Recombinant Fusion Proteins / Q000378:metabolism / Q000494:pharmacology*; D013329:Structure-Activity Relationship; D013601:T-Lymphocytes / Q000276:immunology / Q000378:metabolism; D037521:Virulence Factors; D000097668:Pseudomonas aeruginosa Exotoxin A
D036002:ADP Ribose Transferases; D000246:Adenosine Diphosphate Ribose; D001427:Bacterial Toxins; D004247:DNA; D005098:Exotoxins; D007376:Interleukin-2; D009841:Oligonucleotides; D010835:Phytohemagglutinins; D021382:Protein Sorting Signals; D011993:Recombinant Fusion Proteins; D037521:Virulence Factors; D000097668:Pseudomonas aeruginosa Exotoxin A