title: An anthrax toxin variant with an improved activity in tumor targeting.
abstract: Anthrax lethal toxin (LT) is an A-B type toxin secreted by Bacillus anthracis, consisting of the cellular binding moiety, protective antigen (PA), and the catalytic moiety, lethal factor (LF). To target cells, PA binds to cell-surface receptors and is then proteolytically processed forming a LF-binding competent PA oligomer where each LF binding site is comprised of three subsites on two adjacent PA monomers. We previously generated PA-U2-R200A, a urokinase-activated PA variant with LF-binding subsite II residue Arg200 mutated to Ala, and PA-L1-I210A, a matrix metalloproteinase-activated PA variant with subsite III residue Ile210 mutated to Ala. PA-U2-R200A and PA-L1-I210A displayed reduced cytotoxicity when used singly. However, when combined, they formed LF-binding competent heterogeneous oligomers by intermolecular complementation, and achieved high specificity in tumor targeting. Nevertheless, each of these proteins, in particular PA-L1-I210A, retained residual LF-binding ability. In this work, we screened a library containing all possible amino acid substitutions for LF-binding site to find variants with activity strictly dependent upon intermolecular complementation. PA-I207R was identified as an excellent replacement for the original clockwise-side variant, PA-I210A. Consequently, the new combination of PA-L1-I207R and PA-U2-R200A showed potent anti-tumor activity and low toxicity, exceeding the performance of the original combination, and warranting further investigation.
authors.affiliation: Microbial Pathogenesis Section, Laboratory of Parasitic Diseases, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, Maryland, USA.
authors.forename: Alexander N
authors.initials: AN
authors.lastname: Wein
authors.affiliation: Proteases and Tissue Remodeling Section, Oral and Pharyngeal Cancer Branch, National Institute of Dental and Craniofacial Research, National Institutes of Health, Bethesda, Maryland, USA.|Program of Pharmacology and Experimental Therapeutics, Tufts University School of Medicine, Boston, MA, USA.
authors.forename: Diane E
authors.initials: DE
authors.lastname: Peters
authors.affiliation: Microbial Pathogenesis Section, Laboratory of Parasitic Diseases, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, Maryland, USA.
authors.forename: Zaheer
authors.initials: Z
authors.lastname: Valivullah
authors.affiliation: Microbial Pathogenesis Section, Laboratory of Parasitic Diseases, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, Maryland, USA.
authors.forename: Benjamin J
authors.initials: BJ
authors.lastname: Hoover
authors.affiliation: Microbial Pathogenesis Section, Laboratory of Parasitic Diseases, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, Maryland, USA.
authors.forename: Aparna
authors.initials: A
authors.lastname: Tatineni
authors.affiliation: Microbial Pathogenesis Section, Laboratory of Parasitic Diseases, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, Maryland, USA.
authors.forename: Qian
authors.initials: Q
authors.lastname: Ma
authors.affiliation: Microbial Pathogenesis Section, Laboratory of Parasitic Diseases, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, Maryland, USA.
authors.forename: Rasem
authors.initials: R
authors.lastname: Fattah
authors.affiliation: Proteases and Tissue Remodeling Section, Oral and Pharyngeal Cancer Branch, National Institute of Dental and Craniofacial Research, National Institutes of Health, Bethesda, Maryland, USA.
authors.forename: Thomas H
authors.initials: TH
authors.lastname: Bugge
authors.affiliation: Microbial Pathogenesis Section, Laboratory of Parasitic Diseases, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, Maryland, USA.
authors.forename: Stephen H
authors.initials: SH
authors.lastname: Leppla
authors.affiliation: Microbial Pathogenesis Section, Laboratory of Parasitic Diseases, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, Maryland, USA.
authors.forename: Shihui
authors.initials: S
authors.lastname: Liu
mesh_terms: D000818:Animals; D000942:Antigens, Bacterial / Q000235:genetics / Q000494:pharmacology*; D001427:Bacterial Toxins / Q000235:genetics / Q000494:pharmacology*; D002460:Cell Line; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects; D004305:Dose-Response Relationship, Drug; D005260:Female; D008264:Macrophages / Q000166:cytology / Q000187:drug effects*; D008297:Male; D008810:Mice, Inbred C57BL; D050505:Mutant Proteins / Q000235:genetics / Q000494:pharmacology*; D020125:Mutation, Missense; D009374:Neoplasms, Experimental / Q000188:drug therapy* / Q000473:pathology; D013997:Time Factors; D047368:Tumor Burden / Q000187:drug effects
chemical_list: D000942:Antigens, Bacterial; D001427:Bacterial Toxins; D050505:Mutant Proteins; C030325:anthrax toxin