Biophysical and biochemical characterization of a liposarcoma‐derived recombinant MnSOD protein acting as an anticancer agent

• Published in: International journal of cancer 2008-12, Vol.123 (11), p.2684-2695
• Main Authors: Mancini, Aldo, Borrelli, Antonella, Schiattarella, Antonella, Aloj, Luigi, Aurilio, Michela, Morelli, Franco, Pica, Alessandra, Occhiello, Antonella, Lorizio, Roberto, Mancini, Roberto, Sica, Alessandro, Mazzarella, Lelio, Sica, Filomena, Grieco, Paolo, Novellino, Ettore, Pagnozzi, Daniela, Pucci, Piero, Rommelaere, Jean
• Format: Article
• Language: English
• Subjects: Amino Acid Sequence; Animals; anticancer agent; Antineoplastic Agents - pharmacology; Antineoplastic Agents - therapeutic use; antioxidant; Base Sequence; Biochemical Phenomena; Biochemistry; Biological and medical sciences; Biophysical Phenomena; Biophysics; Cell Line, Tumor; Chromatography, Affinity; Circular Dichroism; Dermatology; Estradiol - pharmacology; free radicals; Health; Humans; Hydrogen Peroxide - metabolism; Liposarcoma - drug therapy; Liposarcoma - enzymology; Liposarcoma - pathology; Medical sciences; Mice; Mice, Nude; Molecular Sequence Data; Peptide Mapping; Recombinant Proteins - chemistry; Recombinant Proteins - isolation & purification; Recombinant Proteins - metabolism; Recombinant Proteins - pharmacology; rMnSOD; Sequence Analysis; Spectrometry, Mass, Electrospray Ionization; Superoxide Dismutase - chemistry; Superoxide Dismutase - isolation & purification; Superoxide Dismutase - metabolism; Superoxide Dismutase - pharmacology; Tumors; Tumors of the skin and soft tissue. Premalignant lesions; Xenograft Model Antitumor Assays
• ISSN: 0020-7136; 1097-0215
• DOI: 10.1002/ijc.23791

A recombinant MnSOD (rMnSOD) synthesized by specific cDNA clones derived from a liposarcoma cell line was shown to have the same sequence as the wild‐type MnSOD expressed in the human myeloid leukaemia cell line U937, except for the presence of the leader peptide at the N‐terminus. These results were fully confirmed by the molecular mass of rMnSOD as evaluated by ES/MS analysis (26662.7 Da) and the nucleotide sequence of the MnSOD cDNA. The role of the leader peptide in rMnSOD was investigated using a fluorescent and/or 68Gallium‐labelled synthetic peptide. The labelled peptide permeated MCF‐7 cells and uptake could be inhibited in the presence of an excess of oestrogen. In vivo it was taken up by the tumour, suggesting that the molecule can be used for both therapy and diagnosis. The in vitro and in vivo pharmacology tests confirmed that rMnSOD is only oncotoxic for tumour cells expressing oestrogen receptors. Pharmacokinetic studies in animals performed with 125I‐ and 131I‐labelled proteins confirmed that, when administered systemically, rMnSOD selectively reached the tumour, where its presence was unambiguously demonstrated by scintigraphic and PET scans. PCR analysis revealed that Bax gene expression was increased and the Bcl2 gene was down regulated in MCF7 cells treated with rMnSOD, which suggests that the protein induces a pro‐apoptotic mechanism. © 2008 Wiley‐Liss, Inc.