title: Formulation and antitumor efficacy of liposomal-caprylated-TNF-SAM2.
abstract: The tumor necrosis factor (TNF) mutant TNF-SAM2 has previously been shown to have a therapeutic profile superior to parental TNF. To initially evaluate the characteristics of liposomal formulations of TNF-SAM2, it was modified with the N-hydroxysuccinimide ester of caprylic acid to increase its hydrophobic binding to multilamellar and small unilamellar vesicles (MLVs and SUVs). Native PAGE and fluorescamine analysis of acetylated parental TNF and TNF-SAM2 indicated that these proteins both displayed trimeric structures based on crosslinking/SDS-PAGE analysis and behaved similarly with respect to reactivity of their amino functions. Limited N-terminal sequencing analysis of partially acetylated (approx 3 acetyl groups per trimer) TNF-SAM2 indicated that the N-terminal Val was not modified; this was also concluded based on HPLC/mass spectrometric (LC-MS) analysis of Glu C digests. LC-MS analysis of tryptic digests of the acetylated TNF-SAM2 indicated that Lys-98 was unreactive. Molecular ions corresponding to acetylated Lys-containing peptides for all five other Lys residues could be detected; none appeared hyperreactive, but Lys-11 appeared hyporeactive. MLVs composed of DMPC/DMPG (7:3) and SUVs composed of DPPC/DSPC (1:1) displayed high capacity for binding to caprylated TNF-SAM2. These formulations of caprylated TNF-SAM2 displayed tumor necrotizing and growth-inhibitory activity in a syngeneic tumor model, and may be candidates for clinical development.
authors.affiliation: Department of Tumor Biology, University of Texas MD Anderson Cancer Center, Houston 77381, USA.
authors.forename: K
authors.initials: K
authors.lastname: Akimaru
authors.forename: E
authors.initials: E
authors.lastname: Auzenne
authors.forename: Y
authors.initials: Y
authors.lastname: Akimaru
authors.forename: M E
authors.initials: ME
authors.lastname: Leroux
authors.forename: A C
authors.initials: AC
authors.lastname: Hayman
authors.forename: T
authors.initials: T
authors.lastname: Utsumi
authors.forename: G
authors.initials: G
authors.lastname: Soma
authors.forename: J
authors.initials: J
authors.lastname: Klostergaard
mesh_terms: D015060:1,2-Dipalmitoylphosphatidylcholine; D000818:Animals; D002152:Calorimetry, Differential Scanning; D002851:Chromatography, High Pressure Liquid; D004134:Dimyristoylphosphatidylcholine; D004337:Drug Carriers; D008081:Liposomes; D013058:Mass Spectrometry; D051379:Mice; D008807:Mice, Inbred BALB C; D010446:Peptide Fragments / Q000737:chemistry; D010715:Phosphatidylglycerols; D012513:Sarcoma, Experimental / Q000628:therapy*; D014409:Tumor Necrosis Factor-alpha / Q000008:administration & dosage* / Q000737:chemistry / Q000627:therapeutic use*
chemical_list: D004337:Drug Carriers; D008081:Liposomes; D010446:Peptide Fragments; D010715:Phosphatidylglycerols; C109083:TNF-SAM2; D014409:Tumor Necrosis Factor-alpha; D015060:1,2-Dipalmitoylphosphatidylcholine; C002773:dimyristoylphosphatidylglycerol; D004134:Dimyristoylphosphatidylcholine