1-Aminocyclobutanecarboxylic Acid Derivatives as Novel Structural Elements in Bioactive Peptides:  Application to Tuftsin Analogs

Abstract

Four novel 2,4-methano amino acids (MAAs, 1-aminocyclobutane-1-carboxylic acids) were synthesized. These include the basic MAA analogs of lysine (16), ornithine (5), and arginine (6) and the neutral methanovaline (22), related to proline. The above MAAs, as well as the MAA analog of homothreonine (7), were incorporated into the peptide chain of the immunomodulatory peptide tuftsin, Thr-Lys-Pro-Arg, known to enhance several biological activities mediated by phagocytic cells. The synthetic methano tuftsin analogs were assayed for their ability to stimulate interleukin-6 (IL-6) secretion by mouse peritoneal macrophages and for their stability in human serum toward enzymatic degradation. It was found that, at 2 x 10(-7) M, [MThr1]tuftsin (24) and an isomer of [MVal3]tuftsin (27a) were considerably more active than the parent peptide in augmentation of cytokine release. [MOrn2]Tuftsin (25) was equally potent. The analogs [MThr1]tuftsin (24) and [MOrn2]tuftsin (25), both pertaining to the proteolytically sensitive Thr-Lys bond of tuftsin, exhibited high resistance to enzymatic hydrolysis as compared to tuftsin. Using specific rabbit anti-tuftsin antibodies in a competitive enzyme-linked immunosorbent assay (ELISA) revealed that none of the MAA analogs can cross-react with tuftsin. It may indicate that the peptides assume global structures different than that of tuftsin.

Tuftsin derivatives of FITC, Tb–DOTA or Gd–DOTA as potential macrophage‐specific imaging biomarkers

“…These data show that the cells incubated with Gd-DOTA-tuftsin were magnetically labelled and detectable by MRI, tuftsin retaining its binding activity after being conjugated with Gd-complex, which confirms the targeting ability of the vectorized MRI probe. The peptide could be substantially more resistant to enzymatic degradation after linking with the Gd chelate (49,50), which may modify its in vivo biodistribution profile, rate and mode of clearance from the body, bioavailability and pharmacokinetics (51).…”

Synthesis of the Four Stereoisomers of 1‐Amino‐2‐(hydroxymethyl)cyclobutanecarboxylic Acid and Their Biological Evaluation as Ligands for the Glycine Binding Site of the NMDA Receptor

“…The combined organic layers were dried (Na 2 SO 4 ) and concentrated in vacuo. The crude product was purified by CC (ethyl acetate, R f ϭ 0.20) to give 800 mg (84%) of (ent)-3aϪd as a colorless oil (ratio of isomers determined by 1 (3S,6R)-6-tert-Butyl-3-[(R)-4-(tert-butyldimethylsilanyloxy)-3-hydroxybutyl]-5-methoxy-6-methyl-3,6-dihydro-2H-1,4-oxazin-2-one (4a), (3S,6R)-6-tert-Butyl-3-[(S)-4-(tert-butyldimethylsilanyloxy)-3-hydroxybutyl]-5-methoxy-6-methyl-3,6-dihydro-2H-1,4-oxazin-2-one (4b), (3R,6R)-6-tert-Butyl-3-[(S)-4-(tert-butyldimethylsilanyloxy)-3-hydroxybutyl]-5-methoxy-6-methyl-3,6-dihydro-2H-1,4-oxazin-2-one (4c), and (3R,6R)-6-tert-Butyl-3-[(R)-4-(tert-butyldimethylsilanyloxy)-3-hydroxybutyl]-5-methoxy-6-methyl-3,6-dihydro-2H-1,4- (3S,6R)-6-tert-Butyl-3-[(R)-4-(tert-butyldimethylsilanyloxy)-3-iodobutyl]-5-methoxy-6-methyl-3,6-dihydro-2H-1,4-oxazin-2-one (5a), (3S,6R)-6-tert-Butyl-3-[(S)-4-(tert-butyldimethylsilanyloxy)-3-iodobutyl]-5-methoxy-6-methyl-3,6-dihydro-2H-1,4-oxazin-2-one (5b), (3R,6R)-6-tert-Butyl-3-[(S)-4-(tert-butyldimethylsilanyloxy)-3-iodobutyl]-5-methoxy-6-methyl-3,6-dihydro-2H-1,4-oxazin-2-one (5c), and (3R,6R)-6-tert-Butyl-3-[(R)-4-(tert-butyldimethylsilanyloxy)-3-iodobutyl]-5-methoxy-6-methyl-3,6-dihydro-2H-1,4-oxazin-2-one (5d): Imidazole (407 mg, 5.95 mmol, 2.2 equiv. ), PPh 3 (782 mg, 2.98 mmol, 1.1 equiv.…”

“…The workup was performed as described for (ent)-5aϪd. CC (petroleum ether/ethyl acetate, 95:5; R f ϭ 0.32) yielded 5aϪd (1.25 g, 90%) as a colorless oil (ratio of isomers determined by 1 (23 mL), and the mixture was stirred for 1.5 h at 0°C. The reaction mixture was then stirred for 1.5 h at room temperature, after which solid Na 2 S 2 O 3 ·5H 2 O (1 g) and phosphate buffer (pH ϭ 7, 5 mL) were added.…”

“…The combined organic layers were dried (MgSO 4 ) and concentrated in vacuo. The crude product was purified by CC (petroleum ether/ethyl acetate, 90:10; R f ϭ 0.62) to give (ent)-5aϪd (1.067 g, 90%) as a colorless oil (ratio of isomers determined by 1 [18] Modulation of [ 3 H]MK-801 binding to pig cortical brain membranes in the presence of -glutamate (100 µM) under non-equilibrium conditions was studied as described previously. [20] K i values for test compounds were calculated from competition experiments with at least six concentrations of test compounds, by the use of Prism 2.01 (GraphPad Software, San Diego, CA).…”

“…[1,2] Although numerous asymmetric syntheses of 1-aminocyclopropane-and 1-aminocyclopentanecarboxylic acids have been developed, no asymmetric syntheses of 1-aminocyclobutanecarboxyclic acids are yet known, according to a recent review and to the best of our knowledge. [3] Moreover, only a few racemic syntheses of 1-aminocyclobutanecarboxylic acids have been published, [4Ϫ6] this being indicative of the unfavorable situation in four-membered ring syntheses with respect both to small-angle strain and to entropy.…”