Evaluation of the effect of D-amino acid incorporation into amyloid-reactive peptides.

BACKGROUND: Systemic amyloidoses comprise diseases characterized by the deposition of proteinaceous material known as amyloid. Currently, without performing multiple biopsies, there is no way to ascertain the extent of amyloid deposition in patients-a critical piece of information that informs prognosis and therapeutic strategies. We have developed pan-amyloid-targeting peptides for imaging amyloid and recently have adapted these for use as pre-targeting agents in conjunction with immunotherapy. Incorporation of D-amino acids in these peptides may enhance serum half-life, which is an important characteristic of effective peptide therapeutics. Herein, we assess the effects of partial incorporation of D-amino acids into the amyloidophilic peptide p5 on in vivo amyloid reactivity.

METHODS: Peptides, referred to as AQA p5 (d) , aqa p5, and AQA p5, were radiolabeled with iodine-125 and the tissue biodistribution (% injected dose/gram) measured in healthy mice at multiple time points post-injection. Microscopic distribution of the peptides was further visualized using microautoradiography (ARG). Peptides aqa p5 and AQA p5 were injected into healthy and amyloid-laden mice and evaluated by using SPECT/CT imaging at 1, 4 and 24 h post injection.

RESULTS: Biodistribution data and ARG revealed persistent retention of [125 I]AQA p5 (d) in the liver and kidneys of healthy mice for at least 24 h. In contrast, peptides [125 I]aqa p5 and [125 I]AQA p5 did not bind these organs and was significantly lower than [125 I]AQA p5 (d) at 24 h post injection (p < 0.0001). SPECT/CT imaging of amyloid-laden mice revealed accumulation of both [125 I]aqa p5 and [125 I]AQA p5 in amyloid-affected organs; whereas, in healthy mice, [125 I]aqa p5 was observed in the kidneys and liver at early time points, and free radioiodide liberated during catabolism of [125 I]AQA p5 was seen in the stomach and thyroid. Autoradiography confirmed that both [125 I]aqa p5 and [125 I]AQA p5 peptides specifically bound amyloid with no off-target binding to healthy organs.

CONCLUSION: Incorporation of D-amino acids in amyloid-binding regions of amyloidophilic peptides resulted in off-target binding; however, N-terminus placement retained amyloid-specificity and evasion of deiodinases. Peptide aqa p5, or similar reagents, may prove useful in novel immunotherapy strategies as well as for imaging renal, gastric and pancreatic amyloidosis.