Stabilization of neurotensin analogues: effect on peptide catabolism, biodistribution and tumor binding

Bruehlmeier, Matthias; Garayoa, Elisa Garcia; Blanc, Alain; Holzer, Barbara; Gergely, Suzanne; Tourwé, Dirk; Schubiger, P. August; Bläuenstein, Peter

doi:10.1016/s0969-8051(01)00304-3

Cited by 58 publications

(115 citation statements)

References 6 publications

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“…The finding of new specifically targeted radiopharmaceuticals is an important challenge in cancer research, where tumor-specific small peptides may have major potential applications, provided that they are stabilized to overcome their degradation by proteases and peptidases. Currently, analogues of NT and particularly of NT-(8 -13) are under investigation to target a variety of neuroendocrine human tumors overexpressing NT receptors (12)(13)20). We demonstrated here that NT and NT-(8 -13), once synthesized in a MAP form, fully retain their biological activity and become much more resistant to blood proteases.…”

Section: Discussionmentioning

confidence: 85%

Synthetic Peptides in the Form of Dendrimers Become Resistant to Protease Activity

Bracci

Falciani

Lelli

et al. 2003

Journal of Biological Chemistry

156

162

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In previous papers, we observed that dendrimers of peptide mimotopes of the nicotinic receptor ligand site are strong antidotes against the lethality of the nicotinic receptor ligand ␣-bungarotoxin. Although their in vitro activity is identical to that of dendrimers, the corresponding monomeric peptide mimotopes are not effective in vivo. Because the higher in vivo efficiency of dendrimers could not in this case be related to polyvalent interaction, the stability to blood protease activity of monomeric versus tetrabranched dendrimeric mimotope peptides was compared here by incubating three different mimotopes with human plasma and serum. Unmodified peptides and cleaved sequences were followed by high pressure liquid chromatography and mass spectrometry. Tetrabranched peptides were shown to be much more stable in plasma and also in serum. To assess the notable stability of multimeric peptides, different bioactive neuropeptides, including enkephalins, neurotensin and nociceptin, were synthesized in monomeric and tetrabranched forms and incubated with human plasma and serum and with rat brain membrane extracts. All the tetrabranched neuropeptides fully retained biological activity and generally showed much greater stability to blood and brain protease activity. Some tetrabranched peptides were also resistant to trypsin and chymotrypsin. Our findings provide new insights into the possible therapeutic use of bioactive peptides.Hundreds of peptides with potential therapeutic activities have been identified. These include naturally occurring peptide hormones and neurotransmitters, which influence and control series of vital functions, such as cell proliferation, tissue development, metabolism, immune defense, perception of pain, reproduction, behavior, and blood pressure. Selective agonists or antagonists of these natural peptides are extremely useful for the investigation of peptidergic systems and are also potential therapeutic agents (1). Moreover, several peptide fragments or mimotopes derived from potential therapeutic proteins show promising biological activity (2).However, the use of peptides as therapeutic drugs has largely been limited by their short half-life in vivo. Because peptides are mainly broken down by proteases and peptidases, peptide delivery is the bottleneck in the development of new peptide drugs. To increase peptide half-life, many strategies involving different levels of chemical modification are possible (3, 4). The introduction of D-amino acids, or pseudo amino acids, and peptide cyclization are the most common strategies to increase peptide stability. However, these modifications may profoundly alter peptide activity. Alternatively, peptidomimetic molecules can be developed by the synthesis of conformationally restricted compounds, in which the peptide is locally or globally constrained in order to reproduce the active conformation. The resulting structures are mostly non-peptide molecules, more resistant to degrading enzymes.In general, peptide molecules have the advantage of good specifici...

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Section: Discussionmentioning

confidence: 85%

Synthetic Peptides in the Form of Dendrimers Become Resistant to Protease Activity

Bracci

Falciani

Lelli

et al. 2003

Journal of Biological Chemistry

156

162

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“…The solution was cooled and neutralised (pH 6.5). This solution [17][18][19]23] was mixed with the NT-XIX analogue (0.02 μmol) and heated at 75°C for 1 h. The final product was analysed and purified with high-performance liquid chromatography (HPLC) to eliminate the excess of free peptide and other impurities.…”

Section: Synthesis Of Nt-xix and Radiolabellingmentioning

confidence: 99%

“…In blockade experiments, mice received 3.5-4 MBq of 99m Tc-NT-XIX co-injected i.v. with 0.3 mg/mouse of NT-XI administered intravenously, a doubly stabilised analogue of the same series with good affinity for NTR1 [18]. Animals were sacrificed at 1.5 h p.i., the different organs/tissues were removed and the radioactivity was measured in a γ-counter.…”

Section: Biodistribution Studiesmentioning

confidence: 99%

“…The three cleavage sites in the metabolic deactivation of NT (8)(9)(10)(11)(12)(13) [16]. Modification of Arg 8 -Arg 9 and Tyr 11 -Ile 12 led to increased plasma stability, high affinity for NTR1 and also good in vivo tumour uptake [17][18][19]. With one analogue (NT-XI), visualisation of the tumour of a patient with ductal pancreatic adenocarcinoma was possible, which demonstrates the potential utility of NT analogues for imaging of these tumours [20].…”

Section: Introductionmentioning

confidence: 99%

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A stable neurotensin-based radiopharmaceutical for targeted imaging and therapy of neurotensin receptor-positive tumours

García-Garayoa

Bläuenstein

Blanc

et al. 2008

Eur J Nucl Med Mol Imaging

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Purpose Neurotensin (NT) and its high affinity receptor (NTR1) are involved in several neoplastic processes. Thus, NT-based radiopharmaceuticals are potential tracers for targeted diagnosis and therapy of NTR-positive tumours. A new analogue based on NT(8-13), NT-XIX, with the three enzymatic cleavage sites stabilised, was synthesised and tested. Methods The synthesis was performed by Boc strategy. Labelling with 99m Tc/ 188Re was performed using the tricarbonyl technique. Metabolic stability was tested in vitro and in vivo. NT-XIX was further characterised in vitro in HT-29 cells and in vivo in nude mice with HT-29 xenografts. Results NT-XIX showed much longer half-lives than nonstabilised analogues. Binding to NTR1 was highly specific, although the affinity was lower than that of natural NT. Bound activity rapidly internalised into HT-29 cells and 50% remained trapped after 24 h. In the time-course biodistribution, the highest uptake was found in the tumour at all p.i. times. In vivo uptake was specific, and accumulation of activity in the kidneys was low. Radioactivity clearance from healthy organs was faster than that from the tumour, resulting in improved tumour-to-tissue ratios and good SPECT/CT imaging. Treatment with 188 Re-NT-XIX (30 MBq, in three or four fractions) decreased tumour growth by 50% after 3 weeks. Conclusion The high in vivo stability and the favourable in vivo behaviour makes NT-XIX an excellent candidate for the imaging and therapy of NTR1-positive tumours.

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“…There are a number of examples of N-methyl amino acid containing peptides with a range of bioactivities, including antibiotic (18), anti-viral (19), anti-cancer (20), and immunosuppressant activities (21). Importantly, modification of biologically active peptides by the inclusion of N-methyl amino acids into an original sequence has been shown to enhance the potency (22), change the receptor subtype selectivity (23), and stabilize a peptide toward proteolytic degradation in a biological system (24). This approach therefore offers the potential to optimize multiple parameters simultaneously.…”

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confidence: 99%

Rapid Optimization of a Peptide Inhibitor of Malaria Parasite Invasion by Comprehensive N-Methyl Scanning

Harris

Casey

Coley

et al. 2009

Journal of Biological Chemistry

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Apical membrane antigen 1 (AMA1) of the malaria parasite Plasmodium falciparum has been implicated in the invasion of host erythrocytes and is an important vaccine candidate. We have previously described a 20-residue peptide, R1, that binds to AMA1 and subsequently blocks parasite invasion. Because this peptide appears to target a site critical for AMA1 function, it represents an important lead compound for anti-malarial drug development. However, the effectiveness of this peptide inhibitor was limited to a subset of parasite isolates, indicating a requirement for broader strain specificity. Furthermore, a barrier to the utility of any peptide as a potential therapeutic is its susceptibility to rapid proteolytic degradation. In this study, we sought to improve the proteolytic stability and AMA1 binding properties of the R1 peptide by systematic methylation of backbone amides (N-methylation). The inclusion of a single N-methyl group in the R1 peptide backbone dramatically increased AMA1 affinity, bioactivity, and proteolytic stability without introducing global structural alterations. In addition, N-methylation of multiple R1 residues further improved these properties. Therefore, we have shown that modifications to a biologically active peptide can dramatically enhance activity. This approach could be applied to many lead peptides or peptide therapeutics to simultaneously optimize a number of parameters.

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Stabilization of neurotensin analogues: effect on peptide catabolism, biodistribution and tumor binding

Cited by 58 publications

References 6 publications

Synthetic Peptides in the Form of Dendrimers Become Resistant to Protease Activity

Synthetic Peptides in the Form of Dendrimers Become Resistant to Protease Activity

A stable neurotensin-based radiopharmaceutical for targeted imaging and therapy of neurotensin receptor-positive tumours

Rapid Optimization of a Peptide Inhibitor of Malaria Parasite Invasion by Comprehensive N-Methyl Scanning

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