Key findings
- Substituting norleucine at position 4 and D-phenylalanine at position 7 produced a peptide resistant to degradation by serum enzymes with prolonged activity, and 26 times as potent as the parent hormone in a melanoma adenylate cyclase assay. [1]
- Forced degradation under acidic, basic, neutral and oxidative stress, ultraviolet light and elevated temperature produced fourteen separable degradation products, with truncation, methylation, deacetylation and oxidation identified as the pathways. [2]
- A derivative of this analogue prepared for a lead chelator showed sub-nanomolar binding affinity for melanocortin receptor 1 with high selectivity against the closely related melanocortin receptor 4. [3]
- In human articular chondrocyte cultures the analogue partly reduced doxorubicin driven senescence markers, significantly reduced reactive oxygen and nitrogen species, and lowered interleukin 6 secretion without changing matrix metalloproteinase secretion. [4]
Identity and structure
- Composition
- A synthetic 13 amino acid peptidomimetic of alpha-melanocyte stimulating hormone carrying norleucine at position 4 and D-phenylalanine at position 7 in place of methionine and L-phenylalanine [2][1]
- A linear analogue
- This is a linear peptide; the cyclic lactam melanotropins were designed later using the acetylated form of this analogue as a starting structure and are different molecules [5]
- Enzymatic resistance
- Resistant to enzymatic degradation by serum enzymes in the cited characterisation, which is the property the two substitutions were introduced to produce [1]
- Derivatised constructs
- Chelator-bearing and radiolabelled derivatives of this analogue circulate in the imaging literature and are different materials from the unmodified peptide [3]
- Form as supplied
- Sterile lyophilized powder
Mechanism as studied
The design is a stability argument rather than a receptor one. Heat and alkali treatment of the parent hormone prolonged its activity, and gas chromatographic analysis of the hydrolysed product showed partial racemisation concentrated at the methionine at position 4 and the phenylalanine at position 7. Building those two changes in deliberately, as norleucine and D-phenylalanine, reproduced the prolonged activity without the treatment and gave resistance to serum enzymes. [1]
Receptor level work places the analogue at melanocortin receptor 1. Competition binding against a series of derivatives identified one built on this analogue as a sub-nanomolar melanocortin receptor 1 ligand with high selectivity against melanocortin receptor 4, and cell work in chondrocytes reports readouts downstream of that receptor, with a receptor-deficient animal line used as the comparison. [3][4]
Outside receptor binding, the peptide interacts with lipid bilayers. Conjugated by an N-terminal spacer to the outer surface of liposomes, it triggered vesicle aggregation, partial leakage and limited fusion, and tryptophan fluorescence indicated it partitions into the bilayer core. The effect appeared only when the peptide was covalently attached, not when it was free in solution. [6]
Research findings
In vitro
- System
- Frog skin bioassay preparations, mouse melanoma cell adenylate cyclase and tyrosinase assays, and serum enzyme incubations, comparing the parent hormone, its heat-alkali treated form, the norleucine analogue and the doubly substituted analogue
- Measured
- Melanosome dispersion in frog skin, quantitative gas chromatographic analysis of the hydrolysed heat-alkali treated peptide for racemisation, melanoma adenylate cyclase activation, melanoma tyrosinase stimulation, and resistance to serum enzymes
- Reported
- Racemisation after heat-alkali treatment was concentrated at positions 4 and 7. The synthesised norleucine and D-phenylalanine analogue gave prolonged activity identical to the treated peptide, was resistant to enzymatic degradation by serum enzymes, and was 26 times as potent as the parent hormone in the adenylate cyclase assay, with raised activity also in the frog skin and tyrosinase assays. [1]
Analytical
- System
- The peptide held under acidic, basic, neutral and oxidative stress, ultraviolet light exposure and 60 °C, following the relevant stability testing guidelines
- Measured
- Separation of degradation products by gradient reversed phase HPLC on a C18 column, and structural elucidation by UHPLC with high resolution tandem mass spectrometry using collision-induced dissociation
- Reported
- The peptide degraded under every applied stress condition, generating fourteen different degradation products which were separated and characterised. The identified pathways were truncation, methylation, deacetylation and oxidation. [2]
In vitro
- System
- Preformed liposomes containing thiol-reactive lipid derivatives, with the analogue or its 4 to 10 fragment extended at the N-terminus by a thiol-functionalised spacer and conjugated to the vesicle surface
- Measured
- Vesicle aggregation, leakage of inner contents, membrane lipid mixing by resonance energy transfer, mixing of inner contents, and tryptophan fluorescence emission, anisotropy and iodide quenching
- Reported
- Covalent conjugation to the liposome surface caused vesicle aggregation and partial leakage of inner contents, correlated with limited fusion by both lipid mixing and content mixing. The effect appeared only for covalently conjugated peptide. A 10 nanometre blue shift in tryptophan emission with raised anisotropy and altered iodide accessibility indicated partitioning into the bilayer core. [6]
Analytical
- System
- A series of compounds derived from alpha-melanocyte stimulating hormone including one built on this analogue and carrying a lead chelator, rather than the unmodified peptide, tested in competition binding and in melanoma bearing and healthy mice
- Measured
- Competition binding affinity at melanocortin receptor 1, selectivity against melanocortin receptor 4, and biodistribution of the labelled compound across tumour and normal organs
- Reported
- The derivative of this analogue showed sub-nanomolar binding affinity for melanocortin receptor 1 and high selectivity against melanocortin receptor 4. Its organ distribution in the animal arm was the most tumour-specific of the series, with the lowest hepatic and splenic accumulation. [3]
In vitro
- System
- Human articular chondrocytes from donors with and without joint degeneration, driven into senescence with doxorubicin
- Measured
- Senescence-associated beta-galactosidase, CDKN2A and CDKN1A, reactive oxygen and nitrogen species, metabolic activity, matrix metalloproteinase secretion and interleukin 6 secretion
- Reported
- The analogue partly mitigated the doxorubicin driven senescence markers and significantly reduced reactive oxygen and nitrogen species in the chondrocytes. It also attenuated interleukin 6 secretion without affecting matrix metalloproteinase secretion, and the effects differed by donor sex and by whether the cells came from degenerated tissue. [4]
Preclinical in vivo
- System
- Wild type and melanocortin receptor 1 signalling deficient mice aged up to 18 months, the animal arm of the same paper as the chondrocyte work above
- Measured
- Knee joint chondrocyte melanocortin receptor 1 expression and apoptosis, subchondral bone architecture, and synovial immune cell populations in male and female animals
- Reported
- Loss of melanocortin receptor 1 signalling worsened the spontaneous age related changes in a sex dependent way, most clearly in female animals, and altered both subchondral bone parameters and synovial immune cell profiles. [4]
Preclinical in vivo
- System
- Zebrafish larvae, comparing synthetic and naturally occurring alpha-melanocyte stimulating hormone variants from several species with the human form as the standard and the cyclic analogue melanotan II as a comparator
- Measured
- Degree of melanosome dispersion in the larvae under each peptide variant
- Reported
- Unique variants from several fish species produced differing degrees of melanophore dispersion, and against the human form as standard the assay identified derivatives with greater physiological effect, with the cyclic analogue melanotan II showing a higher capacity for melanophore dispersion than human alpha-melanocyte stimulating hormone. [7]
Preclinical in vivo
- System
- Chitosan nanoparticles loaded with the analogue and surface-modified with polysorbate 80, rather than the free peptide, characterised in vitro and then followed in Wistar rats
- Measured
- Particle morphology by scanning electron microscopy, colloidal properties by dynamic light scattering, structure by ATR-FTIR spectroscopy, and distribution of fluorescently labelled and peptide-loaded particles in Wistar rats
- Reported
- Polysorbate 80 formed a flattened layer on the particle surface. Both the fluorescent and the peptide-loaded particles crossed the blood-brain barrier, reached the brain and accumulated in CA1 neurons of the dorsal hippocampus within two hours, which the authors present as a route around the enzymatic stability and distribution limits of the free peptide. [8]
Handling for in-vitro work
- Light and oxidation
- Degrades under ultraviolet light and under oxidative stress in the cited forced degradation work, alongside acidic, basic, neutral and thermal stress; protect reconstituted material from light and from oxidants [2]
- Serum enzymes
- Resistant to degradation by serum enzymes in the cited characterisation, unlike the parent hormone [1]
- Storage
- Lyophilized at -20 °C, dark and dry; reconstituted aliquots kept cold and used promptly
Open questions
- This linear analogue and the cyclic lactam melanotropins share a family and part of a name but are different molecules, and the cited work does not treat them as interchangeable.
- The binding work in the cited set used a chelator-bearing derivative rather than the unmodified peptide, so its affinity and selectivity numbers belong to that derivative.
- One cited animal study used a nanoparticle formulation, so its distribution result describes the particle rather than the free peptide.
- The forced degradation work lists four degradation pathways under stress, and the cited set does not establish which of them limits shelf life under ordinary storage.
A clinical literature on this analogue exists and concerns a finished formulation; it is out of scope for a research material profile.
Lot records
Check the record for the exact material you order. A published paper and a batch certificate answer different questions.
- RV-24-0026-1 ↗Melanotan-1 (Afamelanotide) · 99.40% HPLC2026-09-16
References
- Sawyer TK, Sanfilippo PJ, Hruby VJ, et al. 4-Norleucine, 7-D-phenylalanine-alpha-melanocyte-stimulating hormone: a highly potent alpha-melanotropin with ultralong biological activity. Proceedings of the National Academy of Sciences of the United States of America. 1980.
- Chawathe A, Sharma N Investigation of the stability profile of therapeutic α-MSH analogue: Insights from liquid chromatography-high resolution mass spectrometry analysis of afamelanotide. Journal of pharmaceutical and biomedical analysis. 2026.
- Scaffidi-Muta JM, Boucher D, Li KC, et al. Preclinical evaluation of MC1R targeted radiopharmaceuticals using the 203/212Pb theranostic pair. European journal of medicinal chemistry. 2026.
- Schäfer N, Kalke P, Mayakrishnan R, et al. The α-MSH-MC1R Axis Modulates Sex-Specific Senescence and Inflammation Processes in Human Articular Chondrocytes and Mice Knee Joints. Aging and disease. 2026.
- Al-Obeidi F, Castrucci AM, Hadley ME, et al. Potent and prolonged acting cyclic lactam analogues of alpha-melanotropin: design based on molecular dynamics. Journal of medicinal chemistry. 1989.
- de Souza DL, Frisch B, Duportail G, et al. Membrane-active properties of alpha-MSH analogs: aggregation and fusion of liposomes triggered by surface-conjugated peptides. Biochimica et biophysica acta. 2002.
- Hong TI, Hwang KS, Choi TI, et al. Zebrafish Bioassay for Screening Therapeutic Candidates Based on Melanotrophic Activity. International journal of molecular sciences. 2021.
- Herrera G, Scimonelli T, Lasaga M, et al. Polysorbate 80 coated chitosan nanoparticles for delivery of α-melanocyte stimulating hormone analog (NDP-MSH) to the brain reverse cognitive impairment related to neuroinflammation produced by a high-fat diet (HFD). Neuropharmacology. 2024.
Publication records fetched from PubMed on 2026-09-20. Profile text reviewed 2026-09-20.