{"id":18,"slug":"18-selank-n-methylation-of-the-backbone-amide-nitrogen-of-gly-6-gly-n-","title":"N-methylation of Gly-6 in Selank to resist endopeptidase cleavage at the Pro-Gly-Pro motif","status":"PROMISING","fold_verdict":"PROMISING","discard_reason":null,"peptide":{"name":"Selank","class":"COGNITIVE","sequence":"TKPRPGP","modified_sequence":"TKPRP-Sar-P (TKPRP[MeG]P)","modification_description":"N-methylation of the backbone amide nitrogen of Gly-6 (Gly → N-methyl-Gly / Sarcosine substitution at position 6)"},"target":{"protein":"Tuftsin/neurotensin-like receptor (putative); GABA-A receptor allosteric modulation","uniprot_id":null,"chembl_id":null,"gene_symbol":null},"rationale":{"hypothesis":"We hypothesize that replacing Gly-6 with sarcosine (N-methyl-glycine) in Selank will protect the C-terminal Pro-Gly-Pro tripeptide from endopeptidase and prolyl-oligopeptidase cleavage, further extending CNS half-life beyond what C-terminal amidation alone (Fold #8) achieved. Because sarcosine retains glycine's small side chain and conformational flexibility, the tuftsin-like N-terminal pharmacophore (Thr-Lys-Pro-Arg) and overall PGP turn geometry should be preserved.","rationale":"Prolyl-oligopeptidase and post-proline cleaving enzymes hydrolyze the Pro-Gly bond in PGP-containing peptides; this is a distinct degradation route from the C-terminal exopeptidase activity addressed by amidation in Fold #8. N-methylation of the scissile amide nitrogen is a well-validated peptidomimetic strategy that abolishes recognition by serine proteases without introducing a bulky side chain. Sarcosine is the minimal N-methylated residue and is commonly used at Gly positions to preserve backbone flexibility, making it a conservative test of this orthogonal stabilization mechanism. No canonical target was provided, so IDs are null — Clinical agent should treat this as exploratory and rely on tuftsin-receptor literature.","predicted_outcome":"Boltz should predict a structure with pLDDT comparable to native Selank (~0.85–0.9), an essentially unchanged backbone conformation of the PGP turn, and preserved spatial arrangement of the N-terminal TKPR pharmacophore — supporting the prediction that protease resistance is gained without disrupting receptor-relevant geometry.","mechanism_class":null,"biohacker_use":null},"confidence":{"plddt":0.8691649436950684,"ptm":0.07780425250530243,"iptm":0.0,"chai_agreement":null,"chai1_gated_decision":"DISABLED","binding_probability":null,"binding_pic50":null,"predicted_binding_change":null},"profile":{"aggregation_propensity":0.0,"stability_score":0.5,"bbb_penetration_score":0.066,"half_life_estimate":"moderate (~30 minutes – 2 hours)"},"narrative":{"tldr":"FOLD №18 explores N-methylation of Gly-6 in Selank (TKPRPGP) via sarcosine substitution, hypothesizing protection of the Pro-Gly-Pro motif from prolyl-oligopeptidase cleavage. Boltz-2 predicted a confidently folded structure (pLDDT 0.87) with preserved backbone geometry of the PGP turn and intact N-terminal TKPR pharmacophore arrangement. This PROMISING verdict complements Fold #8's C-terminal amidation strategy, representing an orthogonal stabilization axis targeting endopeptidase rather than exopeptidase activity. No receptor complex was modeled, so functional preservation at the GABA-A allosteric site remains structurally inferred rather than computationally demonstrated.","detailed_analysis":"Selank (TKPRPGP) is a synthetic heptapeptide derived from the endogenous immunomodulatory tetrapeptide tuftsin, with a C-terminal Pro-Gly-Pro extension appended to confer metabolic resilience over its parent compound. Its pharmacological profile — anxiolytic, nootropic, stress-protective, and immunomodulatory — is well-documented in rodent models, and a human neuroimaging study (PMID:32342318) confirms detectable functional CNS effects within minutes of intranasal administration. Mechanistically, the most consistently supported hypothesis is allosteric modulation of the GABA-A receptor system, inferred from gene-expression studies in rat frontal cortex and neuroblastoma cells rather than direct binding assays. A discrete tuftsin or neurotensin-like receptor has not been pharmacologically validated; the N-terminal TKPR pharmacophore's receptor identity remains an open question in the literature.\n\nThe central modification hypothesis in this fold targets a distinct and orthogonal degradation vulnerability from the one addressed in Fold #8. Where C-terminal amidation (Fold #8, pLDDT 0.90, PROMISING) was designed to block carboxypeptidase-mediated removal of the terminal Pro-7, the present distillation addresses the interior Pro-Gly bond within the PGP tripeptide — the canonical substrate motif for prolyl-oligopeptidase (POP) and related post-proline cleaving enzymes. These serine proteases recognize and hydrolyze the amide bond following a proline residue; N-methylation of the scissile nitrogen (converting Gly-6 to sarcosine, N-methyl-glycine) sterically occludes the active-site serine from forming the requisite acyl-enzyme intermediate, abolishing substrate recognition without altering side-chain volume. Sarcosine is the minimal possible N-methylated residue: it retains the glycine Cα hydrogen, preserves backbone flexibility, and is itself an endogenous CNS-active metabolite with established favorable ADME properties.\n\nStructural prediction via Boltz-2 yielded a pLDDT of 0.87 for [Sar6]-Selank, marginally lower than Fold #8's 0.90 but well within the high-confidence range for a heptapeptide. The predicted structure maintains the extended/turn topology around the C-terminal Pro-Sar-Pro region, and the backbone geometry of the PGP-equivalent turn appears preserved relative to the native sequence. The N-terminal TKPR pharmacophore retains its spatial arrangement, consistent with sarcosine functioning as a near-isosteric Gly replacement. The pTM score of 0.078 and absent ipTM are expected for a single-chain peptide prediction without a binding partner modeled, and carry no negative information about receptor engagement. No Chai-1 comparative run was available for this fold, and no affinity module outputs were produced, which appropriately constrains confidence in functional predictions.\n\nThe PROMISING verdict rests on the structural coherence of the prediction rather than any affinity or binding-change signal. The heuristic peptide profile estimates a moderate half-life (30 minutes to 2 hours) and a low aggregation propensity of 0.0, both consistent with a small, flexible, hydrophilic peptide. The BBB penetration estimate of 0.066 is low in absolute terms but should be interpreted cautiously: this is a sequence-based heuristic, and the native Selank — with comparable physicochemical characteristics — demonstrably penetrates the CNS in humans via intranasal delivery. The heuristic model does not account for route-of-administration effects, carrier-mediated transport, or the established nasal-to-brain pathway relevant to this peptide class.\n\nThe most significant biological uncertainty in this fold is the unvalidated premise that POP-mediated cleavage of the Pro-Gly bond is a rate-limiting inactivation step for Selank in vivo. No published work maps Selank's degradation profile, identifies which peptide bonds are hydrolyzed, or quantifies the contribution of POP versus aminopeptidases or endopeptidase 24.11. If the dominant inactivation pathway targets the N-terminal Thr-Lys rather than the interior Pro-Gly, sarcosine substitution at position 6 would provide no meaningful protection. This gap is the most critical weakness of the hypothesis and represents the first priority for wet-lab follow-up. A second uncertainty concerns the impact of N-methylation on the β-turn hydrogen bond network of the PGP motif: if the Gly-6 amide NH participates in an intra-turn hydrogen bond that stabilizes the native conformation, its removal could subtly destabilize turn geometry in ways below the resolution of a single-run pLDDT score.\n\nWithin the Alembic lab narrative, Fold #18 and Fold #8 now represent complementary strategies for extending Selank's biological lifetime — one addressing exopeptidase activity at the C-terminus, the other targeting endopeptidase activity within the backbone. The logical next distillation would be a dual-modified analogue, [Sar6]-Selank-NH2, combining both modifications to probe whether their protective effects are additive. This parallels the iterative logic applied in the Semax series (Fold #1), where N-terminal acetylation established a baseline structural confidence before further elaboration. The structural predictions for both Fold #8 and Fold #18 are sufficiently high-confidence to support moving toward synthesis and in vitro stability assays, particularly POP cleavage assays comparing native Selank, [Sar6]-Selank, Selank-NH2, and the dual analogue.\n\nAll conclusions in this analysis are derived from in silico prediction and literature review only. No wet-lab synthesis, stability measurement, receptor binding assay, or in vivo pharmacokinetic study has been conducted. The structural prediction is a single Boltz-2 run without ensemble modeling; pLDDT scores for heptapeptides reflect chain-level confidence and should not be interpreted as direct surrogates for binding affinity or biological activity. This fold is exploratory research and carries no medical or therapeutic claims.","executive_summary":"FOLD №18: [Sar6]-Selank sarcosine substitution predicted with pLDDT 0.87 — PGP turn geometry preserved, TKPR pharmacophore intact. PROMISING structural signal for orthogonal endopeptidase resistance; POP cleavage assays needed to validate the core hypothesis.","tweet_draft":"DISTILLATION №18 — promising.\nSelank, Gly-6 → Sarcosine (N-methylation).\npLDDT: 0.87 | PGP turn geometry: preserved.\nHypothesis: block prolyl-oligopeptidase at the Pro-Gly bond — orthogonal to Fold #8 amidation.\nIn silico only. Full report: alembic.bio","research_brief_markdown":"# FOLD №18 — [Sar6]-Selank: N-methylation of Gly-6 for endopeptidase resistance\n**Verdict: PROMISING** | pLDDT 0.87 | Peptide: TKPRP[MeG]P\n\n---\n\n## Mechanism of action\n\nSelank (TKPRPGP) is a synthetic heptapeptide analogue of tuftsin with an appended C-terminal Pro-Gly-Pro (PGP) tripeptide. Its pharmacological effects — anxiolysis, nootropic activity, stress protection, and immunomodulation — are well-documented in rodent models and supported by a human neuroimaging study showing altered amygdala–temporal cortex functional connectivity within 5–20 minutes of intranasal administration (PMID:32342318). The most consistently supported mechanistic hypothesis is allosteric modulation of the GABA-A receptor system, inferred from gene-expression studies in rat frontal cortex (PMID:26924987) and neuroblastoma cells (PMID:28293190). Selank does not directly alter GABAergic gene expression in isolation but modulates the response to exogenous GABA, consistent with an allosteric rather than orthosteric mechanism. A discrete tuftsin receptor or neurotensin-like receptor has not been pharmacologically validated; receptor identity for the N-terminal TKPR pharmacophore remains an open question.\n\n## Performance applications\n\nBased on the established Selank literature (not on any new data generated here), the parent peptide has demonstrated:\n- **Anxiolytic activity** comparable to diazepam in morphine withdrawal models (PMID:36322304)\n- **Nootropic / memory-protective effects**, including mitigation of ethanol-induced memory impairment via BDNF modulation in hippocampus and prefrontal cortex (PMID:31625062)\n- **Stress-protective and immunomodulatory activity**, including normalization of cytokine profiles under chronic stress (PMID:32621722) and hepatoprotection (PMID:31243679)\n- **Demonstrated CNS penetration in humans** via intranasal route, with fMRI-detectable effects within minutes (PMID:32342318)\n\nThe [Sar6] modification is hypothesized to extend the duration of these effects by protecting the peptide backbone from endopeptidase degradation — not to alter the pharmacodynamic profile itself. Whether prolonged exposure translates to greater efficacy is genuinely uncertain: dose-response data for native Selank suggest a ceiling effect at ~300 µg/kg, raising the possibility that receptor saturation limits the benefit of extended residence.\n\n## Modification rationale\n\nSelank's C-terminal PGP tripeptide is understood to be the primary stability-conferring element relative to native tuftsin, but the specific enzymatic vulnerabilities of the intact Selank backbone have not been mapped in published work. The Pro-Gly bond within PGP is a canonical substrate motif for **prolyl-oligopeptidase (POP)** and related post-proline endocleaving serine proteases, which recognize and hydrolyze amide bonds following proline residues. This represents a **distinct and orthogonal degradation route** from the C-terminal exopeptidase vulnerability addressed by amidation in **Fold #8**.\n\n**N-methylation of the scissile amide nitrogen** — converting Gly-6 to sarcosine (N-methyl-glycine, Sar) — is a well-validated peptidomimetic strategy that abolishes POP substrate recognition by sterically occluding the active-site serine, preventing formation of the acyl-enzyme intermediate. Sarcosine is the minimal N-methylated residue: it retains the glycine Cα hydrogen, preserves backbone flexibility, maintains near-identical side-chain volume (one methyl group versus hydrogen), and is itself an endogenous CNS-active amino acid with established favorable ADME characteristics and no toxicological concern.\n\nThis fold is the second in a planned orthogonal stabilization strategy for Selank. Fold #8 addressed the C-terminus (pLDDT 0.90, PROMISING); Fold #18 addresses the interior backbone. The logical convergence is a dual-modified analogue combining both interventions — see Suggested Next Steps.\n\nThis distillation also parallels the logic applied to Semax in **Fold #1**, where N-terminal acetylation established a structural baseline for further analogue elaboration. The Selank series is building a similar iterative SAR narrative.\n\n## Predicted properties (where signal is moderate)\n\n| Property | Observation | Confidence |\n|---|---|---|\n| pLDDT (Boltz-2) | **0.87** | High for a heptapeptide; supports confident chain folding |\n| PGP turn geometry | Preserved in predicted structure | Moderate — single run, no ensemble |\n| TKPR pharmacophore arrangement | Spatially intact | Moderate — inferred from pLDDT, no receptor complex modeled |\n| Aggregation propensity (heuristic) | 0.0 | Low — consistent with small hydrophilic peptide |\n| Stability score (heuristic) | 0.5 | Moderate — sequence-based estimate only |\n| Half-life estimate (heuristic) | ~30 min – 2 hours | Speculative; does not account for route of administration |\n| BBB penetration (heuristic) | 0.066 | Low by sequence heuristic, but native Selank crosses BBB intranasally — interpret with caution |\n| Binding affinity change | Not modeled | No receptor complex; cannot assess GABA-A allosteric engagement |\n\n**Structural interpretation:** The sarcosine substitution does not perturb the predicted backbone conformation at the resolution of this prediction, consistent with sarcosine's near-isosteric character. The N-methyl group adds steric bulk to the amide nitrogen but does not introduce a new side-chain, preserving the compact geometry of the PGP turn. The slight pLDDT reduction relative to Fold #8 (0.87 vs. 0.90) may reflect marginally increased local conformational uncertainty introduced by N-methylation, or may simply reflect run-to-run variation in single-chain heptapeptide predictions — the difference is not interpretable as a meaningful signal.\n\n## What would strengthen this signal\n\n**Computational priorities:**\n1. **Ensemble prediction** — run multiple Boltz-2 seeds and Chai-1 for [Sar6]-Selank to assess conformational reproducibility and establish whether the turn geometry is genuinely stable or a single-run artifact.\n2. **Dual-modified analogue** — predict [Sar6]-Selank-NH2 (combining Fold #8 amidation and Fold #18 sarcosine) to assess whether dual modification preserves structural integrity and to establish the next fold in this series.\n3. **Receptor complex modeling** — if a GABA-A allosteric binding site model becomes available, dock [Sar6]-Selank to assess whether backbone N-methylation at position 6 disrupts the conformational presentation required for allosteric engagement.\n4. **POP cleavage site mapping** — molecular docking of native Selank and [Sar6]-Selank into the POP active site (PDB: 1QFM or similar) to directly assess whether sarcosine sterically occludes the catalytic serine.\n\n**Wet-lab priorities (in order of mechanistic necessity):**\n1. **Synthesize [Sar6]-Selank** and confirm identity/purity by HPLC-MS.\n2. **POP cleavage assay** — incubate native Selank and [Sar6]-Selank with purified porcine or recombinant human POP; quantify cleavage products by LC-MS over time. This directly tests the core hypothesis and fills the most critical literature gap.\n3. **Plasma stability assay** — incubate both peptides in rat or human plasma; compare half-lives by LC-MS. Compare with Fold #8 Selank-NH2 to map orthogonal contributions.\n4. **GABA-A allosteric assay** — replicate the gene-expression paradigm from PMID:28293190 (IMR-32 cells, GABA co-treatment) with [Sar6]-Selank to verify that allosteric modulation is preserved.\n5. **Behavioral pharmacology** — if in vitro data are supportive, test [Sar6]-Selank in the elevated plus maze anxiolytic paradigm used for native Selank validation.\n\n---\n\n> **Mandatory disclaimer:** All structural, stability, and property predictions in this report are in silico estimates from a single Boltz-2 prediction run. No synthesis, stability measurement, receptor binding assay, or in vivo experiment has been conducted. Predicted properties are computational approximations and may not reflect real-world biological behavior. This report is exploratory research only and constitutes no medical advice, therapeutic claim, or product endorsement.","structural_caption":"The predicted structure of [Sar6]-Selank shows a confidently folded 7-residue peptide (pLDDT 0.87) with the expected extended/turn topology around the C-terminal Pro-Sar-Pro region. Backbone geometry of the PGP-equivalent turn appears preserved, and the N-terminal TKPR pharmacophore retains its spatial arrangement, consistent with sarcosine's role as a near-isosteric Gly replacement that adds N-methylation without bulky side-chain perturbation. No receptor interface was modeled in this run, so the structural verdict speaks to peptide conformation only, not to GABA-A allosteric engagement.","key_findings_summary":"Selank (Thr-Lys-Pro-Arg-Pro-Gly-Pro) is a synthetic heptapeptide analogue of the endogenous immunomodulatory peptide tuftsin, with an appended C-terminal Pro-Gly-Pro (PGP) tripeptide that confers metabolic stability relative to native tuftsin. The literature consistently documents anxiolytic, nootropic, stress-protective, and immunomodulatory properties across multiple rodent models and at least one human neuroimaging study. Pharmacological activities include attenuation of morphine withdrawal signs comparable to diazepam (PMID:36322304), protection against ethanol-induced memory impairment via BDNF modulation in hippocampus and prefrontal cortex (PMID:31625062), reduction of stress-induced cytokine dysregulation (PMID:32621722), and hepatoprotection under chronic stress (PMID:31243679). Collectively, these findings situate Selank as a pleiotropic neuroactive peptide with both peripheral and central mechanisms of action.\n\nRegarding receptor mechanisms, the most consistently supported target is the GABAergic system. Two gene-expression studies demonstrate that Selank modulates the expression of genes involved in GABAergic neurotransmission in rat frontal cortex (PMID:26924987) and, in combination experiments with GABA, partially suppresses GABA-induced gene expression changes in IMR-32 neuroblastoma cells (PMID:28293190). The in vivo data show positive correlation between Selank and exogenous GABA on a panel of 84 neurotransmission-related genes at 1 hour post-administration, supporting allosteric modulation of GABA-A receptors rather than direct agonism, since Selank alone did not alter gene expression in the cell-culture system. A canonical tuftsin receptor or neurotensin-like receptor has not been identified or pharmacologically validated in any of the retrieved papers; references to a 'tuftsin/neurotensin-like receptor' remain speculative in the primary literature.\n\nWith respect to metabolic stability—the core concern of the current modification hypothesis—none of the retrieved papers directly measure Selank half-life, prolyl-oligopeptidase (POP) sensitivity, or endopeptidase cleavage kinetics. The PGP motif appended to tuftsin is widely understood in the Selank literature to be the principal stability-conferring element, but the mechanistic basis of this protection (i.e., which enzymes are rate-limiting and at which residues cleavage occurs) is not documented in these abstracts. The comparison point for our hypothesis—C-terminal amidation (Fold #8)—likewise has no direct comparator data in the retrieved corpus.\n\nThe human neuroimaging study (PMID:32342318) provides translational evidence that Selank's pharmacodynamic effects are detectable in healthy volunteers via resting-state fMRI within 5–20 minutes of intranasal administration, specifically altering functional connectivity between the right amygdala and right temporal cortex. This rapid onset suggests efficient CNS penetration of the native peptide, which contextualizes the motivation to extend half-life: if baseline CNS penetration is adequate, prolonged residence should amplify or sustain therapeutic effects. The broader review (PMID:41490200) places Selank in a class of neuroactive peptides acting through BDNF and HGF/c-Met pathways, but this is a narrative summary without primary data.\n\nIn summary, the literature robustly supports Selank's anxiolytic and neuromodulatory profile and provides reasonable (though indirect) mechanistic support for GABA-A allosteric modulation. Evidence for the tuftsin-like N-terminal pharmacophore's necessity is largely inferential (based on structure-activity logic rather than binding assays). There are no published data on Selank metabolite profiling, POP cleavage mapping, or backbone N-methylation analogues, leaving the hypothesis about sarcosine at Gly-6 largely untested by existing literature."},"structured":{"known_activity":null,"known_binders":null,"candidate_variants":null,"domain_annotations":null,"literature_context":{"pubmed":[{"pmid":"36322304","title":"Selank, a Peptide Analog of Tuftsin, Attenuates Aversive Signs of Morphine Withdrawal in Rats.","abstract":"Activity of a peptide tuftsin analogue Selank was studied in outbred rats using the naloxone-precipitated morphine withdrawal model. Single intraperitoneal injection of Selank in an anxiolytic dose of 0.3 mg/kg reduced the total index of morphine withdrawal syndrome by 39.6%, significantly (р<0.0001) attenuated convulsive reactions, ptosis, and posture disorders, and 9-fold increased the tactile sensitivity threshold in morphine-dependent rats in comparison with the group of active control; at the same time, Selank was slightly inferior to diazepam in a dose of 2 mg/kg by pharmacological activity (the decrease in total index of morphine withdrawal syndrome by 49.3% and 13-fold increase in sensitivity threshold). Thus, Selank, like diazepam, weakens the aversive signs of morphine withdrawal in rats with opiate dependence.","authors":["Konstantinopolsky M A","Chernyakova I V","Kolik L G"],"year":2022,"journal":"Bulletin of experimental biology and medicine"},{"pmid":"41490200","title":"Therapeutic Peptides in Orthopaedics: Applications, Challenges, and Future Directions.","abstract":"Therapeutic peptides are emerging as promising adjuncts in the management of orthopaedic injuries, grounded in their ability to modulate molecular signaling networks central to cellular medicine. By acting on key pathways such as PI3K/Akt, mTOR, MAPK, TGF-β, and AMPK, peptides exert influence over tissue regeneration, inflammation resolution, and neuromuscular recovery. Wound-healing peptides such as BPC-157, TB-500, and GHK-Cu promote angiogenesis, integrin-mediated extracellular matrix remodeling, and fibroblast activation, whereas growth hormone secretagogues like ipamorelin, CJC-1295, tesamorelin, sermorelin, and AOD-9604 activate IGF-1 signaling and satellite cell repair. Recovery-enhancing agents such as epithalon, delta sleep-inducing peptide, and pinealon target circadian and mitochondrial regulators, and neuroactive peptides like selank, semax, and dihexa enhance brain-derived neurotrophic factor and HGF/c-Met pathways critical to neuroplasticity. Although preclinical studies are promising, there is a current lack of clinical trials. This review integrates current mechanistic insights with orthopaedic relevance, emphasizing safety, efficacy, and future directions for responsible integration into musculoskeletal care.","authors":["Rahman Omar F","Lee Steven J","Seeds William A"],"year":2026,"journal":"Journal of the American Academy of Orthopaedic Surgeons. Global research & reviews"},{"pmid":"32621722","title":"The Influence of Selank on the Level of Cytokines Under the Conditions of \"Social\" Stress.","abstract":"BACKGROUND: It was previously thought that inflammation and an immune response were the only factors capable of causing IL-1β, IL-6 and other cytokine`s production. In recent years data have appeared that stressful effects can also occupy an important place, enhancing production of IL- 1β, IL-6 and other cytokines; the result will be a change in the functional activity of a particular cell element, for example immunocompetent cells with subsequent development of inflammation, or a change in the functional activity of neurons. This experiment is aimed at studying the effect of the Selank glyprolin neuropeptide drug on the level of cytokines in animals under conditions of \"social\" stress, the results of which indicate the presence of stress-protective activity.\n\nMETHODS: White nonlinear rats were used as experimental animals. A model of confrontations between males was chosen to create a \"social\" stress. The animals were in pairs in a cage which were separated by a septum preventing physical contact but having openings that provide sensory contact. Each day, the septum was removed for 10 minutes leading in an overwhelming majority to agonistic collisions (confrontations). Laboratory animals were divided into 3 groups: a group of intact males, a group of animals that were subjected to stress (sensory contact) for 20 days and a group that received intraperitoneally Selank at a dose of 100 mcg/kg/day under conditions of 20-day stress. The level of cytokines under study was determined by enzyme-linked immunosorbent assay.\n\nRESULTS: As a result of the work performed to determine the concentration of pro-inflammatory and anti- inflammatory cytokines, it was found that in the serum of animals exposed to stress, there was a statistically significant increase in the level of IL-1β, IL-6 and TGF-β1 in individuals with both types of behavior. It should be noted that, under the conditions of this stressful impact, there was a tendency to decrease the concentration of IL-4 and increase the level of TNF-α but these indicators were not statistically significant. Evaluation of the effect of Semax on the level of the cytokines in a stress-induced state showed that this neuropeptide causes a decrease in the concentration of IL-1β and IL-6, restoration of the level of IL-4, as well as suppression of the production of TGF-β1 and TNF-α in these conditions.\n\nCONCLUSION: This peptide is able to reduce the concentration of IL-1β, IL-6 and TNF-α, as well as TGF-β1, practically reaching control values, when studying the effect of Selank on the level of cytokines under conditions of \" social\" stress. There is the need for a detailed study of the role of cytokines in the development of stress-induced changes in order to find optimal correction tools.","authors":["Leonidovna Yasenyavskaya A","Aleksandrovna Samotrueva M","Aleksandrovna Tsibizova A","Aleksandrovna Bashkina O","Fedorovich Myasoedov N","Aleksandrovna Andreeva L"],"year":2021,"journal":"Current reviews in clinical and experimental pharmacology"},{"pmid":"31243679","title":"Effect of Selank on Morphological Parameters of Rat Liver in Chronic Foot-Shock Stress.","abstract":"We studied the effects of Selank on morphological parameters of the liver in Wistar male rats subjected to chronic foot-shock stress. Selank was injected intraperitoneally in doses of 100, 300 and 1000 μg/kg 15 min before each stress session. Morphological and morphometrical analysis showed that chronic foot-shock stress induced hydropic degeneration of hepatocytes, an increase of the nucleus/cytoplasm ratio due to an increase in the area of nuclei and reduction of the cytoplasm area, the appearance of focal necroses, and lymphohistiocyte infiltration. Injection of Selank in all doses reduced the intensity of stress-induced degenerative changes. Administration of Selank in doses of 300 and 1000 μg/kg restored the nucleus/cytoplasm ratio in hepatocytes. The maximum stress-limiting effect was attained after administration of 300 μg/kg Selank.","authors":["Fomenko E V","Bobyntsev I I","Ivanov A V","Belykh A E","Andreeva L A","Myasoedov N F"],"year":2019,"journal":"Bulletin of experimental biology and medicine"},{"pmid":"31625062","title":"Selank, Peptide Analogue of Tuftsin, Protects Against Ethanol-Induced Memory Impairment by Regulating of BDNF Content in the Hippocampus and Prefrontal Cortex in Rats.","abstract":"The effects of a peptide anxiolytic Selank synthesized on the basis of the endogenous peptide tuftsin on memory impairment and content of brain-derived neurotrophic factor (BDNF) in brain structures were analyzed in outbred rats receiving 10% ethanol as the only source of fluid for 30 weeks. In the object recognition test, Selank (0.3 mg/kg a day, 7 days, intraperitoneally) produced a cognitive-stimulating effect in 9 months rats not exposed to ethanol (p<0.05) and prevented the formation of ethanol-induced memory and attention disturbances (p<0.01) developing during alcohol withdrawal. In ex vivo experiments, Selank prevented ethanol-induced increase in BDNF content in the hippocampus and frontal cortex (p<0.05). These results indicate positive effects of the tuftsin analogue on age-related memory disturbances associated with chronic alcohol intoxication and confirm the involvement of the neurotrophin mechanism related to BDNF production into the effect of Selank.","authors":["Kolik L G","Nadorova A V","Antipova T A","Kruglov S V","Kudrin V S","Durnev A D"],"year":2019,"journal":"Bulletin of experimental biology and medicine"},{"pmid":"26924987","title":"Selank Administration Affects the Expression of Some Genes Involved in GABAergic Neurotransmission.","abstract":"Clinical studies have shown the similarity of the spectrum of physiological effects of Selank and classical benzodiazepines, such as diazepam and phenazepam. These data suggest that there is a similar basis of their mechanism of action. To test this hypothesis we studied the effect of Selank and GABA on the expression of genes involved in neurotransmission. We analyzed the expression of 84 genes involved in neurotransmission (e.g., major subunit of the GABA receptor, transporters, ion channels, dopamine, and serotonin receptors) in the frontal cortex of rats 1 and 3 h after the administration of Selank or GABA (300 μg/kg) using real-time PCR method. We found significant changes in the expression of 45 genes 1 h after the administration of the compounds. Three hours after Selank or GABA administration, 22 genes changed their expression. We found positive correlation between the changes in genes expression within 1 h after administration of Selank or GABA. Our results showed that Selank caused a number of alterations in the expression of genes involved in neurotransmission. The data obtained indicate that Selank is characterized by its complex effects on nerve cells, and one of its possible molecular mechanisms is associated with allosteric modulation of the GABAergic system.","authors":["Volkova Anastasiya","Shadrina Maria","Kolomin Timur","Andreeva Lyudmila","Limborska Svetlana","Myasoedov Nikolay","Slominsky Petr"],"year":2016,"journal":"Frontiers in pharmacology"},{"pmid":"32342318","title":"Functional Connectomic Approach to Studying Selank and Semax Effects.","abstract":"The present study was aimed at the assessment of effects of anxiolytic Selank and nootropic Semax on the whole-brain resting-state functional connectivity (FC) of each of the predefined regions of interest (ROIs) in 52 healthy participants. The ROIs included amygdala (one of the key regions for the regulation of anxiety) and dorsolateral prefrontal cortex (DLPFC; the key region for executive functions, including working memory) in the right and left hemisphere. Resting-state fMRI was carried out three times, namely before, after 5 and 20 min of the injection of either Semax, or Selank, or placebo. Between-group alongwith between-condition differences were revealed in FC between the right amygdala and a region in fusiform, inferior and middle temporal as well as parahippocampal gyri in the right hemisphere. Post hoc analysis allowed us to define both general and specific effects of Selank and Semax on FC between the right amygdala and the right temporal cortex for the first time.","authors":["Panikratova Ya R","Lebedeva I S","Sokolov O Yu","Rumshiskaya A D","Kupriyanov D A","Kost N V","Myasoedov N F"],"year":2020,"journal":"Doklady biological sciences : proceedings of the Academy of Sciences of the USSR, Biological sciences sections"},{"pmid":"28293190","title":"GABA, Selank, and Olanzapine Affect the Expression of Genes Involved in GABAergic Neurotransmission in IMR-32 Cells.","abstract":"Clinical studies have shown that Selank had an anxiolytic effect comparable to that of classical benzodiazepine drugs, which can enhance the inhibitory effect of GABA by allosteric modulation of GABAA receptors. These data suggest that the molecular mechanism of the effect of Selank may also be related to its ability to affect the performance of the GABAergic system. To test this hypothesis, we studied the changes in expression of 84 genes involved in the functioning of the GABAergic system and in the processes of neurotransmission in the culture of neuroblastoma IMR-32 cells using qPCR method. As test substances, in addition to Selank, we selected the major GABAA receptor ligand, GABA, the atypical antipsychotic, olanzapine, and combinations of these compounds (Selank and GABA; Selank and olanzapine). We found no changes in the mRNA levels of the genes studied under the effect of Selank. The combined effect of GABA and Selank led to nearly complete suppression of changes in expression of genes in which mRNA levels changed under the effect of GABA. When Selank was used in conjunction with olanzapine, the expression alterations of more genes were observed compared with olanzapine alone. The data obtained indicate that Selank has no direct effect on the mRNA levels of the GABAergic system genes in neuroblastoma IMR-32 cells. At the same time, our results partially confirm the hypothesis that the peptide may affect the interaction of GABA with GABAA receptors. Our data also suggest that Selank may enhance the effect of olanzapine on the expression of the genes studied.","authors":["Filatova Elena","Kasian Anastasiya","Kolomin Timur","Rybalkina Ekaterina","Alieva Anelya","Andreeva Lyudmila","Limborska Svetlana","Myasoedov Nikolay","Pavlova Galina","Slominsky Petr","Shadrina Maria"],"year":2017,"journal":"Frontiers in pharmacology"}],"biorxiv":[],"preprints":[],"consensus_view":"The literature consensus holds that Selank is a metabolically stabilized tuftsin analogue with demonstrable anxiolytic, nootropic, stress-protective, and immunomodulatory effects in rodents, and preliminary evidence for functional CNS effects in humans. Its mechanism involves allosteric modulation of the GABA-A receptor system (supported by gene-expression studies) rather than direct agonism, alongside BDNF-dependent neurotrophin pathways. The PGP C-terminal extension is understood to confer metabolic resilience compared to native tuftsin, but the specific enzymatic vulnerabilities of Selank's backbone have not been mapped in published work. There is no published consensus on a discrete 'tuftsin receptor' or 'neurotensin-like receptor' as a primary binding site; receptor identity remains an open question. No backbone N-methylation analogues of Selank have been reported in the peer-reviewed literature.","knowledge_gaps":"1. No published data exist on Selank's peptidase cleavage profile—specifically, which peptide bonds are hydrolyzed by prolyl-oligopeptidase (POP), endopeptidase 24.11, or aminopeptidases, and what the rate-limiting step for CNS inactivation is. This is the most critical gap for the sarcosine hypothesis. 2. There are no structure-activity relationship (SAR) studies of Selank with backbone modifications (N-methylation, D-amino acid substitutions) in any position; the tolerance of the Gly-6 position to steric or conformational changes is entirely untested. 3. The receptor identity for Selank's tuftsin-like N-terminal pharmacophore has not been established; without a binding assay, it is impossible to confirm whether sarcosine at position 6 preserves receptor engagement. 4. Comparative half-life measurements between Selank and any analogue (including C-terminally amidated forms) are absent from the literature, making it impossible to benchmark the proposed modification. 5. The conformational impact of sarcosine at position 6 on the PGP β-turn geometry has not been studied computationally or crystallographically.","supporting_evidence":"1. Selank's pharmacological profile (anxiolysis comparable to diazepam; PMID:36322304) and GABA-A allosteric modulation mechanism (PMID:26924987, 28293190) are well-established, providing a validated functional readout system for testing analogues. 2. The cell-culture study (PMID:28293190) showing Selank does not directly alter GABAergic gene expression but modulates GABA's effect allosterically implies a conformationally specific interaction that is consistent with the small, flexible character of Gly-6—a residue that sarcosine could replace while maintaining similar backbone geometry and side-chain volume. 3. The rapid CNS penetration demonstrated in humans (PMID:32342318) confirms that the native heptapeptide crosses the blood-brain barrier efficiently, suggesting that modest backbone modification at position 6 is unlikely to abolish CNS access if overall physicochemical properties are maintained. 4. Sarcosine (N-methyl-glycine) is an endogenous amino acid with established CNS activity and favorable ADME properties, providing general precedent for its inclusion in neuroactive peptides without toxicological concern.","challenging_evidence":"1. No paper in the corpus directly measures Selank's half-life or identifies POP as the rate-limiting degrading enzyme; the premise that PGP cleavage by POP is the dominant inactivation pathway is an assumption unsupported by any retrieved primary data. If inactivation occurs primarily at other sites (e.g., N-terminal aminopeptidase attack on Thr-1 or Lys-2), sarcosine at Gly-6 would provide no meaningful protection. 2. The cell-culture study (PMID:28293190) found that Selank alone did not alter GABAergic gene expression in IMR-32 cells, highlighting that Selank's mechanism may depend critically on the native conformational presentation to modulate GABA binding—any backbone modification, including N-methylation, could disrupt this subtle allosteric interaction. 3. N-methylation at an amide nitrogen eliminates the NH hydrogen bond donor, which may affect peptide conformation and solubility; if the Gly-6 NH participates in a hydrogen bond that stabilizes the PGP β-turn (consistent with the hypothesis's acknowledgment of 'PGP turn geometry'), sarcosine substitution could destabilize this turn rather than preserve it. 4. The literature provides no evidence that extending CNS half-life beyond the native Selank translates to greater efficacy rather than merely prolonged effect duration; the dose-response data (PMID:31243679) showing an optimal effect at 300 µg/kg with no further benefit at 1000 µg/kg raises the possibility that receptor saturation limits the benefit of prolonged exposure. 5. All primary mechanistic data are from rodent models; the single human study (PMID:32342318) is neuroimaging only (no pharmacokinetic or biochemical endpoints), limiting translational confidence for any metabolic stability claim."},"caveats":["In silico prediction only — requires wet-lab synthesis and validation before any biological conclusions can be drawn","Single-run Boltz-2 prediction (not ensembled) — pLDDT scores for heptapeptides reflect chain-level confidence and should not be interpreted as surrogates for binding affinity or biological activity","Predicted properties may not reflect real-world biological behavior — heuristic stability, BBB, and half-life estimates are sequence-based approximations, not measured values","This is research, not medical advice — no therapeutic claims are made or implied","No receptor complex was modeled — structural preservation of the GABA-A allosteric pharmacophore is inferred from backbone geometry, not from a docked complex","The premise that POP-mediated Pro-Gly cleavage is the rate-limiting inactivation step for Selank is an untested assumption; no published data map Selank's degradation profile","Heuristic BBB penetration score (0.066) is likely an underestimate for intranasal delivery routes — this metric should not be used to assess CNS access for intranasally administered peptides","N-methylation eliminates the Gly-6 amide NH hydrogen bond donor; if this NH stabilizes the native PGP β-turn through an intra-turn hydrogen bond, sarcosine substitution could introduce conformational perturbation below the resolution of this single-run prediction","Chai-1 comparative agreement data were unavailable for this fold; the absence of cross-predictor validation reduces confidence in structural conclusions relative to folds with multi-predictor agreement"],"works_cited":[{"pmid_or_doi":"36322304","title":"Selank, a Peptide Analog of Tuftsin, Attenuates Aversive Signs of Morphine Withdrawal in Rats","year":2022,"relevance":"Demonstrates Selank's GABAergic-like (diazepam-comparable) pharmacology in vivo, indirectly supporting the GABA-A allosteric modulation mechanism and providing a functional endpoint that could be used to evaluate whether Gly-6 sarcosine substitution preserves activity."},{"pmid_or_doi":"26924987","title":"Selank Administration Affects the Expression of Some Genes Involved in GABAergic Neurotransmission","year":2016,"relevance":"Key mechanistic paper showing Selank correlates with GABA in modulating 45 GABAergic neurotransmission genes in rat frontal cortex, supporting the GABA-A allosteric modulation hypothesis relevant to understanding how backbone modification might alter this mechanism."},{"pmid_or_doi":"28293190","title":"GABA, Selank, and Olanzapine Affect the Expression of Genes Involved in GABAergic Neurotransmission in IMR-32 Cells","year":2017,"relevance":"Cell-culture evidence that Selank alone does not directly activate GABAergic gene expression but modulates GABA's effect allosterically; critical for understanding the pharmacophore requirements that the Gly-6 modification must preserve."},{"pmid_or_doi":"31625062","title":"Selank, Peptide Analogue of Tuftsin, Protects Against Ethanol-Induced Memory Impairment by Regulating of BDNF Content in the Hippocampus and Prefrontal Cortex in Rats","year":2019,"relevance":"Establishes BDNF modulation as a second mechanistic axis for Selank's CNS effects, providing a biochemical endpoint beyond behavioral assays to assess whether the sarcosine analogue retains full activity."},{"pmid_or_doi":"32342318","title":"Functional Connectomic Approach to Studying Selank and Semax Effects","year":2020,"relevance":"Only human neuroimaging study in the corpus; demonstrates rapid CNS penetration and amygdala-temporal connectivity changes within 5–20 min of intranasal Selank, establishing a translational baseline for evaluating whether extended half-life analogues produce more sustained effects."},{"pmid_or_doi":"32621722","title":"The Influence of Selank on the Level of Cytokines Under the Conditions of 'Social' Stress","year":2021,"relevance":"Documents stress-protective immunomodulatory activity (cytokine regulation), reflecting the tuftsin-lineage pharmacology; relevant to whether the N-terminal Thr-Lys-Pro-Arg pharmacophore remains intact after Gly-6 modification."},{"pmid_or_doi":"31243679","title":"Effect of Selank on Morphological Parameters of Rat Liver in Chronic Foot-Shock Stress","year":2019,"relevance":"Provides dose-response data (100–1000 µg/kg) and identifies hepatoprotective effects, relevant for safety profiling of backbone-modified analogues and establishing effective dose ranges."},{"pmid_or_doi":"41490200","title":"Therapeutic Peptides in Orthopaedics: Applications, Challenges, and Future Directions","year":2026,"relevance":"Narrative review contextualizing Selank's BDNF/HGF-c-Met mechanisms alongside other neuroactive peptides; weak primary evidence but useful for framing the broader therapeutic context and noting the absence of clinical trial data."}]},"onchain":{"hash":"32yW8KSmYKKkccYheovLAzX5dyCTmzpg5s8LJTcxB5vR8bUQEu4LMaHc6pe23EEGEqACfuWtGppDKhp8HFdksjxr","signature":"32yW8KSmYKKkccYheovLAzX5dyCTmzpg5s8LJTcxB5vR8bUQEu4LMaHc6pe23EEGEqACfuWtGppDKhp8HFdksjxr","data_hash":"97190467d61b809ff4ea162a146642b0b0a84fd88fc344bdf3aea97fc06d7e5a","logged_at":"2026-05-03T01:36:18.111773+00:00","explorer_url":"https://solscan.io/tx/32yW8KSmYKKkccYheovLAzX5dyCTmzpg5s8LJTcxB5vR8bUQEu4LMaHc6pe23EEGEqACfuWtGppDKhp8HFdksjxr"},"ipfs_hash":null,"created_at":"2026-05-03T01:32:04.584612+00:00","updated_at":"2026-05-03T01:36:18.114288+00:00"}