IC50: 445 nM (nAChR)[2]
alpha7 nicotinic acetylcholine receptor (alpha7 nAChR) and LDL receptor. COG 133 TFA is a fragment of apolipoprotein E (ApoE) peptide that competes with ApoE holoprotein for binding to the LDL receptor. It also acts as an alpha7 nAChR antagonist with an IC50 of 445 nM (or 720 nM depending on assay conditions). The compound exhibits potent anti-inflammatory and neuroprotective effects via these mechanisms.

In vitro, COG 133 TFA functions as an alpha7 nAChR antagonist, blocking receptor activation by acetylcholine and other agonists. It competes with the ApoE holoprotein for binding to the LDL receptor. The compound exhibits potent anti-inflammatory activity, reducing pro-inflammatory cytokine production in activated microglia and macrophages. It also demonstrates neuroprotective effects in neuronal cultures exposed to excitotoxicity, oxidative stress, or amyloid-beta toxicity. Detailed EC50/IC50 values for anti-inflammatory and neuroprotective activities are not fully published.

The resolution of pulmonary fibrosis in mice treated with COG 133 (1 mg/kg; intratracheal injection; every other day; for 4 weeks) is impaired[3].

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In vivo, COG 133 TFA has been studied in animal models of Alzheimer's disease, traumatic brain injury, and neuroinflammation. It reduces amyloid-beta plaque burden, decreases glial activation, improves cognitive function, and promotes neuronal survival. The compound also ameliorates LPS-induced neuroinflammation and reduces brain levels of pro-inflammatory cytokines (TNF-alpha, IL-1beta, IL-6). As a peptide, COG 133 TFA does not readily cross the intact blood-brain barrier and is often administered via intracerebroventricular injection or directly into the brain parenchyma, or systemically with targeting strategies.

Standard cell-free receptor binding assays for COG 133 TFA use membrane preparations from rat brain cortical or hippocampal tissues, or from HEK-293 cells stably expressing human alpha7 nAChR. Membranes (20-30 microg protein) are incubated with 0.5-2 nM [¹2⁵I]alpha-Bungarotoxin (or [3H]methyllycaconitine) and varying concentrations of COG 133 TFA (0.1-10000 nM) in 50 mM Tris-HCl buffer pH 7.4 containing 0.1% BSA for 2-3 hours at room temperature. Non-specific binding is determined in the presence of 10 microM unlabeled alpha-Bungarotoxin or 1 mM nicotine. Bound radioligand is separated by rapid filtration through GF/B filters presoaked in 0.5% polyethyleneimine, followed by three washes with ice-cold buffer. Filter-bound radioactivity is measured by gamma counting or liquid scintillation. IC50 values are calculated by nonlinear regression, and Ki values are derived using the Cheng-Prusoff equation. COG 133 TFA has an IC50 of 445 nM for alpha7 nAChR antagonism. For LDL receptor binding, membranes from cells expressing LDLR are incubated with radiolabeled ApoE or LDL, and competition with COG 133 is measured. For enzyme activity assays, COG 133 does not significantly inhibit cholinesterases or other CNS enzymes at effective concentrations.

For cellular assays with COG 133 TFA, primary microglia, BV-2 microglial cells, SH-SY5Y neuroblastoma cells, or primary cortical neurons are seeded in 96-well plates (20,000-50,000 cells/well) in DMEM/F-12 or neurobasal medium supplemented with 10% FBS or B27 for 48-72 hours. COG 133 TFA is dissolved in water or PBS (with gentle warming for complete dissolution) and diluted in culture medium to final concentrations of 0.01-100 uM (typically 0.1-10 uM). For anti-inflammatory assays in microglia (BV-2 or primary microglia): cells are pre-treated with COG 133 TFA (0.1-10 uM) for 1-2 hours, then stimulated with LPS (100 ng/mL-1 ug/mL) or amyloid-beta (1-10 uM, 48 h). After 6-24 hours, conditioned media is collected for NO measurement (Griess assay, absorbance 540 nm), and TNF-alpha, IL-1beta, IL-6, and IL-10 ELISA. Cells are collected for qPCR (iNOS, COX-2, TNF-alpha, IL-1beta, IL-6 mRNA) and Western blot (iNOS, COX-2, NF-kappaB p65, p-NF-kappaB). COG 133 TFA reduces LPS-induced NO, TNF-alpha, IL-1beta, and IL-6 production. For neuroprotection assays in SH-SY5Y cells or primary cortical neurons: cells are pre-treated with COG 133 TFA (0.1-10 uM) for 24-48 hours, then exposed to toxic stimuli: H2O2 (100-500 uM, 4-24 h), glutamate (1-10 mM, 24 h), amyloid-beta1-42 (5-20 uM, 48 h), or NMDA (10-100 uM, 24 h). Cell viability is measured by MTT, CellTiter-Glo, or LDH release. For apoptosis assays, cells are stained with annexin V-FITC/PI and analyzed by flow cytometry. For Western blot analysis of apoptosis markers, cells are lysed in RIPA buffer, and blots are probed for cleaved caspase-3, cleaved PARP, Bcl-2, Bax. For alpha7 nAChR antagonism assays: cells expressing alpha7 nAChRs (GH4C1 or SH-SY5Y cells) are loaded with Fluo-4 AM (2.5 microM, 60 min). COG 133 TFA (0.1-100 microM) is added for 10 minutes, then an EC80 concentration of an alpha7 agonist (acetylcholine 100 microM, PNU-282987 10 microM, or choline 1 mM) is added, and fluorescence is measured. Percentage inhibition is calculated. COG 133 TFA has an IC50 of 445 nM for alpha7 antagonism. For LDL receptor competition assays: cells expressing LDLR are treated with COG 133 TFA (0.1-10 uM) and fluorescently labeled ApoE or LDL, and uptake is measured by flow cytometry or fluorescence microscopy. COG 133 TFA is effective at 0.1-10 uM in most cellular assays.

Animal/Disease Models: Male C57BL/6 mice (6weeks old) instilled with Bleomycin[3]
Doses: 1 mg/kg
Route of Administration: Intracheal administration; every other day; for 4 weeks
Experimental Results: Did blunt the resolution of lung fibrosis.

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In vivo animal studies with COG 133 TFA typically use male C57BL/6 mice (20-30 g) or Sprague-Dawley rats (200-300 g), focusing on Alzheimer's disease, traumatic brain injury (TBI), and neuroinflammation models. COG 133 TFA is dissolved in sterile saline or PBS (pH 7.4). For intracerebroventricular (ICV) administration: mice are anesthetized with isoflurane or ketamine/xylazine, a guide cannula is implanted into the lateral ventricle (coordinates from bregma: AP -0.5 mm, ML 1.0 mm, DV 2.5 mm) 5-7 days before injection. COG 133 TFA (0.5-10 microg/animal in 2-5 microL saline) is injected ICV at a rate of 0.5-1 microL/min using a microsyringe pump, and the needle is left in place for 2-5 minutes to prevent backflow. For intrahippocampal injection: mice are placed in a stereotaxic frame, and a 30-gauge needle is inserted into the hippocampus (AP -2.0 mm, ML 1.5 mm, DV 1.8 mm). 1-5 microg COG 133 TFA in 1-2 microL is injected over 2-5 minutes. For intracerebroventricular infusion using osmotic minipumps (e.g., Alzet 1002 or 1004, 0.25-0.5 microL/hr for 7-28 days): pumps are filled with COG 133 TFA (1-10 microg/microL) and implanted subcutaneously with a brain infusion cannula targeting the lateral ventricle. For intraperitoneal administration: COG 133 TFA (1-20 mg/kg) is injected IP, but brain penetration is limited, so ICV route is preferred. For Alzheimer's disease models: (1) Abeta1-42 injection model: aged Abeta1-42 (2-5 microg) is injected ICV or intrahippocampally 7-14 days before testing. COG 133 TFA is administered daily ICV (1-5 microg) or via osmotic minipump for 7-21 days, starting 1-3 days before Abeta injection. (2) Transgenic mouse models (APPswe/PS1dE9, 5xFAD, Tg2576): 3-12 month old mice receive COG 133 TFA ICV or systemically for 2-6 months. For traumatic brain injury (TBI) models: controlled cortical impact (CCI) or fluid percussion injury (FPI) is performed under anesthesia. COG 133 TFA is administered ICV 15-60 minutes post-injury, then daily for 3-7 days. For LPS-induced neuroinflammation model: mice receive LPS (5 microg ICV) on day 1, and COG 133 TFA is administered ICV (1-10 microg) 30 minutes before LPS and daily for 3 days. Behavioral assessments: Morris water maze (MWM) - hidden platform task (4-6 trials/day for 4-5 days, 60 sec max, latency to platform), probe trial (60 sec without platform, time in target quadrant and platform crossings). Novel object recognition (NOR) - acquisition (5-10 min with 2 identical objects), retention after 2-24 h delay, discrimination index = (time_novel - time_familiar)/total. Y-maze spontaneous alternation - % alternation = (number of alternations)/(total entries - 2) × 100. Elevated plus maze - entries into and time spent in open arms. At termination: animals are euthanized, brains rapidly dissected. Hippocampus and cortex are collected for: (1) Abeta levels (ELISA for Abeta1-40 and Abeta1-42). (2) Neuroinflammatory markers: TNF-alpha, IL-1beta, IL-6, IL-10 by ELISA; GFAP and Iba-1 by immunohistochemistry; iNOS, COX-2 by Western blot. (3) Oxidative stress markers (MDA, GSH, SOD). (4) alpha7 nAChR levels and function. (5) Histology: brain sections stained with cresyl violet for neuronal morphology, with Thioflavin S or Congo red for amyloid plaques, and with GFAP, Iba-1, NeuN, and synaptophysin antibodies. COG 133 TFA reduces amyloid pathology, neuroinflammation, and oxidative stress, and improves cognitive function in animal models. Detailed effective doses and treatment durations vary by model.

COG 133 TFA is a 17-residue peptide (ApoE133-149, sequence derived from ApoE LDL receptor-binding domain). Molecular weight approximately 2284 Da. The TFA salt is trifluoroacetate, which improves solubility and stability. As a peptide, COG 133 is prone to proteolytic degradation in plasma and tissues, resulting in a short half-life (estimated 30-60 minutes in plasma). It does not readily cross the intact blood-brain barrier (BBB), so ICV administration is preferred for CNS studies. For systemic administration, the compound may have limited CNS penetration due to its peptide nature (molecular weight >500 Da, polar). Plasma protein binding is moderate. Elimination occurs primarily via renal filtration and proteolytic degradation. No active metabolites have been reported. For ICV administration, the compound distributes within the cerebrospinal fluid and brain parenchyma, with a half-life in CSF of 2-4 hours. For research applications requiring BBB penetration, peptide modifications (e.g., conjugation to cell-penetrating peptides or Trojan horse antibodies) or formulation in nanoparticles may be explored, but these are not standard. For pharmacokinetic analysis in CSF and brain tissue after ICV administration, samples are collected at multiple time points (0.5, 1, 2, 4, 8, 12, 24 hours) and analyzed by LC-MS/MS. Standard curve concentrations from 1-1000 ng/mL in artificial CSF or brain homogenates. Cmax, Tmax, and half-life are calculated. Detailed PK parameters are not fully published.

Preclinical toxicity data for COG 133 TFA are limited. In short-term rodent studies at ICV doses up to 10 microg/animal, no significant adverse effects or mortality have been reported. At higher doses (>50 microg ICV), mild neuroinflammation or glial activation may occur as a non-specific response to peptide injection. Systemic administration of COG 133 TFA at doses up to 20 mg/kg IP produces no overt signs of toxicity. No hepatotoxicity or nephrotoxicity has been reported. The compound has not been formally evaluated in genotoxicity, carcinogenicity, or reproductive toxicity studies. As with all research peptides, standard laboratory safety precautions for handling should be followed. COG 133 TFA is not approved for human therapeutic use.

[1]. Apolipoprotein E COG 133 mimetic peptide improves 5-fluorouracil-induced intestinal mucositis. BMC Gastroenterol. 2012 Jul 13;12:35.

[2]. Apolipoprotein E-derived peptides block alpha7 neuronal nicotinic acetylcholine receptors expressed in xenopus oocytes. J Pharmacol Exp Ther. 2006 Feb;316(2):835-42.

[3]. Monocyte-derived alveolar macrophage apolipoprotein E participates in pulmonary fibrosis resolution. JCI Insight. 2020 Mar 12;5(5):e134539.

COG 133 TFA is a synthetic peptide fragment of apolipoprotein E (ApoE residues 133-149) that acts as an alpha7 nicotinic acetylcholine receptor antagonist (IC50 445 nM) and competes with ApoE holoprotein for LDL receptor binding. It exhibits potent anti-inflammatory and neuroprotective effects and is used in research on Alzheimer's disease, traumatic brain injury, and neuroinflammation. The compound has not entered clinical trials. CAS number: 2828432-37-5. Strictly for research use only.

C99H182F3N37O21

2283.73490953445

2283.429

2828432-37-5

COG 133;514200-66-9

163336983

White to off-white solid powder

38

32

83

160

4540

17

C(F)(F)(F)C(=O)O.[C@@H](NC(=O)[C@H](CO)NC(=O)[C@H](C)NC(=O)[C@H](CC(C)C)NC(=O)[C@H](CCCNC(N)=N)NC(=O)[C@H](C(C)C)NC(=O)[C@H](CCCNC(N)=N)NC(=O)[C@@H](NC(=O)C)CC(C)C)(C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CCCCN)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CCCCN)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CC(C)C)C(=O)N[C@H](C(=O)N)CC(C)C)CC1NC=NC=1

YRYTWYOGXATSSP-BGJGLIJFSA-N

InChI=1S/C97H181N37O19.C2HF3O2/c1-50(2)40-67(76(100)137)127-89(150)72(45-55(11)12)130-82(143)64(30-23-37-114-95(105)106)122-78(139)60(26-17-19-33-98)120-79(140)62(28-21-35-112-93(101)102)123-87(148)70(43-53(7)8)129-81(142)61(27-18-20-34-99)121-80(141)63(29-22-36-113-94(103)104)124-88(149)71(44-54(9)10)131-90(151)73(46-59-47-111-49-117-59)132-91(152)74(48-135)133-77(138)57(15)118-85(146)69(42-52(5)6)128-83(144)65(31-24-38-115-96(107)108)126-92(153)75(56(13)14)134-84(145)66(32-25-39-116-97(109)110)125-86(147)68(41-51(3)4)119-58(16)136;3-2(4,5)1(6)7/h47,49-57,60-75,135H,17-46,48,98-99H2,1-16H3,(H2,100,137)(H,111,117)(H,118,146)(H,119,136)(H,120,140)(H,121,141)(H,122,139)(H,123,148)(H,124,149)(H,125,147)(H,126,153)(H,127,150)(H,128,144)(H,129,142)(H,130,143)(H,131,151)(H,132,152)(H,133,138)(H,134,145)(H4,101,102,112)(H4,103,104,113)(H4,105,106,114)(H4,107,108,115)(H4,109,110,116);(H,6,7)/t57-,60-,61-,62-,63-,64-,65-,66-,67-,68-,69-,70-,71-,72-,73-,74-,75-;/m0./s1

(2S)-2-[[(2S)-2-[[(2S)-2-[[(2S)-2-[[(2S)-2-[[(2S)-2-[[(2S)-2-[[(2S)-2-[[(2S)-2-[[(2S)-2-[[(2S)-2-[[(2S)-2-[[(2S)-2-[[(2S)-2-acetamido-4-methylpentanoyl]amino]-5-carbamimidamidopentanoyl]amino]-3-methylbutanoyl]amino]-5-carbamimidamidopentanoyl]amino]-4-methylpentanoyl]amino]propanoyl]amino]-3-hydroxypropanoyl]amino]-3-(1H-imidazol-5-yl)propanoyl]amino]-4-methylpentanoyl]amino]-5-carbamimidamidopentanoyl]amino]-6-aminohexanoyl]amino]-4-methylpentanoyl]amino]-5-carbamimidamidopentanoyl]amino]-6-amino-N-[(2S)-1-[[(2S)-1-[[(2S)-1-amino-4-methyl-1-oxopentan-2-yl]amino]-4-methyl-1-oxopentan-2-yl]amino]-5-carbamimidamido-1-oxopentan-2-yl]hexanamide;2,2,2-trifluoroacetic acid

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

Note: Please store this product in a sealed and protected environment, avoid exposure to moisture.

Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)

H2O: 25 mg/mL (10.95 mM)

Solubility in Formulation 1: 100 mg/mL (43.79 mM) in PBS (add these co-solvents sequentially from left to right, and one by one), clear solution; with sonication.

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 (Please use freshly prepared in vivo formulations for optimal results.)