| Size | Price | Stock | Qty |
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| 5mg |
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| 10mg |
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| 50mg |
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| 100mg |
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| 250mg | |||
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| Targets |
Procaspase-3 (interacts directly, inducing autoprocessing and activation). [1]
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| ln Vitro |
Stretch-induced IKK activation and IL-6 mRNA expression were eliminated by RGD peptide (GRGDNP) (50 μM; preincubation for 3 h before stretching). In unstretched HUVEC, it has little influence on the expression of IL-6 mRNA and IKK activity [2]. Oxygen glucose deprivation (OGD)-treated primary hippocampal neurons, HT22 cell line, and their sEVs totally reverse increased FN1 expression when exposed to RGD peptide (GRGDNP) at 300 μg/mL for six hours [3].
The peptide GRGDNP induced rapid, dose-dependent apoptosis in resting peripheral blood T lymphocytes. The half-maximal effect (EC50) was observed at a concentration of 250 μM. Apoptosis was confirmed using standard methods. [1] GRGDNP also induced apoptosis in a variety of other leukocyte cell types, including CD4+ T-cell lines, leukemic T-cell lines (Jurkat, Molt-4), Epstein-Barr virus-transformed B cell lines (LCL), and the erythroleukaemic cell line K562, indicating a broad pro-apoptotic effect. [1] The pro-apoptotic effect of GRGDNP was independent of integrin-mediated cell clustering or signaling, as demonstrated by experiments using peptide-coated beads or cells cultured on integrin ligand-coated plates, which did not replicate the apoptosis induced by the free peptide. [1] GRGDNP entered cells, as shown by internalization of biotinylated peptide. [1] GRGDNP directly induced the autoprocessing and activation of procaspase-3. Incubation of purified procaspase-3 with 1 mM GRGDNP led to its cleavage into active fragments. This process was specific, as the control peptide GRADSP had no effect. [1] Caspase-3 activity was significantly increased in cytoplasmic extracts from cells treated with GRGDNP, as measured by a fluorometric assay. This activation was inhibited by the caspase-3 inhibitor DEVD-CHO, but not by the caspase-1 inhibitor YVAD-CHO. [1] Western blot analysis confirmed the processing of procaspase-3 into its active p17 and p12 subunits in cells treated with GRGDNP. [1] The pro-apoptotic effect of GRGDNP was absent in MCF-7 breast carcinoma cells, which have a functional deletion of the caspase-3 gene, confirming that caspase-3 is required for GRGDNP-mediated cell death. [1] |
| Enzyme Assay |
Caspase-3 Activity Assay: Enzymatic caspase-3 activity was measured using a commercial fluorometric assay system. Cytoplasmic extracts were prepared from treated cells. The assay was performed according to the manufacturer's protocol. Caspase activation was expressed as the ratio of activity in the sample to that in control (untreated) samples. Background activity remaining after inhibition by the caspase-3 specific inhibitor DEVD-CHO was subtracted. [1]
Procaspase-3 Autoprocessing Assay: Procaspase-3 was purified from Molt-4 cell lysates by immunoprecipitation using an anti-caspase-3 antibody. The immunoprecipitated material was washed and resuspended in caspase assay buffer. It was then incubated with 1 mM GRGDNP or control peptide GRADSP for 10 minutes at 30°C. The reaction was stopped by adding Laemmli buffer, followed by boiling. The samples were then analyzed by SDS-PAGE and Western blotting to detect the cleavage fragments of procaspase-3 using the same anti-caspase-3 antibody. [1] |
| Cell Assay |
Apoptosis Induction Assay (Lymphocytes/Leukocytes): Resting peripheral blood T cells or other leukocyte cell types were cultured in serum-free media with GRGDNP at concentrations ranging from 0.1 to 1 mM (typically 1 mM) for 18 to 24 hours. Cell survival was assessed by propidium iodide exclusion and by forward- and side-scatter characteristics using flow cytometry. Results were expressed as the percentage of live cells remaining compared to the starting cell number. [1]
Intracellular Peptide Uptake Assay: Molt-4 cells were cultured with biotinylated GRGDNP for 18 hours. Cells were then either fixed and permeabilized to assess total (surface and cytoplasmic) peptide staining, or stained prior to fixation to assess surface-only staining. Staining was performed with fluorescently labeled streptavidin, and cells were analyzed by confocal microscopy. Propidium iodide was used as a counterstain. [1] Caspase-3 Processing (Western Blot): Whole-cell lysates were prepared from treated cells using RIPA buffer containing protease inhibitors. Protein concentration was determined. Lysates were boiled in Laemmli buffer, resolved by SDS-PAGE (15% gel), and transferred to a membrane for Western blot analysis. Antibodies against caspase-3 and β-actin (loading control) were used for detection. [1] Integrin-Independence Assay: To test if apoptosis required integrin clustering, two approaches were used: 1) Resting peripheral blood lymphocytes were cultured with beads coated with GRGDNP or control proteins/peptides. Free peptides were also added in some conditions. Survival was assessed. 2) Molt-4 cells were cultured on plastic coated with an integrin ligand (ProNectin-F) or standard tissue culture plates, with or without free GRGDNP. Survival was assessed. [1] |
| References |
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| Additional Infomation |
GRGDNP is a synthetic peptide containing an arginine-glycine-aspartic acid (RGD) motif. RGD sequences are known to mediate cell adhesion through integrin binding. [1]
This study discovered a novel, integrin-independent mechanism of action for RGD peptides. GRGDNP can enter cells and interact directly with procaspase-3, inducing its self-processing and activation, ultimately leading to apoptosis. [1] Sequence alignment results in this study showed that the procaspase-3 sequence contains an RGD motif, and there is also a potential RGD binding motif (aspartic acid-aspartic acid-methionine, DDM) near the cleavage site. The authors speculate that GRGDNP may trigger a conformational change in procaspase-3 through RGD-DDM interaction, thereby inducing apoptosis. [1] The results explain the pro-apoptotic properties of RGD peptides in angiogenesis, inflammation, and cancer metastasis models at the molecular level. [1] |
| Molecular Formula |
C23H38N10O10
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|---|---|
| Molecular Weight |
614.60900
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| Exact Mass |
614.277
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| CAS # |
114681-65-1
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| Related CAS # |
RGD peptide (GRGDNP) (TFA)
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| PubChem CID |
9895205
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| Appearance |
White to off-white solid powder
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| Density |
1.7±0.1 g/cm3
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| Index of Refraction |
1.696
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| LogP |
-3.41
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| Hydrogen Bond Donor Count |
10
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| Hydrogen Bond Acceptor Count |
12
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| Rotatable Bond Count |
18
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| Heavy Atom Count |
43
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| Complexity |
1110
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| Defined Atom Stereocenter Count |
4
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| SMILES |
C1C[C@H](N(C1)C(=O)[C@H](CC(=O)N)NC(=O)[C@H](CC(=O)O)NC(=O)CNC(=O)[C@H](CCCN=C(N)N)NC(=O)CN)C(=O)O
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| InChi Key |
CWAHAVYVGPRZJU-XUXIUFHCSA-N
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| InChi Code |
InChI=1S/C23H38N10O10/c24-9-16(35)30-11(3-1-5-28-23(26)27)19(39)29-10-17(36)31-12(8-18(37)38)20(40)32-13(7-15(25)34)21(41)33-6-2-4-14(33)22(42)43/h11-14H,1-10,24H2,(H2,25,34)(H,29,39)(H,30,35)(H,31,36)(H,32,40)(H,37,38)(H,42,43)(H4,26,27,28)/t11-,12-,13-,14-/m0/s1
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| Chemical Name |
(2S)-1-[(2S)-4-amino-2-[[(2S)-2-[[2-[[(2S)-2-[(2-aminoacetyl)amino]-5-(diaminomethylideneamino)pentanoyl]amino]acetyl]amino]-3-carboxypropanoyl]amino]-4-oxobutanoyl]pyrrolidine-2-carboxylic acid
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| HS Tariff Code |
2934.99.9001
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| Storage |
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. |
| Shipping Condition |
Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)
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| Solubility (In Vitro) |
May dissolve in DMSO (in most cases), if not, try other solvents such as H2O, Ethanol, or DMF with a minute amount of products to avoid loss of samples
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| Solubility (In Vivo) |
Note: Listed below are some common formulations that may be used to formulate products with low water solubility (e.g. < 1 mg/mL), you may test these formulations using a minute amount of products to avoid loss of samples.
Injection Formulations
Injection Formulation 1: DMSO : Tween 80： Saline = 10 : 5 : 85 (i.e. 100 μL DMSO stock solution → 50 μL Tween 80 → 850 μL Saline)(e.g. IP/IV/IM/SC) *Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH ₂ O to obtain a clear solution. Injection Formulation 2: DMSO : PEG300 ：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL DMSO → 400 μLPEG300 → 50 μL Tween 80 → 450 μL Saline) Injection Formulation 3: DMSO : Corn oil = 10 : 90 (i.e. 100 μL DMSO → 900 μL Corn oil) Example: Take the Injection Formulation 3 (DMSO : Corn oil = 10 : 90) as an example, if 1 mL of 2.5 mg/mL working solution is to be prepared, you can take 100 μL 25 mg/mL DMSO stock solution and add to 900 μL corn oil, mix well to obtain a clear or suspension solution (2.5 mg/mL, ready for use in animals). View More
Injection Formulation 4: DMSO : 20% SBE-β-CD in saline = 10 : 90 [i.e. 100 μL DMSO → 900 μL (20% SBE-β-CD in saline)] Oral Formulations
Oral Formulation 1: Suspend in 0.5% CMC Na (carboxymethylcellulose sodium) Oral Formulation 2: Suspend in 0.5% Carboxymethyl cellulose Example: Take the Oral Formulation 1 (Suspend in 0.5% CMC Na) as an example, if 100 mL of 2.5 mg/mL working solution is to be prepared, you can first prepare 0.5% CMC Na solution by measuring 0.5 g CMC Na and dissolve it in 100 mL ddH2O to obtain a clear solution; then add 250 mg of the product to 100 mL 0.5% CMC Na solution, to make the suspension solution (2.5 mg/mL, ready for use in animals). View More
Oral Formulation 3: Dissolved in PEG400 (Please use freshly prepared in vivo formulations for optimal results.) |
| Preparing Stock Solutions | 1 mg | 5 mg | 10 mg | |
| 1 mM | 1.6270 mL | 8.1352 mL | 16.2705 mL | |
| 5 mM | 0.3254 mL | 1.6270 mL | 3.2541 mL | |
| 10 mM | 0.1627 mL | 0.8135 mL | 1.6270 mL |
*Note: Please select an appropriate solvent for the preparation of stock solution based on your experiment needs. For most products, DMSO can be used for preparing stock solutions (e.g. 5 mM, 10 mM, or 20 mM concentration); some products with high aqueous solubility may be dissolved in water directly. Solubility information is available at the above Solubility Data section. Once the stock solution is prepared, aliquot it to routine usage volumes and store at -20°C or -80°C. Avoid repeated freeze and thaw cycles.
Calculation results
Working concentration： mg/mL；
Method for preparing DMSO stock solution： mg drug pre-dissolved in μL DMSO (stock solution concentration mg/mL). Please contact us first if the concentration exceeds the DMSO solubility of the batch of drug.
Method for preparing in vivo formulation:：Take μL DMSO stock solution, next add μL PEG300, mix and clarify, next addμL Tween 80, mix and clarify, next add μL ddH2O，mix and clarify.
(1) Please be sure that the solution is clear before the addition of next solvent. Dissolution methods like vortex, ultrasound or warming and heat may be used to aid dissolving.
(2) Be sure to add the solvent(s) in order.
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