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| 5mg |
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| Targets |
LY311727 targets human non-pancreatic secretory phospholipase A2 (hnps-PLA2, Group II sPLA2). IC50 (mole fraction, X(50)) = 0.000019 in chromogenic assay. [1] For porcine pancreatic s-PLA2 (Group I), X(50) = 0.029 (1,500-fold less potent). [1] Apparent dissociation constant (KB) on guinea pig lung pleural strips against hnps-PLA2-induced contractions = 0.27 ± 0.05 μM. [1] No cyclooxygenase (CO) activity. [1]
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| ln Vitro |
Human non-pancreatic secretory phospholipase A2 (hnps-PLA2)-induced contractile responses are inhibited in a concentration-dependent manner by LY-311727 (0.1-10 μM) [1]. At 1 μM, LY-311727 nearly completely reduced the hnps-PLA2 response, but it was unable to suppress the porcine pancreatic PLA2 concentration response curve [1]. A 1,500-fold selectivity against porcine pancreas s-PLA2 was revealed by LY-311727 [1].
[1] LY311727 (indole 10) displayed an X(50) of 0.000019 mole fraction in chromogenic assays, representing a 737-fold improvement over the initial lead indole 1 (X(50)=0.014). The ethyl group at R2 (compared to methyl in indole 8) improved potency ~3-fold; a propyl group (indole 11) resulted in >250-fold loss of activity. A phenyl substitution at the para position of the benzyl ring (indole 12) showed >100-fold lower activity than LY311727. The compound showed no cyclooxygenase (CO) activity in the arachidonic acid (AA) tissue bath assay. [1] [2] In transgenic mice overexpressing human sPLA2, LY311727 significantly suppressed circulating sPLA2 catalytic activity in a dose-dependent manner. At 10 mg/kg i.v., inhibition was >80% at 30 min. At 30 mg/kg i.v., inhibition remained >80% over 4 h. A close structural analog LY314024 (which is ~250-fold less potent in vitro) showed no significant inhibition at 10 mg/kg compared to vehicle. [2] |
| ln Vivo |
In metallothionein promoter-human secretory PLA2 minigene (Mt-sPLA2) transgenic mice, intravenous (iv) administration of LY-311727 (3-30 mg/kg) dramatically reduces circulating enzyme activity [2].
[1] LY311727 inhibited hnps-PLA2-induced contractile responses on guinea pig lung pleural strips in a concentration-related manner (0.1–10 μM), with an apparent dissociation constant (KB) of 0.27 ± 0.05 μM. At 10 μM, LY311727 nearly abolished hnps-PLA2 responses while failing to suppress porcine pancreatic PLA2 concentration-response curves at the same concentration. Contractions induced by arachidonic acid were not inhibited, confirming no cyclooxygenase activity. [1] [2] In Mt-sPLA2 transgenic mice (C57BL/6 background, sPLA2-deficient), intravenous administration of LY311727 at 3, 10, and 30 mg/kg dose-dependently suppressed serum PLA2 catalytic activity. At 30 mg/kg, inhibition exceeded 80% over 4 hours post-dose. At 10 mg/kg, inhibition was >80% at 30 minutes; the less potent analog LY314024 showed no effect at the same dose. [2] |
| Enzyme Assay |
[1] hnps-PLA2 inhibition was evaluated using a modified chromogenic assay with a thiol substrate analogue. The assay contained final concentrations of 0.96 mM racemic 1,2-bis(thioheptanoyl)-1,2-dideoxyphosphatidylcholine (PC), 0.27 mM Triton X-100, and 0.12 mM 5,5'-dithiobis(2-nitrobenzoic acid). Concentration-response curves were generated with 16 nM recombinant hnps-PLA2 for 30 min at 40 °C in a microtiter plate format. Mole fractions (X(50) = [I]/([I]+[PC]+[T])) for 50% inhibition were determined in triplicate; standard deviations were +/- 10-50%. [1]
[2] Serum PLA2 catalytic activity in transgenic mice was assayed using a modified phosphatidylcholine/deoxycholate mixed micelle assay containing 3 mM sodium deoxycholate and 1 mM 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine. [2] [1] Crystallographic studies: crystals of hnps-PLA2 complexed with LY311727 (and related indoles) were grown by vapor diffusion from solutions containing 10 mg/mL protein in 50 mM buffer (MES or MOPS), pH 6.6–7.5, 80–92% saturated sodium chloride, 1% pyridine, with 1.5 molar equivalents of inhibitor. X-ray diffraction data were collected using imaging plate detector and rotating anode source (CuKa, λ=1.542 Å). Structures were solved by molecular replacement using X-PLOR and refined with PROLSQ. [1] |
| Cell Assay |
[1] Guinea pig lung pleural strips were challenged with hnps-PLA2 or arachidonic acid. LY311727 was incubated with tissues for 30 min prior to starting PLA2 concentration-response curves. Data were expressed as percentage of maximal KCl (40 mM) responses. [1]
[2] PLA2 catalytic activity in plasma/serum of transgenic mice was measured using an E. coli membrane assay; the amount of enzyme present was calculated from a standard curve generated using purified recombinant human sPLA2. PLA2 protein levels were quantitated by ELISA capture assay using a mouse monoclonal anti-human sPLA2 antibody as capture and a horseradish peroxidase-conjugated monoclonal antibody as detection reagent. [2] |
| Animal Protocol |
Animal/Disease Models: C57BL/6J mice, Mt-sPLA2 transgenic mouse model [2]
Doses: 3 mg/kg, 10 mg/kg, 30 mg/kg Route of Administration: intravenous (iv) (iv)injection Experimental Results: Significant and dose-dependent Inhibits PLA2 activity in serum. [1] For guinea pig pleural strip assays: tissues were prepared and LY311727 incubated for 30 min prior to starting PLA2 concentration-response curves. Data pooled from individual experiments expressed as percentage of maximal KCl (40 mM) responses (means ± sem). [1] [2] Transgenic mice (Mt-sPLA2 on C57BL/6J background, which are sPLA2-deficient due to a natural mutation) were used. Mice were bled retro-orbitally prior to drug or vehicle administration and at 30 min, 2 h, and 4 h thereafter. Compounds were administered as an intravenous bolus via the tail vein in a volume of 0.15 mL. Vehicle composition: 5% dimethylsulfoxide, 5% ethanol, and 30% polyethylene glycol 300 in H2O. Three to six mice were used per dose. PLA2 catalytic activity in serum was assayed and values graphed as percentage change (± S.E.M.) from time zero (pre-treated) serum sample. [2] |
| References |
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| Additional Infomation |
[1] LY311727 is the first potent and selective inhibitor of hnps-PLA2. It was developed through structure-based drug design from a screening lead (indole 1) identified from large-scale screening. The optimization involved converting the 3-acetate to an acetamide (providing hydrogen bond to His 48 and calcium coordination), adding a 5-carboxylate or phosphonate (mimicking phosphate interactions), and optimizing the 2-position with an ethyl group. The compound's 1,500-fold selectivity over porcine pancreatic PLA2 was demonstrated both in biochemical assays and on guinea pig lung tissue (natural membrane substrate). The compound shows 50% inhibition of substrate hydrolysis at a concentration 20,000 times less than the phospholipid substrate concentration. [1]
[2] LY311727 was used to demonstrate that inhibition of circulating human sPLA2 in transgenic mice occurs through specific interaction with the catalytic domain, as the structurally similar but less potent analog LY314024 (which is ~250-fold less active in vitro) showed no effect. The transgenic model (C57BL/6 background, sPLA2-deficient due to a natural mutation) expresses human sPLA2 under the inducible mouse metallothionein promoter, with circulating levels inducible to those observed in patients with systemic inflammatory response. [2] |
| Molecular Formula |
C22H27N2O5P
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|---|---|
| Molecular Weight |
430.4408
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| Exact Mass |
430.166
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| CAS # |
164083-84-5
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| PubChem CID |
155922
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| Appearance |
White to off-white solid powder
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| Density |
1.32g/cm3
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| Boiling Point |
758.2ºC at 760mmHg
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| Flash Point |
412.3ºC
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| Vapour Pressure |
3.69E-24mmHg at 25°C
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| Index of Refraction |
1.619
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| LogP |
4.376
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| Hydrogen Bond Donor Count |
3
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| Hydrogen Bond Acceptor Count |
5
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| Rotatable Bond Count |
10
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| Heavy Atom Count |
30
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| Complexity |
606
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| Defined Atom Stereocenter Count |
0
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| InChi Key |
OPWQYOUZRHDKBR-UHFFFAOYSA-N
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| InChi Code |
InChI=1S/C22H27N2O5P/c1-2-20-19(14-22(23)25)18-13-17(29-11-6-12-30(26,27)28)9-10-21(18)24(20)15-16-7-4-3-5-8-16/h3-5,7-10,13H,2,6,11-12,14-15H2,1H3,(H2,23,25)(H2,26,27,28)
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| Chemical Name |
Phosphonic acid, (3-((3-(2-amino-2-oxoethyl)-2-ethyl-1-(phenylmethyl)-1H-indol-5-yl)oxy)propyl)-
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| Synonyms |
LY 311727 LY311727
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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 |
| 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 | 2.3232 mL | 11.6160 mL | 23.2320 mL | |
| 5 mM | 0.4646 mL | 2.3232 mL | 4.6464 mL | |
| 10 mM | 0.2323 mL | 1.1616 mL | 2.3232 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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