p70S6K (IC50 = 4 nM)

LY-2584702 (LY2584702), having an IC50 of 0.1-0.24 μM, suppresses the phosphorylation of S6 ribosomal protein (pS6) in HCT116 colon cancer cells [1]. The S6K1 enzyme test yields an IC50 value of 2 nM for LY-2584702 (LY2584702). In cells, IC50=100 nM for pS6 inhibition. At high concentrations (IC50=58-176 nM in the enzyme assay), LY-2584702 exhibits specific action against the S6K-related kinases MSK2 and RSK. The phosphorylation of LY-2584702's downstream effector S6 determines how dose-dependently it suppresses S6K activity in EOMA cells [2]. When administered at 0.1 μM for more than 24 hours, LY-2584702 (LY2584702) can considerably limit the proliferation of A549 (P<0.05); additional time under treatment and/or higher drug concentrations make the downward trend more pronounced (both P<0.05). In SK-MES-1, comparable outcomes were noted, but at 0.6 μM Significantly more potent than A549, LY-2584702 showed an inhibitory effect (P<0.05)[3].

Under 2.5 mg/kg twice daily (BID) and 12.5 mg/kg BID, LY-2584702 showed noteworthy single-agent activity in HCT116 colon cancer and U87MG glioblastoma xenograft models. At TMED50 (threshold minimum effective dose 50%) (2.3 mg/kg) and TMED90 (10 mg/kg), LY-2584702 demonstrated statistically significant tumor growth decrease in the HCT116 colon cancer xenograft model [1]. LY-2584702 or rapamycin were administered to shAkt3-expressing EOMA cells implanted into nu/nu mice for a period of 14 days in order to investigate the function of S6K in vivo. LY-2584702 suppressed S6 phosphorylation nearly as well as rapamycin, according to analysis of tumors removed after 14 days. In contrast to pLKO, tumor development was enhanced by Akt3 loss. The growth of pLKO tumors was not substantially impacted by LY-2584702 treatment alone. LY-2584702, however, dramatically slowed the growth of shAkt3 tumors [2].

[1]. A phase I trial of LY2584702 tosylate, a p70 S6 kinase inhibitor, in patients with advanced solid tumors. Eur J Cancer. 2014 Mar;50(5):867-75.

[2]. Akt1 and akt3 exert opposing roles in the regulation of vascular tumor growth. Cancer Res. 2015 Jan 1;75(1):40-50.

[3]. Hyperphosphorylation of RPS6KB1, rather than overexpression, predicts worse prognosis in non-small cell lung cancer patients. PLoS One. 2017 Aug 9;12(8):e0182891.

Vascular tumors are endothelial cell neoplasms whose mechanisms of tumorigenesis are poorly understood. Moreover, current therapies, particularly those for malignant lesions, have little beneficial effect on clinical outcomes. In this study, we show that endothelial activation of the Akt1 kinase is sufficient to drive de novo tumor formation. Mechanistic investigations uncovered opposing functions for different Akt isoforms in this regulation, where Akt1 promotes and Akt3 inhibits vascular tumor growth. Akt3 exerted negative effects on tumor endothelial cell growth and migration by inhibiting activation of the translation regulatory kinase S6-Kinase (S6K) through modulation of Rictor expression. S6K in turn acted through a negative feedback loop to restrain Akt3 expression. Conversely, S6K signaling was increased in vascular tumor cells where Akt3 was silenced, and the growth of these tumor cells was inhibited by a novel S6K inhibitor. Overall, our findings offer a preclinical proof of concept for the therapeutic utility of treating vascular tumors, such as angiosarcomas, with S6K inhibitors.[2]

---

RPS6KB1 is the kinase of ribosomal protein S6 which is 70 kDa and is required for protein translation. Although the abnormal activation of RPS6KB1 has been found in types of diseases, its role and clinical significance in non-small cell lung cancer (NSCLC) has not been fully investigated. In this study, we identified that RPS6KB1 was over-phosphorylated (p-RPS6KB1) in NSCLC and it was an independent unfavorable prognostic marker for NSCLC patients. In spite of the frequent expression of total RPS6KB1 and p-RPS6KB1 in NSCLC specimens by immunohistochemical staining (IHC), only p-RPS6KB1 was associated with the clinicopathologic characteristics of NSCLC subjects. Kaplan-Meier survival analysis revealed that the increased expression of p-RPS6KB1 indicated a poorer 5-year overall survival (OS) for NSCLC patients, while the difference between the positive or negative RPS6KB1 group was not significant. Univariate and multivariate Cox regression analysis was then used to confirm the independent prognostic value of p-RPS6KB1. To illustrate the underlying mechanism of RPS6KB1 phosphorylation in NSCLC, LY2584702 was employed to inhibit the RPS6KB1 phosphorylation specifically both in lung adenocarcinoma cell line A549 and squamous cell carcinoma cell line SK-MES-1. As expected, RPS6KB1 dephosphorylation remarkably suppressed cells proliferation in CCK-8 test, and promoted more cells arresting in G0-G1 phase by cell cycle analysis. Moreover, apoptotic A549 cells with RPS6KB1 dephosphorylation increased dramatically, with an elevating trend in SK-MES-1, indicating a potential involvement of RPS6KB1 phosphorylation in inducing apoptosis. In conclusion, our data suggest that RPS6KB1 is over-activated as p-RPS6KB1 in NSCLC, rather than just the total protein overexpressing. The phosphorylation level of RPS6KB1 might be used as a novel prognostic marker for NSCLC patients.[3]

C21H20CLF4N7

481.877016067505

481.14

1082948-81-9

LY-2584702 tosylate salt;1082949-68-5;LY-2584702 free base;1082949-67-4

66650363

Typically exists as solid at room temperature

2

9

3

33

644

0

CN1C=C(C2=CC=C(F)C(C(F)(F)F)=C2)N=C1C3CCN(C4=C5C(NN=C5)=NC=N4)CC3.[H]Cl

GDGYRKDHQORLNT-UHFFFAOYSA-N

InChI=1S/C21H19F4N7.ClH/c1-31-10-17(13-2-3-16(22)15(8-13)21(23,24)25)29-19(31)12-4-6-32(7-5-12)20-14-9-28-30-18(14)26-11-27-20;/h2-3,8-12H,4-7H2,1H3,(H,26,27,28,30);1H

4-[4-[4-[4-fluoro-3-(trifluoromethyl)phenyl]-1-methylimidazol-2-yl]piperidin-1-yl]-1H-pyrazolo[3,4-d]pyrimidine;hydrochloride

LY-2584702 hydrochloride; 1082948-81-9; LY-2584702 (hydrochloride); 4-(4-(4-(4-Fluoro-3-(trifluoromethyl)phenyl)-1-methyl-1H-imidazol-2-yl)piperidin-1-yl)-1H-pyrazolo[3,4-d]pyrimidine hydrochloride; 4-[4-[4-[4-fluoro-3-(trifluoromethyl)phenyl]-1-methylimidazol-2-yl]piperidin-1-yl]-1H-pyrazolo[3,4-d]pyrimidine;hydrochloride; 4-{4-[4-fluoro-3-(trifluoromethyl)phenyl]-1-methyl-1H-imidazol-2-yl}-1-{1H-pyrazolo[3,4-d]pyrimidin-4-yl}piperidine hydrochloride; LY2584702 Hydrochloride; SCHEMBL312564;

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

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

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

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 Formulation 1: DMSO : Tween 80： Saline = 10 : 5 : 85 (i.e. 100 μL DMSO stock solution → 50 μL Tween 80 → 850 μL Saline)
*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)]
*Preparation of 20% SBE-β-CD in Saline (4°C，1 week): Dissolve 2 g SBE-β-CD in 10 mL saline to obtain a clear solution.
Injection Formulation 5: 2-Hydroxypropyl-β-cyclodextrin : Saline = 50 : 50 (i.e. 500 μL 2-Hydroxypropyl-β-cyclodextrin → 500 μL Saline)
Injection Formulation 6: DMSO : PEG300 : castor oil : Saline = 5 : 10 : 20 : 65 (i.e. 50 μL DMSO → 100 μLPEG300 → 200 μL castor oil → 650 μL Saline)
Injection Formulation 7: Ethanol : Cremophor : Saline = 10： 10 : 80 (i.e. 100 μL Ethanol → 100 μL Cremophor → 800 μL Saline)
Injection Formulation 8: Dissolve in Cremophor/Ethanol (50 : 50), then diluted by Saline
Injection Formulation 9: EtOH : Corn oil = 10 : 90 (i.e. 100 μL EtOH → 900 μL Corn oil)
Injection Formulation 10: EtOH : PEG300：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL EtOH → 400 μLPEG300 → 50 μL Tween 80 → 450 μL Saline)

---

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
Oral Formulation 4: Suspend in 0.2% Carboxymethyl cellulose
Oral Formulation 5: Dissolve in 0.25% Tween 80 and 0.5% Carboxymethyl cellulose
Oral Formulation 6: Mixing with food powders

---

Note: Please be aware that the above formulations are for reference only. InvivoChem strongly recommends customers to read literature methods/protocols carefully before determining which formulation you should use for in vivo studies, as different compounds have different solubility properties and have to be formulated differently.

---

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