FTS also suppressed CER-induced ERK activation in the kidney. In vitro treatment of the established cell line, LLC-PK1 cells, with FTS significantly ameliorated CER-induced cell injury, as measured by lactate dehydrogenase (LDH) leakage. Our results, taken together with our previous report that MEK inhibitors ameliorated CER-induced renal cell injury and ERK activation induced by CER, suggest that FTS participates in protection from CER-induced nephrotoxicity by suppressing ERK activation induced by CER.[2]

FTS also suppressed CER-induced ERK activation in the kidney. In vitro treatment of the established cell line, LLC-PK1 cells, with FTS significantly ameliorated CER-induced cell injury, as measured by lactate dehydrogenase (LDH) leakage. Our results, taken together with our previous report that MEK inhibitors ameliorated CER-induced renal cell injury and ERK activation induced by CER, suggest that FTS participates in protection from CER-induced nephrotoxicity by suppressing ERK activation induced by CER.[2]

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AZD9291 demonstrated potent inhibition of EGFR phosphorylation in cell lines harboring EGFR mutations. In H1975 cells (DM: L858R/T790M), the phosphorylation IC₅₀ was 15 nM. In PC9 cells (AM: exon 19 deletion), the phosphorylation IC₅₀ was 17 nM. In contrast, it showed much weaker activity in wild-type EGFR cell lines like LoVo (IC₅₀ >480 nM) and Calu-3 (antiproliferative GI₅₀ = 264 nM), indicating significant selectivity for mutant over wild-type EGFR. [1]
In cellular antiproliferation assays, AZD9291 showed potent growth inhibition against mutant EGFR-driven cells. The GI₅₀ was 24 nM in H1975 cells (DM), 23 nM in PC9 cells (AM), and 264 nM in Calu-3 cells (WT). [1]
Kinase selectivity profiling against a panel of 270 kinases (Millipore panel) at 1 µM showed that AZD9291 is highly selective, with significant inhibition primarily confined to a small number of tyrosine kinases including EGFR mutants, IGF1R, and INSR. [1]

Serum thymic factor (FTS), a thymic peptide hormone, has been reported to increase superoxide disumutase (SOD) levels in senescence-accelerated mice.That CER led to extracellular signal-regulated protein kinase (ERK) activation in the rat kidney. So, we also investigated whether FTS has an effect on ERK activation induced by CER. Treatment of male Sprague-Dawley rats with intravenous CER (1.2 g/kg) for 24 h markedly increased BUN and plasma creatinine levels and urinary excretion of glucose and protein, decreased creatinine clearance and also led to marked pathological changes in the proximal tubules, as revealed by electron micrographs. An increase in phosphorylated ERK (pERK) was detected in the nuclear fraction prepared from the rat kidney cortex 24 h after CER injection. Pretreatment of rats with FTS (50 microg/kg, i.v.) attenuated the CER-induced renal dysfunction and pathological damage.[2]

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The effect of serum thymic factor (FTS) on the D-variant of encephalomyocarditis (EMC-D) virus-induced diabetes and myocarditis in BALB/cAJcl mice was investigated. Mice pretreated with 50 or 10 micrograms of FTS were infected with 10 or 10(3) PFU of EMC-D virus. In the mice inoculated with 10 PFU of virus, 40% developed diabetes on post-infection day (PID) 14, whereas those treated with FTS (50 micrograms/administration) on day 2 and 1 before infection did not develop diabetes. FTS (10 micrograms)-pretreated mice developed diabetes. In histological observation, FTS non-treated mice which developed diabetes showed severe necrosis and inflammation of mononuclear cells in the islets of Langerhans and myocardia on 19 PID. Mice pretreated with 50 micrograms of FTS, however, manifested mild islet degeneration without any myocardial inflammation. Furthermore, in FTS non-treated mice, immunohistological staining showed a loss of insulin granules. This loss was markedly reversed and insulin granules remained largely intact in FTS-pretreated mice. Viral titers in pancreas of FTS-pretreated mice approximated well to those of non-treated mice on PID 4, 7 and 19. In mice inoculated with higher titer of EMC-D virus (10(3) PFU), however, 50 micrograms of FTS pretreatment did not change the course of these acute pathological developments (diabetes and myocarditis observed from PID 4).[3]

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In mouse xenograft models, oral administration of AZD9291 at 5 and 10 mg/kg once daily for 7 days caused significant tumor growth inhibition and regression in models harboring EGFR mutations. In the H1975 (DM) model, it induced 148% tumor growth inhibition (TGI) at 10 mg/kg/day. In the PC9 (AM) model, it induced 140% TGI at 10 mg/kg/day and 122% TGI even at the lower 5 mg/kg/day dose. In contrast, it had minimal effect on the A431 xenograft model which expresses wild-type EGFR. [1]
A single oral dose of AZD9291 (10 mg/kg) in mice bearing H1975 or PC9 xenografts led to profound and sustained inhibition of EGFR phosphorylation (p-EGFR) as a pharmacodynamic biomarker, with near-complete inhibition observed for up to 24 hours. [1]

The enzymatic activity of AZD9291 against IGF1R and INSR was determined using standard kinase assays. The compound's potency is reported as an IC₅₀ value. [1]
The inhibition of the hERG potassium channel was assessed using an Ionworks assay platform, determining the concentration at which 50% of the channel current is inhibited (IC₅₀). [1]

Cellular potency (IC₅₀) for EGFR phosphorylation inhibition was measured using specific cell lines. For double mutant EGFR, the H1975 human lung cancer cell line (harboring L858R/T790M) was used. For activating mutant EGFR, the PC9 human lung cancer cell line (harboring exon 19 deletion) was used. For wild-type EGFR, the LoVo human colon cancer cell line or the Calu-3 human lung cancer cell line was used. Cells were treated with compound for 2 hours, after which EGFR phosphorylation levels were quantified, typically using ELISA methods. IC₅₀ values are the geometric mean of at least two independent measurements. [1]
Cellular antiproliferative activity (GI₅₀) was assessed using the same panel of cell lines (H1975, PC9, Calu-3). Cells were exposed to the compound, and cell viability/growth was measured after a designated period to determine the concentration causing 50% growth inhibition. [1]
Kinase selectivity was profiled using a commercial kinase panel (Millipore panel of 270 kinases). Compounds were tested at a single concentration of 1 µM, and the percentage inhibition of each kinase activity was measured. [1]

For in vivo efficacy studies in mouse xenograft models, female immune-compromised (SCID or nude) mice were implanted subcutaneously with tumor cells (H1975, PC9, or A431). When tumors reached a specified volume, mice were randomized into groups. AZD9291 (free base or mesylate salt) was formulated as a suspension in 0.5% hydroxypropyl methylcellulose (HPMC) / 0.1% Tween 80 in water. The compound was administered orally (po) once daily (q.d.) at doses of 5 or 10 mg/kg for 7 to 28 days. Tumor volumes and body weights were monitored regularly. [1]
For pharmacodynamic (PD) studies, mice bearing H1975 or PC9 xenografts received a single oral dose of AZD9291 (10 mg/kg). Tumors were harvested at various time points (e.g., 1, 6, 16, 24, 30 hours post-dose), and levels of phosphorylated EGFR (p-EGFR) were measured in tumor lysates using ELISA kits. [1]
For toxicology and toxicokinetic studies in rats, male Han Wistar rats received a single oral dose of AZD9291 (50 or 200 mg/kg) as a suspension. Blood samples were collected over 24 hours to measure plasma drug concentrations, glucose levels, and insulin levels. [1]

In preclinical animal models, AZD9291 showed moderate to high clearance rates: 77 mL/min/kg in mice, 22 mL/min/kg in rats, and 16 mL/min/kg in dogs after intravenous injection. [1] Oral bioavailability varied among individuals and was dose-dependent. Bioavailability was 13% in rats at a dose of 5 mg/kg; 31% in mice at a dose of 10 mg/kg; and 67% in dogs at a dose of 5 mg/kg. Higher doses resulted in increased drug exposure, suggesting that the clearance pathway may be saturated. [1] AZD9291 is metabolized in vivo. In rodents and humans, the main circulating metabolites are metabolite 27 (M27) via N-demethylation of the indole group and metabolite 28 (M28) via dealkylation of the side chain. In addition, a small amount of N-oxide metabolite was also detected. [1]
In the first-in-human phase I study, patients with EGFR mutation-positive non-small cell lung cancer (NSCLC) were given 20 mg of mesylate orally once daily. AZD9291 and its metabolites M27 and M28 were detected in plasma with low inter-patient variability. [1]
AZD9291 has a logD value of 3.4 at pH 7.4. Its aqueous solubility at pH 7.4 is 7 µM. It has high plasma protein binding, with a free fraction of 2.9% in human plasma. [1]

In vitro experiments showed that the IC₅₀ of AZD9291 for inhibiting hERG potassium channels was 16.2 µM, suggesting a possible risk of QT interval prolongation, but the risk was relatively low. [1] In a rat toxicology study, a single oral high dose (200 mg/kg) of the early lead compounds (20, 21, 22) strongly inhibited IGF1R/INSR, resulting in significant hyperglycemia and hyperinsulinemia. In contrast, the same dose of AZD9291 had no significant effect on blood glucose or insulin levels, consistent with its weaker IGF1R/INSR inhibitory effect. [1] In a guinea pig cardiovascular safety model, AZD9291 did not cause significant QT interval prolongation at free plasma concentrations significantly higher than the predicted effective human exposure, supporting its higher safety profile compared to other lead compounds. [1]
AZD9291 has a high plasma protein binding rate; its free fraction in human plasma was only 2.9% as determined by equilibrium dialysis. [1]

[1]. Role of thymulin or its analogue as a new analgesic molecule. Ann N Y Acad Sci. 2006 Nov;1088:153-63.

[2]. Protective effect of serum thymic factor, FTS, on cephaloridine-induced nephrotoxicity in rats. Biol Pharm Bull. 2005 Nov;28(11):2087-91.

[3]. In vivo administration of serum thymic factor (FTS) prevents EMC-D virus-induced diabetes and myocarditis in BALB/cAJcl mice. Arch Virol. 1996;141(1):73-83.

[4]. Physiology, molecular biology and therapeutic potential of the thymic peptide thymulin. Physiological Mini Reviews (PMR), 2012, 6.

thymus-dependent nonapeptide present in normal blood. It stimulates the formation of E-rosettes and is thought to be involved in T-cell differentiation.

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AZD9291 (osimertinib) is a third-generation irreversible EGFR tyrosine kinase inhibitor (TKI) designed to overcome resistance mediated by the T790M mutation in patients with non-small cell lung cancer (NSCLC) who have previously received first-generation EGFR TKIs (such as gefitinib and erlotinib). [1]
Its mechanism of action involves covalently binding to the cysteine 797 residue at the ATP-binding site of the mutated EGFR, resulting in irreversible inhibition. Based on published EGFR structural models, its key interaction with the hinge region (M793) and the orientation of its indole group are crucial. [1]
Preliminary clinical data from a phase I clinical trial (AURA) in patients with T790M-positive, EGFR-TKI-resistant non-small cell lung cancer (NSCLC) showed encouraging efficacy. Two representative patients experienced significant tumor shrinkage (39-63% reduction according to RECIST 1.1 criteria) and improved clinical symptoms after receiving once-daily doses of 20 mg. Treatment was well tolerated, with no severe rash reported in early-stage patients; only mild diarrhea was reported. [1]

C33H54N12O15

858.86

858.383

63958-90-7

3085284

Typically exists as solid at room temperature

1.417 g/cm33

1658.9ºC at 760 mmHg

957.1ºC

1.575

-10.9

15

16

28

60

1610

7

C[C@H](NC([C@@H]1CCC(N1)=O)=O)C(N[C@H](C(N[C@H](C(N[C@H](C(NCC(NCC(N[C@H](C(N[C@H](C(O)=O)CC(N)=O)=O)CO)=O)=O)=O)CCC(N)=O)=O)CO)=O)CCCCN)=O

LIFNDDBLJFPEAN-BPSSIEEOSA-N

InChI=1S/C33H54N12O15/c1-15(39-29(55)18-6-8-24(50)40-18)27(53)42-16(4-2-3-9-34)30(56)45-21(14-47)32(58)43-17(5-7-22(35)48)28(54)38-11-25(51)37-12-26(52)41-20(13-46)31(57)44-19(33(59)60)10-23(36)49/h15-21,46-47H,2-14,34H2,1H3,(H2,35,48)(H2,36,49)(H,37,51)(H,38,54)(H,39,55)(H,40,50)(H,41,52)(H,42,53)(H,43,58)(H,44,57)(H,45,56)(H,59,60)/t15-,16-,17-,18-,19-,20-,21-/m0/s1

(2S)-4-amino-2-[[(2S)-2-[[2-[[2-[[(2S)-5-amino-2-[[(2S)-2-[[(2S)-6-amino-2-[[(2S)-2-[[(2S)-5-oxopyrrolidine-2-carbonyl]amino]propanoyl]amino]hexanoyl]amino]-3-hydroxypropanoyl]amino]-5-oxopentanoyl]amino]acetyl]amino]acetyl]amino]-3-hydroxypropanoyl]amino]-4-oxobutanoic acid

thymulinSerum thymic factorNonathymulin

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.

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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).

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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)

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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).

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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

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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.

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