mTOR (IC50 = 1.76 μM)

In the absence of FKBP12, Temsirolimus potently inhibits mTOR kinase activity with IC50 of 1.76 μM, similar to that of rapamycin with IC50 of 1.74 μM. Temsirolimus treatment at nanomolar concentrations (10 nM to <5 μM) exhibits a modest and selective antiproliferative activity via FKBP12-dependent mechanism, but at low micromolar concentrations (5-15 μM), it can completely inhibit the proliferation of a wide panel of tumor cells by suppressing mTOR signaling in a manner that is FKBP12-independent. Treatment with temsirolimus at micromolar (20 μM), but not nanomolar, concentrations results in a marked reduction in overall protein synthesis and polyribosome disassembly, which is accompanied by a sharp rise in the phosphorylation of the translation elongation factor eEF2 and the translation initiation factor eIF2A.[1] Temsirolimus inhibits cell growth and clonogenic survival in both cells in a concentration-dependent manner, but more potently in PTEN-positive DU145 cells than in PTEN-negative PC-3 cells. It also inhibits the phosphorylation of ribosomal protein S6. [2] Primary human lymphoblastic leukemia (ALL) cells are potently inhibited from proliferating and are induced to undergo apoptosis by temsirolimus (100 ng/mL).[3]

In the NOD/SCID xenograft models with human ALL, Temsirolimus treatment at 10 mg/kg/day produces a decrease in peripheral blood blasts and in splenomegaly.[3] Temsirolimus (20 mg/kg i.p. 5 days/week) significantly slows down the growth of DAOY xenografts compared to controls, delaying it by 160% after 1 week and 240% after 2 weeks. One week of treatment with a single high-dose of temsirolimus (100 mg/kg i.p.) causes a 37% reduction in tumor volume. The growth of rapamycin-resistant U251 xenografts is also 148% delayed by temsirolimus treatment for 2 weeks.[4] Temsirolimus's inhibition of mTOR enhances performance on four distinct behavioral tasks and reduces aggregate formation in a mouse model of Huntington disease. [5] Temsirolimus administration results in significant dose-dependent, antitumor responses against subcutaneous growth of 8226, OPM-2, and U266 xenografts, with ED50 values of 20 mg/kg and 2 mg/kg for 8226 and OPM-2, respectively. These responses are linked to decreased tumor cell growth and inhibition of angiogenesis as well as increased apoptosis and inhibition of proliferation.[6]

Transiently transfecting HEK293 cells with Flag-tagged wild-type human mTOR (Flag-mTOR) DNA constructs. 48 hours later, Flag-mTOR protein is extracted and purified. Purified Flag-mTOR in vitro kinase assays are carried out in 96-well plates in the presence of various Temsirolimus concentrations without FKBP12, and the results are detected using the dissociation-enhanced lanthanide fluorescent immunoassay (DELFIA) method with His6-S6K1 as the substrate. Enzymes are first diluted in kinase assay buffer (10 mM Hepes (pH 7.4), 50 mM NaCl, 50 mM β-glycerophosphate, 10 mM MnCl2, 0.5 mM DTT, 0.25 μM microcystin LR, and 100 μg/mL BSA). 12 μL of the diluted enzyme and 0.5 μL of temsirolimus are quickly combined in each well. The kinase reaction is started by adding 12.5 μL of ATP and His6-S6K-containing kinase assay buffer to create a final reaction volume of 25 μL that contains 800 ng/mL FLAG-mTOR, 100 μM ATP, and 1.25 μM His6-S6K. The reaction plate is incubated for 2 hours (linear at 1-6 hours) at room temperature with gentle shaking before being stopped by adding 25 μL of stop buffer (20 mM Hepes (pH 7.4), 20 mM EDTA, and 20 mM EGTA). A monoclonal anti-P(T389)-p70S6K antibody labeled with Europium-N1-ITC (Eu) (10.4 Eu per antibody) is used for the DELFIA detection of the phosphorylated (Thr-389) His6-S6K at room temperature. Transfer 45 μL of the terminated kinase reaction mixture to a MaxiSorp plate containing 55 μL PBS. Eu-P(T389)-S6K antibody is added to 100 μL of DELFIA buffer at a concentration of 40 ng/mL. With minimal agitation, the antibody binding is continued for an additional hour. The wells are then aspirated and cleaned using PBS with 0.05% Tween 20 (PBST). Each well receives 100 L of DELFIA Enhancement solution before the plates are read using a PerkinElmer Victor model plate reader.

Temsirolimus is applied to cells in a range of concentrations for 72 hours. Viable cell densities are assessed following treatment using the CellTiter AQ assay kit to measure MTS dye conversion.
In cell culture studies, CCI-779 at the commonly used nanomolar concentrations generally confers a modest and selective antiproliferative activity. Here, we report that, at clinically relevant low micromolar concentrations, CCI-779 completely suppressed proliferation of a broad panel of tumor cells. This "high-dose" drug effect did not require FKBP12 and correlated with an FKBP12-independent suppression of mTOR signaling. An FKBP12-rapamycin binding domain (FRB) binding-deficient rapamycin analogue failed to elicit both the nanomolar and micromolar inhibitions of growth and mTOR signaling, implicating FRB binding in both actions. Biochemical assays indicated that CCI-779 and rapamycin directly inhibited mTOR kinase activity with IC(50) values of 1.76 +/- 0.15 and 1.74 +/- 0.34 micromol/L, respectively. Interestingly, a CCI-779-resistant mTOR mutant (mTOR-SI) displayed an 11-fold resistance to the micromolar CCI-779 in vitro (IC(50), 20 +/- 3.4 micromol/L) and conferred a partial protection in cells exposed to micromolar CCI-779. Treatment of cancer cells with micromolar but not nanomolar concentrations of CCI-779 caused a marked decline in global protein synthesis and disassembly of polyribosomes. The profound inhibition of protein synthesis was accompanied by rapid increase in the phosphorylation of translation elongation factor eEF2 and the translation initiation factor eIF2 alpha. These findings suggest that high-dose CCI-779 inhibits mTOR signaling through an FKBP12-independent mechanism that leads to profound translational repression. This distinctive high-dose drug effect could be directly related to the antitumor activities of CCI-779 and other rapalogues in human cancer patients.[1]

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Researchers study the rapamycin analogue CCI-779, alone or with chemotherapy, as an inhibitor of proliferation of the human prostate cancer cell lines PC-3 and DU145. The PTEN and phospho-Akt/PKB status and the effect of CCI-779 on phosphorylation of ribosomal protein S6 were evaluated by immunostaining and/or Western blotting. Expression of phospho-Akt/PKB in PTEN mutant PC-3 cells and xenografts was higher than in PTEN wild-type DU145 cells. Phosphorylation of S6 was inhibited by CCI-779 in both cell lines. Cultured cells were treated weekly with mitoxantrone or docetaxel for two cycles, and CCI-779 or vehicle was given between courses. Growth and clonogenic survival of both cell lines were inhibited in a dose-dependent manner by CCI-779, but there were minimal effects when CCI-779 was given between courses of chemotherapy. [2]

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Lymphoblasts from adult patients with precursor B ALL were cultured on bone marrow stroma and were treated with CCI-779, a second generation MTI. Treated cells showed a dramatic decrease in cell proliferation and an increase in apoptotic cells, compared to untreated cells. We also assessed the effect of CCI-779 in a NOD/SCID xenograft model. We treated a total of 68 mice generated from the same patient samples with CCI-779 after establishment of disease. Animals treated with CCI-779 showed a decrease in peripheral-blood blasts and in splenomegaly. In dramatic contrast, untreated animals continued to show expansion of human ALL. We performed immunoblots to validate the inhibition of the mTOR signaling intermediate phospho-S6 in human ALL, finding down-regulation of this target in xenografted human ALL exposed to CCI-779. We conclude that MTIs can inhibit the growth of adult human ALL and deserve close examination as therapeutic agents against a disease that is often not curable with current therapy.[3]

Cells are implanted in matrigel for the creation of xenografts; matrigel is stored at −20°C and thawed on ice at 4°C for 3 hours prior to use. After being gently resuspended in 1 mL of PBS, the cells are incubated for 5 minutes on ice. Cells are transferred to the tube containing 1 mL of matrigel using a prechilled pipette, and the cell concentration is adjusted to 3×107/mL. Using a 25-gauge needle, the cells (3×106 in 0.1 mL) are injected s.c. into the mice's flanks. When xenografts grew to a size of about 5 mm in diameter, animals are assorted randomLy into groups of 10 mice. The following experiments are conducted: Mice bearing PC-3 tumors are treated with CCI-779 (1, 5, 10, and 20 mg per kg per day), or vehicle solution for 3 or 5 days per week for 3 weeks. Mice bearing DU145 tumors are only treated with CCI-779 (20 mg per kg per day) or vehicle solution for 3 weeks. Mice bearing PC-3 tumors receive the following treatments: (a) control, vehicle solution for CCI-779; (b) chemotherapy alone, mitoxantrone 1.5 mg/kg or docetaxel 10 mg/kg is injected i.p. weekly for 3 doses; (c) CCI-779 alone, 5 or 10 mg/kg is injected i.p. daily, three times a week for 3 weeks; (4) chemotherapy followed by CCI-779.

Absorption, Distribution and Excretion
Infused intravenous over 30 - 60 minutes. Cmax is typically observed at the end of infusion
Excreted predominantly in feces (76%), 4.6% of drug and metabolites recovered in urine. 17% of drug was not recovered by either route following a 14-day sample collection.
172 L in whole blood of cancer patients; both temsirolimus and sirolimus are extensive distributed partitioned into formed blood elements
16.2 L/h (22%)
Following administration of a single 25 mg dose of temsirolimus in patients with cancer, mean temsirolimus Cmax in whole blood was 585 ng/mL (coefficient of variation, CV =14%), and mean AUC in blood was 1627 ng.hr/mL (CV=26%). Typically Cmax occurred at the end of infusion. Over the dose range of 1 mg to 25 mg, temsirolimus exposure increased in a less than dose proportional manner while sirolimus exposure increased proportionally with dose. Following a single 25 mg intravenous dose in patients with cancer, sirolimus AUC was 2.7-fold that of temsirolimus AUC, due principally to the longer half-life of sirolimus.
Following a single 25 mg intravenous dose, mean steady-state volume of distribution of temsirolimus in whole blood of patients with cancer was 172 liters. Both temsirolimus and sirolimus are extensively partitioned into formed blood elements.
Following a single 25 mg dose of temsirolimus in patients with cancer, temsirolimus mean (CV) systemic clearance was 16.2 (22%) L/hr.
It is not known whether temsirolimus is excreted into human milk...
Following IV administration of a single radiolabeled dose of temsirolimus, approximately 78% of the total radioactivity is recovered in feces and 4.6% in urine within 14 days.

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Metabolism / Metabolites
Primarily metabolized by cytochrome P450 3A4 in the human liver. Sirolimus, an equally potent metabolite, is the primary metabolite in humans following IV infusion. Other metabolic pathways observed in in vitro temsirolimus metabolism studies include hydroxylation, reduction and demethylation.
Sirolimus, an active metabolite of temsirolimus, is the principal metabolite in humans following intravenous treatment. The remainder of the metabolites account for less than 10% of radioactivity in the plasma.
Temsirolimus is metabolized by hydrolysis to sirolimus, the principal active metabolite. Both temsirolimus and sirolimus also are metabolized by cytochrome P-450 (CYP) isoenzyme 3A4. Although temsirolimus is metabolized to sirolimus, temsirolimus itself exhibits antitumor activity and is not considered a prodrug.
The in vitro metabolism of temsirolimus, (rapamycin-42-[2,2-bis-(hydroxymethyl)]-propionate), an antineoplastic agent, was studied using human liver microsomes as well as recombinant human cytochrome P450s, namely CYP3A4, 1A2, 2A6, 2C8, 2C9, 2C19, and 2E1. Fifteen metabolites were detected by liquid chromatography (LC)-tandem mass spectrometry (MS/MS or MS/MS/MS). CYP3A4 was identified as the main enzyme responsible for the metabolism of the compound. Incubation of temsirolimus with recombinant CYP3A4 produced most of the metabolites detected from incubation with human liver microsomes, which was used for large-scale preparation of the metabolites. By silica gel chromatography followed by semipreparative reverse-phase high-performance liquid chromatography, individual metabolites were separated and purified for structural elucidation and bioactivity studies. The minor metabolites (peaks 1-7) were identified as hydroxylated or desmethylated macrolide ring-opened temsirolimus derivatives by both positive and negative mass spectrometry (MS) and MS/MS spectroscopic methods. Because these compounds were unstable and only present in trace amounts, no further investigations were conducted. Six major metabolites were identified as 36-hydroxyl temsirolimus (M8), 35-hydroxyl temsirolimus (M9), 11-hydroxyl temsirolimus with an opened hemiketal ring (M10 and M11), N- oxide temsirolimus (M12), and 32-O-desmethyl temsirolimus (M13) using combined LC-MS, MS/MS, MS/MS/MS, and NMR techniques. Compared with the parent compound, these metabolites showed dramatically decreased activity against LNCaP cellular proliferation.

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Biological Half-Life
Temsirolimus exhibits a bi-exponential decline in whole blood concentrations and the mean half-lives of temsirolimus and sirolimus were 17.3 hr and 54.6 hr, respectively.
Temsirolimus exhibits a bi-exponential decline in whole blood concentrations and the mean half-lives of temsirolimus and sirolimus were 17.3 hr and 54.6 hr, respectively.

Hepatotoxicity
Serum aminotransferase elevations occur in 30% to 40% and alkaline phosphatase in 60% to 70% of patients receiving temsirolimus, but the abnormalities are usually mild, asymptomatic and self-limiting, rarely requiring dose modification or discontinuation. Elevations of liver enzymes above 5 times the upper limit of normal occur in only 1% to 3% of patients. Since approval and wide spread clinical use, there have been no case reports of clinically apparent liver injury attributed to temsirolimus use. Temsirolimus, like sirolimus, is immunosuppressive, and reactivation of hepatitis B is considered a possible complication of therapy. Yet despite more than 10 years of clinical use, there have been no reports of reactivation of hepatitis B attributed to temsirolimus therapy. Thus, acute liver injury with jaundice due to temsirolimus is probably quite rare, if it occurs at all. Hypersensitivity reactions to temsirolimus infusions are not uncommon (for which reason premedication with an antihistamine is recommended) and instances of Stevens Johnson syndrome have been reported.
Likelihood score: E* (unproven but suspected rare cause of clinically apparent liver injury).

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Effects During Pregnancy and Lactation
◉ Summary of Use during Lactation
Temsirolimus is a prodrug of sirolimus. Because no information is available on the use of temsirolimus or sirolimus during breastfeeding, an alternate drug may be preferred, especially while nursing a newborn or preterm infant. The manufacturer recommends that breastfeeding be discontinued during temsirolimus therapy and for 3 weeks following the last dose.
◉ Effects in Breastfed Infants
Relevant published information was not found as of the revision date.
◉ Effects on Lactation and Breastmilk
Relevant published information was not found as of the revision date.

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Protein Binding
87% bound to plasma proteins in vitro at a concentration of 100 ng/ml

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Interactions
CYP3A4 inhibitors: Potential pharmacokinetic interaction (increased plasma concentrations of the principal active metabolite sirolimus). Concomitant use with a potent CYP3A4 inhibitors should be avoided; if no alternative is available, consideration should be given to temsirolimus dosage adjustment.
CYP3A4 inducers: Potential pharmacokinetic interaction (decreased plasma concentrations of the principal active metabolite sirolimus). Concomitant use with potent CYP3A4 inducers should be avoided; if no alternative is available, consideration should be given to temsirolimus dosage adjustment.
Angioedema-type reactions observed during concomitant therapy with angiotensin-converting enzyme (ACE) inhibitors. Caution is advised.
Increased risk of intracerebral bleeding in patients receiving concomitant therapy. Caution is advised.
For more Interactions (Complete) data for Temsirolimus (17 total), please visit the HSDB record page.

[1]. A new pharmacologic action of CCI-779 involves FKBP12-independent inhibition of mTOR kinase activity and profound repression of global protein synthesis. Cancer Res, 2008, 68(8), 2934-2943.

[2]. Effects of the mammalian target of rapamycin inhibitor CCI-779 used alone or with chemotherapy on human prostate cancer cells and xenografts. Cancer Res, 2005, 65(7), 2825-2831.

[3]. The mTOR inhibitor CCI-779 induces apoptosis and inhibits growth in preclinical models of primary adult human ALL. Blood, 2006, 107(3), 1149-1155.

[4]. Antitumor activity of the rapamycin analog CCI-779 in human primitive neuroectodermal tumor/medulloblastoma models as single agent and in combination chemotherapy. Cancer Res, 2001, 61(4), 1527-1532.

[5]. Inhibition of mTOR induces autophagy and reduces toxicity of polyglutamine expansions in fly and mouse models of Huntington disease. Nat Genet. 2004 Jun;36(6):585-95. Epub 2004 May 16.

[6]. In vivo antitumor effects of the mTOR inhibitor CCI-779 against human multiple myeloma cells in a xenograft model. Blood. 2004 Dec 15;104(13):4181-7. Epub 2004 Aug 10.

[7]. A case study of an integrative genomic and experimental therapeutic approach for rare tumors: identification of vulnerabilities in a pediatric poorly differentiated carcinoma. Genome Med. 2016 Oct 31;8(1):116.

[8]. Temsirolimus activates autophagy and ameliorates cardiomyopathy caused by lamin A/C gene mutation. Sci Transl Med. 2012 Jul 25; 4(144): 144ra102.

Therapeutic Uses
Temsirolimus is indicated for the treatment of advanced renal cell carcinoma. /Included in US product label/

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Drug Warnings
Anaphylaxis, dyspnea, flushing, and chest pain have been reported. Temsirolimus should be used with caution in patients with known hypersensitivity to the drug or its metabolites (eg, sirolimus), polysorbate 80, or any other ingredient in the formulation.
Pretreatment with an antihistamine prior to each dose of temsirolimus is recommended to prevent hypersensitivity reactions. Temsirolimus should be used with caution in patients with known hypersensitivity to antihistamines or with conditions requiring avoidance of antihistamines.
The safety and pharmacokinetics of temsirolimus were evaluated in a dose escalation phase 1 study in 110 patients with normal or varying degrees of hepatic impairment. Patients with baseline bilirubin >1.5 x ULN experienced greater toxicity than patients with baseline bilirubin /= grade 3 adverse reactions and deaths, including deaths due to progressive disease, were greater in patients with baseline bilirubin >1.5 x ULN. temsirolimus is contraindicated in patients with bilirubin >1.5 x ULN due to increased risk of death. Use caution when treating patients with mild hepatic impairment. Concentrations of temsirolimus and its metabolite sirolimus were increased in patients with elevated AST or bilirubin levels. If temsirolimus must be given in patients with mild hepatic impairment (bilirubin >1 - 1.5 x ULN or AST >ULN but bilirubin No clinical studies were conducted with temsirolimus in patients with decreased renal function. Less than 5% of total radioactivity was excreted in the urine following a 25 mg intravenous dose of (14)C-labeled temsirolimus in healthy subjects. Renal impairment is not expected to markedly influence drug exposure, and no dosage adjustment of temsirolimus is recommended in patients with renal impairment.
For more Drug Warnings (Complete) data for Temsirolimus (29 total), please visit the HSDB record page.

C56H87NO16

1030.29

1029.602

C, 65.28; H, 8.51; N, 1.36; O, 24.85

162635-04-3

162635-04-3

6918289

White to off-white solid powder

1.2±0.1 g/cm3

1048.4±75.0 °C at 760 mmHg

99-101ºC

587.8±37.1 °C

0.0±0.6 mmHg at 25°C

1.554

2.96

4

16

11

73

2010

15

O(C([H])([H])[H])[C@@]1([H])[C@@]([H])(C([H])([H])C([H])([H])[C@@]([H])(C([H])([H])[C@@]([H])(C([H])([H])[H])[C@]2([H])C([H])([H])C([C@@]([H])(C([H])=C(C([H])([H])[H])[C@]([H])([C@]([H])(C([C@]([H])(C([H])([H])[H])C([H])([H])[C@]([H])(C([H])([H])[H])C([H])=C([H])C([H])=C([H])C([H])=C(C([H])([H])[H])[C@]([H])(C([H])([H])[C@]3([H])C([H])([H])C([H])([H])[C@@]([H])(C([H])([H])[H])[C@@](C(C(N4C([H])([H])C([H])([H])C([H])([H])C([H])([H])[C@@]4([H])C(=O)O2)=O)=O)(O[H])O3)OC([H])([H])[H])=O)OC([H])([H])[H])O[H])C([H])([H])[H])=O)C1([H])[H])OC(C(C([H])([H])[H])(C([H])([H])O[H])C([H])([H])O[H])=O |c:35,66,70,t:62|

CBPNZQVSJQDFBE-FUXHJELOSA-N

InChI=1S/C56H87NO16/c1-33-17-13-12-14-18-34(2)45(68-9)29-41-22-20-39(7)56(67,73-41)51(63)52(64)57-24-16-15-19-42(57)53(65)71-46(30-43(60)35(3)26-38(6)49(62)50(70-11)48(61)37(5)25-33)36(4)27-40-21-23-44(47(28-40)69-10)72-54(66)55(8,31-58)32-59/h12-14,17-18,26,33,35-37,39-42,44-47,49-50,58-59,62,67H,15-16,19-25,27-32H2,1-11H3/b14-12+,17-13+,34-18+,38-26+/t33-,35-,36-,37-,39-,40+,41+,42+,44-,45+,46+,47-,49-,50+,56-/m1/s1

[(1R,2R,4S)-4-[(2R)-2-[(1R,9S,12S,15R,16E,18R,19R,21R,23S,24E,26E,28E,30S,32S,35R)-1,18-dihydroxy-19,30-dimethoxy-15,17,21,23,29,35-hexamethyl-2,3,10,14,20-pentaoxo-11,36-dioxa-4-azatricyclo[30.3.1.04,9]hexatriaconta-16,24,26,28-tetraen-12-yl]propyl]-2-methoxycyclohexyl] 3-hydroxy-2-(hydroxymethyl)-2-methylpropanoate

CCI-779; CCI779; Temsirolimus; Torisel; 162635-04-3; 624KN6GM2T; DTXSID2040945; UNII-624KN6GM2T; WAY-CCI 779; CCI 779; NSC 683864; NSC683864; NSC-683864; Temsirolimus; 624KN6GM2T; DTXSID2040945; UNII-624KN6GM2T; WAY-CCI 779; Brand name: Torisel

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

Note: (1). This product requires protection from light (avoid light exposure) during transportation and storage.  (2). Please store this product in a sealed and protected environment (e.g. under nitrogen), 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)

DMSO: ~75 mg/mL (~72.8 mM)
Water: <1 mg/mL
Ethanol: ~75 mg/mL (~72.8 mM)

Solubility in Formulation 1: ≥ 5 mg/mL (4.85 mM) (saturation unknown) in 10% EtOH + 40% PEG300 + 5% Tween80 + 45% Saline (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 50.0 mg/mL clear EtOH stock solution to 400 μL PEG300 and mix evenly; then add 50 μL Tween-80 to the above solution and mix evenly; then add 450 μL normal saline to adjust the volume to 1 mL.
Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH₂ O to obtain a clear solution.

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Solubility in Formulation 2: ≥ 5 mg/mL (4.85 mM) (saturation unknown) in 10% EtOH + 90% Corn Oil (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 50.0 mg/mL clear EtOH stock solution to 900 μL of corn oil and mix well.

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Solubility in Formulation 3: ≥ 2.08 mg/mL (2.02 mM) (saturation unknown) in 10% DMSO + 40% PEG300 + 5% Tween80 + 45% Saline (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 20.8 mg/mL clear DMSO stock solution to 400 μL PEG300 and mix evenly; then add 50 μL Tween-80 to the above solution and mix evenly; then add 450 μL normal saline to adjust the volume to 1 mL.
Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH₂ O to obtain a clear solution.

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Solubility in Formulation 4: ≥ 2.08 mg/mL (2.02 mM) (saturation unknown) in 10% DMSO + 90% Corn Oil (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 20.8 mg/mL clear DMSO stock solution to 900 μL corn oil and mix evenly.

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Solubility in Formulation 5: 30% PEG400+0.5% Tween80+5% propylene glycol:10mg/mL

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