HDAC1 ( IC50 = 36 nM ); HDAC2 ( IC50 = 47 nM ); HDAC4 ( IC50 = 510 nM ); HDAC6 ( IC50 = 14000 nM )

Romidepsin treatment potently inhibits the neovascularization of chick embryos and adult mice in the Matrigel plug assay. AThe median survival times of mice with U-937 lymphoma are 30.5 (0.56 mg/kg) and 33 days (0.32 mg/kg), respectively, after receiving romidepsin at 0.1–1 mg/kg twice a week (compared to 20 days in control mice).

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FK228 showed potent antiangiogenic activities in vivo [4]
The antiangiogenic activity of FK228 was also investigated in vivo, using a CAM assay. As shown in Figure 3, the FK228 produced a significant decrease in the developmental of angiogenesis in a chick embryo without any sign of thrombosis and hemorrhage. The inhibition of the angiogenesis of RA used as the positive control was 70.83 ± 6.22% (n = 32) and that of empty coverslips was 4.43 ± 4.43% (n = 45). However, FK228 completely inhibited the neovascularization of chick embryo (100 ± 0.00%, n = 16) without affecting any preexisting vessels. The antiangiogenic activity of FK228 was also confirmed by performing an established in vivo angiogenesis model, the Matrigel plug assay in an adult mouse.29 The Matrigel was injected subcutaneously into C57/BL6J mice, solidified in vivo and removed from the mice at 5 days after implantation. For histologic examination, image analysis was performed. As shown in Figure 4, control plugs in which the Matrigel was injected with the heparin alone had little vessel, but bFGF (100 ng/ml) enriched abundant vessels inside the plugs. However, FK228 strongly inhibited the bFGF-induced angiogenesis. Hb contents of the Matrigel plugs were also measured to quantify the functional vasculature. Control plugs showed 0.55 ± 0.44 g/dl of Hb (n = 5) and bFGF-containing Matrigel showed 12.94 ± 2.82 g/dl (n = 6). However, FK228 markedly reduced the Hb levels to 1.36 ± 0.25 g/dl (n = 5). These results demonstrated that FK228 potently inhibits angiogenesis in vivo and in vitro.

In the enzyme assay, 90 μL of the HDAC enzyme fraction extracted from 293T cells overexpressing HDAC1 or HDAC2 in the presence of increasing concentrations of Romidepsin is mixed with 10 μL of [³H]acetyl-labeled histones (25,000 cpm/10 μg). The mixture is then incubated at 37 °C for 15 minutes. For a minimum of 60 minutes, the enzyme reaction is linear. The addition of 10 μL of concentrated HCl stops the reaction. For the purpose of determining radioactivity, 0.9 mL of the solvent layer is added to 5 mL of aqueous counting scintillant II solution after the released [³H]acetic acid is extracted with 1 mL of ethylacetate. At least three separate independent dose-response curves are used to calculate the IC50 values.

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[3H]-Methylthymidine assay [4]
To measure cell proliferation, the BAECs were placed in a 24-well culture plates at a density of 3 × 104 cells. Then, Romidepsin (FK228) was added and incubated under normoxic or hypoxic condition for 24 hr. Then 1 μCi [3H]-methylthymidine (25 mCi/mmol) was added at the final 4 hr before the assay. The cells were washed with PBS and fixed with methanol on ice for 30 min. Unincorporated [3H]-methylthymidine was removed by washing with 10% trichloroacetic acid (TCA). After the overnight incubation with 10% TCA, the cells were solubilized in 0.2 M NaOH and 0.1% SDS at 37°C for 1 hr. The counts per minute were determined with a liquid scintillation counter and the values were expressed as mean cpm (of triplicate). Each sample was assayed in triplicate and the experiment was repeated twice independently.

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HDAC activity assay [4]
The BAECs (1 × 107 to 5 × 107) treated with Romidepsin (FK228) (1–10 ng/ml) for 24 hr were harvested and lysed in 1 ml HDAC buffer (50 mM Tris-HCl, pH 8.0, 150 mM NaCl, 10% glycerol, 0.5% Triton X-100). [3H]-Acetylated histones were incubated with 100 μl of prepared crude cell extract for 2 hr at 37°C. The reaction was quenched with 1 M HCl and 0.16 M acetic acid. Released [3H]acetate was extracted with 600 μl of ethyl acetate and quantified by a scintillation counter.

In 96-well plates, cells are exposed to different doses of romidepsin for a duration of 72 hours. For four hours, 20 μL of a 5 mg/mL MTT solution in PBS is added to each well. To dissolve the formazan crystals, 170 μL of DMSO is added to each well after the medium has been removed. At 540 nm, the absorbance is calculated. Moreover, trypan blue is added to the cells, and the quantity of transparent (living) and blue (dead) cells is counted in a hemocytometer. Cells are incubated in a propidium iodide staining solution containing 0.05 mg/mL propidium iodide, 1 mM EDTA, 0.1% Triton X-100, and 1 mg/mL RNase A in PBS for 30 minutes in order to perform a cell cycle analysis. The suspension is then examined using a Becton Dickinson FACScan after being run through a nylon mesh filter.

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Cell growth assay [4]
The endothelial cell growth was determined by 3-(4,5-diphenylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay. The BAECs were grown at a density of 1.5 × 10~4 cells in a 24-well culture plates and incubated for 24 hr for stabilization. The medium was replaced with 1.5 ml fresh medium and Romidepsin (FK228) was added. After 24 and 72 hr, the medium was aspirated and the MTT assay was performed. The amount of MTT-formazan was determined as absorbance at 540 nm. Each sample was assayed in triplicate and the experiment was repeated twice independently.

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Tube formation assay [4]
Matrigel (250 μl in a concentration of 10 mg/ml) was placed in a 24-well culture plate and polymerized for 30 min at 37°C. The BAECs (5 × 10~5 cells) were seeded on the surface of the Matrigel. Then, the Romidepsin (FK228) (10 ng/ml) was added and incubated for 24 hr under normoxic or hypoxic condition. The morphologic changes in the cells were observed under a microscope and photographed at a ×40 magnification using ImagePro Plus software. Each sample was assayed in duplicate and the experiment was repeated twice independently.

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Chemomigration assay [4]
Chemomigration of the BAECs was performed using a 24-well transwell culture chamber with 8.0-μm-pore polycarbonate filter inserts. The lower side of the filter was coated with 10 μl type IV collagen (3 mg/ml) and air-dried for 1 hr. The BAECs (3 × 104 cells) were placed in the upper part of the filters and Romidepsin (FK228) (10 ng/ml) was applied to both sides of the filter. The cells were incubated at 37°C for 8 hr under normoxic or hypoxic condition and then fixed with methanol and stained with hematoxylin and eosin. The cells on the upper surface were carefully removed by rubbing with a cotton swab and mounted on slide glasses. The cell migration was determined by counting the whole cell numbers on a single filter using optical microscopy at a ×40 magnification. Each sample was assayed in duplicate and the experiment was repeated twice independently.

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Chemoinvasion assay [4]
The invasiveness of the BAECs was performed in vitro using a transwell chamber system with 8.0-μm-pore polycarbonate filter inserts. The lower side of the filter was coated with 10 μl of type IV collagen (3 mg/ml), whereas the upper side was coated with 10 μμl Matrigel (0.5 mg/ml). The BAECs (3 × 10~4 cells) were placed in the upper part of the filters and Romidepsin (FK228) (10 ng/ml) was treated in both parts. The cells were incubated at 37°C for 18 hr under normoxic or hypoxic condition and then fixed with methanol and stained with hematoxylin and eosin. The cell invasion was determined by counting the whole cell numbers in a single filter using optical microscopy at a ×40 magnification. Each sample was assayed in duplicate and the experiment was repeated twice independently.

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Cell adhesion assay [4]
A cell adhesion assay was performed as described below. Each well of a 24-well culture plate was coated with type IV collagen (5 μg/ml), vitronectin (1 μg/ml), fibronectin (1 μg/ml) or type I collagen (5 μg/ml) and incubated for 2 hr at 37°C. After washing with PBS, 3.0% BSA was added to each well for 1 hr to prevent nonspecific attachment. The BAECs (3 × 10~4 cells) suspended in serum-free media was added to each coated well and Romidepsin (FK228) (10 ng/ml) was added. After incubation at 37°C for 1 hr under normoxic or hypoxic condition, non-adherent cells were washed off by streaming PBS over the plate 3 times. The remaining adherent cells were stained with 1% crystal violet and washed with PBS several times to remove the excess dye. The stained crystal violet was eluted with 10% acetic acid and determined by scanning with an ELISA reader with a 600-nm filter. Each sample was assayed in triplicate and the experiment was repeated twice independently.

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Chorioallantoic membrane (CAM) assay [4]
Fertilized chick eggs were kept in a humidified incubator at 37°C for 3 days. About 2 ml of egg albumin was then removed with a hypodermic needle allowing the CAM and yolk sac to drop away from the shell membrane. On day 3.5, the shell was punched out and removed and the shell membrane peeled away. At the stage of a 4.5-day old chick embryo, an Romidepsin (FK228)-loaded (50 ng/egg) Thermanox coverslip was air-dried and applied to the CAM surface. Two days later, 2 ml of 10% fat emulsion was injected into the chorioallantois and the CAM was observed under a microscope. Because retinoic acid (RA) is known as an antiangiogenic compound, 1 μg/egg RA was used as a positive control for antiangiogenic responses. When the CAM treated with Romidepsin (FK228) had an avascular zone to a similar degree as RA-treated CAM, the response was scored as positive and calculated based on the percentage of positive eggs to the total number of eggs tested. The experiment was repeated 3 times and more than 20 eggs were used each time.

Male scid mice inoculated i.p. with U-937 cells
~1 mg/kg once or twice a week
Treated i.p.

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In vivo mouse Matrigel plug assay [4]
The mouse Matrigel plug assay was performed as previously described. Briefly; C57BL/6J mice were injected subcutaneously with 0.5 ml Matrigel containing the indicated amount of Romidepsin (FK228) (0.7 μg/ml) with bFGF (100 ng/ml) and heparin (10 U/ml). The injected Matrigel rapidly formed a single, solid gel plug. After 5 days, mice were sacrificed and the Matrigel plug was removed, fixed with 3.7% formaldehyde/PBS and embedded in paraffin. The plugs were then sectioned and examined with Masson-Trichrome staining for microscopic observation. To quantify the formation of new blood vessel, the amount of hemoglobin (Hb) was measured using Drabkin reagent kit 525. The concentration of Hb was calculated from a known amount of Hb provided by the kit in parallel according to the supplier's protocol. The experiment was repeated 3 or 4 times, independently.

Absorption, Distribution and Excretion
Romidepsin exhibited linear pharmacokinetics at standard doses.
44.5L
8.4L/h

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Metabolism / Metabolites
Romidepsin undergoes extensive hepatic metabolism in vitro primarily by CYP3A4 with minor contribution from CYP3A5, CYP1A1, CYP2B6 and CYP2C19.

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Biological Half-Life
Approximately 3 hours

Hepatotoxicity
In clinical trials of romidepsin in patients with CTLC and PTLC, the rates of serum enzyme elevations during therapy ranged from 7% to 20%, but the abnormalities were usually transient and mild and did not require dose modifications. Serum ALT elevations above 5 times ULN occurred in 6% of patients. In the preregistration clinical trials of romidepsin, there were no reports of hepatitis, jaundice or clinically apparent liver injury among the treated subjects. Romidepsin has had limited clinical use, but there is no evidence that it is associated with significant liver injury.
Romidepsin also has immunomodulatory activities and has been reported to cause reactivation of latent DNA viruses including Epstein-Barr, varicella zoster and hepatitis B virus. Reactivation of hepatitis B occurred in a patient who was initially negative for HBsAg, but reactive for anti-HBc and anti-HBs. Nevertheless, the clinical features of hepatitis B reactivation were mild and responded to oral antiviral therapy. In patients with EBV associated lymphoma, romidepsin has been associated with severe reactivation of EBV infection and acute hepatitis that can be severe and even fatal.
Likelihood score: C (probable cause of clinically apparent liver injury, which can be due to reactivation of hepatitis B or EBV infection).

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Protein Binding
Highly protein bound in plasma (92%-94%)

[1]. Cancer Res. 2002 Sep 1;62(17):4916-21.

[2]. Br J Cancer. 2000 Sep;83(6):817-25.

[3]. Mol Cancer Ther. 2002 Sep;1(11):937-41.

[4]. Int J Cancer. 2002 Jan 20;97(3):290-6.

[5]. Biochem Pharmacol. 2002 Oct 1;64(7):1079-90.

[6]. Blood. 2003 Jul 15;102(2):652-8.

[7]. Cancer Res. 2006 Jul 15;66(14):7317-25.

[8]. Clin Cancer Res. 2008 Jan 15;14(2):549-58.

[9]. Clin Cancer Res. 2010 Jan 15;16(2):554-65.

Romidepsin is a cyclodepsipeptide consisting of the cyclic disulfide of (2Z)-2-aminobut-2-enoyl, L-valyl, (3S,4E)-3-hydroxy-7-sulfanylhept-4-enoyl, D-valyl and D-cysteinyl residues coupled in sequence and cyclised head-to tail. It has a role as an antineoplastic agent and an EC 3.5.1.98 (histone deacetylase) inhibitor. It is a cyclodepsipeptide, an organic disulfide and a heterocyclic antibiotic.
Romidepsin is a drug that has been approved by the U.S. Food and Drug Administration (FDA) under the brand name Istodax for the treatment of a certain type of cancer. Romidepsin is also being studied as an investigational drug as part of a strategy to cure HIV infection.As an HIV investigational drug, romidepsin belongs to a group of drugs called latency-reversing agents.
Romidepsin is a selective inhibitor of histone deacetylase, approved in the US in 2009 for the treatment of cutaneous T-cell lymphoma (CTCL) in patients who have received at least one prior systemic therapy.
Romidepsin is a Histone Deacetylase Inhibitor. The mechanism of action of romidepsin is as a Histone Deacetylase Inhibitor, and Bile Salt Export Pump Inhibitor, and Organic Anion Transporting Polypeptide 1B1 Inhibitor, and Organic Anion Transporting Polypeptide 1B3 Inhibitor, and Organic Anion Transporter 1 Inhibitor, and Organic Cation Transporter 2 Inhibitor.
Romidepsin is an intravenously administered histone deacetylase inhibitor and antineoplastic agent that is approved for use in refractory or relapsed cutaneous and peripheral T cell lymphomas. Romidepsin is associated with modest rate of minor serum enzyme elevations during therapy but has not been linked to cases of clinically apparent liver injury, although it has been reported to cause reactivation of hepatitis B.
Istodax has been reported in Humicola fuscoatra and Chromobacterium violaceum with data available.
Romidepsin is a bicyclic depsipeptide antibiotic isolated from the bacterium Chromobacterium violaceum with antineoplastic activity. After intracellular activation, romidepsin binds to and inhibits histone deacetylase (HDAC), resulting in alterations in gene expression and the induction of cell differentiation, cell cycle arrest, and apoptosis. This agent also inhibits hypoxia-induced angiogenesis and depletes several heat shock protein 90 (Hsp90)-dependent oncoproteins.

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Drug Indication
Romidepsin is indicated for the treatment of cutaneous T-cell lymphoma (CTCL) in adult patients who have received at least one prior systemic therapy.
FDA Label
treatment of peripheral T-cell lymphoma (PTCL),
Treatment of peripheral T-cell lymphoma (nodal, other extranodal and leukaemic/disseminated)

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Mechanism of Action
Romidepsin is a prodrug, where it becomes active once taken up into the cell. The active metabolite has a free thiol group, which interacts with zinc ions in the active site of class 1 and 2 HDAC enzymes, resulting in inhibition of its enzymatic activity. Certain tumors have over expressed HDACs and downregulated/mutated histone acetyltransferases. This imbalance of HDAC relative to histone acetyltransferase can lead to a decrease in regulatory genes, ensuing tumorigenesis. Inhibition of HDAC may restore normal gene expression in cancer cells and result in cell cycle arrest and apoptosis.

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FK228 is a histone deacetylase (HDAC) inhibitor, the molecular mechanism of inhibition of which has been unknown. Here we show that reduction of an intramolecular disulfide bond of FK228 greatly enhanced its inhibitory activity and that the disulfide bond was rapidly reduced in cells by cellular reducing activity involving glutathione. Computer modeling suggests that one of the sulfhydryl groups of the reduced form of FK228 (redFK) interacts with the active-site zinc, preventing the access of the substrate. HDAC1 and HDAC2 were more strongly inhibited by redFK than HDAC4 and HDAC6. redFK was less active than FK228 in inhibiting in vivo HDAC activity, due to rapid inactivation in medium and serum. Thus, FK228 serves as a stable prodrug to inhibit class I enzymes and is activated by reduction after uptake into the cells. The glutathione-mediated activation also implicates its clinical usefulness for counteracting glutathione-mediated drug resistance in chemotherapy. [1]

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Depsipeptide, FR901228, a novel cyclic peptide inhibitor of histone deacetylase with a unique cytotoxicity profile is currently in phase I clinical trials. Here we demonstrate that, in addition to G2/M arrest, FR901228 causes G1 arrest with Rb hypophosphorylation. In vitro kinase assays demonstrated no direct inhibition of CDK activity, however, an inhibition was observed in CDKs extracted from cells exposed to FR901228. Cyclin D1 protein disappeared between 6 and 12 hours after treatment with FR901228, whereas cyclin E was upregulated. While it did not induce wt p53, FR901228 did induce p21(WAF1/CIP1)in a p53-independent manner. Cell clones lacking p21 were not arrested in G1 phase, but continued DNA synthesis and were arrested in G2/M phase following FR901228 treatment. Finally, FR901228 blunted ERK-2/MAPK activation by EGF whereas early signal transduction events remained intact since overall cellular tyrosine phosphorylation after EGF stimulation was unaffected. Thus, FR901228, while not directly inhibiting kinase activity, causes cyclin D1 downregulation and a p53-independent p21 induction, leading to inhibition of CDK and dephosphorylation of Rb resulting in growth arrest in the early G1 phase. In contrast to the G1 arrest, the G2/M arrest is p21-independent, but is associated with significant cytotoxicity. [2]

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By preventing deacetylation of histones, histone deacetylase inhibitors (HDIs) transcriptionally induce p21. Here we show that the HDIs sodium butyrate (Bu), trichostatin A (TSA) and depsipeptide (FR901228) all induced p21, but only TSA and FR901228 caused mitotic arrest (in addition to arrest in G1 and G2). The ability to cause mitotic arrest correlated with the higher cytotoxicity of these compounds. Although causing mitotic arrest, TSA and FR901228 (unlike paclitaxel) did not affect tubulin polymerization. Unlike FR9012208, TSA caused acetylation of tubulin at lysine 40; both soluble tubulin and microtubules were acetylated. Whereas the induction of p21 reached a maximum by 8 h, tubulin was maximally acetylated after only 1 h of TSA treatment. Tubulin acetylation was detectable after treatment with 12-25 ng/ml TSA although acetylation plateaued at 50 ng/ml TSA, coinciding with G2-M arrest, appearance of cells with a sub-2N DNA content, poly(ADP-ribose) polymerase cleavage, and rapid cell death. We conclude that HDIs have differential effects on non-histone deacetylases and that rapid acetylation of tubulin caused by TSA is a marker of nontranscriptional effects of TSA. [3]

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FK228 (formerly FR901228) was recently isolated from Chromobacterium violaceum as a potent antitumor agent and its biologic target protein was identified as histone deacetylase (HDAC). Because of its unique chemical structure (i.e., bicyclic depsipeptide) and activity profile in the National Cancer Institute's developmental therapeutics program, FK228 is currently in a phase I clinical trial for cancer therapy. In the present study, we investigated the antiangiogenic activity of FK228 in vivo and in vitro. FK228 potently blocked the hypoxia-stimulated proliferation, invasion, migration, adhesion and tube formation of bovine aortic endothelial cells at the same concentration at which the agent inhibited the HDAC activity of cells. In addition, FK228 inhibited the neovascularization of chick embryo and that of adult mice in the Matrigel plug assay. Interestingly, the expression of angiogenic-stimulating factors such as vascular endothelial growth factor or kinase insert domain receptor were suppressed by FK228, whereas that of angiogenic-inhibiting factors such as von Hippel Lindau and neurofibromin2 were induced, suggesting that a gene-transcription effect was involved in the inhibition of angiogenesis by FK228. These results indicate that FK228 is a novel antiangiogenic agent and may suppress tumor expansion, at least in part, by the inhibition of neovascularization. [4]

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FK228 [(E)-(1S,4S,10S,21R)-7-[(Z)-ethylidene]-4,21-diisopropyl-2-oxa-12,13-dithia-5,8,20,23-tetraazabicyclo-[8,7,6]-tricos-16-ene-3,6,9,19,22-pentanone; FR901228, depsipeptide] is a novel histone deacetylase inhibitor that shows therapeutic efficacy in Phase I trials of patients with malignant lymphoma. However, its mechanism of action has not been characterized. In this study, we examined the in vitro and in vivo effects of FK228 on human lymphoma U-937 cells. FK228 very strongly inhibited the growth of U-937 cells with an IC(50) value of 5.92 nM. In a scid mouse lymphoma model, mice treated with FK228 once or twice a week survived longer than control mice, with median survival times of 30.5 (0.56 mg/kg) and 33 days (0.32 mg/kg), respectively (vs. 20 days in control mice). Remarkably, 2 out of 12 mice treated with FK228 (0.56 mg/kg once or twice a week) survived past the observation period of 60 days. The apoptotic population of U-937 cells time-dependently increased to 37.7% after 48 hr of treatment with FK228. In addition, FK228 induced G1 and G2/M arrest and the differentiation of U-937 cells to the CD11b(+)/CD14(+) phenotype. Expression of p21(WAF1/Cip1) and gelsolin mRNA increased up to 654- and 152-fold, respectively, after 24hr of treatment with FK228. FK228 caused histone acetylation in p21(WAF1/Cip1) promoter regions, including the Sp1-binding sites. In conclusion, (i) FK228 prolonged the survival time of scid mice in a lymphoma model, and (ii) the beneficial effects of FK228 on human lymphoma may be exerted through the induction of apoptosis, cell cycle arrest, and differentiation via the modulation of gene expression by histone acetylation. [5]

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Depsipeptide is in clinical trials for chronic lymphocytic leukemia (CLL) on the basis of earlier observations demonstrating selective in vitro activity in CLL. We sought to determine the relationship of histone H3 and H4 acetylation, inhibition of histone deacetylase, and apoptosis observed in CLL cells to justify a pharmacodynamic end point in these clinical trials. We demonstrate that in vitro depsipeptide induces histone H3 and H4 acetylation and histone deacetylase enzyme inhibition at concentrations corresponding to the LC50 (concentration producing 50% cell death) for cultured CLL cells (0.038 microM depsipeptide). The changes in histone acetylation are lysine specific, involving H4 K5, H4 K12, and H3 K9, and to a lesser extent H4 K8, but not H4 K16 or H3 K14. Depsipeptide-induced apoptosis is caspase dependent, selectively involving the tumor necrosis factor (TNF) receptor (extrinsic pathway) initiating caspase 8 and effector caspase 3. Activation of caspase 8 was accompanied by the down-regulation of cellular FLICE-inhibitory protein (c-FLIP, I-FLICE) without evidence of Fas (CD95) up-regulation. Changes in other apoptotic proteins, including Bcl-2, Bax, Mcl-1, and X-linked inhibitor of apoptosis (XIAP), were not observed. Our results demonstrate a relationship between target enzyme inhibition of histone deacetylase, histone H3 and H4 acetylation, and apoptosis involving the TNF-receptor pathway of apoptosis that is not used by other therapeutic agents in CLL. These data suggest use of histone H3 and H4 acetylation, inhibition of histone deacetylase, and down-regulation of [6]

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The proteasome inhibitor, bortezomib, and the histone deacetylase inhibitor, depsipeptide (FK228), up-regulate tumor death receptors. Therefore, we investigated whether pretreatment of malignant cells with these agents would potentiate natural killer (NK)-mediated tumor killing. NK cells isolated from healthy donors and patients with cancer were expanded in vitro and then tested for cytotoxicity against tumor cell lines before and after exposure to bortezomib or depsipeptide. In 11 of 13 (85%) renal cell carcinoma cell lines and in 16 of 37 (43%) other cancer cell lines, exposure to these drugs significantly increased NK cell-mediated tumor lysis compared with untreated tumor controls (P < 0.001). Furthermore, NK cells expanded from patients with metastatic renal cell carcinoma were significantly more cytotoxic against autologous tumor cells when pretreated with either bortezomib or depsipeptide compared with untreated tumors. Tumors sensitized to NK cell cytotoxicity showed a significant increase in surface expression of DR5 [tumor necrosis factor-related apoptosis-inducing ligand (TRAIL)-R2; P < 0.05]; in contrast, surface expression of MHC class I, MIC-A/B, DR4 (TRAIL-R1), and Fas (CD95) did not change. The enhanced susceptibility to NK cell killing was completely abolished by blocking TRAIL on NK cells, and partially abolished by blocking DR5 on tumor cells. These findings show that drug-induced sensitization to TRAIL could be used as a novel strategy to potentiate the anticancer effects of adoptively infused NK cells in patients with cancer. [7]

C24H36N4O6S2

540.7

540.207

C, 53.31; H, 6.71; N, 10.36; O, 17.75; S, 11.86

128517-07-7

5352062

White to off-white solid powder

1.2±0.1 g/cm3

942.8±65.0 °C at 760 mmHg

219-224°C

524.0±34.3 °C

0.0±0.3 mmHg at 25°C

1.529

0.95

4

8

2

36

905

4

S1C([H])([H])[C@]2([H])C(N([H])/C(=C(/[H])\C([H])([H])[H])/C(N([H])C([H])(C(=O)O[C@]([H])(C([H])=C([H])C([H])([H])C([H])([H])S1)C([H])([H])C(N([H])[C@@]([H])(C(N2[H])=O)C([H])(C([H])([H])[H])C([H])([H])[H])=O)C([H])(C([H])([H])[H])C([H])([H])[H])=O)=O |c:27|

OHRURASPPZQGQM-GCCNXGTGSA-N

InChI=1S/C24H36N4O6S2/c1-6-16-21(30)28-20(14(4)5)24(33)34-15-9-7-8-10-35-36-12-17(22(31)25-16)26-23(32)19(13(2)3)27-18(29)11-15/h6-7,9,13-15,17,19-20H,8,10-12H2,1-5H3,(H,25,31)(H,26,32)(H,27,29)(H,28,30)/b9-7+,16-6-/t15-,17-,19-,20+/m1/s1

(1S,4S,7Z,10S,16E,21R)-7-ethylidene-4,21-di(propan-2-yl)-2-oxa-12,13-dithia-5,8,20,23-tetrazabicyclo[8.7.6]tricos-16-ene-3,6,9,19,22-pentone

NSC 630176;FK228; FK 228; Romidepsin; Chromadax; 128517-07-7; Antibiotic FR 901228; FK-228; FR901228; FR-901228; FR 901228; NSC-630176; NSC-630176; depsipeptide; US trade name: Istodax.

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

Note: This product is not stable in solution, please use freshly prepared working solution for optimal results.

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

Solubility in Formulation 1: ≥ 2.08 mg/mL (3.85 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 2: 2.08 mg/mL (3.85 mM) in 10% DMSO + 90% (20% SBE-β-CD in Saline) (add these co-solvents sequentially from left to right, and one by one), suspension solution; with ultrasonication.
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 of 20% SBE-β-CD physiological saline solution and mix evenly.
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.

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Solubility in Formulation 3: ≥ 2.08 mg/mL (3.85 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 of corn oil and mix evenly..

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Solubility in Formulation 4: 1% DMSO+30% polyethylene glycol+1% Tween 80: 18mg/mL

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