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Treosulfan (NSC 39069; Treosulphan) is a novel and potent DNA alkylating agent with activity in ovarian cancer and other solid tumor types. Treosulfan is the byproduct of an immunosuppressive, myeloablative, and antineoplastic bifunctional sulfonate alkylating agent. Treosulfan undergoes a nonenzymatic conversion to L-diepoxybutane via a monoepoxide intermediate in a physiological setting. DNA fragmentation and apoptosis are caused by the production of DNA interstrand crosslinks and alkylation of DNA at guanine residues by the monoepoxide intermediate and L-diepoxybutane. At higher dosages, this substance also exhibits myeloablative and immunosuppressive effects.
Physicochemical Properties
| Molecular Formula | C6H14O8S2 |
| Molecular Weight | 278.30056 |
| Exact Mass | 278.013 |
| Elemental Analysis | C, 25.89; H, 5.07; O, 45.99; S, 23.04 |
| CAS # | 299-75-2 |
| Related CAS # | 299-75-2 (Treosulfan); 55-98-1 (Busulfan); 52-24-4 (Thiotepa, Girostan; AI3-24916; NSC-6396) |
| PubChem CID | 9882105 |
| Appearance | White to off-white solid powder |
| Density | 1.6±0.1 g/cm3 |
| Boiling Point | 607.0±55.0 °C at 760 mmHg |
| Melting Point | 216 °F (NTP, 1992) |
| Flash Point | 320.9±31.5 °C |
| Vapour Pressure | 0.0±3.9 mmHg at 25°C |
| Index of Refraction | 1.518 |
| LogP | -1.64 |
| Hydrogen Bond Donor Count | 2 |
| Hydrogen Bond Acceptor Count | 8 |
| Rotatable Bond Count | 7 |
| Heavy Atom Count | 16 |
| Complexity | 345 |
| Defined Atom Stereocenter Count | 2 |
| SMILES | O[C@H]([C@@H](O)COS(C)(=O)=O)COS(C)(=O)=O |
| InChi Key | YCPOZVAOBBQLRI-WDSKDSINSA-N |
| InChi Code | InChI=1S/C6H14O8S2/c1-15(9,10)13-3-5(7)6(8)4-14-16(2,11)12/h5-8H,3-4H2,1-2H3/t5-,6-/m0/s1 |
| Chemical Name | [(2S,3S)-2,3-dihydroxy-4-methylsulfonyloxybutyl] methanesulfonate |
| Synonyms | NSC-39069; Treosulfan; NSC 39069; Treosulphan; Ovastat; Dihydroxybusulfan; threosulphan; Treosulfano; Treosulfanum; NSC39069; (2S,3S)-2,3-Dihydroxybutane-1,4diyl dimethanesulfonate |
| HS Tariff Code | 2934.99.9001 |
| 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) |
Biological Activity
| Targets | DNA Alkylator |
| ln Vitro | Treosulfan is an agent that alkylates. Treosulfan exhibits nearly 100% cytotoxicity at 100 μg/mL on a number of cancer cell lines, including Panc-1, Miapaca-2, and Capan-2 cells, with IC50s of 3.6 μg/mL, 1.8 μg/mL, and 2.1 μg/mL, respectively. When combined with LY 188011, treosulfan (0.1-100 μg/mL) shows increased activity against cancer cells. On the other hand, Treosulfan (1, 2.5, and 5 μg/ml) in combination with 5-FU (0.1, 0.25, and 0.5 μg/ml) exhibits antagonistic effects on Miapaca-2 cells at all doses and on Panc-1 cells at intermediate and high concentrations[1]. Treosulfan (800 µg/mL) significantly lowers erythrocyte forward scatter and raises the proportion of ROS, [Ca2+]i, and annexin-V-binding cells. When extracellular Ca2+ is removed, Treosulfan's effect on annexin-V binding is negated[2]. |
| ln Vivo | Treosulfan (1.5 g/kg/day) causes mice to rapidly undergo myeloablation and lose all of their splenic B and T cells. Treosulfan (1.5 g/kg/day) briefly increases the production of olny interleukin-2 in the spleen cells without clearly having a major impact on the synthesis of tumor necrosis factor-α and/or IFN-γ in mice[3]. |
| Cell Assay | In tissue culture plates with 96 wells, the cells are grown in 100 μL volume per well and plated at 1×104 cells/mL for cytotoxicity assays. After allowing the cells to adhere for a full night, they are cultured with varying concentrations of either Treosulfan alone or in conjunction with LY 188011. The drug combination is introduced either sequentially—the second drug is added 12 hours after the first—or simultaneously to the cell cultures. Alamar Blue® solution is added to the wells following a 72-hour incubation period before an additional overnight incubation. Next, absorbance is determined using a spectrophotometer, and drug cytotoxicity and cell proliferation are computed. Additionally, in certain experiments, trypan blue exclusion is used to determine proliferation and cytotoxicity, and an improved Neubauer hemocytometer is used to count cells. Cell viability is evaluated by staining the cells with 7-amino-actinomycin D (final concentration 200 μg/mL) and Annexin-V, followed by flow cytometry analysis using an FACS Scan flow cytometer[1]. |
| Animal Protocol | Mice: At 10 to 12 weeks of age, female BALB/c mice weighed about 20 g. Standard pelleted feed and unlimited water are provided to the animals. They are kept in a climate-controlled room with a 12-hour light/dark cycle. There are four groups that they are split up into: one group gets treated with liposomal NCI C01592 (37 mg/kg/day) for four days straight; another group receives NSC-26271 (0.1 g/kg/day) for two days straight; a control group does not receive any treatment. To sustain the animals' survival in the absence of bone marrow support, sublethal doses of NSC-26271, NCI C01592, and treosulfan are administered. Days 1, 3, 6, 9, and 12 following the final treatment dose are dedicated to animal sacrifice, during which the femurs and spleen are extracted. Two control and six treated animals are included at each time point [3]. |
| ADME/Pharmacokinetics |
Absorption, Distribution and Excretion In a pharmacologic study of the bioavailability of treosulfan in a capsule formulation, patients with relapsed ovarian carcinoma were treated with alternating doses of oral and intravenous (i.v.) treosulfan of 1.5 or 2.0 g daily for 5 to 8 days. ... The bioavailability ratio (f) of oral to i.v. administration was calculated as 0.97 + or - 0.18 (mean + or - SD) using the values AUC oral = 82.1 + or- 39.4 ug/ml hr and AUC i.v. = 85.4 + or - 30.3 ug/ml hr. The peak plasma concentration cmax (29 + or - 14 ug/ml vs 65 + or - 23 ug/ml) was significantly (P < 0.01) higher after i.v. administration and the tmax after oral administration was 1.5 + or - 0.34 hr. The terminal half-life of treosulfan was about 1.8 hr. The mean urinary excretion of the parent compound was about 15% of the administered total dose over 24 hr (range 6-26%). ... A feasible and reliable oral treosulfan formulation could provide the basis for the development of long-term low-dose outpatient treatment of patients with malignant diseases. |
| Toxicity/Toxicokinetics |
dog LDLo intravenous 222 mg/kg GASTROINTESTINAL: OTHER CHANGES; BLOOD: LEUKOPENIA; BLOOD: OTHER CHANGES Cancer Chemotherapy Reports, Part 2., 2(203), 1965 monkey LDLo intravenous 222 mg/kg BLOOD: LEUKOPENIA; BLOOD: AGRANULOCYTOSIS; BLOOD: OTHER CHANGES Cancer Chemotherapy Reports, Part 2., 2(203), 1965 Interactions L-buthionine-[S,R]-sulfoximine had minor effects on the toxicity of doxorubicin, ACNU (1-[(4-amino-2-methyl-5-pyrimidinyl)methyl]-3-(2-chloroethyl)-3-nitrosou rea, nimustine) and vincristine. L-buthionine-[S,R]-sulfoximine failed to alter teniposide or cytarabine toxicity. L-buthionine-[S,R]-sulfoximine induced prominent sensitization to the alkylating agent, treosulfan, in both cell lines, as assessed by viability assays, in situ DNA end labeling and quantitative DNA fragmentation. Treosulfan is thought to mediate toxicity via formation of reactive epoxides. PMID:9484802 Antidote and Emergency Treatment Basic treatment: Establish a patent airway. Suction if necessary. Watch for signs of respiratory insufficiency and assist ventilations if needed. Administer oxygen by nonrebreather mask at 10 to 15 L/min. Monitor for pulmonary edema and treat if necessary ... . Monitor for shock and treat if necessary ... . Anticipate seizures and treat if necessary ... . For eye contamination, flush eyes immediately with water. Irrigate each eye continuously with normal saline during transport ... . Do not use emetics. For ingestion, rinse mouth and administer 5 ml/kg up to 200 ml of water for dilution if the patient can swallow, has a strong gag reflex, and does not drool ... . Cover skin burns with dry sterile dressings after decontamination ... . /Poison A and B/ Advanced treatment: Consider orotracheal or nasotracheal intubation for airway control in the patient who is unconscious, has severe pulmonary edema, or is in respiratory arrest. Positive pressure ventilation techniques with a bag valve mask device may be beneficial. Monitor cardiac rhythm and treat arrhythmias as necessary ... . Start an IV with D5W /SRP: "To keep open", minimal flow rate/. Use lactated Ringer's if signs of hypovolemia are present. Watch for signs of fluid overload. Consider drug therapy for pulmonary edema ... . For hypotension with signs of hypovolemia, administer fluid cautiously. Watch for signs of fluid overload ... . Treat seizures with diazepam (Valium) ... . Use proparacaine hydrochloride to assist eye irrigation ... . /Poison A and B/ View More
Human Toxicity Excerpts
Non-Human Toxicity Excerpts The cytotoxicity and mutagenicity of the human carcinogen, treosulphan, and its hydrolysis product, dl-1,2:3,4-diepoxybutane (DEB), were studied in Chinese hamster ovary, AS52, cells. Treosulphan (0.1-1.0 mM) is toxic and mutagenic at the gpt locus. A strong pH dependence was noted. dl-1,2:3,4-diepoxybutane is cytotoxic and mutagenic at a much lower dose (0.025 mM), but these effects were not affected by pH. The results suggest that the toxic and mutagenic effects of treosulphan are mediated by its hydrolysis product DEB, and that the conversion of treosulphan to DEB is highly pH-dependent. PMID:8419160 Two human carcinogens, 4-aminobiphenyl (4AB) and treosulphan (Treo), were tested in male B6C3F1 mice for the induction of micronuclei in bone marrow and peripheral blood cells by 1-, 2- and 3-exposure protocols. Both compounds tested positive. The magnitude of response with respect to the incidence of micronucleated polychromatic erythrocytes by 2- and 3-exposure protocols was considerably higher than by the single-exposure protocol. The peripheral blood results for Treo were as typically seen with a 24-h delay when compared to the bone marrow. The peripheral blood results for 4AB, however, differed from those expected. The incidence of MN-PCE in peripheral blood of animals exposed to 4AB was significantly greater than seen in the bone marrow in 2- and 3-exposure protocols. There was also an increase in the % PCE at the 60 mg/kg dose level as a function of time. Based on these studies, it is concluded that a step-wise scoring scheme may be the best protocol for rodent micronucleus assay, involving a 3-exposure protocol with single sampling of bone marrow (24 h after the last treatment) and two samplings of peripheral blood (24 h and 48 h after the first treatment). This approach is cost-effective, it limits the number of animals required and provides maximum sensitivity. |
| References |
[1]. Synergistic cytotoxic activity of treosulfan and LY 188011 in pancreatic cancer cell lines. Anticancer Res. 2014 Apr;34(4):1779-84. [2]. Programmed erythrocyte death following in vitro Treosulfan treatment. Cell Physiol Biochem. 2015;35(4):1372-80. [3]. Myeloablative and immunosuppressive properties of treosulfan in mice. Exp Hematol. 2006 Jan;34(1):115-21. |
| Additional Infomation |
Treosulfan can cause cancer according to California Labor Code. Treosulphan is an odorless white crystalline powder. (NTP, 1992) Treosulfan is a methanesulfonate ester. Treosulfan is under investigation in Allogeneic Haematopoietic Stem Cell Transplantation. Treosulfan has been investigated for the treatment of Lymphoblastic Leukemia, Acute, Childhood. Treosulfan is the prodrug of a bifunctional sulfonate alkylating agent with myeloablative, immunosuppressive, and antineoplastic activities. Under physiological conditions, treosulfan converts nonenzymatically to L-diepoxybutane via a monoepoxide intermediate. The monoepoxide intermediate and L-diepoxybutane alkylate DNA at guanine residues and produce DNA interstrand crosslinks, resulting in DNA fragmentation and apoptosis. In escalated doses, this agent also exhibits myeloablative and immunosuppressive activities. Drug Indication Treosulfan in combination with fludarabine is indicated as part of conditioning treatment prior to allogeneic haematopoietic stem cell transplantation (alloHSCT) in adult patients and in paediatric patients older than one month with malignant and non-malignant diseases. Conditioning treatment prior to haematopoietic-progenitor-cell transplantation Mechanism of Action The anti-tumour drug treosulfan (L-threitol 1,4-bismethanesulphonate, Ovastat) is a prodrug for epoxy compounds by converting non-enzymatically to L-diepoxybutane via the corresponding monoepoxide under physiological conditions. The present study supports the hypothesis that this conversion of treosulfan is required for cytotoxicity in vitro. DNA alkylation and interstrand cross-linking of plasmid DNA is observed after treosulfan treatment, but this is again produced via the epoxide species. Alkylation occurs at guanine bases with a sequence selectivity similar to other alkylating agents such as the nitrogen mustards. In treosulfan-treated K562 cells, cross-links form slowly, reaching a peak at approximately 24 h. Incubation of K562 cells with preformed epoxides shows faster and more efficient DNA cross-linking. |
Solubility Data
| Solubility (In Vitro) |
DMSO: ~56 mg/mL (~201.2 mM) Water: ~56 mg/mL |
| Solubility (In Vivo) |
Solubility in Formulation 1: ≥ 2.5 mg/mL (8.98 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 25.0 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. Solubility in Formulation 2: ≥ 2.5 mg/mL (8.98 mM) (saturation unknown) in 10% DMSO + 90% (20% SBE-β-CD in 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 25.0 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. Solubility in Formulation 3: ≥ 2.5 mg/mL (8.98 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 25.0 mg/mL clear DMSO stock solution to 900 μL of corn oil and mix evenly. Solubility in Formulation 4: 16.67 mg/mL (59.90 mM) in PBS (add these co-solvents sequentially from left to right, and one by one), clear solution; with ultrasonication. (Please use freshly prepared in vivo formulations for optimal results.) |
| Preparing Stock Solutions | 1 mg | 5 mg | 10 mg | |
| 1 mM | 3.5932 mL | 17.9662 mL | 35.9324 mL | |
| 5 mM | 0.7186 mL | 3.5932 mL | 7.1865 mL | |
| 10 mM | 0.3593 mL | 1.7966 mL | 3.5932 mL |