Lomeguatrib
(Synonyms: 罗米鲁曲,PaTrin-2) 目录号 : GC11778
Inactivator of O6-methylguanine-DNA methyltransferase
Cas No.:192441-08-0
Sample solution is provided at 25 µL, 10mM.
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Quality Control & SDS
- View current batch:
- Purity: >99.50%
- COA (Certificate Of Analysis)
- SDS (Safety Data Sheet)
- Datasheet
Kinase experiment: | Briefly, 200 μg of extracted cellular protein from HeLaS3 cells in 200 μL of 70 mM HEPES buffer (with 1 mM dithiothreitol (DTT), 5 mM EDTA, pH 7.8) is incubated at 37°C with a defined concentration of Lomeguatrib (added as a DMSO solution). After 30 min an excess of [3H]-methylated DNA (120 000 cpm) is added, and the incubation is continued for an additional 90 min. The reaction is stopped by the addition of 400 μL TCA (13%), and the DNA is hydrolyzed by heating the reaction mixture for 30 min at 98°C. The precipitated protein is washed three times with 400-μL portions of 5% TCA, solubilized in 0.1 N NaOH, and analyzed by liquid scintillation counting using the cocktail Rotiszint eco plus and a TRI-CARB. Enzyme activity is expressed as fmol of [3H]methyl transferred to TCA-insoluble protein material per mg of total cellular protein. Percent inhibition is calculated relative to untreated control samples. Each assay is repeated three times, and IC50 values are determined graphically from plots of percent inhibition vs inhibitor concentration[1]. |
Cell experiment: | To determine toxicity, the MTT growth inhibition assay is employed. Cells (1000 per well) are plated into a 96-well plate and following a 24 h attachment period, Lomeguatrib is added to the cells. After 2 h incubation with Lomeguatrib (10 μM) at 37°C, 5% CO2, increasing doses of temozolomide or vehicle are added and the cells are incubated for a further 4-5 days. At the end of the exposure period, 150 μg MTT is added to each well and plates are incubated for 3 h at 37°C, 5% CO2. The media are removed and the formazan crystals formed in the viable cells are solubilised in 200 μL DMSO. The absorbances at 540 and 690 nm are determined using a ELISA plate reader and growth inhibition calculated as a percentage of the A540-A690 of untreated wells[2]. |
Animal experiment: | Mice[2]To assess the ability of Lomeguatrib to sensitise human breast tumour xenografts to the tumour growth inhibitory effects of temozolomide, groups of at least six nude mice are treated as follows: the vehicle control group are given corn oil then 20% DMSO in PBS; the temozolomide only group are given corn oil then temozolomide (100 mg/kg/day); the Lomeguatrib only group are given Lomeguatrib (20 mg/kg/day) then DMSO in PBS, and the Lomeguatrib plus temozolomide group are given Lomeguatrib (20 mg/kg/day) then temozolomide (100 mg/kg/day). Drugs or vehicles are administered i.p. once daily for 5 days with a separation of 1 h. Up to 10 and at least six animals are assigned to each group, and mean tumour volume is standardised across the groups at the start of the experiment: thus the control, Lomeguatrib, temozolomide and Lomeguatrib plus temozolomide groups had mean tumour volumes of 29.8±7.6 (range 19.0-38.7), 33.2±14.7 (range 16.5-58.7), 35.1±10.9 (range 20.9-52.4) and 30.3±10.0 (range 20.7-44.5) mm3, respectively[2]. |
References: [1]. Reinhard J, et al. Monosaccharide-linked inhibitors of O(6)-methylguanine-DNA methyltransferase (MGMT): synthesis, molecular modeling, and structure-activity relationships. J Med Chem. 2001 Nov 22;44(24):4050-61. | |
Lomeguatrib is an O6-methylguanine-DNA-methyl-transferase (MGMT) inhibitor [1], with an IC50 value of about 6 nM to inactivate MGMT in MCF-7 cells, effectively [2].
MGMT activity is closely related to MTIC (a metabolite of dacarbazine)-mediated DNA damage [1].
Lomeguatrib sensitized MGMT-activity-bearing A375P cells to temozolomide (TMZ), but it failed to affect the effect of dacarbazine (DTIC). In other mutBRAF cells and several mutNRAS cell lines such as WM1361, similar results were obtained [1].
With one exception, patients treated with lomeguatrib showed no active or very low MGMT in PBMCs. Lomeguatrib at a dose of 20 mg resulted in 16.7 fmol/μg DNA active MGMT in a CNS-tumor-bearing patient. This patient showed a percentage of 25% for inactive tumor MGMT. This percentage was lower than that in the other two CNS patients with lomeguatrib at the same dose. Different tumor types showed remarkable differences in total tumor MGMT. Prostate cancers had the highest (554 ± 404 fmol/mg protein), CNS tumors had the lowest (89.9 ± 44.5 fmol/mg protein), and colorectal tumors had intermediate levels of total protein (244 ± 181 fmol/mg protein). In the colorectal cancer, the primary CNS tumor, and the prostate cancer of patients, increasing lomeguatrib doses resulted in increasing inactive MGMT proportions [3].
References:
[1]. Imanol Arozarena, Ibai Goicoechea, Oihane Erice, et al. Differential chemosensitivity to antifolate drugs between RAS and BRAF melanoma cells. Molecular Cancer, 2014, 13:154.
[2]. M Clemons, J Kelly, AJ Watson, et al. O6-(4-bromothenyl)guanine reverses temozolomide resistance in human breast tumour MCF-7 cells and xenografts. British Journal of Cancer, 2005, 93:1152-1156.
[3]. Amanda J. Watson, Ami Sabharwal, Mary Thorncroft, et al. Tumor O6-methylguanine-DNA Methyltransferase Inactivation by Oral Lomeguatrib. Clinical Cancer Research, 2010, 16(2):743-9.
| Cas No. | 192441-08-0 | SDF | |
| 别名 | 罗米鲁曲,PaTrin-2 | ||
| 化学名 | 6-[(4-bromothiophen-2-yl)methoxy]-7H-purin-2-amine | ||
| Canonical SMILES | C1=C(SC=C1Br)COC2=NC(=NC3=C2NC=N3)N | ||
| 分子式 | C10H8BrN5OS | 分子量 | 326.17 |
| 溶解度 | ≥ 50mg/mL in DMSO with gentle warming | 储存条件 | Store at -20°C |
| General tips | 请根据产品在不同溶剂中的溶解度选择合适的溶剂配制储备液；一旦配成溶液，请分装保存，避免反复冻融造成的产品失效。 储备液的保存方式和期限：-80°C 储存时，请在 6 个月内使用，-20°C 储存时，请在 1 个月内使用。 为了提高溶解度，请将管子加热至37℃，然后在超声波浴中震荡一段时间。 |
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| Shipping Condition | 评估样品解决方案：配备蓝冰进行发货。所有其他可用尺寸：配备RT，或根据请求配备蓝冰。 | ||
| 制备储备液 | |||
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1 mg | 5 mg | 10 mg |
| 1 mM | 3.0659 mL | 15.3294 mL | 30.6589 mL |
| 5 mM | 0.6132 mL | 3.0659 mL | 6.1318 mL |
| 10 mM | 0.3066 mL | 1.5329 mL | 3.0659 mL |
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| 给药剂量 | mg/kg |  |
动物平均体重 | g | |
每只动物给药体积 | ul | |
动物数量 | 只 |
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| % DMSO % % Tween 80 % saline | ||||||||||
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工作液浓度： mg/ml；
DMSO母液配制方法： mg 药物溶于 μL DMSO溶液（母液浓度 mg/mL，
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Lomeguatrib Increases the Radiosensitivity of MGMT Unmethylated Human Glioblastoma Multiforme Cell Lines
Background: Treatment resistance of glioblastoma multiforme to chemo- and radiotherapy remains a challenge yet to overcome. In particular, the O6-methylguanine-DNA-methyltransferase (MGMT) promoter unmethylated patients have only little benefit from chemotherapy treatment using temozolomide since MGMT counteracts its therapeutic efficacy. Therefore, new treatment options in radiotherapy need to be developed to inhibit MGMT and increase radiotherapy response. Methods: Lomeguatrib, a highly specific MGMT inhibitor, was used to inactivate MGMT protein in vitro. Radiosensitivity of established human glioblastoma multiforme cell lines in combination with lomeguatrib was investigated using the clonogenic survival assay. Inhibition of MGMT was analyzed using Western Blot. Cell cycle distribution and apoptosis were investigated to determine the effects of lomeguatrib alone as well as in combination with ionizing radiation. Results: Lomeguatrib significantly decreased MGMT protein and reduced radiation-induced G2/M arrest. A radiosensitizing effect of lomeguatrib was observed when administered at 1 ?M and increased radioresistance at 20 ?M. Conclusion: Low concentrations of lomeguatrib elicit radiosensitization, while high concentrations mediate a radioprotective effect.
Temozolomide: mechanisms of action, repair and resistance
Glioblastoma multiforme is the most common aggressive adult primary tumour of the central nervous system. Treatment includes surgery, radiotherapy and adjuvant temozolomide (TMZ) chemotherapy. TMZ is an alkylating agent prodrug, delivering a methyl group to purine bases of DNA (O6-guanine; N7-guanine and N3-adenine). The primary cytotoxic lesion, O6-methylguanine (O6-MeG) can be removed by methylguanine methyltransferase (MGMT; direct repair) in tumours expressing this protein, or tolerated in mismatch repair-deficient (MMR-) tumours. Thus MGMT or MMR deficiency confers resistance to TMZ. Inherent- and acquired resistance to TMZ present major obstacles to successful treatment. Strategies devised to thwart resistance and enhance response to TMZ, including inhibition of DNA repair mechanisms which contribute to TMZ resistance, are under clinical evaluation. Depletion of MGMT prior to alkylating agent chemotherapy prevents O6-MeG repair; thus, MGMT pseudosubstrates O6-benzylguanine and lomeguatrib are able to sensitise tumours to TMZ. Disruption of base excision repair (BER) results in persistence of potentially lethal N7- and N3- purine lesions contributing significantly to TMZ cytoxicity particularly when O6-MeG adducts are repaired or tolerated. Several small molecule inhibitors of poly(ADP-ribose)polymerase-1 (PARP-1), a critical BER protein are yielding promising results clinically, both in combination with TMZ and as single agent chemotherapy in patients whose tumours possess homologous recombination DNA repair defects. Another validated, but as yet preclinical protein target, mandatory to BER is abasic (AP) endonuclease-1 (APE-1); in preclinical tests, APE-1 inhibition potentiates TMZ activity. An alternative strategy is synthesis of a molecule, evoking an irrepairable cytotoxic O6-G lesion. Preliminary efforts to achieve this goal are described.
Preparation of PEGylated liposomes incorporating lipophilic lomeguatrib derivatives for the sensitization of chemo-resistant gliomas
Liposomal delivery is a well-established approach to increase the therapeutic index of drugs, mainly in the field of cancer chemotherapy. Here, we report the preparation and characterization of a new liposomal formulation of a derivative of lomeguatrib, a potent O6-methylguanine-DNA methyltransferase (MGMT) inactivator. The drug had been tested in clinical trials to revert chemoresistance, but was associated with a low therapeutic index. A series of lomeguatrib conjugates with distinct alkyl chain lengths - i.e. C12, C14, C16, and C18 - was synthesized, and the MGMT depleting activity as well as cytotoxicity were determined on relevant mouse and human glioma cell lines. Drug-containing liposomes were prepared and characterized in terms of loading and in vitro release kinetics. The lipophilic lomeguatrib conjugates did not exert cytotoxic effects at 5 μM in the mouse glioma cell line and exhibited a similar MGMT depleting activity pattern as lomeguatrib. Overall, drug loading could be improved by up to 50-fold with the lipophilic conjugates, and the slowest leakage was achieved with the C18 derivative. The present data show the applicability of lipophilic lomeguatrib derivatization for incorporation into liposomes, and identify the C18 derivative as the lead compound for in vivo studies.
Lomeguatrib, a potent inhibitor of O6-alkylguanine-DNA-alkyltransferase: phase I safety, pharmacodynamic, and pharmacokinetic trial and evaluation in combination with temozolomide in patients with advanced solid tumors
Purpose: A major mechanism of resistance to temozolomide involves the DNA repair protein O6-alkylguanine-DNA-alkyltransferase (ATase). The main aims of this phase I trial were to determine an ATase-depleting dose (ADD) of lomeguatrib, a potent pseudosubstrate inhibitor, and to define a suitable dose of temozolomide to be used in combination with lomeguatrib in patients with advanced cancer.
Experimental design: Lomeguatrib was administered at dose levels of 10 to 40 mg/m2 days 1 to 5, as a single agent, and also in combination with temozolomide. Once the ADD of lomeguatrib was identified, the dose of temozolomide in combination was increased, in successive patient cohorts, from 50 to 175 mg/m2 on days 1 to 5 of a 28-day cycle to define the maximal tolerated dose and dose-limiting toxicity of the combination.
Results: Thirty-eight patients with advanced solid tumors were enrolled. More than 95% ATase depletion within 4 hours of the first dose was achieved in peripheral blood mononuclear cells at lomeguatrib doses of > or =10 mg/m2/d i.v. or > or =20 mg/m2/d orally, and tumor biopsies showed > or =92% ATase depletion. At the ADD of lomeguatrib i.v., the maximal tolerated dose of temozolomide in combination was 150 mg/m2 days 1 to 5. The dose limiting toxicity of the combination of lomeguatrib and temozolomide was myelosuppression. The toxicity of lomeguatrib alone was minimal. In 23 patients with measurable disease, one complete response was seen and 12 patients had stable disease for at least 3 months.
Conclusion: This first administration of lomeguatrib to man successfully established an oral ADD of lomeguatrib and identified a combination regimen with temozolomide suitable for future clinical evaluation.
Effect of lomeguatrib-temozolomide combination on MGMT promoter methylation and expression in primary glioblastoma tumor cells
Temozolomide (TMZ) is commonly used in the treatment of glioblastoma (GBM). The MGMT repair enzyme (O (6)-methylguanine-DNA methyltransferase) is an important factor causing chemotherapeutic resistance. MGMT prevents the formation of toxic effects of alkyl adducts by removing them from the DNA. Therefore, MGMT inhibition is an interesting therapeutic approach to circumvent TMZ resistance. The aim of the study was to investigate the effect of the combination of lomeguatrib (an MGMT inactivator) with TMZ, on MGMT expression and methylation. Primary cell cultures were obtained from GBM tumor tissues. The sensitivity of primary GBM cell cultures and GBM cell lines to TMZ, and to the combination of TMZ and lomeguatrib, was determined by a cytotoxicity assay (MTT). MGMT and p53 expression, and MGMT methylation were investigated after drug application. In addition, the proportion of apoptotic cells and DNA fragmentation was analyzed. The combination of TMZ and lomeguatrib in primary GBM cell cultures and glioma cell lines decreased MGMT expression, increased p53 expression, and did not change MGMT methylation. Moreover, apoptosis was induced and DNA fragmentation was increased in cells. In addition, we also showed that lomeguatrib-TMZ combination did not have any effect on the cell cycle. Finally, we determined that the sensitivity of each primary GBM cells and glioma cell lines to the lomeguatrib-TMZ combination was different and significantly associated with the structure of MGMT methylation. Our study suggests that lomeguatrib can be used with TMZ for GBM treatment, although further clinical studies will be needed so as to determine the feasibility of this therapeutic approach.