| Size | Price | Stock | Qty |
|---|---|---|---|
| 1mg |
|
||
| 5mg |
|
||
| 10mg | |||
| Other Sizes |
Purity: ≥98%
SJG-136 (NSC694501), a novel sequence-selective pyrrolobenzodiazepine (PBD) dimer, is a novel and potent DNA cross-linking/intercalating agent with an XL50 of 45 nM for pBR322 DNA; SJG-136 has potent anticancer activity and can be potentially used for the treatment of ovarian cancer and leukemia.
| Targets |
SJG-136 (SG2000) targets DNA minor groove, forming covalent adducts with guanine N2 positions.
- DNA interstrand cross-linking: XL50 = 0.045 μM (pBR322 DNA, 2 h incubation) [1]
- Thermal denaturation (calf thymus DNA, 5:1 DNA/ligand ratio, 18 h): ΔTm = 33.6 °C [1] |
|---|---|
| ln Vitro |
- DNA cross-linking efficiency: In linearized pBR322 DNA, SJG-136 produced measurable cross-linking above background at 0.001 μM, with 100% cross-linking at doses >0.3 μM. XL50 = 0.045 μM, which is 440-fold more potent than melphalan (XL50 = 20 μM). [1]
- Thermal denaturation (DNA stabilization): Using calf thymus DNA at a 5:1 DNA/ligand ratio and 18 h incubation at 37 °C, SJG-136 increased Tm by 33.6 °C. At 72 h, ΔTm reached 34.4 °C. At a 100:1 ratio, ΔTm was 9.1 °C after 18 h. [1] - Cell growth inhibition (canine cancer cell lines): SJG-136 was tested against 12 canine tumour cell lines (1 h exposure + 96 h drug-free, or continuous 96 h exposure). GI50 values after 1 h exposure ranged from 0.33 nM (C2 mast cell) to >100 nM (several lines); after continuous exposure, GI50 ranged from <0.03 nM (C2, KMeC) to 28.33 nM (MDCK normal kidney). Normal canine cells (CF35MG mammary normal, MDCK) had GI50 >100 nM (1 h) and 6.0–28.33 nM (continuous). [1] - Relationship between ICL and cytotoxicity: In six canine cancer lines and one human melanoma line, correlation R² = 0.7362 (or 0.9131 excluding CMeC-2). [1] - Persistence of DNA ICLs: In CMeC-1 cells treated with 53 nM SJG-136 for 1 h, ICLs persisted without removal for up to 48 h post-incubation. [1] - Cell cycle effects: In four canine melanoma lines (CMeC-1, CMeC-2, KMeC, LMeC) treated at GI50 doses for 1 h, accumulation in G2/M was observed in LMeC, KMeC, and CMeC-1, while CMeC-2 showed S-phase accumulation. [1] - γ-H2AX foci formation (in vitro): In CMeC-1 cells treated with 10 nM SJG-136 for 1 h, γ-H2AX foci were slow to form, becoming evident at 4 h, peaking at 24 h (mean foci/cell increased from ~2 to ~25), and gradually declining by 48 h. Dose-dependent increase in foci was observed from 0.1 to 100 nM in CMeC-1 and LMeC cells at 24 h post-treatment. [1] DNA cross-linker SJG-136 (dimer 5) has an XL50 (reagent concentration needed for 50% cross-linking of pBR322 DNA) of 45 nM. A2780 (IC50, 22.5 pM), A2780cisR (IC50, 24 pM), CH1 (IC50, 0.12 nM), CH1cisR (IC50, 0.6 nM), and SKOV-3 (IC50, 9.1 nM) are among the ovarian cell lines that are cytotoxic to SJG-136 [1]. A set of canine cancer cells is likewise less viable when exposed to SJG-136 (SG2000); GI50 values range from 0.33 to >100 nM after an hour of treatment and from <0.03 to 17.33 nM when exposed continuously [2]. |
| ln Vivo |
- Antitumor activity in canine melanoma xenografts (LMeC model): SJG-136 given i.v. at 0.15 or 0.30 mg/kg (single dose or weekly × 3). Dose-dependent growth inhibition observed. Single dose 0.30 mg/kg caused significant delay (p<0.05); weekly ×3 at 0.30 mg/kg gave >40 days growth delay. Median weight loss: 0% (0.15 mg/kg single), 2.45% (0.30 mg/kg single), 1.9% (0.15 mg/kg weekly ×3), 15.81% (0.30 mg/kg weekly ×3). [1]
- Antitumor activity in canine melanoma xenografts (CMeC-1 model): 0.30 mg/kg (single or weekly ×3) gave significantly greater effect than 0.15 mg/kg (p<0.01). Weekly ×3 schedule produced growth delays >100 days at both doses. Mean survival increased by 90 days over controls at 0.30 mg/kg weekly ×3. Weight loss: 4.4% (0.15 mg/kg weekly ×3), 12.6% (0.30 mg/kg weekly ×3). [1] - In vivo DNA ICL formation (comet assay): In LMeC tumours, at 2 h post-dose, % decrease in tail moment was dose-dependent (higher at 0.30 mg/kg than 0.15 mg/kg). ICLs persisted at 24 h. In CMeC-1 tumours, similar dose-dependent ICL at 2 h and persistence at 24 h. ICL in peripheral blood mononuclear cells was lower than in tumours. [1] - Repeated dosing ICL accumulation: In peripheral blood mononuclear cells, ICL levels increased after the second and third weekly doses of SJG-136 (0.15 or 0.30 mg/kg), indicating persistence of unrepaired cross-links. [1] - In vivo γ-H2AX foci response: In LMeC and CMeC-1 tumours, γ-H2AX foci were higher at 24 h than at 2 h post-dose, and higher at 0.30 mg/kg than 0.15 mg/kg. Foci levels were greater in tumours than in peripheral blood mononuclear cells. [1] - Immunohistochemistry for γ-H2AX in tumours: In formalin-fixed paraffin-embedded sections, SJG-136 induced a dose- and time-dependent increase in γ-H2AX-positive cells (higher at 24 h than 2 h; higher at 0.30 mg/kg than 0.15 mg/kg). More positive cells in CMeC-1 than in LMeC. [1] SJG-136-induced H2AX phosphorylation shows good correspondence but is less sensitive than focal measurements. SJG-136 at 0.30 mg/kg had a more potent antitumor effect on CMeC-1 tumors than SJG-136 at 0.15 mg/kg, either as a single dose or once weekly, administered by intravenous injection, for three weeks [2]. |
| Enzyme Assay |
No classical enzyme assays were reported for SJG-136 (SG2000). Instead, DNA binding and cross-linking assays are described:
- Thermal denaturation (DNA melting) assay: Calf thymus DNA (100 μM) was incubated with SJG-136 at various [DNA]/[ligand] ratios (5:1, 50:1, 100:1) in 10 mM sodium phosphate buffer (pH 7.00 ± 0.01) containing 1 mM EDTA, at 37.0 ± 0.1 °C for 0–18 h. Samples were heated from 45 to 98 °C at 1 °C/min. Absorbance at 260 nm was recorded. Tm was determined as the midpoint of the normalized melting profile. ΔTm = Tm(DNA+ligand) – Tm(DNA alone). Free DNA Tm = 67.83 ± 0.06 °C. [1]
- DNA cross-linking assay (gel electrophoresis): Linearized pBR322 DNA (5′-end labeled with ³²P) was incubated with SJG-136 (0.001–10 μM) in 25 mM triethanolamine/1 mM EDTA (pH 7.2) at 37 °C for 2 h. Reactions were stopped with 0.6 M NaOAc/20 mM EDTA/100 μg/mL tRNA, and DNA precipitated with ethanol. Samples were denatured (90 °C, 2 min) in 30% DMSO/1 mM EDTA, chilled, and run on a 0.8% neutral agarose gel at 40 V for 16 h. Gels were dried and autoradiographed. XL50 was the concentration producing 50% double-stranded (cross-linked) DNA. [1] |
| Cell Assay |
- Cell growth inhibition (SRB assay for adherent cells): Cells were seeded in 96-well plates and allowed to attach overnight. SJG-136 was added at concentrations from 0.03 nM to 1000 nM for either 1 h (followed by 96 h in drug-free medium) or continuous 96 h exposure. Cells were fixed with 10% trichloroacetic acid, stained with 0.4% sulforhodamine B in 1% acetic acid, and absorbance read at 540 nm. GI50 was calculated. [1]
- Cell growth inhibition (MTT assay for suspension cells): Used for C2 cell line. Following drug exposure, MTT was added, and formazan absorbance measured. [1] - Comet assay for DNA ICL: Cells (2.5×10⁴/mL) were irradiated (15 Gy) to introduce random strand breaks, embedded in 1% agarose on slides, lysed (100 mM EDTA, 2.5 M NaCl, 10 mM Tris-HCl pH 10.5, 1% Triton X-100) for 1 h, washed, and incubated in alkali buffer (50 mM NaOH, 1 mM EDTA, pH 12.5) for 45 min. Electrophoresis was at 18 V (0.6 V/cm, 250 mA) for 25 min. Slides were neutralized, stained with propidium iodide (2.5 μg/mL), and analyzed by fluorescence microscopy. Tail moment was calculated, and % decrease in tail moment = [1 – ((TMDi – TMCu)/(TMCi – TMCu))] × 100. [1] - Cell cycle analysis by flow cytometry: Cells (2×10⁶/mL) were treated with SJG-136 at GI50 doses for 1 h, then incubated in drug-free medium for up to 72 h. Harvested cells were fixed in 70% ethanol, stained with propidium iodide (0.05 mg/mL) and RNase A (0.5 mg/mL), and analyzed by flow cytometry. Percentages of cells in G1/G0, S, and G2/M were determined. [1] - γ-H2AX immunofluorescence (in vitro): Cells grown on chamber slides were treated with SJG-136 (0.1–100 nM) for 1 h, then fixed at various times (up to 48 h) with ice-cold methanol/acetone (1:1). Cells were permeabilized (0.5% Triton X-100), blocked (0.1% Triton X-100, 0.2% skim milk), and incubated with anti-phospho-histone H2AX (Ser139) primary antibody (1:1000) overnight at 4 °C, then with Alexa Fluor 488 secondary antibody (1:1000) for 4 h. Nuclei were counterstained with propidium iodide. Foci were counted in 50 cells per time point. [1] |
| Animal Protocol |
- Xenograft model (LMeC and CMeC-1 canine melanoma): Female CD1 Nu/Nu immunocompromised mice were injected subcutaneously with 5×10⁶ tumour cells in serum-free RPMI. When tumours were established, mice were divided into groups (n=10 per group, 6 for efficacy, 4 for pharmacodynamics). SJG-136 was dissolved in 0.9% NaCl containing 1% DMSO and administered intravenously via tail vein at 0.15 or 0.30 mg/kg (volume 0.1 mL/10 g body weight). Dosing schedules: single dose or once weekly for three weeks. Control mice received vehicle only. Tumour volumes (ellipsoid formula) and body weights were measured every 3–4 days. [1]
- Tissue sampling for pharmacodynamics: Tumours were dissected, minced to single-cell suspension, and stored in freezing medium (FCS + 10% DMSO) at –80 °C. Peripheral blood mononuclear cells were isolated from blood by Ficoll density gradient centrifugation (450×g, 20 min) and stored similarly. [1] - Immunohistochemistry for γ-H2AX in tumour tissue: Tumour tissues were fixed in 10% formalin for 24 h, paraffin-embedded, sectioned, de-waxed, rehydrated, and treated with 3% H₂O₂ to block endogenous peroxidase. Antigen retrieval was performed with 10 mM citrate buffer (pH 6) at 800 W microwave for 30 min. Sections were blocked with 3% normal goat serum, incubated with anti-phospho-histone H2AX (Ser139) primary antibody (1:200) for 1 h, then with avidin-biotin-peroxidase complex for 30 min, and developed with diaminobenzidine. Counterstaining with hematoxylin. Positive cells were counted per microscopic field (×20). [1] |
| Toxicity/Toxicokinetics |
- Body weight loss as toxicity indicator in mice: In LMeC xenograft model, single doses of 0.15 and 0.30 mg/kg caused median weight loss of 0% and 2.45%, respectively. Weekly ×3 doses caused 1.9% (0.15 mg/kg) and 15.81% (0.30 mg/kg) weight loss. In CMeC-1 model, weekly ×3 caused 4.4% (0.15 mg/kg) and 12.6% (0.30 mg/kg) weight loss. Weight regain occurred within 14 days after the final dose at higher dose levels. [1]
- Selectivity for tumour cells over normal cells: In vivo, DNA ICL levels were lower in peripheral blood mononuclear cells than in tumour cells, suggesting some selectivity. [1] |
| References |
|
| Additional Infomation |
SJG-136 has been used in therapeutic trials for various diseases, including recurrent fallopian tube cancer, secondary acute myeloid leukemia, new-onset myelodysplastic syndrome, recurrent epithelial ovarian cancer, and secondary myelodysplastic syndrome.
The DNA minor groove binder SG2000 is a sequence-selective pyrrolobenzodiazepine (PBD) dimer with potential antitumor activity. After intravenous injection, SG2000 preferentially covalently binds to the purine-GATC-pyrimidine sequence, and the imine/methanolamine moiety of SG2000 binds to the N2 site of guanine on both strands of DNA. This induces interstrand crosslinking, inhibiting DNA replication and gene transcription, thereby suppressing cell growth. The SG2000 adduct preferentially binds to the purine-GATC-pyrimidine sequence and therefore appears to be less susceptible to p53-mediated DNA excision repair. - Mechanism: SJG-136 is a PBD dimer that binds covalently to the N2 positions of guanine on opposite DNA strands, forming interstrand cross-links spanning four base pairs. It can also form intrastrand cross-links and monoadducts depending on sequence. The C2/C2′-exo-methylene groups flatten the C-rings, allowing a snug isohelical fit in the minor groove and producing non-distorting adducts that may evade DNA repair. [1] - In vitro potency comparison: The tetralactam analogue (21) lacking the imine moieties showed no covalent DNA interaction, only non-covalent (ΔTm = 0.78 °C), and had IC50 = 0.57 μM in CH1 cells, demonstrating the necessity of the imine for potent cytotoxicity. [1] - Clinical status: SJG-136 has entered human Phase I and Phase II clinical trials. Dose-limiting toxicities in humans included edema, dyspnea, fatigue, and delayed liver toxicity; no significant myelotoxicity was observed. [1] |
| Molecular Formula |
C31H32N4O6
|
|---|---|
| Molecular Weight |
556.6090
|
| Exact Mass |
556.232
|
| Elemental Analysis |
C, 63.26; H, 5.48; N, 9.52; O, 21.74
|
| CAS # |
232931-57-6
|
| PubChem CID |
393111
|
| Appearance |
Off-white to yellow solid powder
|
| Density |
1.36g/cm3
|
| Boiling Point |
805.5ºC at 760 mmHg
|
| Flash Point |
441ºC
|
| Vapour Pressure |
5.52E-26mmHg at 25°C
|
| Index of Refraction |
1.66
|
| LogP |
3.271
|
| Hydrogen Bond Donor Count |
0
|
| Hydrogen Bond Acceptor Count |
8
|
| Rotatable Bond Count |
8
|
| Heavy Atom Count |
41
|
| Complexity |
1020
|
| Defined Atom Stereocenter Count |
2
|
| SMILES |
COC1=C(C=C2C(=C1)C(=O)N3CC(=C)C[C@H]3C=N2)OCCCOC4=C(C=C5C(=C4)N=C[C@@H]6CC(=C)CN6C5=O)OC
|
| InChi Key |
RWZVMMQNDHPRQD-SFTDATJTSA-N
|
| InChi Code |
InChI=1S/C31H32N4O6/c1-18-8-20-14-32-24-12-28(26(38-3)10-22(24)30(36)34(20)16-18)40-6-5-7-41-29-13-25-23(11-27(29)39-4)31(37)35-17-19(2)9-21(35)15-33-25/h10-15,20-21H,1-2,5-9,16-17H2,3-4H3/t20-,21-/m0/s1
|
| Chemical Name |
(6aS)-3-[3-[[(6aS)-2-methoxy-8-methylidene-11-oxo-7,9-dihydro-6aH-pyrrolo[2,1-c][1,4]benzodiazepin-3-yl]oxy]propoxy]-2-methoxy-8-methylidene-7,9-dihydro-6aH-pyrrolo[2,1-c][1,4]benzodiazepin-11-one
|
| Synonyms |
SJG-136; NSC-694501; SJG136; NSC694501; SJG 136; NSC 694501;
|
| 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)
|
| Solubility (In Vitro) |
DMSO : ~33.33 mg/mL (~59.88 mM)
|
|---|---|
| Solubility (In Vivo) |
Solubility in Formulation 1: ≥ 2.5 mg/mL (4.49 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 2: 2.08 mg/mL (3.74 mM) in 10% DMSO + 40% PEG300 + 5% Tween80 + 45% 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 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. View More
Solubility in Formulation 3: ≥ 2.08 mg/mL (3.74 mM) (saturation unknown) in 10% DMSO + 90% Corn Oil (add these co-solvents sequentially from left to right, and one by one), clear solution. |
| Preparing Stock Solutions | 1 mg | 5 mg | 10 mg | |
| 1 mM | 1.7966 mL | 8.9830 mL | 17.9659 mL | |
| 5 mM | 0.3593 mL | 1.7966 mL | 3.5932 mL | |
| 10 mM | 0.1797 mL | 0.8983 mL | 1.7966 mL |
*Note: Please select an appropriate solvent for the preparation of stock solution based on your experiment needs. For most products, DMSO can be used for preparing stock solutions (e.g. 5 mM, 10 mM, or 20 mM concentration); some products with high aqueous solubility may be dissolved in water directly. Solubility information is available at the above Solubility Data section. Once the stock solution is prepared, aliquot it to routine usage volumes and store at -20°C or -80°C. Avoid repeated freeze and thaw cycles.
Calculation results
Working concentration： mg/mL；
Method for preparing DMSO stock solution： mg drug pre-dissolved in μL DMSO (stock solution concentration mg/mL). Please contact us first if the concentration exceeds the DMSO solubility of the batch of drug.
Method for preparing in vivo formulation:：Take μL DMSO stock solution, next add μL PEG300, mix and clarify, next addμL Tween 80, mix and clarify, next add μL ddH2O，mix and clarify.
(1) Please be sure that the solution is clear before the addition of next solvent. Dissolution methods like vortex, ultrasound or warming and heat may be used to aid dissolving.
(2) Be sure to add the solvent(s) in order.
Products are for research use only; We do not sell to patients
Copyright 2020 InvivoChem LLC | All Rights Reserved
COA