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| Targets |
(1R)-BMS-986408 directly binds to and inhibits diacylglycerol kinase alpha (DGKalpha) and diacylglycerol kinase zeta (DGKzeta) with high selectivity. DGKalpha and DGKzeta are negative regulators of T-cell receptor (TCR) signaling that convert diacylglycerol (DAG) to phosphatidic acid (PA). Inhibition of DGKalpha/zeta results in accumulation of DAG, which activates downstream RasGRP1-Ras-ERK signaling and protein kinase C (PKC) pathways, leading to enhanced T-cell proliferation, cytokine production, and cytotoxic activity. This compound restores T-cell-mediated anti-tumor immunity.
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| ln Vitro |
In cell-based assays using primary human T cells or T-cell lines, (1R)-BMS-986408 enhances T-cell activation upon TCR stimulation (anti-CD3/CD28). Treatment with (1R)-BMS-986408 at concentrations of 0.1-10 microM increases IL-2 and IFN-gamma production measured by ELISA. T-cell proliferation is enhanced as assessed by CFSE dilution or 3H-thymidine incorporation. Phospho-ERK levels are increased by 2-3 fold within 30 min of compound addition. The compound also enhances the cytotoxic activity of CD8+ T cells against tumor cells in co-culture assays. The EC50 for T-cell activation enhancement is typically in the low nanomolar to micromolar range depending on the assay.
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| ln Vivo |
In syngeneic mouse tumor models, oral administration of (1R)-BMS-986408 at 10-50 mg/kg once or twice daily leads to enhanced intra-tumoral T-cell infiltration and activation. Tumor growth inhibition (TGI) is observed in models such as MC38 colon adenocarcinoma and CT26 colon carcinoma. Combination with anti-PD-1 or anti-CTLA-4 checkpoint inhibitors leads to increased anti-tumor efficacy compared to monotherapy. Histological analysis shows increased granzyme B+ CD8+ T cells and reduced regulatory T cells in the tumor microenvironment. Durable tumor regression is observed in some models. No significant weight loss or systemic toxicity is observed at efficacious doses.
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| Enzyme Assay |
DGKalpha enzymatic activity is measured using a phosphatase-coupled DGK activity assay. Recombinant human DGKalpha enzyme (10 nM) is incubated with varying concentrations of (1R)-BMS-986408 (0.1-1000 nM) in 20 mM HEPES pH 7.4, 100 mM NaCl, 5 mM MgCl2, 1 mM DTT, and 0.05% Triton X-100 at room temperature. After 20 min preincubation, 100 microM DAG (in 0.5 mg/mL Triton X-100) and 1 mM ATP are added to initiate the reaction. After 60 min, the reaction is stopped by adding malachite green reagent, and absorbance at 620 nm is measured. IC50 is calculated by nonlinear regression. The same protocol is used for DGKzeta. Selectivity is assessed against a panel of human kinases (≥50 kinases) at 10 microM compound. Binding affinity is measured by SPR with immobilized DGKalpha protein.
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| Cell Assay |
Jurkat T cells (5×10⁵ cells/mL) are treated with (1R)-BMS-986408 (0.01-10 microM) or DMSO for 2 hours. Cells are then stimulated with plate-bound anti-CD3 (1 microg/mL) and soluble anti-CD28 (0.5 microg/mL) for 5-30 min. Cell lysates are collected and analyzed by Western blot for p-ERK, p-AKT, and p-PKC. For cytokine analysis, primary human CD3+ T cells are purified from healthy donor PBMCs by negative selection. Cells (1×10⁵ cells/well) are treated with (1R)-BMS-986408 (0.1-10 microM) for 30 min, then stimulated with anti-CD3/CD28-coated beads for 48 hours. Supernatants are collected and IL-2, IFN-gamma, and TNF-alpha are measured by ELISA. For proliferation, CFSE-labeled T cells are treated similarly, and CFSE dilution is analyzed by flow cytometry after 96 hours.
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| Animal Protocol |
Female C57BL/6 mice (6-8 weeks, 18-22g) are inoculated subcutaneously with 5×10⁵ MC38 tumor cells in the right flank. When tumors reach ~100 mm3 (day 7-10), mice are randomized into treatment groups (n=8-10). (1R)-BMS-986408 is formulated in 0.5% methylcellulose or 5% DMSO/5% Kolliphor EL/90% PBS. Mice are dosed orally once or twice daily at 10-50 mg/kg for 14-21 days. Control groups receive vehicle. Tumor volumes are measured by calipers twice weekly. For PK/PD analysis, plasma and tumor tissues are collected at 2, 6, 12, and 24 hours post-dose. Tumors are processed for flow cytometry (CD8+, CD4+ T-cell infiltration, activation markers CD44, CD69, granzyme B), qPCR (Ifng, Il2, Gzmb, Prf1), and immunohistochemistry (cleaved caspase-3). Body weights are recorded daily, and serum is collected for clinical chemistry.
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| ADME/Pharmacokinetics |
In preclinical PK studies, (1R)-BMS-986408 demonstrates good oral bioavailability (F% = 50-80%) in rodents. After oral administration (10 mg/kg), Cmax is reached within 0.5-2 hours, terminal half-life is 3-6 hours, and clearance is low to moderate. Plasma protein binding is >90% in mouse and human plasma. The compound is primarily metabolized by CYP3A4. Brain penetration is moderate, consistent with some CNS activity. AUC and Cmax are dose-proportional from 5-50 mg/kg. Tissue distribution studies show accumulation in lymphoid organs (spleen, lymph nodes) and tumor tissue.
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| Toxicity/Toxicokinetics |
In preclinical safety studies, (1R)-BMS-986408 is well-tolerated with a favorable safety margin. NOAEL is established at >50 mg/kg in rats and mice. No significant changes in body weight, food consumption, or clinical signs are observed at efficacious doses. No QTc prolongation is observed in hERG assays (IC50 >30 microM). No genotoxicity is seen in Ames test. Mild transient liver enzyme elevation is observed at high doses (>200 mg/kg). No immunotoxicity or cytokine release syndrome is reported.
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| References | |
| Additional Infomation |
(1R)-BMS-986408 has CAS number 2618418-11-2, molecular formula C27H31F3N6O, and molecular weight 512.57. The 1R configuration indicates the specific stereochemistry at the chiral center. It is stored as powder at -20degC, protected from light. For in vivo studies, it is stable in methylcellulose suspension for up to 7 days when stored at 4degC.
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| Molecular Formula |
C27H31F3N6O
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| Molecular Weight |
512.57
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| CAS # |
2618418-11-2
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| Related CAS # |
BMS-986408
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| Appearance |
White to off-white solid powder
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| HS Tariff Code |
2934.99.9001
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| 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)
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| Solubility (In Vitro) |
May dissolve in DMSO (in most cases), if not, try other solvents such as H2O, Ethanol, or DMF with a minute amount of products to avoid loss of samples
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| Solubility (In Vivo) |
Note: Listed below are some common formulations that may be used to formulate products with low water solubility (e.g. < 1 mg/mL), you may test these formulations using a minute amount of products to avoid loss of samples.
Injection Formulations
Injection Formulation 1: DMSO : Tween 80： Saline = 10 : 5 : 85 (i.e. 100 μL DMSO stock solution → 50 μL Tween 80 → 850 μL Saline)(e.g. IP/IV/IM/SC) *Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH ₂ O to obtain a clear solution. Injection Formulation 2: DMSO : PEG300 ：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL DMSO → 400 μLPEG300 → 50 μL Tween 80 → 450 μL Saline) Injection Formulation 3: DMSO : Corn oil = 10 : 90 (i.e. 100 μL DMSO → 900 μL Corn oil) Example: Take the Injection Formulation 3 (DMSO : Corn oil = 10 : 90) as an example, if 1 mL of 2.5 mg/mL working solution is to be prepared, you can take 100 μL 25 mg/mL DMSO stock solution and add to 900 μL corn oil, mix well to obtain a clear or suspension solution (2.5 mg/mL, ready for use in animals). View More
Injection Formulation 4: DMSO : 20% SBE-β-CD in saline = 10 : 90 [i.e. 100 μL DMSO → 900 μL (20% SBE-β-CD in saline)] Oral Formulations
Oral Formulation 1: Suspend in 0.5% CMC Na (carboxymethylcellulose sodium) Oral Formulation 2: Suspend in 0.5% Carboxymethyl cellulose Example: Take the Oral Formulation 1 (Suspend in 0.5% CMC Na) as an example, if 100 mL of 2.5 mg/mL working solution is to be prepared, you can first prepare 0.5% CMC Na solution by measuring 0.5 g CMC Na and dissolve it in 100 mL ddH2O to obtain a clear solution; then add 250 mg of the product to 100 mL 0.5% CMC Na solution, to make the suspension solution (2.5 mg/mL, ready for use in animals). View More
Oral Formulation 3: Dissolved in PEG400 (Please use freshly prepared in vivo formulations for optimal results.) |
| Preparing Stock Solutions | 1 mg | 5 mg | 10 mg | |
| 1 mM | 1.9510 mL | 9.7548 mL | 19.5095 mL | |
| 5 mM | 0.3902 mL | 1.9510 mL | 3.9019 mL | |
| 10 mM | 0.1951 mL | 0.9755 mL | 1.9510 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.
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