NCT-503
目录号 : GC15943
A PHGDH inhibitor
Cas No.:1916571-90-8
Sample solution is provided at 25 µL, 10mM.
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Quality Control & SDS
- View current batch:
- Purity: >98.00%
- COA (Certificate Of Analysis)
- SDS (Safety Data Sheet)
- Datasheet
| Cell experiment [1]: | |
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Cell lines |
MDA-MB-468 cells |
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Preparation Method |
Cells were pretreated with 10 μM compound or an equivalent volume of DMSO in RPMI for 1 h. |
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Reaction Conditions |
10 μM NCT-503,1h |
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Applications |
NCT-503 treatment did not change intracellular glucose concentration |
| Animal experiment [2]: | |
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Animal models |
Female NOD.CB17-Prkdcscid/J mice, 6–8 weeks old |
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Preparation Method |
NCT-503 was prepared in a vehicle of 5% ethanol, 35% PEG 300, and 60% of an aqueous 30% hydroxypropyl-β-cyclodextrin (Sigma) solution, and injected intraperitoneally once daily. |
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Dosage form |
30 mg/kg NCT-503 injected intraperitoneally once daily |
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Applications |
NCT-503 treatment reduced the growth and weight of PHGDH-dependent MDA-MB-468 xenografts but did not affect those of PHGDH-independent MDA-MB-231 xenografts. PHGDH inhibition(NCT-503) also selectively increased necrosis in MDA-MB-468 but not MDA-MB-231 xenografts . Importantly, mice treated with the compound did not lose weight during the 24d treatment. Levels of NCT-503 in tumors were 3 μM at the conclusion of the experiment, validating exposure of the tumor to compound. |
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References: [1]. Pacold ME, Brimacombe KR,et,al. A PHGDH inhibitor reveals coordination of serine synthesis and one-carbon unit fate. Nat Chem Biol. 2016 Jun;12(6):452-8. doi: 10.1038/nchembio.2070. Epub 2016 Apr 25. Erratum in: Nat Chem Biol. 2016 Jul 19;12 (8):656. PMID: 27110680; PMCID: PMC4871733. |
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NCT-503 is a phosphoglycerate dehydrogenase (PHGDH) inhibitor with an IC50 of 2.5 μM[1]
Treatment of three PHGDH-independent cell lines and five PHGDHdependent cell lines with NCT-503 demonstrated that NCT-503 had EC50 values of 8-16 μM for the PHGDH-dependent cell lines, and no toxicity toward other PHGDH-independent cell lines [1]. When determined the GI50 of NCT-503 in the MHCC97L cell line and found that treatment of NCT-503 significantly reduced the relative ratio of NADPH/NADP+ in cells. NCT-503 could double the number of the apoptotic cells induced by Sorafenib[2]
Primary MM cells are sensitive to doses of the PHGDH inhibitor NCT-503, that are tolerated by PBMCs[4].PHGDH level was significantly increased in the lung adenocarcinoma PC9ER4 cells that acquired resistance to erlotinib. Perturbation of PHGDH by NCT-503, augmented the tumoricidal effect and restored sensitivity to erlotinib in cell lines and xenografts. ROS stress and DNA damage marker γH2AX were enhanced by NCT-503 [3].The combination treatment of NCT-503 and Physcion substantially inhibited hepatocellular carcinoma growth in vitro and in vivo[7]
In mice, NCT-503 exhibits favorable absorption, distribution, metabolism and excretion (ADME) properties. NCT-503 has good exposure, half-life (2.5 hr) and Cmax (20 uM in plasma) following intraperitoneal administration with significant partitioning into the liver and brain. NCT-503 treatment reduces the growth and weight of PHGDH-dependent MDA-MB-468 xenografts but does not affect the growth or weight of PHGDH-independent MDA-MB-231 xenografts[1].In C57BL/KaLwRij mice were injected with 5T33MM cells model, NCT-503 treatment did not reduce tumor load at the doses used but reduced tumor growth in combination with bortezomib[4,5].Adding NCT-503 to the diet of mice increased body weight and liver weight, and increased triglyceride content in liver. NCT-503 supplementation significantly inhibited PHGDH activity and decreased the serine content in the liver[6].Treatment of murine and human lung fibroblasts with NCT-503 decreased TGF-β-induced collagen protein synthesis. Mice treated with the PHGDH inhibitor NCT-503 beginning 7 days after intratracheal instillation of bleomycin had attenuation of lung fibrosis[8]
References:
[1]: Pacold ME, Brimacombe KR, et,al. A PHGDH inhibitor reveals coordination of serine synthesis and one-carbon unit fate. Nat Chem Biol. 2016 Jun;12(6):452-8. doi: 10.1038/nchembio.2070. Epub 2016 Apr 25. Erratum in: Nat Chem Biol. 2016 Jul 19;12 (8):656. PMID: 27110680; PMCID: PMC4871733.
[2]: Wei L, Lee D, et,al. Genome-wide CRISPR/Cas9 library screening identified PHGDH as a critical driver for Sorafenib resistance in HCC. Nat Commun. 2019 Oct 15;10(1):4681. doi: 10.1038/s41467-019-12606-7. PMID: 31615983; PMCID: PMC6794322.
[3]: Dong JK, Lei HM, et,al. Overcoming erlotinib resistance in EGFR mutation-positive lung adenocarcinomas through repression of phosphoglycerate dehydrogenase. Theranostics. 2018 Feb 12;8(7):1808-1823. doi: 10.7150/thno.23177. Erratum in: Theranostics. 2021 Feb 9;11(8):3963. PMID: 29556358; PMCID: PMC5858502.
[4]: Elsaadi S, Steiro I, et,al. Targeting phosphoglycerate dehydrogenase in multiple myeloma. Exp Hematol Oncol. 2021 Jan 4;10(1):3. doi: 10.1186/s40164-020-00196-w. PMID: 33397437; PMCID: PMC7784327.
[5]: Dong JK, Lei HM, et,al. Overcoming erlotinib resistance in EGFR mutation-positive lung adenocarcinomas through repression of phosphoglycerate dehydrogenase. Theranostics. 2018 Feb 12;8(7):1808-1823. doi: 10.7150/thno.23177. Erratum in: Theranostics. 2021 Feb 9;11(8):3963. PMID: 29556358; PMCID: PMC5858502.
[6]: He L, Liu Y, et,al. Exogenous and Endogenous Serine Deficiency Exacerbates Hepatic Lipid Accumulation. Oxid Med Cell Longev. 2021 Oct 19;2021:4232704. doi: 10.1155/2021/4232704. PMID: 34712382; PMCID: PMC8548146.
[7]: Dewdney B, Alanazy M, et,al. The effects of fructose and metabolic inhibition on hepatocellular carcinoma. Sci Rep. 2020 Oct 7;10(1):16769. doi: 10.1038/s41598-020-73653-5. PMID: 33028928; PMCID: PMC7541473.
[8]:Hamanaka RB, Nigdelioglu R, et,al. Inhibition of Phosphoglycerate Dehydrogenase Attenuates Bleomycin-induced Pulmonary Fibrosis. Am J Respir Cell Mol Biol. 2018 May;58(5):585-593. doi: 10.1165/rcmb.2017-0186OC. PMID: 29019702; PMCID: PMC5946329
NCT-503 是一种磷酸甘油酸脱氢酶 (PHGDH) 抑制剂，IC50 为 2.5 μM[1]
用 NCT-503 处理三种 PHGDH 非依赖性细胞系和五种 PHGDH 依赖性细胞系表明，NCT-503 对 PHGDH 依赖性细胞系的 EC50 值为 8-16 μM，并且对其他 PHGDH 非依赖性细胞无毒性[1] 行。当测定 MHCC97L 细胞系中 NCT-503 的 GI50 时，发现 NCT-503 的处理显着降低了细胞中 NADPH/NADP+ 的相对比例。 NCT-503可使索拉非尼诱导的凋亡细胞数量增加一倍[2]
原代 MM 细胞对 PBMC 耐受的 PHGDH 抑制剂 NCT-503 剂量敏感[4]。在对厄洛替尼产生耐药性的肺腺癌 PC9ER4 细胞中，PHGDH 水平显着升高. NCT-503 对 PHGDH 的扰动增强了肿瘤杀伤作用并恢复了细胞系和异种移植物对厄洛替尼的敏感性。 NCT-503 [3]增强了 ROS 应激和 DNA 损伤标志物 γH2AX。NCT-503 和 Physcion 的联合治疗在体外和体内显着抑制肝细胞癌的生长[7]< /sup>
在小鼠中，NCT-503 表现出良好的吸收、分布、代谢和排泄 (ADME) 特性。 NCT-503 在腹膜内给药后具有良好的暴露、半衰期（2.5 小时）和 Cmax（血浆中 20 uM），并显着分配到肝脏和大脑中。 NCT-503 处理可降低 PHGDH 依赖性 MDA-MB-468 异种移植物的生长和重量，但不影响 PHGDH 非依赖性 MDA-MB-231 异种移植物的生长或重量[1]。在 C57BL 中/KaLwRij 小鼠被注射 5T33MM 细胞模型，NCT-503 治疗在所用剂量下并未降低肿瘤负荷，但与硼替佐米[4,5] 联合治疗时可降低肿瘤生长。将 NCT-503 添加到小鼠的饮食增加了体重和肝脏重量，并增加了肝脏中甘油三酯的含量。补充 NCT-503 可显着抑制 PHGDH 活性并降低肝脏中的丝氨酸含量[6]。用 NCT-503 处理小鼠和人肺成纤维细胞可降低 TGF-β 诱导的胶原蛋白合成。在气管内滴注博来霉素后 7 天开始使用 PHGDH 抑制剂 NCT-503 治疗的小鼠肺纤维化有所减轻[8]
| Cas No. | 1916571-90-8 | SDF | |
| 化学名 | N-(4,6-dimethyl-2-pyridinyl)-4-[[4-(trifluoromethyl)phenyl]methyl]-1-piperazinecarbothioamide | ||
| Canonical SMILES | S=C(N1CCN(CC2=CC=C(C(F)(F)F)C=C2)CC1)NC3=NC(C)=CC(C)=C3 | ||
| 分子式 | C20H23F3N4S | 分子量 | 408.5 |
| 溶解度 | ≥ 40.9mg/mL in DMSO | 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 | 2.448 mL | 12.2399 mL | 24.4798 mL |
| 5 mM | 0.4896 mL | 2.448 mL | 4.896 mL |
| 10 mM | 0.2448 mL | 1.224 mL | 2.448 mL |
| 第一步：请输入基本实验信息（考虑到实验过程中的损耗，建议多配一只动物的药量） | ||||||||||
| 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 |
| 第二步：请输入动物体内配方组成（配方适用于不溶于水的药物；不同批次药物配方比例不同，请联系GLPBIO为您提供正确的澄清溶液配方） | ||||||||||
| % DMSO % % Tween 80 % saline | ||||||||||
| 计算重置 | ||||||||||
计算结果:
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，
体内配方配制方法：取 μL DMSO母液，加入 μL PEG300，混匀澄清后加入μL Tween 80，混匀澄清后加入 μL saline，混匀澄清。
1. 首先保证母液是澄清的；
2.
一定要按照顺序依次将溶剂加入，进行下一步操作之前必须保证上一步操作得到的是澄清的溶液，可采用涡旋、超声或水浴加热等物理方法助溶。
3. 以上所有助溶剂都可在 GlpBio 网站选购。
Genome-wide CRISPR/Cas9 library screening identified PHGDH as a critical driver for Sorafenib resistance in HCC
Sorafenib is the standard treatment for advanced hepatocellular carcinoma (HCC). However, the development of drug resistance is common. By using genome-wide CRISPR/Cas9 library screening, we identify phosphoglycerate dehydrogenase (PHGDH), the first committed enzyme in the serine synthesis pathway (SSP), as a critical driver for Sorafenib resistance. Sorafenib treatment activates SSP by inducing PHGDH expression. With RNAi knockdown and CRISPR/Cas9 knockout models, we show that inactivation of PHGDH paralyzes the SSP and reduce the production of αKG, serine, and NADPH. Concomitantly, inactivation of PHGDH elevates ROS level and induces HCC apoptosis upon Sorafenib treatment. More strikingly, treatment of PHGDH inhibitor NCT-503 works synergistically with Sorafenib to abolish HCC growth in vivo. Similar findings are also obtained in other FDA-approved tyrosine kinase inhibitors (TKIs), including Regorafenib or Lenvatinib. In summary, our results demonstrate that targeting PHGDH is an effective approach to overcome TKI drug resistance in HCC.
Inhibiting PHGDH with NCT-503 reroutes glucose-derived carbons into the TCA cycle, independently of its on-target effect
The small-molecule inhibitor of phosphoglycerate dehydrogenase, NCT-503, reduces incorporation of glucose-derived carbons into serine in vitro. Here we describe an off-target effect of NCT-503 in neuroblastoma cell lines expressing divergent phosphoglycerate dehydrogenase (PHGDH) levels and single-cell clones with CRISPR-Cas9-directed PHGDH knockout or their respective wildtype controls. NCT-503 treatment strongly reduced synthesis of glucose-derived citrate in all cell models investigated compared to the inactive drug control and independent of PHGDH expression level. Incorporation of glucose-derived carbons entering the TCA cycle via pyruvate carboxylase was enhanced by NCT-503 treatment. The activity of citrate synthase was not altered by NCT-503 treatment. We also detected no change in the thermal stabilisation of citrate synthase in cellular thermal shift assays from NCT-503-treated cells. Thus, the direct cause of the observed off-target effect remains enigmatic. Our findings highlight off-target potential within a metabolic assessment of carbon usage in cells treated with the small-molecule inhibitor, NCT-503.
Overcoming erlotinib resistance in EGFR mutation-positive lung adenocarcinomas through repression of phosphoglycerate dehydrogenase
How to improve the efficacy and reverse the resistance to epidermal growth factor receptor tyrosine kinase inhibitors (EGFR-TKIs), such as erlotinib, remains a major challenge in the targeted therapy of lung adenocarcinoma with EGFR-activating mutation. Phosphoglycerate dehydrogenase (PHGDH) is the key enzyme of de novo serine biosynthesis over-expressed in various types of cancer including lung cancer. Elevated PHGDH expression is correlated with a worse overall survival in clinical lung adenocarcinoma patients. Here we investigated the role of PHGDH in lung adenocarcinoma with the acquisition of resistance to erlotinib. Methods: The necessary genes required for the acquired erlotinib resistance in lung adenocarcinoma cells were screened out by RNA-Seq analysis. Then the protein and mRNA levels of PHGDH were confirmed by immunoblotting and qRT-PCR in the erlotinib resistant cells. The effects of PHGDH inhibition or overexpression on erlotinib resistance were examined using cell culture and tumor xenograft mouse models respectively. To explore mechanism, the ROS level and DNA damage marker, γH2AX, were tested by DCFH-DA staining and immunofluorescence after PHGDH inhibition. Results: We found that PHGDH level was significantly increased in the lung adenocarcinoma PC9ER4 and HCC827ER9 cells that acquired resistance to erlotinib. Perturbation of PHGDH by siPHGDH transfection or NCT-503, a small molecular PHGDH inhibitor, synergistically augmented the tumoricidal effect and restored sensitivity to erlotinib in cell lines and xenografts. Over-expression of PHGDH caused xenografts resistant to erlotinib. Furthermore, multiple DNA damage repair pathways related genes were changed by PHGDH depletion specifically in erlotinib resistant cells. ROS stress and DNA damage marker γH2AX were enhanced by siPHGDH and NCT-503, which was reversed by NAC. Conclusion: Our study indicated that PHGDH inhibition has potential therapeutic value in lung adenocarcinoma with the acquired resistance to EGFR-TKIs.
Targeting phosphoglycerate dehydrogenase in multiple myeloma
Background: Multiple myeloma (MM) is a hematological malignancy characterized by the clonal expansion of plasma cells in the bone marrow. To date, this disease is still incurable and novel therapeutic approaches are required. Phosphoglycerate dehydrogenase (PHGDH) is the first and rate-limiting enzyme in the de novo serine synthesis pathway, and it has been attributed to bortezomib-resistance in MM.
Methods: Two different PHGDH inhibitors, CBR5884 and NCT-503, were tested against human myeloma cell lines, primary MM cells from patients, and peripheral blood mononuclear cells isolated from healthy donors. The PHGDH inhibitors were then tested in combination with proteasome inhibitors in different MM cell lines, including proteasome-resistant cell lines. Furthermore, we confirmed the effects of PHGDH inhibition through knocking down PHGDH and the effect of NCT-503 in vivo in the 5T33MM mouse model.
Results: All the tested myeloma cell lines expressed PHGDH and were sensitive to doses of NCT-503 that were tolerated by peripheral blood mononuclear cells isolated from healthy donors. Upon testing bortezomib in combination with NCT-503, we noticed a clear synergy in several HMCLs. The sensitivity to bortezomib also increased after PHGDH knockdown, mimicking the effect of NCT-503 treatment. Interestingly, targeting PHGDH reduced the intracellular redox capacity of the cells. Furthermore, combination treatment with NCT-503 and bortezomib exhibited a therapeutic advantage in vivo.
Conclusions: Our study shows the therapeutic potential of targeting PHGDH in MM, and suggest it as a way to overcome the resistance to proteasome inhibitors.
Exogenous and Endogenous Serine Deficiency Exacerbates Hepatic Lipid Accumulation
Serine is involved in the regulation of hepatic lipid metabolism. However, whether exogenous or endogenous serine deficiency affects lipid accumulation in the liver and related mechanisms is unclear. Here, we investigated the effects of serine deficiency on hepatic fat accumulation in mice fed a serine-deficient diet or in mice supplemented with the D-3-phosphoglycerate dehydrogenase (PHGDH) inhibitor NCT-503. Both treatments produced an increase in body weight and liver weight and higher triglyceride content in the liver. Both treatments also exacerbated hepatic inflammatory responses and oxidative stress. Importantly, NCT-503 supplementation significantly inhibited PHGDH activity and decreased the serine content in the liver. Dietary serine deficiency significantly affected the colonic microbiota, characterized by a decreased ratio of Firmicutes/Bacteroidetes and decreased proportion of Bifidobacterium. Dietary serine deficiency additionally resulted in significantly decreased colonic and serum acetate and butyrate levels. The collective results indicate that NCT-503 supplementation may contribute to overaccumulation of hepatic lipid, by causing hepatic serine deficiency, while dietary serine deficiency may produce similar outcomes by affecting the gut-microbiota-liver axis.