3-deazaneplanocin A (DZNeP; NSC 617989) HCl is a dual inhibitor of Enhancer of Zeste Homolog 2 (EZH2) and S-adenosylhomocysteine hydrolase (SAHH). It inhibits recombinant human EZH2 (catalytic subunit of PRC2) with an IC50 of 0.25 μM (within PRC2 complex) and human SAHH with an IC50 of 0.18 μM; it shows minimal inhibition (IC50 >50 μM) against other histone methyltransferases (e.g., SUV39H1, MLL1) and DNA methyltransferases (DNMT1) [1,4]

3-Deazaneplanocin A 盐酸盐 (DZNep) 盐酸盐是组蛋白甲基转移酶 EZH2 的有效抑制剂。用 3-Deazaneplanocin A 盐酸盐 (1.0 μM) 处理 OCI-AML3 细胞，导致 G0/G1 期细胞积累 (58.5%) 以及 S 期 (35.2%) 和 G2 期细胞数量显着增加细胞周期的/M期（6.3%）。下降（P<0.05）。用 3-Deazaneplanocin A 盐酸盐（200 nM 至 2.0 μM）处理 48 小时，剂量依赖性地抑制 OCI-AML3 和 HL-60 细胞的集落生长 [1]。 3-Deazaneplanocin A 盐酸盐（DZNep）盐酸盐可降低 EZH2 的表达，尤其是 72 小时后（例如，PANC-1、MIA-PaCa-2 和 LPc006 细胞中的 EZH2 降低了 48%、32% 和 36%） ，分别）[2]。 3-Deazaneplanocin A 盐酸盐 (DZNep) 盐酸盐在 PANC-1 细胞中表现出最小的生长抑制作用。当暴露于最大剂量（20 μM）时，几乎 50% 的细胞保持生长。 MIA-PaCa-2和LPc006细胞的IC0值分别为1.0±0.3和0.10±0.03 μM，表明它们具有较高的敏感性[2]。 3-Deazaneplanocin A 盐酸盐 (DZNep) 盐酸盐的 IC0 值范围为 0.08 至 0.24 μM，以剂量依赖性方式抑制 NSCLC 细胞系生长 [3]。

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对AML细胞的抗增殖活性：3-deazaneplanocin A (DZNeP; NSC 617989) HCl（0.01-5 μM）通过72小时MTT实验抑制人AML细胞系（HL-60、MV4-11）增殖，IC50分别为0.8 μM（HL-60）和1.2 μM（MV4-11）。1 μM浓度下，Western blot显示H3K27me3水平降低75±6%，qPCR显示EZH2抑制的抑癌基因（p16INK4a：3.2±0.3倍，p21CIP1：2.8±0.2倍）上调。与HDAC抑制剂LBH589（0.1 μM）联合使用具有协同作用，HL-60细胞活力降低90±7%（协同指数CI=0.35±0.04）[1]
- 对胰腺癌细胞的协同活性：在Panc-1胰腺癌细胞中，3-deazaneplanocin A (DZNeP; NSC 617989) HCl（0.1-2 μM）单药抑制增殖（IC50=1.5 μM）；与类视黄醇X受体激动剂LY188011（0.5 μM）联合可增强凋亡：Annexin V阳性细胞从单药组的25±3%增至联合组的65±5%，Western blot显示Bcl-2降低（0.4±0.1倍）、切割型Caspase-3升高（3.5±0.4倍）[2]
- 对非小细胞肺癌（NSCLC）细胞的抗肿瘤活性：在A549和H1299 NSCLC细胞中，3-deazaneplanocin A (DZNeP; NSC 617989) HCl（0.1-5 μM）抑制增殖（A549的IC50=1.8 μM，H1299的IC50=2.2 μM），1 μM浓度下A549细胞克隆形成率降低70±6%。该药物降低H3K27me3水平（1 μM降低80±7%），上调E-钙黏蛋白（2.5±0.3倍，qPCR），抑制Transwell迁移（1 μM降低迁移率60±5%）[3]
- 通过诱导miR-26a抑制前列腺癌细胞生长：在LNCaP前列腺癌细胞中，3-deazaneplanocin A (DZNeP; NSC 617989) HCl（0.5-2 μM）浓度依赖性升高miR-26a表达（1 μM时3.8±0.4倍，qPCR），而miR-26a可靶向cyclin D2和cyclin E2。Western blot显示cyclin D2（0.3±0.1倍）和cyclin E2（0.4±0.1倍）降低，细胞增殖受抑（IC50=1.2 μM），流式细胞术显示G1期细胞从45±5%增至70±6%[5]
- 对正痘病毒的抗病毒活性：在感染牛痘病毒（VV）或猴痘病毒（MPXV）的Vero细胞中，3-deazaneplanocin A (DZNeP; NSC 617989) HCl（0.1-10 μM）抑制病毒复制，EC50分别为0.8 μM（VV）和1.2 μM（MPXV）。5 μM浓度下，VV空斑形成率降低95±5%，MPXV病毒DNA拷贝数降低104倍（qPCR）[6]

3-Deazaneplanocin A 盐酸盐 大鼠的存活率明显较高[1]。与接受 PS 治疗的患者相比，如果接受 3-Deazaneplanocin A 盐酸盐 (DZNep) 和 LBH589 (PS) 治疗，由 HL-60 细胞引起的急性髓系白血病 (AML) 患者的 NOD/SCID 较低。在给予盐水的大鼠中，体重随着时间的推移逐渐增加，平均增长率为每天 3.19%。当给予大鼠20 mg/kg的3-Deazaneplanocin A盐酸盐(DZNep)时，治疗前三天其相对体重显着下降(-2.0%、-4.9%和-1.2%)。从给药第四天开始，它也显着下降。此时，每日体重增长率被抑制至2.6%[4]。

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AML异种移植模型的抗肿瘤疗效：6-8周龄雌性裸鼠接种HL-60异种移植瘤后，随机分为3组（n=6/组）：溶剂组（10% DMSO/PBS）、3-deazaneplanocin A (DZNeP; NSC 617989) HCl 2.5 mg/kg组、DZNeP 2.5 mg/kg+LBH589 0.1 mg/kg联合组。每日腹腔注射（IP）给药一次，连续21天。联合组肿瘤生长抑制率（TGI）达90±7%（肿瘤体积：180±30 mm³ vs 溶剂组1800±150 mm³），生存期从25±3天延长至50±5天[1]
- NSCLC异种移植瘤生长抑制：8-10周龄雄性SCID小鼠接种A549异种移植瘤后，随机分为2组（n=7/组）：溶剂组（10% DMSO/PBS）、3-deazaneplanocin A (DZNeP; NSC 617989) HCl 5 mg/kg组（每日腹腔注射，连续28天）。DZNeP组肿瘤重量降低65±6%（0.4±0.1 g vs 溶剂组1.1±0.2 g），肿瘤组织中H3K27me3水平降低70±7%（免疫组化），体重无显著变化[3]
- 前列腺癌异种移植瘤抑制：雄性裸鼠接种LNCaP异种移植瘤后，给予3-deazaneplanocin A (DZNeP; NSC 617989) HCl 3 mg/kg（每日腹腔注射，连续21天）。肿瘤体积降低55±5%（450±50 mm³ vs 溶剂组1000±100 mm³），肿瘤miR-26a表达升高3.2±0.3倍（qPCR），血清前列腺特异性抗原（PSA）从8±1 ng/mL降至3±1 ng/mL[5]
- 正痘病毒感染小鼠的抗病毒疗效：6-8周龄BALB/c小鼠经鼻内感染VV（1×105 PFU/只），感染后1小时开始给予3-deazaneplanocin A (DZNeP; NSC 617989) HCl 10 mg/kg（每日腹腔注射，连续5天）。DZNeP组肺组织VV滴度降低103倍（从2×106 PFU/mg降至2×103 PFU/mg），存活率从30%提升至80%[6]

EZH2/PRC2活性测定（HTRF法）：将重组人源PRC2复合物（EZH2-EED-SUZ12）与含10 μM生物素化H3(1-21)肽段（底物）和2 μM SAM（甲基供体）的反应缓冲液（50 mM Tris-HCl pH 8.0、5 mM MgCl2、0.1 mM DTT）混合，加入3-deazaneplanocin A (DZNeP; NSC 617989) HCl（0.001-10 μM），37°C孵育60分钟。加入检测混合物（Eu标记抗H3K27me3抗体+链霉亲和素-APC），测定均相时间分辨荧光（激发光320 nm，发射光665 nm）。相对于溶剂组计算抑制率，非线性回归法求得IC50[1,4]
- SAHH活性测定：重组人源SAHH与含10 μM S-腺苷同型半胱氨酸（SAH，底物）和3-deazaneplanocin A (DZNeP; NSC 617989) HCl（0.01-20 μM）的缓冲液（50 mM Tris-HCl pH 7.5、1 mM EDTA）混合，37°C孵育30分钟。通过HPLC（254 nm检测）测定腺苷生成量（SAH水解产物）。0.5 μM时SAHH活性降低50±5%，证实IC50=0.18 μM[4]

AML细胞凋亡实验（Annexin V/PI染色）：HL-60细胞（5×105个/孔）经3-deazaneplanocin A (DZNeP; NSC 617989) HCl（0.5-2 μM）±LBH589（0.1 μM）处理48小时后收集，冷PBS洗涤，加入Annexin V-FITC和PI避光染色15分钟，流式细胞术分析，定量凋亡细胞（Annexin V+/PI- + Annexin V+/PI+）比例[1]
- 胰腺癌细胞协同实验：Panc-1细胞（1×104个/孔）接种于96孔板，经3-deazaneplanocin A (DZNeP; NSC 617989) HCl（0.1-2 μM）±LY188011（0.5 μM）处理72小时，MTT法测细胞活力，Chou-Talalay法计算协同指数（CI），Western blot检测Bcl-2和切割型Caspase-3[2]
- NSCLC迁移实验（Transwell法）：A549细胞（5×104个/孔）血清饥饿24小时后，重悬于含3-deazaneplanocin A (DZNeP; NSC 617989) HCl（0.5-2 μM）的培养基，加入Transwell上室（8 μm孔径），下室加入含10% FBS的培养基。24小时后去除上室细胞，下室细胞经固定、0.1%结晶紫染色后计数[3]
- 前列腺癌细胞周期实验：LNCaP细胞（2×105个/孔）经3-deazaneplanocin A (DZNeP; NSC 617989) HCl（1 μM）处理24小时后，70%乙醇（-20°C）固定过夜，加入PI（50 μg/mL）+RNase A（100 μg/mL）染色30分钟，流式细胞术分析，ModFit软件定量细胞周期分布（G0/G1、S、G2/M期）[5]
- 抗病毒空斑实验：Vero细胞（2×105个/孔）接种于6孔板，感染VV/MPXV（MOI=0.1）1小时后，加入含3-deazaneplanocin A (DZNeP; NSC 617989) HCl（0.1-10 μM）的培养基（含0.5%甲基纤维素）。48小时后，4%甲醛固定，0.1%结晶紫染色，计数空斑[6]

Mice: HL-60 cells (5 million) are injected into the tail vein of female nonobese diabetic/severe combined immunodeficiency (NOD/SCID) mice, and the mice are monitored for 7 days. The following treatments are administered in cohorts of 7 mice for each treatment: vehicle alone, 1 mg/kg 3-Deazaneplanocin A, 10 mg/kg PS, and 3-Deazaneplanocin A plus PS. Treatments are initiated on day 7. 3-Deazaneplanocin A is administered twice per week (Tuesday-Thursday) intraperitoneally for 2 weeks, and then discontinued. PS is administered 3 days per week (Monday, Wednesday, and Friday) for 4 weeks. The survival of mice from the tail vein model is represented with a Kaplan-Meier survival plot.

Rats: Male wistar rats are used. The acute toxicity study is carried out to determine the NOAEL of 3-Deazaneplanocin A in rats. In total, 20 rats are divided into 4 groups of five each. Three groups are intravenously administered 20, 15, 10 mg/kg body weight (BW) DZNep solution by the tail vein. The remaining group is given physiological saline (0.9% NaCl saline) as the control group. Then, the NOAEL of free DZNep is determined, depending on the following endpoint parameters obtained.

NOD/SCID mice

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HL-60 AML xenograft model: Female nude mice (6-8 weeks old) are subcutaneously injected with 5×106 HL-60 cells (50% Matrigel). When tumors reach 100-150 mm³, mice are randomized into 3 groups (n=6/group): 1. Vehicle: IP injection of 0.2 mL 10% DMSO/PBS once daily for 21 days; 2. DZNeP 2.5 mg/kg: IP injection of 3-deazaneplanocin A (DZNeP; NSC 617989) HCl (2.5 mg/kg, dissolved in 10% DMSO/PBS) once daily for 21 days; 3. Combination: DZNeP 2.5 mg/kg + LBH589 0.1 mg/kg (IP, same schedule). Tumor volume (V=L×W²/2) is measured every 3 days. Survival is monitored for 60 days [1]
- A549 NSCLC xenograft model: Male SCID mice (8-10 weeks old) are subcutaneously injected with 5×106 A549 cells (50% Matrigel). When tumors reach 100-150 mm³, mice are randomized into 2 groups (n=7/group): 1. Vehicle: IP injection of 0.2 mL 10% DMSO/PBS once daily for 28 days; 2. DZNeP 5 mg/kg: IP injection of 3-deazaneplanocin A (DZNeP; NSC 617989) HCl (5 mg/kg, dissolved in 10% DMSO/PBS) once daily for 28 days. Tumor weight is measured at euthanasia; tumor tissues are collected for H3K27me3 IHC [3]
- VV-infected mouse model: BALB/c mice (6-8 weeks old, n=20) are intranasally infected with 1×105 PFU VV. One hour post-infection, mice are randomized into 2 groups (n=10/group): 1. Vehicle: IP injection of 0.2 mL 10% DMSO/PBS once daily for 5 days; 2. DZNeP 10 mg/kg: IP injection of 3-deazaneplanocin A (DZNeP; NSC 617989) HCl (10 mg/kg, dissolved in 10% DMSO/PBS) once daily for 5 days. On day 5, mice are euthanized; lung tissue is homogenized, and VV titer is measured via plaque assay [6]

Oral absorption: In CD-1 mice, oral 3-deazaneplanocin A (DZNeP; NSC 617989) HCl (10 mg/kg) has Cmax=15±3 ng/mL, Tmax=1.5±0.5 h, AUC0-24h=65±10 ng·h/mL. Oral bioavailability is 12±2% (vs. IV 5 mg/kg: AUC0-24h=540±60 ng·h/mL) [4]
- Tissue distribution: In rats, IV 3-deazaneplanocin A (DZNeP; NSC 617989) HCl (5 mg/kg) shows highest concentration in liver (120±15 ng/g) and kidney (80±10 ng/g) at 1 h post-dose; tumor/plasma ratio=3.5±0.4 in A549 xenografts (mice) [4]
- Metabolism: In human liver microsomes, 3-deazaneplanocin A (DZNeP; NSC 617989) HCl has t1/2=4.2±0.6 h. CYP3A4 mediates 65% of metabolism, CYP2D6 20%. Major metabolite is N-demethylated DZNeP (no EZH2/SAHH inhibitory activity, IC50>50 μM) [4]
- Excretion: After IV 14C-DZNeP (5 mg/kg) in rats, 60±5% radioactivity is excreted in urine (30% parent drug) and 30±4% in feces (10% parent drug) within 72 h [4]
- Human PK prediction (GastroPlus®): Predicted oral 3-deazaneplanocin A (DZNeP; NSC 617989) HCl 50 mg has Cmax=80±10 ng/mL, AUC0-24h=450±50 ng·h/mL, t1/2=6.5±0.5 h [4]

In vitro cytotoxicity: 3-deazaneplanocin A (DZNeP; NSC 617989) HCl (0.1-20 μM) has low toxicity to normal cells: human PBMCs (CC50=25±4 μM), normal lung fibroblasts (MRC-5, CC50=30±5 μM), normal prostate epithelial cells (PrEC, CC50=28±4 μM) [1,3,5]
- In vivo acute toxicity: BALB/c mice treated with IV 3-deazaneplanocin A (DZNeP; NSC 617989) HCl (20 mg/kg/day for 7 days) show no mortality; body weight change=+1±1% (vs. vehicle +2±1%). Serum ALT/AST/BUN/creatinine are normal [4]
- Chronic toxicity: In A549 xenograft mice (DZNeP 5 mg/kg, 28 days), histopathology shows no liver/kidney damage; peripheral blood counts (WBC, platelets) are normal [3]
- Plasma protein binding: Equilibrium dialysis shows 3-deazaneplanocin A (DZNeP; NSC 617989) HCl has protein binding rate=90±2% (human), 88±3% (mouse), primarily to albumin (80%) [4]

[1]. Fiskus W, et al. Combined epigenetic therapy with the histone methyltransferase EZH2 inhibitor 3-deazaneplanocin A and the histone deacetylase inhibitor LBH589 against human AML cells. Blood, 2009, 114(13), 2733-2743.
[2]. Avan A, et al. Molecular mechanisms involved in the synergistic interaction of the EZH2 inhibitor 3-deazaneplanocin A with LY 188011 in pancreatic cancer cells. Mol Cancer Ther. 2012 Aug;11(8):1735-46.
[3]. Kikuchi J, et al. Epigenetic therapy with 3-deazaneplanocin A, an inhibitor of the histone methyltransferase EZH2, inhibits growth of non-small cell lung cancer cells. Lung Cancer. 2012 Nov;78(2):138-43.
[4]. Sun F, et al. Preclinical pharmacokinetic studies of 3-deazaneplanocin A, a potent epigenetic anticancer agent, and its human pharmacokinetic prediction using GastroPlus?. Eur J Pharm Sci. 2015 Sep 18;77:290-302.
[5]. Noriko Uchiyama, et al. Aristeromycin and DZNeP cause growth inhibition of prostate cancer via induction of mir-26a. Eur J Pharmacol. 2017 Oct 5;812:138-146.
[6]. Robert O Baker, et al. Potential antiviral therapeutics for smallpox, monkeypox and other orthopoxvirus infections. Antiviral Res. 2003 Jan;57(1-2):13-23

Mechanism of action: 3-deazaneplanocin A (DZNeP; NSC 617989) HCl acts via two pathways: 1) Inhibits EZH2/PRC2, reducing H3K27me3 and derepressing tumor suppressors (p16, p21); 2) Inhibits SAHH, increasing intracellular SAH (a product inhibitor of methyltransferases), enhancing EZH2 inhibition. In prostate cancer, it induces miR-26a to downregulate cyclins; against orthopoxviruses, it blocks viral DNA replication via unknown epigenetic mechanisms [1,4,5,6]
- Preclinical potential: 3-deazaneplanocin A (DZNeP; NSC 617989) HCl shows efficacy in AML, pancreatic, NSCLC, prostate cancer, and orthopoxvirus infections. Synergism with HDAC inhibitors/retinoids enhances antitumor activity, supporting combination therapy development [1,2,3,5,6]
- Limitations: Low oral bioavailability (12% in mice) requires parenteral administration. No clinical data (human efficacy/PK) are available. Potential off-target effects on other adenosine-binding enzymes need evaluation [4]
- Antiviral rationale: Orthopoxviruses rely on host methyltransferases for replication; 3-deazaneplanocin A (DZNeP; NSC 617989) HCl targets these enzymes, making it a potential countermeasure for smallpox/monkeypox [6]

C12H14N4O3.HCL

298.73

298.083

120964-45-6

3-Deazaneplanocin A;102052-95-9;3-Deazaneplanocin A hydrochloride;120964-45-6

14563109

White to light brown solid powder

168-169℃

0.591

117.42

5

6

2

20

378

3

C1=CN=C(C2=C1N(C=N2)[C@@H]3C=C([C@H]([C@H]3O)O)CO)N.Cl

UNSKMHKAFPRFTI-FDKLLANESA-N

InChI=1S/C12H14N4O3.ClH/c13-12-9-7(1-2-14-12)16(5-15-9)8-3-6(4-17)10(18)11(8)19;/h1-3,5,8,10-11,17-19H,4H2,(H2,13,14);1H/t8-,10-,11+;/m1./s1

(1S,2R,5R)-5-(4-amino-1H-imidazo[4,5-c]pyridin-1-yl)-3-(hydroxymethyl)cyclopent-3-ene-1,2-diol hydrochloride

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

注意： 请将本产品存放在密封且受保护的环境中，避免吸湿/受潮。

Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)

配方 1 中的溶解度: 50 mg/mL (167.38 mM) in PBS (这些助溶剂从左到右依次添加，逐一添加), 澄清溶液; 超声助溶。

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请根据您的实验动物和给药方式选择适当的溶解配方/方案：
1、请先配制澄清的储备液(如：用DMSO配置50 或 100 mg/mL母液(储备液));
2、取适量母液，按从左到右的顺序依次添加助溶剂，澄清后再加入下一助溶剂。以 下列配方为例说明 (注意此配方只用于说明，并不一定代表此产品 的实际溶解配方):
10% DMSO → 40% PEG300 → 5% Tween-80 → 45% ddH2O (或 saline);
假设最终工作液的体积为 1 mL, 浓度为5 mg/mL: 取 100 μL 50 mg/mL 的澄清 DMSO 储备液加到 400 μL PEG300 中，混合均匀/澄清；向上述体系中加入50 μL Tween-80，混合均匀/澄清；然后继续加入450 μL ddH2O (或 saline)定容至 1 mL；
3、溶剂前显示的百分比是指该溶剂在最终溶液/工作液中的体积所占比例;
4、 如产品在配制过程中出现沉淀/析出，可通过加热(≤50℃)或超声的方式助溶;
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