E7046

EP4 ( IC50 = 13.5 nM ); EP4 ( Ki = 23.14 nM )

Palupiprant (E7046)通过选择性EP4的选择性抗氧化作用逆转PGE2对人骨髓细胞活化和活化的免疫抑制作用[2]。

在 CT-26 肿瘤模型中，Palupiprant/RT 组合引起 9 只动物的肿瘤抗记忆反应。在 4T1 模型中，与单独的负荷处理相比，Palupiprant 和 RT 的组合也产生了显着更好的肿瘤生长该组合通过抑制4T1肿瘤血管的自发性肺漂移显着提高生存率[1]。Palupiprant (150 mg/kg)抑制多种同基因肿瘤模型的生长。EP4信号缓冲器的阻断可促进抗肿瘤DC 窒息并窒息肿瘤的生长。Palupiprant 处理依赖于骨髓和 CD8C T 细胞的方式减少生长甚至底层已建立的肿瘤。另外，Palupiprant 和 E7777 (一种优先杀死 Treg 的 IL-2-白喉毒素融合蛋白)的共同产品良好地破坏了骨髓和Treg免疫抑制网络，从而在小鼠肿瘤模型中产生有效和持久的抗免疫反应[2]。

用无菌 CM 冲洗 BALB/c 小鼠的股骨以提取骨髓 (BM) 细胞。收获后，添加新鲜分离的 (0.5×10⁶) 小鼠 GM-CSF 重组体，并在 37°C 下使用 20 ng/mL ±PGE2 (10 nM ）。在第 3 天和第 6 天，添加新的 GM-CSF ± PGE2。体外分化后使用流式细胞术检查细胞。将 CT26、4T1 细胞上清液和/或 1 mM EP1 (SC-57089)、EP2 (ER-880696)、EP3 (L-798106) 或 EP4 (E7046) 拮抗剂添加到 BM 细胞中进行特定实验。为了评估分化的骨髓细胞对 T 细胞增殖的影响，将用 CFSE (1 mM) 染色并用抗 CD3/CD28 Dynabeads 刺激的 T 细胞与小鼠骨髓细胞共培养 72 小时。 CFSE 稀释液用于流式细胞术测量 T 细胞增殖。

In order to conduct tumor isograft efficacy studies, 6-week-old female BALB/c mice are implanted with 1×10⁵ CT26 or 4T1 cells, 8×10⁵ H22 cells per mouse s.c., or 1×10⁵ EMT6 cells in the mammary fat pad. Single-cell suspensions of 1x106 Pan02 cells per mouse and 2-3 mm³ SAI/N tumor fragments per mouse are injected into C57BL/6 and A/J mice, respectively. One thousand five hundred CT26 cells are injected subcutaneously into 6-week-old female nude mice, who do not have T cells, in order to study the function of T cells in the anti-tumor response. Tumor-bearing mice are randomly allocated to vehicle or treatment groups, and treatment regimens start when tumor sizes reach between 50 and 100 mm³. E7046 is taken orally (p.o.) as a suspension of 100 or 150 mg/kg in 0.5% MC every day for 21 days (QDx21). In the context of combination studies, E7777 is injected intravenously (i.v.) in saline at a dose of 2.5 mg/mouse in two to three injections spaced one week apart (Q7Dx2-3). Body weights and tumor volumes are measured two or three times per week. Mice are assigned to vehicle control, E7046 C E7777, anti-PD1 antibodies, or anti-mouse PD-1 C antimouse CTLA4 antibodies treatment groups in order to compare with current immunotherapies. For a total of 300 mg each, anti-PD-1 and anti-CTLA-4 antibodies (1 mg/mL) are given intraperitoneally (i.p.) in 100 mL, three times, four days apart (Q4Dx3). The control group receives isotype controls intraperitoneally at a dose of 1 mg/mL. Antimouse CD4 or anti-mouse CD8 antibodies, or their isotype controls, are injected intraperitoneally (i.p.) into a 100 mL volume every 4 days at a dose of 2.5 mg/mL, for a total of 4 injections per mouse (1 mg) in order to deplete CD4CT and CD8CT lymphocytes.

[1]. Oncoimmunology . 2017 Jun 28;6(8):e1338239.

Palupiprant is an orally bioavailable antagonist of the prostaglandin E2 (PGE2) receptor type 4 (EP4; EP-4), with potential immunomodulating and antineoplastic activities. Upon oral administration, palupiprant selectively targets, binds to and blocks the activity of immunosuppressive tumor-associated myeloid cells (TAMCs) in the microenvironment. This abolishes TAMC-dependent immunosuppression and reduces tumor cell proliferation. The presence of immunosuppressive myeloid cells in certain tumors is associated with a poor prognosis.

C22H18F5N3O4

483.395

483.121

C, 54.66; H, 3.75; F, 19.65; N, 8.69; O, 13.24

1369489-71-3

1369489-71-3

56944705

White to off-white solid powder

1.4±0.1 g/cm3

594.8±50.0 °C at 760 mmHg

313.5±30.1 °C

0.0±1.8 mmHg at 25°C

1.564

3.81

93.4

2

10

7

34

717

1

O=C(O)C1=CC=C([C@@H](NC(C2=C(OC3=CC=CC(C(F)(F)F)=C3)N(C)N=C2C(F)F)=O)C)C=C1

MKLKAQMPKHNQPR-NSHDSACASA-N

InChI=1S/C22H18F5N3O4/c1-11(12-6-8-13(9-7-12)21(32)33)28-19(31)16-17(18(23)24)29-30(2)20(16)34-15-5-3-4-14(10-15)22(25,26)27/h3-11,18H,1-2H3,(H,28,31)(H,32,33)/t11-/m0/s1

4-[(1S)-1-[[3-(difluoromethyl)-1-methyl-5-[3-(trifluoromethyl)phenoxy]pyrazole-4-carbonyl]amino]ethyl]benzoic acid

E7046; E-7046; E 7046

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

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

DMSO: ~100 mg/mL (~206.9 mM)

配方 1 中的溶解度: ≥ 2.08 mg/mL (4.30 mM) (饱和度未知) in 10% DMSO + 40% PEG300 + 5% Tween80 + 45% Saline (这些助溶剂从左到右依次添加，逐一添加), 澄清溶液。
例如，若需制备1 mL的工作液，可将100 μL 20.8 mg/mL澄清DMSO储备液加入400 μL PEG300中，混匀；然后向上述溶液中加入50 μL Tween-80，混匀；加入450 μL生理盐水定容至1 mL。
*生理盐水的制备：将 0.9 g 氯化钠溶解在 100 mL ddH₂O中，得到澄清溶液。

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配方 2 中的溶解度: ≥ 2.08 mg/mL (4.30 mM) (饱和度未知) in 10% DMSO + 90% (20% SBE-β-CD in Saline) (这些助溶剂从左到右依次添加，逐一添加), 澄清溶液。
例如，若需制备1 mL的工作液，可将 100 μL 20.8 mg/mL澄清DMSO储备液加入900 μL 20% SBE-β-CD生理盐水溶液中，混匀。
*20% SBE-β-CD 生理盐水溶液的制备（4°C，1 周）：将 2 g SBE-β-CD 溶解于 10 mL 生理盐水中，得到澄清溶液。

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配方 3 中的溶解度: ≥ 2.08 mg/mL (4.30 mM) (饱和度未知) in 10% DMSO + 90% Corn Oil (这些助溶剂从左到右依次添加，逐一添加), 澄清溶液。
例如，若需制备1 mL的工作液，可将 100 μL 20.8 mg/mL 澄清 DMSO 储备液加入到 900 μL 玉米油中并混合均匀。

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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;
5、为保证最佳实验结果，工作液请现配现用！
6、如不确定怎么将母液配置成体内动物实验的工作液，请查看说明书或联系我们;
7、 以上所有助溶剂都可在 Invivochem.cn网站购买。