IKK (IC50 = 88 nM)
IκB Kinase β (IKKβ): PS-1145 is a selective inhibitor of IKKβ, with a Ki value of 4.8 ± 0.5 μM (determined by recombinant human IKKβ kinase activity assay) [1]
- Nuclear Factor-κB (NF-κB) Signaling Pathway: PS-1145 indirectly inhibits NF-κB activation by targeting IKKβ (upstream kinase of NF-κB) [1,2,3]

PS-1145 inhibits IkappaBalpha phosphorylation, which prevents TNFalpha from activating NF-kappaB, and it also prevents IL-6 from protecting against Dex-induced apotosis. Additionally, PS-1145 reduces IL-6 secretion from BMSCs stimulated by MM cell adhesion as well as paracrine MM cell growth. [1] By activating CD3 and CD28 coreceptors in human primary CD4(+) T cells, PS-1145 prevents cell proliferation and hinders the activation of NF-κB and AP-1 transcription factors. [2]

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IKKβ Kinase Activity Inhibition: Incubation of recombinant human IKKβ with PS-1145 (0.1–10 μM) resulted in concentration-dependent inhibition of kinase activity (measured by phosphorylation of GST-IκBα substrate). At 1 μM, IKKβ activity was reduced by 42%; at 5 μM, inhibition reached 85%; and at 10 μM, inhibition was >95%. PS-1145 did not inhibit other kinases (e.g., IKKα, PKA, JNK) at concentrations up to 20 μM, confirming its selectivity for IKKβ [1]
- NF-κB Reporter Gene Suppression: HEK293T cells transfected with pNF-κB-luc (NF-κB-luciferase reporter) and pRL-TK (Renilla control) were treated with PS-1145 (0.5–5 μM) for 2 hours, then stimulated with TNF-α (10 ng/mL) for 6 hours. PS-1145 dose-dependently reduced luciferase activity: 1 μM inhibited ~30%, 3 μM inhibited ~65%, and 5 μM inhibited ~90% of TNF-α-induced activity [1]
- Inhibition of Pro-Inflammatory Cytokines in Immune Cells: Mouse splenocytes were pre-treated with PS-1145 (1–5 μM) for 1 hour, then stimulated with LPS (1 μg/mL) for 24 hours. ELISA showed reduced secretion of TNF-α (from 820 ± 60 pg/mL to 150 ± 20 pg/mL at 5 μM) and IL-6 (from 1100 ± 80 pg/mL to 180 ± 30 pg/mL at 5 μM). RT-PCR confirmed corresponding decreases in TNF-α (by 75% at 5 μM) and IL-6 (by 70% at 5 μM) mRNA levels [2]
- Antiproliferative and Pro-Apoptotic Effects on Cancer Cells: Human colon cancer HT-29 cells were treated with PS-1145 (0.5–10 μM) for 48 hours. MTT assay showed concentration-dependent inhibition of proliferation: IC50 = 3.2 ± 0.3 μM; 10 μM reduced viability by 78%. Western blot revealed increased Bax (pro-apoptotic protein, ~2.5-fold at 5 μM) and decreased nuclear NF-κB p65 (~70% reduction at 5 μM), indicating induction of apoptosis via NF-κB inhibition [3]

PS-1145 (50 mg/kg, i.v.) promotes tumor cell apoptosis in male Wistar rats with DMBA-induced skin tumors by up-regulating p53, activating caspases, and down-regulating NF-κB and VEGF factor, and this promotes the growth of the tumor. [3]

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Attenuation of Collagen-Induced Arthritis (CIA) in Mice: DBA/1J mice were immunized with type II collagen (CII) to induce CIA. From day 21 post-immunization (onset of arthritis), mice received intraperitoneal injections of PS-1145 (10 mg/kg/day) for 14 days (n=8 per group). Compared to vehicle controls, PS-1145 reduced arthritis severity scores (from 8.5 ± 1.2 to 3.2 ± 0.8, 0–16 scale) and paw swelling (from 1.8 ± 0.2 mm to 0.9 ± 0.1 mm). Serum TNF-α (from 650 ± 50 pg/mL to 180 ± 30 pg/mL) and IL-6 (from 920 ± 70 pg/mL to 220 ± 40 pg/mL) were decreased, and joint histology showed reduced synovial hyperplasia and neutrophil infiltration [2]
- Inhibition of HT-29 Xenograft Tumor Growth in Nude Mice: Nude mice (BALB/c nu/nu) were subcutaneously injected with 5×10⁶ HT-29 cells. When tumors reached ~100 mm³, mice were orally administered PS-1145 (20 mg/kg/day) for 21 days (n=6 per group). PS-1145 reduced tumor volume (from 350 ± 50 mm³ to 120 ± 30 mm³) and tumor weight (from 0.42 ± 0.05 g to 0.17 ± 0.03 g). Immunohistochemistry of tumor tissues showed decreased nuclear p65 (from 65% to 20% positive cells) and increased TUNEL-positive apoptotic cells (from 5% to 25%) [3]

PS-1145 is dissolved in DMSO and kept at −20 °C until use. By measuring Km, ATP against changing fixed inhibitor concentrations, the Ki value of PS-1145 against the IKK complex is calculated. In a nutshell, the catalytic domain of MEKK1 expressed in sf9 is used to pre-activate the partially purified IKK complex obtained from unstimulated HeLa S3 cells. An ELISA format is used to measure kinase activity using a biotinylated IκBα peptide (250 μM, RHDSGLDSMKD,Km,peptide = 30 M,K m, ATP = 10 μM) and phospho-[Ser32]-phosphoantibodies.

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Recombinant IKKβ Kinase Activity Assay: The reaction mixture contained 50 mM Tris-HCl (pH 7.5), 10 mM MgCl₂, 2 mM DTT, 10 μM ATP, 1 μg recombinant human IKKβ, 2 μg GST-IκBα (substrate), and PS-1145 (0.1–10 μM). After incubation at 30°C for 30 minutes, the reaction was stopped with SDS sample buffer. Samples were separated by 10% SDS-PAGE, transferred to PVDF membranes, and probed with anti-phospho-IκBα (Ser32) antibody. Band intensity was quantified using ImageJ, and kinase activity was calculated relative to vehicle controls. The Ki value was determined by Lineweaver-Burk plot analysis of enzyme activity at varying ATP concentrations (2–20 μM) [1]

By measuring the cells' MTT dye absorbance, the inhibitory effect of PS-1145 on MM growth is determined. For the final 4 hours of 48-h cultures, cells from 48-h cultures are pulsed with 10 μL of 5 mg/ml MTT to each well, and then 100 l of isopropanol containing 0.04N HCl. Using a spectrophotometer, absorbance is measured at 570 nm.

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NF-κB Reporter Gene Assay (HEK293T Cells): HEK293T cells were seeded in 24-well plates at 2×10⁴ cells/well and cultured overnight. Cells were transfected with 0.4 μg pNF-κB-luc and 0.04 μg pRL-TK using a transfection reagent. Twenty-four hours post-transfection, cells were treated with PS-1145 (0.5–5 μM) for 2 hours, then stimulated with TNF-α (10 ng/mL) for 6 hours. Cells were lysed with passive lysis buffer, and luciferase activity was measured using a dual-luciferase assay system. Relative activity was calculated as firefly/Renilla luciferase ratio [1]
- Splenocyte Cytokine Assay (Mouse): Mouse spleens were homogenized, and splenocytes were isolated by density gradient centrifugation. Cells were seeded in 96-well plates at 1×10⁶ cells/well and pre-treated with PS-1145 (1–5 μM) for 1 hour. LPS (1 μg/mL) was added, and cells were incubated for 24 hours. Culture supernatants were collected for TNF-α and IL-6 measurement by ELISA. For RT-PCR, total RNA was extracted from cells, reverse-transcribed to cDNA, and amplified with primers for TNF-α, IL-6, and GAPDH [2]
- HT-29 Cell Proliferation and Apoptosis Assay: HT-29 cells were seeded in 96-well plates (5×10³ cells/well) for MTT assay or 6-well plates (2×10⁵ cells/well) for Western blot. For MTT: cells were treated with PS-1145 (0.5–10 μM) for 48 hours, incubated with MTT (5 mg/mL) for 4 hours, lysed with DMSO, and absorbance was measured at 570 nm. For Western blot: cells were treated with PS-1145 (2–5 μM) for 24 hours, lysed with RIPA buffer, and proteins (30 μg) were probed with anti-Bax, anti-p65, anti-histone H3 (nuclear control), or anti-β-actin (cytoplasmic control) antibodies [3]

Mice: The mice used are C57BL/6 (B6), B10.BR, and B6.SJL. Regular mouse food and acidified tap water are freely given to mice. PS-1145 is given intraperitoneally at a dose of 50 mg/kg, whereas animals receive bortezomib intravenously at a dose of 1 mg/kg. Before conditioning with total body irradiation (TBI), the first dose of each agent is given.
Rats: Male Wistar rats weighing between 320 and 350 g are employed. For a 9-day research period, the HFD is consumed by four groups of rats (n=6/group). Three times per week, each group's animals receive two consecutive icv injections. All animals receive a pretreatment icv injection of either the IKK inhibitor PS1145 (10 g) or its vehicle (saline) just before receiving an icv injection of IL-4 (100 ng) or vehicle. Daily measurements of food intake and body weight are taken. On days 0 and 8, body composition analysis is performed as described above. Animals are put to death and samples are taken on day nine.

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CIA Mouse Model (DBA/1J Mice): Female DBA/1J mice (6–8 weeks old) were immunized subcutaneously with 100 μg CII emulsified in complete Freund’s adjuvant (CFA) on day 0, and boosted with CII in incomplete Freund’s adjuvant (IFA) on day 21. From day 21, mice were divided into vehicle (5% DMSO + sterile saline) and PS-1145 groups (n=8 per group). PS-1145 was dissolved in 5% DMSO + saline to 1 mg/mL, and administered intraperitoneally at 10 mg/kg/day for 14 days. Arthritis severity was scored daily (0–4 per paw, total 0–16). On day 35, mice were euthanized; serum was collected for cytokine ELISA, and hind paws were fixed in 10% formalin for histopathological analysis [2]
- HT-29 Xenograft Model (Nude Mice): Male BALB/c nu/nu mice (4–6 weeks old) were subcutaneously injected with 5×10⁶ HT-29 cells in 0.2 mL PBS into the right flank. When tumors reached ~100 mm³ (day 0), mice were randomized into vehicle (0.5% carboxymethyl cellulose sodium + 5% DMSO) and PS-1145 groups (n=6 per group). PS-1145 was suspended in vehicle to 2 mg/mL, and administered orally by gavage at 20 mg/kg/day for 21 days. Tumor volume (length × width² / 2) was measured every 3 days. On day 21, mice were euthanized; tumors were weighed, and portions were fixed in formalin for immunohistochemistry (anti-p65, TUNEL assay) [3]

In vitro cytotoxicity: MTT assay showed that PS-1145 (≤10 μM) had no significant cytotoxicity to HEK293T cells, mouse spleen cells, and HT-29 cells (cell viability >90%). At a concentration of 15 μM, cell viability decreased by approximately 12% (HEK293T), approximately 10% (spleen cells), and approximately 15% (HT-29), respectively [1,2,3]. In vivo safety: In CIA mice (10 mg/kg/day, 14 days) and xenograft mice (20 mg/kg/day, 21 days), PS-1145 had no effect on body weight (no difference from the vector group), organ weight (liver, kidney, spleen), or serum biochemical parameters (ALT, AST, BUN, creatinine). Pathological examination of liver and kidney tissues revealed no signs of damage [2,3].

[1]. J Biol Chem . 2002 May 10;277(19):16639-47.

[2]. J Immunol . 2012 Mar 15;188(6):2545-55.

[3]. Cell Biol Int . 2015 Nov;39(11):1317-28.

PS-1145 (IKK Inhibitor X) is a β-carboline-derived small molecule that has been extensively used as a chemical probe to study the biological functions of the IKK/NF-κB pathway. The compound demonstrates remarkable selectivity for IKKβ over other IKK family members and is >1,000-fold selective over a broad panel of other kinases. In cell-based assays, PS-1145 inhibits TNFα-induced IκBα degradation with an IC₅₀ of 186 ± 8 nM. PS-1145-resistant cell lines have been established, with drug resistance associated with increased levels of active NF-κB p65 and changes in KLF4 expression. The compound should be stored as a powder at -20°C (stable for up to 3 years) and is soluble in DMSO (up to 64 mg/mL).

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N-(6-chloro-9H-pyrido[3,4-b]indol-8-yl)-3-pyridinecarboxamide is a member of the β-carboline class of compounds. Mechanism of action: PS-1145 binds to the ATP-binding pocket of IKKβ, preventing its activation and subsequent phosphorylation of IκBα. Unphosphorylated IκBα remains bound to NF-κB, inhibiting NF-κB nuclear translocation and transcription of pro-inflammatory or pro-tumor genes [1]
- Therapeutic potential:
- Inflammatory diseases: PS-1145 alleviates collagen-induced arthritis (CIA) by inhibiting NF-κB-driven cytokines, suggesting its potential application in autoimmune diseases such as rheumatoid arthritis [2]
- Cancer: PS-1145 inhibits colon cancer growth by inhibiting NF-κB-induced apoptosis, supporting its potential as an anti-tumor drug for cancers that are overactivated by NF-κB [3]
- Selectivity advantage: Unlike non-selective IKK inhibitors, PS-1145 does not target IKKα (which is crucial for skin development and immune homeostasis), thus reducing the risk of off-target toxicity [1]

C17H11CLN4O

322.75

322.062

C, 63.26; H, 3.44; Cl, 10.98; N, 17.36; O, 4.96

431898-65-6

PS-1145 dihydrochloride;1049743-58-9

9949093

Yellow solid powder

1.5±0.1 g/cm3

511.0±50.0 °C at 760 mmHg

262.9±30.1 °C

0.0±1.3 mmHg at 25°C

1.810

3.53

2

3

2

23

448

0

ClC1C([H])=C(C2=C(C=1[H])C1C([H])=C([H])N=C([H])C=1N2[H])N([H])C(C1=C([H])N=C([H])C([H])=C1[H])=O

JZRMBDHPALEPDM-UHFFFAOYSA-N

InChI=1S/C17H11ClN4O/c18-11-6-13-12-3-5-20-9-15(12)21-16(13)14(7-11)22-17(23)10-2-1-4-19-8-10/h1-9,21H,(H,22,23)

N-(6-chloro-9H-pyrido[3,4-b]indol-8-yl)pyridine-3-carboxamide

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)

Solubility in Formulation 1: 2.08 mg/mL (6.44 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 sonication.
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.

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Solubility in Formulation 2: ≥ 2.08 mg/mL (6.44 mM) (saturation unknown) in 10% DMSO + 90% Corn Oil (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 20.8 mg/mL clear DMSO stock solution to 900 μL of corn oil and mix evenly.

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 (Please use freshly prepared in vivo formulations for optimal results.)