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Fig. 1 · Structure
Cat. No.
A11041
CAS No.
941678-49-5
Formula
C17H18N6
Mol. Weight
306.4

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Ruxolitinib (INCB018424)

JAK1/2 Inhibitor

INCB018424 is a potent and selective inhibitor of Janus-associated kinase (JAK) 1 and 2, with IC50 to be 2.7, 4.5 and 332 nM for JAK1, JAK2 and JAK3 respectively;

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Publications for Ruxolitinib

7 publications citing this product

  1. Meenu Kesarwani, .et al. , Blood Adv, 2024, Jun 11;8(11):2765-2776 PMID: 38531054
  2. Aya Hasan Alshammari, .et al. , Biochem Pharmacol, 2022, Mar;197:114914 PMID: 35041812
  3. Daniel Doheny, .et al. , Oncogene, 2020, Oct;39(42):6589-6605 PMID: 32929154
  4. Rasmus Siersbek, .et al. , Cancer cell, 2020, Jul 6;S1535-6108(20)30311-1 PMID: 32679107
  5. Brittany M. Duggan, .et al. , Sci Rep, 2017, 7: 1578 PMID: 28484277
  6. Fujita KI, .et al. , J Pharm Sci, 2017, Sep;106(9):2632-2641 PMID: 28479358
  7. Kagoya Y, .et al. , Blood., 2014, Nov 6;124(19):2996-3006 PMID: 25217696
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Biological Activity

Bioactivity / Mechanism

INCB018424 (ruxolitinib) inhibited recombinant JAK1 and JAK2 with IC50 values of 3.3 ± 1.2 and 2.8 ± 1.2 nM, respectively (mean ± SD). In the same assay, TYK2 and JAK3 IC50 values were 19 ± 3.2 and 428 ± 243 nM. These results support preferential JAK1/2 inhibition within the tested JAK family. [1]

Literature-reported observations and conditions; not Adooq-tested protocols or general treatment recommendations.

Target & Activity

Target Activity Value
JAK2 IC50 2.8 nM
JAK1 IC50 3.3 nM
Tyk2 IC50 19 nM
JAK3 IC50 428 nM

Cellular Effect

Cell Line Activity Value Description Reference
BaF3 IC50 126 nM Cytotoxicity against mouse BAF3 cells expressing JAK2 V617F mutant after 48 hrs by CellTiterGlo assay 10.1039/C1MD00175B
HCT-116 IC50 > 10 μM Antiproliferative activity against human HCT116 cells after 72 hrs by MTT assay [PMID: 28953386]
HEL IC50 > 100 nM Inhibition of JAK-mediated STAT5 phosphorylation in HEL cells by Western blotting analysis 10.1039/C1MD00175B
HEL IC50 0.3 μM Synergistic antiproliferative activity against HEL cells harboring JAK2 V617F mutant assessed as reduction in cell viability after 48 hrs in presence of SAHA by MTT assay [PMID: 30901208]
HEL IC50 1.4 μM Antiproliferative activity against human HEL cells harboring JAK2 V617F mutant assessed as inhibition of cell growth incubated for 72 hrs by MTT assay [PMID: 33689932]
HEL IC50 18.6 μM Antiproliferative activity against HEL cells harboring JAK2 V617F mutant assessed as reduction in cell viability after 48 hrs by MTT assay [PMID: 30901208]
HEL IC50 2.62 μM Antiproliferative activity against HEL cells harboring JAK2 V617F mutant after 48 hrs by MTT assay [PMID: 27774135]
HEL IC50 7.639 μM Antiproliferative activity against HEL cells harboring JAK2 V617F mutant measured after 3 days by CCK8 assay [PMID: 30981578]
HEL 92.1.7 IC50 > 4 μM Antiproliferative activity against human HEL 92.1.7 cells after 36 hrs by PrestoBlue dye based assay [PMID: 28953386]
HEL 92.1.7 IC50 > 4 μM Antiproliferative activity against HEL 92.1.7 cells harboring JAK2 V617F mutant after 36 hrs by PrestoBlue dye based assay [PMID: 27541357]
HEL 92.1.7 IC50 14.7 μM Antiproliferative activity against HEL 92.1.7 cells assessed as viable cells measured after 3 days by WST-1 assay [PMID: 27555284]
HEL 92.1.7 IC50 18.06 μM Cytotoxicity against human HEL 92.1.7 cells expressing JAK2 assessed as reduction in cell viability incubated for 72 hrs by presto blue reagent assay [PMID: 34046625]
Jurkat IC50 > 5 μM Antiproliferative activity against human Jurkat cells assessed as reduction in cell viability after 48 hrs by MTT assay [PMID: 30901208]
K562 IC50 1.03 μM Synergistic antiproliferative activity against human K562 cells assessed as reduction in cell viability after 48 hrs in presence of SAHA by MTT assay [PMID: 30901208]
K562 IC50 10.3 μM Antiproliferative activity against human K562 cells after 48 hrs by MTT assay [PMID: 27774135]
K562 IC50 23.2 μM Antiproliferative activity against human K562 cells assessed as reduction in cell viability after 48 hrs by MTT assay [PMID: 30901208]
KMS-12-BM IC50 > 10 μM Antiproliferative activity against human KMS-12-BM cells after 48 hrs by CellTiter-Glo luminescent assay [PMID: 28953386]
MCF7 IC50 > 10 μM Antiproliferative activity against human MCF7 cells after 72 hrs by MTT assay [PMID: 28953386]
MCF7 IC50 > 5 μM Antiproliferative activity against human MCF7 cells after 48 hrs by MTT assay [PMID: 27774135]
MDA-MB-231 IC50 > 10 μM Antiproliferative activity against human MDA-MB-231 cells after 72 hrs by MTT assay [PMID: 28953386]

20 records · Scroll to view more

In Vitro Studies

After 48 hours, INCB018424 reduced viable cell number in Ba/F3-EpoR-JAK2V617F cells (IC50 126 nM in the main Results) and HEL cells (EC50 186 nM). HEL growth was incompletely inhibited, and BCR-ABL-expressing comparators showed no significant viability response. [1]

In AB555P breast-tumor explants, 500 nM ruxolitinib for two days reduced pSTAT3 staining and MYC/BCL3 mRNA. Ki67 and ELF3 decreases were nonsignificant (p=0.099 and p=0.098). Samples were pieces from one PDX tumor cultured on the same sponge. [2]

JAK-family biochemical inhibition

Biochemical assay · Recombinant human JAK1, JAK2, JAK3 and TYK2 kinase domains [1]

INCB018424 inhibited JAK1 and JAK2 more potently than JAK3 in the same biochemical assay. TYK2 inhibition was also observed.

Measured endpoints — not treatment concentrations
EndpointReported resultReference
JAK1 kinase inhibitionIC50 (nM; mean ± SD) 3.3 ± 1.2 nM; N = 7[1]
JAK2 kinase inhibitionIC50 (nM; mean ± SD) 2.8 ± 1.2 nM; N = 8[1]
JAK3 kinase inhibitionIC50 (nM; mean ± SD) 428 ± 243 nM; N = 5[1]
TYK2 kinase inhibitionIC50 (nM; mean ± SD) 19 ± 3.2 nM; N = 8[1]

Literature-Reported Treatment Conditions

Treatment concentration
Concentration-response assay; tested concentration series not specified in the reviewed assay description. IC50 values are endpoints, not treatment doses.
Exposure duration
1 hour
Method
Homogeneous time-resolved fluorescence kinase assay
Vehicle
2.0% DMSO
Co-treatment
None
Reported limits
Exact chemical salt, analytical stereochemistry, concentration series and identity of control condition beyond assay control were not specified in reviewed assay text.
Methods and source locations

Compound reported in the paper: INCB018424 (ruxolitinib); Compound label in the paper: INCB018424

Evidence for the named compound in this model; salt/form and lot equivalence to SKU A11041 were not analytically established.

Table 1, enzyme-assay rows; Methods: Biochemical assays

JAK1 kinase inhibition: Homogeneous time-resolved fluorescence peptide assay; ATP 1 mM; 1 hour; signal inhibition versus control. — Table 1, JAK1 row; Methods: Biochemical assays

JAK2 kinase inhibition: Homogeneous time-resolved fluorescence peptide assay; ATP 1 mM; 1 hour; signal inhibition versus control. — Table 1, JAK2 row; Methods: Biochemical assays

JAK3 kinase inhibition: Homogeneous time-resolved fluorescence peptide assay; ATP 1 mM; 1 hour; signal inhibition versus control. — Table 1, JAK3 row; Methods: Biochemical assays

TYK2 kinase inhibition: Homogeneous time-resolved fluorescence peptide assay; ATP 1 mM; 1 hour; signal inhibition versus control. — Table 1, TYK2 row; Methods: Biochemical assays

Viable-cell response with BCR-ABL comparators

Cell-based study · Ba/F3-EpoR-JAK2V617F and HEL cells; Ba/F3-BCR-ABL and TF-1-BCR-ABL comparators [1]

INCB018424 reduced viable cell number in JAK2V617F-bearing cells after 48 hours. HEL growth was incompletely inhibited; BCR-ABL comparators showed no significant viability effect.

Measured endpoints — not treatment concentrations
EndpointReported resultReference
Ba/F3-EpoR-JAK2V617F viable cell numberIC50 126 nM[1]
HEL viable cell numberEC50 186 nM[1]
Response limits across cell linesQualitative HEL proliferation not completely inhibited; no significant effect in BCR-ABL comparators[1]

Literature-Reported Treatment Conditions

Treatment concentration
Increasing concentrations of INCB018424; exact complete dose series not tabulated. Do not interpret IC50/EC50 as fixed treatment concentrations.
Exposure duration
48 hours
Method
Cellular ATP determination (Cell-Titer Glo) or viable-cell counting; exact readout allocation by cell line not specified in shared methods.
Vehicle
0.2% final DMSO
Co-treatment
None
Reported limits
Replicate number and error-bar definition for Figure 1D not specified in its caption. Salt and analytical stereochemistry not given.
Methods and source locations

Compound reported in the paper: INCB018424 (ruxolitinib); Compound label in the paper: INCB018424

Evidence for the named compound in this model; salt/form and lot equivalence to SKU A11041 were not analytically established.

Figure 1D and caption; Results: Selective inhibition of JAK2V617F signaling, proliferation, and survival in cells, viability paragraph; Methods: Cell proliferation assay

Ba/F3-EpoR-JAK2V617F viable cell number: 48-hour cellular ATP determination or viable-cell counting; normalized to DMSO vehicle; nonlinear regression. — Results: viability paragraph linked to Figure 1D; Methods: Cell proliferation assay

HEL viable cell number: 48-hour viability assay; same shared proliferation-assay methods. — Results: viability paragraph; Figure 1D

Response limits across cell lines: Figure 1D concentration-response comparison; no exact values inferred from curves. — Figure 1D image and caption; Results: viability paragraph

Ruxolitinib response in AB555P breast-tumor explants

Cell-based study · ER-positive/PR-negative AB555P patient-derived breast tumor explant pieces cultured ex vivo [2]

Two-day treatment reduced pSTAT3 staining and MYC/BCL3 expression. Ki67 and ELF3 were lower numerically but did not reach p < 0.05. Pieces came from one PDX tumor and were cultured on the same sponge.

Measured endpoints — not treatment concentrations
EndpointReported resultReference
pSTAT3 Y705-positive stainingDirection; two-sided t-test p Decreased; p = 1.1e-06; n = 9–10 tumor pieces; error bars SEM[2]
Ki67-positive stainingDirection; two-sided t-test p Numerically decreased; p = 0.099; n = 9–10 tumor pieces; error bars SEM[2]
MYC, ELF3 and BCL3 mRNADirection; two-sided t-test p Decreased numerically: MYC p = 0.0074; ELF3 p = 0.098; BCL3 p = 0.00013; n = 7–8 pieces; SEM[2]

Literature-Reported Treatment Conditions

Treatment concentration
500 nM ruxolitinib
Exposure duration
2 days
Method
AB555P tumor pieces on media-soaked sponges; treatment versus no ruxolitinib; IHC and RNAscope.
Vehicle
Control labeled vehicle in Figure 6; solvent identity not specified for AB555P in reviewed methods.
Co-treatment
None; no exogenous IL6 treatment in the AB555P protocol.
Reported limits
Solvent identity and analytical chemical form not specified for AB555P. Tumor pieces are within-tumor samples, not independent patient replicates. STG335 conditions were not imported.
Methods and source locations

Compound reported in the paper: INCB018424 (ruxolitinib); Compound label in the paper: Ruxolitinib

Evidence for the named compound in this model; salt/form and lot equivalence to SKU A11041 were not analytically established.

Figure 6B-D and caption; Methods: Ex vivo explant experiments; corresponding Results paragraph

pSTAT3 Y705-positive staining: IHC quantified by Halo image analysis. — Figure 6B,C image and caption

Ki67-positive staining: IHC quantified by Halo; nonsignificant at 0.05. — Figure 6B,C image and caption

MYC, ELF3 and BCL3 mRNA: RNAscope dots per cell; TBP housekeeping control. ELF3 nonsignificant at 0.05. — Figure 6D image and caption

In Vivo Studies

In an endocrine-resistant breast PDX model, ruxolitinib reduced local invasion while producing only modest primary-tumor growth inhibition (~20% reported). Fulvestrant alone did not reduce invasion; the combination also reduced invasion. The retrieved main manuscript does not specify a ruxolitinib mass dose. [2]

In CSF3R-driven leukemia mouse models, ruxolitinib suppressed white-cell counts without eliminating the marked leukemic population. Adding BCI did not improve the reported white-cell or leukemic-burden responses over ruxolitinib alone, illustrating that blood-count control need not indicate clone eradication. [3]

Endocrine-resistant breast PDX response

Animal study · Female NSG mice bearing endocrine-resistant ER-positive/PR-positive intraductal patient-derived breast tumors [2]

Ruxolitinib produced modest primary-growth reduction and reduced invasion into surrounding tissue. Fulvestrant alone did not reduce invasion; the combination also reduced invasion. Tumor growth and invasion were distinct endpoints.

Measured endpoints — not treatment concentrations
EndpointReported resultReference
Primary tumor growth relative to vehicleApproximate reduction reported in text ~20% with ruxolitinib; ~30% with fulvestrant or combination[2]
Invasive area as percentage of total tumor areaQualitative comparison Reduced by ruxolitinib and combination; not reduced by fulvestrant alone[2]
Study groups and reported outcomes
Group / doseComparatorObserved resultReference
Ruxolitinib aloneVehicleModest growth reduction (~20% reported); reduced invasion.[2]
Fulvestrant alone
160 mg/kg		VehicleModest primary growth reduction (~30% reported); no reduction in invasion.[2]
Ruxolitinib + fulvestrant
Ruxolitinib: mass dose not found; fulvestrant: 160 mg/kg		Vehicle and monotherapy arms~30% growth reduction reported; invasion reduced. Synergy is not established by these observations.[2]
Ruxolitinib alone
Model: Female NSG mice bearing endocrine-resistant ER-positive/PR-positive intraductal patient-derived breast tumors
Route: Oral gavage
Frequency: Once daily
Treatment course: Start one week after tumor injection; duration not specified in reviewed text.
Vehicle: 0.5% methylcellulose in water
Reported limits: Ruxolitinib mg/kg dose not found in retrieved main manuscript methods or Figure 6 caption; volume 10 mL/kg is not a mass dose. Supplement was not acquired. Exact treatment duration and meaning of n=4–5 for histology are not specified in caption.
Fulvestrant alone
Model: Female NSG mice bearing endocrine-resistant ER-positive/PR-positive intraductal patient-derived breast tumors
Route: Subcutaneous injection
Frequency: Weekly
Treatment course: Start one week after tumor injection; last dose 3 days before sacrifice.
Vehicle: 4% DMSO in castor oil
Reported limits: Treatment end date not specified; one fulvestrant invasion outlier removed (Q=2%).
Ruxolitinib + fulvestrant
Model: Female NSG mice bearing endocrine-resistant ER-positive/PR-positive intraductal patient-derived breast tumors
Route: Ruxolitinib: oral gavage; fulvestrant: subcutaneous
Frequency: Ruxolitinib daily; fulvestrant weekly
Treatment course: Start one week after tumor injection; duration not specified in reviewed text.
Vehicle: Ruxolitinib: 0.5% methylcellulose in water; fulvestrant: 4% DMSO-castor oil
Reported limits: Ruxolitinib mass dose and exact course unavailable in retrieved main text.
Methods and source locations

Compound reported in the paper: INCB018424 (ruxolitinib); Compound label in the paper: Ruxolitinib

Evidence for the named compound in this model; salt/form and lot equivalence to SKU A11041 were not analytically established.

Figure 6E-I and caption; Methods: Mouse intraductal xenografts experiments; corresponding Results paragraph

Primary tumor growth relative to vehicle: Radiance-based growth; Figure 6E: 19, 22, 22 and 16 tumors across four arms, from 8 mice per arm; SEM; two-way ANOVA/Tukey. Tumors within mice are not independent animals. — Results: paragraph following Figure 6; Figure 6E caption

Invasive area as percentage of total tumor area: Histological quantification, automated cell detection with manual curation; n = 4–5, SEM; one-way ANOVA/Tukey. One fulvestrant outlier excluded using Prism Q=2%. — Figure 6I image and caption

Ruxolitinib alone · Figure 6E-I; Methods: Mouse intraductal xenografts experiments

Fulvestrant alone · Figure 6E-I; Methods: Mouse intraductal xenografts experiments

Ruxolitinib + fulvestrant · Figure 6E-I; Methods: Mouse intraductal xenografts experiments

Ruxolitinib with or without BCI in CSF3R-driven leukemia

Animal study · Mice with transplanted wild-type marrow Kit-positive cells expressing CSF3R T618I or T618I/Q741* [3]

Ruxolitinib lowered white-cell counts but did not eliminate the marked leukemic population. Adding BCI showed no improvement over ruxolitinib in the reported blood-count and leukemic-burden comparisons.

Measured endpoints — not treatment concentrations
EndpointReported resultReference
Peripheral white blood cell countDirection Suppressed with ruxolitinib; BCI addition did not improve the reported response[3]
Venus-positive cells in peripheral blood (leukemic-burden surrogate)Direction Leukemic cells persisted with ruxolitinib; no added reduction with BCI[3]

Conditions shared by the study groups

Treatment course
Begin 2 weeks after transplantation; stop date not specified.
Vehicle
DMSO stock diluted in PBS
Study groups and reported outcomes
Group / doseComparatorObserved resultReference
Ruxolitinib alone
50 mg/kg		Vehicle; BCI and combination comparatorsWBC suppression without eradication of marked leukemic cells.[3]
BCI + ruxolitinib
Ruxolitinib 50 mg/kg; BCI 10 mg/kg		Ruxolitinib alone, BCI alone and vehicleNo improvement over ruxolitinib in reported WBC and leukemic-burden comparisons.[3]
Ruxolitinib alone
Model: Mice with transplanted wild-type marrow Kit-positive cells expressing CSF3R T618I or T618I/Q741*
Route: Oral gavage
Frequency: Twice daily
Reported limits: Exact course and final vehicle composition not specified.
BCI + ruxolitinib
Model: Mice with transplanted wild-type marrow Kit-positive cells expressing CSF3R T618I or T618I/Q741*
Route: Ruxolitinib oral gavage; BCI intraperitoneal
Frequency: Ruxolitinib twice daily; BCI described as twice, interval not specified.
Reported limits: BCI interval, exact treatment stop date and final DMSO fraction not specified.
Methods and source locations

Compound reported in the paper: INCB018424 (ruxolitinib); Compound label in the paper: Ruxolitinib

Evidence for the named compound in this model; salt/form and lot equivalence to SKU A11041 were not analytically established.

Figure 3A-F; Results: Off-target inhibition of DUSP6 abrogates the antileukemic response to DUSP1 inhibition by BCI; Methods: Plasmids and inhibitors; Drug preparation and in vivo treatments

Peripheral white blood cell count: Figure 3A,D longitudinal WBC curves; 3 mice/group, representative of 2 independent experiments; mean ± SD. — Figure 3A,D and caption; corresponding Results paragraph

Venus-positive cells in peripheral blood (leukemic-burden surrogate): Figure 3C,F longitudinal marked-cell measurements; 3 mice/group, representative of 2 independent experiments; mean ± SD. — Figure 3C,F and caption; corresponding Results paragraph

Ruxolitinib alone · Figure 3A,C,D,F; Methods: Drug preparation and in vivo treatments

BCI + ruxolitinib · Figure 3A,C,D,F; Methods: Drug preparation and in vivo treatments

Specifications

Manufacturer Adooq Bioscience
Actions Inhibitor
CAS Number 941678-49-5
Applications JAK1/2 Inhibitor
Formula C17H18N6
Molecular Weight 306.4
Synonyms INCB18424; INCB018424
SMILES
C1CCC(C1)[C@@H](CC#N)N2C=C(C=N2)C3=C4C=CNC4=NC=N3

Storage & Solubility

In Vitro Solubility

Reported Solubility

SolventReported Solubility
DMSO 59 mg/mL (192.57 mM)
Water Insoluble
Ethanol 59 mg/mL (192.57 mM)

Preparing Stock Solutions

Stock preparation is unavailable for this product molecular weight.

Calculated stock concentrations do not establish solvent solubility. Refer to the reported solubility data for experimental guidance.

In Vivo Preparation

Concentration
4 mg/mL
Vehicle
2% DMSO+30% PEG 300+ddH2O

Storage

Store powder at -20°C, keep desiccated.
In powder form, the chemical is stable for 36 months.
In solution, store at -20°C and use within 1 month to prevent loss of potency. Aliquot to avoid multiple freeze/thaw cycles.

Research References

  1. [1] Quintás-Cardama A, Vaddi K, Liu P, et al. Preclinical characterization of the selective JAK1/2 inhibitor INCB018424: therapeutic implications for the treatment of myeloproliferative neoplasms. Blood (2010). PMID: 20130243 DOI: 10.1182/blood-2009-04-214957 Source material
  2. [2] Siersbæk R, Scabia V, Nagarajan S, et al. IL6/STAT3 Signaling Hijacks Estrogen Receptor α Enhancers to Drive Breast Cancer Metastasis. Cancer cell (2020). PMID: 32679107 DOI: 10.1016/j.ccell.2020.06.007 Source material
  3. [3] Kesarwani M, Kincaid Z, Azhar M, et al. Enhanced MAPK signaling induced by CSF3R mutants confers dependence to DUSP1 for leukemic transformation. Blood advances (2024). PMID: 38531054 DOI: 10.1182/bloodadvances.2023010830 Source material

Publications

7 publications citing this product

  1. Meenu Kesarwani, .et al. , Blood Adv, 2024, Jun 11;8(11):2765-2776 PMID: 38531054
  2. Aya Hasan Alshammari, .et al. , Biochem Pharmacol, 2022, Mar;197:114914 PMID: 35041812
  3. Daniel Doheny, .et al. , Oncogene, 2020, Oct;39(42):6589-6605 PMID: 32929154
  4. Rasmus Siersbek, .et al. , Cancer cell, 2020, Jul 6;S1535-6108(20)30311-1 PMID: 32679107
  5. Brittany M. Duggan, .et al. , Sci Rep, 2017, 7: 1578 PMID: 28484277
  6. Fujita KI, .et al. , J Pharm Sci, 2017, Sep;106(9):2632-2641 PMID: 28479358
  7. Kagoya Y, .et al. , Blood., 2014, Nov 6;124(19):2996-3006 PMID: 25217696

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