Biological Activity

Bioactivity / Mechanism

PCI-32765 inhibits BTK-linked B-cell receptor signaling. In primary human B lymphocytes stimulated with anti-IgM plus hydrogen peroxide, reported cellular IC50 values were 96.4 nM for BTK Y551 phosphorylation and 9.5 nM for ERK1/2 phosphorylation. These assay-specific values describe cellular pathway responses. [1]

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

Target & Activity

Target	Activity	Value
BTK	IC50	0.5nM
BLK	IC50	0.5nM
Bmx	IC50	0.8nM
CSK	IC50	2.3nM
FGR	IC50	2.3nM
BRK	IC50	3.3nM
Hck	IC50	3.7nM
EGFR	IC50	5.6nM
YES	IC50	6.5nM
ErbB2	IC50	9.4nM
ITK	IC50	10.7nM
JAK3	IC50	16.1nM
FRK	IC50	29.2nM
LCK	IC50	33.2nM
RET	IC50	36.5nM
FLT3	IC50	73nM
TEC	IC50	78nM
Abl	IC50	86nM
Fyn	IC50	96nM
RIPK2	IC50	152nM
c-Src	IC50	171nM
Lyn	IC50	200nM
PDGFRα	IC50	718nM
Fms	IC50	5.545μM
FER	IC50	8.07μM
JAK1	IC50	>10μM
JAK2	IC50	>10μM
NEK2	IC50	>10μM
p38	IC50	>10μM
PI3K	IC50	>10μM
PLK1	IC50	>10μM
RSK1	IC50	>10μM
Syk	IC50	>10μM

33 targets · Scroll to view more

Cellular Effect

Cell Line	Activity	Value	Description	Reference
A2780	EC50	20.1 μM	Cytotoxicity against human A2780 cells assessed as cell growth inhibition after 3 days by presto blue dye based plate reader method	[PMID: 27077228]
A-431	GI50	> 10 μM	Antiproliferative activity against human A431 cells after 72 hrs by CellTiter-Glo assay	[PMID: 28628824]
A-431	EC50	2.38 μM	Antiproliferative activity against human A431 cells expressing wild type EGFR incubated for 96 hrs measured on day 5 by CellTiterGlo assay	[PMID: 28853575]
A549	GI50	> 10 μM	Antiproliferative activity against human A549 cells after 72 hrs by CellTiter-Glo assay	[PMID: 28628824]
A549	IC50	21.79 μM	Antiproliferative activity against human A549 cells harboring wild type EGFR after 72 hrs by MTT assay	[PMID: 28734581]
B cell	IC50	8 nM	Antiproliferative activity against anti-IgM-stimulated human KB cells assessed as cell growth inhibition pretreated for 30 mins followed by anti-IgM stimulation for 72 hrs by celltiter-glo reagent based assay	[PMID: 33540357]
BaF3	IC50	> 1 μM	Antiproliferative activity against mouse BA/F3 cells expressing TEL-JAK3 after 72 hrs by cell titer glo assay	[PMID: 26258521]
BaF3	GI50	> 10 μM	Cytotoxicity against mouse BAF3 cells assessed as cell proliferation after 72 hrs by CelltiterGlo assay	[PMID: 26630553]
BaF3	GI50	> 10 μM	Antiproliferative activity against mouse BAF3 cells after 72 hrs by CellTiter-Glo assay	[PMID: 28956923]
BaF3	GI50	> 10 μM	Antiproliferative activity against mouse BA/F3 cells after 72 hrs by CellTiter-Glo assay	[PMID: 28628824]
BaF3	GI50	> 10 μM	Growth inhibition of mouse BAF3 cells after 72 hrs by CellTiter-Glo assay	[PMID: 28282122]
BaF3	GI50	0.12 μM	Inhibition of FLT3 ITD mutant (unknown origin) expressed in BaF3 cells assessed as inhibition of cell proliferation after 72 hrs by CelltiterGlo assay	[PMID: 26630553]
BaF3	GI50	0.12 μM	Antiproliferative activity against mouse BAF3 cells harboring FLT3-ITD mutation after 72 hrs by CellTiter-Glo assay	[PMID: 28956923]
BaF3	GI50	2.5 μM	Antiproliferative activity against mouse BAF3 cells harboring FLT3-ITD-F691L mutation after 72 hrs by CellTiter-Glo assay	[PMID: 28956923]
CMK	GI50	> 10 μM	Antiproliferative activity against human CMK cells after 72 hrs by CellTiter-Glo assay	[PMID: 28628824]
DOHH-2	IC50	0.023 μM	Cytotoxicity against human DOHH-2 cells expressing BTK and PI3kdelta assessed as inhibition of cell growth incubated for 72 hrs by celltiter-glo assay	[PMID: 35247756]
DOHH-2	IC50	0.0334 μM	Antiproliferative activity against human DOHH2 cells after 72 hrs by CellTiter-Glo luminescent cell viability assay	[PMID: 30522954]
DOHH-2	GI50	0.41 μM	Cytotoxicity against human DOHH2 cells assessed as growth inhibition after 72 hrs by CellTiter-Glo luminescent cell viability assay	[PMID: 24915291]
DOHH-2	IC50	0.5062 μM	Antiproliferative activity against human DOHH-2 cells incubated for 72 hrs by CellTiter-Glo assay	[PMID: 35939993]
DU-145	GI50	> 10 μM	Antiproliferative activity against human DU145 cells after 72 hrs by CellTiter-Glo assay

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In Vitro Studies

In isolated mouse ventricular myocytes, 1 μmol/L ibrutinib attenuated IGF-1-dependent calcium-transient activation after 24 ± 2 h. Ibrutinib alone left measured calcium-transient amplitude and decay unchanged. The extent of inhibition differed between independent experimental series; these acute cellular findings do not establish an organ-level or human effect. [2]

BTK-pathway phosphorylation in primary human B cells

Cell-based study · Primary human peripheral B lymphocytes from healthy volunteers; CD20-positive flow-cytometry gate [1]

PCI-32765 reduced BTK Y551 and ERK1/2 phosphorylation under anti-IgM/H2O2 stimulation. These are cellular pathway endpoints, with different reported IC50 values; Y551 is not described here as a direct biochemical autophosphorylation assay.

Measured endpoints — not treatment concentrations
Endpoint	Reported result	Reference
Intracellular BTK Y551 phosphorylation	IC50 96.4 nM	[1]
Intracellular ERK1/2 phosphorylation	IC50 9.5 nM	[1]

Literature-Reported Treatment Conditions

Treatment concentration: Serial dilutions of PCI-32765; endpoint IC50 values are not treatment doses.
Exposure duration: 30 min pretreatment; 10 min BCR stimulation at 37°C
Method: Anti-IgM F(ab′)2, 10 μg/mL; 3.3 mM hydrogen peroxide also added; fixation, permeabilization and intracellular phosphoflow.
Reported limits: Vehicle identity/concentration is not specified in the selected phosphoflow Methods. Biological donor number is not stated there. No washout/irreversibility inference is made.

Methods and source locations

Compound reported in the paper: PCI-32765; Compound label in the paper: PCI-32765

Findings apply to the named paper material and experimental conditions; they do not validate an Adooq catalog lot or establish clinical efficacy or safety.

Figure 3a (image and caption); Results, primary B-lymphocyte paragraph in “PCI-32765 directly affects the function of multiple immune cells in vitro”; Methods, Phosphoflow assays

Intracellular BTK Y551 phosphorylation: Phosphoflow; 10,000 CD20-positive events per well, triplicate wells. Event count is not biological n. — Figure 3a (image and caption); Results, primary B-lymphocyte paragraph in “PCI-32765 directly affects the function of multiple immune cells in vitro”; Methods, Phosphoflow assays

Intracellular ERK1/2 phosphorylation: Phosphoflow; pT202/204 antibody. Figure 3a includes anti-IgM alone and H2O2-alone controls as well as combined stimulation. — Figure 3a (image and caption); Results, primary B-lymphocyte paragraph in “PCI-32765 directly affects the function of multiple immune cells in vitro”; Methods, Phosphoflow assays

IGF-1-dependent calcium responses in isolated ventricular myocytes

Cell-based study · Isolated adult (10–12 weeks) C57BL6/J mouse ventricular myocytes; 24 ± 2 h culture [2]

Ibrutinib attenuated IGF-1-evoked calcium-transient activation in the first experimental series. Ibrutinib alone left measured amplitude and decay kinetics unchanged versus vehicle. A distinct independent comparator series is recorded separately and limits generalization of complete antagonism.

Measured endpoints — not treatment concentrations
Endpoint	Reported result	Reference
Ca-transient amplitude, IGF-1 + ibrutinib versus IGF-1 alone, Figure 1B	Mean ± SEM 0.19 ± 0.01 a.u. (n = 73) versus 0.40 ± 0.03 a.u. (n = 71); P < 0.05.	[2]
Ca-transient amplitude and decay kinetics with ibrutinib alone	Negative/contextual control Unchanged versus vehicle-treated cells (Results, Figure 1).	[2]

Literature-Reported Treatment Conditions

Treatment concentration: Ibrutinib 1 μmol/L; IGF-1 10 nmol/L where indicated.
Exposure duration: 24 ± 2 h
Method: Four main conditions: vehicle, ibrutinib alone, IGF-1 alone, IGF-1 + ibrutinib. Fura-2 AM epifluorescence during 0.5 Hz field stimulation.
Vehicle: Vehicle-treated control cells are stated; vehicle solvent/composition is not specified in the selected Methods.
Co-treatment: IGF-1 is a signaling stimulus. The IGF-1 + ibrutinib result is not an ibrutinib-alone effect.
Reported limits: Drug salt, catalog code and lot are not given in the selected Methods. Figure parentheses are retained in review notes without assigning an unreported n definition.

Methods and source locations

Compound reported in the paper: Ibrutinib; Compound label in the paper: Ibrutinib

Findings apply to the named paper material and experimental conditions; they do not validate an Adooq catalog lot or establish clinical efficacy or safety.

Figure 1A–D (viewed image/caption); Methods, Isolation and culture, Epifluorescence and Statistical analysis; Results, first subsection; Limitations

Ca-transient amplitude, IGF-1 + ibrutinib versus IGF-1 alone, Figure 1B: Text-reported values; F340/F380 fluorescence-ratio amplitude; one-way ANOVA with Tukey post-hoc test. n refers to measured myocytes in Results, not n independent animals. — Figure 1A–D (viewed image/caption); Methods, Isolation and culture, Epifluorescence and Statistical analysis; Results, first subsection; Limitations

Ca-transient amplitude and decay kinetics with ibrutinib alone: Fura-2 AM epifluorescence; comparison is within the first experimental series. — Figure 1A–D (viewed image/caption); Methods, Isolation and culture, Epifluorescence and Statistical analysis; Results, first subsection; Limitations

Independent calcium-response series with acalabrutinib comparator

Cell-based study · Isolated adult (10–12 weeks) C57BL6/J mouse ventricular myocytes; 24 ± 2 h culture [2]

In an explicitly independent experimental series, both ibrutinib and acalabrutinib attenuated the IGF-1-dependent calcium response but neither fully antagonized its amplitude. These are separate inhibitor treatments, not combination therapy.

Measured endpoints — not treatment concentrations
Endpoint	Reported result	Reference
Independent series comparing ibrutinib and acalabrutinib, Figure 5	Reproducibility/extent limitation Both attenuated IGF-1-induced calcium-transient activation; neither fully antagonized the amplitude response in this series.	[2]

Literature-Reported Treatment Conditions

Treatment concentration: IGF-1 10 nmol/L; ibrutinib 1 μmol/L or the separate acalabrutinib comparator 10 μmol/L.
Exposure duration: 24 ± 2 h
Method: Independent series: vehicle, IGF-1 alone, IGF-1 + ibrutinib, IGF-1 + acalabrutinib; Fura-2 AM epifluorescence at 0.5 Hz.
Vehicle: Vehicle-treated control cells are stated; vehicle solvent/composition is not specified in the selected Methods.
Co-treatment: IGF-1 is a signaling stimulus. The IGF-1 + ibrutinib result is not an ibrutinib-alone effect. Acalabrutinib was tested separately, not combined with ibrutinib.
Reported limits: Drug salt, catalog code and lot are not given in the selected Methods. Figure parentheses are retained in review notes without assigning an unreported n definition.

Methods and source locations

Compound reported in the paper: Ibrutinib; Compound label in the paper: Ibrutinib

Findings apply to the named paper material and experimental conditions; they do not validate an Adooq catalog lot or establish clinical efficacy or safety.

Figure 5A–C (viewed image/caption); Methods, Isolation and culture, Epifluorescence and Statistical analysis; Results, final subsection; Limitations

Independent series comparing ibrutinib and acalabrutinib, Figure 5: Independent experiment set, as specified in Methods. No exact amplitude is inferred from Figure 5 bars; qualitative comparison only. — Figure 5A–C (viewed image/caption); Methods, Isolation and culture, Epifluorescence and Statistical analysis; Results, final subsection; Limitations

In Vivo Studies

Oral PCI-32765 at 1.56–12.5 mg/kg/day for 18 days reduced clinical scores in mice with established collagen-induced arthritis. Higher doses produced clinical regression. At 12.5 mg/kg, splenic BTK occupancy was reported as 85–90% three hours after the final dose and returned to baseline by 24 hours. Synovial IL-10 was not significantly reduced, illustrating endpoint-specific effects. [1]

Therapeutic oral PCI-32765 in established mouse arthritis

Animal study · Male DBA1/1OlaHsd mice with established collagen-induced arthritis [1]

Once-daily oral treatment reduced clinical arthritis scores in a dose-dependent manner. The highest dose produced pronounced clinical regression, but splenic BTK occupancy was transient across the 24-hour dosing interval. Synovial IL-10 was not significantly reduced.

Measured endpoints — not treatment concentrations
Endpoint	Reported result	Reference
Daily clinical arthritis score	Treatment response Dose-dependent reduction; all four doses reported significantly different from vehicle (Figure 1a: *** P < 0.001; Mann–Whitney U-test); n = 12.	[1]
Splenic BTK occupancy after final 12.5 mg/kg dose	Target occupancy/time course 85–90% at 3 h; returned to baseline by 24 h (Results); Figure 1b n = 6 at each time point.	[1]
Synovial fluid IL-10 at 12.5 mg/kg/day	Limiting result Not significantly reduced versus vehicle; Figure 1e n = 12.	[1]

Conditions shared by the study groups

Route: Oral (PO)
Frequency: Once daily
Treatment course: 18 days
Vehicle: 1% methylcellulose, 0.4% Cremephor EL and 98.09% water, as printed in Drug formulation; percentages are not normalized here.

Study groups and reported outcomes
Group / dose	Comparator	Observed result	Reference
PCI-32765 1.56 mg/kg/day	Vehicle-treated CIA group; Figure 1a also includes dexamethasone 0.2 mg/kg as a positive comparator.	Clinical scores were reduced versus vehicle, but substantial residual disease remained in Figure 1a.	[1]
PCI-32765 3.125 mg/kg/day	Vehicle-treated CIA group; Figure 1a also includes dexamethasone 0.2 mg/kg as a positive comparator.	Clinical scores were reduced versus vehicle; symptoms were not fully reversed in Figure 1a.	[1]
PCI-32765 6.25 mg/kg/day	Vehicle-treated CIA group; Figure 1a also includes dexamethasone 0.2 mg/kg as a positive comparator.	Results report significant disease regression from day 2 to day 18.	[1]
PCI-32765 12.5 mg/kg/day	Vehicle-treated CIA group; Figure 1a also includes dexamethasone 0.2 mg/kg as a positive comparator.	Results report reversal of clinical arthritis symptoms after 10 days. At 3 h after the final dose, reported splenic BTK occupancy was 85–90%; occupancy returned to baseline by 24 h.	[1]

PCI-32765 1.56 mg/kg/day: Model: Male DBA1/1OlaHsd mice with established collagen-induced arthritis; enrollment clinical score 1.5 on a 0–5 scale; Reported limits: Figure 1a reports n = 12; final-dose occupancy subsets in Figure 1b report n = 6 at each time point. The retrieved caption does not define the clinical-curve error-bar type.

PCI-32765 3.125 mg/kg/day: Model: Male DBA1/1OlaHsd mice with established collagen-induced arthritis; enrollment clinical score 1.5 on a 0–5 scale; Reported limits: Figure 1a reports n = 12; final-dose occupancy subsets in Figure 1b report n = 6 at each time point. The retrieved caption does not define the clinical-curve error-bar type.

PCI-32765 6.25 mg/kg/day: Model: Male DBA1/1OlaHsd mice with established collagen-induced arthritis; enrollment clinical score 1.5 on a 0–5 scale; Reported limits: Figure 1a reports n = 12; final-dose occupancy subsets in Figure 1b report n = 6 at each time point. The retrieved caption does not define the clinical-curve error-bar type.

PCI-32765 12.5 mg/kg/day: Model: Male DBA1/1OlaHsd mice with established collagen-induced arthritis; enrollment clinical score 1.5 on a 0–5 scale; Reported limits: Figure 1a reports n = 12; final-dose occupancy subsets in Figure 1b report n = 6 at each time point. The retrieved caption does not define the clinical-curve error-bar type.

Methods and source locations

Compound reported in the paper: PCI-32765; Compound label in the paper: PCI-32765

Findings apply to the named paper material and experimental conditions; they do not validate an Adooq catalog lot or establish clinical efficacy or safety.

Figure 1a,b,e (image and caption); Results, CIA subsection; Methods, Drug formulation, CIA model, Probe assays, Synovial fluid cytokine measurements and Statistics

Daily clinical arthritis score: Daily clinical scoring; no exact curve values digitized. — Figure 1a,b,e (image and caption); Results, CIA subsection; Methods, Drug formulation, CIA model, Probe assays, Synovial fluid cytokine measurements and Statistics

Splenic BTK occupancy after final 12.5 mg/kg dose: PCI-33380 affinity-probe assay; Figure 1b plots unoccupied BTK, not occupied BTK. — Figure 1a,b,e (image and caption); Results, CIA subsection; Methods, Drug formulation, CIA model, Probe assays, Synovial fluid cytokine measurements and Statistics

Synovial fluid IL-10 at 12.5 mg/kg/day: Multiplex immunoassay of knee synovial-fluid lavage. Other measured inflammatory mediators decreased, but no exact concentrations are inferred from bars. — Figure 1a,b,e (image and caption); Results, CIA subsection; Methods, Drug formulation, CIA model, Probe assays, Synovial fluid cytokine measurements and Statistics

PCI-32765 1.56 mg/kg/day · Figure 1a,b,e (image and caption); Results, CIA subsection; Methods, Drug formulation, CIA model, Probe assays, Synovial fluid cytokine measurements and Statistics

PCI-32765 3.125 mg/kg/day · Figure 1a,b,e (image and caption); Results, CIA subsection; Methods, Drug formulation, CIA model, Probe assays, Synovial fluid cytokine measurements and Statistics

PCI-32765 6.25 mg/kg/day · Figure 1a,b,e (image and caption); Results, CIA subsection; Methods, Drug formulation, CIA model, Probe assays, Synovial fluid cytokine measurements and Statistics

PCI-32765 12.5 mg/kg/day · Figure 1a,b,e (image and caption); Results, CIA subsection; Methods, Drug formulation, CIA model, Probe assays, Synovial fluid cytokine measurements and Statistics

Specifications

Manufacturer	Adooq Bioscience
Actions	Inhibitor
CAS Number	936563-96-1
Applications	Bruton tyrosine kinase inhibitor
Formula	C₂₅H₂₄N₆O₂
Molecular Weight	440.5
Synonyms	PCI-32765
SMILES	C=CC(=O)N1CCC[C@H](C1)N2C3=C(C(=N2)C4=CC=C(C=C4)OC5=CC=CC=C5)C(=NC=N3)N

Storage & Solubility

In Vitro Solubility

Reported Solubility

Solvent	Reported Solubility
DMSO	81 mg/mL (183.88 mM)
Water	Insoluble
Ethanol	41 mg/mL (93.07 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: 9 mg/mL
Vehicle: 5% DMSO+30% PEG 300+5% Tween 80+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] Chang BY, Huang MM, Francesco M, et al. The Bruton tyrosine kinase inhibitor PCI-32765 ameliorates autoimmune arthritis by inhibition of multiple effector cells Arthritis research & therapy (2011). PMID: 21752263 DOI: 10.1186/ar3400 Source material
[2] Tarnowski D, Feder AL, Trum M, et al. Ibrutinib impairs IGF-1-dependent activation of intracellular Ca handling in isolated mouse ventricular myocytes Frontiers in cardiovascular medicine (2023). PMID: 37655217 DOI: 10.3389/fcvm.2023.1190099 Source material

Publications

6 publications citing this product

Daniel Tarnowski, .et al. , Front Cardiovasc Med, 2023, Aug 15:10:119009 PMID: 37655217
Marta G Fuster, .et al. , Pharmaceutics, 2023, Apr 7;15(4):1186 PMID: 37111671
Simonowski A, .et al. , Biochim Biophys Acta Mol Cell Res, 2019, Dec 11;1867(4):118622 PMID: 31837347
Carolin N. Zorn, .et al. , Sci Rep, 2018, 8: 15467 PMID: 30341350
Li T, .et al. , Oncogene, 2018, Nov;37(47):6180-6194 PMID: 30013190
Brittany M. Duggan, .et al. , Sci Rep, 2017, 7: 1578 PMID: 28484277

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Ibrutinib (PCI-32765)

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Biological Activity

Bioactivity / Mechanism

Target & Activity

Cellular Effect

In Vitro Studies

BTK-pathway phosphorylation in primary human B cells

Literature-Reported Treatment Conditions

IGF-1-dependent calcium responses in isolated ventricular myocytes

Literature-Reported Treatment Conditions

Independent calcium-response series with acalabrutinib comparator

Literature-Reported Treatment Conditions

In Vivo Studies

Therapeutic oral PCI-32765 in established mouse arthritis

Conditions shared by the study groups

Specifications

Storage & Solubility

In Vitro Solubility

Reported Solubility

Preparing Stock Solutions

In Vivo Preparation

Storage

Research References

Publications

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