IC50: Tyk2: 0.5 nM; JAK2: 6 nM; JAK3: 8 nM ; JAK1:12 nM; Syk:32 nM; MST1:4 nM; ARK5:4 nM; MLK1:5 nM ; FMS:5 nM; AMPK:6 nM; TBK1:10 nM; MARK1:10 nM; PAR1B-a:13 nM; TSSK:14 nM; MST2:15 nM ; GCK:18 nM; JNK3:18 nM; Rsk2:20 nM; Rsk4:28 nM; CHK1:42 nM; Flt4:51 nM; Flt3:90 nM; Ret:105 nM ; Itk:194 nM

The inhibitory impact of cerulelatinib ranges from 0.37 to 10.02 µM on 60 CLL. Cerdulatinib promotes apoptosis in CLL in conjunction with PARP breakage and MCL-1 down-regulation. Cerdulatinib (2µM) can cause CLL cell death by overriding the microenvironment's support. Primary CLL cells that are both ibrutinib-sensitive and -resistant are prevented from proliferating by cerulelatinib (250–500 nM). In addition, ceruleanib inhibits the growth of BTKC481S-transfected cell lines, ibrutinib-sensitive and ibrutinib-resistant primary CLL cells, as well as the BCR and JAK-STAT signaling pathways. Moreover, cerdulatinib's suppression of SYK and JAK results in the downstream inhibition of ERK and AKT. The NF-kB pathway is inhibited by cerulelatinib[1]. The early activation marker CD69's (IC50=0.11 µM) capacity to increase its cell-surface expression is diminished by PRT062070 in activated B cells. Differential efficacy against cytokine JAK/STAT signaling pathways is demonstrated by PRT062070. BCR-signaling competent NHL cell lines undergo apoptosis when exposed to 1 or 3 µM of PRT062070[2]. Cerdulatinib exhibits inhibitory action against DLBCL cells of both the ABC and GCB classes. Through caspase 3 and PARP cleavage, ceruleanib also causes apoptosis in the DLBCL cell lines belonging to the GCB and ABC classes. Additionally, cerdulatinib inhibits the cell cycle in the DLBCL ABC and GCB subtypes by downregulating cyclin E and RB phosphorylation. In all DLBCL cell lines, ceruleaninb causes apoptosis and cell cycle arrest in response to BCR stimulation. Moreover, cerdulatinib inhibits BCR and JAK/STAT signaling in GCB DLBCL and ABC cell lines. Cerdulatinib causes primary human DLBCL samples to undergo cell death[3]. In a dose-dependent manner, cerulelatinib mostly suppresses BCR-induced signals between 0.3 and 1 μM. and especially in IGHV-unmutated samples that express larger levels of sIgM, CD49d+, or ZAP70+, or that have stronger BCR signaling capacity and responsiveness to IL4. By inhibiting the induction of MCL-1 and BCL-XL, cerulelatinib circumvents the protective effects of anti-IgM, IL4/CD40L, or NLC; BCL-2 expression remains unchanged. Moreover, cerdulatinib and venetoclax work together in vitro to elicit more apoptosis in samples treated with IL4/CD40L than either medication alone[4].

While there is a nonstatistically significant trend toward decreased ankle inflammation with PRT062070 (0.5 mg/kg), the 1.5, 3, and 5 mg/kg doses result in significant reductions in inflammation. The production of anticollagen antibodies is impacted by PRT062070. Following oral administration in mice, PRT062070 (15 mg/kg) inhibits BCR signaling and activation in the spleen and suppresses upregulation of splenic B-cell surface CD80/86 and CD69[2].

The heterogeneity and severity of certain autoimmune diseases and B-cell malignancies warrant simultaneous targeting of multiple disease-relevant signaling pathways. Dual inhibition of spleen tyrosine kinase (SYK) and Janus kinase (JAK) represents such a strategy and may elicit several benefits relative to selective kinase inhibition, such as gaining control over a broader array of disease etiologies, reducing probability of selection for bypass disease mechanisms, and the potential that an overall lower level suppression of individual targets may be sufficient to modulate disease activity. To this end, we provide data on the discovery and preclinical development of PRT062070 [4-(cyclopropylamino)-2-({4-[4-(ethylsulfonyl)piperazin-1-yl]phenyl}amino)pyrimidine-5-carboxamide hydrochloride], an orally active kinase inhibitor that demonstrates activity against SYK and JAK. Cellular assays demonstrated specific inhibitory activity against signaling pathways that use SYK and JAK1/3. Limited inhibition of JAK2 was observed, and PRT062070 did not inhibit phorbol 12-myristate 13-acetate–mediated signaling or activation in B and T cells nor T-cell antigen receptor–mediated signaling in T cells, providing evidence for selectivity of action[2].

Cell metabolic activity, cell growth and viability determination[1]
DLBCL cell lines were treated with various concentrations of cerdulatinib for up to 72 h. The metabolic activities of cells were determined with MTT assay according to manufacturer's instruction at 72 h time point. IC50 was calculated using the Sigma Plot generated with GraphPad Prism 6 software. Cell growth was measured at every 24 h counting live cells with flow cytometry as previously described. Cells were collected every 24 h and cell viability was determined by the PE Annexin V Apoptosis Detection Kit I.

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Cell cycle analysis[1]
DLBCL cells were treated with various concentrations of cerdulatinib for 48 h. Cells were incubated with 10 μM BrdU at 37°C for 2 h, and stained with PE conjugated anti-BrdU antibody (BD Biosciences) according to the supplier's manual. The percentage of cell cycle distribution was analyzed with FlowJo.

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Intracellular phosphospecific flow cytometry assays[1]
DLBCL cells were treated with cerdulatinib for 6 h followed by stimulation with 5 μg/mL of goat F (ab')2 anti-human IgM and IgG antibodies at 37°C for 15 min. Cells were fixed in 4% formaldehyde at room temperature for 10 min, and permeabilized with 100% methanol on ice for 20 min before flow cytometric analyses.

[1]. Guo A, et al. Dual SYK/JAK inhibition overcomes ibrutinib resistance in chronic lymphocytic leukemia: Cerdulatinib, but not ibrutinib, induces apoptosis of tumor cells protected by the microenvironment. Oncotarget. 2017 Feb 21;8(8):12953-12967.
[2]. Coffey G, et al. The novel kinase inhibitor PRT062070 (Cerdulatinib) demonstrates efficacy in models of autoimmunity and B-cell cancer. J Pharmacol Exp Ther. 2014 Dec;351(3):538-48.
[3]. Ma J, et al. Cerdulatinib, a novel dual SYK/JAK kinase inhibitor, has broad anti-tumor activity in both ABC and GCB types of diffuse large B cell lymphoma. Oncotarget. 2015 Dec 22;6(41):43881-96.
[4]. Blunt MD, et al. The Dual Syk/JAK Inhibitor Cerdulatinib Antagonizes B-cell Receptor and Microenvironmental Signaling in Chronic Lymphocytic Leukemia. Clin Cancer Res. 2017 May 1;23(9):2313-2324

B-cell receptor (BCR) and JAK/STAT pathways play critical roles in diffuse large B-cell lymphoma (DLBCL). Herein, we investigated the anti-lymphoma activity of cerdulatinib, a novel compound that dually targets SYK and JAK/STAT pathways. On a tissue microarray of 62 primary DLBCL tumors, 58% expressed either phosphorylated SYK or STAT3 or both. SYK and STAT3 are also phosphorylated in a panel of eleven DLBCL cell lines although ABC and GCB subtypes exhibited different JAK/STAT and BCR signaling profiles. In both ABC and GCB cell lines, cerdulatinib induced apoptosis that was associated with caspase-3 and PARP cleavage. The compound also blocked G1/S transition and caused cell cycle arrest, accompanied by inhibition of RB phosphorylation and down-regulation of cyclin E. Phosphorylation of BCR components and STAT3 was sensitive to cerdulatinib in both ABC and GCB cell lines under stimulated conditions. Importantly, JAK/STAT and BCR signaling can be blocked by cerdulatinib in primary GCB and non-GCB DLBCL tumor cells that were accompanied by cell death. Our work provides mechanistic insights into the actions of cerdulatinib, suggesting that the drug has a broad anti-tumor activity in both ABC and GCB DLBCL, at least in part by inhibiting SYK and JAK pathways.[3]

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At concentrations achievable in patients, cerdulatinib inhibited BCR- and IL4-induced downstream signaling in CLL cells using multiple readouts and prevented anti-IgM- and nurse-like cell (NLC)-mediated CCL3/CCL4 production. Cerdulatinib induced apoptosis of CLL cells, in a time- and concentration-dependent manner, and particularly in IGHV-unmutated samples with greater BCR signaling capacity and response to IL4, or samples expressing higher levels of sIgM, CD49d+, or ZAP70+ Cerdulatinib overcame anti-IgM, IL4/CD40L, or NLC-mediated protection by preventing upregulation of MCL-1 and BCL-XL; however, BCL-2 expression was unaffected. Furthermore, in samples treated with IL4/CD40L, cerdulatinib synergized with venetoclax in vitro to induce greater apoptosis than either drug alone.Conclusions: Cerdulatinib is a promising therapeutic for the treatment of CLL either alone or in combination with venetoclax, with the potential to target critical survival pathways in this currently incurable disease.[4]

C20H27N7O3S

445.54

445.18960

C, 53.92; H, 6.11; N, 22.01; O, 10.77; S, 7.20

1198300-79-6

Cerdulatinib hydrochloride;1369761-01-2

44595079

Typically exists as light gray to khaki solids at room temperature

1.4±0.1 g/cm3

741.9±70.0 °C at 760 mmHg

402.5±35.7 °C

0.0±2.5 mmHg at 25°C

1.683

0.37

141.93

O=C(C1=CN=C(NC2=CC=C(N3CCN(S(=O)(CC)=O)CC3)C=C2)N=C1NC4CC4)N

BGLPECHZZQDNCD-UHFFFAOYSA-N

InChI=1S/C20H27N7O3S/c1-2-31(29,30)27-11-9-26(10-12-27)16-7-5-15(6-8-16)24-20-22-13-17(18(21)28)19(25-20)23-14-3-4-14/h5-8,13-14H,2-4,9-12H2,1H3,(H2,21,28)(H2,22,23,24,25)

4-(cyclopropylamino)-2-[4-(4-ethylsulfonylpiperazin-1-yl)anilino]pyrimidine-5-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: ≥ 2.5 mg/mL (5.61 mM) (saturation unknown) in 10% DMSO + 40% PEG300 + 5% Tween80 + 45% Saline (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 25.0 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: ≥ 2.5 mg/mL (5.61 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 25.0 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.)