BTK (IC50 = 1.3 nM); BMX (IC50 = 1.0 nM); ITK (IC50 = 440 nM); TEC (IC50 = 0.8 nM); RLK (IC50 = 1.2 nM); BLK (IC50 = 6.3 nM); EGFR (IC50 = 520 nM); ERBB2 (IC50 = 3900 nM); ERBB4 (IC50 = 11.3 nM)

With an IC50 of 1.3±0.5 nM, rilzabrutinib is a reversible covalent inhibitor of Bruton's tyrosine kinase (BTK). Additionally, when rilzabrutinib was evaluated against a panel of 251 additional kinases, it was shown to be extremely selective. Rilzabrutinib, which targets the cysteine of BTK, caused a delayed dissociation rate; 18 hours after the chemical was washed away in vitro, 79±2% of bound BTK was still present in PBMC. Complete reversibility of covalent cysteine binding occurs upon target denaturation. Rilzabrutinib suppressed human B cell proliferation (10% serum) produced by anti-IgM and B cell CD69 expression, with IC50 values of 5±2.4 nM and 123±38 nM, respectively [2].

Following the drug's removal from the bloodstream, rilzabrutinib continues to have pharmacodynamic effects that are consistent with a prolonged target residence duration. Additionally, in a dose-dependent manner, rilzabrutinib reversed and totally suppressed collagen-induced arthritis in rats, which associated target occupancy with the alleviation of the disease [2].

In this study, researchers assessed the effects of selective Btk inhibitors PRN1008 (rilzabrutinib) and PRN473 on platelet signalling and function mediated by CLEC-2 and GPVI. They used healthy donor and XLA platelets to determine off-target inhibitor effects. Inferior vena cava (IVC) stenosis and Salmonella infection mouse models were used to assess antithrombotic effects of PRN473 in vivo. PRN1008 and PRN473 potently inhibited CLEC-2-mediated platelet activation to rhodocytin. No off-target inhibition of SFKs was seen. PRN1008 treatment of Btk-deficient platelets resulted in minor additional inhibition of aggregation and tyrosine phosphorylation, likely reflecting inhibition of Tec. No effect on GPCR-mediated platelet function was observed. PRN473 significantly reduced the number of thrombi in podoplanin positive vessels following Salmonella infection and the presence of IVC thrombosis following vein stenosis. The potent inhibition of human platelet CLEC-2, and reduced thrombosis in in vivo models, together with the lack of off-target SFK inhibition and absence of bleeding reported in rilzabrutinib treated immune thrombocytopenia patients, suggest Btk inhibition as a promising antithrombotic strategy.[Selective Btk inhibition by PRN1008/PRN473 blocks human CLEC-2 & PRN473 reduces venous thrombosis formation in mice. Blood Adv. 2024 Jul 5:bloodadvances.2024012713]

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10, 20, 40 mg/kg

Rats with collagen-induced arthritis (CIA) model

Cmax was highly correlated with the magnitude of BTK occupancy 4 h post‐PRN1008 dosing (Figure 4). A sigmoidal Emax model, with a fitted γ and E0 provided the best fit of PRN1008 Cmax vs. occupancy data. The parameter estimates (% coefficient of variation) were: Emax 76.7 (13) %; EC50 46.5 (15) ng ml–1; E0 17.8 (42) %, and γ 2.1 (31). These results suggest a fairly steep exposure–response relationship, with low variability, and with 80% of maximal occupancy achieved at Cmax concentrations of approximately 100 ng ml–1 and above. The E0 value of 17.8% is consistent with the lower quantifiable range of the assay.
The robust relationship between PRN1008 Cmax and 4‐h occupancy, along with the very consistent occupancy decay rate of ~1.6% h–1 should allow for design of dosing regimens (dose and dose intervals) which precisely target various levels of occupancy over the course of a dose interval. It is noted that the nature of the PK and PK/PD relationships of PRN1008 may differ between healthy volunteers and patient populations, and should be further evaluated in future studies in patients.[1]

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This single-center, open-label, non-randomized, two-part, phase I study was conducted (1) to evaluate the absolute oral bioavailability of rilzabrutinib 400 mg tablet following an i.v. microtracer dose of ~100 μg [14C]-rilzabrutinib (~1 μCi) and single oral dose of 400 mg rilzabrutinib tablet (part 1), and (2) to characterize the absorption, metabolism, and excretion (AME) of 14C-radiolabeled rilzabrutinib following single oral dose (300 mg) of [14C]-rilzabrutinib (~1000 μCi; administered as a liquid) in healthy male participants (part 2). A total of 18 subjects were enrolled (n = 8 in part 1; n = 10 in part 2). The absolute bioavailability of 400 mg rilzabrutinib oral tablet was low (<5%). In part 1, rilzabrutinib was absorbed rapidly after single oral dose of rilzabrutinib 400 mg tablet with a median (range) time to maximum concentration (Tmax ) value of 2.03 h (1.83-2.50 h). The geometric mean (coefficient of variation) terminal half-life following the oral dose and i.v. microtracer dose of ~100 μg [14C]-rilzabrutinib, were 3.20 (51.0%) and 1.78 (37.6%) h, respectively. In part 2, rilzabrutinib was also absorbed rapidly following single oral dose of 300 mg [14C]-rilzabrutinib solution with a median (range) Tmax value of 1.00 h (1.00-2.00 h). The majority of total radioactivity was in the feces for both non-bile collection subjects (92.9%) and bile collection subjects (87.6%), and ~5% of radioactivity was recovered in urine after oral administration. Urinary excretion of unchanged rilzabrutinib was low (3.02%). The results of this study advance the understanding of the absolute bioavailability and AME of rilzabrutinib and can help inform its further investigation.Clin Transl Sci. 2023 Jul;16(7):1210-1219.

Data from all 80 enrolled subjects who received study drug (either PRN1008 or placebo) were included in the safety population. All participants were assessed for AEs for the duration of the study. No serious AEs or deaths were reported during the study, and no participants discontinued treatment due to an AE in either Part A or Part B. At PRN1008 single doses of 50 to 600 mg, safety and tolerability was similar to placebo. In these four cohorts, there was only one subject of the six treated in each cohort who experienced a treatment‐emergent AE (TEAE). Of these four TEAEs, only one was considered related to study drug (nausea in Cohort A4), and one was graded as moderate (toothache in Cohort A2, unrelated to study drug). This compares with two TEAEs reported in two of the 10 subjects who received placebo (both graded as mild, not drug related). In Cohort A5 (1200 mg), the primary AEs observed were gastrointestinal (GI) in nature, and were reported by each of the six subjects receiving PRN1008. The drug‐related AEs included diarrhoea (coded as loose stools; n = 6, 3 mild and 3 moderate severity), nausea (n = 3, 2 mild and 1 moderate severity), vomiting (n = 1), throat irritation (n = 3), and oropharyngeal discomfort (n = 1). There was no apparent relationship between GI AEs and PRN1008 pharmacokinetics. As described above, both Cmax and area under the concentration–time curve for PRN1008 were similar at the 600 mg and 1200 mg doses. Despite similar plasma PK, the increase in GI AEs for the 600 mg vs. 1200 mg dose levels would suggest a localized effect related to total administered dose, and not to plasma exposure. Following 10 or 11 days of dosing, PRN1008 was generally safe and well tolerated. TEAEs were reported in 7/8, 4/8, 8/8, 7/8 and 4/8 subjects in the 300 mg QD, 300 mg twice daily (BID), 600 mg QD, 450 mg BID and placebo groups, respectively. TEAEs classified as treatment‐related appeared to be more frequent in PRN1008 receiving subjects, reported in 6/8, 3/8, 8/8, 6/8 and 1/8 subjects in the 300 mg QD, 300 mg BID, 600 mg QD, 450 mg BID and placebo groups, respectively. All TEAEs classified as related to study drug were mild in intensity, with the exception of one subject in the 450 mg BID cohort who reported moderate diarrhoea. There were no clinically significant or dose‐dependent changes observed in haematology, biochemistry or coagulation laboratory parameters in either Part A or Part B of the study. Similarly, no clinically significant changes were observed in vital signs, ECGs or urinalysis evaluations.[1]

[1]. A phase I trial of PRN1008, a novel reversible covalent inhibitor of Bruton's tyrosine kinase, in healthy volunteers. Br J Clin Pharmacol. 2017 Nov;83(11):2367-2376.

[2]. Hill RJ, Bradshaw JM, Bisconte A, Tam D, Owens TD, Brameld KA, Smith PF, Funk JO, Goldstein DM, Nunn PA. Preclinical Characterization of PRN1008, a Novel Reversible Covalent Inhibitor of BTK that Shows Efficacy in a RAT Model of Collagen-Induced Arthritis. Annals of the Rheumatic Diseases 2015; 74(Suppl 2): 216.

Rilzabrutinib is an oral, reversible covalent inhibitor of Bruton's tyrosine kinase being investigated for the treatment of immune disorders, such as immune thrombocytopenic purpura.
Rilzabrutinib is an orally bioavailable reversible covalent inhibitor of Bruton's tyrosine kinase (BTK), with potential immunomodulatory and anti-inflammatory activities. Upon oral administration, rilzabrutinib inhibits the activity of BTK. This prevents the activation of the B-cell antigen receptor (BCR) signaling pathway, and the resulting immune activation and inflammation. BTK, a cytoplasmic tyrosine kinase and member of the Tec family of kinases, plays an important role in B-lymphocyte development, activation, signaling, proliferation and survival. In addition to B-cells, BTK is also expressed in other cells of hematopoietic origin, including monocytes, macrophages, neutrophils, mast cells, eosinophils and platelets, and plays an important role in both adaptive and innate immune responses.

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Drug Indication
Treatment of immune thrombocytopenia

C36H40FN9O3

665.7597

665.32

C, 64.95; H, 6.06; F, 2.85; N, 18.93; O, 7.21

1575596-29-0

1575596-77-8;1575596-29-0;1575591-66-0

73388818

White to off-white solid powder

3.4

1

11

8

49

1230

1

CC(C)(/C=C(\C#N)/C(=O)N1CCC[C@H](C1)N2C3=NC=NC(=C3C(=N2)C4=C(C=C(C=C4)OC5=CC=CC=C5)F)N)N6CCN(CC6)C7COC7

LCFFREMLXLZNHE-GBOLQPHISA-N

InChI=1S/C36H40FN9O3/c1-36(2,45-15-13-43(14-16-45)26-21-48-22-26)18-24(19-38)35(47)44-12-6-7-25(20-44)46-34-31(33(39)40-23-41-34)32(42-46)29-11-10-28(17-30(29)37)49-27-8-4-3-5-9-27/h3-5,8-11,17-18,23,25-26H,6-7,12-16,20-22H2,1-2H3,(H2,39,40,41)/b24-18+/t25-/m1/s1

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 (3.12 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 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 (3.12 mM) (saturation unknown) in 10% DMSO + 90% (20% SBE-β-CD in 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 20.8 mg/mL clear DMSO stock solution to 900 μL of 20% SBE-β-CD physiological saline solution and mix evenly.
Preparation of 20% SBE-β-CD in Saline (4°C，1 week): Dissolve 2 g SBE-β-CD in 10 mL saline to obtain a clear solution.

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Solubility in Formulation 3: ≥ 2.08 mg/mL (3.12 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.)