DISCOVERY AND PRECLINICAL PROPERTIES OF JNJ-64264681, A POTENT AND SELECTIVE COVALENT BTK INHIBITOR FOR THE TREATMENT OF B CELL MALIGNANCIES
Author(s): ,
Ulrike Philippar
Affiliations:
Janssen Research & Development,Beerse,Belgium
,
Navin Rao
Affiliations:
Janssen Research & Development,Spring House,United States
,
Haopeng Rui
Affiliations:
Janssen Research & Development,Shanghai,China
,
Mark Tichenor
Affiliations:
Janssen Research & Development,San Diego,United States
,
Marcello Gaudiano
Affiliations:
Janssen Research & Development,Beerse,Belgium
,
Mariette Bekkers
Affiliations:
Janssen Research & Development,Beerse,Belgium
,
Ivo Cornelissen
Affiliations:
Janssen Research & Development,Beerse,Belgium
,
Kathryn Packman
Affiliations:
Janssen Research & Development,Spring House,United States
,
Sriram Balasubramanian
Affiliations:
Janssen Research & Development,San Diego,United States
,
John Gerecitano
Affiliations:
Janssen Research & Development,Spring House,United States
,
Jacqueline Bussolari
Affiliations:
Janssen Research & Development,Spring House,United States
Jennifer Venable
Affiliations:
Janssen Research & Development,San Diego,United States
EHA Library. Philippar U. 06/09/21; 325627; EP869
Ulrike Philippar
Ulrike Philippar
Contributions
Abstract
Presentation during EHA2021: All e-poster presentations will be made available as of Friday, June 11, 2021 (09:00 CEST) and will be accessible for on-demand viewing until August 15, 2021 on the Virtual Congress platform.

Abstract: EP869

Type: E-Poster Presentation

Session title: Lymphoma Biology & Translational Research

Background
Bruton’s tyrosine kinase (BTK), a member of the Tec family of tyrosine kinases, plays a critical role in B-cell activation and oncogenic signaling via the B cell receptor. BTK inhibitors have been widely studied in B-cell hematologic malignancies and 3 inhibitors, including the first-in-class BTK inhibitor ibrutinib, are currently approved. While BTK inhibitors have been paradigm-changing in Chronic lymphocytic leukemia, Mantle cell lymphoma, and Waldenstroms macroglobulinemia, combination therapies with other agents may be required to reach optimal response rates and cures in other histologies (eg, Diffuse large B-cell lymphoma [DLBCL]).

Aims
Describe the discovery and preclinical properties of a novel, highly selective, small-molecule covalent BTK inhibitor, JNJ-64264681, designed to enable combination therapies.

Methods
The covalent inhibitor potency was determined by kinact/KI ratios. Inhibition of B-cell activation was assessed in mouse primary cells. The proportion of free and modified BTK was determined in a covalent adduct assay. Dose response of BTK occupancy was assessed in Ramos B cells. A quantitative mechanistic model simulated and used human B cell BTK occupancy to estimate covalent binding. Target engagement and pharmacokinetics/pharmacodynamics studies were conducted in rats and tumor-bearing mice. The effect of JNJ-64264681 on cancer cell proliferation was assessed in B cell lymphoma cell lines, and tumor growth inhibition was studied in cell line and lymphoma patient-derived xenograft mouse models (OCI-Ly10, LY2298).

Results
JNJ-64264681 was developed to obtain high selectivity and ideal physicochemical properties. A series of optimizations resulted in improved microsomal stability, high pH 2 solubility, measurable pH 7 solubility, and double-digit nM potency in rat and human whole blood. Cocrystallization of JNJ-64264681 with BTK confirmed covalent bond formation with Cys481, and in line with this, the adduct was resistant to denaturation. JNJ-64264681 was highly selective against a diverse panel of receptors, enzymes, ion channels, and human kinases. BTK occupancy was dose proportional in vivo, and full occupancy was maintained even after plasma drug levels decreased. JNJ-64264681 showed potent antiproliferative activity (IC50 <100 nM) in CD79b-mutant activated B cell (ABC)-DLBCL cell lines, but not in cell lines which are independent of NF-κB signaling for survival, highlighting its selectivity. Treatment with JNJ-64264681 led to potent in vivo BTK occupancy, inhibition of specific cytokines and tumor growth inhibition in a CD79b-mutant ABC-DLBCL xenograft model. Furthermore, JNJ-64264681 showed significant tumor growth inhibition in the ABC-DLBCL-like LY2298 patient-derived xenograft model. Similarly, treatment with the combination of JNJ-64264681 and the MALT1 inhibitor JNJ-67856633 resulted in increased tumor growth inhibition compared with single agents and tumor regression in both CD79b mutant mouse lymphoma models.

Conclusion
The preclinical results confirm that JNJ-64264681 is a highly selective and potent BTK inhibitor and provide proof-of-concept for human trials in patients with B cell malignancies driven by the classical NF-κB pathway. JNJ-64264681 is currently in human clinical trials (NCT04210219 and NCT04657224) as single agent to establish dose and pharmacokinetics and to evaluate efficacy in combination with the first-in-class MALT1 inhibitor JNJ-67856633.

Keyword(s): B cell lymphoma, Malignant lymphoma

Presentation during EHA2021: All e-poster presentations will be made available as of Friday, June 11, 2021 (09:00 CEST) and will be accessible for on-demand viewing until August 15, 2021 on the Virtual Congress platform.

Abstract: EP869

Type: E-Poster Presentation

Session title: Lymphoma Biology & Translational Research

Background
Bruton’s tyrosine kinase (BTK), a member of the Tec family of tyrosine kinases, plays a critical role in B-cell activation and oncogenic signaling via the B cell receptor. BTK inhibitors have been widely studied in B-cell hematologic malignancies and 3 inhibitors, including the first-in-class BTK inhibitor ibrutinib, are currently approved. While BTK inhibitors have been paradigm-changing in Chronic lymphocytic leukemia, Mantle cell lymphoma, and Waldenstroms macroglobulinemia, combination therapies with other agents may be required to reach optimal response rates and cures in other histologies (eg, Diffuse large B-cell lymphoma [DLBCL]).

Aims
Describe the discovery and preclinical properties of a novel, highly selective, small-molecule covalent BTK inhibitor, JNJ-64264681, designed to enable combination therapies.

Methods
The covalent inhibitor potency was determined by kinact/KI ratios. Inhibition of B-cell activation was assessed in mouse primary cells. The proportion of free and modified BTK was determined in a covalent adduct assay. Dose response of BTK occupancy was assessed in Ramos B cells. A quantitative mechanistic model simulated and used human B cell BTK occupancy to estimate covalent binding. Target engagement and pharmacokinetics/pharmacodynamics studies were conducted in rats and tumor-bearing mice. The effect of JNJ-64264681 on cancer cell proliferation was assessed in B cell lymphoma cell lines, and tumor growth inhibition was studied in cell line and lymphoma patient-derived xenograft mouse models (OCI-Ly10, LY2298).

Results
JNJ-64264681 was developed to obtain high selectivity and ideal physicochemical properties. A series of optimizations resulted in improved microsomal stability, high pH 2 solubility, measurable pH 7 solubility, and double-digit nM potency in rat and human whole blood. Cocrystallization of JNJ-64264681 with BTK confirmed covalent bond formation with Cys481, and in line with this, the adduct was resistant to denaturation. JNJ-64264681 was highly selective against a diverse panel of receptors, enzymes, ion channels, and human kinases. BTK occupancy was dose proportional in vivo, and full occupancy was maintained even after plasma drug levels decreased. JNJ-64264681 showed potent antiproliferative activity (IC50 <100 nM) in CD79b-mutant activated B cell (ABC)-DLBCL cell lines, but not in cell lines which are independent of NF-κB signaling for survival, highlighting its selectivity. Treatment with JNJ-64264681 led to potent in vivo BTK occupancy, inhibition of specific cytokines and tumor growth inhibition in a CD79b-mutant ABC-DLBCL xenograft model. Furthermore, JNJ-64264681 showed significant tumor growth inhibition in the ABC-DLBCL-like LY2298 patient-derived xenograft model. Similarly, treatment with the combination of JNJ-64264681 and the MALT1 inhibitor JNJ-67856633 resulted in increased tumor growth inhibition compared with single agents and tumor regression in both CD79b mutant mouse lymphoma models.

Conclusion
The preclinical results confirm that JNJ-64264681 is a highly selective and potent BTK inhibitor and provide proof-of-concept for human trials in patients with B cell malignancies driven by the classical NF-κB pathway. JNJ-64264681 is currently in human clinical trials (NCT04210219 and NCT04657224) as single agent to establish dose and pharmacokinetics and to evaluate efficacy in combination with the first-in-class MALT1 inhibitor JNJ-67856633.

Keyword(s): B cell lymphoma, Malignant lymphoma

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