Optimization of the In Vivo Potency of Pyrazolopyrimidine MALT1 Protease Inhibitors by Reducing Metabolism and Increasing Potency in Whole Blood

Abstract

The paracaspase MALT1 has gained increasing interest as a target for the treatment of subsets of lymphomas as well as autoimmune diseases, and there is a need for suitable compounds to explore the therapeutic potential of this target. Here, we report the optimization of the in vivo potency of pyrazolopyrimidines, a class of highly selective allosteric MALT1 inhibitors. High doses of the initial lead compound led to tumor stasis in an activated B-cell-like (ABC) diffuse large B-cell lymphoma (DLBCL) xenograft model, but this compound suffered from a short in vivo half-life and suboptimal potency in whole blood. Guided by metabolism studies, we identified compounds with reduced metabolic clearance and increased in vivo half-life. In the second optimization step, masking one of the hydrogen-bond donors of the central urea moiety through an intramolecular interaction led to improved potency in whole blood. This was associated with improved in vivo potency in a mechanistic model of B cell activation. The optimized compound led to tumor regression in a CARD11 mutant ABC-DLBCL lymphoma xenograft model.

TRAF6 prevents fatal inflammation by homeostatic suppression of MALT1 protease

“…In line with previous reports, MLT-985 was slightly more potent (two- to threefold) than MLT-943 in inhibiting constitutive MALT1 protease activity in TRAF6 KO Jurkat T cells (Fig. 7A) ( 37 ). Compared with MLT-943, MLT-985 displayed superior pharmacokinetics and improved bioavailability in mice ( 37 ) and therefore was chosen to achieve effective inhibition of chronic MALT1 activity in vivo.…”

“…We asked whether pharmacologic inhibition of constitutive MALT1 protease activation is able to ameliorate uncontrolled immune activation resulting from T cell–specific TRAF6 ablation. We evaluated inhibition of constitutive MALT1 protease activity by MLT-943 and MLT-985, two potent MALT1 inhibitors ( 28 , 37 ). In line with previous reports, MLT-985 was slightly more potent (two- to threefold) than MLT-943 in inhibiting constitutive MALT1 protease activity in TRAF6 KO Jurkat T cells (Fig.…”

A20 and ABIN-1 cooperate in balancing CBM complex-triggered NF-κB signaling in activated T cells

“…For stimulation of primary T cells, cells were transferred to anti-CD3 (0.5 µg/ml) pre-coated 96-well plates and stimulated in the presence of soluble anti-CD28 and anti-mouse IgG1/IgG2a as described above. Inhibitor treatment was done with 25 μM proteasome inhibitor MG132 (1 h pre-treatment; Calbiochem), 1 μM MALT1 inhibitor MLT-985 [ 32 ] (4 h pre-treatment), 75 μM MALT1 inhibitor Z-VRPR-FMK (Enzo Life Sciences) (3 h pre-treatment) and 2.5 μM cIAP1/2 inhibitor birinapant (10 min pre-treatment; BioCat) solved in dimethylsulphoxide (DMSO).…”

Identification of Novel Fused Heteroaromatics-Based MALT1 Inhibitors by High-Throughput Screening to Treat B Cell Lymphoma

“…However, the structures of these two compounds are very similar to that of 4 . Among these two compounds, MLT-985 effectively led to tumor regression in an ABC-DLBCL lymphoma xenograft model. , In 2018, Lorena et al reported a z-VRPR-FMK derivative 5 as a substrate-mimetic peptidic covalent MALT1 inhibitor. This compound showed maximal inhibition of MALT1 activity in in vitro and cell-based assays.…”