Resistance Mutations to BTK Inhibitors Originate From the NF-κB but Not From the PI3K-RAS-MAPK Arm of the B Cell Receptor Signaling Pathway

Abstract

Since the first clinical report in 2013, inhibitors of the intracellular kinase BTK (BTKi) have profoundly altered the treatment paradigm of B cell malignancies, replacing chemotherapy with targeted agents in patients with chronic lymphocytic leukemia (CLL), mantle cell lymphoma (MCL), and Waldenström’s macroglobulinemia. There are over 20 BTKi, both irreversible and reversible, in clinical development. While loss-of-function (LoF) mutations in the BTK gene cause the immunodeficiency X-linked agammaglobulinemia, neither inherited, nor somatic BTK driver mutations are known. Instead, BTKi-sensitive malignancies are addicted to BTK. BTK is activated by upstream surface receptors, especially the B cell receptor (BCR) but also by chemokine receptors, and adhesion molecules regulating B cell homing. Consequently, BTKi therapy abrogates BCR-driven proliferation and the tissue homing capacity of the malignant cells, which are being redistributed into peripheral blood. BTKi resistance can develop over time, especially in MCL and high-risk CLL patients. Frequently, resistance mutations affect the BTKi binding-site, cysteine 481, thereby reducing drug binding. Less common are gain-of-function (GoF) mutations in downstream signaling components, including phospholipase Cγ2 (PLCγ2). In a subset of patients, mechanisms outside of the BCR pathway, related e.g. to resistance to apoptosis were described. BCR signaling depends on many proteins including SYK, BTK, PI3K; still based on the resistance pattern, BTKi therapy only selects GoF alterations in the NF-κB arm, whereas an inhibitor of the p110δ subunit of PI3K instead selects resistance mutations in the RAS-MAP kinase pathway. BTK and PLCγ2 resistance mutations highlight BTK’s non-redundant role in BCR-mediated NF-κB activation. Of note, mutations affecting BTK tend to generate clone sizes larger than alterations in PLCγ2. This infers that BTK signaling may go beyond the PLCγ2-regulated NF-κB and NFAT arms. Collectively, when comparing the primary and acquired mutation spectrum in BTKi-sensitive malignancies with the phenotype of the corresponding germline alterations, we find that certain observations do not readily fit with the existing models of BCR signaling.

The TKI Era in Chronic Leukemias

“…Another mechanism of resistance to irreversible BTKi, although less common and BTK mutation, is the acquisition of gain of function muta-tions in PLC γ2 protein (R665W and L845F), leading to autonomous BCR activity [108]. Subsequently, other mutations have been discovered and reported in Figure 3 [109].…”

“…Another mechanism of resistance to irreversible BTKi, although less common and BTK mutation, is the acquisition of gain of function mutations in PLC γ2 protein (R665W and L845F), leading to autonomous BCR activity [108]. Subsequently, other mutations have been discovered and reported in Figure 3 [109]. The reversible BTKi (pirtobrutinib, vecabrutinib, ARQ531, fenebrutinib), potentially overcomes C481 mutations by very strong non-covalent interactions (hydrogen, ionic bonds, and hydrophobic interactions) with BTK that block ATP binding and do not require C481 [82].…”

Bruton Tyrosine Kinase Inhibitors in B-Cell Malignancies: Their Use and Differential Features

“…Relapse or disease progression during treatment with BTK inhibitors in patients with B-cell malignancies is commonly associated with acquired resistance. The frequency with which acquired resistance develops varies between different B-cell malignancy subtypes but appears to be higher among patients with MCL and high-risk CLL/SLL [ 29 ]. The most commonly observed mutations conferring resistance to first- and second-generation BTK inhibitors are mutations at the Cys-481 residue in the BTK active site [ 30 ].…”

“…Other mutations leading to acquired resistance to BTK inhibitors include gain-of-function mutations resulting in the increased activity of downstream kinases (e.g. phospholipase C gamma 2) despite inhibition of BTK [ 29 ], with several other mechanisms of acquired resistance also observed (albeit less commonly) [ 32 ].…”

Structure-Function Relationships of Covalent and Non-Covalent BTK Inhibitors

“…A drawback of irreversible inhibitors is that drug resistance in malignant diseases can develop when BTK variations at the catalytic site and the gatekeeper are not able to bind efficiently to irreversible inhibitors in treated patients ( 40 , 41 ). This is a rather common event in patients treated with irreversible inhibitors and who have a relapse.…”