Pirtobrutinib Resistance in CLL Can Arise From Secondary BTK Mutations

By Cecilia Brown - February 28, 2023

Resistance to non-covalent Bruton’s tyrosine kinase (BTK) inhibitors in patients with chronic lymphocytic leukemia (CLL) can arise from secondary mutations in BTK.

Aishath Naeem, PhD, of the Broad Institute of the Massachusetts Institue of Technology and Harvard, and colleagues conducted the research because while covalent BTK inhibitors “transformed the therapy” for CLL, “continuous therapy has been complicated by the development of resistance.”

For example, a mutation in BTK that leads to alteration of the protein’s 481 cysteine residue (C481S), which is where BTK inhibitors form covalent bonds with the protein, is the most common resistance mechanism in patients who have disease progression on covalent BTK inhibitors.

However, pirtobrutinib is a highly selective, non-covalent BTK inhibitor that has “substantial clinical activity” regardless of BTK mutation status in patients who experience disease progression on a covalent BTK inhibitor, Dr. Naeem and colleagues wrote.

The study’s investigators used in vitro models of ibrutinib resistance and cells from patients with CLL to study pirtobrutinib, demonstrating the drug “potently inhibits BTK-mediated functions including [B-cell receptor] signaling, cell viability and CCL3/CCL4 chemokine production in both BTK wild-type and C481S mutant CLL cells.”

Primary CLL cells from patients who responded to pirtobrutinib had reduced B-cell receptor signaling, cell survival, and CCL3/CCL4 chemokine secretion. However, the primary CLL cells upon disease progression showed “increasing resistance to pirtobrutinib in signaling inhibition, cell viability and cytokine production,” Dr. Naeem and colleagues wrote.

The researchers used longitudinal whole exome sequencing on samples from two patients with CLL who had disease progression on pirtobrutinib, determining BTK mutations that impact the gatekeeper residue T474—as well as second-site kinase dead mutations in BTK—can cause pirtobrutinib resistance.

“In summary, we show that pirtobrutinib is a potent BTK inhibitor in vitro and in vivo, regardless of mutations at C481, with novel multiclonal second site BTK mutations leading to resistance among patients with pre-existing BTK C481 mutations,” Dr. Naeem and colleagues concluded.


Naeem A, Utro F, Wang Q, et al. Pirtobrutinib targets BTK C481S in ibrutinib-resistant CLL but second-site BTK mutations lead to resistance. Blood Adv. 2022. doi:10.1182/bloodadvances.2022008447

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