LPA81: Pioneering a Highly Effective PROTAC for Targeting BCR-ABL1 in Chronic Myeloid Leukemia

Chronic Myeloid Leukemia (CML) is a condition arising from the BCR-ABL1 fusion gene, which is an active tyrosine kinase due to a chromosomal translocation. The use of imatinib and other TKIs has improved survival rates for patients in the chronic phase of CML. However, challenges persist, with only a quarter of patients achieving treatment-free remission and a significant number progressing to the blast phase.

A new approach involves the use of PROTACs, which are molecules designed to degrade target proteins through the proteasome pathway. A library of 52 asciminib-based PROTACs was synthesized, with the aim of targeting both native and mutant forms of BCR-ABL1. A Nanoluc protein complementation assay was developed to screen these compounds, identifying LPA81 as the most effective.

LPA81 was found to degrade over 90% of BCR-ABL1 within 12 hours at a concentration of 0.5 μM and showed high potency against single mutants at the same concentration. Notably, LPA81 also degraded compound mutant BCR-ABL1. Immunoblotting confirmed LPA81's degradation activity in a dose- and time-dependent manner, with degradation observed as early as 4 hours and at concentrations as low as 10nM. LPA81's degradation efficiency was high, with a maximal degradation level of 98% within 24 hours.

The degradation process was confirmed to occur via the 26s-proteasome, as it was reversed by the proteasome inhibitor MG132 but not by the lysosomal inhibitor chloroquine. LPA81 demonstrated selectivity and induced cytotoxicity in tested human CML cell lines with IC50 values below 1μM, without toxicity to BaF3 cells up to 10μM.

A NanoBRET competitive binding assay was conducted to assess LPA81's binding affinity to ABL1, revealing a weaker binding affinity compared to asciminib or a reduced cellular uptake.

The findings indicate that LPA81 is a potent PROTAC that can degrade both native and mutant BCR-ABL1, leading to the death of CML cells. Further studies on cellular uptake and efficacy in primary cells are ongoing and will provide additional insights into LPA81's potential as a treatment for CML.