Important Compound Classes
Title
Proteolysis Targeting Chimera Compounds and Methods of Preparing and using same.
Patent Publication Number
US 2020/0325130 A1
Publication Date
October 15, 2020
Priority Application
US 16/848,290
Priority Date
April 14, 2020
Inventors
Crews, C.; Toure, M.; Ko, E.; Jaime-Figueroa, S.
Assignee Company
Yale University, New Haven, CT, United State.
Disease Area
Cancer
Biological Target
Fusion Protein BCR-ABL
Summary
The fusion of the BCR and ABL1 genes on the Philadelphia chromosome (Ph), arising from a reciprocal translocation between chromosomes 9 and 22 in hematological progenitor cells, is found in more than 90% of patients with chronic myelogenous leukemia (CML) and some with adult acute lymphoblastic leukemia (ALL). The oncogenic BCR-ABL protein exhibits constitutively activated ABL tyrosine kinase activity that drives downstream signaling pathways, resulting in uncontrolled proliferation of CML, dysregulated differentiation, growth, and survival of leukemic cells. Therapeutic intervention by inhibiting the kinase activity of BCR-ABL1 involves targeting its ATP-binding site with drugs such as imatinib (Gleevec), nilotinib (Tasigna), and dasatinib (Sprycel). These compounds have been found to greatly reduce leukemic tumor and increase the overall survival rate of CML patients; however, the majority of chronic phase CML patients undergoing continuous treatment have a normal life expectancy. Furthermore, some patients suffer from either poor initial response, loss of response, or tolerability issues, which can arise when clones harboring mutations in the BCR-ABL1 kinase (BCR-ABL1mut) that hinder the tyrosine kinase inhibitor (TKI) from binding and result in further drive of the disease. This raises the possibility that knockdown of BCR-ABL protein might have potential therapeutic benefit for CML treatment.
Current approved TKIs for the treatment of CML target the catalytic ATP binding site of ABL1, either in a type-I or type-II binding mode, which bind to either the catalytically active or catalytically inactive conformation of the SH1 domain. For example, imatinib mesylate, the first TKI developed against BCR-ABL, binds competitively at the ATP-binding site of c-ABL and inhibits both c-ABL and the oncogenic fusion protein BCR-ABL. The need arises for the second generation TKIs such as dasatinib and bosutinib to treat CML patients with acquired resistance to imatinib. A major drawback to the remarkable success of BCR-ABL TKIs is all CML patients must remain on lifelong treatment since the TKIs are not curative due to persistent leukemic stem cells (LSCs). While approved drugs such as nilotinib and imatinib are more selective than ponatinib and the type-I TKIs, bosutinib, and dasatinib, all of these inhibitors have distinct profiles but also inhibit many other kinases and are susceptible to resistance mutations. The T3151 mutation is at the gatekeeper in the ATP-binding pocket of BCR-ABL and, common in advanced phase of CML, tends to disrupt vital contact points between the inhibitors and the enzyme, which then leads to one of the main causes of resistance.
This Patent Highlight illustrates the proteolysis-targeting chimeric (PROTAC) technology, which has attracted attention as a pharmacological strategy for abrogating post-translational protein functions. The discovery of several small molecular E3 ligands, include cereblon (CRBN) ligand pomalidomide, Von Hippel–Lindau (VHL) ligand, mouse double minute 2 homologue (MDM2) ligand nutlin-3a, and inhibitor of apoptosis protein (IAP) bestatin. In general, PROTAC is composed of target protein ligand and E3 ubiquitin ligase ligand connected by flexible linker and the chimeric molecules recruit target protein into spatial proximity with an E3 ubiquitin ligase, which induces the target ubiquitination and subsequent degradation by the proteasome. There are many successful applications to several target proteins, including bromodomain containing 4 (BRD4), estrogen receptor (ER), and androgen receptor (AR). In this Highlight, the TKI is capable of binding to c-ABL and/or BCR-ABL and simultaneously binds a tyrosine kinase and a ubiquitin ligase, and the tyrosine kinase is ubiquitinated by the ubiquitin ligase. The E3 ubiquitin ligase comprises a Von Hippel–Lindau (VHL) E3 ubiquitin ligase or a cereblon (CRBN) E3 ligase. The representation TKI-L-(ULM)k illustrates TKI as tyrosine kinase inhibitor; L a linker; ULM a ubiquitin ligase binder; and k is an integer ranging from 1 to 4 and also the TKI is covalently linked to L and ULM, which in turn is covalently linked to L. Furthermore, the TKI is selected from a group of compounds consisting of imatinib, saracatinib, dasatinib, danusertib, ponatinib, nilotinib, erlotinib, bafetinib, and so forth.
Key Structures
Figure 1.
Biological Assay
For Western blotting using standard electrophoresis, the separated proteins were incubated with the respective antibodies. In vitro kinase binding affinity was determined using the KinomeScan platform and cell viability assay using the CellTiterGlo luminescent cell viability assay.
Biological Data
Table 1 shows selected PROTAC affinities for the ABL kinase domain. DAS-CRBN PROTAC induced BCR-ABL degradation of >60% at 1 μM and >80% at 2.5 μM. In the cell viability assay, DAS-6-2-2-6-CRBN was active against BCR-ABL driven K562 cell line with a half-maximal response concentration (EC50) of 4.4 ± 2.1 nM.
Table 1.
Recent Review Articles
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The author declares no competing financial interest.