Development and comparison of single FLT3-inhibitors to dual FLT3/TAF1-inhibitors as an anti-leukemic approach
Robert S. Leigh, Bogac L. Kaynak, Heikki Ruskoaho, Mika J. Välimäki
Abstract
Acute myeloid leukemia (AML) is characterized by several recurrent mutations that affect disease biology and phenotype, response to therapy and risk of subsequent relapse. Though tyrosine kinase inhibitors have gained regulatory approval for the treatment of AML, it is unclear whether single drugs targeting a specific genomic alteration will be sufficient to eradicate disease. Fortuitously, kinase/bromodomain inhibitors allow targeting of downstream transcriptional effectors of oncogenic pathways, allowing impediment of drug resistance at the transcriptional level. Successful development of combinatorial therapeutic strategies to inhibit both upstream oncogenic pathways and their downstream effectors could thus impede the onset of resistant disease.
Introduction
Acute myeloid leukemia (AML) is a malignant disorder of hematopoietic stem cells characterized by clonal expansion of abnormally differentiated blasts of the myeloid lineage [1]. AML is generally treated using chemotherapeutic drugs (e.g., combination of cytarabine and an anthracycline). Unfortunately, with standard chemotherapy, long-term survival of patients with AML is achieved in only 35–45% of those younger than 60 years and 10–15% of those aged 60 years and older [2]. New drugs have recently been developed which act on genes necessary for AML cancer cell survival, such as FMS-like tyrosine kinase 3 (FLT3) and isocitrate dehydrogenase (IDH) 1/2 inhibitors [3–5].
Materials and method
Computational chemistry
The commercial modelling package MOE 2022.02 (Chemical Computing Group Inc., Montreal, Canada) was utilized for protein modelling and docking experiments. The docking protocol was carried out with a protein structure of FLT3 (PDB; 4XUF, type II) and TAF1(2) (PDB; 5I29) [35,36]. An Amber12:EHT forcefield was applied for the molecule parametrization and protein structure preparation. On-flight generated ligand conformations were placed in the cavity of the ATP binding site of FLT3 and the second bromodomain of TAF1(2) with the Triangle Matcher method and ranked with the London dG scoring function. Subsequently, the 30 highest ranked poses were applied for a refinement procedure containing the energy minimization and rescoring with the Generalized-Born Volume Integral/Weighted Surface area (GBVI/WSA dG) scoring function.
Results
FLT3/TAF1 bromodomain inhibitors with a novel chemical scaffold
Our aim was to produce a novel series of multitarget compounds for testing in AML assays and to provide insights on the molecular mechanisms of action and synergistic effects in treating multifactorial disease. We assumed that simultaneously targeting both FLT3 kinase and TAF1-bromodomains may potentially have a lower propensity to result in drug-resistant cancers. In our previous screening campaign, we identified a drug-like chemical probe, 3i-1103, as a modulator of atrial and ventricular reporter gene expression [37]. Preliminary structural analyses and ligand binding evaluations suggested a methoxyphenyl moiety of 3i-1103 as a structural feature (warhead) linked to TAF1 bromodomain and FLT3 kinase specificity (Fig 1).
Discussion
By using a combination of high-throughput cell-based screening assays and structure-based design, we have uncovered a novel molecular scaffold against AML. At low concentrations, these 3i-compounds efficiently inhibit the growth of AML cell lines. Interestingly, bromodomain and kinase screening results indicated that 3i-compounds act on a set of clinically relevant AML drug targets: FLT3, TAF1, c-KIT and PDGFRB. By novelly targeting both FLT3 kinase and the TAF1 bromodomain, we anticipated that these novel agents may have a lower propensity to result in drug-resistant cancers. Additionally, using a pluripotent stem cell-based toxicological screening assay, these novel 3i-compounds are less toxic to stem cells than tool compound BAY299 [26].
Acknowledgments
We are grateful to the High Throughput Biomedicine Unit at the Finnish Institute for Molecular Medicine (University of Helsinki) for assistance with in vitro cancer cell efficacy assays.
Citation: Leigh RS, Kaynak BL, Ruskoaho H, Välimäki MJ (2025) Development and comparison of single FLT3-inhibitors to dual FLT3/TAF1-inhibitors as an anti-leukemic approach. PLoS ONE 20(3): e0320443. https://doi.org/10.1371/journal.pone.0320443
Editor: Francesco Bertolini, European Institute of Oncology, Italy
Received: November 21, 2024; Accepted: February 19, 2025; Published: March 28, 2025
Copyright: © 2025 Leigh et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
Data Availability: All relevant data are within the manuscript and its Supporting Information files.
Funding: This work was supported by the Jane and Aatos Erkko Foundation (H.R, grant number -). The funder had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. This work was supported by the Sigrid Juselius Foundation (H.R, grant number -). The funder had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. This work was supported by the Finnish Cultural Foundation (R.S.L, grant number -). The funder had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. This work was supported by the Finnish Foundation for Cardiovascular Research (R.S.L, grant number -). The funder had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. There was no additional external funding received for this study.
Competing interests: The authors have declared that no competing interests exist.
