Identifying the mechanistic differences between hypomethylating agents for the treatment of peripheral T cell lymphoma

Introduction: Peripheral T-cell lymphomas (PTCL) are a rare and heterogeneous group of malignancies that share unifying feature epigenetic dysregulation. Recent evidence suggests PTCL, unlike other forms cancer or even lymphoma, exhibits marked vulnerability to hypomethylating agents (HMAs) alone especially in combination with HDAC inhibitors. HMAs like azacitidine (AZA) decitabine (DAC) have been shown reverse transcriptional repression secondary hypermethylation. Cladribine (CLAD) is FDA-approved for the treatment hairy cell leukemia postulated inhibit both DNA histone methylation. Here, we sought define mechanistic differences between different order inform rationale on optimal combinations might exploit vulnerabilities PTCL. Methods: Cell viability, caspase activity assay, western blotting (WB) were performed panel lymphoma lines identify effects AZA/DAC/CLAD survival, apoptosis, methyltransferase (DNMT) levels. Liquid chromatography-based mass spectrometry (LC-MS) was done quantify 5-methyl cytosine (5-mC) Results: viability analysis confirmed dose time-dependent sensitivity six PTCL differed following exposure AZA/DAC/CLAD, where CLAD showed lowest IC50 value (0.1–1 µM) across all lines, followed by AZA (IC50: 1–10 µM), DAC had least effect >100 µM, except T-ALL). A differential threshold activation observed most superior leading highest apoptotic potential. However, WB after 24 hours treatment, DNMT1 DNMT3A protein levels significantly depleted at low concentrations (<0.01 compared whereas did not affect LC-MS-based quantification 5-mC similar relationship methylation, initial changes doses >0.01 μM >0.1 AZA/CLAD. Taking short half-life AZA/DAC into account, assays daily addition increased cytotoxicity treated samples, which comparable CLAD. Ongoing analyses gene expression metabolism universal methyl donor S-adenosylmethionine (SAM) will help understand difference these HMAs. Conclusions: These data suggest distinct mechanism action hypomethylation. While induces hypomethylation depletion DNMTs, acts DNMT depletion-independent pathway. understanding how best use them clinic as well assist strategic development biological correlates further support their therapeutic application. The research funded by: Translational Orphan Blood Cancer Research Initiative Fund RO1 # FD-R-006814-01. Keywords: Genomics, Epigenomics, Other -Omics, Molecular Targeted Therapies No conflicts interests pertinent abstract.