Edo-S101, a Bendamustine (BDM)/ Histone-Deacetylase Inhibitor (HDACi) Fusion Molecule, Demonstrates Potent Preclinical Activity Against T-Cell Malignancies and Overcomes BDM-Resistance

Introduction: T-cell and natural killer (NK)-cell lymphomas encompass a heterogeneous spectrum of malignancies with variable degrees of biological and clinical aggressiveness and different patterns of nodal/extra nodal dissemination. Despite advances in understanding of their genetic landscape, treatment options for these tumors are limited and survival remains dismal especially in patients (pts.) who relapse/progress after frontline treatment. Therefore, development of new active agents is a pressing medical need in this setting. The unconventional alkylator BDM and HDACi are clinically effective in pts. with recurring disease and the combination of these agents was suggested as a way to overcome early resistance and improve response and survival rates in T-cell tumors. Since EDO-S101 is a fusion molecule that combines the active structures of BDM and the pan-HDACi vorinostat to simultaneously damage DNA and block damage repair, we investigated its antitumor activity in preclinical models of human T- and NK- lymphomas.

Methods: We assessed the patterns of EDO-S101 cytotoxicity and its regulatory effects on genes involved in DNA-damage/repair, apoptosis and drug resistance on a large panel of cell lines representative of most T-lymphoma subtypes, including precursor T-cell lymphoma/leukemia (pTCL; MOLT-4, KE-37, Jurkat, CCRF-CEM), anaplastic large T-cell lymphoma (ALCL; SR-786, SUDHL1) and cutaneous T-cell lymphoma (CTCL; HUT78, MYLA, HH, PB2B, MAC-2A). As a further model, we exploited an ALCL cell line (SUDHL1-R100) selected for resistance to BDM. SUDHL1-R100 display a growth pattern indistinguishable from parental cells in the presence of BDM (100 μmol/L).

Results: EDO-S101 induced a significant time- and dose-dependent inhibition of growth and survival in all T-cell lines, with a IC50 at 48 hrs (0.33 to 0.68 μmol/L), that was at least 90% and 50% lower than IC50 of BDM (10.5 to 23.67 μmol/L) and vorinostat (0.42 to 1.17 μmol/L) in the same cell lines. No significant differences emerged across various cell lines as to sensitivity to EDO-S101. Notably, growth inhibitory effects of EDO-S101 were 35% to 40% higher than those achieved by the concomitant exposure of tumor cells to BDM and vorinostat at equimolar concentrations. A significant induction of the DNA double strand breaks marker pγH2AX, along with decreased levels of Thr-1989 pATR and Polo-like kinase 1 (PLK1) were consistently observed in pTCL, ALCL and CTCL cell lines, indicating that EDO-S101 triggered inappropriate mitotic entry with unrepaired DNA damage. In addition, qRT-PCR results showed upregulation of the proapototic genes NOXA, p21 and/or p27 in pTCL, CTCL and ALCL cells. Among DNA repair genes, EDO-S101-induced downregulation of ATM, ATR and EXO1 was paralleled by an increase of MGMT transcripts. Most intriguingly, BDM-resistant ALCL cells (SUDHL1-R100) were highly sensitive to EDO-S101, with an IC50 of only 0.72 μmol/L, but less responsive to vorinostat (IC50: 2.17 μmol/L) than parental cells (IC50: 1.46 μmol/L). Exposure to EDO-S101 also resulted in transcriptional regulation of components of the PI3K pathway, mainly the PI3Kα and δ isoforms and mTOR. These findings, prompted experiments showing that EDO-S101 at sublethal concentrations was highly synergic with the predominant PI3Kα/δ inhibitor copanlisib in inducing cell death of tumor T- and NK-cells. Finally, exposure to EDO-S101 (1.0 μmol/L) for 48 hrs resulted in a sustained upregulation of surface PD1 and PD-L1 in most tumor T- and NK-cell lines, as a possible result of its HDACi component.

Conclusions: We have shown for the first time that the BDM/vorinostat fusion molecule EDO-S101 is very active in preclinical models of T/NK cell lymphomas. It shows a superior activity towards different types of tumor T- and NK-cells than its single components administered either individually or in combination by also overcoming acquired resistance to BDM. Data indicate that EDO-S101 simultaneously damages DNA and interferes on its repair pathways in tumor T-cells. We also showed synergy between EDO-S101 and PI3Kα/δ inhibitors possibly due to regulatory effects on specific components of the PI3K pathway as well as a potential for this agent to enhance PD1-blockade strategies. A clinical trial is ongoing to evaluate efficacy and safety of EDO-S101 in pts. with relapsed/refractory hemopoietic malignancies including T-cell lymphomas.