Abstract 4555: C8-linked pyrrolobenzodiazepine (PBD)-benzofused hybrids as transcription factor inhibitors
Authors: Corcoran David B. , Jackson Paul J. M. , Ajaz Ambereen , Lewis Thomas , Pepper Chris , Thurston David E. , Rahman Khondaker M. 
Published: August 2015

Abstract
The pyrrolo[2,1-c][1,4]benzodiazepines (PBDs) have been extensively investigated as DNA-interactive agents since their discovery in 1963. The significant cytotoxicity of the naturally derived PBDs has led to the search for synthetic analogues that retain the cytotoxicity of the parent structure while providing an enhanced therapeutic index. There is growing evidence that PBD monomers inhibit the binding of transcription factors to gene promoter regions, blocking their effects on gene expression. For example, GWL-78, a C8-linked PBD-Py-Py (Py = pyrrole) conjugate, has been shown to block interaction of the transcription factor NF-Y, and KMR-28-39, a C8-linked PBD-MPB conjugate inhibits the activity of the transcription factor NF-κB. As up-regulation of NF-κB has long been implicated in the development and progression of several cancer types, this may account for the femtomolar potency observed for PBD monomers such as KMR-28-39.
The initial synthesis of a small library of C8-PBD conjugates with benzofused C8-substituents provided notable activity against a panel of leukaemia cell lines. In addition to exhibiting low picomolar IC50 values in a number of cell lines, these compounds also showed selectivity for binding to NF-κB consensus sequences. This small library was used as the starting point for the synthesis of three analogous libraries of PBD C8-conjugates designed to explore the SARs of such structures, with a view to selecting a candidate for pre-clinical development. The design rationale for the novel conjugates involved modification of the shape of the C8-linked substituent with the introduction of specific functional groups in an effort to induce sequence-selective DNA interactivity.
The sequence selectivity of the novel benzofused PBD conjugates was investigated using a FRET-based assay in which ligand binding to several different transcription factor binding sequences implicated in cancer was explored (i.e., AP-1, NF-κB1, NF-κB2, STAT3 and EGR). Differential DNA stabilisation compared to control was observed for all analogues, with ΔTm values between 23.4 and 3.4oC at 1 μM ligand concentration (Ligand:DNA = 1:5) for these sequences, indicating the molecules had preferential binding to certain transcription factor binding sequences. Stabilisation was especially pronounced for the AP-1 consensus sequence with two lead compounds (DC-3-28-33 and DC-1-113) providing a high degree of stabilisation (23.9oC and 26oC, respectively). Preliminary cytotoxicity screening studies were carried out in Primary CLL and a Myeloma cell line (JJN3), and low nanomolar IC50 values were observed in the CLL cells for several of the synthesised compounds. These included DC-1-92 (9.7nM), DC-1-115 (14nM), DC-1-36 (16nM), DC-1-170 (17nM), and DC-1-113 (18nM). Control screening in normal lymphocytes is currently underway in order to establish the selective toxicity profiles of these compounds.
Citation Format: David B. Corcoran, Paul J. M. Jackson, Ambereen Ajaz, Thomas Lewis, Chris Pepper, David E. Thurston, Khondaker M. Rahman. C8-linked pyrrolobenzodiazepine (PBD)-benzofused hybrids as transcription factor inhibitors. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 4555. doi:10.1158/1538-7445.AM2015-4555