An Extended Pyrrolobenzodiazepine–Polyamide Conjugate with Selectivity for a DNA Sequence Containing the ICB2 Transcription Factor Binding Site

Abstract

The binding of nuclear factor Y (NF-Y) to inverted CCAAT boxes (ICBs) within the promoter region of DNA topoisomerase IIα results in control of cell differentiation and cell cycle progression. Thus, NF-Y inhibitory small molecules could be employed to inhibit the replication of cancer cells. A library of pyrrolobenzodiazepine (PBD) C8-conjugates consisting of one PBD unit attached to tri-heterocyclic polyamide fragments was designed and synthesized. The DNA-binding affinity and sequence selectivity of each compound were evaluated in DNA thermal denaturation and DNase I footprinting assays, and the ability to inhibit binding of NF-Y to ICB1 and ICB2 was studied using an electrophoretic mobility shift assay (EMSA). 3a was found to be a potent inhibitor of NF-Y binding, exhibiting a 10-fold selectivity for an ICB2 site compared to an ICB1-containing sequence, and showing low nanomolar cytotoxicity toward human tumor cell lines. Molecular modeling and computational studies have provided details of the covalent attachment process that leads to formation of the PBD-DNA adduct, and have allowed the preference of 3a for ICB2 to be rationalized.

Synthesis and Biological Evaluation of a Novel C8-Pyrrolobenzodiazepine (PBD) Adenosine Conjugate. A Study on the Role of the PBD Ring in the Biological Activity of PBD-Conjugates

“…The C8-position of the A ring of PBD is the most preferred point of attachment of substituents within the compound's framework. Several chemical scaffolds, including heterocyclic polyamides [13][14][15], biaryl-units [16], benzofused rings [17] and quinazolinone [18] rings have been linked to the C8 position of PBD monomer producing compounds with improved DNA-sequence selectivity [13,16], and antimicrobial [19,20], anticancer [16] and antitubercular [21][22][23] activities compared to the PBD unit alone. The shape and physicochemical properties of C8-substituents have a direct effect on the cytotoxicity of the PBD-conjugates and their ability, or inability, to interact with the DNA-minor groove and inhibit transcription factors activity [13].…”

The use of molecular dynamics simulations to evaluate the DNA sequence-selectivity of G–A cross-linking PBD–duocarmycin dimers

“…Based on the results of a thermallyinduced cleavage assay, they reported that 3 forms cross-links at the 5'-AATTTTCC(G)-3' sequence. During the past six years we have developed molecular dynamics simulation methodologies [11,13,48] that allow us to accurately predict and interpret how both mono-alkylating and DNA cross-linking agents of this type interact in the minor groove of DNA. As the hybrid dimer 3 reported by Lee [47] had a significantly lower DNA cross-linking potency and in vitro cytotoxicity compared to the Hurley (1) and Denny (2) dimers, we decided to use our simulation methodologies to compare the interaction of the three molecules at their proposed DNA recognition sites.…”

“…Through the years, many novel cross-linking agents have been designed, synthesized and evaluated [4] , [5] in an attempt to (a) improve on cross-linking efficiency [6], (b) modify the sequence-selectivity of cross-linking [7], (c) change the distribution of inter-versus intrastrand cross-links [8][9][10], and/or (d) target different base pair combinations [11][12][13]. For example, the naturally occurring pyrrolobenzodiazepine (PBD) molecules, derived from Streptomyces species [14][15][16][17][18], are characterized by an N10-C11 imine group that enables formation of a covalent bond to the C2-amino group of a guanine base [19], and these have been successfully incorporated into PBD dimer structures such as DSB-120 [20] and SJG-136 [21] (Figure 1A) that can form sequence-selective G-G cross-links in the DNA minor groove.…”

Pyrrolobenzodiazepines (PBDs) Do Not Bind to DNA G-Quadruplexes

“…, isohelicity) to fit perfectly into the DNA minor groove. PBD/DNA adduct formation has been shown to inhibit a number of biological processes, including the binding of transcription factors to DNA [9], [10], [11], [12] and the function of enzymes such as endonucleases [13], [14] and RNA polymerase [15]. Many PBD molecules also have significant antimicrobial activity [16], [17], [18], [19], [20], [21].…”