Biological activity of quinazoline analogues and molecular modeling of their interactions with G-quadruplexes
Graphical abstract
Introduction
Numerous plant and marine medicines with quinazoline components have been known to Asian folk practitioners for time immemorial. Presently, more than 220 natural alkaloids possess a quinazoline or quinazolinone fragment, and the interest in this class of compounds is steadily growing due to their broad and diverse biological activity and medicinal applicability [[1], [2], [3]]. Quinazoline heterocycles are structurally closely similar to a number of biologically active compounds, extending to the basic carriers of biological hereditary information, namely purines and pyrimidines as nucleic acid (NA) bases [1,2]. Quinazolin-4-ones are among the conveniently accessible heterocycles, frequently using the long known Niementowski synthesis [[1], [2], [3], [4]]. The latter heterocyclic molecules provide sufficient variability in the search for biologically potent compounds of prospective medicinal interest [[1], [2], [3], [4], [5]]. We have used aldol type conversions of 2-alkylsubstituted quinazolin-4-ones [6] looking for structural and topological similarities with nucleic acid base pairs [[5], [6], [7], [8], [9]], so that enhanced activity of potential novel derivatives might be expected beforehand.
There are a couple of important details of structural requirements to potentially biologically active molecules related to NA base pairs. First, it is the capability to form intra- and inter- molecular hydrogen bonds like the in-plane interactions of NA base pairs. The second requirement is the capability to participate in dispersion interactions of the type of stacking interactions between NA base pairs within the NA helix. With heterocyclic 2-substituted quinazolin-4-one analogues, both requirements are easy to satisfy, as is obvious with planar heterocyclic compounds 1–6 shown in the Scheme 1 below, possessing at least one proton donor – acceptor pair of atoms including nitrogen.
The naturally occurring schizocommunin 3 has been extensively studied aiming first at the clarification of its structure as a quinazoline derivative [7]. More recently, 3 and its derivatives have been found to react specifically with DNA in telomeres and thus to affect a deepest mechanism of cell proliferation and apoptosis by stabilization of guanine quadruplexes, G4, and thus inhibiting telomerase [8]. Pyridine and quinoline substituted quinazolin-4-ones 5 and 6 are known [5], and have also been reported as physiologically active, interacting with another guanine-quadruplex related enzyme, topoisomerase [[9], [10], [11]]. In this study, we design and synthesize the novel molecules 1 and 2, as well as 4, which is an aza-derivative of 3. Then we use the opportunity to investigate the biological activities of these four as well as of the two relatively well-known compounds, 5, and 6, in relation to their primary structural characteristics. Our results indicate that all listed compounds induce DNA damage of different extent depending on their structure on human breast cancer cells. In addition, the studied quinazoline analogues bring changes in the overall cellular morphology of the tested cells. We further use computational modeling of shown quinazolin-4-one derivatives 1–6, and attempt to find connections of theoretical quantities to the biological activity of the compounds. As far as mechanisms of the observed biological effects of 3, 5, and 6 have already been discussed, [[7], [8], [9], [10], [11], [12]] this might open possibilities to develop potential drugs based on heterocyclic compounds designed preliminarily. On the other hand, correspondence of model expectations for chosen heterocycles and experimentally registered effects may in turn contribute to understanding of biochemical mechanisms, and possibly direct further design of promising heterocyclic structures.
Our chosen selection logic for molecules, capable of hydrogen bonding and NA stacking, limits the range of eligible structures to aromatic or quasi-aromatic heterocycles while evidently including the growing set of natural aromatic alkaloids, known for many useful biological effects. [12]
Section snippets
2-Substituted analogues of quinazolin-4-one induce genotoxic stress on MDA cells
Human breast cancer cells of the MDA cell line have been used as a model to study the bioactivity of the tested compounds. The cells have been treated with compounds 1 to 6, Scheme 1, for 4 h at 37 °C. To test the genotoxic potential of the six tested substances we have performed the method of Comet assay, also called single-cell gel electrophoresis (SCGE). SCGE sensitively and precisely detects all kinds of damages in DNA including single-strand DNA breaks, double-strand DNA breaks and
Mechanism of action
So far, we have considered effects of structural variations of quinazoline analogues on their biological activity. To understand the reasons for the observed differences, we need to consider the interactions of studied molecules with their presumed biochemical counterparts. This is another more complicated level of molecular organization, which must include both studied heterocyclic molecules, and biological components. For this purpose, we attempt molecular dynamics simulation, MD, for the
Conclusions and outlook
Anticancer activity exhibited by natural alkaloids and synthetic heterocycles, including quinazoline analogues, may in a number of cases be due to their contribution to the stabilization of four-stranded G-quadruplexes, observed in guanine-rich NA sequences. This work uses a model of G-quadruplex stabilization by heterocyclic molecules stacking to its bottom. On the basis of this model, and without preliminary expectations of possible activity and/or biochemical mechanism, we find correlations
Chemistry of 2-substituted quinazolin-4-one analogues
An efficient two step aldol type synthesis of 2-substituted-quinazolinones involves double lithiation of 2-methyl-4(3H)-quinazolin-4-one and subsequent in situ trapping with variety of electrophiles.3 The same approach is applied for the preparation of 2-((3-oxoisoindolin-1-ylidene)methyl)quinazolin-4(3H)-one 1 (Scheme 3). Thus, lithiation of 2-methyl-quinazolin-4-one with 3 equivalents of LDA in THF at −78 °C followed by addition of phthalimide furnished the key intermediate 1a in 66% yield.
Author statement
Jose Kaneti: The G-quadruplex model; Computational modeling; Writing and editing of the computational part.
Milena Georgieva: Conceptualization, Methodology and Data analysis and Curation for genotoxicity results of the studied materials, Writing and Editing of the biological part.
Miroslav Rangelov: MD simulations.
Irena Philipova: Synthesis and characterization of the studied compounds.
Ivan Angelov: Fluorescence experiments.
Bela Vasileva: Methodology and Experimental design of genotoxicity
Declaration of Competing Interest
None declared.
Acknowledgements
This work has been supported by the Bulgarian Science Fund, via Grant Number DN 19/11 of Dec. 10, 2017.The funding organization, www.fni.bg, has not been otherwise involved in the study design, data collection and report writing. Most reported computations have been carried out on the AVITOHOL HPC cluster facility, https://www.top500.org/system/178609; last accessed Sep. 01, 2020.
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