A New Family of Jumonji C Domain-Containing KDM Inhibitors Inspired by Natural Product Purpurogallin

Abstract

Aberrant epigenetic modifications are involved in cancer development. Jumonji C domain-containing histone lysine demethylases (KDMs) are found mainly up-regulated in breast, prostate, and colon cancer. Currently, growing interest is focusing on the identification and development of new inhibitors able to block the activity of KDMs and thus reduce tumor progression. KDM4A is known to play a role in several cellular physiological processes, and was recently found overexpressed in a number of pathological states, including cancer. In this work, starting from the structure of purpurogallin 9aa, previously identified as a natural KDM4A inhibitor, we synthesized two main sets of compound derivatives in order to improve their inhibitory activity against KDM4A in vitro and in cells, as well as their antitumor action. Based on the hypothetical biogenesis of the 5-oxo-5H-benzo[7]annulene skeleton of the natural product purpurogallin (Salfeld, 1960; Horner et al., 1961; Dürckheimer and Paulus, 1985; Tanaka et al., 2002; Yanase et al., 2005) the pyrogallol and catechol units were first combined with structural modifications at different positions of the aryl ring using enzyme-mediated oxidative conditions, generating a series of benzotropolone analogs. Two of the synthetic analogs of purpurogallin, 9ac and 9bc, showed an efficient inhibition (50 and 80%) of KDM4A in enzymatic assays and in cells by increasing levels of its specific targets, H3K9me3/2 and H3K36me3. However, these two compounds/derivatives did not induce cell death. We then synthesized a further set of analogs of these two compounds with greater structural diversification. The most potent of these analogs, 9bf, displayed the highest KDM4A inhibitory enzymatic activity in vitro (IC 50 of 10.1 and 24.37 µM) in colon cancer cells, and the strongest antitumor action in several solid and hematological human cancer cell lines with no toxic effect in normal cells. Our findings suggest that further development of this compound and its derivatives may lead to the identification of new therapeutic antitumor agents acting through inhibition of KDM4A.

Histone Methylation Related Therapeutic Challenge in Cardiovascular Diseases

“…LSD1 is the first identified histone demethylase ( 50 ). HDMs in JHDMs include Fe 2+ - and α-ketoglutarate-dependent hydroxylases, and seven phylogenetically distinct subfamilies were identified in this family ( 51 ).…”

Continuous‐Flow Preparation of Benzotropolones: Combined Batch and Flow Synthesis of Epigenetic Modulators of the (JmjC)‐Containing Domain

“…[6] Representative examples of secondary metabolites belonging to this group are purpurogallin 1, [7,8] theaflavin 2, [9,10] fomentariol 3, [11] aurantricholone 4, [12] goupiolone A 5 [13] and crocipodin 6 [14] (Figure 1). Furthermore, we have showed that properly functionalized benzotropolone 7 act as very potent inhibitors of lysine demethylase JMJD2 A, [15] a subtype of the Jumonji C (JmjC)containing domain family of enzymes, [16][17][18] responsible of the demethylation of N-methylated histone lysine side chains. This epigenetic modification has shown dramatic effects in cell proliferation causing, among other activities, endocrine deregulation and cancer development.…”

“…[31][32][33] Following our interest in the development of continuous flow synthetic methodologies for the synthesis of biologically active molecules for further characterization of their activity, [34,35] we describe our work in the development of a continuous synthetic procedure for the preparation of benzotropolone core ring present in this family of compounds, and further derivatization to render the recently described epigenetic modulator 7. [15] In order to develop an efficient methodology that eventually allows, not only the isolation of larger quantities of compound 7, [15] but also the preparation of other derivatives in a reproducible manner, we started to study the adequacy of the benzotropolone core ring formation reaction under continuous flow. Besides the different methodologies described in literature for the preparation of benzotropolones, [36][37][38][39] peroxidase-mediated condensation reaction of catechol and pyrogallol derivatives stands out as the most frequently used methodology for the formation of benzotropolone ring, [6,14,15,40] so these reaction conditions were chosen as starting point.…”

“…[15] In order to develop an efficient methodology that eventually allows, not only the isolation of larger quantities of compound 7, [15] but also the preparation of other derivatives in a reproducible manner, we started to study the adequacy of the benzotropolone core ring formation reaction under continuous flow. Besides the different methodologies described in literature for the preparation of benzotropolones, [36][37][38][39] peroxidase-mediated condensation reaction of catechol and pyrogallol derivatives stands out as the most frequently used methodology for the formation of benzotropolone ring, [6,14,15,40] so these reaction conditions were chosen as starting point.…”

“…[43] To the best of our knowledge, there is only one precedent in literature, reported by the Ley group, describing the synthesis of targeted molecules using horseradish peroxidase adsorbed onto silica. [44] In view to all that, and considering that the insolubility of our benzotropolones under previously described reaction conditions using soluble horseradish peroxidase [15] could eventually lead to packed-bed reactor clogging, we started our screening by mixing two independent solutions, one containing commercially available 8 a, 9 a, and the enzyme and the other containing the oxidative solution. Under these condition reactor blockage was observed despite the internal diameter of the coiled reactor was increased (1-2 mm i.d.)…”