Abstract C214: Disorazol Z: A highly cytotoxic natural compound with antitumor properties.

Abstract

In search of novel antitumor agents natural macrocyclic polyketides called Disorazols have been isolated from myxobacterium Sorangium cellulosum[1,2,3]. Here we show that Disorazol Z specifically produced by strain So ce427, possesses outstanding cytotoxicity with single digit to even subnanomolar EC50 values in a highly diverse panel of more than 60 different tumor cell lines. Interestingly, the maximum efficacy by which Disorazol Z inhibited cell growth increased with the incubation time, e.g. in the endometrium carcinoma cell line Hec1A reaching a maximum efficacy of about 25% after 24 hours, but about 75% efficacy after 72 hours at a saturating concentration of 100 nM, being approximately 100–200 fold above the respective EC50 values. Based on this observation, it was speculated that the mechanism of action of Disorazol Z is dependent on progressing through cell cycle. Indeed, this was supported by showing that in the cell line RKOp27Kip inducible expression of the cell cycle inhibitor p27Kip led to a complete loss of Disorazol Z cytotoxicity up to a concentration of 100 nM. In contrast, without cell cycle arrest Disorazol Z demonstrated high cytotoxic activity with an EC50 value of 0.54 nM in the same cell line. Furthermore, cell cycle analysis revealed that Disorazol Z arrested KB/HeLa cells in the G2/M phase of the cell cycle with an IC50 value of 0.8 nM. As expected this Disorazol Z-induced G2/M arrest induced apoptotic events, as demonstrated by Caspase 3/7 activation in HCT-116 cells with an EC50 value of 0.25 nM. In agreement with the above described behaviour Disorazol Z could be identified as tubulin binding agent by inhibiting in vitro tubulin polymerisation with a mean IC50 value of 3.3 μM as well as inducing multipolar spindle formation at low nanomolar concentrations in U-2 OS cells. The high divergence of the in vitro tubulin activity compared to the cellular efficacy may point to additional mechanisms of action for Disorazol Z. This working hypothesis may be supported by the fact that synergistic cytotoxic action of Disorazol Z and the tubulin binding agent Colchicin could be observed with combination indices down to about 0.5. Neither Taxol nor Vinblastine showed such synergism, both tubulin binders behaved nearly additive in combination with Disorazol Z. Currently, experiments are under way to identify the tubulin binding site for Disorazol Z. Additional ongoing studies focus on the evaluation of the mechanisms of action of this novel highly potent agent with antitumor properties. Further, we aim at evaluating the utility of Disorazol Z as cytotoxic component in a drug-targeting approach utilizing GPCR ligands as the targeting moieties for the treatment of GPCR overexpressing cancers. References: 1. Jansen et al. (1994), Liebigs Ann. Chem. 1994, 759–773 2. Irschik et al., (1995), Journal of Antibiotics 48, 31–35 2. Elnakady et al. (2004), Biochemical Pharmacology 67, 927–935 Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2011 Nov 12-16; San Francisco, CA. Philadelphia (PA): AACR; Mol Cancer Ther 2011;10(11 Suppl):Abstract nr C214.

A fluorescence anisotropy assay to discover and characterize ligands targeting the maytansine site of tubulin

“…4a, b ). Likewise, the potent cytotoxin disorazole Z 29 , which belongs to the same family of myxobacterial natural products as the vinblastine-site ligand disorazole C 1 30 , but whose binding to tubulin had not been investigated previously, interacts with the maytansine site with a K d of 576 ± 49 nM (mean ± SEM; n = 3) (Fig. 4c, d ).…”

Disorazoles Block Group A Streptococcal Invasion into Epithelial Cells Via Interference with the Host Factor Ezrin

“…To our knowledge, disorazoles have never been tested in an infection context. Two compounds from the disorazole family (disorazole A [Jansen et al, 1994] and disorazole Z [Aicher et al, 2011]) were chosen for further investigation as they differ slightly in their chemical structure (Figures 1A and 1B). It is worth mentioning that the myxobacterial compound library contains additional secondary metabolites active against tubulin assembly, such as the tubulysins (Khalil et al, 2006) and epothilones (Gerth et al, 1996;Tré dan et al, 2015), none of which showed similar activity to the disorazoles.…”

Synthesis and Biological Evaluation of C(13)/C(13′)‐Bis(desmethyl)disorazole Z**

“…As an additional distinguishing feature, the disorazole Z side chain contains two stereocenters, due to oxidation of one of the methyl groups of the characteristic gem-dimethyl motif in all other disorazoles to the carboxylic acid oxidation state. [15] As other members of the disorazole family, disorazole Z (1) (also referred to in the literature as disorazol Z) has been reported to inhibit tubulin polymerization in vitro [16] and to exhibit nanomolar to sub-nanomolar antiproliferative activity against different cancer cell lines. [16,17] The compound was also shown to block group A Staphylococcus (GAS) invasion into epithelial cells by binding to the host factor ezrin.…”

“…[15] As other members of the disorazole family, disorazole Z (1) (also referred to in the literature as disorazol Z) has been reported to inhibit tubulin polymerization in vitro [16] and to exhibit nanomolar to sub-nanomolar antiproliferative activity against different cancer cell lines. [16,17] The compound was also shown to block group A Staphylococcus (GAS) invasion into epithelial cells by binding to the host factor ezrin. [18] Most recently, the molecular interactions of disorazole Z (1) with tubulin have been characterized by X-ray crystallography at 2.1 Å resolution, [19] which revealed that binding occurs at the maytansine site on β-tubulin.…”