Proansamycin B derivatives from the post-PKS modification gene deletion mutant of Amycolatopsis mediterranei S699.

Ten new proansamycin B congeners (1-10) together with one known (11) were isolated and characterized on the basis of 1D and 2D NMR spectroscopic and HRESIMS data from the Amycolatopsis mediterranei S699 ΔPM::rifR+rif-orf19 mutant. Compounds 8 and 9 featured with six-membered ring and five-membered ring hemiketal, respectively. Compounds 1, 2, and 9 displayed antibacterial activity against MRSA (methicillin-resistant Staphylococcus aureus), with the MIC (minimal inhibitory concentration) values of 64, 8, and 128 µg/mL, respectively. Compound 1 showed significant cytotoxicity against MDA-MB-231, HepG2 and Panc-1 cell lines with IC50 (half maximal inhibitory concentration) values of 2.3 ± 0.2, 2.5 ± 0.3 and 3.8 ± 0.5 µM, respectively.

Deciphering the Timing of Naphthalenic Ring Formation in the Biosynthesis of 8-Deoxyrifamycins.

Although the biosynthesis of rifamycin has been studied for three decades, the biosynthetic formation of the naphthalenic ring remains unclear. In this study, by deletion of all post-PKS modification genes, we identified macrolactam precursors released from rif PKS. Isolated prorifamycins (M3 and M4) have a benzenic chromophore and exist in two sets of macrocyclic atropisomers. The transformation from prorifamycins to benzenoid (5) and naphthalenoid (6) was suggested to be a non-enzymatic process, which is an off-PKS assembly.

Hybrid Histone Deacetylase Inhibitor: An Effective Strategy for Cancer Therapy.

Inhibition of histone deacetylases (HDACs) has proven to be an effective strategy for cancer therapy. To date, five histone deacetylase inhibitors (HDACis) have been approved for cancer treatment, and numerous others are undergoing clinical trials. An agent that can simultaneously and effectively inhibit two or more targets may offer greater therapeutic benefits over single-acting agents in preventing resistance to treatment and potentiating synergistic effects. A prime example of a bifunctional agent is the hybrid HDACi. Representative classes of reported hybrid HDACis are reviewed here to shed light on the design of novel hybrid HDACis for cancer therapy.

Discovery of 1,2-diphenylethene derivatives as human DNA topoisomerase II catalytic inhibitors and antitumor agents.

Human DNA topoisomerase II (TopoII) is highly correlated with cell proliferation, and involved in tumor biogenesis and development. The classic chemotherapeutic agents etoposide (VP-16) and adriamycin (ADR) targeting TopoII are wildly used in clinical applications. Herein, fifty-eight pinosylvin (1,2-diphenylethene) derivatives as TopoII inhibitors were designed and synthesized through three generations of structural optimizations on the basis of the structure of the initial hit A1 from in-house chemical library. The most potent compound F2 showed high in vitro inhibitory efficacy against TopoII (IC50 α 3.8, ß 10.1 µM), compared to that of VP-16 (IC50 α 110.0, ß 36.1 µM) for pBR322 DNA relaxation with no evident TopoII poisons in DNA cleavage assay. Meanwhile, F2 exhibited strong antitumor activities against human cancer cell lines HeLa, HCT-116, PC-3, MDA-MB-231, HepG2 and A549 (IC50 0.1-0.3 µM), compared to that of VP-16 (IC50 1.5-15.1 µM). F2 showed less cytotoxicity against normal murine cell line CCL-226 (IC50 > 50 µM) than that of VP-16 (IC50 20.8 µM). The selectivity index of F2 and VP-16 are larger than 52.1 and 1.3-26.2 in cell lines, respectively. Additionally, F2 exhibited potent potency in apoptosis induction and cell cycle arrest in HepG2 cells. These results provide a promising strategy and good starting point for the development of potent TopoII inhibitors as antitumor agents.

D319 induced antifungal effects through ROS-mediated apoptosis and inhibited isocitrate lyase in Candida albicans.

BACKGROUND: Candida albicans (C. albicans) is an opportunistic pathogen that can cause superficial and life-threatening systemic infections in immunocompromised patients. However, the available clinically antifungals are limited. Therefore, the development of effective antifungal agents and therapies is urgently needed. Quinoline type of compounds were reported to possess potent anti-fungal effect. A series of quinoline derivatives were synthesized. Moreover their inhibitory activities and mechanisms on C. albicans were evaluated in this study. METHODS: The structure of D319 was identified by extensive spectroscopic analysis. The antifungal activity of D319 on C. albicans was evaluated using conventional methods, including the inhibition zone diameters with filter paper, Clinical Laboratory Standard Institute (CLSI) broth microdilution method in vitro, and in a murine model in vivo. Flow cytometry, fluorescence microscopy, western blot, knockout mutant and revertant strain techniques, and molecular modeling were applied to explore the mechanism of action of D319 in anti-Candida. RESULTS: D319 exhibited potent anti-Candida activity with Minimum Inhibitory Concentration value of 2.5 µg/mL in vitro. D319 significantly improved survival rate and reduced fungal burden compared to vehicle control in a murine model in vivo. The treatment of C. albicans with D319 resulted in fungal apoptosis through reactive oxygen species (ROS) accumulation in C. albicans. Furthermore, D319 inhibited the glyoxylate enzyme isocitrate lyase (ICL) of C. albicans, which was also confirmed by docking analysis. CONCLUSIONS: Quinoline compound D319 exhibited strong anti-Candida activities in vitro and in vivo models through inhibiting ICL activity and ROS accumulation in C. albicans. GENERAL SIGNIFICANCE: This study showed that compound D319 as a novel isocitrate lyase inhibitor, would be a promising anti-Candida lead compound, which provided a potential application of this type of compounds in fighting clinical fungal infections. Furthermore, this study also supported ICL as a potential target for anti-Candida drug discovery.

Cost-effectiveness of fuzuloparib compared to routine surveillance, niraparib and olaparib for maintenance treatment of patients with germline BRCA1/2 mutation and platinum-sensitive recurrent ovarian carcinoma in China.

Background: Maintenance therapy with the poly (ADP-ribose) polymerase inhibitors (PARPis) for platinum-sensitive recurrent ovarian carcinoma (OC) have proven to be effective compared with placebo. We aimed to evaluate the cost-effectiveness (CE) of maintenance fuzuloparib compared to routine surveillance (RS), niraparib and olaparib for platinum-sensitive recurrent OC from the Chinese healthcare systems. Method: A partitioned survival model with three-state (progression-free, progressed, death) was constructed utilizing TreeAge Pro 2011 software to evaluate the economic value of fuzuloparib, niraparib and olaparib maintenance treatment for platinum-sensitive recurrent OC based on the clinical data derived from FZOCUS-2, ENGOT-OV16/NOVA and ENGOT-Ov21/SOLO2. Transition probabilities were estimated from the reported survival probabilities in those trials. Cost and health preference data were derived from the literature. The quality-adjusted life-years (QALYs) and lifetime costs were measured for this analysis. A 5 years horizon and 5%/year discount rates were used. One-way analysis, and probabilistic sensitivity analysis (PSA) were performed to explore the model uncertainties. Results: Total cost of fuzuloparib, niraparib and olaparib were $31628.10, $48183.48 and $54605.54, whereas they had an incremental cost-utility ratio of $31992.69, $32216.08 and $23359.26 per additional progression-free survival (PFS) QALYs gained compared with RS, relatively. Model showed that maintenance fuzuloparib achieved at least an 85.5% probability of CE at the threshold of $37654.50/QALY. One-way sensitivity analysis revealed that the results were sensitive to the PFS and the price of medicines. Conclusion: Fuzuloparib was less cost-effective for patients with germline BRCA1/2 mutation and platinum-sensitive recurrent OC compared to olaparib, but was superior to niraparib from the Chinese healthcare systems perspective.

HSAF-induced antifungal effects in Candida albicans through ROS-mediated apoptosis.

Heat-stable antifungal factor (HSAF) belongs to polycyclic tetramate macrolactams (PTMs), which inhibits many fungal pathogens and is effective in inhibiting Candida albicans (C. albicans). In this study, we found that HSAF induced the apoptosis of Candida albicans SC5314 through inducing the production of reactive oxygen species (ROS). Nevertheless, we validated the efficacy of HSAF against candidiasis caused by C. albicans in a murine model in vivo, and HSAF significantly improved survival and reduced fungal burden compared to vehicles. A molecular dynamics (MD) simulation was also investigated, revealing the theoretical binding mode of HSAF to the ß-tubulin of C. albicans. This study first found PTMs-induced fungal apoptosis through ROS accumulation in C. albicans and its potential as a novel agent for fungicides.

Alteramide B is a microtubule antagonist of inhibiting Candida albicans.

BACKGROUND: Alteramide B (ATB), isolated from Lysobacter enzymogenes C3, was a new polycyclic tetramate macrolactam (PTM). ATB exhibited potent inhibitory activity against several yeasts, particularly Candida albicans SC5314, but its antifungal mechanism is unknown. METHODS: The structure of ATB was established by extensive spectroscopic analyses, including high-resolution mass spectrometry, 1D- and 2D-NMR, and CD spectra. Flow cytometry, fluorescence microscope, transmission electron microscope, molecular modeling, overexpression and site-directed mutation studies were employed to delineate the anti-Candida molecular mechanism of ATB. RESULTS: ATB induced apoptosis in C. albicans through inducing reactive oxygen species (ROS) production by disrupting microtubules. Molecular dynamics studies revealed the binding patterns of ATB to the ß-tubulin subunit. Overexpression of the wild type and site-directed mutants of the ß-tubulin gene (TUBB) changed the sensitivity of C. albicans to ATB, confirming the binding of ATB to ß-tubulin, and indicating that the binding sites are L215, L217, L273, L274 and R282. In vivo, ATB significantly improved the survival of the candidiasis mice and reduced fungal burden. CONCLUSION: The molecular mechanism underlying the ATB-induced apoptosis in C. albicans is through inhibiting tubulin polymerization that leads to cell cycle arrest at the G2/M phase. The identification of ATB and the study of its activity provide novel mechanistic insights into the mode of action of PTMs against the human pathogen. GENERAL SIGNIFICANCE: This study shows that ATB is a new microtubule inhibitor and a promising anti-Candida lead compound. The results also support ß-tubulin as a potential target for anti-Candida drug discovery.

Iterative assembly of two separate polyketide chains by the same single-module bacterial polyketide synthase in the biosynthesis of HSAF.

Antifungal HSAF (heat-stable antifungal factor, dihydromaltophilin) is a polycyclic tetramate macrolactam from the biocontrol agent Lysobacter enzymogenes. Its biosynthetic gene cluster contains only a single-module polyketide synthase-nonribosomal peptide synthetase (PKS-NRPS), although two separate hexaketide chains are required to assemble the skeleton. To address the unusual biosynthetic mechanism, we expressed the biosynthetic genes in two "clean" strains of Streptomyces and showed the production of HSAF analogues and a polyene tetramate intermediate. We then expressed the PKS module in Escherichia coli and purified the enzyme. Upon incubation of the enzyme with acyl-coenzyme A and reduced nicotinamide adenine dinucleotide phosphate (NADPH), a polyene was detected in the tryptic acyl carrier protein (ACP). Finally, we incubated the polyene-PKS with the NRPS module in the presence of ornithine and adenosine triphosphate (ATP), and we detected the same polyene tetramate as that in Streptomyces transformed with the PKS-NRPS alone. Together, our results provide evidence for an unusual iterative biosynthetic mechanism for bacterial polyketide-peptide natural products.

Juanlimycins A and B, ansamycin macrodilactams from Streptomyces sp.

Ansamycins are a family of macrolactams characterized by an aromatic chromophore with an aliphatic chain (ansa chain) connected back to a nonadjacent position through an amide bond. This family has shown a high degree of druggability exemplified by rifamycins, maytansinoids, and geldanamycins. In this study, the isolation of two novel ansamycin macrodilactams with unprecedented features, juanlimycins A (1) and B (2), from Streptomyces sp. LC6 were reported. The structures of 1 and 2 were assigned on the basis of analysis of NMR spectroscopic data and X-ray single crystal diffraction.

Identification and characterization of the cuevaene A biosynthetic gene cluster in streptomyces sp. LZ35.

Genome sequence analysis of Streptomyces sp. LZ35 has revealed a large number of secondary metabolite pathways, including one encoded in an orphan type I polyketide synthase gene cluster that contains a putative chorismatase/3-hydroxybenzoate synthase gene. Mutagenesis and comparative metabolic profiling led to the identification of cuevaene A as the metabolic product of the gene cluster, thus making it the first 3-HBA containing polyketide biosynthetic gene cluster described to date. Cuv10 was proven to be responsible for the conversion of chorismate into 3-HBA; Cuv18 is speculated to be responsible for the 6-hydroxylation of 3-HBA during polyketide chain elongation. Additionally, several pathway-specific regulatory factors that affect the production of cuevaene A were identified. Our results indicate that targeted inactivation of a gene followed by comparative metabolic profiling is a useful approach to identify and characterize cryptic biosynthetic gene clusters.