Inhibiting androgen receptor nuclear entry in castration-resistant prostate cancer

Clinical resistance to the second-generation antiandrogen enzalutamide in castration resistant prostate cancer (CRPC), despite persistent androgen receptor (AR) activity in tumors, highlights the unmet medical need for next generation antagonists. We have identified and characterized tetra-aryl cyclobutanes (CBs) as a new class of competitive AR antagonists that exhibit a unique mechanism of action. These CBs are structurally distinct from current antiandrogens (hydroxyflutamide, bicalutamide, and enzalutamide), and inhibit AR-mediated gene expression, cell proliferation, and tumor growth in several models of CRPC. Conformational profiling revealed that CBs stabilize an AR conformation resembling an unliganded receptor. Using a variety of techniques, it was determined that the AR:CB complex was not recruited to AR-regulated promoters and, like apo AR, remains sequestered in the cytoplasm bound to heat shock proteins. Thus, we have identified third generation AR antagonists whose unique mechanism of action suggests that they may have therapeutic potential in CRPC

Testing a global standard for quantifying species recovery and assessing conservation impact.

Recognizing the imperative to evaluate species recovery and conservation impact, in 2012 the International Union for Conservation of Nature (IUCN) called for development of a "Green List of Species" (now the IUCN Green Status of Species). A draft Green Status framework for assessing species' progress toward recovery, published in 2018, proposed 2 separate but interlinked components: a standardized method (i.e., measurement against benchmarks of species' viability, functionality, and preimpact distribution) to determine current species recovery status (herein species recovery score) and application of that method to estimate past and potential future impacts of conservation based on 4 metrics (conservation legacy, conservation dependence, conservation gain, and recovery potential). We tested the framework with 181 species representing diverse taxa, life histories, biomes, and IUCN Red List categories (extinction risk). Based on the observed distribution of species' recovery scores, we propose the following species recovery categories: fully recovered, slightly depleted, moderately depleted, largely depleted, critically depleted, extinct in the wild, and indeterminate. Fifty-nine percent of tested species were considered largely or critically depleted. Although there was a negative relationship between extinction risk and species recovery score, variation was considerable. Some species in lower risk categories were assessed as farther from recovery than those at higher risk. This emphasizes that species recovery is conceptually different from extinction risk and reinforces the utility of the IUCN Green Status of Species to more fully understand species conservation status. Although extinction risk did not predict conservation legacy, conservation dependence, or conservation gain, it was positively correlated with recovery potential. Only 1.7% of tested species were categorized as zero across all 4 of these conservation impact metrics, indicating that conservation has, or will, play a role in improving or maintaining species status for the vast majority of these species. Based on our results, we devised an updated assessment framework that introduces the option of using a dynamic baseline to assess future impacts of conservation over the short term to avoid misleading results which were generated in a small number of cases, and redefines short term as 10 years to better align with conservation planning. These changes are reflected in the IUCN Green Status of Species Standard

Long-Term Stable and Tightly Controlled Expression of Recombinant Proteins in Antibiotics-Free Conditions

Plasmid-based gene expression is a fundamental tool in the field of biotechnology. However, overexpression of genes of interest with multi-copy plasmids often causes detrimental effects on host cells. To overcome this problem, chromosomal integration of target genes has been used for decades; however, insufficient protein expression occurred with this method. In this study, we developed a novel cloning and expression system named the chromosomal vector (ChroV) system, that has features of stable and high expression of target genes on the F' plasmid in the Escherichia coli JM109(DE3) strain. We used an RMT cluster (KCTC 11994BP) containing a silent cat gene from a previous study to clone a gene into the F' plasmid. The ChroV system was applied to clone two model targets, GFPuv and carotenoids gene clusters (4 kb), and successfully used to prove the inducible tightly regulated protein expression in the F' plasmid. In addition, we verified that the expression of heterologous genes in ChroV system maintained stably in the absence of antibiotics for 1 week, indicating ChroV system is applicable to antibiotics-free production of valuable proteins. This protocol can be widely applied to recombinant protein expression for antibiotics-free, stable, and genome-based expression, providing a new platform for recombinant protein synthesis in E. coli. Overall, our approach can be widely used for the economical and industrial production of proteins in E. coli

Discovery of ODM-201, a new-generation androgen receptor inhibitor targeting resistance mechanisms to androgen signaling-directed prostate cancer therapies

Activation of androgen receptor (AR) is crucial for prostate cancer growth. Remarkably, also castration-resistant prostate cancer (CRPC) is dependent on functional AR, and several mechanisms have been proposed to explain the addiction. Known causes of CRPC include gene amplification and overexpression as well as point mutations of AR. We report here the pharmacological profile of ODM-201, a novel AR inhibitor that showed significant antitumor activity and a favorable safety profile in phase 1/2 studies in men with CRPC. ODM-201 is a full and high-affinity AR antagonist that, similar to second-generation antiandrogens enzalutamide and ARN-509, inhibits testosterone-induced nuclear translocation of AR. Importantly, ODM-201 also blocks the activity of the tested mutant ARs arising in response to antiandrogen therapies, including the F876L mutation that confers resistance to enzalutamide and ARN-509. In addition, ODM-201 reduces the growth of AR-overexpressing VCaP prostate cancer cells both in vitro and in a castration-resistant VCaP xenograft model. In contrast to other antiandrogens, ODM-201 shows negligible brain penetrance and does not increase serum testosterone levels in mice. In conclusion, ODM-201 is a potent AR inhibitor that overcomes resistance to AR-targeted therapies by antagonizing both overexpressed and mutated ARs. ODM-201 is currently in a phase 3 trial in CRPC

ERF mutations reveal a balance of ETS factors controlling prostate oncogenesis

Half of all prostate cancers are caused by the TMPRSS2-ERG gene-fusion, which enables androgens to drive expression of the normally silent E26 transformation-specific (ETS) transcription factor ERG in prostate cells. Recent genomic landscape studies of such cancers have reported recurrent point mutations and focal deletions of another ETS member, the ETS2 repressor factor ERF. Here we show these ERF mutations cause decreased protein stability and mostly occur in tumours without ERG upregulation. ERF loss recapitulates the morphological and phenotypic features of ERG gain in normal mouse prostate cells, including expansion of the androgen receptor transcriptional repertoire, and ERF has tumour suppressor activity in the same genetic background of Pten loss that yields oncogenic activity by ERG. In the more common scenario of ERG upregulation, chromatin immunoprecipitation followed by sequencing indicates that ERG inhibits the ability of ERF to bind DNA at consensus ETS sites both in normal and in cancerous prostate cells. Consistent with a competition model, ERF overexpression blocks ERG-dependent tumour growth, and ERF loss rescues TMPRSS2-ERG-positive prostate cancer cells from ERG dependency. Collectively, these data provide evidence that the oncogenicity of ERG is mediated, in part, by competition with ERF and they raise the larger question of whether other gain-of-function oncogenic transcription factors might also inactivate endogenous tumour suppressors