Designed ligands that inhibit hypoxia-inducible gene expression could offer new tools for genomic research and, potentially, drug discovery efforts for the treatment of neovascularization in cancers. We report a stabilized alpha-helix designed to target the binding interface between the C-terminal transactivation domain (C-TAD) of hypoxia-inducible factor 1alpha (HIF-1alpha) and cysteine-histidine rich region (CH1) of transcriptional coactivator CBP/p300. The synthetic helix disrupts the structure and function of this complex, resulting in a rapid downregulation of two hypoxia-inducible genes (VEGF and GLUT1) in cell culture.

The comprehensive understanding of cellular signaling pathways remains a challenge due to multiple layers of regulation that may become evident only when the pathway is probed at different levels or critical nodes are eliminated. To discover regulatory mechanisms in canonical WNT signaling, we conducted a systematic forward genetic analysis through reporter-based screens in haploid human cells. Comparison of screens for negative, attenuating and positive regulators of WNT signaling, mediators of R-spondin-dependent signaling and suppressors of constitutive signaling induced by loss of the tumor suppressor adenomatous polyposis coli or casein kinase 1α uncovered new regulatory features at most levels of the pathway. These include a requirement for the transcription factor AP-4, a role for the DAX domain of AXIN2 in controlling β-catenin transcriptional activity, a contribution of glycophosphatidylinositol anchor biosynthesis and glypicans to R-spondin-potentiated WNT signaling, and two different mechanisms that regulate signaling when distinct components of the β-catenin destruction complex are lost. The conceptual and methodological framework we describe should enable the comprehensive understanding of other signaling systems.DOI: http://dx.doi.org/10.7554/eLife.21459.001

Selective blockade of gene expression by designed small molecules is a fundamental challenge at the interface of chemistry, biology, and medicine. Transcription factors have been among the most elusive targets in genetics and drug discovery, but the fields of chemical biology and genetics have evolved to a point where this task can be addressed. Herein we report the design, synthesis, and in vivo efficacy evaluation of a protein domain mimetic targeting the interaction of the p300/CBP coactivator with the transcription factor hypoxia-inducible factor-1α. Our results indicate that disrupting this interaction results in a rapid down-regulation of hypoxia-inducible genes critical for cancer progression. The observed effects were compound-specific and dose-dependent. Gene expression profiling with oligonucleotide microarrays revealed effective inhibition of hypoxia-inducible genes with relatively minimal perturbation of nontargeted signaling pathways. We observed remarkable efficacy of the compound HBS 1 in suppressing tumor growth in the fully established murine xenograft models of renal cell carcinoma of the clear cell type. Our results suggest that rationally designed synthetic mimics of protein subdomains that target the transcription factor-coactivator interfaces represent a unique approach for in vivo modulation of oncogenic signaling and arresting tumor growth.helix mimetics | synthetic inhibitors of transcription | rational design | hydrogen bond surrogate helices

Age-related macular degeneration (AMD) is associated with dysfunction and death of retinal pigment epithelial (RPE) cells. Cell-based approaches using RPE-like cells derived from human pluripotent stem cells (hPSCs) are being developed for AMD treatment. However, most efficient RPE differentiation protocols rely on complex, stepwise treatments and addition of growth factors, whereas small-moleculeonly approaches developed to date display reduced yields. To identify new compounds that promote RPE differentiation, we developed and performed a high-throughput quantitative PCR screen complemented by a novel orthogonal human induced pluripotent stem cell (hiPSC)-based RPE reporter assay. Chetomin, an inhibitor of hypoxiainducible factors, was found to strongly increase RPE differentiation; combination with nicotinamide resulted in conversion of over onehalf of the differentiating cells into RPE. Single passage of the whole culture yielded a highly pure hPSC-RPE cell population that displayed many of the morphological, molecular, and functional characteristics of native RPE.retinal pigment epithelium | pluripotent stem cells | high-throughput screening | differentiation | age-related macular degeneration A ge-related macular degeneration (AMD) is the leading cause of irreversible vision loss and blindness among the elderly in industrialized countries. Dysfunction of the retinal pigment epithelium (RPE) is an early event associated with AMD. The RPE, a monolayer of pigmented cells directly abutting the photoreceptor cell layer, plays many important roles in vision and in maintaining the health and integrity of the retina (1). As the RPE deteriorates, there is progressive degeneration of photoreceptor cells.Successful antiangiogenesis treatments have been developed for the neovascular, or "wet," form of AMD. However, there are no Food and Drug Administration-approved treatment options available for the majority of AMD patients, who suffer from the more common nonneovascular, or "dry," form of the disease. In the past few years, however, transplantation of human pluripotent stem cellderived RPE (hPSC-RPE) has emerged as a promising new therapy for dry AMD. A Phase I clinical trial of human embryonic stem (hES)-derived RPE cells recently reported some preliminary encouraging results (2). Additionally, a Phase I trial that will use RPE cells generated from human induced pluripotent stem cells (hiPSCs) reprogrammed from the patients' own skin cells recently injected their first patient (3).If the promise of hiPSC-based approaches for AMD is to be translated into the clinic, each patient would require individualized generation of RPE cells from his or her stem cells, thereby necessitating the development of simple, efficient, safe, and affordable protocols for RPE generation. Although highly efficient protocols have been established, they rely upon mixtures of growth factors (4-6) with the use of complex biologics derived from animal cells or bacteria, presenting potential clinical challenges. As an alternative approach, use...