A novel complexity-to-diversity strategy for the diversity-oriented synthesis of structurally diverse and complex macrocycles from quinine.

Recent years have witnessed a global decline in the productivity and advancement of the pharmaceutical industry. A major contributing factor to this is the downturn in drug discovery successes. This can be attributed to the lack of structural (particularly scaffold) diversity and structural complexity exhibited by current small molecule screening collections. Macrocycles have been shown to exhibit a diverse range of biological properties, with over 100 natural product-derived examples currently marketed as FDA-approved drugs. Despite this, synthetic macrocycles are widely considered to be a poorly explored structural class within drug discovery, which can be attributed to their synthetic intractability. Herein we describe a novel complexity-to-diversity strategy for the diversity-oriented synthesis of novel, structurally complex and diverse macrocyclic scaffolds from natural product starting materials. This approach exploits the inherent structural (including functional) and stereochemical complexity of natural products in order to rapidly generate diversity and complexity. Readily-accessible natural product-derived intermediates serve as structural templates which can be divergently functionalized with different building blocks to generate a diverse range of acyclic precursors. Subsequent macrocyclisation then furnishes compounds that are each based around a distinct molecular scaffold. Thus, high levels of library scaffold diversity can be rapidly achieved. In this proof-of-concept study, the natural product quinine was used as the foundation for library synthesis, and six novel structurally diverse, highly complex and functionalized macrocycles were generated.The research leading to these results has received funding from the European Research Council under the European Union’s Seventh Framework Programme (FP7/2007- 2013)/ERC grant agreement no [279337/DOS]. In addition, the group research was supported by grants from the Engineering and Physical Sciences Research Council, Biotechnology and Biological Sciences Research Council, Medical Research Council, Wellcome Trust and AstraZeneca

Concise synthesis of rare pyrido[1,2-a]pyrimidin-2-ones and related nitrogen-rich bicyclic scaffolds with a ring-junction nitrogen.

Pyrido[1,2-a]pyrimidin-2-ones represent a pharmaceutically interesting class of heterocycles. The structurally related pyrido[1,2-a]pyrimidin-4-ones are associated with a broad range of useful biological properties. Furthermore, quinolizinone-type scaffolds of these sorts with a bridgehead nitrogen are expected to display interesting physico-chemical properties. However, pyrido[1,2-a]pyrimidin-2-ones are largely under-represented in current small molecule screening libraries and the physical and biological properties of the pyrido[1,2-a]pyrimidin-2-one scaffold have been poorly explored (indeed, the same can be said for unsaturated bicyclic compounds with a bridgehead nitrogen in general). Herein, we report the development of a new strategy for the concise synthesis of substituted pyrido[1,2-a]pyrimidin-2-ones from readily available starting materials. The synthetic route involved the acylation of the lithium amide bases of 2-aminopyridines with alkynoate esters to form alkynamides, which were then cyclised under thermal conditions. The use of lithium amide anions ensured excellent regioselectivity for the 2-oxo-isomer over the undesired 4-oxo-isomer, which offers a distinct advantage over some existing methods for the synthesis of pyrido[1,2-a]pyrimidin-2-ones. Notably, different aminoazines could also be employed in this approach, which enabled access to several very unusual bicyclic systems with higher nitrogen contents. This methodology thus represents an important contribution towards the synthesis of pyrido[1,2-a]pyrimidin-2-ones and other rare azabicycles with a ring-junction nitrogen. These heterocycles represent attractive structural templates for drug discovery.The research leading to these results has received funding from the European Research Council under the European Union's Seventh Framework Programme (FP7/2007-2013) / ERC grant agreement n° [279337/DOS]. The authors also thank AstraZeneca, the European Union (EU), the Engineering and Physical Sciences Research Council (EPSRC), the Biotechnology and Biological Sciences Research Council (BBSRC), the Medical Research Council (MRC), and the Wellcome Trust for funding. Data accessibility: all data supporting this study are provided as Supplementary Information accompanying this paper.This is the final version of the article. It was first available from Royal Society of Chemistry via http://dx.doi.org/10.1039/c5ob01784

An expedient strategy for the diversity-oriented synthesis of macrocyclic compounds with natural product-like characteristics

Naturally-derived macrocyclic compounds are associated with a diverse range of biological activities, including antibacterial effects, and there are over 100 marketed macrocycle drugs derived from natural products. However, synthetic macrocycles are widely considered to be poorly explored in antibiotic development (indeed, within drug discovery in general). This has been attributed to challenges associated with the generation of such compounds. Whilst there are synthetic methods that can produce large collections of structurally similar macrocycles (i.e., compounds with varying appendages based around similar core macrocyclic ring architectures) there is a relative dearth of strategies for the efficient generation of more structurally diverse macrocycle collections in which there is greater variation in the nature of macrocyclic scaffolds present. Such macrocycle collections should contain compounds with a broad range of biological activities (including antibacterial activities) and the requisite robust synthetic methodology useful for analogue synthesis and lead optimization once an active compound has been identified in a biological screen. Herein, we describe a new and expedient diversity-oriented synthesis (DOS) strategy for the generation of a library of novel structurally diverse macrocyclic compounds with a high level of scaffold diversity. The strategy is concise, proceeds from readily-available starting materials, is modular in nature and features a variety of macrocyclisation techniques. In this proof-of-concept study, the synthesis of several previously unreported macrocyclic compounds was achieved. Each of these macrocycles was based around a distinct molecular scaffold and contained natural product-like structural features (e.g., three-dimensionality and multiple hydrogen bond donors and acceptors) as well as synthetic handles for potential further elaboration. The successful generation of these macrocycles demonstrates the feasibility of the new DOS strategy as a synthetic platform for library generation.The research leading to these results has received funding from the European Research Council under the European Union’s Seventh Framework Programme (FP7/2007-2013)/ERC grant agreement no [279337/DOS]. In addition, the group research was supported by grants from the Engineering and Physical Sciences Research Council, Biotechnology and Biological Sciences Research Council, Medical Research Council and Welcome Trust

ETS1 Mediates MEK1/2-Dependent Overexpression of Cancerous Inhibitor of Protein Phosphatase 2A (CIP2A) in Human Cancer Cells

EGFR-MEK-ERK signaling pathway has an established role in promoting malignant growth and disease progression in human cancers. Therefore identification of transcriptional targets mediating the oncogenic effects of the EGFR-MEK-ERK pathway would be highly relevant. Cancerous inhibitor of protein phosphatase 2A (CIP2A) is a recently characterized human oncoprotein. CIP2A promotes malignant cell growth and is over expressed at high frequency (40–80%) in most of the human cancer types. However, the mechanisms inducing its expression in cancer still remain largely unexplored. Here we present systematic analysis of contribution of potential gene regulatory mechanisms for high CIP2A expression in cancer. Our data shows that evolutionary conserved CpG islands at the proximal CIP2A promoter are not methylated both in normal and cancer cells. Furthermore, sequencing of the active CIP2A promoter region from altogether seven normal and malignant cell types did not reveal any sequence alterations that would increase CIP2A expression specifically in cancer cells. However, treatment of cancer cells with various signaling pathway inhibitors revealed that CIP2A mRNA expression was sensitive to inhibition of EGFR activity as well as inhibition or activation of MEK-ERK pathway. Moreover, MEK1/2-specific siRNAs decreased CIP2A protein expression. Series of CIP2A promoter-luciferase constructs were created to identify proximal −27 to −107 promoter region responsible for MEK-dependent stimulation of CIP2A expression. Additional mutagenesis and chromatin immunoprecipitation experiments revealed ETS1 as the transcription factor mediating stimulation of CIP2A expression through EGFR-MEK pathway. Thus, ETS1 is probably mediating high CIP2A expression in human cancers with increased EGFR-MEK1/2-ERK pathway activity. These results also suggest that in addition to its established role in invasion and angiogenesis, ETS1 may support malignant cellular growth via regulation of CIP2A expression and protein phosphatase 2A inhibition

Vandetanib (ZD6474), an inhibitor of VEGFR and EGFR signalling, as a novel molecular-targeted therapy against cholangiocarcinoma

Cholangiocarcinoma is an intractable cancer, with no effective therapy other than surgical resection. Elevated vascular endothelial growth factor (VEGF) and epidermal growth factor receptor (EGFR) expressions are associated with the progression of cholangiocarcinoma. We therefore examined whether inhibition of VEGFR and EGFR could be a potential therapeutic target for cholangiocarcinoma. Vandetanib (ZD6474, ZACTIMA), a VEGFR-2/EGFR inhibitor, was evaluated. Four human cholangiocarcinoma cell lines were molecularly characterised and investigated for their response to vandetanib. In vitro, two cell lines (OZ and HuCCT1), both of which harboured KRAS mutation, were refractory to vandetanib, one cell line (TGBC24TKB) was somewhat resistant, and another cell line (TKKK) was sensitive. The most sensitive cell line (TKKK) had EGFR amplification. Vandetanib significantly inhibited the growth of TKKK xenografts at doses ⩾12.5 mg kg−1 day−1 (P<0.05), but higher doses (50 mg kg−1 day−1, P<0.05) of vandetanib were required to inhibit the growth of OZ xenografts. Vandetanib (25 mg kg−1 day−1) also significantly (P=0.006) prolonged the time to metastasis in an intravenous model of TKKK metastasis. Inhibiting both VEGFR and EGFR signalling appears a promising therapeutic approach for cholangiocarcinoma. The absence of KRAS mutation and the presence of EGFR amplification may be potential predictive molecular marker of sensitivity to EGFR-targeted therapy in cholangiocarcinoma

Inhibition of epidermal growth factor receptor signalling reduces hypercalcaemia induced by human lung squamous-cell carcinoma in athymic mice

The purpose of this study was to evaluate the role of the epidermal growth factor receptor (EGFR) in parathyroid hormone-related protein (PTHrP) expression and humoral hypercalcaemia of malignancy (HHM), using two different human squamous-cell carcinoma (SCC) xenograft models. A randomised controlled study in which nude mice with RWGT2 and HARA xenografts received either placebo or gefitinib 200 mg kg−1 for 3 days after developing HHM. Effectiveness of therapy was evaluated by measuring plasma calcium and PTHrP, urine cyclic AMP/creatinine ratios, and tumour volumes. The study end point was at 78 h. The lung SCC lines, RWGT2 and HARA, expressed high levels of PTHrP mRNA as well as abundant EGFR protein, but very little erbB2 or erbB3. Both lines expressed high transcript levels for the EGFR ligand, amphiregulin (AREG), as well as, substantially lower levels of transforming growth factor-α (TGF-α), and heparin binding-epidermal growth factor (HB-EGF) mRNA. Parathyroid hormone-related protein gene expression in both lines was reduced 40–80% after treatment with 1 μM of EGFR tyrosine kinase inhibitor PD153035 and precipitating antibodies to AREG. Gefitinib treatment of hypercalcaemic mice with RWGT2 and HARA xenografts resulted in a significant reduction of plasma total calcium concentrations by 78 h. Autocrine AREG stimulated the EGFR and increased PTHrP gene expression in the RWGT2 and HARA lung SCC lines. Inhibition of the EGFR pathway in two human SCC models of HHM by an anilinoquinazoline demonstrated that the EGFR tyrosine kinase is a potential target for antihypercalcaemic therapy

A novel complexity-to-diversity strategy for the diversity-oriented synthesis of structurally diverse and complex macrocycles from quinine.

Recent years have witnessed a global decline in the productivity and advancement of the pharmaceutical industry. A major contributing factor to this is the downturn in drug discovery successes. This can be attributed to the lack of structural (particularly scaffold) diversity and structural complexity exhibited by current small molecule screening collections. Macrocycles have been shown to exhibit a diverse range of biological properties, with over 100 natural product-derived examples currently marketed as FDA-approved drugs. Despite this, synthetic macrocycles are widely considered to be a poorly explored structural class within drug discovery, which can be attributed to their synthetic intractability. Herein we describe a novel complexity-to-diversity strategy for the diversity-oriented synthesis of novel, structurally complex and diverse macrocyclic scaffolds from natural product starting materials. This approach exploits the inherent structural (including functional) and stereochemical complexity of natural products in order to rapidly generate diversity and complexity. Readily-accessible natural product-derived intermediates serve as structural templates which can be divergently functionalized with different building blocks to generate a diverse range of acyclic precursors. Subsequent macrocyclisation then furnishes compounds that are each based around a distinct molecular scaffold. Thus, high levels of library scaffold diversity can be rapidly achieved. In this proof-of-concept study, the natural product quinine was used as the foundation for library synthesis, and six novel structurally diverse, highly complex and functionalized macrocycles were generated.The research leading to these results has received funding from the European Research Council under the European Union’s Seventh Framework Programme (FP7/2007- 2013)/ERC grant agreement no [279337/DOS]. In addition, the group research was supported by grants from the Engineering and Physical Sciences Research Council, Biotechnology and Biological Sciences Research Council, Medical Research Council, Wellcome Trust and AstraZeneca

Targeting vascular endothelial growth factor receptor-1 and -3 with cediranib (AZD2171): effects on migration and invasion of gastrointestinal cancer cell lines.

The effect of vascular endothelial growth factor (VEGF) ligands and cediranib on tumor cell proliferation, migration, and invasion was determined. It has recently been suggested that autocrine signaling through the VEGF receptor (VEGFR) pathway may play a role in tumor cell survival, invasion, and migration. The purpose of the present study was to determine the expression of VEGFRs and VEGFR ligands in a panel of gastrointestinal carcinoma cells. Additionally, we evaluated the effects of VEGF autocrine signaling on tumor cell proliferation, migration, and invasion utilizing cediranib (AZD2171), a pan-VEGFR inhibitor. Five colorectal, three pancreatic, and two hepatocellular carcinoma cell lines were screened for VEGFR and VEGF expression by several methods. Expression of VEGFR-1 and VEGFR-3 was cell line-dependent, whereas VEGFR-2 was not detected. Secretion of VEGF-A was detected in the supernatants of all cell lines whereas VEGF-C secretion was detected in the Panc-1, MiaPaca2, and Hep1 cells only. Tumor cells showed increased migratory activity, but not proliferation, when stimulated with VEGFs. The pan-VEGFR inhibitor cediranib (100 nmol/L) inhibited tumor cell migration and invasion, with no effects on proliferation. Cediranib decreased VEGFR-1 and VEGFR-3 phosphorylation as well as activation of downstream effectors. VEGFR-1 and VEGFR-3 expression was detected in all the gastrointestinal carcinoma cells evaluated. Although activation of the VEGF pathway did not affect cell proliferation, our data indicate that this pathway seems to play a role in tumor cell migration and invasion in these cell lines. Therefore, inhibition of VEGFR by cediranib may represent a clinically relevant treatment option for gastrointestinal tumors