The endothelin peptides have an important role in the cancer-stromal interactions that promote tumour growth. Endothelin-1 (ET-1), clinically the most investigated endothelin, is a vital agent in the growth and progression of several tumours including prostate, ovarian, colorectal, bladder, breast and lung carcinomas. ET-1 exerts its effects through the activation of two distinct receptors, ET A and ET B . Once activated, these receptors transmit signals via numerous intracellular signalling pathways. The effects of ET receptor stimulation in cancer cells or cancer-associated cells include proliferation, resistance to apoptosis, angiogenesis, migration and subsequent invasion. At present, the manipulation of the endothelin axis within the pre-clinical setting is the subject of intense investigation. Recent studies into ET receptor antagonism have produced interesting results highlighting the fact that these receptors may provide novel targets for a new generation of chemotherapeutic agents in a variety of cancers.

Endothelin-1 (ET-1) is produced by and stimulates colorectal cancer cells. Fibroblasts produce tumour stroma required for cancer development. We investigated whether ET-1 stimulated processes involved in tumour stroma production by colonic fibroblasts. Primary human fibroblasts, isolated from normal tissues adjacent to colon cancers, were cultured with or without ET-1 and its antagonists. Cellular proliferation, migration and contraction were measured. Expression of enzymes involved in tumour stroma development and alterations in gene transcription were determined by Western blotting and genome microarrays. ET-1 stimulated proliferation, contraction and migration (p < 0.01 v control) and the expression of matrix degrading enzymes TIMP-1 and MMP-2, but not MMP-3. ET-1 upregulated genes for profibrotic growth factors and receptors, signalling molecules, actin modulators and extracellular matrix components. ET-1 stimulated colonic fibroblast cellular processes in vitro that are involved in developing tumour stroma. Upregulated genes were consistent with these processes. By acting as a strong stimulus for tumour stroma creation, ET-1 is proposed as a target for adjuvant cancer therapy.Colorectal cancer accounts for almost a million cancer cases a year, and the incidence is rising worldwide. The initiation and progression of epithelial cancer types, including colorectal, is dependent on the development of a supportive tumour stroma. This is a glycoprotein-rich extracellular matrix (ECM) containing blood vessels and nonepithelial cells. It provides a scaffold and source of nutrients to allow tumours to grow and invade.

Endothelin 1 (ET-1) is overexpressed in cancer, contributing to disease progression. We previously showed that ET-1 stimulated proliferative, migratory, and contractile tumorigenic effects via the ET A receptor. Here, for the first time, we evaluate zibotentan, a specific ET A receptor antagonist, in the setting of colorectal cancer, in cellular models. Pharmacologic characteristics were further determined in patient tissues. Colorectal cancer lines (n ¼ 4) and fibroblast strains (n ¼ 6), isolated from uninvolved areas of colorectal cancer specimens, were exposed to ET-1 and/or ET A/B receptor antagonists. Proliferation (methylene blue), migration (scratch wounds), and contraction (gel lattices) were assessed. Receptor distribution and binding characteristics (K d , B max ) were determined using autoradiography on tissue sections and homogenates and cytospun cells, supported by immunohistochemistry. Proliferation was inhibited by ET A (zibotentan > BQ123; P < 0.

Panagiotis Vlachostergios highlights the importance of evaluating clinically the targeted therapeutic zibotentan.Despite the wide overexpression of endothelin-1 (ET-1) and endothelin A receptors in the majority of carcinomas (1), establishing clinical efficacy of receptor antagonism has proved complex. Numerous factors predispose to discrepancies between preclinical data and clinical outcomes, such as evaluating antagonists in "isolated" cancer cell line cultures. Recently, interest has grown in evaluating drugs within more complex environments involving signaling pathways associated with stromal components: cancer-associated fibroblasts and neovascularization. The importance of fibroblasts in cancer was highlighted many years ago (2).Our group further defined, at mRNA and protein levels, how exposure to and antagonism of ET axis molecules affected both cancer cells, which define cancer parenchyma, and fibroblasts, which populate and refashion cancer stroma. Specifically, zibotentan treatment opposed antiapoptotic pathways, invasive and neovascularization properties, and reduced chemoresistance (3). This we believe justifies its use as additional first-line therapy and points to multitherapy agents for targeting colorectal cancer.To overcome limitations of two-dimensional (2D) cultures, we recently developed a three-dimensional (3D) tissue-engineered in vitro cancer model with distinct compartments (cancer and stroma) that exhibits central hypoxia (4); we are currently developing this as a platform technology for drug testing. Recent studies demonstrate that MCF7 breast cancer cells in 3D collagen I matrices have an altered (reduced) phosphoinositide 3-kinase (PI3K) pathway response when exposed to chemotherapeutics, compared with 2D models (5). Hence, moving to 3D models may present us with more relevant laboratory models for drug evaluations.Clinical trials often start at advanced stages of disease or following previous therapies. This may lead to altered drug-induced molecular signaling away from classic tumorigenic pathways (studied laboratory settings), drug resistance via MDR upregulation, switching high-dimensional cancer attractors, more aggressive dedifferentiated tumors, and resilient types due to Darwinian selection or Lamarckian induction.In terms of molecular mechanisms, we demonstrated in colorectal cancer cells that ET-1 activates numerous tumorigenic signal transduction mechanisms, including PI3K. Inhibiting ET-1 is an initially attractive option; however, this inhibition may result in detrimental systemic effects, especially within vascular beds causing hypotension, renal, and neurologic dysfunction.We are considering interfering RNA approaches integrated in a nanodrug delivery platform, carrying multiple anticancer agents. We suggest that this approach may optimize targeted delivery while effectively targeting multiple pathways.In terms of markers of response, we agree that ET axis molecules are inappropriate; therefore, more conventional markers need to remain in use. However, this does not de...

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