The Glial cell line-derived neurotrophic Family Ligands (GFL) are soluble neurotrophic factors that are required for development of multiple human tissues, but which are also important contributors to human cancers. GFL signaling occurs through the transmembrane RET receptor tyrosine kinase, a well-characterized oncogene. GFL-independent RET activation, through rearrangement or point mutations occurs in thyroid and lung cancers. However, GFL-mediated activation of wildtype RET is an increasingly recognized mechanism promoting tumor growth and dissemination of a much broader group of cancers. RET and GFL expression have been implicated in metastasis or invasion in diverse human cancers including breast, pancreatic, and prostate tumors, where they are linked to poorer patient prognosis. In addition to directly inducing tumor growth in these diseases, GFL-RET signaling promotes changes in the tumor microenvironment that alter the surrounding stroma and cellular composition to enhance tumor invasion and metastasis. As such, GFL RET signaling is an important target for novel therapeutic approaches to limit tumor growth and spread and improve disease outcomes.

Here, we describe a rapid and flexible protocol for the formation of 3D cell spheroids through cell aggregation. This is easily adapted ...

RET encodes a receptor tyrosine kinase that is essential for spermatogenesis, development of the sensory, sympathetic, parasympathetic, and enteric nervous systems and the kidneys, as well as for maintenance of adult midbrain dopaminergic neurons. RET is alternatively spliced to encode multiple isoforms that differ in their C-terminal amino acids. The RET9 and RET51 isoforms display unique levels of autophosphorylation and have differential interactions with adaptor proteins. They induce distinct gene expression patterns, promote different levels of cell differentiation and transformation, and play unique roles in development. Here we present a comprehensive study of the subcellular localization and trafficking of RET isoforms. We show that immature RET9 accumulates intracellularly in the Golgi, whereas RET51 is efficiently matured and present in relatively higher amounts on the plasma membrane. RET51 is internalized faster after ligand binding and undergoes recycling back to the plasma membrane. This differential trafficking of RET isoforms produces a more rapid and longer duration of signaling through the extracellular-signal regulated kinase/mitogen-activated protein kinase pathway downstream of RET51 relative to RET9. Together these differences in trafficking properties contribute to some of the functional differences previously observed between RET9 and RET51 and establish the important role of intracellular trafficking in modulating and maintaining RET signaling.

The RET receptor tyrosine kinase has essential roles in cell survival, differentiation, and proliferation. Oncogenic activation of RET causes the cancer syndrome multiple endocrine neoplasia type 2 (MEN 2) and is a frequent event in sporadic thyroid carcinomas. However, the molecular mechanisms underlying RET's potent transforming and mitogenic signals are still not clear. Here, we show that nuclear localization of β-catenin is frequent in both thyroid tumors and their metastases from MEN 2 patients, suggesting a novel mechanism of RET-mediated function through the β-catenin signaling pathway. We show that RET binds to, and tyrosine phosphorylates, β-catenin and show that the interaction between RET and β-catenin can be direct and independent of cytoplasmic kinases, such as SRC. As a result of RET-mediated tyrosine phosphorylation, β-catenin escapes cytosolic down-regulation by the adenomatous polyposis coli/Axin/glycogen synthase kinase-3 complex and accumulates in the nucleus, where it can stimulate β-catenin–specific transcriptional programs in a RET-dependent fashion. We show that down-regulation of β-catenin activity decreases RET-mediated cell proliferation, colony formation, and tumor growth in nude mice. Together, our data show that a β-catenin–RET kinase pathway is a critical contributor to the development and metastasis of human thyroid carcinoma. [Cancer Res 2008;68(5):1338–46]

Multiple endocrine neoplasia type 2 is an inherited cancer syndrome characterized by tumors of thyroid and adrenal tissues. Germline mutations of the REarranged during Transfection (RET) proto-oncogene, leading to its unregulated activation, are the underlying cause of this disease. Multiple endocrine neoplasia type 2 has been a model in clinical cancer genetics, demonstrating how knowledge of the genetic basis can shape the diagnosis and treatment of the disease. Here, we discuss the nature and effects of the most common recurrent mutations of RET found in multiple endocrine neoplasia type 2. Current understanding of the molecular mechanisms of RET mutations and how they alter the structure and function of the RET protein leading to its aberrant activation, and the effects on RET localization and signaling are described.