Neurotrophins mediate their diverse functions through two structurally distinct classes of transmembrane receptors; the ‘common’ p75 neurotrophin receptor (p75^NTR) which binds all of the neurotrophins with approximately equal affinity, and specific receptor tyrosine kinase receptors called tropomyosin related kinases (Trks) that exhibit specificity in neurotrophin binding (Figure 1) [18,25].

p75^NTR is a member of the death-promoting tumor necrosis factor receptor (TNF-R) superfamily, which also includes the characteristic death receptors TNF-R apoptosis-inducing ligand (TRAIL)-R and Fas/CD95 [26,27]. p75^NTR is a 75 kDa glycoprotein with four extracellular cysteine rich repeats required for ligand binding [28,29]. It is a single pass type I transmembrane receptor, with an intracellular domain containing a juxtamembrane region and a type II consensus death domain (DD) sequence [18]. Although p75 binds dimeric neurotrophin ligands, there is some controversy over the oligomeric status of p75 and evidence indicates that it may signal as a monomer or as a dimer [30]. Recently, it was shown that p75 can form covalent homodimers through a disulphide bond in the transmembrane region. In this case, neurotrophin binding is understood to induce a conformational change in the receptor, such that it pivots on this disulfide bond at Cys257 to permit access of intracellular adaptor proteins to the intracellular domain in what is termed a snail-tong model [30-32]. The receptor does not possess intrinsic enzymatic activity and transduces signals through recruitment of a variety of adaptor proteins to the intracellular domain, leading to proliferation, survival, or cell death [18,33]. p75^NTR-mediated downstream signal transduction pathways are extremely diverse, being heavily dependent on the cell context, availability of intracellular adaptor molecules and expression of interacting transmembrane co-receptors [34]. Interestingly, p75^NTR also serves as a receptor for immature pro-neurotrophins which induce cell death in a manner that is dependent on binding to a co-receptor, sortilin [35]. Furthermore, p75^NTR is capable of eliciting cellular signals in the absence of bound ligand [31]. p75^NTR signaling can also lead to downstream activation of nuclear factor-kappa B (NF-κB) which promotes cell survival through upregulation of anti-apoptotic genes such as cFLIP, which interferes with the activation of initiator caspase-8, the Bcl-2 family member Bcl-X_L, and inhibitor of apoptosis proteins (IAPs) XIAP and cIAP1/2 [36-38]. Together with the pro-apoptotic effects of p75, these findings show that p75^NTR can act as a bifunctional switch directing the cell down opposing paths of cell death or survival.

There is a wide array of proteins that have been demonstrated to interact with the intracellular domain of p75^NTR. These include neurotrophin receptor-interacting MAGE homolog (NRAGE), p75^NTR-associated cell death executor (NADE) and neurotrophin receptor-interacting factor (NRIF), which are all involved in mediating pro-apoptotic effects; FAP-1, receptor interacting protein 2 (RIP2) and tumor necrosis factor receptor associated factor 6 (TRAF6), which are involved in mediating pro-survival effects, while Schwann cell factor-1 (SC-1) is involved in mediating cell cycle effects and RhoA affects neuritogenesis [18]. In addition, p75 has been reported to interact with other TRAF family members, particularly TRAF2 and TRAF4, eliciting different effects on NF-κB activation and apoptosis [39].

Trk receptors are structurally unrelated to p75^NTR and are receptor tyrosine kinases [40]. In contrast to p75^NTR, Trk receptors are selective in which neurotrophins they will bind, with TrkA preferring NGF, TrkB preferring BDNF and NT-4/5 and TrkC preferring NT-3 [40]. Binding of their dimeric ligands induces receptor homodimerization and trans-autophosphorylation in the intracellular domain leading to activation of three well characterized intracellular signal transduction pathways, including Ras/mitogen-activated protein kinase (MAPK) signaling, phosphatidylinositol 3-kinase (PI3K)/Akt and phospholipase Cγ (PLCγ) [28,41]. These pathways are best studied in neuronal cells, where they are involved in neuronal growth, survival and differentiation [40]. In non-neuronal cells, activation of Trk receptors is linked to proliferation, survival, migration and increased invasiveness of cancer cells (Figure 1) [42,43].

In addition, TrkA signaling has also been linked to induction of autophagy in cancer cells. For example, autophagy was identified as a novel mechanism of NGF-induced cell death via TrkA in the human glioblastoma cell line, G55 [44]. Several characteristics of autophagy were observed, including autophagic vacuoles, acidic vesicular organelles (which could be prevented by bafilomycin A1), increased processing of LC3, lack of detection of caspase activation, and vacuolation was prevented by the autophagy inhibitor 3-methyladenine [44]. The authors also found that activation of ERK and c-Jun N-terminal kinase, but not p38, were involved in autophagic vacuolation [44]. Very recently, TrkA has been shown to induce cell death in medulloblastoma Daoy cells without caspase activation and demonstrating several clear signs of autophagy [45]. However, siRNA silencing of four proteins essential to autophagy (beclin-1, Atg5, LC3 and Atg9) neither blocked NGF-induced vacuole formation or cell death [45]. Instead, the authors revealed that it is the hyperstimulation of macropinocytosis, a form of bulk fluid endocytosis [46], that is responsible for their death and that it requires TrkA-dependent activation of casein kinase 1 [45]. In both of these studies, TrkA was ectopically expressed in the cells. In fact, a series of studies by Kim and colleagues demonstrates that overexpression of TrkA in U20S osteosarcoma and SK-N-MC neuroblastoma cells induces cell death even in the absence of NGF and that, at least in U2OS, a large portion of this cell death is caused by induction of autophagy [47]. Together, these studies suggest that TrkA expression might confer a growth disadvantage to certain cancerous cells, such as glioblastoma [44], medulloblastoma [45] and osteosarcoma [47]. However, further research is required to ascertain the precise mode of death. Nevertheless, TrkA activation may potentially provide a therapeutic avenue, particularly as several current chemotherapeutics also induce autophagic cell death, e.g., [48].

Breast cancer is the most common malignancy in women, and is a major cause of cancer-related death in women, second in mortality only to lung cancer [80]. Advances in understanding of the disease, and development of therapeutics targeting estrogen and epidermal growth factor signaling, correlate with a notable reduction in breast cancer mortality levels during the past 10 years, with adjuvant chemotherapeutic agents such as anthracyclines, taxanes and cyclophosphamides also becoming intrinsic to cancer treatment regimes. Normal breast epithelial cells express TrkA and p75^NTR [55]. The function of these receptors in breast development and physiology has not been studied and in fact, ligands for these receptors are not normally expressed by adult breast cells, although it is not clear whether the receptors are activated by NGF secreted in a paracrine fashion. In contrast, breast tumors have been shown to express NGF. For example, immunohistochemical analysis has shown that NGF is expressed in up to 80% of breast cancer tumor biopsies [81]. This expression does not discriminate between breast tumor subtype and NGF mRNA levels are expressed at levels comparable to those seen in the MDA-MB-231 breast cancer cell line [81]. In another study, investigation of the expression of NGF and its receptors in breast cancer cells in effusions and solid tumors revealed that activated TrkA (i.e., phosphorylated TrkA) is upregulated in effusions compared to primary breast tumors and lymph node metastasis, with downregulation of p75 in effusions [82]. Another recent immunohistochemical analysis has shown expression of NGF in fat tissue extracts from high risk patients [83]. Thus far, NGF expression has not been associated with expression of any known prognostic factor, such as estrogen receptor (ER), progesterone receptor (PR) or human epidermal growth factor receptor 2 (HER2). Anti-estrogens such as tamoxifen are used in approximately two thirds of breast cancers, while HER2 is overexpressed in 25–30% of human metastatic breast cancers [84]. Since NGF is expressed in 80% of breast cancers, this suggests that NGF signaling has a potentially broader target range than current therapies, which are directed against either estrogen signaling or HER2. Importantly, NGF can stimulate mitogenesis and pro-survival signaling in triple negative breast cancer cells (MDA-MB-231) which lack ER, PR and HER2 and are the most difficult type of breast tumor to treat [42]. Importantly, a xenograft model of breast cancer using MDA-MB-231 cells is responsive to anti-NGF treatments, including antibody and siRNA downregulation of NGF [81].

NGF signaling has been implicated in the proliferation and survival of breast cancer cells and more recently, migration [42,43,85]. The proliferative effect is via TrkA signaling and can be inhibited by tyrosine kinase inhibitors such as the indolocarbazole K252a [42]. Interestingly, tamoxifen has also been reported to inhibit NGF mediated TrkA phosphorylation in an estrogen receptor-independent manner [86]. TrkA-induced proliferation of breast cancer cells proceeds through activation of MAP kinase signaling leading to ERK phosphorylation and an NGF-dependent decrease in cell cycle duration (Figure 2) [42]. In contrast, NGF does not activate MAP kinases in normal breast epithelial cells and is not mitogenic for these cells [55]. The reason for this lack of MAP kinase activation in normal breast epithelial cells is not clear, since these cells express TrkA [55]. Recently, TrkA signaling was also shown to play a role in breast cancer metastasis and angiogenesis, which expands the role of NGF signaling in this disease [43,58].

NGF exerts pro-survival effects on a wide range of breast cancer cell lines (MCF-7, MDA-MB-231, T-47D and BT-20) and can protect them from C2-ceramide induced apoptosis [42,61]. In contrast to mitogenic signaling, NGF pro-survival effects in breast cancer cells are mediated by the p75^NTR receptor [42]. The precise mechanism by which NGF signaling through p75^NTR protects breast cancer cells is largely unknown but functional studies have shown that it is mediated by activation of NF-κB [42]. In breast cancer cells the adaptors that link NGF/p75^NTR receptor activation to NF-κB and pro-survival signaling are largely unknown. In MCF-7 cells neurotrophin-dependent recruitment of TNF-R-associated death domain (TRADD) to p75^NTR has been shown to be required for activation of NF-κB and pro-survival signaling (Figure 2) [60]. Other death receptors, such as TNF-R1 and TRAIL, which recruit TRADD to their DDs, use it as a platform for the recruitment of additional adaptor proteins including receptor-interacting protein 1 (RIP1), TNF-R-associated factor 2 (TRAF2) and Fas-associated death domain (FADD) to promote either cell survival or death [87]. In fact, knockdown of TRADD causes cells to be deficient in both TNFα-induced NF-κB activation and in caspase-8-dependent apoptosis [88]. Notably, there is currently no evidence that TRADD associated with p75^NTR can recruit these or other adaptors in breast cancer cells or other cells. Instead, in Schwann cells, RIP2 has been shown to interact directly with the DD of p75^NTR and to stimulate pro-survival signaling through activation of NF-κB [89]. In addition, certain members of the TRAF family (TRAF2, 4, 6) have been reported to interact directly with the p75^NTR intracellular domain [39,90-92]. The role of TRAF proteins in p75^NTR-mediated pro-survival signaling in breast cancer is not known and may be an avenue worthy of investigation.

In breast cancer cells, a novel adaptor Bex2 has recently been shown to be required for NGF/p75^NTR-dependent NF-κB activation and prosurvival effects (Figure 2) [61,63]. BEX2 is a 15 kDa protein which is over-expressed in ER-positive breast cancer cells and is suggested to be an estrogen-regulated gene [61]. Although it has not been shown whether or not BEX2 interacts directly with p75^NTR, it is reported to be necessary and sufficient for the anti-apoptotic function of NGF in breast cancer cells [61,62]. Furthermore, it has been shown to protect breast cancer cells from mitochondrial-mediated apoptosis, through modulation of Bcl-2 family member proteins, including up-regulation of anti-apoptotic Bcl-2 and down-regulation of pro-apoptotic members Bad, Bak and PUMA [62]. Interestingly, the homologues BEX1 and BEX3 have also been shown to interact with p75^NTR, and in neural tissues BEX1 inhibits NF-κB, links neurotrophin signaling to the cell cycle and may sensitize cells to apoptosis [93,94]. BEX2 has also been shown to be required for progression of MCF-7 breast cancer cells through the G₁ phase of the cell cycle via its regulation of cyclin D1 and p21 [62,63].

Prostate cancer is the most commonly diagnosed solid tumor in men and the second leading cause of male cancer related deaths [80]. Its incidence significantly increases with age, regardless of variables such as diet, occupation and life style [95]. The normal adult prostate is composed of ducts with epithelial cells associated with stromal cells. The epithelial cells are of three types; secretory luminal cells which line the lumen of the duct and are surrounded by a continuous layer, formed mainly by basal epithelial cells and a scattering of neuroendocrine cells. The stroma is separated from the epithelial cells by the basal lamina, and is mainly composed of smooth muscle cells. In human prostate cancer, there is progressive disorganization of luminal and basal epithelial cells, leading to breakdown of the basal lamina and eventual metastasis of the cells to other body sites [54]. While the tumor cells are initially dependent on androgen for their survival, this dependence is progressively lost leading to development of resistance to androgen-targeting therapies. Interestingly, the secretory luminal cells are androgen-sensitive, while the basal epithelial cells express low levels of androgen receptor and the neuroendocrine cells are androgen insensitive, [54].

After the central nervous system the human prostate is the next most abundant source of NGF [96], where it plays a role in normal prostate development [97]. Stromal cells secrete NGF which binds to TrkA and p75^NTR present on prostate epithelial cells stimulating their growth [98-100]. It is of interest to note that BDNF, which can bind to p75^NTR, has also been shown to be expressed by normal prostate stromal cells [101].

Progression of prostate cancer is accompanied by modifications in the expression of neurotrophins, including NGF, and neurotrophin receptors [95,102]. The most striking and important change in this regard is a reduction in p75^NTR expression by the epithelial cells [73]. While there is no change in the expression of NGF or TrkA, the epithelial layer exhibits a reduction in p75^NTR expression [73]. Immunocytochemical staining of both normal and malignant prostate epithelial tissue shows progressive loss of p75^NTR expression is associated with tumor development [98]. Importantly, the loss of p75^NTR is directly related to the grade of malignancy with early stages (prostatic intraepithelial neoplasia) still showing p75^NTR, and poorly differentiated carcinomas having undetectable levels [103]. This is also seen in prostate cancer cell lines, where p75^NTR is absent from cell lines derived from advanced metastatic prostate cancer [73,74,98]. These studies also show that TrkA receptor expression is unchanged, resulting in an overall decrease in the p75^NTR:TrkA ratio during prostate cancer progression. This is also accompanied by an increase in the in vitro invasive capacity of human prostatic cancer cells in xenograft models [104]. This has been reported to involve increased expression of heparanase [103]. In contrast, the mitogenic action of NGF on prostate cancer lines is mediated by TrkA [105].

There is also some evidence to suggest that during progression of prostate cancer epithelial cells acquire the ability to express neurotrophins [101]. For example, the non-metastatic LNCaP cell line does not express neurotrophins, while metastatic lines DU145 and PC-3 secrete measurable amounts of NGF [99,104] and PC-3 secretes both NGF and BDNF [99]. This switch from paracrine to autocrine control of neurotrophin activity, as well as loss of p75^NTR, could facilitate survival and proliferation of these cells upon metastasis to other regions of the body.

This acquired ability of prostatic cancer cells to evade cell death has lead to p75^NTR being proposed as a tumor suppressor in prostate cancer cells [71,72,106]. In the absence of p75^NTR, prostatic cancer cells respond to proliferative signals mediated by TrkA activation and proliferate. In fact, treatment of prostate tumor cells with pharmacological inhibitors of TrkA signaling, including K252a and CEP-701, reduces proliferation induced by NGF and leads to increased cell death [107,108]. When p75^NTR is artificially reintroduced into prostate cancer cells, the cells exhibit cell cycle arrest, accumulating in S phase, and undergo an increase in spontaneous apoptosis [71,105,109]. The mechanism of induction of apoptosis by NGF/p75^NTR has not been fully elucidated. However, reintroduction of p75^NTR expression into PC-3 cells is accompanied by a decrease in activation of NF-κB and c-Jun N-terminal kinase (JNK), suggesting these signals mediate the anti-apoptotic effect in the absence of p75^NTR [110]. Although pro-apoptotic signaling by p75^NTR in prostate cells requires the DD [71], it does not involve activation of initiator caspases-8 or -10 which are activated by other death receptors such as TNF-R, TRAIL receptors and Fas [111,112].

The loss of p75^NTR in prostate cancer does not appear to be due to errors or mutations in the genetic transcript coding for p75^NTR. Rather, it has been linked to instability of the mRNA, which may be due to changes in the 3′-untranslated region (3′-UTR) of the p75^NTR gene [73]. The 3′-UTR of genes generally contains several regulatory sequences involved in poly-adenylation, mRNA stability and binding sites for micro-RNAs (miRNAs) which are involved in controlling mRNA stability and translation [113,114]. Thus, for example, it is possible that enhanced expression of miRNAs that target the 3′-UTR of p75^NTR could be responsible for downregulation of expression of the protein. Interestingly, gonadotrophin releasing hormone (GnRH) analogue therapy induces a significant increase in p75^NTR protein expression (with no increase in TrkA expression) [115], prompting the question as to whether GnRH analogue increases the stability of p75^NTR mRNA.