Pro-nerve Growth Factor Induces Autocrine Stimulation of Breast Cancer Cell Invasion through Tropomyosin-related Kinase A (TrkA) and Sortilin Protein

Abstract

The precursor of nerve growth factor (proNGF) has been described as a biologically active polypeptide able to induce apoptosis in neuronal cells, via the neurotrophin receptor p75 NTR and the sortilin receptor. Herein, it is shown that proNGF is produced and secreted by breast cancer cells, stimulating their invasion. Using Western blotting and mass spectrometry, proNGF was detected in a panel of breast cancer cells as well as in their conditioned media. Immunohistochemical analysis indicated an overproduction of proNGF in breast tumors, when compared with benign and normal breast biopsies, and a relationship to lymph node invasion in ductal carcinomas. Interestingly, siRNA against proNGF induced a decrease of breast cancer cell invasion that was restored by the addition of noncleavable proNGF. The activation of TrkA, Akt, and Src, but not the MAP kinases, was observed. In addition, the proNGF invasive effect was inhibited by the Trk pharmacological inhibitor K252a, a kinase-dead TrkA, and siRNA against TrkA sortilin, neurotensin, whereas siRNA against p75 NTR and the MAP kinase inhibitor PD98059 had no impact. These data reveal the existence of an autocrine loop stimulated by proNGF and mediated by TrkA and sortilin, with the activation of Akt and Src, for the stimulation of breast cancer cell invasion. Nerve growth factor (NGF),5 the prototypical member of the neurotrophin family of polypeptides, is essential for the survival and differentiation of central and peripheral neurons, and its role in the development and regeneration of the sympathetic and sensory nervous systems has been extensively described (1). NGF binds to the tropomyosin-related kinase A (TrkA) receptor, a tyrosine kinase receptor, and to the p75 neurotrophin receptor (p75 NTR ), a member of the tumor necrosis factor receptor family, to induce its neurotrophic effects. NGF is synthesized as a 25-kDa precursor protein, named proNGF, that yields the mature NGF polypeptide of 13.5 kDa and an inactive prosegment of 11.5 kDa, released from the N terminus intracellularly by furin, or extracellularly by plasmin as well as by several matrix metalloproteases (2). Importantly, proNGF can be secreted without being processed to mature NGF and can have its own biological effects (3). As more than just a source for NGF, proNGF was shown to induce neuronal death by apoptosis where mature NGF induced survival and differentiation (4, 5). For inducing its proapoptotic effect on neurons, proNGF forms a trimeric complex with two plasma membrane receptors: p75 NTR and sortilin (4). Sortilin, a 95-kDa type I receptor,

NGF and ProNGF: Regulation of neuronal and neoplastic responses through receptor signaling

“…As described above, breast cancers express and respond to NGF, and therefore the discovery that proNGF is secreted by tumor cells was not overly surprising (Demont et al ., 2012). However the determination that it stimulates their migration/invasion through an autocrine loop mediated by TrkA and sortilin was unexpected.…”

“…In addition, a comparison between proNGF levels and clinicopathological parameters revealed a correlation with lymph node invasion. In invasive ductal carcinomas, which represent the majority of breast cancers, there was no correlation with histological grade, tumor value, axillary lymph node status, age and presence of estrogen receptors, although a statistically significant association was obtained between the quantity of proNGF and lymph node invasion, suggesting a link to metastasis (Demont et al ., 2012). Indeed, proNGF may serve as a biomarker of metastasis and possibly as a therapeutic target in breast cancer.…”

Nerve–Cancer Cell Cross-talk: A Novel Promoter of Tumor Progression

“…This has been particularly well described for neurotrophins in breast cancer (22). For instance, NGF and proNGF are expressed in breast cancer and stimulate tumor cell growth and invasion (23,24), including the stem cell compartment (25). In prostate cancer, changes in the expression of NGF and its Trk receptor contribute to tumor cell growth and dissemination through a variety of kinasebased signaling pathways, and the inhibition of Trk receptors decreases the growth of prostatic cancer cells (26).…”

p75 neurotrophin receptor and pro-BDNF promote cell survival and migration in clear cell renal cell carcinoma

“…We have previously described another function of sortilin, as a crucial protein to form a complex with two receptors containing a tyrosine kinase domain, TrkB and EGFR, in lung cancer [48]. Sortilin complexed with TrkA, another tyrosine kinase receptor for NGF, was also identified in breast cancer, promoting cell invasion through pro-NGF binding to this complex [49]. Similar results were reported in melanoma, expressing Trks, p75 NTR and sortilin, wherein cell migration and invasion induced by pro-NGF were depending on p75 NTR [50].…”