Nerve Growth Factor (NGF) Encourages the Neuroinvasive Potential of Pancreatic Cancer Cells by Activating the Warburg Effect and Promoting Tumor Derived Exosomal miRNA-21 Expression

Abstract

Background. It has been reported that signaling from the nerve growth factor (NGF) pathway associated with peripheral nerves is able to contribute to perineural invasion (PNI) of pancreatic cancer (PC). Nevertheless, the underlying mechanism by which NGF leads to PNI remained poorly understood. Methods. Western blotting was employed to determine NGF level in PC and paracarcinoma tissues and in PC cell lines as well as pancreatic ductal epithelial cells. MiaPaCa-2 and CFPAC-1 cells were treated with 100 ng/ml of NGF or the NGF inhibitor Tanezumab for 24 h, CCK-8 and Transwell assays were employed to test cell proliferation, invasion, and migration, respectively. TrkA expression was knocked down in MiaPaCa-2 and dorsal root ganglion (DRG) cells treated with NGF to determine its effect on the Warburg effect. To reveal that the NGF-TrkA signaling pathway was closely associated with PC PNI, in vitro neuroinvasion model was established by using MiaPaCa-2 cells via coculturing DRG cells in Matrigel. Further, exosomes were extracted from PC cells and identified by examining the levels of specific markers for exosomes. Then RT-qPCR was applied to test miR-21-5p level in tumor derived exosomal (TDE-miR-21-5p). RIP assay was performed to validate NGF and miR-21 binding ability in MiaPaCa-2 cells. Rescue experiments were performed by using coprocessing of Tanezumab and miR-21-5p mimic on MiaPaCa-2 cells, followed by coculture with DRG cells. Subsequently, we used a model of neuroinvasion in nude mice to assess the effect of NGF in vivo on tumor nerve invasion as well as on nociceptive transmission. Results. NGF level was preeminently higher in PC tissues and cell lines than in paracarcinoma tissues and normal pancreatic epithelial cell lines. NGF promoted MiaPaCa-2 and CFPAC-1 cell invasion and migration, while Tanezumab treatment showed the opposite results. Besides, NGF binding to TrkA receptors encouraged the intracellular Warburg effect in PC and DRG cells. TrkA blocking-up could restrain NGF induced PC cell migration and neural invasion. Mechanistically, NGF could upregulate TDE-miR-21-5p levels, and DRG cells took up TDE to activate the Warburg effect and stimulate nociceptor gene expression. miR-21-5p inhibitor could abolish the facilitative effect of NGF on PNI in MiaPaCa-2 cells. In vivo tumorigenesis experiments, Tanezumab markedly alleviated nerve invasion of PC cells as well as relieved nociceptive conduction in animal models. Conclusions. These findings displayed that NGF/TrkA encouraged the neuroinvasive potential of PC cells by activating the Warburg effect in DRG cells through upregulation of TDE-miR-21-5p expression.

Lingual Denervation Improves the Efficacy of Anti-PD-1 Immunotherapy in Oral Squamous Cell Carcinomas by Downregulating TGFβ Signaling

“… 55 ), anti-NGF mAb (tanezumab; ref. 56 ), and CGRP inhibitors (eptinezumab, erenumab; ref. 57 ) have been approved by FDA and are potential to be repurposed as adjuvant therapies to regulate the intratumoral neural component.…”

Causal inference of the effect of inflammatory cytokines on the risk of oral cancer：two-sample Mendelian randomization

“…Studies have shown elevated NGF expression in oral cancer tissues compared to healthy mucosa. Higher NGF levels have also been associated with increased proliferation and migration in oral cancer cell lines.In pancreatic cancer,NGF could encourage the neuroinvasive potential of pancreatic cancer cells [34] ,and treatment targeting nerve growth factor (NGF) and autophagy pathways may inhibit peripheral nerve in ltration and provide new research directions and an experimental basis for understanding and treating this process [35] . A study [36] initially examinied the low-glucose environment of oral mucosa carcinomas, we discovered that cancer cells employ ROS-triggered activation of c-Jun to secrete NGF, which conditions nociceptive nerves for calcitonin gene-related peptide (CGRP)…”

The contribution of the nervous system in the cancer progression

“…NGF is a neurotrophic growth factor that controls neuronal survival and neurite outgrowth mainly via its receptors, tropomyosin receptor kinase A (TrkA) with high affinity or p75 neurotrophin receptor (p75NTR) with low affinity ( 15 ). Last decades, numerous studies have been conducted to inhibit tumor growth by regulating the NGF-TrkA signaling ( 16 - 18 ). In tumor tissues obtained from patients with breast cancer, significantly increased NGF levels were observed in nerve fibers + breast carcinoma compared to that in nerve fibers − breast carcinoma suggesting an association between the growth of nerve-innervating tumor and NGF expression ( 19 ).…”