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Background: Perineural invasion (PNI) is a typical pathological characteristic of salivary adenoid cystic carcinoma (SACC) and other neurotrophic cancers. The mechanism of the neural microenvironment controlling tumor progression during the PNI process is unclear. In the present study, we investigated the role and molecular mechanisms of nerve-derived neuropeptide galanin (GAL) and its receptor (GALR2) in the regulation of PNI in SACC., Methods: Immunohistochemistry staining and clinical association studies were performed to analyze the expression of GAL and GALR2 in SACC tissues and their clinical value. Dorsal root ganglion or SH-SY5Y cells were co-cultured with SACC cells in vitro to simulate the interactions between the neural microenvironment and tumor cells, and a series of assays including transcriptome sequencing, Western blot, and Transwell were performed to investigate the role and molecular mechanism of GAL and GALR2 in the regulation of SACC cells. Moreover, both the in vitro and in vivo PNI models were established to assess the potential PNI-specific therapeutic effects by blocking the GAL/GALR2 axis., Results: GAL and GALR2 were highly expressed in SACC tissues, and were associated with PNI and poor prognosis in SACC patients (p < 0.05). Nerve-derived GAL activated GALR2 expression in SACC cells and induced epithelial-to-mesenchymal transition (EMT) in SACC cells. Adding human recombinant GAL to the co-culture system promoted the proliferation, migration, and invasion of SACC cells significantly, but inhibited the apoptosis of SACC cells. Adding M871, a specific antagonist of GALR2, significantly blocked the above effects (p < 0.05) and inhibited the PNI of SACC cells in vitro and in vivo (p < 0.05)., Conclusions: This study demonstrated that nerve-derived GAL activated GALR2 expression, and promoted EMT in SACC cells, thereby enhancing the PNI process. Interruption of the GAL/GALR2 axis might be a novel strategy for anti-PNI therapy for SACC., (© 2022 The Authors. Cancer Medicine published by John Wiley & Sons Ltd.)

Salivary adenoid cystic carcinoma (SACC) is a malignant tumor that is characterized by perineural invasion (PNI). p53 is an essential tumor-suppressor gene and p53 mutations play a critical role in tumor occurrence and progression (e.g., pancreatic, prostate and head and neck cancer). However, the regulatory role of the p53 gene in SACC and the PNI process remains unknown. In the present study, we employed RNA interference technique to downregulate p53 gene expression in SACC-83 cells to explore the role of p53 in the PNI process. Our results showed that the downregulation of the p53 gene induced significant 'epithelial-mesenchymal transition (EMT)-like changes' in SACC-83 cells, including decreased expression levels of epithelial markers (E-cadherin, EMA and CK5) and increased expression levels of mesenchymal markers (vimentin, N-cadherin and C-cadherin). The downregulation of p53 also caused a lower apoptotic index of Annexin V-FITC/PI and a lower number of SACC-83 cells in the second G0/G1 phase of the cell cycle. Furthermore, the downregulation of the p53 gene resulted in a significant increase in PNI activity in the SACC-83 cells. Thus, our findings revealed that downregulation of p53 promoted in vitro PNI activity through 'EMT-like changes' in SACC-83 cells. The present study suggests the essential regulatory role of p53 in the PNI activity of SACC cells, and implies that p53 may be a new target gene for the clinical treatment of SACC.

Salivary adenoid cystic carcinoma (SACC) has a unique tendency for perineural invasion (PNI), which results in tumor recurrence and poor prognosis. Recent studies have shown that the chemokine CCL5 and its receptor CCR5 play important roles in tumor invasion and metastasis. However, the role of the CCL5/CCR5 axis in the PNI of SACC has not been studied to date. In the present study, we evaluated the expression of CCL5 and CCR5 in SACC cases and nerve tissues, and performed a series of in vitro assays with the SACC cell line, SACC-83, to indicate the role of the CCL5/CCR5 axis in the PNI of SACC. We found that CCL5 (35.9%; 23/64) and CCR5 (70.3%; 45/64) were positively expressed in SACC cases, and the expression of CCR5 was significantly associated with the PNI of SACC (P<0.05). We also found that SACC-83 cells expressed the functional receptor, CCR5, for the chemokine CCL5, as demonstrated by calcium mobilization and actin polymerization assays. Furthermore, we found that exogenous CCL5 significantly facilitated the migration, invasion and PNI activity of SACC-83 cells in vitro (P<0.05). Further study showed that the CCR5 inhibitor (maraviroc) effectively blocked the migration, invasion and PNI activity of SACC-83 cells with or without CCL5 stimulation (P<0.05). These results indicate that the CCL5/CCR5 axis plays a critical role in the PNI of SACC, and that antagonists against CCR5 may be an effective anti-PNI agent for SACC therapy.