Advances in understanding cancer-associated neurogenesis and its implications on the neuroimmune axis in cancer

Abstract

Nerves and immunologic mediators play pivotal roles in body homeostasis by interacting with each other through diverse mechanisms. The spread of nerves in the tumor microenvironment increases tumor cell proliferation and disease progression, and this correlates with poor patient outcomes. The effects of sympathetic and parasympathetic nerves on cancer regulation are being investigated. Recent findings demonstrate the possibility of developing therapeutic strategies that target the tumor microenvironment and its components such as immune cells, neurotransmitters, and extracellular vesicles. Therefore, examining and understanding the mechanisms and pathways associated with the sympathetic and parasympathetic nervous systems, neurotransmitters, cancer-derived mediators and their interactions with the immune system in the tumor microenvironment may lead to the development of new cancer treatments. This review discusses the effects of nerve cells, immune cells, and cancer cells have on each other that regulate neurogenesis, cancer progression, and dissemination.

1. Introduction

Cancer is an extremely complex disease to fight because of its social and psychological aspects as well as the physical pain and destruction that it causes, and its incidence is increasing worldwide. In 2020, about 10 million deaths worldwide were cancer deaths (Sung et al., 2021). In clinical management of cancer, the most widely preferred treatment methods are insufficient to meet patient needs because of several limitations such as associated toxicity, adverse effects, high cost, and drug resistance. Therefore, discovering new mechanisms and biomarkers to aid in the diagnosis and treatment of advanced cancers is needed to achieve better patient outcomes.

Cancer cells divide and grow uncontrollably and spread to other parts of the body. This can result from many mechanisms, such as the activation of proto-oncogenes by translocations and point mutations and the inactivity of genes that normally limit cell proliferation. Although previous research focused on the ways in which tumor cell-intrinsic pathways (e.g., deregulation of proliferative signals, activation of oncogenes) contribute to the progression of cancer, growing evidence suggests that tumor cell-extrinsic mechanisms heavily influence the tumor microenvironment, which is an essential but complex setting for cancer cell dissemination that includes nerves and specific molecular signals (Amit, Na’ara, & Gil, 2016). Studies conducted in recent years have shown that tumor development is supported by factors such as the immune system and perineural invasion (Liebig, Ayala, Wilks, Berger, & Albo, 2009) that are significant players in the tumor microenvironment. Neurons that interact with immune and structural cells in the perineural niche drive tumorigenesis for several cancer types, including pancreatic, gastric, hematopoietic, and non-melanoma skin cancers (Hunt, Andujar, Silverman, & Amit, 2021). Immune cells such as lymphocytes, macrophages, and natural killer (NK) cells have a prominent role in antitumor responses and markedly regulate tumor proliferation and cancer metabolism (Leone & Powell, 2020). Therefore, the role of the neuroimmune axis in cancer is an intriguing research field.

Recent studies have shown that the relationship between nerves and cancer is not unilateral but rather bidirectional. Although the sympathetic and parasympathetic nervous systems play a role in the behavior of tumor cells, investigators have shown that neurotrophic growth factor, axon guidance molecules, vascular endothelial growth factor (VEGF), and chemical messengers secreted by tumor cells increase neoneurogenesis, axonogenesis, and angiogenesis (O’Donnell, Chance, & Bashaw, 2009; Palm & Entschladen, 2007; Zhang et al., 2021). The tumor-infiltrating components of tissues include dense inflammatory molecules and various cell types such as endothelial cells, immune cells, smooth muscle cells, epithelial cells, adipocytes, and fibroblasts, that maintain homeostasis in the tumor microenvironment or promote tumor development (Kamiya, Hiyama, Fujimura, & Yoshikawa, 2021; Scheff & Saloman, 2021). This neuroimmune interaction plays a significant role in the mechanisms of treatment resistance, growth of cancer cell population, and angiogenesis (Kamiya et al., 2021; Scheff & Saloman, 2021).

In this review article, we discuss recent developments in the relationship between neuroimmune cells and cancer by focusing on the mechanisms of the sympathetic and parasympathetic nervous systems, neurotransmitters, cancer-derived mediators, and the role of the neuroimmune axis in cancer progression.

2. Sympathetic nervous system

The sympathetic nervous system stimulates tumor growth and metastasis (Cole, Nagaraja, Lutgendorf, Green, & Sood, 2015) via epinephrine/norepinephrine, and catecholamines act directly on tumors via endothelial cells, immune cells, and fibroblasts. Likewise, sympathetic innervation plays a role in early cancer metastasis by activating endoneurial macrophage infiltration in the tumor microenvironment (Hunt et al., 2021; Jobling et al., 2015). Researchers have shown that the neurotransmitters of the sympathetic system promote the progression of prostate (Magnon et al., 2013; Zahalka et al., 2017), stomach (Liao et al., 2010; Lu et al., 2017), pancreatic (Ceyhan et al., 2009; Kim-Fuchs et al., 2014; Renz et al., 2018), breast (Campbell et al., 2012; Kamiya et al., 2019; Setordzi, Chang, Liu, Wu, & Zuo, 2021; Sloan et al., 2010; Tanaka & Sakaguchi, 2017; Yu et al., 2021), ovarian (Armaiz-Pena et al., 2013; Huang et al., 2016; Kang et al., 2016; Thaker et al., 2006), skin (Calvani et al., 2018; Calvani et al., 2019; Dal Monte et al., 2014; Vander Heiden, Cantley, & Thompson, 2009), brain (He et al., 2017; Venkatesh et al., 2015; Venkatesh et al., 2017), head and neck (Amit et al., 2020; Bernabe, Tamae, Biasoli, & Oliveira, 2011; Bravo-Calderon et al., 2020; Lopes-Santos, Bernabe, Miyahara, & Tjioe, 2021; Yamanaka et al., 2002), and colon (Chin et al., 2016; Ciurea et al., 2017; Hicks, Murray, Powe, Hughes, & Cardwell, 2013; Zhou et al., 2018) cancers.

Magnon et al., by performing surgical or chemical sympathectomy in a mouse model, showed that adrenergic signals have an important role in prostate cancer metastasis and tumor development. Loss of (β2-adrenergic receptors (β2-AR) and (β3-adrenergic receptors (β3-AR) causes decreased tumor development and dissemination (Magnon et al., 2013). Furthermore, (β2-AR signals maintain endothelial cell metabolism and promote angiogenesis by supporting aerobic glycolysis rather than oxidative phosphorylation (Zahalka et al., 2017). In gastric cancer cells, cyclooxygenase 2 (COX-2), VEGF, and matrix metalloprotease-2 and -9 (MMP-2/–9) levels were downregulated by exposure to the non-selective (β-adrenergic antagonist, propranolol, with inhibited expression of nuclear factor-κB (Liao et al., 2010). Using a human gastric cancer cell line, surgically resected tissue specimens, and a mouse model, researchers showed that isoprenaline increases neovascularization in gastric cancers by upregulating the expression of plexin-1 and VEGF receptor-2 (Lu et al., 2017).

Investigators found that neural density, neural hypertrophy, and norepinephrine levels were higher in pancreatic cancer tissue than in normal pancreatic tissue (Ceyhan et al., 2009). An increased norepinephrine level causes an increase in neurotrophic growth factor levels and induces axonogenesis via tropomyosin receptor kinase (Trk) receptors. Inhibition of (β-AR expression interrupts bidirectional interaction between cancer-derived neurotrophic factors and adrenergic signaling, reduces pancreatic tumorigenesis, and contributes to the prolongation of survival (Renz, Takahashi, et al., 2018). Another study showed that isoproterenol increased cyclic adenosine monophosphate (cAMP) levels in tumor cells, whereas MMP-2/–9 levels were increased twofold and fourfold, respectively, by β-adrenoceptor signaling (Kim-Fuchs et al., 2014).

In breast cancer studies, Kamiya et al. found that neurotransmitters secreted from sympathetic nerves and signals of β2-AR increased cancer progression using newly generated retrograde adeno-associated virus vector-based genetic modeling (Kamiya et al., 2019). Although sympathetic nerve denervation did not change the numbers of CD4⁺ or CD8⁺ T cells, it increased the expression of programmed death ligand 1 (PD-L1) (Setordzi et al., 2021) and IFN-γ (Setordzi et al., 2021) in these cells and increased the expression of forkhead box protein 3 (Tanaka & Sakaguchi, 2017) in CD4⁺ T cells. These immune checkpoint molecules and cytokines play important roles in immunosuppression in the tumor microenvironment and will be influential in developing new immunotherapies to slow cancer progression. In addition to the development of new treatment methods that aim to slow growth of tumor, the prevention of breast cancer-related metastasis plays an important role in stopping the progression of cancer. Studies using beta blockers demonstrated that sympathetic nerves increase cancer cell migration into tissues by increasing tumor-associated macrophage (TAM) infiltration and receptor activator of nuclear factor kappa-B ligand expression (Campbell et al., 2012; Sloan et al., 2010). This stimulatory effect may arise directly from tumor cells or indirectly from the sites of metastases (Campbell et al., 2012).

Studies of ovarian cancers have had results similar to those of prostate, pancreatic, and breast cancers, suggesting that the sympathetic nervous system affects cancer. Thaker et al. showed a positive correlation between tumor weight and nodal metastasis number with increasing duration of exposure to chronic stress, which was also shown by using β-AR⁺ ovarian cancer cell lines. Researchers have not shown this correlation between tumor progression and metastasis in β-AR-null ovarian cancer cell lines. However, they did show that β-AR exerts its effects via the cAMP/PKA signaling pathway and increases tumor vascularization by increasing VEGF, MMP-2, and MMP-9 expression (Thaker et al., 2006). Furthermore, norepinephrine induces Src activation via the cAMP/PKA signaling pathway, and this activation increases tumor growth, migration, and invasion by activating Rap1 and inhibiting extracellular signal-regulated kinase (ERK) in cancer cells (Armaiz-Pena et al., 2013).

In another study of 237 ovarian cancer cases, AR expression was high in patients experiencing depression and anxiety. In the same study, more specimens with poorly differentiated histology and higher tumor stages were found in β2-AR⁺ tumors than in β2-AR⁻ tumors (T. Huang et al., 2016). Also, Kang et al. showed that norepinephrine induces expression of dual-specificity phosphatase 1 (DUSP1), which inhibits the c-Jun/c-Jun N-terminal kinase (JNK) pathway, which is important for cancer cell apoptosis (Kang et al., 2016).

β3-AR, which is involved in tumor progression and metastasis, activates glycolytic enzymes by inducing uncoupling protein 2 expression, which mediates Warburg metabolism (Vander Heiden et al., 2009) in melanoma cells (Calvani et al., 2018). Whereas low blood or oxygen supply conditions activate β3-AR diminished levels of β3-AR contribute to apoptosis of melanoma cells by downregulating nitric oxide synthesis (Dal Monte et al., 2014). Moreover, this reduction in β3-AR levels was correlated with increased numbers of NK and CD8⁺ cells as well as reduced numbers of regulatory T cells and myeloid-derived suppressor cells (MDSCs) in melanoma cases (Calvani et al., 2019).

Isoproterenol causes an increase in the number of glioblastoma cells through the ERK1/2 pathway. β-AR activation has enhanced invasion and metastasis by increasing the expression of metalloproteinases (MMP-2 and MMP-9) in glioblastomas, which is consistent with the effect of this activation on other cancer types (He et al., 2017). Venkatesh et al. showed that neuroligin-3, a synaptic protein that acts as a mitogen, stimulates the phosphatidylinositol-3-kinase (PI3K)/mammalian target of rapamycin pathway in high-grade glioma cells and that neuroligin-3 expression correlates with poor prognosis (Venkatesh et al., 2015). In addition, blocking the release of neuroligin-3 inhibits high-grade glioma growth (Venkatesh et al., 2017).

We showed that molecules that spread from cancer cells regulate adrenergic differentiation in nerves and that the resulting newly formed adrenergic nerves increase the progression of oral squamous cell cancer. We also found a positive correlation between nerve density and aggressiveness of oral cavity cancers by examining specimens from patients with oral squamous cell cancer (Amit et al., 2020).

In another study, solute carrier family 6 member 2, which encodes norepinephrine transport protein, played a role in β-adrenergic signaling in head and neck squamous cell carcinoma cases (Lopes-Santos et al., 2021). β-adrenergic signals markedly increase the expression of metalloproteinases and laminins and reduce the levels of adhesion molecules such as epithelial cell adhesion molecule, vascular cell adhesion molecule 1, and intercellular adhesion molecule 4. Investigators showed that β2-AR increases the expression of proinflammatory genes and decreases the expression of genes related to cancer cell stemness (Lopes-Santos et al., 2021). Also, norepinephrine increased the release of interleukin(IL)-6 in oral squamous cell cancer cells, which indirectly increased the growth of cancer cells (Bernabe et al., 2011). Contrary to this finding, researchers showed that the increased effect of β2-AR in oral squamous cell cancers conversely affects the migration of tumor cells (Bravo-Calderon et al., 2020; Yamanaka et al., 2002). In view of these findings, we can say that the adrenergic system can even differentially regulate the same cancer types. However, confirming this requires further study.

Contrary to that in patients with other cancer types, researchers detected sympathetic nerves in patients with colon cancers in the early stages of the disease and found that it was associated with a good prognosis and reduced lymph node metastasis (Zhou et al., 2018). Hicks et al. reported that postdiagnostic beta blocker use had no effect on mortality in their case-control study of 4794 colorectal cancer patients (Hicks et al., 2013). In contrast, in a study of 48 patients, Ciurea et al. showed a direct correlation of expression of β-AR with tumor progression, invasion, and lymph node metastasis (Ciurea et al., 2017). Similarly, Chin et al. in their study of the mechanism of the pro-tumorigenic effect of β-AR, noted that β2-AR blockade suppressed the development of colorectal cancer cells (Chin et al., 2016). Inhibition of β2-AR signaling increased the expression of G1-checkpoint cyclin-dependent kinase (CDK) inhibitors such as p21 and p27 and decreased the formation of the cyclin D1/cyclin-dependent kinase 44 complex. This prevented tumor cells from transitioning to the G2/M phase by reducing histone H3 phosphorylation and increasing histone H2A formation. In addition, β2-AR inhibition increased caspase-3, caspase-8, caspase-9, and PARP cleavage; downregulated Bcl-2, BcL-xL, and Mcl-l; and upregulated Bcl-2 Associated X-Protein (Bax), Bcl-2 homologous antagonist/killer (Bak), p53 upregulated modulator of apoptosis (Puma), and Bcl-2 like protein 11 (Bim). Through these mechanisms, β2-AR inhibition increased apoptosis of colorectal cancer cells (Chin et al., 2016) (Fig. 1).

Fig. 1.

Schematic showing how inducers or blockers of the sympathetic nervous system affect tumor progression, metastasis, and apoptosis. Propranolol causes cell apoptosis through caspases or the mitochondrial cell death pathway. Norepinephrine induces expression of the DUSP1, which inhibits cell apoptosis through regulation of the JNK pathway in the nucleus and through the mitochondrial cell death pathway. As shown, isoprenaline and chronic stress promote tumor progression, angiogenesis, and metastasis. The figure was created with BioRender.com.

3. Parasympathetic nervous system

The vagus nerve plays an important role in parasympathetic innervation of the subdiaphragmatic organs. Moreover, it affects tumorigenesis and cancer stem cells directly or indirectly, which results in dissimilar consequences for diverse types of cancers. The direct effect of vagotomy in gastric cancer patients is reduction of tumorigenesis by inhibition of cholinergic receptor muscarinic M3 (CHRM3) (Wang et al., 2018) through Wnt signaling (Zhao et al., 2014). The vagotomy outcome in colorectal cancer patients is similar to that in gastric cancer patients: decreased tumor proliferation. In contrast, vagotomy promotes pancreatic tumorigenesis by increasing proliferation of tumors (Renz et al., 2018). In light of these findings, the effects of the vagus nerve on tumorigenesis are diverse depending on the type of cancer.

The vagus nerve reduces cancer proliferation by inhibiting the sympathetic nervous system in the tumor microenvironment (Clancy et al., 2014). Consistently, the vagus nerve has promoted tumor progression by inducing β-adrenergic signaling indirectly (Renz, Takahashi, et al., 2018). Studies involving denervation of the vagus nerve showed that vagotomy increased the invasiveness of tumors, increased secretion of cytokines such as tumor necrosis factor-α (TNF-α), and decreased survival rates. Antitumor effects of the vagus nerve can occur as a result of TNF-α release from TAMs (Hutchings, Phillips, & Djamgoz, 2020). Also, loss of vagal signals around the cancer cells has increased the number of TAMs, resulting in increased in TNF-α levels (Partecke et al., 2017). Renz et al. showed that after parasympathetic denervation, expression of cholinergic receptor muscarinic type 1 receptor (Chrm1) in epithelial cells in pancreatic tumors increased cancer development by suppressing the epidermal growth factor receptor/mitogen-activated protein kinase (EGFR/MAPK) and PI3K/AKT cascades, whereas treatment with bethanechol after vagotomy reduced tumorigenesis and CD44 expression. They also showed that inhibition of CD11b⁺ myeloid cells suppressed pancreatic carcinogenesis (Renz, Tanaka, et al., 2018). Hence, vagotomy has substantial effects on pancreatic cancer growth and spread, altering tumor morphology and shortening survival (Partecke et al., 2017).

In an in vivo study, Magnon et al. showed that prostate cancer metastasis to lymph nodes increased via Chrm1 by using carbachol, a non-selective acetylcholine (ACh) receptor agonist. They showed that when Chrm1 was absent from the tumor microenvironment, no effects of carbachol on metastasis were observed, and carbachol had no effect on tumor growth. Similarly, giving pirenzepine (a Chrm1-specific antagonist) reduced metastasis of prostate cancer cells (Magnon et al., 2013). In contrast, Coarfa et al. by using chemical and physical denervation, found that bilateral botulinum toxin (Botox) application and spinal cord injury had the same effect on prostate tumor progression. They also found that the incidence of prostate tumors and overall prostate weights were reduced. Furthermore, they showed that the rate of apoptosis of tumor cells was higher in prostate cancer specimens taken from patients injected with Botox than in those from control patients (Coarfa et al., 2018).

In studies conducted with genetic manipulation in breast cancer cases, activation of the parasympathetic nervous system reduced tumor growth and distant metastasis by decreasing the expression of programmed cell death protein 1 (PD-1) and PD-L1 (Kamiya et al., 2019). Whereas the parasympathetic nervous system, like the sympathetic nervous system, did not cause any change in the number of CD4⁺ or CD8⁺ tumor-infiltrating lymphocytes, it decreased the expression of PD-1 in CD4⁺ T cells and the expression of PD-1 ligands in tumor tissue. It increased the release of IFN-γ from CD4⁺ and CD8⁺ tumor-infiltrating lymphocytes (Kamiya et al., 2019). Erin et al. showed an increase in the incidence of distant metastasis of breast cancers to the adrenal glands after vagotomy (Erin, Barkan, & Clawson, 2013).

The effects of vagotomy on gastric cancers were believed to be cancer-inducing in the 1980s. Researchers showed that vagotomy increased gastric pH levels, increased the atypical glandular hyperplasia level, and increased the incidence of adenocarcinomas (Tatsuta et al., 1985; Tatsuta, Iishi, Yamamura, Baba, & Taniguchi, 1988). In a rat study, the effect on tumors of vagotomy performed before carcinogen administration was unclear, but performing vagotomy after carcinogen administration did cause a significant increase in the number of gastric tumors (Tatsuta et al., 1985; Tatsuta et al., 1988). However, as in recent studies, vagotomy or treatment with Botox increased the effectiveness of chemotherapy while prolonging survival (Zhao et al., 2014).

Authors reported that vagal nerve denervation reduced the expression of Wnt signaling, a regulator of tumorigenesis (Zhao et al., 2014) (Fig. 2). They also showed that gastric tumorigenesis was suppressed by pharmacological inhibition and genetic knockout of the M3 receptor. In addition, Hayakawa et al. found that tuft cells secreted ACh, which is involved in early tumor growth and neuron formation around cancer cells. They showed that the positive-feedback loop of ACh and neurotrophic growth factor was responsible for innervation in the tumor microenvironment. The researchers further showed that tyrosine kinase is a primary receptor in this loop. In addition, they reported that muscarinic ACh receptor M3 (M3R) activates Wnt with Yes-associated protein (YAP), which is a transcriptional coactivator for Wnt/β-catenin signaling (Hayakawa et al., 2017). Another study showed that ACh, through the M3 receptor, increased gastric cancer cell proliferation and induced the phosphorylated ERK1/2 and AKT pathways. Moreover, the M3-selective antagonist 1,1-dimethyl-4-diphenylacetoxypiperidinium iodide (4-DAMP) reduced the phosphorylation of ERK and AKT. ACh also activated the EGFR signal via M3R while promoting cell proliferation, whereas blocking the EGFR signal reduced cell proliferation (Yu et al., 2017).

Fig. 2.

Schematic showing how parasympathetic nervous system activation reduces tumor growth and metastasis via decreased expression of PD-1 and PD-L1 while increasing release of IFN-γ from CD4⁺ and CD8⁺ tumor-infiltrating lymphocytes. As shown, denervation of the parasympathetic system or vagotomy decreases gastric tumorigenesis through Wnt signaling and causes a similar outcome in colorectal cancer cases. Increased cytokines, TNF-α, and TAMs; high invasiveness; and decreased survival rates are the results of vagotomy in patients with other cancers such as prostate, pancreatic, and breast cancer. The figure was created with BioRender.com.

Similar studies have shown that the mechanisms found in gastric cancers are also found in gastrointestinal system cancers (Belo et al., 2011; Cheng et al., 2008). Cheng et al. reported that ACh-dependent cell proliferation in colon cancers is mediated by M3R and includes an autocrine mechanism (Cheng et al., 2008). Binding of ACh to CHRM3 activates MMP-7 and increases cell proliferation, cell survival, and migration. This occurs via MAPK/ERK1/2 and PI3K/AKT pathways, which are regulated by epidermal growth factor receptor (Belo et al., 2011). Likewise, the use of scopolamine butylbromide, a muscarinic receptor antagonist, has reduced the numbers and sizes of tumors in small intestinal neoplasms (Raufman et al., 2011) (Fig. 3).

Fig. 3.

Schematic showing how acetylcholine and its antagonist affect cell apoptosis, cell proliferation, and tumorigenesis. Ach binding to muscarinic acetylcholine receptor M1 (M1R) induces EGFR-mediated tumorigenesis via MMPs and enhances cell proliferation. Muscarinic acetylcholine receptor M3 (M3R) promotes tumorigenesis through the ERK1/2 and AKT pathways. M3R increases cell proliferation by activating MMP7, which induces the ERK1/2 and PI3K/AKT pathways regulated by the EGFR signal. 4-DAMP, an M3R antagonist, promotes cell apoptosis via caspase-3 and Bax in mitochondria by inhibiting the ERK and AKT pathways. Tuft cells and nerves release ACh, upregulating NGF expression, which promotes cancer cell proliferation in gastric epithelium. The figure was created with BioRender.com.

Furthermore, investigators demonstrated that parasympathetic nerve fiber density increased the lymph node metastasis of colon cancers (Zhou et al., 2018). They showed that patients with parasympathetic nerve inhibition had a better prognosis. Also, the expression of α9 nicotinic ACh receptor (nAChR) was directly proportional to the number of parasympathetic nerves, with higher numbers found in those with lymph node metastases, and tumors with α9 nAChR were more advanced (Zhou et al., 2018). The presence of α9 nAChR has also been associated with increased metastasis with activation of vimentin and fibronectin in breast cancers cases (Lee et al., 2010). In addition, α7 nAChR has contributed to pancreatic and lung tumor development via the PI3K/AKT, nuclear factor-κB, and signal transducer and activator of transcription (STAT) pathways (Zhou et al., 2018).

4. Neurotransmitters

Neurotransmitters secreted from neurons and other cells in the tumor microenvironment link intratumoral nerves and cancer cells, thus affecting cancer invasion, metastasis, and progression. One of these neurotransmitters, glutamate, which plays a critical role in nutrition, metabolism, and signaling, is one of the most abundant amino acids (Brosnan & Brosnan, 2013). Within the central nervous system, glutamate is the major excitatory neurotransmitter, and its product, gamma-aminobutyric acid (GABA), is the major inhibitory neurotransmitter (Brosnan & Brosnan, 2013). Within cancer cells, glutamate is a potential growth factor and energy source for tumor progression (Reitzer, Wice, & Kennell, 1979; Stepulak, Rola, Polberg, & Ikonomidou, 2014; Yu, Wall, Wangari-Talbot, & Chen, 2017). Glutamate plays a role in the Krebs cycle and lipogenesis pathway, which are important sources of energy for cancer cells (Budczies et al., 2015; Koochekpour, 2013). Researchers have thought that glutamate can be used as a diagnostic biomarker because of its higher levels in cancer cells comparing with normal tissues (Budczies et al., 2015; Koochekpour, 2013). In a study of primary prostate cancer, serum glutamate levels have correlated directly with Gleason scores and tumor aggressiveness (Koochekpour, 2013). Decreased glutamate levels decrease prostate tumor growth, and invasion; and stimulate apoptosis of prostate cancer cells (Koochekpour, 2013; Koochekpour et al., 2012). Similarly, authors reported that glutamate levels were much higher in breast tumors than in normal breast tissue, whereas glutamine levels did not differ notably (Budczies et al., 2015). Based on these findings, a high glutamate/glutamine ratio, which was thought to be a positive prognostic factor, was associated with prolonged survival of breast cancer (Mohammadpour et al., 2019).

Glutamate receptors consist of two main groups: G-protein-coupled metabotropic glutamate receptors (mGluRs) and ionotropic glutamate receptors (iGluRs) (Stepulak et al., 2014). Recently, researchers found both receptors in peripheral tissues, such as the lung, breast, pancreas, bone, and skin, in addition to the central nervous system (Du, Li, & Li, 2016; Yi, Talmon, & Wang, 2019). Investigators detected increased or abnormal expression of mGluRs, which causes tumor growth via autocrine or paracrine signaling, in several cancers. Metabotropic glutamate receptor 1 can be oncogenic in epithelial cells by activating multiple cancer-signaling pathways, such as the MAPK and AKT pathways (Martino et al., 2013).

In a study of non-small cell carcinomas, Tamura et al. found aberrant expression of methylated N-methyl-d-aspartate receptor type 2B (NMDAR2B) to be an important prognostic factor. The presence of NMDAR2B has been shown associated with a better prognosis, especially for squamous cell carcinoma (Tamura et al., 2011). In a genetic study of esophageal squamous cell carcinoma, NMDAR2B was more methylated in tumors than in normal esophageal mucosa. Ifenprodil, a specific NMDAR2B inhibitor, blocks NMDAR-mediated apoptosis (Kim et al., 2006). In addition, GluN2B-mediated NMDAR signaling has been involved in metastasis of breast tumors to the brain and has been associated with poor prognosis (Zeng et al., 2019). Several studies showed that human T-cell leukemia and human T-cell lymphoma cell lines express AMPA GluR3, mGluR5, and mGluR1 (Ganor & Levite, 2014). Pharmacological inhibition of AMPA receptors in pancreatic adenocarcinoma cases reduces the invasion and migration of cancer cells and inhibits pancreatic tumor growth (Herner et al., 2011). In addition, activation of the AMPA receptor induces K-ras activity and the MAPK pathway (Herner et al., 2011).

GABA, which is primarily synthesized from glutamate, is a main inhibitory neurotransmitter in the central nervous system (Boonstra et al., 2015; Brzozowska, Burdan, Duma, Solski, & Mazurkiewicz, 2017). Similar to glutamate receptors, GABA exerts its effects on ionotropic (GABA-A or GABA-C) and metabotropic (GABA-B) GABA receptors (Brzozowska et al., 2017). GABA increases intracellular calcium levels and activates the MAPK/Erk cascade (Takehara et al., 2007). Induction of GABA-B receptor expression increases the migration of cancer cells by increasing the phosphorylation of ERK1/2 and expression of MMP-2 (Zhang et al., 2014). In a study of breast cancers, a high GABA level was directly proportional to a prolonged median overall survival duration(Brzozowska et al., 2017). GABA secreted from cancer cells increases beta-catenin signaling, which causes proliferation of tumor cells and suppression of intratumoral infiltration of CD8⁺T cells (Huang et al., 2022). Additionally, secretion of GABA vesicles from neuroendocrine prostate cancer cells was mediated by GABA type B receptor subunit 1 which was implicated to have a role in promotion of metastasis (Solorzano et al., 2018).

Dopamine is a monoamine catecholamine neurotransmitter that regulates tumor angiogenesis and vasculogenesis. In addition, it increases the effectiveness of anti-cancer drugs (Lan et al., 2017). Dopamine exerts its effects by using G-protein-coupled dopamine receptors (Lan et al., 2017). Dopamine inhibits tumor growth (Senogles, 2007), reduces vascular permeability and microvessel density (by decreasing VEGF receptor-2, focal adhesion kinase, and MAPK phosphorylation) (Sarkar et al., 2004; Sarkar, Chakroborty, Chowdhury, Dasgupta, & Basu, 2008), and reduces hypoxia-inducible factor-1α expression in cancer cases (Qin et al., 2015). When used with anticancer drugs, dopamine has prolonged life and reduced cancer progression (Sarkar et al., 2008). Dopamine inhibits insulin-like growth factor-1-induced gastric cancer cell proliferation via D2 receptors (Ganguly et al., 2010). In another study, dopamine inhibited tumor growth and neovascularization via D2 receptors by suppressing VEGFA-induced ERK1/ERK2 phosphorylation and MMP-9 synthesis (Chakroborty et al., 2008) (Table 1).

Table 1.

Neurotransmitters that promote tumor growth and dissemination.

Neurotransmitter	Receptors	Cancer Cell Types	References
Glutamate	G-protein coupled metabotropic glutamate receptors (mGluRs)	Melanoma	(Martino et al., 2013)
	Ionotropic glutamate receptors (iGluRs)	Non-small cell lung carcinoma Esophageal squamous cell carcinoma Breast cancer cell-brain metastasis Human T-leukemia and T-lymphoma Pancreatic adenocarcinoma	(Tamura et al., 2011) (Kim et al., 2006) (Zeng et al., 2019) (Ganor & Levite, 2014) (Herner et al., 2011)
GABA	Metabotropic receptor (GABA-B)	Breast cancer Colorectal cancer Prostate cancer	(Zhang et al., 2014) (Huang et al., 2022) (Solorzano et al., 2018)
	Ionotropic receptors (GABA-A or GABA-C)	Pancreatic ductal adenocarcinoma	(Takehara et al., 2007)
Dopamine	G-protein-coupled dopamine receptors (DRs)	Malignant glioma Small cell lung cancer Breast and colon cancer Glioma Gastric cancer Sarcoma	(Lan et al., 2017) (Senogles, 2007) (Sarkar et al., 2004) (Qin et al., 2015) (Ganguly et al., 2010) (Chakroborty et al., 2008)

5. Cancer-derived mediators

Besides cancer cells being affected by neural regulation in the tumor microenvironment, mediators released from cancer cells, for instance, extracellular vesicles (EVs), play a critical role in tumor progression and metastasis (Becker et al., 2016). These vesicles have unique compositions and carry distinct molecules, including proteins like transcription factors, enzymes, and receptors; nucleic acids like RNA, DNA, and microRNA (miRNA); and lipids (Raposo & Stoorvogel, 2013; Valadi et al., 2007). Authors reported that mRNA and tetraspanin Tspan8 proteins in tumor-derived exosomes increased angiogenesis in pancreatic tumors (Nazarenko et al., 2010). Hypoxic exosomes that carry highly expressed miR-301a-3p, which activates the PTEN/PI3K pathway and induces M2 polarization of macrophages, have been associated with poor prognosis for pancreatic cancer (Wang et al., 2018). In breast cancer studies, tumor-derived exosomes caused increased cancer cell migration and metastasis (Harris et al., 2015). In addition, an abundance of miRNAs was associated with increased tumorigenesis, invasiveness, and enhanced therapy resistance, and thus was a negative prognostic marker in breast cancer cases (Jia et al., 2017).

In addition to these findings, tumor-derived EVs, particularly exosomes, induced neurite outgrowth in a study using PC12 cells (rat pheochromocytoma cell line) and exosomes derived from head and neck cancer patients. In that study, tumor-derived exosomes with high levels of EphrinB1, an axon-guidance molecule, promoted neuritogenic effects and tumor innervation (Madeo et al., 2018). Consistently, stimulated neuritogenesis also occurred in PC12 cells treated with cervical cancer-derived exosomes (Lucido et al., 2019). Moreover, we showed that miRNAs in EVs regulate neuritogenesis, which was triggered by loss of the miR-34a or the presence of the miR-21 and miR-324 (Amit et al., 2020). These findings show that tumor-derived EVs regulate tumor innervation and progression for different cancer types.

Another group of mediators released from cancer cells, neurotrophins, also contribute to neurogenesis and tumor innervation (Wang et al., 2014). In pancreatic cancer, increased neurturin (NRTN) was positively correlated with an aggressive phenotype and invasiveness, and induced neuroplasticity. In addition, tumor-derived neurotrophic growth factor (NGF) and brain-derived neurotrophic factor have promoted tumor progression by regulating PI3K/AKT and Ras/ERK signaling pathways and axonogenesis via Trk receptors, which drive cancer modulation (Cervantes-Villagrana, Albores-Garcia, Cervantes-Villagrana, & Garcia-Acevez, 2020). Taken together, these results demonstrate that cancer-released mediators like neurotrophins and EVs are promising factors for therapeutic approaches for cancer and should be investigated further.

6. Neuroimmune interactions

Sympathetic nervous system activation enhances metastasis of cancer cells by stimulating macrophage infiltration, inflammation, angiogenesis, epithelial-mesenchymal transition, and tumor invasion and by inhibiting the immune response and apoptosis (Cole et al., 2015).

IL-6, a pro-angiogenic cytokine, plays a role in inflammation, chronic stress, and cancer development. It prevents cancer cell apoptosis via many cellular mechanisms, such as the ERK/MAPK, PI3-K, and JAK/STAT pathways (Nguyen, Li, & Tewari, 2014; Nilsson et al., 2007). Researchers showed that sympathetic nerves induce IL-6 gene expression with β-adrenergic activation or inhibition of GATA1 by using norepinephrine agonists and antagonists, respectively (Cole et al., 2010). Also, β-adrenergic stimulation and norepinephrine increased IL-6 expression via Src, a proto-oncogene product, and the tyrosine kinase signaling pathway in ovarian cancer cells (Nilsson et al., 2007). Furthermore, stimulation of norepinephrine in gastric cancers caused an increase in IL-6 expression via the cAMP/PKA signaling pathway (Yang, Lin, Gao, & Zhang, 2014).

Investigators showed that stress increased metastasis of breast cancer in both in vivo and ex vivo studies (Sloan et al., 2010). Administration of β-AR antagonists had no effect on primary tumor growth or metastatic burden in the control mouse group (non-stressed) but blocked metastasis in the chronically stressed mouse group. Also, stress increased the number of macrophages in the tumor microenvironment and the expression of the Tgfb, Arg1, Cox2, Mmp9, Vegf, and Vcam1 genes while decreasing expression of the Ifnb gene (Sloan et al., 2010). Consistent with these findings, β-adrenergic signals enhanced infiltration of CD14⁺ CD68⁺ cells and macrophages in the tumor microenvironment by increasing production of macrophages and monocyte chemoattractant protein 1 (MCP1) (Armaiz-Pena et al., 2015).

β-adrenergic signaling inhibits DNA damage repair mechanisms and prevents p53-associated apoptosis. In a study using a neuroblastoma line, propranolol upregulated p53 and caused apoptosis; in addition, it made cancer cells sensitive to the topoisomerase inhibitor SN-38 (Wolter et al., 2014). Inhibition of cytotoxic T cells and NK cells is believed to play a role in postoperative metastasis of cancer (Goldfarb et al., 2011; Inbar et al., 2011) and has been associated with increased dissemination of these tumor cells (Goldfarb et al., 2011). β-AR activation has suppressed natural killer activity and cytotoxic T cells in vivo (Inbar et al., 2011). In addition, chronic stress enhanced cancer growth via MDSCs and increased the accumulation of MDSCs in the tumor microenvironment. Furthermore, β2-AR stimulation caused phosphorylation of STAT3, which is involved in the immunosuppressive function of MDSCs, leading to increased numbers of MDSCs in tumors (Mohammadpour et al., 2019) (Fig. 4).

Fig. 4.

Schematic showing how activators and blockers of the sympathetic nervous system affect tumor and immune mediators. Propranolol promotes cell apoptosis by upregulating p53. Norepinephrine increases tumor cell proliferation by inducing expression of IL-6, which is regulated by the cAMP/PKA signaling pathway. Chronic stress promotes tumor growth via phosphorylated STAT3, which induces the numbers of MDSCs. β-adrenergic signals enhance the numbers of CD14⁺ CD68⁺ TAMs and macrophages via increased production of MCP1. Chronic stress and norepinephrine inhibit DNA damage repair mechanisms and prevent p53-associated cell apoptosis. The figure was created with BioRender.com.

Neurons can exert their effects on cancer through the inflammatory reflex. Specifically, they can regulate the level of IL-1β with the receptors in the afferent fibers of the vagus nerve. In response, the vagus nerve releases ACh on macrophages via α7 nAChR, thus inhibiting the release of proinflammatory cytokines and inhibiting high mobility group box protein 1 (HMGB1) secretion (Cortese, Rigamonti, Mantovani, & Marchesi, 2020).

Mediators released from tumors, EVs, not only regulate neurogenesis and cancer progression but also modulate immune cells in the tumor microenvironment. Tumor-infiltrating immune cells, regulatory B cells (Ye et al., 2018), regulatory T (Treg) cells (Wieckowski et al., 2009), TAMs (Chen et al., 2018), neutrophils, and MDSCs (Valenti et al., 2006) were modulated by tumor-derived EVs. Investigators showed that microvesicles isolated from melanoma patients caused immunosuppression by inhibiting monocyte differentiation and by generating MDSCs with tumor growth factory-β-mediated suppressive activity on T-cells (Valenti et al., 2006). Also, EVs released from melanoma cells have induced CD8⁺ T cells apoptosis and promoted Treg cell expansion thus contributing to immune suppression (Wieckowski et al., 2009). In one important study in the field, exosomal PD-L1 suppressed immunity and facilitated immune evasion in patients with metastatic melanoma (Chen et al., 2018). Additionally, exosomal HMGB1 expression led to immune escape via regulatory B cells expansion in hepatocellular carcinoma (Ye et al., 2018). In view of the interaction between cancer cells and immune cells through EVs within the tumor microenvironment, future studies will reveal unexplored areas of cancer.

7. Conclusions

The mechanisms of neural activity that have facilitative effects on cancer metastasis and cause poor patient outcomes are not completely understood. With an emphasis on developments in neuroimmunology, recent studies of tumor proliferation and dissemination have focused on nerves, immune cells, and interaction with cancer. Those studying the tumor microenvironment have an ongoing interest in understanding how tumors interact with other mediators such as nerves, chemokines, immune cells, and signaling molecules. Although these novel approaches highlight many undiscovered mechanisms, neuroimmune interplay in tumor dissemination is a substantial research interest in the field. In this review, we discuss sympathetic and parasympathetic regulation of different cancers, neurotransmitters, tumor-derived mediators, and pathways in the tumor. The effects of the peripheral nervous system on cancer cells and mediators released from tumor cells, such as EVs and neurotrophins, are innovative therapeutic targets for cancer. A comprehensive understanding of cancer and neuroimmune crosstalk will reveal promising treatment opportunities such as overcoming immunotherapy resistance in cancer patients.

Abbreviations:

ACh

acetylcholine

AR

adrenergic receptors

β2-AR

beta 2 adrenergic receptors

β3-AR

beta 3 adrenergic receptors

cAMP

cyclic adenosine monophosphate

Chrm1

cholinergic receptor muscarinic type 1 receptor

COX

cyclooxygenase

DUSP1

dual specificity phosphatase 1

EGFR

epidermal growth factor receptor

ERK

extracellular signal-regulated kinase

EVs

extracellular vesicles

GABA

Gamma aminobutyric acid

HMGB1

high mobility group box protein 1

iGluR

ionotrophic glutamate receptors

JNK

c-Jun N-terminal kinase

MAPK

mitogen-activated protein kinase

MCP1

monocyte chemoattractant protein 1

MDSCs

myeloid-derived suppressor cells

mGluR

metabotropic glutamate receptor

miRNA

microRNA

MMP

matrix metalloproteinase

nAChR

nicotinic acetylcholine receptor

NGF

neurotrophic growth factor

NK

natural killer

NMDAR2B

methylated N-methyl-d-aspartate receptor type 2B

PD-L1

programmed death ligand 1

PI3K

phosphatidylinositol-3-kinase

PKA

protein kinase A

STAT

signal transducer and activator of transcription

TAM

tumor-associated macrophage

TNF-α

tumor necrosis factor α

Treg

T regulatory cell

VEGF

vascular endothelial growth factor

4-DAMP

1,1-dimethyl-4-diphenylacetoxypiperidinium iodide