Introduction

During each step of the metastatic cascade, the highly immunogenic tumor cells should be recognized and eliminated by the host immune system

^[1][2][3]

. In general,

the antigen-presenting cells (APCs) such as dendritic cells (DCs) phagocytose tumor antigens, and the fragmented antigens are generally known to be

presented by major histocompatibility complex (MHC) class I and II to cytotoxic T lymphocytes (CTLs) and cluster of differentiation (CD)4+ T cells, respectively. Subsequently, the activated T cells should infiltrate into tumors in order to kill the cancer cells

^[4][5]

. 

Cellular adhesion molecules such as integrins and the immunoglobulin superfamily may initiate T cell adhesion to APCs

^[6].

Particularly, integrin may mediate the infiltration of immune cells to the tumor site, the activation and proliferation of effector cells, and the formation of immunological synapse between immune cells and tumor cells

. Particularly, integrin may mediate the infiltration of immune cells to the tumor site, the activation and proliferation of effector cells, and the formation of immunological synapse between immune cells and tumor cells

^[6]

. The leukocyte function-associated antigen-1 (LFA-1) is a member of the β2 integrin family of adhesion receptor. It is also the predominant integrin in lymphocytes. It is an important component in T cell trafficking and extravasation

^[7]

by binding to its ligand, the intercellular adhesion molecules-1 (ICAM-1)

^[6]

. The LFA-1/ICAM-1 activates the formation of the immunological synapse via cell–cell interactions between immune effector cells and tumor cells (Figure 1). 

Figure 1: The role of LFA-1/ICAM-1 in T cell activation and tumor eradication^[8]

The small GTPase Ras-associated protein 1 (Rap1) is one of the main components of inside-out signaling cascades that plays a critical role in activating integrin

The small GTPase Ras-associated protein 1 (Rap1) is one of the main components of inside-out signaling cascades that plays a critical role in activating integrin

^[6][7]

. Indeed, Rap1 signaling has been shown to play a role as a potent immunological regulator in modulating LFA-1-mediated adhesion in Jurkat T cells

^[9]

. Activation of Rap1 in the T cells rapidly increased LFA-1 affinity, while overexpression of dominant-negative form of Rap1 (Rap1N17) or signal-induced proliferation-associated gene-1 (SPA-1) completely inhibited LFA-1 activation

^[10]

and reduced adhesiveness and interleukin (IL)-2 production in Jurkat T cells

^[9]

. Conversely, overexpression of Rap1 enhanced T cell activation and augmented IL-2 production

^[9]

. In addition, in vivo study showed that increased active Rap1 in T lymphocytes was associated with enhanced T cell receptor (TCR)-mediated response and increased integrin activation

^[11]

. Hence, these findings clearly indicate that Rap1 plays a crucial role in mediating integrin activation in T cells via inside-out signaling cascades

Biology of Rap1

Rap1 is a member of the Ras-like small GTP binding protein family. Rap1 can bind to either guanosine triphosphate (GTP) or guanosine diphosphate (GDP). Rap1 activity is positively regulated by guanine nucleotide exchange factors (GEFs) and negatively regulated by GTPase-activating proteins (GAPs)

^[12]

. GEFs promote dissociation of GDP, allowing loading of GTP to Rap1, resulting in the active form of GTP-bound Rap1 (Rap1-GTP). Conversely, Rap1-GTP is effectively hydrolyzed into inactive GDP-bound Rap1 (Rap1-GDP) in the presence of GAPs

^[12]

. The well-characterized GEF family includes C3G (RapGEF1), RasGRP2, PDZ-GEF, cAMP and Epac

^[13]

. In contrast to the diverse types of GEFs, there are only two groups of GAPs, Rap1GAP and SPA-1

^[14]

. The regulation of the biological activity of Rap1 is summarized in Figure 2.

Figure 2: Regulation of Rap1 activity^[8]

Rap1 Activation and Cancer

Rap1 Activation and Cancer

Rap1 signalling plays a diverse role in tumor initiation and progression (Figure 3). For instance, Rap1 activation induces tumor initiation and epithelial-mesenchymal transition (EMT) via Notch signaling

Rap1 signalling plays a diverse role in tumor initiation and progression (Figure 3). For instance, Rap1 activation induces tumor initiation and epithelial-mesenchymal transition (EMT) via Notch signaling

^[8]

. EGFR and Src/FAK may be stimulated by the activated Rap1, leading to integrin-mediated cell adhesion in cancer. The adhesion of integrins to the extracellular matrix (ECM) is an essential step for tumor cell invasion and metastasis

^[8]

. Src may activate MAPK/ERK to increase VEGF expression levels in the tumor cells, ultimately leading to angiogenesis. Src may also induce p130Cas and PD-L1 expression in the tumor cells

^[8]

. The upregulation of MMP during Rap1 activation may trigger EMT in cancer and thus enhancing invasiveness and metastasis

^[8]

. Therefore, Rap1 has rendered itself to be an attractive therapeutic target in cancer. However, it remains uncertain whether Rap1 signaling will positively or negatively regulate tumor progression as it showed to demonstrate contrasting effects on a tumor (Table 1).  Therefore, further research is required to delineate its roles in cancers.

Figure 3: Rap1 activation contributes to the acquisition of cancer hallmarks^[8]

Table 1. Roles of Rap1 in tumor progression^[8].

Rap1 and Integrin

Rap1 and Integrin

Integrin, the main cell adhesion receptor for the extracellular matrix (ECM), exists in different conformational states, and thus

leads to the difference in its receptor affinity. An inactivated integrin (bent and closed) is associated with a low affinity for ECM ligand, whereas the active form (open) integrin is capable of binding to the ligand and clustering on the plasma membrane (adhesion)

Integrin, the main cell adhesion receptor for the extracellular matrix (ECM), exists in different conformational states, and thus leads to the difference in its receptor affinity. An inactivated integrin (bent and closed) is associated with a low affinity for ECM ligand, whereas the active form (open) integrin is capable of binding to the ligand and clustering on the plasma membrane (adhesion)

^[8]

. Rap1 mediates integrin-dependent biological processes in cancers, by which integrin adhesion activates signaling pathway such as Src/FAK to promote cytoskeletal remodeling, tumor migration, invasion, and metastasis (Figure 4)

^[8]

. Integrin activation can be inhibited by Rap1GAP directly or indirectly

^[8]

. 

Figure 4: Rap1 regulates integrin-dependent biological processes in cancers^[8]

Rap1 and Cadherin

Rap1 and Cadherin

Rap 1 signalling plays important role in regulating cadherin expression (Figure 5). Rap1 activation leads to the reduction in E-cadherin expression, resulting in the loss of epithelial cell–cell adhesive junctions and contact inhibition of proliferation

Rap 1 signalling plays important role in regulating cadherin expression (Figure 5). Rap1 activation leads to the reduction in E-cadherin expression, resulting in the loss of epithelial cell–cell adhesive junctions and contact inhibition of proliferation

^[8]

. During the restoration of Rap1GAP expression in tumor cells, Rap1 will be inactivated and thus inhibiting tumor progression

^[8]

.

Figure 5: Rap1 promotes tumor progression via disruption of E-cadherin-mediated cell adhesion^[8]

Rap1 Signaling Promotes Angiogenesis, Proliferation, and Survival

Rap1 Signaling Promotes Angiogenesis, Proliferation, and Survival

Rap1 signaling has been implicated in the regulation of angiogenesis, cell proliferation and migration in different cancers. In order to induce angiogenesis, VEGF increased Rap1 activity in an integrin-dependent manner. The use of β1 integrin antibody was shown to inhibit constitutive active Rap1-induced angiogenesis in endothelial cells

Rap1 signaling has been implicated in the regulation of angiogenesis, cell proliferation and migration in different cancers. In order to induce angiogenesis, VEGF increased Rap1 activity in an integrin-dependent manner. The use of β1 integrin antibody was shown to inhibit constitutive active Rap1-induced angiogenesis in endothelial cells

^[23]

. Retroviral silencing of Rap1GAP greatly enhanced ERK/MAPK phosphorylation and activation, induced extensive cytoskeletal reorganization and expression of EMT features, and enhanced invasion in breast ductal carcinoma in situ (DCIS) cells

^[24]

. Enforced re-expression of Rap1GAP in DCIS cells resulted in decreased Rap1 activation and a reversed mesenchymal to epithelial phenotype

^[24]

.

Therapeutic Potential of Rap1 Signaling in Cancers

Therapeutic Potential of Rap1 Signaling in Cancers

Clinical studies have demonstrated that the loss of Rap1GAP expression was correlated with negative clinicopathological outcomes, such as invasion depth, lymph node metastasis, advanced TNM stages and poor overall survival (OS) as determined by Kaplan–Meier survival analyses

Clinical studies have demonstrated that the loss of Rap1GAP expression was correlated with negative clinicopathological outcomes, such as invasion depth, lymph node metastasis, advanced TNM stages and poor overall survival (OS) as determined by Kaplan–Meier survival analyses

^[25][26]

. Rescued expression of Rap1GAP in renal cell carcinoma cell lines using a demethylating drug, decitabine (5-azadC), resulted in a significant reduction in tumor invasiveness

^[27]

. In pancreatic cancer, the use of a novel Epac-specific inhibitor (ESI-09), specifically inhibited Epac1-mediated Rap1 activation, ultimately resulting in impaired migration and invasion capability of cancer cells

^[28]

. Taking all these into account, Rap1GAP holds the high potential to serve as a novel target for therapeutic intervention.