Fibroblast activation protein-α (FAP) is an inducible cell surface glycoprotein originally identified in 1986 in cultured fibroblasts using the monoclonal antibody (mAb) F19. It is a type ii integral serine protease that is specifically expressed by activated fibroblast.
Cancer-associated fibroblasts (CAFs) in the tumor stroma have an abundant and stable expression of FAP, which plays an important role in promoting tumor growth, invasion, metastasis, and immunosupression. For example, in females with a high incidence of breast cancer, CAFs account for 50-70% of the cells in the tumor’s microenvironment.
CAF over expression of FAP promotes tumor development and metastasis by extracellular matrix remodeling, intracellular signaling, angiogenesis, epithelia-to-mesenchymal transition, and immunosuppression.
The structure of Fibroblast activation protein
Fibroblast activation protein-α (FAP) is a type ii transmembrane glycoprotein consisting of 760 amino acids. It belongs to the family of post-proline dipeptidyl aminopeptidase, known as dipeptidyl peptidase-4 activity (DPP4). At the genetic level, human FAP and dipeptidyl peptidase-4 (DPP4) genes share substantial homology. Human FAP gene is located on chromosome 2q23 and contain 26 exons (total length: 73kb) while DPP4 is located on chromosome 2q24.3 and contains 26 exons (total length: 70kb)
The FAP protein is a 170-kda homodimer with two N-terminal glycosylated subunits. The 97-kda type ii transmembrane serine protease is a member of the polylpeptidase family, which includes DPP-4 (most similar to FAP) DPP7, DPP8, DPP9 and prolyl carboxypeptidase. The FAP and DPP-4 proteins have a 70% identity at the amino acid sequence and share a catalytic triad of serine, aspartic acid, and histidine residues.
Serine plays a nucleophilic role, which allows DPP-4 to cleave N-terminal Pro-“X” peptide bond (where “X” is any amino acid except proline or hydroxylproline). Unlike FAP, DPP-4 is expressed in a variety of human tissues under normal conditions and is related to many physiological processes, including glucose homeostasis and T-cell activation.
Enzymatic activity of Fibroblast activation protein-α
The FAP protein has dipeptidyl peptidase and endopeptidase activities, which are sometimes described as gelatinase activity. Although both FAP and DPP-4 have dipeptidase activity, the unique endopeptidase activity of FAP makes it preferentially cleave to the Gly-pro”X” sequence, with the most effective cleavage when “X” is Phe or Met and the least effective one when “X” is His or Glu.
The cleavage activity of FAP can be impaired when P4 and P2 residues are heavily charged amino acids. Therefore, endopeptidase activity can be used to specifically detect FAP and are the basis of nanomaterial treatments that aim to specifically inhibit FAP.
Studies have shown that DPP-4 can cleave neuropeptide Y, peptide YY, Sp (substance P), and brain natriuretic peptide 32, which can also be cleaved by FAP. The known active substrates of endopeptidase include collagens I, ii, and v, as well as α-2-antiplasmin and fibroblast growth factor 2. The ability of FAP o cleave collagen depends on previous matrix metalloproteinase activity o thermal degradation.
Soluble FAP is known as α-2-antiplasmin cleaving enzymes (APCE), which has pro-coagulation poperties. After FAP cleaves α-2-antiplasmin, it is converted into a more effective plasmin inhibitor, which slows the dissolution of the fibrin clot and reduces bleeding during tissue repair.
Expression of Fibroblast activation-α in Tumors
Fibroblast activation protein-α level is generally undetectable in normal tissues. However, FAP is over expressed in many tumor tissues, including breast, pancreatic, lung, colorectal, brain, intrahepatic bile duct, and ovarian cancers. High level of FAP can also be detected in some tumors that are derived from non-epithelial tissues, such mas melanoma and myeloma. In these tumors, FAP over expression is typically observed in the intestitium, which has led to FAP being considered a universal marker for CAFs.
Roles of fibroblast activation protein-α in tumors
Fibroblast activation protein-α expression influences tumor growth by impacting tumor cell proliferation and invasion, angiogenesis, epithelia-to-mesenchymal transition, immunosuppression and drug resistance.
Fibroblast activation protein-α promotion of Tumor cell proliferation and invasion
Fibroblast activation protein promotes tumor cell proliferation, migration and invasion, which ultimately leads to tumor growth. There are two hypothesis regarding the underlying mechanisms. The first hypothesis involves an indirect mechanism, whereby FAP regulates extracellular matrix remodeling that leads to enhanced tumor growth and invasion. It remains unclear whether FAP regulates this extracellular matrix remodeling through its enzymatic or non-enzymatic activity.
The second hypothesis involves a direct mechanism, in which FAP expression influences signaling pathways that control the cell cycle and proliferation, which ultimately promote tumor growth.
In the indirect hypothesis, transfection of small interfering RNA targeting FAP inhibits the proliferation of ovarian CAFs, leading to cell cycle arrest. In squamous lung cancer cell, FAP overexpresion promotes proliferation, migration and invasion, accompanied by upregulaton of the PI3k/protein kinase B and sonic hedgehog/glioma-associated oncogene signaling pathways. Other studies on oral squamous cell carcinoma have indicated that FAP is an upstream regulation of phosphatase and tensin homolog, PI3k/protein kinase B and Ras-ERK signaling pathways.
Fibroblast activation protein as a tumor suppressor
Studies suggest that FAP has tumor suppressive activity and show that this activity is independent of its enzymatic activities. Elevated expression of Fibroblast activation protein in cancer causes dramatic promotion or suppression of tumor growth, depending on the model system investigated. There is however obvious discrepancy between FAP function in tumor promotion and tumor suppression. Some researchers propose that the factor that determines this must reside in the signaling molecules that are available for interaction with FAP on the cell.