Claudin-4 and claudin-18 are highly and differentially expressed in PCa (37). Claudin-4 contributes to the function of anion channels/pores, the transepithelial resistance (TER) was enhanced in a claudin-specific manner. Claudin-4 is highly expressed in pancreaticobiliary ductal PCa (p = 0.015), PanIN, IPMN, MCN, and the major precursor lesions of pancreatic ductal adenocarcinoma (73), and its expression is correlated with the histological tumor grade of IPMN and MCN (11, 23, 34, 57). Similar to claudin-18.2, claudin-4 is also highly positive in liver metastases from PCa (74).

Patrick Michl et al. demonstrated that claudin-4 is a potent inhibitor of the aggressive and metastatic phenotype of PCa cells and a target of transforming growth factor (TGF)-β and the Ras/Raf/Erk pathway. Furthermore, proinvasive transforming factor-β downregulates claudin-4 in PANC-1 cells. Specific inhibitors of dominant negative Ras and downstream effectors of mitogen-activated protein/Erk and phosphatidylinositol 3’-kinase inhibit Ras signaling, thereby decreasing claudin-4 expression and in GC, which further increases PI3K and Akt phosphorylation, thus promoting tumor cell proliferation, migration, invasion, and tumorigenesis, a mechanism that may be similar in PCa (38, 39, 75). In studies of gastric cancer, claudin-4 expression was found to be associated with bivalent histone modifications, and decreased inhibitory histone methylation as well as increased active histone methylation was associated with claudin-4 overexpression in gastric cancer cells. Histone methylation and acetylation are required for claudin-4 alterations. The high expression of claudin-4 in pancreatic cancer needs to be further investigated and validated (76).

In the absence of TGF-β, DEC1 siRNA upregulated claudin-4, claudin-7, and E-cadherin expression and downregulated N-cadherin expression, consequently upregulating claudin-1 and claudin-4 (77). Tsutsumi et al. demonstrated that increased CLDN4 mRNA expression was significantly associated with improved prognosis of PDAC patients and that reduced claudin-4 expression in endothelial cells reduced paracellular resistance and promoted the invasion of epithelial cancer cells through the endothelial cell layer, similar to the mechanism of E-cadherin (78). Histopathological examination revealed that the incidence of lymphovascular (ly), vascular (v), and neural (ne) invasion was higher in the low claudin-4 expression group than in the high claudin-4 protein expression group (P = 0.0014, 0.0133, and 0.0205, respectively) and overexpression of epithelial claudin-4 reduced the invasive and metastatic potential of the PCa cells in vitro (75). Research have shown that, median survival of patients with high claudin-4 expression was 63.0 months, compared with 14.7 months for patients with low claudin-4 expression (P = 0.0067). Immunohistochemistry (IHC) analysis revealed that CLDN4 mRNA expression was significantly correlated with claudin-4 protein expression (P = 0.0168), indicating that increased expression of CLDN4 mRNA can predict a better prognosis of PCa (27). Furthermore, claudin-4 is negatively expressed in reactive mesothelial cells and mesothelioma, differentiated from PCa, while it is significantly positively expressed in plasma membrane metastases and primary carcinoma of PCa. Therefore, claudin-4 can be used as an IHC reagent to exclude mesothelioma diagnosis (79). These results suggest that claudin-4 has great potential as a novel diagnostic biomarker for PCa (80).

Endoscopic ultrasound-guided fine-needle aspiration is a specific and effective marker for the differential diagnosis of pancreatic ductal carcinoma from gastrointestinal contamination and benign pancreatic ductal epithelium (81). claudin-4 co-localization in vitro and in vivo provides a method for early detection imaging diagnostic method for PCa (82). Claudin-4 also has great potential for targeted delivery and imaging, designed to label PCa using quantum dots (QD), as a sensitive optical contrast agent, and to image PCa by using anti-CLDN4 as a targeting ligand. Mintai P et al. developed a carrier protein-based PCa detection platform where any antibody and Ni-NTA-functionalized nanoparticles can bind the target protein via protein G and 6*His-tag, respectively; the carriers are modified by the desferrin gene to form a unique probe, which diagnoses PCa cells by detecting claudin-4 (83). The 18F-labeled CLDN-selective peptide, prepared by linking 5-(18F) fluoro-5-deoxyribose (5-(18F) FDR) to the oxime of the CLDN-selective peptide, provides a new imaging tool for PET-CT diagnosis of PCa. One of the 5-(18F) FDR clones27 with a molar activity of 4.0 GBq/µmol (for a 30 MBq tracer) was isolated at 98% radiochemical purity and 15% radiochemical yield (EOB) in 98 min. Therefore, these tracers contribute to the early diagnosis of pancreatic tumors (84). Thus, the specific expression of claudin-4 in PCa makes it a potential diagnostic target, which can be explored further.

CPE causes diarrhea and causes fluid accumulation in the intestines by altering the permeability of the intestinal epithelial membrane (85). It has an N-terminal cytotoxic region and a C-terminal receptor-binding region, and claudin-4 is sensitive to CPE-mediated cytolysis (86). mAbs can bind to ECL2 of the claudin-4 extracellular structure, which serves as a therapeutic target for tumors. Since CPE is a high-affinity receptor for this ring and causes cytotoxic, the C-CPE (receptor binding domain) was extracted and used in drug delivery and selective therapy of claudin-4-positive tumors (87–89).

A related study utilized translationally optimized CPE vectors (optCPE) for novel suicide pathways in vitro and in vivo, using cell line-derived and patient-derived PCa xenografts (CDX and PDX, respectively). This study demonstrated that apoptosis/necrosis signaling is activated in vitro, while necrosis and tumor cell killing is activated in vivo after implantation of the optCPE gene into PCa cells, which is mediated by pore formation. It was selectively toxic to claudin-3/4 overexpressing PCa cells. Meanwhile, downregulation of claudin-4 expression prevented CPE cytotoxicity (90), and optCPE non-viral in vivo intratumoral injection gene therapy showed targeted anti-tumor effects in different CDX and PDX PCa models, leading to reduced tumor cell viability and induction of tumor necrosis. optCPE treatment in combination with chemotherapy can further enhance tumor necrosis and CPE gene transfer. This approach is superior to other toxins used in cancer therapy because they act on more common non-tumor-specific targets (91). This combination therapy selectively acts on CLDN3/4 overexpressing tumors, in vitro and in vivo, thus improving PCa prognosis and contributing to the local control of PCa, especially in unresectable or refractory PCa (92). CPE is more cytotoxic to hypofractionated PCa than to highly differentiated PCa, and it is closely associated with the expression and localization of claudin-4 and barrier function (93).

The binding domain of claudin-4 and fusion can identify the abnormal localization of claudin in malignant tumors (86), and can be developed as a target for tumor treatment by using the fusion of cCPE and protein synthesis inhibitor (C-CPE-PSIF).The anti-CLDN18.2 mAb (5D12) binds to the second extracellular loop of claudin-4 and human-rat chimeric IgG1 of 5D12 activates the Fc gamma receptor IIIa, thus activating ADCC in CLDN4-expressing cells. It was found that anti-CLDN-4 mAbs significantly inhibited tumor growth in Dutch human CRC and GC mice without significant adverse effects, such as weight loss or liver and kidney damage, in PCa. Although human in vivo experimental effects remain to be proven in further experiments, the anti-propensity therapy of anti-CLDN-4 mAbs against claudin-4 targets is a promising treatment for PCa (94). Suzuki et al. isolated KM3900 (IgG2a), which specifically binds to claudin-4, from BXSB mice immunized with PCa cells. KM3900 recognizes and binds to the ECL2 of claudin-4 4, which can be detected in CLDN4-expressing PCa and OC cells. Mouse-human chimeric IgG1 (KM3934) induced dose-dependent ADCC and CDC in vitro and significantly inhibited tumor growth in ovarian cancer cells(MCAS or CFPAC-1) xenograft SCID mice in vivo ( 87). CLDN4 serves to restrain pro-oncogenic signaling from EphA2 by limiting the activity of E-catenin and PI3K and preventing phosphorylation of EphA2 on S897 by AKT. This suggest that increases CLDN4 level and functional activity can inhibit the progression of cancer (95). These results suggest that claudin-4 is an effective target for cancer therapy and that anti-CLDN-4 targeting antibodies are promising therapeutic candidates.

Bioinformatics analysis suggests that claudin-4 may be regulated by FOX3 or USF2 and plays an important role in acute pancreatitis (96), and the variation in claudin-4 expression in PCa and acute pancreatitis needs to be further investigated.

Claudin-1 is regulated by a variety of transcription factors, growth factors, and cytokines under normal physiological conditions, which maintains its tightly linked gating function (28). In pancreatic cancer, the expression of claudin-1 is increased, especially in metastatic tissues, there is a nuclear cytoplasmic membrane localization error. TNF-a upregulates the expression of claudin-1 in pancreatic cancer by concentration dependence, leading to accelerated tumor cell proliferation (28, 97).The region surrounding the CpG island of the CLDN1 promoter controls transcription through methylation to alter epigenetic regulation. Furthermore, claudin-1 expression was negatively correlated with DNA methylation in short- and long-term survivor (GSE51820) (21) data analysis (98). ZEBI-regulated ZIP4 overexpression in PCa cells decreased the expression of ZO-1 and claudin-1, leading to phosphorylation of FAK and Paxillin, thereby promoting PCa invasion and migration (29). In contrast, ZEP4 knockdown increases claudin-1 expression, thereby reducing cancer cell invasion and metastasis. A study by Gurbuz showed that targeting 5-HT1B and 5-HT1D receptors downregulated ZEB1 and Snail family proteins while upregulating claudin-1 (30, 31). miR-193b expression resulted in increased expression of E-Cadherin and claudin-1 and decreased expression of Snail and TCF8/ZEB1 via the eEF2K and MAPK/Erk axes. Furthermore, miR-193b expression inhibited PDAC cell proliferation, migration, invasion, and EMT by suppressing the eEF2K/MAPK-Erk oncogenic axis (32). Additionally, the expression of claudin-1 gradually increases with the recovery of the disease while exhibit while exhibit opposite expression trend of cell-free DNA, in the mouse model of ulcerative colitis, the relationship between the content of the two is promising in the study of disease diagnosis indicators (99). In the future, targeted molecules related to claudin-1 can be prepared in a similar way to claudin-4 targeted drugs, and are expected to enter clinical practice (28).

Claudin- 5 is widely distributed in the vascular endothelium, where it is associated with adhesion between arterial endothelial cells. The second extracellular structural domain of claudin-5 is capable of dimerizing in vitro ( 100). Additionally, claudin- 5 is the most abundant TJ protein in the human blood-brain barrier and plays an important role in maintaining its integrity. Consequently, its dysfunction is associated with neurodegenerative diseases (Alzheimer’s disease), neuroinflammatory diseases (multiple sclerosis), and psychiatric disorders (depression and schizophrenia) (101), and the symptoms of these diseases can be eliminated to an extent by regulating claudin-5 expression.

Furthermore, claudin- 5 is involved in maintaining cell polarity and intercellular barriers (102). It is expressed in the vascular endothelium and some lymphatic vessels as well as in cells of many vascular and non-vascular tumors (40). It is expressed to a lesser extent in the subepithelial vessels of pancreatic plasmacytoma (SN) than in the interstitial vessels of SN (41). Claudin-5 is significantly positively expressed in pancreatic solid-pseudopapillary tumor cell membranes (SPN), but not in pancreatic neuroendocrine tumors (P-NET), and combined IHC detection of claudin-5 and intercellular adhesion molecule, CD99, is required for the diagnosis of SPN and P-NET (42).

Claudin-7 is expressed in normal and malignant pancreatic tissue. It is expressed in the alveoli and ducts of the normal pancreas, and its expression is decreased in pancreatic ductal adenocarcinoma and is associated with the degree of differentiation of the tumor. In tumor cells, claudin-7 is present in two phases. Only GEM-located claudin-7 (palmitoylation) promotes tumor cell metastasis. Palmitoylated claudin-7 is involved in the development of drug resistance by inhibiting PTEN and subsequently activating the PI3K/Akt pathway (103). Because phosphorylated PTEN is not directed to GEM to compete with non-phosphorylated PTEN, association of cld7mPalm (claudin-7 has a mutated palmitoylation site) with the major PTEN phosphorylation kinase does not restore apoptosis resistance (104). Claudin-7 expression is negatively correlated with the glandular size of the tumor cells, and it is more pronounced in medium and large ductal tumors and well-differentiated tumors (47). Cancer-initiating cell-tumor exosomes (CIC-TEX) reprogram non-CIC into malignant tumors, and claudin-7, a biomarker for CIC, was recovered in gastrointestinal TEX. It was shown that individual cells delivering different microvesicles are particularly sensitive to claudin-7 and that claudin-7 located in the GEM is palmitoylated, which promotes the GEM-integrated plasma membrane and associated recruitment of signaling molecules. Non-palmitoylated claudin-7 recruits transport components, proteins involved in fatty acid metabolism, and TJ proteins into TEX. Claudin-7 also helps TEX recover selected miRNAs. Thus, well-localized claudin-7 affects CIC-TEX composition and claudin-7 may play a unique role in equipping CIC-TEX with transport proteins and regulatory molecules for lipid metabolism (48). CLDN7-associated epithelial cell adhesion molecules (EpCAM) are recruited to TEM, which support proliferation accompanied by sustained extracellular signaling regulating kinase-1/2 phosphorylation, anti-apoptotic protein upregulation, and drug resistance, but not EpCAM-mediated intercellular adhesion. This was only observed in EpCAM-CLDN-7-expressing cells where co-localization of claudin-7 with actin bundles may support enhanced motility. Additionally, the EpCAM-CLDN-7 complex strongly promotes tumorigenicity, accelerates tumor growth, and promotes ascites production and thymic metastasis formation. Therefore, high expression of the tumor marker, EpCAM, is often associated with poor prognosis (105). Claudin-7 functions through integrins to regulate cell proliferation and maintain epithelial cell attachment, and integrin β1 forms a complex of proteins that regulate cell growth and cell cycle progression, which in turn exerts control over tumor cell proliferation (106).

The solid papillary type (SPT) of PCa can be distinguished from the alveolar and neuroendocrine tumors by claudin-7 and claudin-5 immunoreactivity. Furthermore, SPT is positive for claudin-18.2and claudin-7 in a few cases, and pancreatic endocrine tumor (PET), acinar cell carcinoma(ACC), and pancreatoblastoma (PB) show strong expression of claudin-7 and a lack of claudin-5 expression (107).

Testicular pancreatic similar SPN is a recently reported entity that morphologically overlaps with the primary indolent mesenchymal tumor of the testis (PSRSTT), and its immune profile is positive for claudin-7. Based on the histological and genetic similarities found between pancreatic SPN and PSRSTT, it can be assumed that both tumors have the same pathogenesis; therefore, PSRSTT can be considered as a testicular analog of pancreatic SPN, and claudin-7 may play an important role in the diagnosis of this disease (108).

DNA microarray analysis showed that claudin-7 was highly expressed in MIA PaCa-2-A cells, and claudin-7 knockdown in MIA-PaCa-2-A cells significantly inhibited pancreatic tumor proliferation, reduced expression of p-Erk1/2, and inhibited G1 cell cycle arrest. Thus, CLDN7 may be expressed in rapidly proliferating and dominant cell populations in human PCa tissues and could be a novel molecular target for PCa treatment (49).

Claudin-12 plays an oncogenic role in PCa cells and promotes its malignant phenotype. Long intergenic non-coding RNAs (LNC00857) was found to affect the phenotype of PAAD cells by targeting miR-150-5p to regulate claudin-12 expression and by recruiting serine/arginine-rich splicing factor 1 (SRSF1) to promote selective splicing (AS) targeting claudin-12. ZNF460-regulated LINC00857 upregulates claudin-12 expression by sponging miR-1505p and recruiting SRSF1 to promote PAAD cell progression (53).Interleukin (IL)-18 is important in inducing BC cell migration via the downregulation of claudin-12 (52). Additionally, Claudin-12 promotes the proliferation and migration of osteosarcoma cells (109). Recent studies in PCa showed that knockdown of claudin-12 inhibited the proliferation of PANC-1 cells and downregulation of claudin-12 limited cell migration and invasion, further demonstrating that claudin-12inhibited EMT of PANC-1 cells. It is valuable to target miR-150-5p to regulate claudin-12 expression investigate therapeutic targets to inhibit PAAD cell progression and miR-150-5p.

Human CLDN23 mRNA is expressed in germinal center B cells, placenta, stomach, and colon cancer. It is a candidate oncogene associated with intestinal-type GC (7). Wei Wang et al. found that the dissociation factors (DFs) isolated from the medium of highly invasive and metastatic PC-1.0 PCa cells induced dissociation of weakly invasive and migratory PCa cells (PC-1), and the addition of DF-conditioned medium significantly reduced CLDN23 mRNA and protein expression in PC-1 cells. These results suggest that CLDN23 is involved in the regulation of PCa cell isolation through changes in gene expression and intracellular localization. In addition, Claudin-23 expression may be associated with the activation of the MEK signaling pathway during PCa cell isolation. Therefore, Claudin-23 also acts as a signaling molecule by actively translocating from cell–cell adhesion sites to the nucleus while simultaneously disrupting the structure of TJs (58). In colorectal cancer (CRC) studies, Claudin-23 expression was found to be epigenetically regulated and the significant downregulation of claudin-23 expression in tissues was attributed to the occupation of the claudin-23 gene, which encodes a structural component of cell-cell adhesion, by the Enhancer of zeste 2 (EZH2) and significant silencing in CRC tissues. The balance between the claudin-23 motif EZH2 methyltransferase activity and claudin-23 expression in CRC tissues, H3 lysine 27 trimethylation (H3K27me3) and H3K4me2, may underlie the regulation of claudin-23 expression. Existing studies suggest that this down-regulation causes disruption of intercellular adhesion function, providing more oxygen and nutrients to the microenvironment in which cancer cells live and enhancing their ability to metastasize (110). Studies on claudin-23 regulation in pancreatic cancer are scarce, and research on claudin-23 in pancreatic cancer could contribute to new targets for molecular therapy to prevent invasion and metastasis of pancreatic cancer.