Current perspectives and trends of CD39-CD73-eAdo/A2aR research in tumor microenvironment: a bibliometric analysis

The Web of Science Core Collection (WoSCC) database is widely used in bibliometrics. Data is obtained from the WoSCC database from 2015 to March 22nd, 2024. The search formula is ((((((AB=(CD39)) OR AB=(CD73)) OR AB=(adenosine)) OR AB=(ENTPD1)) OR AB=(NT5E)) OR AB=(A2aR)) OR AB=(A2AR) AND ((((AB=(immune microenvironment)) OR AB=(immune)) OR AB=(microenvironment)) OR AB=(tumor microenvironment)) OR AB=(Tumor immune microenvironment) AND (((((((AB=(Tumor)) OR AB=(Neoplasm)) OR AB=(Tumors)) OR AB=(Neoplasia)) OR AB=(Cancer)) OR AB=(Cancers)) OR AB=(Malignancy)) OR AB=(Benign Neoplasm). The publication type included is “article” or “review”, and the language is “English”.

The bibliometric tool based on R (version 4.3.0) and VOS viewer (16) (version 1.6.20) are used to quantify and identify the individual impact and cooperation information by analyzing these articles’ detailed information, such as annual global publications, distribution of countries/regions and institutions, journals, authors, and co-cited authors. Meanwhile, we identify the researching hotspots and trending by analyzing the keywords, co-occurrence author keywords, and the co-cited references of CD39-CD73-eAdo/A2aR research in cancer immunity.

We collected the research results of eAdo synthesis key enzymes (CD39, CD37), adenosine, and A2aR in the field of TME from the Web of Science core collection database, screened the search results, and only included the articles and reviews in the past ten years on March 22nd, 2024. The basic situation of the search results is shown in Table 1. A total of 1,721 eligible papers are found. The number of publications in this field has increased over the past decade (Figure 2). From the perspective of author cooperation, more than 27.08% of the research results involve international cooperation, which reflects that this field is a topic of global concern, and there are frequent international academic exchanges and cooperation. In addition, the average number of co-authors in a single article is nine, which means that the research involved in this field is complex and requires multi-faceted knowledge and techniques. Considering the above data, we speculate that CD39-CD73-eAdo/A2aR are essential molecular markers or therapeutic targets in TME research. With the continued growth in the amount of literature and the high citation rate, this field will likely continue to expand, and research will be further in-depth, possibly discovering new treatments or drugs. The high proportion of international cooperation also indicates that there may be more international cooperation projects and transnational research teams to promote the understanding and application of CD39-CD73-eAdo/A2aR as targets in tumor immunotherapy.

Through the country and institutional analysis of bibliometric analysis, we can understand the research contributions of various countries and institutions in the adenosine axis and TME to navigate better and participate in this field. Table 2 presents the details of the top 10 productive countries/regions. So far, 69 countries have published papers in this field, of which China has the most papers (554), followed by the United States (347) and Italy (106) (Figure 3A). The lead of China in the number of documents published by a single country (448) indicated that China has a strong research capacity and high output in this field. However, the United States leads in multi-country collaborative publications (112), stating its activity and leadership in international scientific research collaboration. The high ratio of multi-country collaborative papers in Iran, Australia, and the United Kingdom suggests that these countries are more likely to collaborate across borders in scientific research, which helps drive broader scientific progress and innovation (Figure 3B). The United States has the highest number of citations (16,902 times), followed by China (8,794 times). Australia and the United States have the highest average number of citations per paper, 57.3 and 48.7 times, respectively, indicating that the above countries have significant influence and recognition in this field (Figure 3C). From the visualization results of international cooperation, we find cooperation between many countries, among which China and the United States have the closest cooperation (Figure 4). Of the institutions that have published the most papers in this field, the United States institutions occupy the top 10 spots, with Harvard University in first place with 167 publications (Table 3). The institutions from France, Iran, and China also make the ranking, with increasing annual publications (Figure 5). Regarding institutional collaboration, the Fudan University and Nanjing Medical University in China cooperate most closely (Supplementary Figure 1).

We can understand which authors influence scientific research activities in the adenosine axis and TME through the author’s bibliometrics analysis. A total of 12,065 authors contribute to the publication of papers in this field, of which 23 published at least eight papers. Figure 6 shows the visual network diagram of the cooperative relationship between the authors of papers in this field. The authors are clustered into six categories, among which the nodes of Stagg J, Smyth MJ, Di Virgilio F, Mueller CE, Figueiro F, and Whiteside TI stand out in each cluster due to their large size, suggesting that they may be relatively active or essential researchers in this field. In addition, the close association between Stagg J and Allard B suggests frequent collaboration between them. Table 4 shows the top 10 authors with the most published papers, among which Stagg J ranks first with 24 published articles and 2,776 citations, indicating his significant academic influence in this field. The results of the author coupling analysis (Supplementary Figure 2) suggest that researchers can be clustered into five categories according to the citation of references. The research directions in the same category are similar, leading them to cite more duplicate references. Figure 7 shows the co-citation analysis results of reference authors co-cited ≥100 times. Co-cited authors can be clustered into three categories. Stagg J, Allard B, Ohta A, and Antonioli, L occupy a central position in the network and have more lines, indicating that their achievements are cited more than those of other authors and the research of these authors has an essential impact on the field, perhaps because of the high quality of their study, the critical topic of their research, or widespread recognition within the academic community.

Through bibliometric analysis of journals, we can identify the academic journals that have played a vital role in the adenosine axis and TME. A total of 579 scholarly journals have published articles in this field. Table 5 lists the top 10 academic journals with the most published articles. The journal FRONTIERS IN IMMUNOLOGY publishes the most significant number of papers, with 97 articles and a total of 2,317 citations, and the number of publications increased year by year (Figure 8A). The journal CANCERS publishes 55 articles with a total of 774 citations. Bradford’s Law helps libraries and researchers identify the most important journals for a particular field. The Bradford’s Law distribution map shows that a few journals constitute the core source of information, which is critical to researchers because they publish most research papers, such as FRONTIERS IN IMMUNOLOGY and CANCERS. (Figure 8B). Supplementary Table 1 presents the details of the top 10 local highly cited journals. Supplementary Figure 3 shows the co-citation analysis results of co-cited journals cited ≥500 times. CANCER RESEARCH, JOURNAL OF IMMUNOLOGY, and NATURE are the core journals in this field with high co-citation frequency. The close association between CELL, NATURE, and SCIENCE, as well as FRONTIERS IN IMMUNOLOGY and CANCER RESEARCH, shows that they form a core cluster group in the research field, and their research results are often co-cited.

Through bibliometric paper analysis, we can identify the core papers that have significantly influenced the adenosine axis in TME research. Through an in-depth analysis of the citation patterns, co-citation relationships, and frequency of occurrence of these critical papers, we can trace the origin and development of the research’s knowledge. 1,721 articles and 74,858 citations in this field currently meet our inclusion criteria. Table 6 presents the field’s top 10 most cited papers. “The ectonucleotidases CD39 and CD73: Novel checkpoint inhibitor targets”, published in IMMUNOLOGICAL REVIEWS by ALLARD B et al., is the most frequently local cited (163 times). Figure 9 shows the paper coupling network analysis. Papers in the same cluster indicate that they are strongly related academically, probably because they study similar topics, methods, or theories, and many of the same articles are in the reference. These closely connected papers may constitute a research cluster or sub-field in this field. Figure 10 shows the co-citation network analysis of references co-cited ≥80 times. Supplementary Table 2 shows the top 10 highly co-cited references. The “A2A adenosine receptor protects tumors from antitumor T cells” was the most co-cited reference (224 times). In this way, researchers can identify critical papers and research trends in the field, which can be very helpful in determining the research directions.

A total of 3,324 author keywords were included in this analysis Figure 11A shows the word cloud of author keywords. It can be seen that immunotherapy occupies a central position in the figure and is shown as the most significant word, indicating that the relationship between the adenosine axis and immunotherapy is a core concept for many researchers in this field. The size of these disease author keywords, such as breast cancer, melanoma, colorectal cancer, lung cancer, ovarian cancer, glioblastoma, pancreatic cancer, and hepatocellular carcinoma, indicates that they are the most frequent cancer types in adenosine-related tumor studies (Figure 11B). Immunotherapy, immunosuppression, immune checkpoint, and immune checkpoint inhibitors are the hot keywords in the research, reflecting the importance of the adenosine axis in tumor immunotherapy. Author keywords related to tumor immunity, such as immunogenic cell death, T cells, Sting, regulatory T cells, innate immunity, and immune infiltration, demonstrate the pathways by which the adenosine axis affects the TME. The combination of these author keywords gives a comprehensive view of the importance of the adenosine axis in the research field of the TME, including its role in disease development, treatment response, and prognostic assessment. Figure 11C is a topic distribution map, and the author keywords in the adenosine axis and TME field are mainly concentrated in the second and fourth quadrants. Pancreatic cancer, lung cancer, innate immunity, Sting, etc., in the second quadrant, are the core topics with high maturity in current research. In the fourth quadrant, immunotherapy, immunogenic cell death, immunosuppression, macrophages, PD-1, breast cancer, ovarian cancer, and glioblastoma are basic topics with low maturity in the current research field, and these topics may become research hotspots or future development trends. This analysis helps investigators decide which areas to focus on and which emerging areas are worth further exploration. Figure 12 presents the author keywords co-occurrence analysis with occurrences ≥15 times. It can be seen that the famous author keywords in recent years have been immunotherapy, immunogenic cell death, inflammation, lung cancer, and gastric cancer.

Cytotoxic T lymphocytes (CTLs) are antigen-specific effector cells that can eliminate cancer cells contact-dependently. Despite the infiltration of tumor-specific CTLs, metabolic disorder impairs the anti-tumor function of tumor-infiltrating CTLs. Restoring the function of these tumor infiltrating CTLs is vital to improving immunotherapy. eAdo weakened the one-to-one binding of activated effector CTLs to target tumor cells, inhibited the sustained anti-tumor effect, and severely impaired the proliferation ability of CTLs. Antagonism of A2aR signaling stabilized and prolonged CTLs-target cell binding and accelerated the delivery of lethal hits to individual contacts and CTLs (17). The combination of A2aR and TIM3 damaged the cytoskeletal polarization of CTLs in vivo and in vitro and inhibited CTLs infiltration. Blockade of A2aR and TIM3 may enhance T-cell activation therapies (such as anti-PD-1 and anti-CTLA-4) (18). Hypoxia-induced eAdo also can lead to inhibitory TME and impede the efficacy of ICIs (19). Studies have found that A2aR blockers improved NK cells’ maturation and cytotoxic function and inhibited tumor metastasis in a perforin-dependent manner (20).

In terms of treatment, the combination of adenosine receptor antagonist and anti-PD-1 prolonged the survival of mice with liver cancer (19). Inhibiting PD-1/PD-L1/L2 combined with inhibiting CD73/A2aR can effectively treat diffuse large B-cell lymphoma (21). The A2aR antagonist ciforadenant has shown therapeutic activity and good tolerance in patients receiving immunotherapy for the first time and in patients resistant or refractory to anti-PD-1/L1 treatment in a phase 1 clinical trial in renal cell carcinoma (22). The novel A2aR antagonist DZD2269 could reverse the immunosuppression induced by high concentration eAdo and play a more significant anti-tumor effect when combined with ICIs, radiotherapy, or chemotherapy (23). Taminadenant (PBF509/NIR178) is also a kind of A2aR antagonist that can activate the anti-tumor immune response. Clinical benefit was observed in all patients treated with taminadenant (24).

A2aR is the key immune regulatory factor that protects normal tissues from inflammatory damage and inhibits CTLs from antitumor function of the body. Lack of oxygen and eAdo-rich TME can inhibit the antitumor function of CTLs (25). Respiratory hyperoxia also promoted the regression of spontaneous metastasis from orthotopically grown breast tumors (25). Sitkovsky and collaborators proposed targeting the hypoxia-adenosine-A2aR pathway as a cancer immunotherapy strategy to prevent suppression of antitumor T cells in the TME (13–15).

Although radiotherapy (RT) has been highly successful in the treatment of non-small cell lung cancer, local recurrence occurs, and distant effects are rare, even when combined with ICBs. The combination of CD39 inhibitor and RT significantly increased the efficacy of RT. The combination treatment significantly inhibited tumor growth and promoted cytotoxic CD8+T infiltration in the tumor and mobilization of CD8+T cell-dependent antitumor immune response (58). RT can induce immunogenic cell death to activate antitumor immune response. In contrast, activating immune escape processes, such as PD-L1 and the up-regulation of CD39 and CD73, can partially counteract the radiation-induced immune response (59, 60). CHEN et al. (60) constructed nano-scale coordination particle AmGd-NPs by assembling high-Z metal gadolinium (Gd) and small-molecule CD73 inhibitor AmPCP. AmGd-NPs can gradually release AmPCP to inhibit the enzyme activity of CD73, driving a pro-inflammatory tumor microenvironment that promoted DC maturation and enhanced CD8+T cell-dependent antitumor immune response. Ionizing radiation activated the STAT1-IRF1-CD39 axis to up-regulate the expression of CD39. Blocking CD39 enabled eATP accumulation, which activated the dendritic cell NLRP3 inflammasome via the P2X7 receptor, thereby promoting radiation-induced ICD (12). Glioma stem cell ICD vaccine induced by radiation and CD39 inhibition can enhance the expansion of CAR-T in peripheral blood, the multifunction of TME, and the antitumor effect in glioma models (12). Combined blocking of CD73 and PD-L1 can significantly enhance the antitumor effects of Stereotactic body radiotherapy (SBRT) and prolong the survival of patients. Triple therapy (SBRT+anti-CD73+anti-PD-L1) modulated tumor-infiltrating immune cells by increasing IFN-γ+CD8+T cells. In addition, triple therapy reprogrammed cytokines/chemokines in the TME to a more immune-stimulating phenotype (59).

The failure of chimeric antigen receptor (CAR) T-cell immunotherapy to treat solid tumors is partly due to the immunosuppressive microenvironment. During the generation of fully human anti-mesothelin CAR T cells (MSLN-CAR T) (cell activation and transduction), A2aR was significantly upregulated in T cells. Targeting the A2aR signaling pathway may be a promising approach to improve the anti-tumor function of CAR-T cells and may improve clinical treatment outcomes (61). The CRISPR/Cas9 strategy targeted A2aR-edited CAR T cells to fight adenosine-mediated immunosuppression significantly, resulting in enhanced production of cytokines (including IFNγ and TNF) and increased expression of genes associated with the JAK-STAT signaling pathway, thereby improving survival in mice. A2aR-deficient CAR T cells were well tolerated and did not induce significant pathological changes in mice, which supported using CRISPR/Cas9 to target A2aR in the clinic to improve CAR T cell function (62). Liu et al. (63) found that shRNA-mediated A2aR interference can also improve the anti-mSLN-CAR T cell anti-tumor effect in vivo and in vitro, and proved that shRNA-mediated gene expression modification may be an excellent strategy for improving CAR T cell function in immunosuppressive TME.

Regulatory T (Treg) cells maintain immune homeostasis by suppressing abnormal/excessive immune responses to self- and non-self-antigens. However, high infiltration of Treg cells is associated with poor survival in various cancers. CD73 can up-regulate CCL5 through the p38-STAT1 axis through the eAdo-A2aR signaling pathway, recruit Treg cells to pancreatic tumors, and cause inhibitory TME (64). Treg cells, which undergo apoptosis due to a weaker NRF2-related antioxidant system and a high vulnerability to free oxygen in TME, can convert large amounts of released eATP to eAdo via CD39 and CD73 to mediate immunosuppression via the eAdo/A2aR pathway. Treg cells in TME can be killed by oxidative stress to maintain and amplify their immunosuppressive capacity (65). CD73+γδTreg cells had more potent immunosuppressive activity than CD4+ or CD8+Tregs (66). Cancer-associated fibroblasts (CAF) derived IL6 induced the differentiation of CD73+γδTreg in normal breast tissue through IL6/STAT3 pathway. CD73+γδTreg, in turn, promoted IL6 secretion by CAFs through eAdo/A2bR/p38MAPK signaling pathway (66). Infiltration of CD73+γδ Treg cells was significantly associated with poor patient prognosis in human breast cancer.

Therefore, there is an urgent need to explore strategies to deplete Treg cells and control their function to increase anti-tumor immune responses in cancer immunotherapy. Targeting adenosine metabolism is an important therapeutic target for Treg cells inhibition.

The original contributions presented in the study are included in the article/. Further inquiries can be directed to the corresponding authors. 1