Soluble CD26: From Suggested Biomarker for Cancer Diagnosis to Plausible Marker for Dynamic Monitoring of Immunotherapy.

Soluble CD26 (sCD26), a glycoprotein with dipeptidyl peptidase (DPP4) enzymatic activity, can contribute to early diagnosis of colorectal cancer and advanced adenomas and has been studied, including for prognostic purposes, across various other types of cancer and disease. The latest research in this field has confirmed that most, though not all, serum/plasma sCD26 is related to inflammation. The shedding and/or secretion of sCD26 from different immune cells are being investigated, and blood DPP4 activity levels do not correlate very strongly with protein titers. Some of the main substrates of this enzyme are key chemokines involved in immune cell migration, and both soluble and cell-surface CD26 can bind adenosine deaminase (ADA), an enzyme involved in the metabolism of immunosuppressor extracellular adenosine. Of note, there are T cells enriched in CD26 expression and, in mice tumor models, tumor infiltrating lymphocytes exhibited heightened percentages of CD26+ correlating with tumor regression. We employed sCD26 as a biomarker in the follow-up after curative resection of colorectal cancer for the early detection of tumor recurrence. Changes after treatment with different biological disease-modifying antirheumatic drugs, including Ig-CTLA4, were also observed in rheumatoid arthritis. Serum soluble CD26/DPP4 titer variation has recently been proposed as a potential prognostic biomarker after a phase I trial in cancer immunotherapy with a humanized anti-CD26 antibody. We propose that dynamic monitoring of sCD26/DPP4 changes, in addition to well-known inflammatory biomarkers such as CRP already in use as informative for immune checkpoint immunotherapy, may indicate resistance or response during the successive steps of the treatment. As tumor cells expressing CD26 can also produce sCD26, the possibility of sorting immune- from non-immune-system-originated sCD26 is discussed.

Improving colorectal cancer screening programs.

In this editorial we comment on the article by Agatsuma et al published in the World Journal of Gastroenterology. They suggest policies for more effective colorectal screening. Screening is the main policy that has led to lower mortality rates in later years among the population that was eligible for screening. Colonoscopy is the gold standard tool for screening and has preventive effects by removing precancerous or early malignant polyps. However, colonoscopy is an invasive process, and fecal tests such as the current hemoglobin immunodetection were developed, followed by endoscopy, as the general tool for population screening, avoiding logistical and economic problems. Even so, participation and adherence rates are low. Different screening options are being developed with the idea that if people could choose between the ones that best suit them, participation in population-based screening programs would increase. Blood tests, such as a recent one that detects cell-free DNA shed by tumors called circulating tumor DNA, showed a similar accuracy rate to stool tests for cancer, but were less sensitive for advanced precancerous lesions. At the time when the crosstalk between the immune system and cancer was being established as a new hallmark of cancer, novel immune system-related biomarkers and information on patients' immune parameters, such as cell counts of different immune populations, were studied for the early detection of colorectal cancer, since they could be effective in asymptomatic people, appearing earlier in the adenoma-carcinoma development compared to the presence of fecal blood. sCD26, for example, detected 80.37% of advanced adenomas. To reach as many eligible people as possible, starting at an earlier age than current programs, the direction could be to apply tests based on blood, urine or salivary fluid to samples taken during routine visits to the primary health system.

Comment on Lai et al. Dipeptidyl Peptidase 4 Stimulation Induces Adipogenesis-Related Gene Expression of Adipose Stromal Cells. Int. J. Mol. Sci. 2023, 24, 16101.

Adiponectin is a circulating hormone secreted by adipose tissue that exerts, unlike other adipokines such as leptin, anti-inflammatory, anti-atherosclerotic and other protective effects on health. Adiponectin receptor agonists are being tested in clinical trials and are expected to show benefits in many diseases. In a recent article, LW Chen's group used monocyte chemoattractant protein-1 (MCP-1/CCL2) to improve plasma levels of adiponectin, suggesting the involvement of dipeptidyl peptidase 4 (DPP4/CD26) in the mechanism. Here, we discuss the significance of the role of DPP4, favoring the increase in DPP4-positive interstitial progenitor cells, a finding that fits with the greater stemness and persistence of other DPP4/CD26-positive cells.

CD26 and Cancer.

This Special Issue presents new knowledge on the complex behaviour of dipeptidyl peptidase 4 (DPP4, EC 3 [...].

Evaluation of Blood Soluble CD26 as a Complementary Biomarker for Colorectal Cancer Screening Programs.

Fecal hemoglobin immunodetection (FIT) in combination with endoscopy has been implemented to reduce mortality from colorectal cancer (CRC), although there are issues that can be improved in relation to participation rates. We studied whether the blood biomarker soluble-CD26 (sCD26), related at least in part to the immune system and inflammation, and/or its dipeptidyl peptidase enzyme activity (DPP4), could help reduce false positives. In a cohort of 1703 individuals who underwent colonoscopy and had a serum sample, sCD26 and DPP4 activity showed statistically significant differences regarding sex and age. According to the colonoscopy findings, sCD26 and DPP4 activity progressively decreased in advanced adenomas and CRC, with statistically significant differences, even between both groups; 918 of them had a FIT result (n = 596 positive cases) with approximately 70% of these (n = 412) false positives. With cut-offs of 440 ng/mL for sCD26, 42 mU/mL for DPP4, and 11 ng/mU for their ratio, the combined information of the three biomarkers (at least positive for one biomarker) identified almost all advanced adenomas and CRC cases in the FIT cohort with approximately half of the false positives compared to FIT. A sequential testing strategy with FIT and our blood biomarker test is proposed.

Study of Plasma Anti-CD26 Autoantibody Levels in a Cohort of Treatment-Naïve Early Arthritis Patients.

In rheumatoid arthritis (RA), the identification of biomarkers to adjust treatment intensity and to correctly diagnose the disease in early stages still constitutes a challenge and, as such, novel biomarkers are needed. We proposed that autoantibodies (aAbs) against CD26 (DPP4) might have both etiological importance and clinical value. Here, we perform a prospective study of the potential diagnostic power of Anti-CD26 aAbs through their quantification in plasmas from 106 treatment-naïve early and undifferentiated AR. Clinical antibodies, Anti-CD26 aAbs, and other disease-related biomarkers were measured in plasmas obtained in the first visit from patients, which were later classified as RA and non-RA according to the American College of Rheumatology criteria. Two different isotype signatures were found among ten groups of patients, one for Anti-CD26 IgA and other for Anti-CD26 IgG and IgM isotypes, both converging in patients with arthritis (RA and Unresolved Undifferentiated Arthritis: UUA), who present elevated levels of all three isotypes. The four UUA patients, unresolved after two years, were ACPA and rheumatic factor (RF) negatives. In the whole cohort, 51.3% of ACPA/RF seronegatives were Anti-CD26 positives, and a similar frequency was observed in the seropositive RA patients. Only weak associations of the three isotypes with ESR, CRP and disease activity parameters were observed. Anti-CD26 aAbs are present in treatment-naïve early arthritis patients, including ACPA and RF seronegative individuals, suggestive of a potential pathogenic and/or biomarker role of Anti-CD26 aAbs in the development of rheumatic diseases.

Distinctive CD26 Expression on CD4 T-Cell Subsets.

Immune system CD4 T-cells with high cell-surface CD26 expression show anti-tumoral properties. When engineered with a chimeric antigen receptor (CAR), they incite strong responses against solid cancers. This subset was originally associated to human CD4 T helper cells bearing the CD45R0 effector/memory phenotype and later to Th17 cells. CD26 is also found in soluble form (sCD26) in several biological fluids, and its serum levels correlate with specific T cell subsets. However, the relationship between glycoprotein sCD26 and its dipeptidyl peptidase 4 (DPP4) enzymatic activity, and cell-surface CD26 expression is not well understood. We have studied ex vivo cell-surface CD26 and in vitro surface and intracellular CD26 expression and secretome's sCD26 in cultured CD4 T cells under different polarization conditions. We show that most human CD26negative CD4 T cells in circulating lymphocytes are central memory (TCM) cells while CD26high expression is present in effector Th1, Th2, Th17, and TEM (effector memory) cells. However, there are significant percentages of Th1, Th2, Th17, and Th22 CD26 negative cells. This information may help to refine the research on CAR-Ts. The cell surface CD45R0 and CD26 levels in the different T helper subsets after in vitro polarization resemble those found ex vivo. In the secretomes of these cultures there was a significant amount of sCD26. However, in all polarizations, including Th1, the levels of sCD26 were lower (although not significantly) compared to the Th0 condition (activation without polarization). These differences could have an impact on the various physiological functions proposed for sCD26/DPP4.

The mechanism of sitagliptin inhibition of colorectal cancer cell lines' metastatic functionalities.

The cell membrane glycoprotein CD26 with peptidase activity (DPP4) and/or its soluble CD26/DPP4 counterpart expression and/or activity are altered in several cancers. Its role in metastasis development was recently highlighted by the discovery of CD26+ cancer stem cell subsets and the fact that clinical DPP4 inhibitors showed antimetastatic effects in animal models. Also, diabetic patients treated with the DPP4 inhibitor sitagliptin showed greater overall survival after colorectal or lung cancer surgery than patients under other diabetic therapies. However, the mechanism of action of these inhibitors in this context is unclear. We studied the role of CD26 and its DPP4 enzymatic activity in malignant cell features such as cell-to-cell homotypic aggregation, cancer cell motility, and invasion in a panel of human colorectal cancer (CRC) cell lines, avoiding models that include the physiological role of DPP4 in chemotaxis. Present results indicate that CD26 participates in the induction of cell invasion, motility, and aggregation of CD26-positive CRC cell lines. Moreover, only invasion and motility assays, which are collagen matrix-dependent, showed a decrease upon treatment with the DPP4 inhibitor sitagliptin. Sitagliptin showed opposite effects to those of transforming growth factor-ß1 on epithelial-to-mesenchymal transition and cell cycle, but this result does not explain its CD26/DPP4-dependent effect. These results contribute to the elucidation of the molecular mechanisms behind sitagliptin inhibition of metastatic traits. At the same time, this role of sitagliptin may help to define areas of medicine where DPP4 inhibitors might be introduced. However, they also suggest that additional tools against CD26 as a target might be used or developed for metastasis prevention in addition to gliptins.

CD26-Related Serum Biomarkers: sCD26 Protein, DPP4 Activity, and Anti-CD26 Isotype Levels in a Colorectal Cancer-Screening Context.

Current screening trials are showing reduction in colorectal cancer incidence and mortality. However, participation rates are often low, and blood-based tests could complement existing screening strategies. CD26 protein (sCD26) and its dipeptidyl peptidase IV (DPP4) enzymatic activity in circulation have been proposed as biomarkers for colorectal cancer and other diseases. However, changes in sCD26 and DPP4 levels show complex degrees of correlation, and their physiological or pathophysiological role is unclear. The aim of this study was to analyse if anti-CD26 autoantibodies are related to sCD26 and DPP4 and to determine their relevance in a context of colorectal cancer screening for complementing the value of sCD26 and DPP4 as biomarkers. These biomarkers were measured in a large prospective cohort (n = 497, except the anti-CD26 antibodies, evaluated in 125 samples) that included a subgroup of individuals that were positive for the faecal immunological occult blood test (FIT) (n = 86) and underwent a colonoscopy (n = 47). We confirmed for the first time higher DPP4 activity in men compared to women (Student's t test, p = 0.002), though this difference between sexes was not seen for serum sCD26 protein. These biomarkers correlated (R = 0.246, p = 0.003) only in women. Correlations were found between anti-CD26 isotypes but not with DPP4 activity or sCD26 concentration, except for a negative correlation only in men between anti-CD26 IgA isotype and sCD26 (R = -0.232, p = 0.044), and an almost significant negative correlation between anti-CD26 IgG and sCD26 limited to FIT-positive men. Interestingly, patients with advanced adenomas displayed the most elevated mean levels of anti-CD26 IgA, IgM, and particularly IgG (Mann-Whitney U test, p = 0.030) in comparison with the other FIT positives without adenomas, and these levels did not correlate with sCD26 or its DPP4 activity. Our preliminary results suggest that the combination of these measures using sex as confounder could perhaps be used as biomarkers for colorectal disease. It also suggests that events affecting the gut influence the levels of anti-CD26 antibodies, which show little or no effect in antigen clearance. These findings should be confirmed in a larger cohort of individuals with colonoscopy. The physiological origin of the sex differences observed should be further addressed.

Surface expression marker profile in colon cancer cell lines and sphere-derived cells suggests complexity in CD26+ cancer stem cells subsets.

Taking advantage of eight established cell lines from colorectal cancer patients at different stages of the disease and the fact that all of them could form spheres, cell surface biomarkers of cancer stem cells and epithelial-mesenchymal transition were tested. The aim was to investigate cancer stem cells and metastatic stem cells in order to provide functional characterization of circulating tumor cells and promote the development of new anti-metastatic therapies. Our model showed an important heterogeneity in EpCAM, CD133, CD44, LGR5, CD26 and E-cadherin expression. We showed the presence of a subset of E-cadherin+ (some cells being E-cadherinhigh) expressing CD26+ (or CD26high) together with the well-known CSC markers LGR5 and EpCAMhigh, sometimes in the absence of CD44 or CD133. The already described CD26+/E-cadherinlow or negative and CD26+/EpCAM-/CD133- subsets were also present. Cell division drastically affected the expression of all markers, in particular E-cadherin, so new-born cells resembled mesenchymal cells in surface staining. CD26 and/or dipeptidyl peptidase 4 inhibitors have already shown anti-metastatic effects in pre-clinical models, and the existence of these CD26+ subsets may help further research against cancer metastasis.

Naturally presented HLA class I-restricted epitopes from the neurotrophic factor S100-ß are targets of the autoimmune response in type 1 diabetes.

Type 1 diabetes (T1D) results from the destruction of pancreatic ß-cells by the immune system, and CD8+ T lymphocytes are critical actors in this autoimmune response. Pancreatic islets are surrounded by a mesh of nervous cells, the peri-insular Schwann cells, which are also targeted by autoreactive T lymphocytes and express specific antigens, such as the neurotrophic factor S100-ß. Previous work has shown increased proliferative responses to whole S100-ß in both human T1D patients and the nonobese diabetic (NOD) mouse model. We describe for the first time naturally processed and presented epitopes (NPPEs) presented by class I human leukocyte antigen-A*02:01 (A2.1) molecules derived from S100-ß. These NPPEs triggered IFN-γ responses more frequently in both newly diagnosed and long-term T1D patients compared with healthy donors. Furthermore, the same NPPEs are recognized during the autoimmune response leading to diabetes in A2.1-transgenic NOD mice as early as 4 wk of age. Interestingly, when these NPPEs are used to prevent diabetes in this animal model, an acceleration of the disease is observed together with an exacerbation in insulitis and an increase in S100-ß-specific cytotoxicity in vaccinated animals. Whether these can be used in diabetes prevention needs to be carefully evaluated in animal models before use in future clinical assays.-Calviño-Sampedro, C., Gomez-Tourino, I., Cordero, O. J., Reche, P. A., Gómez-Perosanz, M., Sánchez-Trincado, J. L., Rodríguez, M. Á., Sueiro, A. M., Viñuela, J. E., Calviño, R. V. Naturally presented HLA class I-restricted epitopes from the neurotrophic factor S100-ß are targets of the autoimmune response in type 1 diabetes.

Oral hygiene might prevent cancer.

Many evidences support that species from the Human Oral Microbiome Database such as Fusobacterium nucleatum or Bacteroides, linked previously to periodontitis and appendicitis, play a role in colorectal cancer (CRC), including metastasis. These typically oral species are invasive anaerobes that form biofilms in their virulent state. Aspirin (a NSAID) has been recently included into routine CRC prevention rationale. NSAIDs can prevent the growth of neoplastic lesions by inhibiting COX enzymes and another set of recently identified COX-independent targets, which include the WNT, AMPK and MTOR signaling pathways, the crosstalk between nucleoli and NF-κB transcriptional activity in apoptosis, and the biochemistry of platelets. These are signaling pathways related to tumor-promoting inflammation. In this process, pathogens or simple deregulation of the microbiota play an important role in CRC. Aspirin and other NSAIDs are efficient inhibitors of biofilm formation and able to control periodontitis development preventing inflammation related to the microbiota of the gingival tissue, so its seems plausible to include this pathway in the mechanisms that aspirin uses to prevent CRC. We propose arguments suggesting that current oral hygiene methods and other future developments against periodontitis might prevent CRC and probably other cancers, alone or in combination with other options; and that the multidisciplinary studies needed to prove this hypothesis might be relevant for cancer prevention.

Anti-CD26 autoantibodies are involved in rheumatoid arthritis and show potential clinical interest.

OBJECTIVES: Rheumatoid arthritis (RA) patients show low serum levels of the Ag dipeptidyl peptidase IV (DPP-IV/CD26), both soluble CD26 (sCD26) concentration and its DPP-IV activity. The aim of this study was to test if anti-DPP-IV/CD26 Abs (Anti-CD26) cleared sCD26. DESIGN & METHODS: Serum Anti-CD26 and Total titers (as comparison) of isotypes IgA, IgM and IgG as well as sCD26 concentration and DPP-IV activity were measured in a cohort of RA patients undergoing different biological and non-biological therapies (n=105) and controls (n=50). RESULTS: Anti-CD26 levels were increased approximately two-fold for each isotype in RA, were not related to the sCD26 clearance, showed several correlations with disease activity parameters, were significantly higher in smokers and they were not ACPA. Anti-CD26 Igs showed high diagnostic power (82% sensitivity and 96% specificity) and their levels differed amongst the different groups of patients stratified by the type of therapy. CONCLUSIONS: As DPP-IV/CD26 is associated to factors triggering RA in the lung and periodontal tissue, these results suggest that Anti-CD26 isotypes may participate in pathogenesis and may be useful as biomarkers for earlier diagnosis and/or precision medicine.

Apportioning Blame: Autoreactive CD4+ and CD8+ T Cells in Type 1 Diabetes.

Type 1 diabetes (T1D) is one of the most studied archetypal organ-specific autoimmune diseases. Although many clinical, epidemiological, and pathological characteristics have been described, there are still important issues which need to be resolved as these will have a major impact on the development of future antigen-specific immunotherapies. An important question relates to T lymphocytes in the development of the disease, in particular their role in the destruction of insulin-producing beta cells. Since the discovery that certain class II histocompatibility complex molecules (HLA) are linked to the development of T1D, much research has focused on CD4+ helper T lymphocytes; however, recent studies highlight class I HLA molecules as an independent risk factor; hence, research into the role played by CD8+ cytotoxic T lymphocytes has gained momentum. In this review, we summarize recent studies clarifying the role played by both sets of autoreactive T lymphocytes in T1D, discuss the targets recognized by these cells and their phenotype in T1D patients. Finally, we will examine the possible generation of regulatory CD8+ T lymphocytes upon different immuno-intervention strategies.

Activity and expression of dipeptidyl peptidase IV on peripheral blood mononuclear cells in patients with early steroid and disease modifying antirheumatic drugs naïve rheumatoid arthritis.

BACKGROUND: Dipeptidyl peptidase IV (DPPIV/CD26) plays an important role in T cell activation and immune regulation, however the role of this enzyme in early rheumatoid arthritis (eRA) has not been clearly defined. The aim of this study was to determine the serum activity of DPPIV, its expression on peripheral blood mononuclear cells (PBMC) and to examine possible correlations with disease activity (DAS28) in untreated patients with eRA. METHODS: The study included 50 patients newly diagnosed with RA, who had not received any corticosteroid or disease modifying antirheumatic drugs (DMARD) therapy and whose conventional radiographs of hands and feet showed no structural damage. The control group consisted of 40 healthy volunteers. Also, 30 patients with chronic RA (cRA) were examined. The serum activity of DPPIV was determined by the direct photometric method, while expression of CD26 on PBMC was determined using flow cytometry. RESULTS: Decreased DPPIV serum activity was detected in patients with eRA and cRA compared to the control group (p=0.024, p<0.0001, respectively). Although, the percentage of overall CD26+ white blood cells (WBC) was significantly decreased in eRA patients (p<0.001), the percentage of CD26+ lymphocytes and monocytes and mean fluorescence intensity of CD26 on these cells in eRA patients showed no significant difference compared to healthy volunteers. DAS28 showed no significant correlation with CD26 expression or DPPIV serum activity, but a significant inverse correlation between the duration of symptoms and DPPIV serum activity was observed. CONCLUSIONS: Our results show that a decrease in DPPIV serum activity, but not CD26 expression, is present in an early stage of rheumatoid arthritis.

CD26 Expression on T Helper Populations and sCD26 Serum Levels in Patients with Rheumatoid Arthritis.

We studied dipeptidyl peptidase IV (DPP-IV, CD26) expression in different T helper cells and serum soluble DPP-IV/sCD26 levels in rheumatoid arthritis (RA) patients, correlated these with disease activity score (DAS), and examined how they were affected by different therapies, conventional or biological (anti-TNF, anti-CD20 and anti-IL6R or Ig-CTLA4). The percentage of CD4+CD45R0+CD26- cells was greatly reduced in patients (up to 50%) when compared with healthy subjects. Three other subsets of CD4 cells, including a CD26high Th1-associated population, changed variably with therapies. Data from these subsets (frequency and staining density) significantly correlated with DAS28 or DAS28 components but different in each group of patients undergoing the different therapies. Th17 and Th22 subsets were implicated in RA as independent CCR4+ and CCR4- populations each, with distinct CD26 expression, and were targeted with varying efficiency by each therapy. Serum DPP-IV activity rather than sCD26 levels was lower in RA patients compared to healthy donors. DPP-IV and sCD26 serum levels were found related to specific T cell subsets but not to disease activity. We conclude that, according to their CD26 expression, different cell subsets could serve to monitor RA course, and an uncharacterized T helper CD26- subset, not targeted by therapies, should be monitored for early diagnosis.

Postoperative serum levels of sCD26 for surveillance in colorectal cancer patients.

One of the main aims of the follow-up after curative resection of colorectal cancer is the early detection and treatment of tumor recurrence. We previously demonstrated decreased preoperative soluble CD26 (sCD26) levels in serum from colorectal cancer patients. We extended now the study to investigate if sCD26 levels in postoperative serum serve as marker of recurrence of the disease during surveillance. Soluble sCD26 was measured in pre- and postoperative serum samples of 43 patients with primary colorectal cancer. Carcinoembryonic antigen, carbohydrate antigen 19.9 and 72.4 levels were also measured during surveillance. The average follow-up period was 41.8 ± 20.8 months. sCD26 levels during follow-up showed well-defined patterns in patients without disease (n = 28), and in patients with tumor persistence (n = 2), local recurrence (n = 3) or distant metastasis (n = 10). Disease-free patients showed stable levels between 460-850 ng/mL during follow-up, while high (over 850 ng/mL) and unstable sCD26 levels were found before recurrence was diagnosed. The mean maximum/minimum sCD26 ratios during surveillance were 1.52, 2.12 and 2.63 for patients with no recurrence, local recurrence and metastasis, respectively (p = 0.005). From the cut-off obtained from a receiver operator characteristics (ROC) curve built with the maximum/minimum sCD26 ratios and the upper and lower cut-offs of sCD26, we were able to discriminate patients with and without recurrent disease. We propose that the measurement of serum sCD26 during the follow-up of patients diagnosed of colorectal cancer could be valuable for the early detection of local and distant recurrence. A large, randomized, prospective trial should be performed to confirm our findings.

Evaluation of pleural effusion sCD26 and DPP-IV as diagnostic biomarkers in lung disease.

In this study, we measured ADA and DPP-IV enzymatic activity and sCD26 concentration in 150 pleural effusion (PE) samples and tested for correlations between these and other cellular and biochemical measures. We found that DPP-IV in particular might improve the specificity (but not the sensitivity) of the ADA test for diagnosis of pulmonary tuberculosis, since half of the false ADA positive results in non-tuberculous PE were also DPP-IV positive. A percentage of patients with malignant PE were sCD26 or DPP-IV positive; however, some patients with benign PE also tested positive. As a pattern associated with DPP-IV (but not the CD26 protein) was observed in PE, we searched for a finding that might increase the value of these biomarkers for diagnosis of malignancy. The observed pattern was related to the presence of leukocytes, as indicated by correlations with the cell count, and to a band of 180 kDa, detected by immunoblotting.