A Predictive Noninvasive Single-Nucleotide Variation-Based Biomarker Signature for Resectable Pancreatic Cancer: Protocol for a Prospective Validation Study.

BACKGROUND: Single-nucleotide variations (SNVs; formerly SNPs) are inherited genetic variants that can be easily determined in routine clinical practice using a simple blood or saliva test. SNVs have potential to serve as noninvasive biomarkers for predicting cancer-specific patient outcomes after resection of pancreatic ductal adenocarcinoma (PDAC). Two recent analyses led to the identification and validation of three SNVs in the CD44 and CHI3L2 genes (rs187115, rs353630, and rs684559), which can be used as predictive biomarkers to help select patients most likely to benefit from pancreatic resection. These variants were associated with an over 2-fold increased risk for tumor-related death in three independent PDAC study cohorts from Europe and the United States, including The Cancer Genome Atlas cohorts (reaching a P value of 1×10-8). However, these analyses were limited by the inherent biases of a retrospective study design, such as selection and publication biases, thereby limiting the clinical use of these promising biomarkers in guiding PDAC therapy. OBJECTIVE: To overcome the limitations of previous retrospectively designed studies and translate the findings into clinical practice, we aim to validate the association of the identified SNVs with survival in a controlled setting using a prospective cohort of patients with PDAC following pancreatic resection. METHODS: All patients with PDAC who will undergo pancreatic resection at three participating hospitals in Switzerland and fulfill the inclusion criteria will be included in the study consecutively. The SNV genotypes will be determined using standard genotyping techniques from patient blood samples. For each genotyped locus, log-rank and Cox multivariate regression tests will be performed, accounting for the relevant covariates American Joint Committee on Cancer stage and resection status. Clinical follow-up data will be collected for at least 3 years. Sample size calculation resulted in a required sample of 150 patients to sufficiently power the analysis. RESULTS: The follow-up data collection started in August 2019 and the estimated end of data collection will be in May 2027. The study is still recruiting participants and 142 patients have been recruited as of November 2023. The DNA extraction and genotyping of the SNVs will be performed after inclusion of the last patient. Since no SNV genotypes have been determined, no data analysis has been performed to date. The results are expected to be published in 2027. CONCLUSIONS: This is the first prospective study of the CD44 and CHI3L2 SNV-based biomarker signature in PDAC. A prospective validation of this signature would enable its clinical use as a noninvasive predictive biomarker of survival after pancreatic resection that is readily available at the time of diagnosis and can assist in guiding PDAC therapy. The results of this study may help to individualize treatment decisions and potentially improve patient outcomes. INTERNATIONAL REGISTERED REPORT IDENTIFIER (IRRID): DERR1-10.2196/54042.

[Expression of CD44 in Tumor Tissue and Serum of Small Cell Lung Cancer and Its Clinical Prognostic Significance].

BACKGROUND: Small cell lung cancer (SCLC) is a highly aggressive malignancy characterized by rapid growth, early metastasis and acquired therapeutic resistance, and the prognosis is extremely poor. Studies have proved that the stem cell marker CD44 is correlated with tumor recurrence and treatment resistance, however, there are limited reports yet concerning on the CD44 expression and its clinical prognostic significance in SCLC patients. The purpose of this study is to investigate the expression of CD44 in tumor tissues as well as serum of SCLC patients and explore its correlation with the clinical characteristics, therapeutic effect and prognosis. METHODS: The tumor tissues and serum samples of 47 newly diagnosed SCLC patients were collected. Immunohistochemistry and enzyme-linked immunosorbent assay were applied to detect CD44. The relationship between CD44 level and the clinical characteristics as well as prognosis of the patients was analyzed. RESULTS: The stem cell marker CD44 was detectable both in serum sample and tumor tissue of SCLC patients. The positive rate of CD44 in tumor tissue was significantly higher in patients with performance status (PS) 2 than that of patients with PS 0-1 (85.71% vs 30%, P=0.017). Patients were divided in to different groups according to the treatment efficacy. The CD44 immunohistochemical score and serum level in the disease progression group were significantly higher than those in the disease control group, and the differences were statistically significant (P=0.006, P=0.034), Univariate analysis depicted that the progression-free survival (PFS) of CD44 positive patients was significantly shorter than that of CD44 negative patients (5.23 mon vs 9.03 mon, P=0.036). CONCLUSIONS: The positive expression of CD44 in tumor tissues of pre-treatment SCLC patients is correlated with poor PFS. The clinical significance of CD44 is worthy to be further studied.

Antihuman CD44 antibody BJ18 inhibits platelet phagocytosis by correcting aberrant FcɣR expression and M1 polarization in immune thrombocytopenia.

BACKGROUND: Immune thrombocytopenia (ITP) is an autoimmune hemorrhagic disease with a low platelet count. CD44 is a pivotal component involved in phagocytosis and inflammation, and monoclonal antibodies (mAbs) against CD44 have been shown to be beneficial in several autoimmune diseases. In the present study, we investigated the correlation between CD44 levels and disease severity in patients with ITP and explored the immunomodulatory mechanisms of the antihuman CD44 mAb BJ18 on platelet phagocytosis mediated by monocytes/macrophages. METHODS: Plasma was collected from 45 participants to measure the circulating concentration of CD44 using ELISA. Peripheral blood mononuclear cells from patients and controls were isolated and induced to differentiate into monocytes/macrophages utilizing cytokines and drugs. CD44 expression on circulating cells and the effects of BJ18 on platelet phagocytosis, Fcɣ receptor (FcɣR) expression and M1/M2 polarization of macrophages were evaluated using flow cytometry and qPCR. RESULTS: CD44 levels of both the soluble form found in plasma and the form expressed on the surface of circulating monocytes/macrophages were significantly elevated in ITP patients. Linear correlations were verified between the CD44 levels and major clinical characteristics. In an in vitro study, BJ18 successfully inhibited platelet phagocytosis by monocytes/macrophages obtained from ITP patients. Further studies indicated that BJ18 corrected abnormal FcγR expression on monocytes/macrophages. Moreover, the polarization of proinflammatory M1 macrophages could also be regulated by BJ18. CONCLUSIONS: Our data indicated that the CD44 level has potential predictive value for disease severity and that the antihuman CD44 mAb BJ18 may be a promising therapy for ITP patients.

Analysis of Serum miRNA in Glioblastoma Patients: CD44-Based Enrichment of Extracellular Vesicles Enhances Specificity for the Prognostic Signature.

Glioblastoma is a devastating disease, for which biomarkers allowing a prediction of prognosis are urgently needed. microRNAs have been described as potentially valuable biomarkers in cancer. Here, we studied a panel of microRNAs in extracellular vesicles (EVs) from the serum of glioblastoma patients and evaluated their correlation with the prognosis of these patients. The levels of 15 microRNAs in EVs that were separated by size-exclusion chromatography were studied by quantitative real-time PCR, followed by CD44 immunoprecipitation (SEC + CD44), and compared with those from the total serum of glioblastoma patients (n = 55) and healthy volunteers (n = 10). Compared to total serum, we found evidence for the enrichment of miR-21-3p and miR-106a-5p and, conversely, lower levels of miR-15b-3p, in SEC + CD44 EVs. miR-15b-3p and miR-21-3p were upregulated in glioblastoma patients compared to healthy subjects. A significant correlation with survival of the patients was found for levels of miR-15b-3p in total serum and miR-15b-3p, miR-21-3p, miR-106a-5p, and miR-328-3p in SEC + CD44 EVs. Combining miR-15b-3p in serum or miR-106a-5p in SEC + CD44 EVs with any one of the other three microRNAs in SEC + CD44 EVs allowed for a prognostic stratification of glioblastoma patients. We have thus identified four microRNAs in glioblastoma patients whose levels, in combination, can predict the prognosis for these patients.

CD44v3 protein-carrying tumor-derived exosomes in HNSCC patients' plasma as potential noninvasive biomarkers of disease activity.

The molecular cargo of tumor-cell-derived exosomes (TEX) mimics that of parental tumor cells. Thus, TEX could potentially serve as noninvasive biomarkers of cancer progression. However, separation of TEX from non-TEX in patients' plasma requires tumor antigen-specific detection reagents. CD44v3 has been of interest as a potential biomarker of disease progression in HNSCC, because its overexpression in tumor cells associates with poor outcome. Here, CD44v3+ TEX immunocaptured from plasma of 44 HNSCC patients and 7 healthy donors (HDs) were evaluated as potential biomarkers of disease activity and stage. Exosomes were isolated from plasma of by size exclusion chromatography. Using anti-CD44v3 or anti-CD3 mAbs on beads, CD44v3+ TEX CD3(-)TEX-enriched exosomes were immunocaptured from supernatants of nonmalignant or HNSCC cell lines and from patients' plasma. On-bead flow cytometry was used for the detection of FAS-L, PD-L1, TGFF-ß. CSPG4 or EGFR on exosomes. The TEX expression profiles were correlated to clinicopathological parameters. Relative florescence intensity (RFI) values for CD44v3 were higher (p < .01) on TEX from HNSCC cell lines or on CD44v3+ CD3(-) plasma-derived exosomes. RFI values of CD44v3 on CD3(-) exosomes were higher (p < .005) in patients than in HDs and correlated (p < .05) with the UICC stage and lymph node metastasis. In HNSCC patients, CD44v3+ exosomes higher levels of immunosuppressive proteins compared to CD44v3(-) exosomes (p < .05-p < .005), and RFI values for these markers correlated with higher disease stages and lymph node metastasis. Isolation of CD44v3+ exosomes by immunocapture allowed for enrichment of TEX which are potentially promising liquid biomarkers of the tumor burden and disease stage in HNSCC.

Single-cell transcriptomics identifies CD44 as a marker and regulator of endothelial to haematopoietic transition.

The endothelial to haematopoietic transition (EHT) is the process whereby haemogenic endothelium differentiates into haematopoietic stem and progenitor cells (HSPCs). The intermediary steps of this process are unclear, in particular the identity of endothelial cells that give rise to HSPCs is unknown. Using single-cell transcriptome analysis and antibody screening, we identify CD44 as a marker of EHT enabling us to isolate robustly the different stages of EHT in the aorta-gonad-mesonephros (AGM) region. This allows us to provide a detailed phenotypical and transcriptional profile of CD44-positive arterial endothelial cells from which HSPCs emerge. They are characterized with high expression of genes related to Notch signalling, TGFbeta/BMP antagonists, a downregulation of genes related to glycolysis and the TCA cycle, and a lower rate of cell cycle. Moreover, we demonstrate that by inhibiting the interaction between CD44 and its ligand hyaluronan, we can block EHT, identifying an additional regulator of HSPC development.

Breast Cancer Cell-Derived Soluble CD44 Promotes Tumor Progression by Triggering Macrophage IL1ß Production.

IL1ß is a central regulator of systemic inflammatory response in breast cancer, but the precise regulatory mechanisms that dictate the overproduction of IL1ß are largely unsolved. Here, we show that IL1ß secretion is increased by the coculture of human monocyte-like cells and triple-negative breast cancer (TNBC) cells. In addition, macrophages robustly produced IL1ß when exposed to the conditioned media of TNBC cells. Consistent with these observations, macrophage depletion decreased serum IL1ß and reduced breast cancer progression in an orthotopic breast cancer mouse model. Profiling the secretome of human breast cancer cells revealed that the CD44 antigen was the most differentially released protein in basal conditions of TNBC cells. Antibody-mediated neutralization of CD44 abrogated IL1ß production in macrophages and inhibited the growth of primary tumors. These results suggest IL1ß-mediated oncogenic signaling is triggered by breast cancer cell membrane-derived soluble CD44 (sCD44) antigen, and targeting sCD44 antigen may provide an alternative therapeutic strategy for breast cancer treatment by modulating inflammatory tumor microenvironment. SIGNIFICANCE: A novel positive feedback loop between IL1ß and CD44 promotes TNBC malignant progression.

Determination of soluble CD44 in serum by using a label-free aptamer based electrochemical impedance biosensor.

CD44 is a promising biomarker in the diagnosis and prognosis of malignancies. The serum CD44 level is closely related to disease progression and metastasis of malignancies. It is of great clinical significance for the detection of serum soluble CD44. In this study, a facile, label-free aptamer based electrochemical impedance sensor for serum CD44 has been proposed. The aptamer showing high affinity to CD44 was immobilized on the gold electrodes through Au-S interaction. The interaction between target CD44 and the immobilized aptamer will cause a complex structure change of the aptamer, which makes the diffusion of [Fe(CN)6]3-/4- toward the electrode surface easy, thus resulting in the decrease of the impedance of the system. The decreased degree of the impedance had a good linear relationship with the logarithm of the CD44 concentration in the range of 0.1-1000 ng mL-1 with a detection limit of 0.087 ng mL-1 (S/N = 3). The developed biosensor has been applied to detect CD44 in serum samples with satisfactory results.

Tumor-suppressive activity of sTRAIL on circulating CD44+ cells in patients with non-small cell lung cancer.

Circulating CD44+ cells have been identified as a prognostic marker for patients with non-small cell lung cancer (NSCLC). Serum tumor necrosis factor-related apoptosis-inducing ligand (sTRAIL) is involved in the pathophysiology of many cancers. However, no previous studies have shown the roles of sTRAIL in circulating CD44+ cells in the blood of NSCLC patients. We detected circulating CD44+ cells and sTRAIL levels in blood samples from NSCLC patients using flow cytometry and an enzyme-linked immunosorbent assay (ELISA). Anti-tumor roles of TRAIL in CD44+ cells were confirmed using a CCK-8 assay and mouse models. A higher number of circulating CD44+ cells were identified in NSCLC patients compared with healthy control individuals. In addition, we confirmed the anti-tumor roles and mechanisms of TRAIL in CD44+ cells both in vitro and in vivo. Our results indicate that (1) there is a negative correlation between sTRAIL and circulating CD44+ cells in NSCLC patients and (2) CD44+ cells have cancer stem cell properties and are more sensitive than CD44- cells to TRAIL.

Validation and evaluation of a common biomarker in human cancers sera protein detected by a monoclonal antibody UNIVmAb.

OBJECTIVE: Management and diagnosis of multiple human cancers remains a challenge and search for a common biomarker is still debatable. In this manuscript we have evaluated the use of monoclonal antibody UNIVmAb, to detect the protein (H11) as a common biomarker for all cancers irrespective of the grade and origin. We have shown by both ELISA and Western Blot that the H11 protein, is a unique hyaluronan binding protein that has not been detected earlier. H11 protein was fractionated in an anion exchange column followed by cibacron blue gel exclusion chromatography. Hyaluronan binding H11 protein reacted with Monoclonal antibody UNIVmAb and b-HA inspite of b-Hyaluronan (biotinylated Hyaluronan) interaction and HA-Oligo (Hyaluronan oligosaccharides) competition from various grades of Human cancers sera. RESULTS: ELISA, Western blot and b-Hyaluronan interactions clearly showed an over-expression of UNIVmAb reacted H11 protein in all fifty cancer's sera when compared with seventy normal sera. UNIVmAb reactive H11 protein can be used as a common biomarker. We believe, UNIVmAb detected H11 protein, is a unique hyaluronan binding protein, that can be used as a common biomarker for all cancers.

Decorin Secreted by Human Umbilical Cord Blood-Derived Mesenchymal Stem Cells Induces Macrophage Polarization via CD44 to Repair Hyperoxic Lung Injury.

Bronchopulmonary dysplasia (BPD), caused by hyperoxia in newborns and infants, results in lung damage and abnormal pulmonary function. However, the current treatments for BPD are steroidal and pharmacological therapies, which cause neurodevelopmental impairment. Treatment with umbilical cord blood-derived mesenchymal stem cells (UCB-MSCs) is an efficient alternative approach. To prevent pulmonary inflammation in BPD, this study investigated the hypothesis that a key regulator was secreted by MSCs to polarize inflammatory macrophages into anti-inflammatory macrophages at inflammation sites. Lipopolysaccharide-induced macrophages co-cultured with MSCs secreted low levels of the inflammatory cytokines, IL-8 and IL-6, but high levels of the anti-inflammatory cytokine, IL-10. Silencing decorin in MSCs suppressed the expression of CD44, which mediates anti-inflammatory activity in macrophages. The effects of MSCs were examined in a rat model of hyperoxic lung damage. Macrophage polarization differed depending on the levels of decorin secreted by MSCs. Moreover, intratracheal injection of decorin-silenced MSCs or MSCs secreting low levels of decorin confirmed impaired alveolarization of damaged lung tissues by down-regulation of decorin. In tissues, a decrease in the anti-inflammatory macrophage marker, CD163, was observed via CD44. Thus, we identified decorin as a key paracrine factor, inducing macrophage polarization via CD44, a master immunoregulator in mesenchymal stem cells.

The Expressions of CD44, CD90 and Alpha Fetoprotein Biomarkers in Indonesian Patients with Advanced Liver Disease: an Observational Study.

BACKGROUND: increased serum alpha fetoprotein (AFP) levels are often found in patients with advanced hepatocellular carcinoma (HCC). Cluster Differentiation 44 (CD44) and CD90 are stem cell biomarkers that have been assumed as the early HCC markers and associated with onset and progressivity of HCC. The study related to HCC stem cell has not been available in Indonesia. The present study aimed to evaluate the expression of cancer stem cell markers (CD44, CD90) and AFP levels in patients with advanced liver disease. METHODS: an observational study was conducted in 41 patients with chronic hepatitis B and/or C infection, liver cirrhosis, and HCC at dr. Saiful Anwar General Hospital. CD44 and CD90 expressions were measured with flow cytometry, and AFP serum levels with ELISA. Data on patient characteristics were evaluated using bivariate and multivariate statistical analysis (One-way ANOVA, Mann-Whitney, Chi-Square, Kruskal-Wallis). Data of CD44, CD90 and AFP were analyzed using Kruskal Wallis test with a significance value of p<0.05, and diagnostic power was analyzed using receiver operating characteristic (ROC). RESULTS: the subjects of our study were 16 patients with chronic hepatitis, 15 patients with liver cirrhosis, and 10 patients with HCC. There was a significant difference regarding CD44+CD90+ and AFP among those three groups (p=0.001; p=0.000) specifically in chronic hepatitis compared to liver cirrhosis (p=0.002; p=0.000) and HCC (p=0.002; p=0.000) respectively. ROC analysis showed the best diagnostic power for the combination of CD44+CD90+ and AFP (AUC=0.981; p=0.000). CONCLUSION: there are higher expressions of CD44+CD90+ and serum AFP levels in patients with HCC compared to the other two groups (those with chronic hepatitis and liver cirrhosis). The combination of both parameters has the best diagnostic power of HCC.

CD44, TGM2 and EpCAM as novel plasma markers in endometrial cancer diagnosis.

BACKGROUND: Endometrial cancer (EC) is the most common malignancy of the female reproductive tract. Despite years of research, the accurate screening strategy is still not available in this disease and it is usually diagnosed only after the clinical signs are present. The recent technological advances in analytical methodologies enabled detection of multiple molecules in one, small sample of biological materials. Such approach was undertaken in the presented study. METHODS: Concentrations of aldehyde dehydrogenase 1 family, member A1 (ALDH1A1), carbonic anhydrase IX (CA9), CD44, epithelial cell adhesion molecule (EpCAM), hepsin, kallikrein-6, mesothelin, midkine, neural cell adhesion molecule L1 (L1CAM), and transglutaminase 2 (TGM2) were measured using MAGPIX®System in plasma samples of 45 EC, 20 healthy controls and 11 patients with endometriosis. RESULTS: Significantly increased concentration in EC as compared to healthy controls were found in case of CD44 (p <  0.001), EpCAM (p = 0.033) and TGM2 (p <  0.001). EpCAM and mesothelin concentrations differed based on FIGO stages. Regression analysis revealed marker panels with high accuracy in detection of EC. The highest AUC 0.937 was attributed to the 3-marker panel of CD44/TGM2/EpCAM (84% sensitivity, 100% specificity), FIGO IA samples were discriminated from more advanced stages of EC with the mesothelin/grade 1 model featuring AUC of 0.911 (95.24% sensitivity, 78.26% specificity). CONCLUSIONS: Novel plasma biomarkers presenting good accuracy in diagnosing EC were found with TGM2 reported for the first time as plasma marker. It was also revealed that endometriosis may share similarities in the pattern of markers alterations characteristic for EC.

Label-Free Electrochemical Sensor for CD44 by Ligand-Protein Interaction.

Detection of biomarkers in biosystems plays a key role in advanced biodiagnostics for research and clinical use. Design of new analytical platforms is challenging and in demand, addressing molecular capture and subsequent quantitation. Herein, we developed a label-free electrochemical sensor for CD44 by ligand-protein interaction. We assembled carbon nanotube composites on the electrode to enhance electronic conductivity by 6.2-fold and reduce overpotential with a shift of 77 mV. We conjugated hyaluronic acid (HA) to the surface of carbon nanotubes via electrostatic interaction between HA and poly(diallyldimethylammonium chloride) (PDDA). Consequently, we performed direct electrochemical sensing of CD44 with a dynamic range of 0.01-100 ng/mL and detection limit of 5.94 pg/mL without any postlabeling for amplification, comparable to the best current results. The sensor also displayed high selectivity, reproducibility with relative standard deviation (RSD, n = 5) of 2.57%, and long-term stability for 14 days. We demonstrated applications of the sensor in detection of human serum and cancer cells. Our work guides the development of more sensor types by ligand-protein interactions and contributes to design of interfaces in given biosystems for diagnosis.

VEGF, Microvessel Density, and CD44 as Inflammation Markers in Peri-implant Healthy Mucosa, Peri-implant Mucositis, and Peri-implantitis: Impact of Age, Smoking, PPD, and Obesity.

Several biologic processes affect the supporting peri-implant tissue leading to implant failure and complications, mainly referred to inflammation that is still poorly investigated in the peri-implant soft tissues. Our aim was to investigate in peri-implant healthy mucosa, peri-implant mucositis, and peri-implantitis the expression of some angiogenesis markers highly associated with inflammation, and evaluate its relationships with age, smoking, peri-implant pocket depth (PPD), and body max index (BMI). Moreover, we wanted to study the impact of these clinical parameters in the disease pathogenesis. Forty-eight total patients were recruited. Sixteen had at least one successfully osteointegrated dental implant (group A) and 32 had at least one osseointegrated implant in need of a peri-implant treatment for inflammatory/infectiveous reasons: precisely 16 for mucositis (group B) and 16 for peri-implantitis (group C). VEGF, CD34, and CD44 immunohistochemical expression was evaluated in the interproximal biopsies of marginal peri-implant tissue and correlated with the clinical parameters. A significant difference between groups in mean PPD was found, while the distribution by age, gender, smoking, and BMI resulted similar. Group C had significantly higher levels of VEGF, CD34, and CD44 expression compared to the other groups. VEGF, CD34, CD44, and peri-implant pocket depth were all positively correlated. Our study revealed that peri-implantitis is a condition characterized by unique and distinctive features. Our results supported that PPD has a great impact on the peri-implantitis and it is closely related to the inflammation marker expression. The identification of specific biomarkers might help in choosing distinct treatment approaches for target individuals.

Cancer stem cell markers ALDH1 and CD44+/CD24- phenotype and their prognosis impact in invasive ductal carcinoma.

Breast cancer is a very heterogeneous disease. The intrinsic molecular subtypes can explain the intertumoral heterogeneity and the cancer stem cell (CSC) hypothesis can explain the intratumoral heterogeneity of this kind of tumor. CD44+/CD24- phenotype and ALDH1 expression are the major CSC markers described in invasive breast cancer. In the present study, 144 samples of invasive breast carcinoma, no special type were distributed in 15 tissue microarrays (TMA) and then evaluated for expression of the CD44+/CD24- phenotype and ALDH1 to understand the importance of these CSC markers and the clinical aspects of breast cancer. The samples were classified into four molecular subtypes according to clinicopathological criteria: Luminal A, Luminal B, HER2, and Basal-like. A statistical association was found between the molecular subtypes and the CSC markers, with HER2 the most frequent subtype for both markers. ALDH1 was also associated with other poor prognostic variables, such as a high histological grade and larger tumors, but it was not associated with the patients' prognosis in this sample and nor was the CD44+/CD24- phenotype in a multivariate analysis. There are still many controversies about the role of these markers in breast cancer molecular subtypes. The identification of these populations of cells, through immunohistochemical markers, can help to better understand the CSC theory in clinical practice and, in the near future, contribute to developing new target therapies.

Prognostic significance of elevated serum CD44 levels in patients with oral squamous cell carcinoma.

BACKGROUND: Availability of reliable methods distinguishing high-risk recurrent tumours from regressive tumours prior to surgery could help in better management of the disease. This study was aimed to estimate pre-surgical serum CD44 concentration and assess the possibility of using it as a non-invasive prognostic tool in oral cancer. METHODS: ELISA was performed on pre-surgical serum samples from 64 primary oral cancer patients and 16 healthy individuals to estimate soluble CD44 levels. Immunohistochemistry was performed on parallel 64 solid tumours and 10 recurrent tumours. All patients clinically followed up for median period of 19.2 months and obtained prognostic information correlated with CD44 concentration in serum as well as in tumours. RESULTS: Serum CD44 concentration was found significantly high in patients as compared to healthy individuals (P < .001) and also in patients whose disease locally recurred as compared to those did not recur (P = 0026). High serum CD44 concentration inversely affected on patients survival (P = .032). CD44v6 staining intensity was detected significantly high in recurrent tumours as compared to primary tumours (P < .001), and it also correlated with poor survival (P < .001). Furthermore, in combination, patients with increased CD44 concentration in serum and CD44v6 expression in tumours significantly correlated with local recurrence (P < .001) and poor survival (P < .001). CONCLUSION: Our data suggest that the ELISA-based estimation of pre-surgical serum CD44 concentration could be a non-invasive reliable method distinguishing high-risk recurrent tumours which can further assist in post-surgery treatment planning.

Glioblastoma-secreted soluble CD44 activates tau pathology in the brain.

During aggressive tumor growth and migration, glioblastoma cells secrete diverse molecules and adhesion proteins to the extracellular matrix. Yet, the biochemical effects of the glioblastoma secretome in the brain remain largely unknown. Here we show that soluble CD44 secreted from glioblastoma cells induces neuronal degeneration through the activation of tau pathology in the brain. Glioblastoma-xenograft tissues showed a number of degenerating neurons bearing highly phosphorylated tau. Through a series of secretome-analyses, we identified that soluble CD44 was the responsible protein inducing tau phosphorylation and aggregation (EC50 = 19.1 ng/mL). The treatment of sCD44 to primary hippocampal neurons-induced tau hyperphosphorylation, leading to neuronal degeneration. Also, the injection of sCD44 into the brains of tau transgenic mice induced tau hyper-phosphorylation in hippocampal neurons. Altogether, our data suggest a neurodegenerative role of sCD44 in promoting tau pathology and serving as a molecular link between glioblastoma and neurodegeneration.

Uncovering the potential of CD44v/SYNE1/miR34a axis in salivary fluids of oral cancer patients.

OBJECTIVES: Late-stage diagnosis is one of the major confounders for poor prognosis of patients with oral cancer owing to lack of a biomarker to diagnose this disease at an early stage. Moreover, till date, invasive biopsies are the only option to assess disease occurrence and progression in this malignancy. Thus, this study aims to identify and assess potential salivary markers in OSCC patients in order to open newer avenues in the field of non-invasive biopsies. METHODOLOGY: Bioinformatic-based analysis was performed to identify potential biomarkers that could be assessed in OSCC patients. The expression patterns of CD44v and its genetic and epigenetic modulators were assessed in saliva of OSCC patients, leukoplakia, and controls using real-time and methylation-specific PCR. Statistical analysis was conducted to understand the significance of these markers in terms of their clinical relevance. RESULTS: CD44v/SYNE1/miR34a axis was identified using bioinformatic analysis, and the expression profile of these markers was assessed in saliva of OSCC patients. CD44v6 and CD44v10 demonstrated a significantly increased expression, whereas SYNE1 and miR34a depicted a significantly decreased expression in OSCC patients. Statistical analysis suggested a probable role of CD44v6, SYNE1, and miR34a in early stages of the malignancy, whereas a strong association was observed between CD44v6, CD44v10, and miR34a expression with locoregional aggressiveness and histopathological conditions. CONCLUSION: Collectively, these findings suggested a plausible role of CD44v/SYNE1/miR34a axis as non-invasive salivary biomarkers to diagnose this disease at an early stage and predict the early onset of metastasis.

Unfavorable clinical implications of peripheral blood CD44+ and CD54+ lymphocytes in patients with lung cancer undergoing chemotherapy.

BACKGROUND: There is an unmet need for identification of additional prognostic markers for lung cancer. The aim of this study was to identify novel clinical and immunological predictors of prognosis in lung cancer patients. METHODS: Lymphocyte subsets CD3+, CD4+, CD8+, CD4+/8+, CD25+, CD69+, CD44+ and CD54+ were quantified in peripheral blood using flow cytometry, for 203 newly diagnosed lung cancer patients and 120 healthy controls. RESULTS: The levels of CD3+, CD4+, CD8+, CD4+/CD8+ and CD69+ lymphocytes were significantly lower in patients with lung cancer compared with the healthy control group, while CD54+ and CD44+ lymphocytes were significantly higher. In stage III/IV patients with lymph node metastasis or distant metastasis, the levels of CD44+ and CD54+ lymphocytes were significantly increased compared with patients with stage I/II disease (p<0.05). The levels of CD44+ and CD54+ lymphocytes markedly reduced after chemotherapy, and follow-up analysis indicated that patients found without increase of CD44+ and CD54+ lymphocytes after chemotherapy had survival advantages. Independent predictors of survival in lung cancer patients included clinical stage (hazard ratio [HR] = 2.791; 95% confidence interval [95% CI], 1.42-3.54, p<0.001), CD44+ lymphocytes (HR = 1.282; 95% CI, 1.02-1.49, p = 0.002) and CD54+ lymphocytes (HR = 1.475; 95% CI, 1.22-1.73, p = 0.003). Elevated levels of CD44+ and CD54+ lymphocytes correlated with poor prognosis in lung cancer patients. CONCLUSIONS: Peripheral blood lymphocyte subsets in patients with lung cancer are different from those in healthy people, and circulating CD44+ and CD54+ lymphocytes seem to be a promising criterion to predict survival in lung cancer patients undergoing chemotherapy.