Interleukin-34 is more suitable than macrophage colony-stimulating factor for predicting liver significant fibrosis in patients with chronic hepatitis B.

AIMS: Interleukin-34 (IL-34) and macrophage colony-stimulating factor (CSF-1) have similar functions, such as promoting the formation of liver fibrosis. This study aimed to evaluate and compare the diagnostic value of serum IL-34 and CSF-1 for significant liver fibrosis in patients with chronic hepatitis B (CHB). METHODS: A total of 369 CHB patients, consisting of 208 HBeAg-negative patients and 161 HBeAg-positive patients, were enrolled in this study. Additionally, 72 healthy individuals served as healthy controls (HCs). Serum levels of IL-34 and CSF-1 were measured using the enzyme-linked immunosorbent assay method. Liver fibrosis grades were assessed using the modified Scheuer scoring system. RESULTS: Serum IL-34 and CSF-1 levels exhibited significant elevation in both HBeAg-negative and HBeAg-positive patients in comparison to HCs (p < 0.001). IL-34 emerged as an independent factor linked to significant liver fibrosis, whereas CSF-1 did not exhibit such an association. Receiver operating characteristic (ROC) analysis indicated higher areas under the curves (AUCs) for IL-34 (0.814, p < 0.001 and 0.673, p < 0.001) when diagnosing significant liver fibrosis in HBeAg-negative and HBeAg-positive patients, respectively, as opposed to CSF-1 (0.602, p < 0.001; 0.619, p = 0.385). Within the HBeAg-negative patient subgroup, the AUC for IL-34 surpassed that of FIB-4 (p = 0.009) and APRI (p = 0.045). CONCLUSION: Serum IL-34 has the potential to be a straightforward and practical biomarker that demonstrates superior performance to serum CSF-1 in the diagnosis of significant liver fibrosis in CHB patients, especially within the HBeAg-negative patients.

The diagnostic accuracy of macrophage colony-stimulating factor for cervical cancer: A systematic review and meta-analysis.

BACKGROUND: Macrophage colony-stimulating factor (M-CSF) overexpression in plasma levels serves as a useful predictor of carcinogenesis and poor prognosis. Thus, we aimed to investigate the diagnostic performance of M-CSF for cervical cancer. METHODS: A comprehensive search of eligible studies was performed in PubMed/MEDLINE, Cochrane Library, Google Scholar, Scopus, Web of Science, and EMBASE published until October 2021. For statistical analysis, Meta-disc software is used. The pooled sensitivity, specificity, positive likelihood ratio, negative likelihood ratio, diagnostic odds ratio, 95% confidence interval (CI), and the area under the curve were estimated. A P value <.05 was considered statistically significant. RESULTS: A total of 5 studies were included in this systematic review and meta-analysis. The result showed that the pooled sensitivity and specificity of M-CSF to diagnose cervical cancer were 70% (95% CI: 66%-74%) and 84% (95% CI: 80%-88%), respectively. The summary positive likelihood ratio and negative likelihood ratio were 4.41 (95% CI: 2.86-6.82) and 0.36 (95% CI: 0.29-0.45), respectively. The area under the curve of the summary receiver operating characteristic curve was 0.86 (95% CI: 0.83-0.88), indicating the excellent diagnostic performance of M-CSF for cervical cancer. CONCLUSIONS: The results of this study showed that M-CSF has diagnostic value for the early detection of cervical cancer. As a result, M-CSF can be utilized in conjunction with existing test platforms to diagnose cervical cancer.

Systemic Immune Dysregulation in Early Breast Cancer Is Associated With Decreased Plasma Levels of Both Soluble Co-Inhibitory and Co-Stimulatory Immune Checkpoint Molecules.

Breast cancer cells exploit the up-regulation or down-regulation of immune checkpoint proteins to evade anti-tumor immune responses. To explore the possible involvement of this mechanism in promoting systemic immunosuppression, the pre-treatment levels of soluble co-inhibitory and co-stimulatory immune checkpoint molecules, as well as those of cytokines, chemokines, and growth factors were measured in 98 newly diagnosed breast cancer patients and compared with those of 45 healthy controls using multiplex bead array and ELISA technologies. Plasma concentrations of the co-stimulatory immune checkpoints, GITR, GITRL, CD27, CD28, CD40, CD80, CD86 and ICOS, as well as the co-inhibitory molecules, PD-L1, CTLA-4 and TIM-3, were all significantly lower in early breast cancer patients compared to healthy controls, as were those of HVEM and sTLR-2, whereas the plasma concentrations of CX3CL1 (fractalkine), CCL5 (RANTES) and those of the growth factors, M-CSF, FGF-21 and GDF-15 were significantly increased. However, when analyzed according to the patients' breast cancer characteristics, these being triple negative breast cancer (TNBC) vs. non-TNBC, tumor size, stage, nodal status and age, no significant differences were detected between the plasma levels of the various immune checkpoint molecules, cytokines, chemokines and growth factors. Additionally, none of these biomarkers correlated with pathological complete response. This study has identified low plasma levels of soluble co-stimulatory and co-inhibitory immune checkpoint molecules in newly diagnosed, non-metastatic breast cancer patients compared to healthy controls, which is a novel finding seemingly consistent with a state of systemic immune dysregulation. Plausible mechanisms include an association with elevated levels of M-CSF and CCL5, implicating the involvement of immune suppressor cells of the M2-macrophage/monocyte phenotype as possible drivers of this state of systemic immune quiescence/dysregulation.

Multi-Modal Characterization of Monocytes in Idiopathic Pulmonary Fibrosis Reveals a Primed Type I Interferon Immune Phenotype.

Idiopathic pulmonary fibrosis (IPF) is the most severe form of chronic lung fibrosis. Circulating monocytes have been implicated in immune pathology in IPF but their phenotype is unknown. In this work, we determined the immune phenotype of monocytes in IPF using multi-colour flow cytometry, RNA sequencing and corresponding serum factors, and mapped the main findings to amount of lung fibrosis and single cell transcriptomic landscape of myeloid cells in IPF lungs. We show that monocytes from IPF patients displayed increased expression of CD64 (FcγR1) which correlated with amount of lung fibrosis, and an amplified type I IFN response ex vivo. These were accompanied by markedly raised CSF-1 levels, IL-6, and CCL-2 in serum of IPF patients. Interrogation of single cell transcriptomic data from human IPF lungs revealed increased proportion of CD64hi monocytes and "transitional macrophages" with higher expression of CCL-2 and type I IFN genes. Our study shows that monocytes in IPF patients are phenotypically distinct from age-matched controls, with a primed type I IFN pathway that may contribute to driving chronic inflammation and fibrosis. These findings strengthen the potential role of monocytes in the pathogenesis of IPF.

Cytokine ranking via mutual information algorithm correlates cytokine profiles with presenting disease severity in patients infected with SARS-CoV-2.

Although the range of immune responses to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is variable, cytokine storm is observed in a subset of symptomatic individuals. To further understand the disease pathogenesis and, consequently, to develop an additional tool for clinicians to evaluate patients for presumptive intervention, we sought to compare plasma cytokine levels between a range of donor and patient samples grouped by a COVID-19 Severity Score (CSS) based on the need for hospitalization and oxygen requirement. Here we utilize a mutual information algorithm that classifies the information gain for CSS prediction provided by cytokine expression levels and clinical variables. Using this methodology, we found that a small number of clinical and cytokine expression variables are predictive of presenting COVID-19 disease severity, raising questions about the mechanism by which COVID-19 creates severe illness. The variables that were the most predictive of CSS included clinical variables such as age and abnormal chest x-ray as well as cytokines such as macrophage colony-stimulating factor, interferon-inducible protein 10, and interleukin-1 receptor antagonist. Our results suggest that SARS-CoV-2 infection causes a plethora of changes in cytokine profiles and that particularly in severely ill patients, these changes are consistent with the presence of macrophage activation syndrome and could furthermore be used as a biomarker to predict disease severity.

Interleukin 6, soluble interleukin 2 receptor alpha (CD25), monocyte colony-stimulating factor, and hepatocyte growth factor linked with systemic hyperinflammation, innate immunity hyperactivation, and organ damage in COVID-19 pneumonia.

BACKGROUND: Patients infected by SARS-CoV-2 can develop interstitial pneumonia, requiring hospitalisation or mechanical ventilation. Increased levels of inflammatory biomarkers are associated with development of acute respiratory distress syndrome (ARDS). The aim of the present study was to determine which cytokines are associated with respiratory insufficiency in patients hospitalised for COVID-19. PATIENTS AND METHODS: Data on 67 consecutive patients were collected between March 8 and March 30, 2020. PaO2/FiO2 ratio (P/F) was calculated at hospital admission. The following cytokines were analysed: interleukin (IL)-6, IL-1α, IL-18, tumour necrosis factor (TNF)-ß, macrophage colony-stimulating factor (M-CSF), macrophage migration inhibitory factor (MIF), soluble IL-2 receptor alpha (sIL-2Rα; CD25), IL-12ß, IL-3, interferon (IFN) α2a, monokine induced by gamma interferon (MIG), monocyte-chemotactic protein 3 (MCP3) and hepatocyte growth factor (HGF). RESULTS: P/F lower than 300 was recorded in 22 out of 67 patients (32.8%). P/F strongly correlated with IL-6 (r = -0.62, P < 0.0001), M-CSF (r = -0.63, P < 0.0001), sIL-2Rα (r = -0.54, P < 0.0001), and HGF (r = -0.53, P < 0.0001). ROC curve analyses for IL-6 (AUC 0.83, 95% CI 0.73-0.93, P < 0.0001), M-CSF (AUC 0.87, 95% CI 0.79-0.96, P < 0.0001), HGF (AUC 0.81, 95% CI 0.70-0.93, P < 0.0001), and sIL-2Rα (AUC 0.80, 95% CI, 0.69-0.90, P < 0.0001) showed that these four soluble factors were highly significant. All four soluble factors correlated with LDH, white blood cell count, neutrophil count, lymphocyte count, and CRP. CONCLUSION: IL-6, M-CSF, sIL-2Rα, and HGF are possibly involved in the main biological processes of severe COVID-19, mirroring the level of systemic hyperinflammatory state, the level of lung inflammation, and the severity of organ damage.

Deficiency of activation-induced cytidine deaminase in a murine model of ulcerative colitis.

Patients with inflammatory bowel disease (IBD) have an increased risk of colorectal cancer, particularly in ulcerative colitis (UC) when the majority of colon epithelial cells may be exposed to inflammation-associated mutagenesis. In addition to mutagenesis generated by oxidative stress, inflammation can induce activation-induced cytidine deaminase (Aicda), a mutator enzyme in the APOBEC family, within colon epithelial cells. This study tested the hypothesis that deletion of the Aicda gene could protect against the development of inflammation-associated colorectal cancers, using a model of UC-like colitis in "T/I" mice deficient in TNF and IL10. Results showed that T/I mice that were additionally Aicda-deficient ("TIA" mice) spontaneously developed moderate to severe UC-like colitis soon after weaning, with histologic features and colon inflammation severity scores similar those in T/I mice. Although the mean survival of TIA mice was decreased compared to T/I mice, multivariable analysis that adjusted for age when neoplasia was ascertained showed a decreased numbers of neoplastic colorectal lesions in TIA mice, with a trend toward decreased incidence of neoplasia. Aicda deficiency increased serum IL1α and slightly decreased IL12p40 and M-CSF, as compared with T/I mice, and led to undetectable levels of IgA, IgG1, IgG2a, IgG2b, and IgG3. Taken together, these studies show that Aicda deficiency can decrease the number of neoplastic lesions but is not sufficient to prevent the risk of inflammation-associated colorectal neoplasia in the setting of severe UC-like inflammation. The TIA model may also be useful for assessing the roles of antibody class-switch recombination deficiency and somatic hypermutation on regulation of microbiota and inflammation in the small intestine and colon, as well as the pathogenesis of colitis associated with hyper-IgM syndrome in humans. Further studies will be required to determine the mechanisms that drive early mortality in TIA mice.

Diagnostic Values of miR-21, miR-124, and M-CSF in Patients With Early Cervical Cancer.

OBJECTIVE: This study aimed to investigate the diagnostic values of microRNA-21, microRNA-124, and macrophage colony-stimulating factor in patients with cervical cancer. METHODS: A total of 68 patients with cervical cancer admitted in our hospital (cervical cancer group) and 57 healthy individuals undergoing physical examinations (healthy group, also control group) were enrolled in this study. The expression of serum microRNA-21 and microRNA-124 was detected by quantitative reverse transcription polymerase chain reaction. The expression of serum macrophage colony-stimulating factor was detected by enzyme-linked immunosorbent assay. The diagnostic values of microRNA-21, microRNA-124, and macrophage colony-stimulating factor in cervical cancer were analyzed. The correlations between the expression of microRNA-21 and microRNA-124 with that of macrophage colony-stimulating factor were also analyzed. RESULTS: Compared to those in the healthy group, patients in the cervical cancer group had a higher expression of microRNA-21 and macrophage colony-stimulating factor (P < .05) but lower expression of microRNA-124 (P < .05). The expression of microRNA-21, microRNA-124, and macrophage colony-stimulating factor in the patients correlated with the tumor size, tumor node metastasis (TNM) staging, tumor differentiation, and the presence or absence of lymph node metastasis and human papillomavirus infection (P < .05). According to the receiver operating characteristic curves, the area under the curve of microRNA-21 for diagnosing cervical cancer was 0.723, the specificity was 58.82%, and the sensitivity was 91.23%. The area under the curve of microRNA-124 was 0.766, the specificity was 94.12%, and the sensitivity was 57.89%. The area under the curve of macrophage colony-stimulating factor was 0.754, the specificity was 64.71%, and the sensitivity was 87.72%. Pearson correlation analysis showed that the expression of microRNA-21 positively correlated with that of macrophage colony-stimulating factor (r = 0.6825, P < .001), and the expression of microRNA-124 negatively correlated with that of macrophage colony-stimulating factor (r = -0.6476, P < .001). CONCLUSION: MicroRNA-21, microRNA-124, and macrophage colony-stimulating factor may be involved in the development and progression of cervical cancer. The detection of serum microRNA-21, microRNA-124, and macrophage colony-stimulating factor has good sensitivity and specificity in the diagnosis of cervical cancer.

Genetic Association and Expression Correlation between Colony-Stimulating Factor 1 Gene Encoding M-CSF and Adult-Onset Still's Disease.

Adult-onset Still's disease (AOSD) is a rare and inflammatory disorder characterized by spiking fever, rash, arthritis, and multisystemic involvement. HLA has been shown to be associated with AOSD; however, it could not explain the innate immunity and autoinflammatory characteristics of AOSD. To assess the genetic susceptibility of AOSD, we conducted a genome-wide association study (GWAS) on a cohort of 70 AOSD cases and 688 controls following a replication study of 36 cases and 200 controls and meta-analysis. The plasma concentrations of associated gene product were determined. The GWAS, replication, and combined sample analysis confirmed that SNP rs11102024 on 5'-upstream of CSF1 encoding macrophage colony-stimulating factor (M-CSF) was associated with AOSD (P = 1.20 × 10-8, OR (95% CI): 3.28 (2.25~4.79)). Plasma levels of M-CSF increased in AOSD patients (n = 82, median: 9.31 pg/mL), particularly in the cases with activity score ≥ 6 (n = 42, 10.94 pg/mL), compared to the healthy donors (n = 68, 5.31 pg/mL) (P < 0.0001). Patients carrying rs11102024TT genotype had higher M-CSF levels (median: 20.28 pg/mL) than those with AA genotype (6.82 pg/mL) (P < 0.0001) or AT genotype (11.61 pg/mL) (P = 0.027). Patients with systemic pattern outcome were associated with elevated M-CSF and frequently observed in TT carriers. Our data suggest that genetic variants near CSF1 are associated with AOSD and the rs11102024 T allele links to higher M-CSF levels and systemic outcome. These results provide a promising initiative for the early intervention and therapeutic target of AOSD. Further investigation is needed to have better understandings and the clinical implementation of genetic variants nearby CSF1 in AOSD.

Diagnostic Power of Cytokine M-CSF, Metalloproteinase 2 (MMP-2) and Tissue Inhibitor-2 (TIMP-2) in Cervical Cancer Patients Based on ROC Analysis.

Macrophage colony-stimulating factor (M-CSF), matrix metalloproteinase-2 (MMP-2) and its specific tissue inhibitor (TIMP-2) may play an important role in the pathogenesis of cancer disease. We investigated the plasma levels and diagnostic power (ROC curve analysis) of M-CSF, MMP-2, TIMP-2 and tumor markers CA 125 and SCC-Ag in cervical cancer (CC) patients as compared to control group. The study included 89 patients with cervical cancer. The control group consisted of 50 healthy, untreated women. The plasma levels of M-CSF, MMP-2 and TIMP-2 were determined using ELISA, CA 125 and SCC-Ag - by CMIA method. The median levels of M-CSF, TIMP-2, SCC-Ag and CA 125 in the entire group of CC were significantly different than compared to the healthy women group. MMP-2 showed the highest value of sensitivity from all examined parameters (in stage I of CC - 93.10%, II - 82.76%, III and IV - 96.88%, total group - 92.05%). The highest specificity was obtained by M-CSF (86%). The area under the ROC curve (AUC) of M-CSF (0.8051) was the largest of all the tested parameters (even higher than commonly used tumor markers) in the group of cervical cancer. The combination of M-CSF, MMP-2 or TIMP-2 with SCC antigen resulted in an increase AUCs in all cases (0.8760;0.7880;0.8081;respectively). The findings of this study suggest the usefulness of all examined parameters in the diagnostics of CC patients. Out of the tested substances, M-CSF also appears to be the best candidate for cancer diagnostics in all stages of the disease, based on ROC analysis.

Plasma Levels and Tissue Expression of Selected Cytokines, Metalloproteinases and Tissue Inhibitors in Patients With Cervical Cancer.

BACKGROUND: Cytokines, metalloproteinases (MMPs) and their tissue inhibitors (TIMPs) take part in many processes involved in tumor progression and invasion such as degradation of the extracellular matrix, influence on immune cells associated with tumor tissue, and angiogenesis. Thus, the aim of this study was to compare the concentration of plasma levels and tissue expression of macrophage colony-stimulating factor (M-CSF), vascular endothelial growth factor (VEGF), matrix metalloproteinases (MMP)-2 and MMP9, and their tissue inhibitors TIMP1 and TIMP2 in patients with cervical cancer, patients with high-grade cervical intraepithelial dysplasia (CIN3) and patients with ectropion. PATIENTS AND METHODS: Concentration and expression of all tested parameters was measured in serum with enzyme-linked immunosorbent assay (ELISA) and in tissue with immunohistochemistry method. RESULTS: The epithelial expression of M-CSF and TIMP1 in cancer tissue was much stronger as compared to that in ectropion and CIN3. In the case of MMP2, lack of or weak expression in epithelial cells was observed in all tested groups. Our studies showed statistical differences of tested parameters in tissue expression and in plasma concentrations in patients with cervical cancer, patients with CIN3 and patients with ectropion. Moreover, data revealed positive correlation between plasma level and cervical cancer cell expression of VEGF. CONCLUSION: Our findings indicate a potential role of all the proteins tested here in cervical cancer diagnosis, especially VEGF. However, further studies will show whether they play a role in the progression of cancerous changes in epithelial tissue of the cervix.

Spring peaks and autumn troughs identified in peripheral inflammatory markers during the peripartum period.

Seasonal variations have recently been described in biomarkers, cell types, and gene expression associated with the immune system, but so far no studies have been conducted among women in the peripartum period. It is of note that pregnancy complications and outcomes, as well as autoimmune diseases, have also been reported to exhibit seasonal fluctuations. We report here a clear-cut seasonal pattern of 23 inflammatory markers, analysed using proximity-extension assay technology, in pregnant women. The inflammatory markers generally peaked in the spring and had a trough in the autumn. During the postpartum period we found seasonality in one inflammatory marker, namely monocyte chemotactic protein 4 (MCP-4). Our findings suggest that seasonal variations in peripheral inflammatory markers are only observed during pregnancy. The results of this study could be valuable to professionals working within the field of immunology-related areas, and provide insight for the understanding of obstetric complications.

Association of plasma macrophage colony-stimulating factor with cardiovascular morbidity and all-cause mortality in chronic hemodialysis patients.

BACKGROUND: Cardiovascular disease (CVD) events are the main cause of death in long-term hemodialysis (HD) patients. Macrophage colony- stimulating factor (M-CSF) is actively involved in the formation of atherosclerosis and causes plaque instability, thrombosis and the development of acute coronary syndromes. However, little information is available on the role of M-CSF in HD patients. We aimed to investigate the association between plasma M-CSF levels and CVD events as well as all-cause mortality in patients undergoing long-term HD. METHODS: Fifty two HD patients and 8 healthy controls were recruited in this study. HD patients were followed up from September 2014 to May 2017. The primary end point was CVD event, the secondary outcome was death from any cause. Patients were divided into two groups with low and high M-CSF levels based on the optimal cut-off value determined by the ROC curve. Cox regression analyses were used to assess the predictive value of plasma M-CSF for CVD events and all-cause mortality in HD patients. We tested the levels of plasma M-CSF and other inflammatory cytokines in surviving HD patients using ELISA or CBA kit. RESULTS: The average plasma level of M-CSF in 52 patients was approximately twice that of healthy controls (992.4 vs. 427.2 pg/mL; p <  0.05). During 32 months of follow-up, 26 patients (50.0%) had at least one CVD event and 8 patients (15.4%) died. The mean plasma M-CSF concentration increased in survivors after follow-up compared to that detected at baseline (1277.8 ± 693.3 vs. 997.2 ± 417.4 pg/mL; p <  0.05). Multivariate Cox regression analysis showed that plasma M-CSF is an independent risk factor for CVD events in HD patients (p <  0.05). In the Cox regression model after adjusting for gender and age, high M-CSF levels were related to an increased risk of all-cause death (p <  0.05). We also found that M-CSF levels were positively correlated with IL-6 and IL-18 levels (both p < 0.05), which are the major pathogentic cytokines that contribute to HD-related CVD events. CONCLUSION: M-CSF is a prognostic factor for CVD events and all-cause mortality in HD patients.

Plasma levels of M-CSF and VEGF in laboratory diagnostics and differentiation of selected histological types of cervical cancers.

BACKGROUND: The search of useful serum biomarkers for the early detection of cervical cancers has been of a high priority. The activation of Macrophage-Colony Stimulating Factor (M-CSF) and Vascular Endothelial Growth Factor (VEGF) is likely involved in the pathogenesis and spread of cancer. We compared the plasma levels of M-CSF and VEGF to the ones of commonly accepted tumor markers CA 125and SCC-Ag in three groups of patients: 1. the cervical cancer group (patients with either squamous cell carcinoma or adenocarcinoma); 2. the cervical dysplasia group; 3. the control group. METHODS: This cohort study included 100 patients with cervical cancer and 55 patients with cervical dysplasia. The control group consisted of 50 healthy volunteers. The plasma levels of VEGF and M-CSF were determined using ELISA, while CA 125 and SCC-Ag concentrations were obtained by the chemiluminescent microparticle immunoassay (CMIA). RESULTS: The median levels of M-CSF and VEGF as well as CA 125 and SCC-Ag in the entire group of cervical cancer patients, were significantly different compared to the healthy women group. In case of both the squamous cell carcinoma and the adenocarcinoma groups, plasma levels of M-CSF and VEGF were higher compared to the control group. No significant differences in the studied parameters between the squamous cell carcinoma and the adenocarcinoma group were observed. The highest sensitivity and specificity were obtained for VEGF (81.18 and 76.00%, respectively) and SCC-Ag (81.18%; 74.00%) in the squamous cell carcinoma group and for VEGF (86.67%; 76.00%) in the adenocarcinoma group. The area under the ROC curve for VEGF was the largest in the adenocarcinoma group followed by the squamous cell carcinoma group (0.9082 and 0.8566 respectively). CONCLUSIONS: Obtained results indicate a possible clinical applicability and a high diagnostic power for the combination of MSC-F, VEGF, CA 125 and SCC-Ag in the diagnosis of both studied types of cervical cancer.

Phase I study of emactuzumab single agent or in combination with paclitaxel in patients with advanced/metastatic solid tumors reveals depletion of immunosuppressive M2-like macrophages.

BACKGROUND: Emactuzumab is a monoclonal antibody against the colony-stimulating factor-1 receptor and targets tumor-associated macrophages (TAMs). This study assessed the safety, clinical activity, pharmacokinetics (PK) and pharmacodynamics (PD) of emactuzumab, as monotherapy and in combination with paclitaxel, in patients with advanced solid tumors. PATIENTS AND METHODS: This open-label, phase Ia/b study comprised two parts (dose escalation and dose expansion), each containing two arms (emactuzumab, every 2 or 3 weeks, as monotherapy or in combination with paclitaxel 80 mg/m2 weekly). The dose-escalation part explored the maximum tolerated dose and optimal biological dose (OBD). The dose-expansion part extended the safety assessment and investigated the objective response rate. A PK/PD analysis of serial blood, skin and tumor biopsies was used to explore proof of mechanism and confirm the OBD. RESULTS: No maximum tolerated dose was reached in either study arm, and the safety profile of emactuzumab alone and in combination does not appear to preclude its use. No patients receiving emactuzumab monotherapy showed an objective response; the objective response rate for emactuzumab in combination with paclitaxel was 7% across all doses. Skin macrophages rather than peripheral blood monocytes or circulating colony-stimulating factor-1 were identified as an optimal surrogate PD marker to select the OBD. Emactuzumab treatment alone and in combination with paclitaxel resulted in a plateau of immunosuppressive TAM reduction at the OBD of 1000 mg administered every 2 weeks. CONCLUSIONS: Emactuzumab showed specific reduction of immunosuppressive TAMs at the OBD in both treatment arms but did not result in clinically relevant antitumor activity alone or in combination with paclitaxel. (ClinicalTrials.gov Identifier: NCT01494688).

Identification of Novel Natural Substrates of Fibroblast Activation Protein-alpha by Differential Degradomics and Proteomics.

Fibroblast activation protein-alpha (FAP) is a cell-surface transmembrane-anchored dimeric protease. This unique, constitutively active serine protease has both dipeptidyl aminopeptidase and endopeptidase activities and can hydrolyze the post-proline bond. FAP expression is very low in adult organs but is upregulated by activated fibroblasts in sites of tissue remodeling, including fibrosis, atherosclerosis, arthritis and tumors. To identify the endogenous substrates of FAP, we immortalized primary mouse embryonic fibroblasts (MEFs) from FAP gene knockout embryos and then stably transduced them to express either enzymatically active or inactive FAP. The MEF secretomes were then analyzed using degradomic and proteomic techniques. Terminal amine isotopic labeling of substrates (TAILS)-based degradomics identified cleavage sites in collagens, many other extracellular matrix (ECM) and associated proteins, and lysyl oxidase-like-1, CXCL-5, CSF-1, and C1qT6, that were confirmed in vitro In addition, differential metabolic labeling coupled with quantitative proteomic analysis also implicated FAP in ECM-cell interactions, as well as with coagulation, metabolism and wound healing associated proteins. Plasma from FAP-deficient mice exhibited slower than wild-type clotting times. This study provides a significant expansion of the substrate repertoire of FAP and provides insight into the physiological and potential pathological roles of this enigmatic protease.

Sera of patients with axial spondyloarthritis (axSpA) enhance osteoclastogenic potential of monocytes isolated from healthy individuals.

BACKGROUND: Axial spondyloarthritis (axSpA) is characterized by significant bone loss caused by dysregulation of physiological bone turnover, possibly resulting from intensified differentiation of osteoclasts. The aim of this study was to reevaluate the levels of osteoclastogenesis-mediating factors: soluble RANKL, M-CSF, OPG and other cytokines in sera of untreated, with sDMARDs and/or bDMARDs, axSpA patients and to test whether these sera influence differentiation of healthy monocytes towards osteoclast lineage. METHODS: Bone remodeling molecules (RANKL, M-CSF, OPG, IL-6, OSM, IL-17A, TGFß, and TNFα) were evaluated in 27 patients with axSpA and 23 age and sex-matched controls. Disease activity (BASDAI, ASDAS) and inflammatory markers (ESR, CRP) were assessed. Monocytes obtained from healthy individuals were cultured in vitro in presence of sera from 11 randomly chosen axSpA patients and 10 controls, with addition of exogenous M-CSF and/or RANKL or without. Osteoclastic differentiation was assessed analyzing osteoclast markers (cathepsin K and RANK at mRNA level) and with osteoclast-specific staining. RESULTS: axSpA patients' sera levels of soluble RANKL were significantly lower and M-CSF, IL-6, OSM, IL-17A and TNFα significantly higher in comparison to controls, whereas of OPG and TGFß were comparable in both groups. Numbers of generated in vitro osteoclasts and cathepsin K mRNA levels did not differ between cultures supplemented with sera of healthy and axSpA patients, both in the absence and presence of M-CSF. Instead, addition of exogenous RANKL boosted osteoclastogenesis, which was significantly higher in cultures with axSpA sera. Furthermore, sera from axSpA patients induced substantially higher levels of RANK mRNA, independently of M-CSF and RANKL stimulation. CONCLUSION: We show that, paradoxically, serum levels of soluble RANKL observed in axSpA are in fact significantly lower in comparison to healthy blood donors. Our results indicate that sera of axSpA patients - in contrary to healthy subjects - contain circulating, soluble factors (presumably IL-6, OSM, IL-17A, TNFα and others) able to stimulate healthy monocytes responsiveness to even relative low RANKL serum levels, by inducing high RANK mRNA expression and - as a net effect - boosting their osteoclastogenic potential. We suggest also that locally produced RANKL in axSpA may induce overactive osteoclasts from their precursors.

Plasma levels and diagnostic utility of macrophage-colony stimulating factor, matrix metalloproteinase-9 and tissue inhibitor of metalloproteinase-1 as tumor markers in cervical cancer patients.

Macrophage-colony stimulating factor, matrix metalloproteinase-9, and tissue inhibitor of metalloproteinase-1 may play an important role in malignant processes. The aim of this study was to investigate the diagnostic power of those parameters (serological biomarkers) in comparison to cancer antigen 125 and squamous cell carcinoma antigen in cervical cancer patients and in relation to the control groups. The study included 100 cervical cancer patients, 50 patients with cervical ectropion and 50 healthy women. Plasma levels of tested parameters were determined by enzyme-linked immunosorbent assay, cancer antigen 125, and squamous cell carcinoma antigen by chemiluminescent microparticle immunoassay. Plasma levels of all parameters in the total cancer group showed statistical significance (in all cases p < 0.05). In stage I of cancer only medial supraclavicular fossa and tissue inhibitor of metalloproteinase-1, in stage II all the tested parameters and cancer antigen 125, and in stage III + IV macrophage-colony stimulating factor, matrix metalloproteinase-9, and cancer antigen 125 showed statistical significance when compared to the healthy volunteers group. Macrophage-colony stimulating factor showed the highest value of sensitivity from all tested parameters (I: 56.25%, II: 72.73%, III + IV: 77.14% and 69% in total cervical cancer group). The highest specificity was obtained by matrix metalloproteinase-9 (94%). Positive predictive values were highest also for matrix metalloproteinase-9 (I: 82.35%, II: 84.21%, III + IV: 88% and 94.55% in total cervical cancer group), negative predictive values for macrophage-colony stimulating factor (I: 75.44%, II: 82.69%, III + IV: 87.5% and 58.11% in total cervical cancer group) and tumor markers. In the total cervical cancer group, all tested parameters showed statistically significant areas under receiver operating characteristic curve, but maximum range was obtained for the combination macrophage-colony stimulating factor + squamous cell carcinoma antigen (0.8723). The combined analysis of tested parameters and tumor markers resulted in an increase in sensitivity and areas under receiver operating characteristic curve values, which provides hope for developing new panel of biomarkers that may be used in the diagnosis of cervical cancer in the future.

Blood CSF1 and CXCL12 as Causal Mediators of Coronary Artery Disease.

BACKGROUND: Identification of biomarkers that cause coronary artery disease (CAD) has led to important advances in prevention and treatment. Epidemiological analyses have identified many biomarker-CAD relationships; however, these associations may arise from reverse causation and/or confounding and therefore may not represent true causal associations. Mendelian randomization (MR) analyses overcome these limitations. OBJECTIVES: This study sought to identify causal mediators of CAD through a comprehensive screen of 237 biomarkers using MR. METHODS: MR was performed by identifying genetic determinants of 227 biomarkers in ORIGIN (Outcome Reduction With Initial Glargine Intervention) trial participants (N = 4,147) and combining these with genetic effects on CAD from the CARDIoGRAM consortium (60,801 cases and 123,504 controls). Blood concentrations of novel biomarkers identified by MR were then tested for association with incident major adverse cardiovascular events in ORIGIN. RESULTS: Six biomarkers were found to be causally linked to CAD after adjustment for multiple hypothesis testing. The causal role of 4 of these is well documented, whereas macrophage colony-stimulating factor 1 (CSF1) and stromal cell-derived factor 1 (CXCL12) have not previously been reported, to the best of our knowledge. MR analysis predicted an 18% higher risk of CAD per SD increase in CSF1 (odds ratio: 1.18; 95% confidence interval: 1.08 to 1.30; p = 2.1 × 10-4) and epidemiological analysis identified a 16% higher risk of major adverse cardiovascular events per SD (hazard ratio: 1.16; 95% confidence interval: 1.09 to 1.23; p < 0.001). Elevated CXCL12 levels were also identified as a causal risk factor for CAD with consistent epidemiological results. Furthermore, genetically predicted CSF1 and CXCL12 levels were associated with CAD in the UK Biobank (n = 343,735). CONCLUSIONS: The study identified CSF1 and CXCL12 as causal mediators of CAD in humans. Understanding the mechanism by which these markers mediate CAD will provide novel insights into CAD and could lead to new approaches to prevention. These results support targeting inflammatory processes and macrophages, in particular, to prevent CAD, consistent with the recent CANTOS (Canakinumab Antiinflammatory Thrombosis Outcome Study). (Outcome Reduction With Initial Glargine Intervention [ORIGIN]; NCT00069784).

T cell-induced CSF1 promotes melanoma resistance to PD1 blockade.

Colony-stimulating factor 1 (CSF1) is a key regulator of monocyte/macrophage differentiation that sustains the protumorigenic functions of tumor-associated macrophages (TAMs). We show that CSF1 is expressed in human melanoma, and patients with metastatic melanoma have increased CSF1 in blood compared to healthy subjects. In tumors, CSF1 expression correlated with the abundance of CD8+ T cells and CD163+ TAMs. Human melanoma cell lines consistently produced CSF1 after exposure to melanoma-specific CD8+ T cells or T cell-derived cytokines in vitro, reflecting a broadly conserved mechanism of CSF1 induction by activated CD8+ T cells. Mining of publicly available transcriptomic data sets suggested co-enrichment of CD8+ T cells with CSF1 or various TAM-specific markers in human melanoma, which was associated with nonresponsiveness to programmed cell death protein 1 (PD1) checkpoint blockade in a smaller patient cohort. Combination of anti-PD1 and anti-CSF1 receptor (CSF1R) antibodies induced the regression of BRAFV600E -driven, transplant mouse melanomas, a result that was dependent on the effective elimination of TAMs. Collectively, these data implicate CSF1 induction as a CD8+ T cell-dependent adaptive resistance mechanism and show that simultaneous CSF1R targeting may be beneficial in melanomas refractory to immune checkpoint blockade and, possibly, other T cell-based therapies.