Combining Magnetic Resonance Imaging with Systemic Monocyte Evaluation for the Implementation of GBM Management.

Magnetic resonance imaging (MRI) is the gold standard for glioblastoma (GBM) patient evaluation. Additional non-invasive diagnostic modalities are needed. GBM is heavily infiltrated with tumor-associated macrophages (TAMs) that can be found in peripheral blood. FKBP51s supports alternative-macrophage polarization. Herein, we assessed FKBP51s expression in circulating monocytes from 14 GBM patients. The M2 monocyte phenotype was investigated by qPCR and flow cytometry using antibodies against PD-L1, CD163, FKBP51s, and CD14. MRI assessed morphologic features of the tumors that were aligned to flow cytometry data. PD-L1 expression on circulating monocytes correlated with MRI tumor necrosis score. A wider expansion in circulating CD163/monocytes was measured. These monocytes resulted in a dramatic decrease in patients with an MRI diagnosis of complete but not partial surgical removal of the tumor. Importantly, in patients with residual tumor, most of the peripheral monocytes that in the preoperative stage were CD163/FKBP51s- had turned into CD163/FKBP51s+. After Stupp therapy, CD163/FKBP51s+ monocytes were almost absent in a case of pseudoprogression, while two patients with stable or true disease progression showed sustained levels in such circulating monocytes. Our work provides preliminary but meaningful and novel results that deserve to be confirmed in a larger patient cohort, in support of potential usefulness in GBM monitoring of CD163/FKBP51s/CD14 immunophenotype in adjunct to MRI.

Role of FK506 Binding Protein on Tacrolimus Distribution in Red Blood Cells.

PURPOSE: Tacrolimus is distributed mainly in red blood cells (RBCs) after transfer into blood. This study aimed to evaluate the effect of FK506-binding proteins (FKBPs) on RBC distribution of tacrolimus in a physiological environment. METHODS: Human RBCs were isolated from fresh blood samples from healthy volunteers. The effect of FKBPs on each process of the RBC distribution of tacrolimus was evaluated in vitro. Effect of intracellular FKBPs was assessed by inhibition experiment with rapamycin, which competitively inhibits the binding of tacrolimus to FKBPs. Effect of extracellular FKBPs was examined by pre-exposure of RBCs to FKBP and preincubation of tacrolimus with FKBP. RESULTS: Pretreatment with rapamycin significantly reduced the rate of tacrolimus distribution in RBCs in a concentration-dependent manner. Pre-exposure of RBCs to FKBP12 followed by exposure to tacrolimus significantly decreased tacrolimus distribution in RBCs in a concentration-dependent manner. In addition, preincubation of tacrolimus with FKBP12 significantly reduced the rate of tacrolimus distribution in RBCs. CONCLUSIONS: FKBP played an important role in the distribution of tacrolimus in RBCs. The effect of intracellular and extracellular FKBPs on RBC distribution of tacrolimus in circulating blood was substantial. FKBP was shown as a potential biomarker for predicting the pharmacokinetics and pharmacodynamics of tacrolimus.

Proteomic analysis of infected primary human leucocytes revealed PSTK as potential treatment-monitoring marker for active and latent tuberculosis.

Markers for monitoring clearance of Mycobacterium tuberculosis (Mtb) infection during anti-TB drug treatment could facilitate management of tuberculosis (TB) treatment, but are lacking. We aimed to screen for Mtb clearance markers from in-vitro-infected leucocytes and to evaluate these markers in followed-up active TB (ATB) patients and latent TB (LTBI) cases after anti-TB drug treatment. Extracellular proteins from primary leucocytes infected with each of the Mtb lineages (East-Asian, Indo-Oceanic, Euro-American and the laboratory strain H37Rv) were screened as possible clearance markers. Leucocytes infected with Staphylococcus aureus acted as controls. The proteomic analysis was performed using GeLC-MS/MS. Several quantitative and qualitative candidate clearance markers were found. These proteins were suppressed during the infection stage of all Mtb lineages and re-expressed after bacillary clearance. PSTK, FKBP8 and MGMT were common clearance markers among the four Mtb lineages in our model. Only PSTK was a potential clearance marker based on western blot validation analysis from culture supernatants. The PSTK marker was further validated with western blot analysis using serum samples (n = 6) from ATB patients and LTBI cases during anti-TB drug treatment, and from healthy controls (n = 3). Time-dependent increase of PSTK was found both in ATB and LTBI patients during the course of anti-TB drug treatment, but not in healthy controls. We have demonstrated that PSTK is a potential treatment-monitoring marker for active and latent TB.

Towards precision medicine for stress disorders: diagnostic biomarkers and targeted drugs.

The biological fingerprint of environmental adversity may be key to understanding health and disease, as it encompasses the damage induced as well as the compensatory reactions of the organism. Metabolic and hormonal changes may be an informative but incomplete window into the underlying biology. We endeavored to identify objective blood gene expression biomarkers for psychological stress, a subjective sensation with biological roots. To quantify the stress perception at a particular moment in time, we used a simple visual analog scale for life stress in psychiatric patients, a high-risk group. Then, using a stepwise discovery, prioritization, validation, and testing in independent cohort design, we were successful in identifying gene expression biomarkers that were predictive of high-stress states and of future psychiatric hospitalizations related to stress, more so when personalized by gender and diagnosis. One of the top biomarkers that survived discovery, prioritization, validation, and testing was FKBP5, a well-known gene involved in stress response, which serves as a de facto reassuring positive control. We also compared our biomarker findings with telomere length (TL), another well-established biological marker of psychological stress and show that newly identified predictive biomarkers such as NUB1, APOL3, MAD1L1, or NKTR are comparable or better state or trait predictors of stress than TL or FKBP5. Over half of the top predictive biomarkers for stress also had prior evidence of involvement in suicide, and the majority of them had evidence in other psychiatric disorders, providing a molecular underpinning for the effects of stress in those disorders. Some of the biomarkers are targets of existing drugs, of potential utility in patient stratification, and pharmacogenomics approaches. Based on our studies and analyses, the biomarkers with the best overall convergent functional evidence (CFE) for involvement in stress were FKBP5, DDX6, B2M, LAIR1, RTN4, and NUB1. Moreover, the biomarker gene expression signatures yielded leads for possible new drug candidates and natural compounds upon bioinformatics drug repurposing analyses, such as calcium folinate and betulin. Our work may lead to improved diagnosis and treatment for stress disorders such as PTSD, that result in decreased quality of life and adverse outcomes, including addictions, violence, and suicide.

Monitoring protein expression by proteomics: human plasma exposed to benzene.

Low levels and long term exposure to benzene is associated with hematotoxicity including aplastic anemia, acute myelogenous leukemia, and lymphoma. Current biomonitoring methods such as urinary phenol, S-phenylmercapturic acid, and trans-trans muconic acid were found to be unreliable as analytical methods to detect benzene exposure. Therefore, to search for a specific protein for biomonitoring benzene exposure, we investigated plasma proteins from workers (n = 50) at a printing company who were exposed to benzene, by two-dimensional gel electrophoresis. The protein profiles are significantly different (p < 0.05) between benzene exposed and unexposed groups, as identified by matrix-assisted laser desorption ionization/time of flight mass spectrometry and confirmed by Western blot analyses. T cell receptor beta chain (TCR beta), FK506-binding protein, and matrix metalloproteinase-13 were expressed only in benzene exposed workers. In addition, interleukin-4 receptor alpha chain and T cell surface glycoprotein CD1b precursor were found to be up-regulated in the plasma of benzene exposed workers. When we treated Jurkat cells with benzene (10 microM-10 mM), TCR beta expression was increased in the membrane more than 6-9-fold compared to untreated cells. In addition, the amount of TCR beta released into the culture media, at benzene concentrations greater than 50 microM, increased up to 10 mM. Therefore, TCR beta levels in plasma could be used as a biomarker and a possible therapeutic target for benzene exposure.