Identification of a molecular profile associated with immune status in first-onset schizophrenia patients.

OBJECTIVES: Alterations in immunological parameters have been reported for schizophrenia although little is known about the effects of inflammatory status on immune-related functional changes at disease onset. Here, we have investigated such T cell-dependent molecular changes in first-onset, antipsychotic-naive schizophrenia patients using a novel ex vivo blood culture system. METHODS: Blood samples from patients (n=17) and controls (n=17) were collected into stimulant-containing or null control TruCulture™ tubes, incubated 24 hours and the concentrations of 107 immune and metabolic molecules measured in the conditioned media using the HumanMAP™ immunoassay system. RESULTS: Nine molecules showed altered release from schizophrenia blood cells compared to those from controls and this was replicated in an independent cohort. In silico pathway analysis showed that these molecules had roles in endothelial cell function, inflammation, acute phase response and fibrinolysis pathways. Importantly, five of these molecules showed altered release only after stimulation. CONCLUSIONS: This study has identified a reproducible peripheral molecular signature associated with altered immune function in first-onset schizophrenia subjects. This suggests that immune status can affect the biomarker profile which could be important for personalized medicine strategies. Furthermore, whole blood culture analysis may be useful in the identification of diagnostic tools or novel treatment strategies due to ease-of-use and clinical accessibility.

Analysis of serum and plasma identifies differences in molecular coverage, measurement variability, and candidate biomarker selection.

PURPOSE: To compare the use of serum and plasma in multiplex immunoassay analyses of 190 proteins and small molecules, and associated molecular pathways. We also tested whether differences between these biofluids can influence the identification of potential biomarkers in a preliminary study comparing bipolar disorder patients with controls. EXPERIMENTAL DESIGN: Using multiplexed immunoassay analyses, we compared the measurement levels and interindividual variation of 190 proteins and small molecules between serum and plasma collected from 21 healthy individuals. We exemplify how this can impact on the outcome of biomarker discovery studies using a case study of 24 patients with bipolar disorder. RESULTS: Detection of analytes was similar for serum and plasma, although there were marked differences in measurement variability for 29 proteins and cortisol. When considering the disease cohort we identified six proteins that changed significantly in serum and ten in plasma with an overlap of two proteins. CONCLUSIONS AND CLINICAL RELEVANCE: In spite of the similarities of coverage on a multiplexed platform for serum and plasma, there were important differences in interindividual variability, which can have significant impact on identifications made in biomarker studies.

Identification of a blood-based biological signature in subjects with psychiatric disorders prior to clinical manifestation.

OBJECTIVES: To determine whether a molecular signature is present in blood of patients with psychiatric disorders before manifestation of symptoms. METHODS: Multiplex immunoassay analyses were carried out using serum obtained from two case-control studies of schizophrenia (n = 75) and bipolar disorder (n = 110) patients and their matched controls. The samples were drawn within 1 month before estimated onset of illness. RESULTS: This led to identification of 20 molecules which were altered in pre-schizophrenia and 14 molecules in pre-bipolar disorder subjects compared to controls. Only two of these molecular changes were identical in both data sets and predictive testing confirmed that the biomarker signatures for pre-schizophrenia and pre-bipolar disorder were dissimilar. CONCLUSION: The present results suggest that there are distinct serum alterations that occur before clinical manifestation of schizophrenia and bipolar disorder. These findings could lead to development of diagnostic tests to help clinical psychiatrists identify and classify vulnerable patients early in the disease process, allowing for earlier and more effective therapeutic intervention.

Altered levels of circulating insulin and other neuroendocrine hormones associated with the onset of schizophrenia.

Recently, we showed that the circulating levels of insulin-related peptides and the secretory granule protein chromogranin A were increased in small cohorts of first onset schizophrenia patients. Assuming that this effect was associated with impaired insulin signalling, we investigated the possibility that secretion of other hormones is also affected in schizophrenia. Multiplex immunoassay analysis of 21 hormones and hormone-related molecules was carried out using sera from 236 first and recent onset schizophrenia patients and 230 matched controls. Serum concentrations of insulin and chromogranin A were increased in schizophrenia subjects, consistent with our previous study. In addition, we found elevated concentrations of pancreatic polypeptide, prolactin, progesterone and cortisol, and decreased levels of growth hormone. We also found that growth hormone levels were decreased in post-mortem pituitaries obtained from chronic schizophrenia patients. It will be important to determine whether any of these molecules are involved in the pathosphysiology of schizophrenia or if they reflect the associated insulin resistance. We conclude that function of multiple components of the hypothalamic-pituitary-adrenal-gonadal axis may be affected in schizophrenia. This could have important implications for future biomarker discovery efforts and personalized medicine strategies based on patient stratification for the treatment of this debilitating disorder.

Validation of a blood-based laboratory test to aid in the confirmation of a diagnosis of schizophrenia.

We describe the validation of a serum-based test developed by Rules-Based Medicine which can be used to help confirm the diagnosis of schizophrenia. In preliminary studies using multiplex immunoassay profiling technology, we identified a disease signature comprised of 51 analytes which could distinguish schizophrenia (n = 250) from control (n = 230) subjects. In the next stage, these analytes were developed as a refined 51-plex immunoassay panel for validation using a large independent cohort of schizophrenia (n = 577) and control (n = 229) subjects. The resulting test yielded an overall sensitivity of 83% and specificity of 83% with a receiver operating characteristic area under the curve (ROC-AUC) of 89%. These 51 immunoassays and the associated decision rule delivered a sensitive and specific prediction for the presence of schizophrenia in patients compared to matched healthy controls.

Key factors in evaluating potential clinical biomarkers.

Clinical biomarkers have important differences from exploratory or experimental biomarkers. The introduction of biomarkers into a true clinical situation requires unique considerations of the potential utility of the markers. In contrast to experimental situations in which various parameters can be tightly controlled, the world of patient care can be characterized as the 'Wild West' - an unknown frontier in which every possible variable may be introduced at some stage. Because a clinical biomarker or pattern of biomarkers arises from a single individual without the benefit of a control group to detect inconsistent sample handling, the biomarker must be, above all else, robust. A biomarker for which even slightly different handling in the clinic results in highly different assay results will have minimal utility, and may even create a dangerous situation for patients. The samples and assays used, the clinical situation and the format of data output are also important considerations in the development of clinical biomarkers.