WO2008107699A1

WO2008107699A1 - Diagnosing psychotic disorders

Info

Publication number: WO2008107699A1
Application number: PCT/GB2008/000826
Authority: WO
Inventors: Sabine Bahn; Rachel Marie Craddock
Original assignee: Cambridge Enterprise Ltd
Current assignee: Cambridge Enterprise Ltd
Priority date: 2007-03-08
Filing date: 2008-03-10
Publication date: 2008-09-12
Anticipated expiration: 2009-09-08

Abstract

A method of diagnosing or monitoring schizophrenia or a major depressive disorder, or predisposition thereto, comprises detecting and/or quantifying, in a sample from a test subject, expression of an isoform of CD45 as a biomarker.

Description

DIAGNOSING PSYCHOTIC DISORDERS Field of the Invention

The invention relates to methods for diagnosing or monitoring psychotic disorders, in particular schizophrenic disorders, using biomarkers. Background of the Invention

Psychosis is a symptom of severe mental illness. Although it is not exclusively linked to any particular psychological or physical state, it is particularly associated with schizophrenia, bipolar disorder (manic depression) and severe clinical depression. These conditions, their characterisation and categorisation, including DSM IV diagnosis criteria, are described in WO2007/045865, the content of which is incorporated herein by reference.

WO01 /63295 describes methods and compositions for screening, diagnosis and determining prognosis of neuropsychiatric or neurological conditions (including bipolar affective disorder, schizophrenia and vascular dementia), for monitoring the effectiveness of treatment in these conditions and for use in drug development.

Other techniques such as magnetic resonance imaging or positron emission tomography based on subtle changes of the frontal and temporal lobes and the basal ganglia are of little value for the diagnosis, treatment, or prognosis of schizophrenic disorders in individual patients, since the absolute size of these reported differences between individuals with schizophrenia and normal comparison subjects has been generally small, with notable overlap between the two groups. The role of these neuroimaging techniques is restricted largely to the exclusion of other conditions which may be accompanied by schizophrenic symptoms, such as brain tumours or haemorrhages. The validation of biomarkers that can detect early changes specifically correlated to reversal or progression of mental disorders is essential for monitoring and optimising interventions. Used as predictors, these biomarkers can help to identify high-risk individuals and disease sub-groups that may serve as target populations for chemo-intervention trials; as surrogate endpoints, biomarkers have the potential for assessing the efficacy and cost effectiveness of preventative interventions at a speed which is not possible at present when the incidence of manifest mental disorder is used as the endpoint.

WO2005/020784 discloses surrogate cell gene expression signatures, by a minimally invasive technique for determining the prognosis of a subject or the subjects susceptibility to a disease, disorder or physical state. It is reported (in Example 2) that various genes are modulated in, inter alia, psychiatric illness. A need exists to identify sensitive and specific methods and biomarkers for diagnosing and monitoring psychotic disorders, such as schizophrenic or bipolar disorders. Additionally, there is a need for methods, models, tests and tools for identification and assessment of existing and new therapeutic agents for the treatment of these disorders and methods for diagnosing psychotic disease.

T-cells are lymphocytes which develop in the thymus and play an important role in the immune system. There are two sub-populations of T-cells: cells with a CD4 marker are called helper T-cells whilst CD8+ cells are cytotoxic T-cells. Both T- cell types have a T-cell receptor (TCR) for antigen recognition. Stimulation or activation of a resting T-cell is initiated by the interaction of the TCR-CD3 complex with antigen-MHC class Il molecules on the surface of an antigen-presenting cell. This interaction initiates a cascade of biochemical events in the T-cell, including activation of gene transcription that eventually results in growth, proliferation and differentiation of the T-cell. WO2007/063333 (unpublished at the first filing date of the present invention, the content of which is incorporated herein by reference) discloses that assays, conducted on stimulated or unstimulated T-cells, can provide valuable information on the condition of a subject. T-cells provide a good model in which to investigate cellular function, as they are relatively easy to isolate with high purity, e.g. from peripheral blood, and can be obtained in a minimally invasive fashion. Summary of the Invention

The present invention is based on the Study, below. This shows that schizophrenia patients have a higher number of naϊve T-cells, by expression of CD45RA and CD45RB at least. An aspect of this invention is a method of diagnosing, predicting or monitoring a psychotic disorder in a subject by assessing the level of circulating naive T-cells and/or their expression of CD45, e.g. one or more of CD45RA, CD45RB and CD45RO. The T-cells are preferably stimulated. Description of Preferred Embodiments

For the avoidance of doubt, terms such as "response", "control" and "sample" as used herein include the possibility of there being more than one such response, control or sample, respectively.

The term "diagnosis" as used herein encompasses identification, confirmation, and/or characterisation of a psychotic disorder, in particular a schizophrenic disorder, bipolar disorder, related psychotic disorder, or predisposition thereto. By predisposition it is meant that a subject does not currently present with the disorder, but is liable to be affected by the disorder in time. Monitoring methods of the invention can be used to monitor onset, progression, stabilisation, amelioration and/or remission of a psychotic disorder.

The term "psychotic disorder" as used herein refers to a disorder in which psychosis is a recognised symptom, this includes neuropsychiatric (psychotic depression and other psychotic episodes) and neurodevelopmental disorders (especially autistic spectrum disorders), neurodegenerative disorders, depression, mania, and in particular, schizophrenic disorders (paranoid, catatonic, disorganized, undifferentiated and residual schizophrenia) and bipolar disorders. Preferably, the invention relates to schizophrenic disorders. T-cell samples are preferably obtained from peripheral blood taken from a subject. Preferably, T-cell samples are freshly isolated, that is they are used immediately following sample collection.

An example of a method for T-cell isolation is described in Example 1 of WO2007/063333. However, the skilled person will appreciate that other methods known in the art for obtaining or isolating T-cells from a biological sample, such as peripheral blood, may also be employed.

The term "stimulus" as used herein refers to a stimulus capable of inducing a response, preferably T-cell proliferation and responses associated with T-cell receptor-triggering. In vitro T-cell stimulation may be used as a method of comparing the functional responses of patient and control cells, firstly in order to investigate peripheral evidence of disease processes in schizophrenia and also to investigate whether global abnormalities or deficits in cell processes, such as cell signalling, gene transcription, protein synthesis and trafficking underlie the pathophysiology of this disorder. In vivo T-cell activation involves ligation of the T-cell receptor (TCR) through interaction with specific antigen presented in association with MHC. The TCR signalling complex is composed of a number of molecules including CD3, which provides the cytoplasmic signalling function of the complex, CD45, involved in de- phosphorylation of inhibitory phosphorylated tyrosine motifs and either CD4 or CD8, which are believed to stabilise the signalling complex. For optimal T-cell responses, co-stimulation is preferred for amplification and regulation of the initial signal. This is provided by molecules such as CD28, CD40, CD80/CD86 and OX40L.

Preferably, stimulation of T-cells is carried out in vitro by mimicking a TCR signal via cross-linking of cell surface CD3, using a monoclonal antibody (anti-CD3). This ultimately results in cell cycle entry and, as T-cell stimulation induces transcription factor activation, gene transcription, protein synthesis and protein trafficking, methods of the invention aim to identify and trace any abnormalities in these physiological processes and any consequences (e.g. differences in response to stimulus which may manifest in differences in mRNA, protein, lipid or other metabolite levels or ratios associated with such abnormalities).

Preferably, the stimulus is anti-CD3 antibody. Stimulation of T-cells may also be carried out using other agents, for example ionomycin and PMA, alone or in combination with CD3.

The term "response" as used herein may thus refer to a response elicited in response to the stimulation/activation of a resting T-cell. Such responses include proliferation, transcription factor activation or deactivation and modulation of one or more of the following: gene expression, protein synthesis, signal transduction, cytokine synthesis, protein trafficking and protein turnover, metabolite or lipid profile. Preferably, the response comprises proliferation, modulation of gene expression, protein synthesis and/or protein turnover.

Identification of differences between responses in T-cell samples from a subject having or being predisposed to a psychotic disorder, and stimulatory response in a normal subject, not affected by or predisposed to a psychotic disorder, can therefore be used to diagnose or monitor psychotic disease. Methods of the invention may comprise comparing a response in a test T-cell sample from a subject with a response to stimulation in a control. Suitable controls include normal controls derived from individuals not unaffected by or predisposed to psychotic disorder and disorder controls derived from individuals with a psychotic disorder preferably a schizophrenic disorder.

Methods of the invention may comprise detecting a difference in a response between the test sample and a control sample. Thus, methods of the invention may involve comparing a response in a test T- cell sample with a response in a normal control T-cell sample, wherein a difference in response is indicative of the presence of or predisposition to a psychotic disorder such as a schizophrenic disorder. Differences in response may be detected as a presence, absence, increase or decrease in a particular response to stimulus. Alternatively or additionally, methods of the invention may comprise a response in a test T-cell sample with a response in a psychotic disorder control T-cell sample, to enable the test T-cell response to be matched to the response characteristic of a particular psychotic disorder; such comparisons are useful for differential diagnosis of psychotic disorders that present with similar or overlapping clinical symptoms.

Following stimulation, T-cells from schizophrenia patients have been found to have significantly lower proliferation compared to healthy controls, as illustrated in Example 2 of WO2007/063333. Thus, in those embodiments where the response is proliferation, a lower proliferation in a T-cell sample from a subject compared to proliferation in a normal control T-cell sample is indicative of a psychotic disorder, in particular schizophrenia, being present. Proliferation may be assessed by ³[H]- thymidine incorporation into progeny cell DNA, as illustrated in Example 1 of WO2007/063333.

Differences in responses of T-cells from individuals having or predisposed to psychotic disorders and those from normal individuals may also be detected by assessing modulation in gene expression in response to exposure to stimulus, preferably in response to exposure to a stimulus for T-cell proliferation. Differences in responses may also be assessed by considering the downstream effects of differential gene expression in subjects having or being predisposed to a psychotic disorder, e.g. differences in metabolic profile, lipid profile, or differences in levels or ratio of biomarkers, compared to those in normal individuals not suffering from or predisposed to a psychotic disorder.

The terms "modulated" and "modulation" are used herein to mean an upregulation or downregulation of expression of a gene or differences in the proteome, for example, an increase or decrease in protein level. Modulation of gene expression can be measured by detecting a variation in mRNA or protein levels. The increase or decrease in protein level may be assessed by simply determining the presence or absence of a protein or by using a quantitative method.

Methods of determining the expression level of a gene are well known in the art. According to the methods of the invention, modulation of expression can be identified by assessing the amount or concentration of mRNA, a nucleic acid derived from mRNA or a protein translated from the mRNA. Gene expression may be measured by assessing mRNA levels using a method including reverse transcription and polymerase chain reactions ("RT-PCR"), such as quantitative PCR (in particular, real-time quantitative PCR), and Northern blotting. In one suitable method for determining the level of mRNA expressed, a total RNA sample is obtained from the cell, cDNA is synthesized from mRNA of the gene or genes of interest, and the cDNA is used for real-time quantitative PCR analysis to determine the level of the mRNA of interest in the sample. Systems and kits for implementing such methods are commercially available.

Arrays may be used to assess expression of a plurality of genes or proteins, for example using weak cation exchange (CM10) chips for SELDI analysis of proteins, or Codelink Bioarrays for gene expression. An example of a method used to assess gene expression is shown in Example 3 of WO2007/063333. Examples of suitable methods for determining the level of protein expression or identifying protein biomarkers include immunological methods, involving an antibody, or an antibody fragment capable of specific binding to the protein of interest. Suitable immunological methods include sandwich immunoassays, such as sandwich ELISA in which detection of the peptide is performed using two antibodies which recognize different epitopes; radioimmunoassays (RIA), direct or competitive enzyme-linked immunosorbent assays (ELISA), enzyme-immuno assays (EIA), Western blotting, immunoprecipitation and any particle-based immunoassay (e.g. using gold, silver, latex or magnetic particles or Q-dots). Immunological methods may be performed, for example, in microtitre plate or strip format.

Other techniques that may be used in the methods of the invention, for example for the detection, identification and/or quantification of a biomarker, e.g. for quantifying the level of a nucleic acid, protein, lipid or metabolite present, include spectral analysis, such as NMR spectroscopy and high resolution NMR spectroscopy (¹H NMR), mass spectrometry, such as Surface Enhanced Laser/Desorption Ionization (SELDI) (-TOF) and/or MALDI (-TOF), 1-D gel-based analysis, 2-D gel- based analysis, LC-MS-based technique or iTRAQ™. An example used to analyse proteins is shown in Example 4 of WO2007/063333. iTRAQ™ technology involves the chemical tagging of N-terminus peptides resulting from protein digestion with trypsin. Up to four labelled samples are combined, fractionated by nano-LC and analysed by tandem mass spectrometry. Protein identification is then achieved by database searching of fragmentation data. Relative quantification of peptides is achieved by fragmentation of the chemical tag, which results in a low molecular weight reporter ion. As samples are labelled after tryptic digestion, analysis of high molecular weight proteins such as trans-membrane receptors is possible and quantification of fragmented tag provides greater confidence in protein identity and quantification.

According to the invention, a suitable cohort of patients and controls may be selected including first onset and/or minimally treated individuals and these will be compared with chronically ill patients having a more established clinical history. This allows comparison of both disease progression and the effects of drug treatment. Membrane-bound and soluble proteins may be prepared from stimulated T-cells. Thus, proteomic profiling of T-cells from psychosis patients and controls may be performed, providing information regarding differing expression of large and small molecular weight proteins, e.g. phosphoproteins, following stimulation.

Methods of the invention may comprise comparing samples by assessing variation in one or more biomarkers in response to stimulation of the sample. The term "biomarker" means a distinctive biological or biologically-derived indicator of a process, event, or condition. Biomarkers can be used in methods of diagnosis (e.g. clinical screening), prognosis assessment; in monitoring the results of therapy, identifying patients most likely to respond to a particular therapeutic treatment, drug screening and development. Preferably, the biomarker is a gene, mRNA, a protein or peptide, lipid, or metabolite. The terms protein and peptide are used interchangeably herein. The biomarker may be quantified. Biomarkers and uses thereof are valuable for identification of new drug treatments and for discovery of new targets for drug treatment. Quantifying the amount of the biomarker present in a sample may include determining the concentration of the peptide biomarker present in the sample. Detecting and/or quantifying may be performed directly on the sample, or indirectly on an extract therefrom, or on a dilution thereof. Detecting and/or quantifying can be performed by any method suitable to identify the presence and/or amount of a specific protein in a biological sample. In one embodiment, the control sample comprises a normal control sample.

In another embodiment, the control sample comprises a psychotic disorder control sample. In another embodiment, the method may also comprise classifying proliferative responses of a sample as having a normal profile, psychotic disorder profile, or psychotic disorder predisposition profile. In methods of the invention, in particular those for diagnosing and monitoring,

T-cell samples may be taken on two or more occasions from a test subject. Stimulatory responses from samples taken on two or more occasions from a test subject can be compared to identify differences between the stimulatory responses in samples taken on different occasions. Methods may include analysis of stimulatory responses from biological samples taken on two or more occasions from a test subject to quantify the level of one or more biomarkers present in the biological samples, and comparing the level of the one or more biomarkers present in samples taken on two or more occasions.

Diagnostic and monitoring methods of the invention are useful in methods of assessing prognosis of a psychotic disorder, in methods of monitoring efficacy of an administered therapeutic substance in a subject having, suspected of having, or of being predisposed to, a psychotic disorder and in methods of identifying an antipsychotic or pro-psychotic substance. Such methods may comprise comparing the level of the one or more biomarkers in a test biological sample taken from a test subject with the level present in one or more samples taken from the test subject prior to administration of the substance, and/or one or more samples taken from the test subject at an earlier stage during treatment with the substance. Additionally, these methods may comprise detecting a change in the level of the one or more biomarkers in biological samples taken from a test subject on two or more occasions.

A method of diagnosis of or monitoring according to the invention may comprise quantifying the one or more biomarkers in a test biological sample taken from a test subject and comparing the level of the one or more biomarkers present in said test sample with one or more controls. The control can be selected from a normal control and/or a psychotic disorder control. The control used in a method of the invention can be selected from: the level of biomarker found in a normal control sample from a normal subject, a normal biomarker level; a normal biomarker range, the level in a sample from a subject with a schizophrenic disorder, bipolar disorder, related psychotic disorder, or a diagnosed predisposition thereto; a schizophrenic disorder marker level, a bipolar disorder marker level, a related psychotic disorder marker level, a schizophrenic disorder marker range, a bipolar disorder marker range and a related psychotic disorder marker range. Detecting differences in responses enables identification of biomarkers for a psychotic disorder. The response may be assessed by any suitable method or combination of methods, for example by considering gene expression, at the mRNA and/or protein level, to detect differential gene expression between disorder and control samples, by considering protein levels (e.g. in cell lysate), lipid profile and/or metabolite profile. The differences may manifest as the presence or absence of a biomarker or a difference (increase or decrease) in level of a biomarker, or in ratios of a biomarker or biomarkers.

Differences in gene expression can be detected by a modulation in mRNA or protein levels. Where the biomarker is a gene, the expression of the gene present in the disorder sample may be modulated compared to the expression of the gene in the control sample, thus different levels of mRNA transcribed from the gene will be detected. For example, the expression may be increased or decreased, or different splice variants or ratios of splice variants of the mRNA may be detected. In another embodiment, the biomarker is a protein and the level of the protein present in the sample differs from the level of the protein present in the control sample. For example, the level may be modulated so that it is increased or decreased, or a difference in protein cleavage products may be found; this may be assessed by a quantitative method or determined by the presence or absence of the protein.

In one embodiment, the level or ratio of one or more biomarkers is detected. This may be carried out using a sensor, e.g. a biosensor comprising one or more enzymes, binding, receptor or transporter proteins, antibody, synthetic receptors or other selective binding molecules for direct or indirect detection of the biomarkers. For detection, the sensor may be coupled to an electrical, optical, acoustic, magnetic or thermal transducer.

The term "antibody" as used in this embodiment includes, but is not limited to, polyclonal, monoclonal, bispecific, humanised or chimeric antibodies, single chain antibodies, Fab fragments and F (ab')₂ fragments, fragments produced by a Fab expression library, anti-idiotypic (anti-Id) antibodies, and epitope-binding fragments of any of the above. The term "antibody" as used herein also refers to immunoglobulin molecules and immunologically-active portions of immunoglobulin molecules, i.e., molecules that contain an antigen binding site that specifically binds an antigen. The immunoglobulin molecules of the invention can be of any class (e. g., IgG, IgE, IgM, IgD and IgA) or subclass of immunoglobulin molecule.

Biomarkers identified using a method of the invention can be used as biomarkers for a psychotic disorder or predisposition thereto. They are thus useful in methods for monitoring or diagnosing psychotic disease. The present invention may be used identify a potential therapeutic agent for the prevention, treatment or amelioration of a psychotic disorder. In one embodiment, the invention comprises comparing a response to stimulation with a response in a control sample. In particular, responses in test and normal control T- cell samples exposed to a candidate therapeutic agent may be compared, identifying the candidate as a potential therapeutic agent if one or more responses in the test T- cell sample are modulated such that a normal response is restored.

According to this aspect, a candidate therapeutic agent is identified if the candidate therapeutic agent is capable of modulating a response in T-cells from a subject having a psychotic disorder, in particular such that one or more responses are restored to the response characteristic of T-cells from normal individuals. Preferably, the response is proliferation or modulation of gene expression, i.e. changes in mRNA or protein levels. The response can be assessed using the methods described herein, in particular by assessing biomarkers of response identified as described herein. Changes in proliferation can be assessed by comparing the proliferation of T-cells in the presence and absence of the candidate therapeutic agent. Modulation of expression of one or more genes can be assessed by comparing the expression level of the gene or genes (at the mRNA or protein level) in the presence and absence of the candidate therapeutic agent. Modulation of protein levels can be assessed by comparing the level of the protein or proteins in the presence and absence of the candidate therapeutic agent. Other suitable biomarkers of response include lipids and metabolites found at different levels in disorder and normal control samples. In another aspect, the invention relates to a diagnostic kit or monitoring kit suitable for performing a method described herein. Kits according to the invention may comprise one or more components selected from: instructions for use of the kit, one or more normal and/or psychotic disorder controls, a sensor or biosensor suitable and/or adapted for detecting a biomarker according to the invention and a ligand, e.g. nucleic acid, antibody, aptamer, or the like, capable of specifically binding a biomarker according to the invention or specifically binding a substance derived from the biomarker or from the action of the biomarker. The ligand may be provided immobilised on a solid support such as bead or surface, for example in the form of an array adapted for use in a method of the invention.

The following Study illustrates the invention. It should be read in conjunction with the drawings, in which each Figure is a graph of expression. Study

The study was conducted in order to investigate mechanisms underlying lower proliferative responses of schizophrenia patient T-cells to stimulation with anti- CD3, by looking at activation subtype of T-cells from schizophrenia patients and controls. The study was conducted as follows:

T-cells from schizophrenia patients and healthy controls (n = 12) were cultured in the presence or absence of anti-CD3 (clone OKT3) in RPMI-1640 medium supplemented with 1% GPS and 10% foetal bovine serum. CD45 expression was assessed by flow cytometry.

The experiments showed that CD45RA expression is significantly higher on CD4 and CD8 T-cells from schizophrenia patients before and after stimulation with OKT3 and is also present on a higher percentage of cells (Figure 1A). CD45RB expression is significantly higher on schizophrenia patient T-cells both before and after stimulation, and is significantly higher on CD8+ cells before stimulation only (Figure 1 B). There is also a trend towards lower expression of CD45RO on schizophrenia patient T-cells (Figure 1C).

CD45 isotypes are indicative of the activation status of T-cell; CD45RA typically determines antigen-naϊve T-cells. Expression of CD45RB increases on CD45RA+ naive cells as they repeatedly encounter antigen, while expression of CD45RA diminishes as CD45RO becomes expressed, to define a population of effecter or memory T-cells. The data suggest that schizophrenia patients have a higher proportion of circulating naϊve T-cells. During this work, it was also found that expression of CD3 and TCR (T-cell receptor) αβ chains was equivalent between patients and controls, ensuring equal stimulation with anti-CD3, and that there was no significant difference in the proportions of CD4+ and CD8+ T-cells between samples (n = 12). Lower T-cell proliferation in schizophrenia patients was not found to result from deficient early tyrosine phosphorylation signaling or lower IL-2 (interleukin-2) production, as these parameters were similar between patients and controls, as was the expression of CD25, the IL-2 receptor α chain. Analysis of CD45 isoforms, however, revealed that patients have a significantly greater percentage of CD8+ and CD4+ CD45RA+ cells before stimulation and significantly higher fluorescence intensity of CD45RA on CD4+ and CD8+ cells before and after stimulation. There was significantly higher expression of CD45 RN on both CD4+ and CD8+ unstimulated cells, with a trend towards lower numbers of CD45RO+ T-cells in patient blood.

Claims

1 A method of diagnosing or monitoring schizophrenia or a major depressive disorder, or predisposition thereto, comprising detecting and/or quantifying, in a sample from a test subject, expression of an isoform of CD45 as a biomarker.

2. A method of monitoring efficacy of a therapy in a subject having, suspected of having, or of being predisposed to schizophrenia or a major depressive disorder, comprising detecting and/or quantifying, in a sample from said subject, CD45 as a biomarker.

3. A method according to claim 2, comprising comparing the amount of the biomarker in said test sample with the amount present in one or more samples taken from said subject prior to commencement of therapy, and/or one or more samples taken from said subject at an earlier stage of therapy.

4. A method according to any preceding claim, which is conducted on samples taken on two or more occasions from a test subject.

5. A method according to any of claims 1 to 4, further comprising comparing the level of the biomarker present in samples taken on two or more occasions.

6. A method according to any of claims 1 to 5, further comprising detecting a change in the amount of the biomarker in samples taken on two or more occasions.

7. A method according to any of claims 1 to 6, comprising comparing the amount of the biomarker present in said test sample with one or more controls.

8. A method of according to claim 7, comprising comparing the amount of the biomarker in a test sample with the amount of the biomarker with the amount present in a sample from a normal subject.

9. A method according to any of claims 1 to 8, wherein samples are taken prior to and/or during and/or following an anti-psychotic therapy.

10. A method according to any of claims 1 to 9, wherein samples are taken at intervals over the remaining life, or a part thereof, of a subject.

11. A method according to any of claims 1 to 10, wherein quantifying is performed by measuring the concentration of the biomarker in the or each sample.

12. A method according to any of claims 1 to 11 , wherein detecting and/or quantifying is performed by one or more methods selected from SELDI (-TOF), MALDI (-TOF), a 1-D gel-based analysis, a 2-D gel-based analysis, Mass spec (MS), reverse phase (RP) LC, size permeation (gel filtration), ion exchange, affinity, HPLC, UPLC or other LC or LC-MS-based technique.

13. A method according to any of claims 1 to 12, wherein detecting and/or quantifying is performed using an immunological method.

14. A method according to any of claims 1 to 13, wherein the detecting and/or quantifying is performed using a biosensor or a microanalytical, microengineered, microseparation or immunochromatography system.

15. A method according to any of claims 1 to 14, wherein the sample is cerebrospinal fluid, whole blood, blood serum, plasma, urine, saliva, or other bodily fluid, or breath, condensed breath, or an extract or purification therefrom, or dilution thereof.

16. A method according to any of claims 1 to 15, wherein the disorder is a major depressive disorder.

17. A method according to any of claims 1 to 16, wherein the biomarker is CD45RA.

18. A method according to any of claims 1 to 16, wherein the biomarker is CD45RB.

19. A method according to any of claims 1 to 16, wherein the biomarker is CD45RO.

20. A biosensor comprising an antibody specific for an isoform of CD45 and means to convert detection of the presence of CD45 into a signal.

21. A biosensor according to claim 20, wherein the antibody is a monoclonal antibody.

22. A biosensor according to claim 20 or claim 21, wherein the antibody is labelled with a detectable marker.

23. A biosensor according to claim 22, wherein the detectable marker is a luminescent, fluorescent or radioactive marker, or an affinity tag.

24. Use of a biosensor according to any of claims 20 to 23, to detect and/or quantify a biomarker as defined in any of claims 1 and 17 to 19.

25. Use according to claim 24, wherein the detection and/or quantification is performed on a sample as defined in claim 15.