WO2024110683A1 - Anticorps antinucléaires anti-ssa en tant que biomarqueurs précoces de la lésion rénale causée par le syndrome de sjörgen - Google Patents

Anticorps antinucléaires anti-ssa en tant que biomarqueurs précoces de la lésion rénale causée par le syndrome de sjörgen Download PDF

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WO2024110683A1
WO2024110683A1 PCT/ES2023/070696 ES2023070696W WO2024110683A1 WO 2024110683 A1 WO2024110683 A1 WO 2024110683A1 ES 2023070696 W ES2023070696 W ES 2023070696W WO 2024110683 A1 WO2024110683 A1 WO 2024110683A1
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ssa
syndrome
kidney damage
urine
sjógren
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Concepción MARAÑÓN LIZANA
María PALOMEQUE SÁNCHEZ
Francisco PÉREZ COZAR
Nieves Yaquelin VARELA HERNÁNDEZ
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Fundación Pública Andaluza Progreso Y Salud
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    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
    • G01N33/564—Immunoassay; Biospecific binding assay; Materials therefor for pre-existing immune complex or autoimmune disease, i.e. systemic lupus erythematosus, rheumatoid arthritis, multiple sclerosis, rheumatoid factors or complement components C1-C9

Definitions

  • Antinuclear anti-SSA antibodies as early biomarkers of kidney damage caused by Sjogren's Syndrome
  • the present invention is within the field of medicine, and relates to biomarkers detectable in the urine of patients to early diagnose kidney damage caused by Sjógren's Syndrome (SjS).
  • SjS Sjógren's Syndrome
  • One of the main functions of the immune system is to distinguish what is its own from what is foreign, which allows maintaining a state of immune tolerance in which it does not react against its own healthy tissues, but does react against foreign antigens.
  • B lymphocytes and T lymphocytes immunoglobulins that recognize the antigen
  • the rearrangement of the genes that determine the variability of the areas that recognize the antigen takes place.
  • the high variability that this process generates gives rise to to combinations of immunoglobulins that recognize its own molecular structures, also known as autoantigens, creating self-reactive lymphocytes.
  • the SI's aversion to acting against its own organism is maintained thanks to central tolerance mechanisms, which take place in the thymus in the case of cells T and in the spleen in the case of B cells, and peripheral tolerance, which occurs in tissues and lymph nodes once the maturation of the lymphocytes has taken place.
  • central tolerance mechanisms which take place in the thymus in the case of cells T and in the spleen in the case of B cells
  • peripheral tolerance which occurs in tissues and lymph nodes once the maturation of the lymphocytes has taken place.
  • Negative selection is a central selection process through which those cells that recognize their own peptides are eliminated, that is, they are autoreactive, so that cases of autoimmunity are avoided.
  • T cells also undergo a second selection in the periphery, known as peripheral tolerance in which these cells can present anergy (cells that have an insufficient concentration of receptors to be functional), be eliminated by apoptosis or suffer suppression by cells. Regulatory tees.
  • AID Autoimmune diseases
  • the adaptive autoimmune response takes place, specific to one or multiple self-antigens.
  • Autoimmune responses are persistent because the antigen is never able to be eliminated, since which is its own (autoantigen), and therefore would entail a complete destruction of the target tissue.
  • the tissue damage resulting from the immune response leads to increased exposure of autoantigens, which causes an increase in the production of autoantibodies, exacerbating the autoimmune response and aggravating tissue damage, leading to a positive feedback loop.
  • the cause of AIS is the loss of tolerance, they share a set of characteristics resulting from the autoimmune response, such as their specificity, chronicity, progressivity, clinical heterogeneity, and greater predisposition in women.
  • SAD systemic autoimmune diseases
  • ANA antinuclear antibodies
  • SLE systemic lupus emthematosus
  • SjS Sjógren's syndrome
  • PAPs antiphospholipid syndrome
  • MCTD mixed connective tissue disease
  • UCTD undifferentiated connective tissue disease
  • IC depositions which cause cellular lesions, exposing DNA and RNA structures.
  • the production of autoantibodies and the tissue damage produced also induces a flow of neutrophils towards these places, where they carry out the process of NETosis (Neutrophil Extracellular Traps), a mechanism by which chromatin exit takes place, together with microbicidal enzymes, of neutrophils. In this way, a self-feeding cycle is generated that causes the damage to worsen over time.
  • NETosis Neurotrophil Extracellular Traps
  • nephritis has the worst prognosis. 30-60% of patients with SLE manifest lupus nephritis (LN).
  • Sjógren's Syndrome SjS is characterized by lymphocytic infiltrates in the exocrine glands, leading to their dysfunction.
  • renal manifestations are minor, being 5%, although in this case tubulointerstitial nephritis is the most common renal condition, and occurs in 75% of renal biopsies, compared to 25% of glomerulonephritis, which It occurs in more severe cases and is indicative of a worse prognosis.
  • tubulointerstitial nephritis cellular infiltrations of T lymphocytes, mostly CD4 + , stand out.
  • hypergammaglobulinemia is common, in which B cells are unable to carry out the isotype change and therefore IgM production is exacerbated, increasing the production of anti-SSA (SjS-related antigen A) autoantibodies. and anti-SSB (SjS-related B antigen).
  • SSA SjS-related antigen A
  • SB SjS-related B antigen
  • PAPs presents as an autoimmune thrombophilia.
  • the damage that occurs in the kidney (2.7-9%) is due to vasculitopathies in the blood vessels of the renal circulation. Manifestations vary from isolated conditions such as hematuria or proteinuria, to causing acute kidney damage or chronic kidney disease. Thrombosis also occurs that can block the blood supply to the kidney, sometimes causing the death of kidney tissue due to lack of oxygen supply.
  • MCTD is defined as a chronic disease that presents overlapping symptoms of SLE, scleroderma, and polymyositis. However, the symptoms it presents are less severe and the kidney involvement is not usually as severe as in SLE, so that the prognosis is, in most cases, more favorable.
  • the detection of anti-U1-RNP (anti-U1 nuclear ribonucleoprotein) antibodies is used along with characteristics of some of the AAI already mentioned above.
  • UCTD ulcerative colitis
  • Nephritis inflammation of the kidney is mostly caused by the accumulation of IC. Inflammation can be found both in the glomeruli (glomerulonef ⁇ tis) and in the tubules of the nephron and surrounding tissue (tubulointerstitial nephritis). Glomerular damage is mostly due to the accumulation of IC as a consequence of the systemic activity of the disease, while tubulointerstitial nephritis is caused by the local response caused in the kidney by immune cell infiltrates. T and B cells can form aggregates, and sometimes germinal centers, in the interstitial tissue of the kidney.
  • Biomarkers are factors that can be measured, whether expression of certain genes, levels of certain proteins or production of chemical substances characteristic of the disease. For a biomarker to be optimal, it must be specific to the disease, represent the activity of the kidney pathology as strictly as possible (and therefore be able to discriminate between activity and chronicity) and be able to predict outbreaks, so it is also important that it does not depend on of other factors such as age, sex or ancestry.
  • kidney biopsy In those cases in which kidney pathologies due to EAS occur, the only method to diagnose them safely is through kidney biopsy. It is a highly invasive technique that carries a series of risks and does not allow patient monitoring because it cannot be performed repeatedly. It is not usually performed until the symptoms are clear, and therefore the disease is advanced. Likewise, the little tissue taken in the sample is not representative of kidney lesions.
  • non-invasive markers that are also used are serum creatinine levels, proteinuria, and active sediment in the urine. These markers are indicative of kidney damage, but are nevertheless not specific enough. This is why the search for new biomarkers related to the pathophysiology of kidney damage is necessary.
  • serum autoantibodies are more stable and the sample is not exposed to the risk of bacterial contamination.
  • serum autoantibodies are representative of disease activity throughout the body and are therefore not specific for kidney damage.
  • blood has a high protein content, which makes analysis difficult.
  • the autoantibody profile in urine is representative of renal inflammation, unlike levels in serum, so that they constitute a viable biomarker to evaluate the deposition of immune complexes (IC) in the kidney, in addition to correlating with the severity of different parameters. kidneys.
  • IC immune complexes
  • the possibility of being able to monitor the patient and recognize early signs of the disease, and therefore be able to carry out an early diagnosis through analysis of the autoantibody profile in urine is a great advance, since it is not an invasive procedure. , so it can be done repeatedly, and it provides us with essential information about the patient's condition. Not only this, but it also allows the administration of the corresponding treatment in the early phases of the disease, avoiding the formation of irreversible damage, and therefore improving the patient's prognosis and reducing the morbidity and mortality effects of EAS.
  • a first aspect of the invention refers to the in vitro use of the level of anti-SSA antinuclear antibodies in urine as an early biomarker to diagnose kidney damage caused by Sjógren's Syndrome.
  • a second aspect of the invention refers to the method of obtaining data useful for the diagnosis of kidney damage caused by Sjógren's Syndrome, which comprises: a) Detecting and quantifying the antinuclear anti-SSA antibody in urine samples previously isolated from individuals, and b ) Compare the values obtained with those of a control group of patients diagnosed with Sjógren's Syndrome who do not present kidney damage.
  • a third aspect of the invention refers to the method for diagnosing kidney damage caused by Sjógren's Syndrome, which includes the method of obtaining useful data described above and which also includes: c) Assigning to the group of individuals who present values of antinuclear antibodies. -SSA higher than the control group, the group of individuals who present kidney damage caused by Sjógren's Syndrome.
  • the detection and quantification of antinuclear antibodies is carried out by multiplexed immunoassays and cytometry. flow.
  • a fourth aspect of the invention refers to the use of a kit or device for detecting and quantifying anti-SSA antinuclear antibodies in urine to carry out the method according to any of claims 2 - 4.
  • the method of the present invention can be applied with samples from individuals of any sex, that is, men or women, and at any age. Preferably applied to samples obtained from women.
  • prognosis is understood as the expected evolution of a disease and refers to the assessment of the probability according to which a subject suffers from a disease, as well as the assessment of its onset, state of development, evolution, or of its regression, and/or the prognosis of the course of the disease in the future.
  • an assessment although preferred, may not normally be correct for 100% of the subjects to be diagnosed. The term, however, requires that a statistically significant portion of subjects can be identified as suffering from the disease or having a predisposition to it.
  • the confidence intervals are at least 90%, at least 95%, at least 97%, at least 98% or at least 99%.
  • the p value is less than 0.1, 0.05, 0.01, 0.005 or 0.0001.
  • the present invention allows the disease to be correctly detected differentially by at least 60%, more preferably by at least 70%, much more preferably by at least 80%, or even much more preferably by at least 90%. of the subjects of a certain group or population analyzed.
  • the method of the invention involves comparing the levels of anti-SSA antinuclear antibodies of a urine sample from a test subject with a reference urine sample or with a median value.
  • reference sample means the sample that is used to determine the variation of the levels of anti-SSA antinuclear antibodies of the present invention.
  • the reference value is obtained from the expression values obtained from a sample with healthy individuals.
  • reference samples are taken from several individuals and combined, so that the reference value reflects the mean value of anti-SSA antinuclear antibodies in urine in healthy individuals or "Reference value" of anti-SSA antinuclear antibodies in a reference sample.
  • the expression profile in the preference reference sample can be generated from a population of two or more people. The population, for example, may contain 3, 4, 5, 10, 15, 20, 30, 40, 50 or more people.
  • anti-SSA antinuclear antibodies can be carried out by any means known in the state of the art.
  • expression levels will give a certain “expression profile”.
  • expression level also called “product amount” or “expression product amount” refers to the biochemical material, specifically anti-SSA antinuclear antibodies.
  • the measurement of the amount or concentration of expression product can be carried out directly or indirectly.
  • Direct measurement refers to the measurement of the amount or concentration of antinuclear antibodies.
  • Said signal (which we can also refer to as intensity signal) can be obtained, for example, by measuring an intensity value of a chemical or physical property of said products.
  • Indirect measurement includes measurement obtained from a secondary component or biological measurement system (e.g. measurement of cellular responses, ligands, "tag” or enzymatic reaction products).
  • Quantity refers to, but is not limited to, the absolute or relative quantity of the expression products, as well as any other value or parameter related thereto or that may be derived. of these.
  • Said values or parameters comprise signal intensity values obtained from any of the physical or chemical properties of said expression products obtained by direct measurement. Additionally, said values or parameters include all those obtained by indirect measurement, for example, any of the measurement systems described elsewhere in this document.
  • comparison refers to, but is not limited to, the comparison of the amount of antinuclear antibodies present in the biological sample to be analyzed, also called biological test sample, with a quantity of antibodies antinuclear anti-SSA of one or several reference samples.
  • the reference sample can be analyzed, for example, simultaneously or consecutively, together with the biological test sample.
  • the method for determining the result that is, anti-SSA antinuclear antibodies, need not be particularly limited.
  • the result can be obtained by any of the available techniques, for example by immunoassay.
  • immunoassay refers to any analytical technique that is based on the conjugation reaction of an antibody with an antigen.
  • immunoassays known in the state of the art are, for example, but not limited to: immunoblot, enzyme-linked immunosorbent assay (ELISA), linear immunoassay (LIA), radioimmunoassay (RIA), immunochromatography, immunofluorescence, x-map or chips of protein.
  • the immunoassay is an enzyme-linked immunosorbent assay or ELISA (Enzyme-Linked ImmunoSorbent Assay).
  • the ELISA is based on the premise that an immunoreagent (antigen or antibody) can be immobilized on a solid support, then placing that system in contact with a fluid phase containing the complementary reagent that can bind to a marker compound.
  • an immunoreagent antigen or antibody
  • the marker compound is selected from the list that includes radioisotopes, enzymes, fluorophores or any molecule capable of being conjugated with another molecule or detected and/or quantified directly.
  • This marker compound can bind to the antibody directly, or through another compound.
  • Some examples of marker compounds that bind directly are, but are not limited to, enzymes such as alkaline phosphatase or peroxidase, radioactive isotopes such as 32P or 35S, fluorochromes such as fluorescein or metallic particles, for direct detection by colorimetry, auto-radiography, fluorimetry , or metallography respectively.
  • the amount of anti-SSA antinuclear antibodies can be normalized by normalization methods well known in the art.
  • the expression is considered increased in a sample of the matter under study when the levels of increase with with respect to the reference sample are at least 5%, at least 10%, at least 15%, at least 20%, at least 25%, at least 30%, at least 35 %, at least 40%, at least 45%, at least 50%, at least 55%, at least 60%, at least at least 65%, at least 70% , at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 100%, at least 1 10%, for at least 120%, at least 130%, at least 140%, at least 150%, or more.
  • the expression is considered decreased when its levels decrease with respect to the reference sample by at least 5%, at least 10%, at least 15%, at least 20%, at least 25%, at least 30%, at least 35%, at least 40%, at least 45%, at least 50%, at least 55%, at least 60%) , at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, for at least 100% (that is, absent).
  • the invention provides a method for assigning a human subject into one of two groups: group, which comprises subjects identifiable by the method of the invention and group 2, which represents the remaining subjects.
  • Another aspect of the invention relates to a computer program comprising instructions for carrying out the procedure according to any of the methods of the invention.
  • the invention encompasses computer programs arranged on or in a carrier.
  • the carrier can be any entity or device capable of supporting the program.
  • the carrier could be an integrated circuit in which the program is included and which has been adapted to execute, or to be used in the execution of, the corresponding processes.
  • the programs could be embedded in a storage medium, such as a ROM memory, a CD ROM memory or a semiconductor ROM memory, a USB memory, or a magnetic recording medium, for example, a floppy disk or a disk. hard.
  • a storage medium such as a ROM memory, a CD ROM memory or a semiconductor ROM memory, a USB memory, or a magnetic recording medium, for example, a floppy disk or a disk. hard.
  • the programs could be supported on a transmissible carrier signal; For example, it could be an electrical or optical signal that could be transported via electrical or optical cable, by radio or by any other means.
  • the invention also extends to computer programs adapted so that any processing means can implement the methods of the invention.
  • Computer programs also include cloud applications based on said procedure.
  • Figure 1 Correlation of autoantibody values between serum and urine. Representation of the levels of anti-Scl70, anti-dsDNA, anti-Jo1, anti-Centromere B, anti-RNP, anti-SSB, anti-SSA and anti-Sm autoantibodies in urine, as a normalized ratio of the median of the fluorescence intensity (MFI), versus blood levels, as U/mL. Values of the correlation coefficient (r) close to 0 indicate that there is no correlation, while those close to 1 indicate a strong correlation in the Spearman test. Graphs created with GraphPad Prism.
  • Figure 2 Representation of the urinary levels of anti-SSA (A), anti-histone (B) and anti-Centromere B (C) in which the median and interquartile range are represented.
  • the horizontal dotted line indicates the value from which a sample is considered positive for ANA, so that those points above this line are positive and those below are negative.
  • this threshold is 0.7638, in the case of anti-histone at 0.4697 and in the case of anti-Centromer B at 0.4484.
  • Urinary ANA values are represented as the normalized ratio of the median fluorescence intensity (MFI). *p ⁇ 0.05 ***p ⁇ 0.001.
  • FIG 3 Representation of anti-SSA levels in patients with Sjógren's Syndrome (SjS). They are represented as the normalized ratio of the median fluorescence intensity (MFI) in different groups that were stratified according to the severity of kidney damage, with C being healthy controls, NOT those patients who suffer from the disease but never presented the damage. renal and finally SI, which are those patients who presented kidney damage in the past or currently present it.
  • the arrow symbolizes that the order of these groups is ascending and the stars the level of significance. of these results according to the Jonckheere-Terpstra test **p ⁇ 0.01 ***p ⁇ 0.001 ****p ⁇ 0.0001.
  • the horizontal line indicates the threshold from which the values are considered positive (0.7638).
  • FIG 4 Representation of anti-histone levels in patients with systemic lupus erythematosus (SLE). They were represented as the normalized ratio of the median fluorescence intensity (MFI) in different groups that were stratified according to the severity of kidney damage, with C- being the control, NOT those patients who suffer from the disease but never presented the damage. renal and finally SI, which are those patients who presented kidney damage in the past or currently present it.
  • MFI median fluorescence intensity
  • the arrow symbolizes that the order of these groups is descending and the stars represent the level of significance of these results according to the Jonckheere-Terpstra test *p ⁇ 0.05.
  • the horizontal line indicates the threshold from which the values are considered positive (0.4697).
  • Figure 5 Representation of the results of the Mann-Whitney test.
  • the groups C- versus NO, NO versus YES and C- versus YES faced each other.
  • the horizontal line indicates the threshold from which the values are considered positive (0.4697).
  • SAD systemic autoimmune diseases
  • SLE systemic lupus erythematosus
  • SjS Sjógren's syndrome
  • PAPs antiphospholipid syndrome
  • MCTD mixed connective tissue disease
  • UCTD undifferentiated disease. connective tissue
  • Table 1 Compilation of the samples to be used grouped according to disease and classified according to sex and age.
  • CTRL Healthy controls.
  • the Age data were grouped into intervals equivalently.
  • Pregnant women, cases of neonatal or drug-induced lupus, or those patients whose status did not allow participation in the study were excluded.
  • certain treatments are also excluded: stable doses of spheroids >15 mg/day or with IV corticosteroids in the 3 months prior to inclusion, immunosuppressive treatment in the previous 3 months (metrotrexate > 25 mg/week; azathiopyrine > 2.5 mg/kg/day; cyclospohna A > 3 mg/kg/day; mycophenolate mofetil > 2 g/day), treatment with cyclophosphamide or belimumab in the last 6 months and patients on combination therapy with two or more immunosuppressants or with biological depleting therapy (e.g. Rituximab) in the previous year.
  • kidney damage parameters to take into account, and the criteria to determine if they are positive, are the following:
  • Abnormal creatinine levels serum creatinine levels equal to or greater than 20% of the normal creatinine range, creatinine levels greater than a pre-established baseline, or reduced glomerular filtration rate below 60 mL/min.
  • Urine samples were taken at the time of inclusion in the study. These samples, with an approximate volume of 50 mL, were centrifuged to eliminate the urinary sediment and the supernatant was aliquoted and frozen at -80°C in the Biobank of Andalusia.
  • the AtheNA Multi-Lyte ANA-II Plus kit from ZEUS Scientific was used, based on Multiplex XMap technology that allows multiplexed immunoassays to be carried out.
  • This technology is based on high-performance screening using polystyrene microspheres conjugated to the antigen complementary to the autoantibody to be studied in the urine, so that if it is present in the patient's urine it will bind to the immobilized antigen.
  • it is combined with optical techniques based on flow cytometry; by knowing which ligand has been used in each of the spheres, it is possible to know the composition of the sample (“XMAP Technology", 2014). This allows us to quantify multiple analytes in a single well with a smaller sample volume, thereby reducing the expense and time involved in executing the protocol.
  • this kit includes a series of internal calibrators that allow the calculation of errors, eliminating the need for duplicates, a technology called Intra-Well Calibration.
  • This includes an internal standard curve within the microsphere suspension, which allows each well to be calibrated automatically internally, adjusting based on the characteristics of the sample (ZEUS Scientific, 2017).
  • the antigens included for the assay are: Scl-70 (topoisomerase I), dsDNA, Jo-1 (aminoacyl-tRNA synthetases), Centromere B, RNP (anti nuclear ribonucleoprotein U 1), SSB, SSA, Histone and Sm. Each of them is a target for an autoantibody that is preferentially associated with a specific disease (Table 2).
  • Table 2 Relationship between the autoantibody present in urine and the disease to which its appearance is associated. Table taken from the ZEUS Scientific protocol. (12-14-2017). AtheNA Multi-Lyte ANA-II Plus Test System. (REF: A21101).
  • the kit contains the microsphere suspension, the goat anti-human IgG conjugate (specific for the y chain) conjugated with phycoerythrin, a negative control and three positive controls of human serum, SAVe Diluent and washing buffer (10X). Additionally, a negative urine control obtained from the PRECISEADS project is used.
  • the urine samples After allowing the urine samples to thaw in the refrigerator, they are centrifuged at 10,000 g for 5 min at room temperature and taking the supernatant, a 1:10 dilution is prepared using SAVe Diluent (1:21 dilution for serum). Next, the microspheres are resuspended, a step of great importance so that their distribution is as homogeneous as possible, and 25 pL are added to a 96-well plate with a conical bottom. To these, 5 pL of the controls and the diluted samples were added and allowed to incubate at room temperature for 30 min while shaking at 500 revolutions per minute (rpm).
  • rpm revolutions per minute
  • the plate was washed with 1X washing buffer and centrifuged at 2000 rpm for 7 min. This process was repeated a total of three times. Afterwards, 75 pL of conjugated anti-human IgG was added, which, being sensitive to light, must be stored in the dark, for which it is covered with aluminum foil. Finally, it is allowed to incubate for 30 min at room temperature while shaking at 500 rpm and the results are read using the Luminex 200 reader. These results are accepted when the number of each of the different beads per well is equal to or greater than 50.
  • the Luminex 200 reader returns information on the fluorescence intensity detected in each population of beads.
  • the normalized ratio of the median fluorescence intensity (MFI) is used, obtained using the Bio-Plex Manager 6.0 software. The data obtained was compiled in an Excel document along with the patient's diagnosis and kidney damage parameters.
  • Non-parametric analyzes were carried out on the collected data, since the data being worked on do not follow a normal distribution. They were analyzed using Spearman's Rho correlation comparing autoantibody levels in serum and urine, the Mann-Whitney U test, which compares whether two independent groups are similar or not, and the Jonckheere-Terpstra test, which indicates whether Different groups present an ascending or descending order, starting from groups they present an a priori order, in this case the severity of the kidney damage. When the p value is less than 0.05 the results are considered statistically significant, increasing their significance the lower the p value.
  • ANA values in serum and urine of those sick patients were correlated to confirm that the urine of patients with systemic autoimmune diseases may contain antinuclear autoantibodies of different types. specificities, and that said concentration is not a reflection of the values circulating in the blood.
  • a Spearman's Rho correlation test was carried out, which is indicative of the dependence shown by two non-parametric groups.
  • ANA levels are represented as the normalized ratio of the median fluorescence intensity (MFI) while in blood they are represented as units per milliliter (U/rnL). ( Figure 1).
  • the group of patients suffering from IICTD also have higher levels, but in this case the difference is less significant (p ⁇ 0.022).
  • anti-histone values p ⁇ 0.035
  • the significant differences between the control group and the group of patients suffering from SLE are due to the fact that in the latter the measured values are lower.
  • the levels of anti-Centromer B in PAPS are slightly elevated, showing a tendency to show significant differences compared to the control.
  • Table 4 Results obtained from the Jonckheere-Terpstra test on kidney damage parameters and anti-SSA levels in SjS. **p ⁇ 0.01;***p ⁇ 0.001;****p ⁇ 0.0001 (from lowest to highest shading intensity).
  • a Mann-Whitney test was also performed on the groups of patients stratified by renal severity, comparing the C- group versus the NO group, the latter in turn with the SI group and C- versus SI ( Figure 5). .
  • the results obtained after analyzing the groups using the Mann-Whitney analysis show significant differences between the healthy control group and the group currently presenting kidney damage for all parameters.
  • the kidney damage parameters that present significant differences between the C- and NO group are only the abnormal lipid profile, nephritis, reduced C3 and 04 levels, and systemic hypertension.
  • kidney damage parameters that present significant differences between the NO and YES groups are abnormal creatinine levels, abnormal urine analysis, proteinuria, abnormal protein/creatinine ratio and reduced 04 levels, this being The latter is the only one that presents significant differences between all groups.
  • the groups stratified for abnormal creatinine levels in SLE did not present significant differences between any of them.
  • SSA is an RNA-binding protein, it binds to a small non-coding RNA known as YRNA, and is believed to be involved in cellular repair mechanisms for damage resulting from exposure to ultraviolet radiation. These YRNA sequences are exposed as a consequence of cellular damage caused by the pro-apoptotic conditions of the autoimmune response.
  • Anti-SSA autoantibodies indicate an intermediate risk of suffering from SjS, with high levels of this being highly predictive for the diagnosis of SjS.
  • positive anti-SSA titers are associated with the presence of rheumatoid factor (RF), which correlates with early stages of SjS in which hypergammaglobulinemia predominates.
  • RF rheumatoid factor
  • it must be taken into account that it is not the only determining factor for the prognosis, and it is not specific to this EAS, but rather positive anti-SSA titers must be present along with dry eyes and mouth and a biopsy that show lymphocytic infiltration (Veenbergen et al., 2022; Zandonella Callegher et al., 2022). No publications on urinary anti-SSA were found in SjS.
  • anti-histone levels show differences according to the Mann-Whitney analysis between the control group and the SLE group, with these levels being decreased in the latter group.
  • Positive anti-histone titers are common in SLE, especially in drug-induced SLE; however, these cases were not included in the study. It seems that serum anti-histone levels correlate with the severity of lupus nephritis (LN), but not with the systemic activity that the disease can present.
  • LN lupus nephritis
  • the results obtained in this work suggest that anti-histone autoantibodies are related to some protective mechanism in SLE, as can be seen in Figure 4. No publications were found on anti-histone studies in the urine of patients with SLE.
  • UCTD undifferentiated connective tissue disease

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Abstract

La présente invention concerne l'utilisation in vitro du niveau d'anticorps antinucléaires dans l'urine, anti-SSA, en tant que biomarqueur précoce pour diagnostiquer et/ou effectuer le pronostic de la lésion rénale causée par le syndrome de Sjörgen.
PCT/ES2023/070696 2022-11-23 2023-11-22 Anticorps antinucléaires anti-ssa en tant que biomarqueurs précoces de la lésion rénale causée par le syndrome de sjörgen WO2024110683A1 (fr)

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ES202231008A ES2975744A1 (es) 2022-11-23 2022-11-23 Metodo de diagnostico de patologias renales causadas por enfermedades autoinmunes
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Title
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