Application of SNCG protein in auxiliary diagnosis of prostate cancer

Abstract

The invention discloses application of SNCG protein in auxiliary diagnosis of prostate cancer, and relates to the technical field of biological medicine. The application way is as follows: the reagent for specifically detecting the expression of the SNCG protein in a biological sample is used for preparing a product for auxiliary diagnosis of the prostate cancer. Proteome analysis on urine proteins of a prostate cancer patient and a normal male shows that the concentration of the SNCG protein in the urine of the prostate cancer patient is obviously higher than that of the normal male group, and immunohistochemical analysis on archive wax blocks and surrounding normal prostate tissues of the prostate cancer patient shows that the SNCG protein can be applied to the prostate cancer patient. The SNCG protein has a certain expression rate in the prostate cancer, which indicates that the SNCG protein has a certain relationship with the occurrence and development of the prostate cancer, and can be used for assisting the diagnosis of the prostate cancer and the diagnosis of difficult cases so as to improve the diagnosis rate and the survival rate of patients.

Description

Technical Field

[0001] The present invention relates to the field of biomedicine technology, and in particular to the application of SNCG protein in auxiliary diagnosis of prostate cancer. Background Art

[0002] Prostate cancer (PCa) is a common malignant tumor in men, with its incidence increasing in recent years. Prostate cancer is typically diagnosed through digital rectal examination and / or prostate-specific antigen (PSA) screening. Elevated serum PSA levels may indicate the presence of PCa. Because PSA is secreted only by prostate cells, it is used as a marker for prostate cancer. The insidious nature of prostate cancer's clinical manifestations makes its early diagnosis and treatment less effective. The importance of early screening, treatment, and prognosis for prostate cancer is self-evident.

[0003] SNCG, or gamma-synuclein, is a member of the synuclein family. Its gene is located at 10q23.2, and the encoded protein is widely distributed throughout the body. Under normal circumstances, SNCG is primarily expressed in the peripheral nervous system, retina, and adipose tissue. Within the nervous system, it plays a role in the integrity of the neurofilament network and may be involved in regulating axonal structure during development and adulthood.

[0004] SNCG is closely associated with a variety of diseases. In terms of neurodegenerative diseases, although it has no obvious relationship with neurodegenerative diseases under normal circumstances, its expression has been observed to change in the retina of patients with Alzheimer's disease. In tumor diseases, SNCG was first discovered in a human breast cancer cDNA library. It is also known as breast cancer susceptibility gene 1 (Liu J, Spence MJ, Zhang YL, Jiang Y, Liu YE, Shi YE. Transcriptional suppression of synuclein gamma (SNCG) expression in human breast cancer cells by the growth inhibitory cytokineoncostatin M. Breast Cancer Res Treat. 2000; 62: 99-107. doi: 10.1023 / A: 1006418219012.). Its expression is elevated in a variety of tumors and can promote tumor cell growth, invasion, and metastasis, interfere with cell apoptosis, and is also associated with tumor stage. It is an independent indicator for judging poor prognosis. To date, according to public reports, for example, (Zhang J, Liu XH, Li C, Wu XX, Chen YL, Li WW, Li X, Gong F, Tang Q, Jiang D. SNCG promotes the progression and metastasis of high-grade serous ovarian cancer via target the PI3K / AKT signaling pathway.J Exp Clin Cancer Res.2020May7;39(1):79.Doi:10.1186 / s13046-020-01589-9.Erratumin:J Exp Clin CancerRes.2020Jun 17;39(1):115.Doi:10.1186 / s13046-020-01596-w.PMID:32381034;PMCID:PMC7204046);Pan Y, Zheng Y, Yang J, Wei Y, Wu H, Liu S, Yin A, Hu J, Zeng YA new biomarker for the early diagnosis of gastric cancer cancer:gastric juice-and serum-derived SNCG.Futu re Oncol.2022Sep;18(28):3179-3190.doi:10.2217 / fon-2022-0253.Epub 2022Aug10.PMID:35947016.). SNCG overexpression has been confirmed in a variety of malignant solid tumors, including breast, ovarian, uterine, liver, and cervical cancers. Currently, there are no reports linking SNCG protein to the development and progression of prostate cancer. Exploring the relationship between SNCG protein and prostate cancer could provide a theoretical basis for prostate cancer screening, diagnosis, and prognosis prediction. Summary of the Invention

[0005] The purpose of the present invention is to overcome the deficiencies of the prior art and provide an application of SNCG protein in the auxiliary diagnosis of prostate cancer.

[0006] The present invention achieves the above-mentioned purpose through the following technical solutions:

[0007] As a first aspect of the present invention, the present invention provides the use of SNCG protein in assisting the diagnosis of prostate cancer.

[0008] A further improvement is that the amino acid sequence of the SNCG protein is shown in SEQ ID No. 1, and the GeneBank accession number of the SNCG protein is O76070.

[0009] A further improvement is that the biological sample is prostate tissue or cells, blood or urine.

[0010] A further improvement is that the detection is performed using an immunochemical method.

[0011] A further improvement is that the immunochemical method is any one of enzyme-linked immunosorbent assay or immunoblotting.

[0012] A further improvement is that the reagent is a binding agent that specifically binds to the SNCG protein, an oligonucleotide probe that specifically recognizes the gene encoding the SNCG protein, or a primer pair that specifically amplifies the gene encoding the SNCG protein.

[0013] A further improvement is that the binding agent is a monoclonal antibody or a polyclonal antibody.

[0014] A further improvement is that the product is any one of a detection kit, a detection test paper, colloidal gold, a color developing card, a detection scale, a chip or a probe.

[0015] The present invention has the following beneficial effects:

[0016] The urine proteome analysis of the present invention found that the content of SNCG protein in the urine of prostate cancer patients was higher than that in the normal male group. Immunohistochemical analysis showed that the basal cells of the prostate acini of prostate cancer patients disappeared, and 26.7% of the prostate cancer patient group were positive, and the positive intensity and breadth were greater than the expression of normal prostate acini (P<0.05).

[0017] The present invention indicates that SNCG protein has a certain relationship with the occurrence and development of prostate cancer, can be used to assist in the diagnosis of prostate cancer, can provide guidance for the treatment of prostate cancer, and has good application prospects. BRIEF DESCRIPTION OF THE DRAWINGS

[0018] Figure 1 The expression of SNCG protein in prostate cancer tissue and prostate acinar provided by the present invention. In the figure, AB shows different areas of prostate cancer tissue from the same patient; C shows a positive area in one patient; D shows basal cell staining of normal prostate gland (red arrow) and localized hyperplasia of basal cells (white arrow). DETAILED DESCRIPTION

[0019] The present application is described in further detail below in conjunction with the accompanying drawings. It is necessary to point out that the following specific implementation methods are only used to further illustrate the present application and cannot be understood as limiting the scope of protection of the present application. Technicians in this field can make some non-essential improvements and adjustments to the present application based on the above application content.

[0020] The experimental methods in the following examples are conventional biochemical methods unless otherwise specified, and the experimental materials used in the following examples are purchased from conventional biochemical reagent stores unless otherwise specified.

[0021] 1. Urine proteome analysis

[0022] In October 2024, 20 ml of morning urine was collected from 10 patients clinically diagnosed with prostate cancer before surgery as the experimental group. 20 ml of urine was collected from 5 normal men as the control group. Proteomics was analyzed using the T test. The analysis process was as follows:

[0023] (1) Protein extraction

[0024] Urine samples were taken out from -80°C, and pH 8.0 Tris-HCl was added to all samples to a final concentration of 50 mM. The samples were centrifuged at 1000g for 5 min, and the supernatant was collected. The samples were centrifuged at 17000g for 10 min, and an equal volume of methanol and 1 / 4 volume of chloroform were added to the supernatant. The samples were shaken for 15 s and then placed at room temperature for 5 min. The samples were centrifuged at 12000g for 15 min at room temperature, and the supernatant was discarded. An equal volume of methanol was added, shaken for 15 s, and the samples were centrifuged at 12000g for 15 min at room temperature. The supernatant was discarded, and 80 μl of lysis buffer (containing 1% SDC) was added for resolubilization. The protein concentration was determined using a BCA kit.

[0025] (2) Enzymatic hydrolysis with pancreatic enzymes

[0026] An equal amount of protein from each sample was enzymatically digested. The volume was adjusted to the same value with lysis buffer, and dithiothreitol (DTT) was added to a final concentration of 5 mM. The cells were reduced at 56°C for 30 minutes. Iodoacetamide (IAA) was then added to a final concentration of 11 mM, and the cells were incubated at room temperature in the dark for 15 minutes. TEAB was added to dilute urea to ensure that the concentration was below 2 M. Trypsin was added at a ratio of 1:50 (protease: protein, m / m) and the cells were digested overnight. Trypsin was then added at a ratio of 1:100 (protease: protein, m / m) and the digestion continued for 4 hours.

[0027] (3) Liquid chromatography-mass spectrometry analysis

[0028] Peptides obtained by trypsin digestion were dissolved in liquid chromatography mobile phase A and separated using a NanoElute ultra-high performance liquid chromatography system. Mobile phase A consisted of 0.1% formic acid and 2% acetonitrile in water; mobile phase B consisted of acetonitrile-water containing 0.1% formic acid. The gradient was set as follows: 6% to 24% B (0-14 min); 24% to 35% B (14-16 min); 35% to 80% B (16-18 min); and 80% B (18-20 min). The flow rate was maintained at 500 nl / min. Following ultra-high performance liquid chromatography separation, the peptides were injected into the Capillary ion source for ionization and then acquired on a timsTOF Pro 2 mass spectrometer. The source voltage was set to 1.75 kV. The peptide precursor ions and their secondary fragments were detected and analyzed using the TOF. The data acquisition mode used was data-independent parallel accumulation serial fragmentation (dia-PASEF) mode. The primary mass spectrometry scan range was set to 300-1500 m / z. After one primary mass spectrum was acquired, 20 PASEF mode acquisitions were performed. The secondary mass spectrometry scan range was 400-850, with every 7 m / z as a window.

[0029] Using urine proteome analysis, a series of analyses including protein extraction, trypsin hydrolysis, and liquid chromatography-mass spectrometry analysis revealed that the concentration of SNCG protein in the urine of prostate cancer patients was significantly higher than that in the normal male group (P=2.45119E-06). The results are shown in Table 1.

[0030] Table 1. Comparison of SNCG protein content in urine of prostate cancer patients and normal male subjects

[0031]

[0032] Note: C1 to C5 are normal male groups; P1 to P10 are prostate cancer groups.

[0033] 2. Immunohistochemical Analysis

[0034] Paraffin blocks from 30 prostate cancer patients between 2019 and 2024, along with 15 adjacent normal prostate tissues, were collected. OriGene's monoclonal antibody, SNCG (clone number: TA423304), was purchased at a dilution of 1:200. Immunohistochemistry was performed using the Envision one-step method, with 10% or more of the cancer cells in the slice being considered positive. SNCG protein was primarily expressed in the cytoplasm, with some nuclear staining. Statistics were performed using SPSS 25.0, using Fisher's exact test.

[0035] Immunohistochemical staining revealed that the basal cells of the prostate acini of prostate cancer patients disappeared, and 26.7% of the prostate cancer patients were positive, with the intensity and breadth of the positive staining greater than that of normal prostate acini (P<0.05). The basal cells of normal prostate acini were stained, as were the proliferating basal cells and the urothelium. In addition, the experiment also observed the staining of the smooth muscle tissue of the normal blood vessel wall, the staining of nerve fibers and ganglion cells, and the staining of vascular endothelial cells ( Figure 1 ).

[0036] The above-described embodiments merely illustrate several implementations of the present invention. While the descriptions are relatively specific and detailed, they should not be construed as limiting the scope of the present invention. It should be noted that a person skilled in the art would be able to make numerous variations and improvements without departing from the spirit of the present invention, and all such variations and improvements fall within the scope of protection of the present invention.

Claims

1. The application of SNCG protein in assisting the diagnosis of prostate cancer is characterized by: The application approach is to use a reagent for specifically detecting SNCG protein expression in a biological sample to prepare a product for assisting in the diagnosis of prostate cancer.

2. The use according to claim 1, characterized in that The amino acid sequence of the SNCG protein is shown in SEQ ID No.

1.

3. The use according to claim 1, characterized in that The biological sample is prostate tissue or cells, blood or urine.

4. The use according to claim 1, characterized in that The detection was performed using an immunochemical method.

5. The use according to claim 4, characterized in that The immunochemical method is any one of enzyme-linked immunosorbent assay or immunoblotting.

6. The use according to claim 1, characterized in that The reagent is a binding agent that specifically binds to the SNCG protein, an oligonucleotide probe that specifically recognizes the gene encoding the SNCG protein, or a primer pair that specifically amplifies the gene encoding the SNCG protein.

7. The use according to claim 6, characterized in that The binding agent is a monoclonal antibody or a polyclonal antibody.

8. The use according to claim 1, characterized in that The product is any one of a detection kit, a detection test paper, colloidal gold, a color developing card, a detection scale, a chip or a probe.

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