Lung cancer bone metastasis marker

Abstract

The invention discloses a lung cancer bone metastasis marker, and belongs to the technical field of biomarkers. The lung cancer bone metastasis marker provided by the invention can realize early-stage, non-invasive and accurate detection of lung cancer bone metastasis, effectively improve the diagnosis sensitivity and specificity, provide reliable basis for clinical prognosis evaluation, relapse monitoring and individualized treatment, has the advantages of simplicity and convenience in operation, good repeatability, high cost effectiveness and the like, and has a wide application prospect. The life quality of a patient can be improved; the medical burden is reduced. According to the invention, the technical blank of lung cancer bone metastasis specific markers is filled, the crossing from image diagnosis to molecular early warning and from empirical treatment to accurate intervention is realized, and the marker has important clinical significance and application value for improving the survival rate and life quality of lung cancer patients.

Description

TECHNICAL FIELD

[0001] The present application relates to the technical field of biomarkers, in particular to a lung cancer bone metastasis marker. BACKGROUND

[0002] Lung cancer is a high-incidence malignant tumor in China and even the world, and its incidence and mortality rate are increasing year by year. Bone metastasis is a common site of extrapulmonary metastasis of lung cancer. Previous studies have found that about 20-30% of lung cancer patients have bone metastasis at the time of diagnosis, and another 35-40% of cases develop bone metastasis during the course of the disease. With the continuous breakthroughs in basic medical research and clinical diagnosis and treatment methods in the field of lung cancer in recent years, molecular targeted therapy, immunotherapy, etc. have gradually become new ways of precise treatment for lung cancer, and have achieved certain clinical efficacy. While the mortality rate of patients is reduced and the survival period is prolonged, the incidence of bone metastasis and skeletal related events (SRE) also potentially increases. Bone pain, pathological fractures, hypercalcemia, spinal metastasis compression, paraplegia, etc. caused by SRE not only reduce the quality of life of patients, limit their mobility, increase treatment costs and death risk, but also have a great damage to the efficacy of tumor treatment for patients. Therefore, it is urgent to explore the etiology and pathogenesis of lung cancer bone metastasis, and it is imminent to seek new prediction targets and treatment entry points.

[0003] Currently, the prediction targets for lung cancer bone metastasis are mostly in the basic research stage, and there is still a distance from clinical wide application. For example, the newly discovered GATA3-driven ceRNA network (such as XLOC_006941 / hsa-miR-543 / NPRL3 axis) and SOSTDC1, MIR4697HG, etc. molecular markers are mostly based on retrospective small sample studies, and their prediction efficiency is generally lack of verification in prospective, large-scale clinical cohorts. Moreover, they are high in cost and not suitable for clinical promotion. In addition, lung cancer bone metastasis involves a complex process of multiple steps and multiple genes, and the molecular mechanisms of different pathological subtypes are different. At the same time, part of the osteoclasts release factors such as osteopontin (OPN) to systematically remodel the immune microenvironment, thereby affecting the treatment response and prognosis. SUMMARY

[0004] The purpose of the present application is to provide a lung cancer bone metastasis marker to solve the problems existing in the prior art.

[0005] To achieve the above purpose, the present application provides the following solutions. One of the technical solutions of the present application is that the levels of CD4, IL-22 and IL-17 are used as detection targets in the preparation of a product for diagnosing whether lung cancer has bone metastasis.

[0006] The second technical scheme of the present application is a product for diagnosing whether lung cancer has bone metastasis, taking CD4, IL-22 and IL-17 levels as detection targets.

[0007] Based on the above technical scheme, the present application has the following technical effects: The lung cancer bone metastasis marker provided by the present application can realize early, non-invasive and accurate detection of lung cancer bone metastasis, effectively improve the diagnostic sensitivity and specificity, provide reliable basis for clinical prognosis evaluation, recurrence monitoring and individualized treatment, has the advantages of simple operation, good repeatability, cost-effectiveness and the like, and helps to improve the quality of life of patients and reduce the medical burden. The present application fills the technical gap of specific markers for lung cancer bone metastasis, realizes the leap from "image diagnosis" to "molecular early warning" and from "experience treatment" to "precise intervention", and has important clinical significance and application value for improving the survival rate and quality of life of lung cancer patients. BRIEF DESCRIPTION OF DRAWINGS

[0008] In order to more clearly illustrate the technical schemes in the embodiments of the present application or the prior art, the drawings needed in the embodiments will be briefly introduced as follows. Obviously, the drawings in the following description are only some embodiments of the present application, and those skilled in the art can obtain other drawings according to these drawings without creative labor.

[0009] Figure 1 The level of Th22 cells in peripheral blood PBMCs of different distant metastasis IV patients. DETAILED DESCRIPTION

[0010] The various exemplary embodiments of the present application will now be described in detail, which should not be considered as limiting the present application, but should be understood as a more detailed description of some aspects, characteristics and embodiments of the present application.

[0011] It should be understood that the terms described in the present application are only for describing the specific embodiments, and are not used to limit the present application. In addition, for the numerical range in the present application, it should be understood that each intermediate value between the upper limit and the lower limit of the range is also specifically disclosed. Each smaller range between any stated value or intermediate value in the range and any other stated value or intermediate value in the range is also included in the present application. The upper limit and the lower limit of these smaller ranges can be independently included or excluded from the range.

[0012] Unless otherwise stated, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art. While only preferred methods and materials have been described herein, any methods and materials similar or equivalent to those described herein may be used in the implementation or testing of this invention. All references to this specification are incorporated by way of citation to disclose and describe methods and / or materials associated with those references. In the event of any conflict with any incorporated reference, the content of this specification shall prevail.

[0013] Various modifications and variations can be made to the specific embodiments described in this specification without departing from the scope or spirit of the invention, as will be apparent to those skilled in the art. Other embodiments derived from this specification will also be obvious to those skilled in the art. This application specification and embodiments are merely exemplary.

[0014] The terms “include,” “including,” “have,” “contain,” etc., used in this article are all open-ended terms, meaning that they include but are not limited to.

[0015] Unless otherwise specified, the technical solutions described in this invention are all conventional solutions in the field, and the reagents or raw materials used are all purchased from commercial channels or are publicly available unless otherwise specified.

[0016] This invention provides the application of CD4, IL-22, and IL-17 levels as detection targets in the preparation of products for diagnosing whether lung cancer has bone metastases.

[0017] In some specific implementations, the product is used to detect the levels of CD4, IL-22, and IL-17 in PBMC cells of patient samples.

[0018] In some specific implementations, the method for diagnosing whether lung cancer has metastasized to bone is as follows: the levels of CD4, IL-22, and IL-17 in the PBMC cells of the patient to be tested are detected and compared with the control sample or reference sample. If the levels of CD4, IL-22, and IL-17 are significantly upregulated, it indicates that the patient's lung cancer has metastasized to bone or has multiple metastases.

[0019] In some specific implementations, the product includes reagents, kits, chips, test strips, or high-throughput sequencing platforms.

[0020] This invention also provides a product for diagnosing whether lung cancer has metastasized to the bone, using CD4, IL-22 and IL-17 levels as detection targets.

[0021] In some specific implementations, the product includes reagents, kits, chips, test strips, or high-throughput sequencing platforms.

[0022] Example 1 1 Sample collection and isolation of peripheral blood mononuclear cells (PBMC) Peripheral blood (from outpatients and inpatients with lung cancer in the First Affiliated Hospital of Zhejiang University School of Medicine from January 2023 to December 2024) was collected with sodium heparin or sodium citrate anticoagulation.

[0023] Inclusion criteria: Collect peripheral blood from healthy controls, patients with newly diagnosed lung cancer without distant metastasis, patients with bone metastasis, and patients with distant metastasis (excluding bone metastasis); Exclusion criteria: 1. Received anti-tumor treatment; 2. Based on a history of autoimmune disease; 2. Combined with other tumors; 3. Combined with tuberculosis / HIV; 4. Combined with acute infectious diseases within 4 weeks; 5. Long-term use of hormone or immunosuppressive therapy; 6. Expected survival less than 3 months.

[0024] PBMCs were isolated using Ficoll density gradient centrifugation. Cells were washed twice with PBS, counted, and adjusted to the concentration for standby.

[0025] 2 Cell stimulation and culture Resuspend PBMCs in complete medium containing stimulants and protein transport inhibitors.

[0026] Stimulants: Use cell stimulation cocktail (such as PMA 20 ng / mL and Ionomycin 1 μg / mL).

[0027] Protein transport inhibitors: Must add monensin 2 μmol / L or brefeldin A 10 μg / ml to prevent cytokine secretion into the extracellular space and accumulate in the intracellular space.

[0028] Conditions: Incubate in a 37°C, 5% CO2 incubator for 4-6 hours. Do not add IL-4 to induce Th2 cell differentiation and interfere with Th22 cell identification.

[0029] 3 Cell surface staining After incubation, transfer the cells to a flow tube and wash with pre-cooled PBS.

[0030] Add 5 μL of anti-human CD4 antibody (usually fluorescently labeled such as FITC or PerCP-Cy5.5), and incubate on ice for 15-30 minutes in the dark. Wash twice with PBS containing 2% fetal bovine serum to remove unbound antibodies.

[0031] Fixation and membrane rupture: add fixative, paraformaldehyde, and incubate at room temperature for 20 minutes in the dark. After washing with PBS, add membrane rupture reagent.

[0032] Intracellular staining: add 10 μL of anti-IL-22 antibody (usually PE or APC labeled) and 10 μL of anti-IL-17 antibody (PE-Cy7 labeled) in membrane rupture buffer.

[0033] Incubate for 30 minutes at room temperature in the dark.

[0034] Wash twice with membrane rupture buffer, and finally resuspend the cells in PBS, ready for machine analysis.

[0035] Flow machine detection and data analysis: finally, in CD4+T cells, analyze the IL-22+IL-17- double positive cell population, which is Th22 cells.

[0036] The results are shown in Table 1. Figure 1 As shown in Table 1, there is a significant difference in CD4+IL-22+IL-17 in PBMC cells between IVa, IVB bone metastasis, IVB-liver metastasis, IVB-brain metastasis, and IVB-multiple metastasis, and the level of CD4+IL-22+IL-17 can be used to determine whether lung cancer has bone metastasis or multiple metastasis.

[0037] Obviously, the above embodiments of the present application are merely examples for clearly illustrating the present application, and are not intended to limit the embodiments of the present application. Based on the above description, other different forms of changes or variations can be made by those of ordinary skill in the art. Here, it is not necessary and impossible to exhaust all embodiments. Any modification, equivalent replacement, and improvement, etc. within the spirit and principles of the present application should be included in the protection scope of the claims of the present application.

Claims

1. Use of CD4, IL-22 and IL-17 levels as detection targets in the preparation of a product for diagnosing whether lung cancer has metastasized to bone.

2. Use according to claim 1, characterized in that, The product is used to detect the levels of CD4, IL-22 and IL-17 in PBMC cells of a patient sample.

3. Use according to claim 1, characterized in that, The method for diagnosing whether lung cancer has metastasized to bone is to detect the levels of CD4, IL-22 and IL-17 in PBMC cells of a patient to be tested, and compared with a control sample or a reference sample, if the levels of CD4, IL-22 and IL-17 are significantly up-regulated, it indicates that the lung cancer of the patient has metastasized to bone or multiple metastasis.

4. Use according to claim 1, characterized in that, The product includes reagents, kits, chips, test papers or high-throughput sequencing platforms.

5. A product for diagnosing whether lung cancer has bone metastasis, characterized by, CD4, IL-22 and IL-17 levels are used as detection targets.

6. The product of claim 5, wherein, The product includes reagents, kits, chips, test papers or high-throughput sequencing platforms.

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