Novel biomarker for early warning of recurrence of chordoma and application thereof

Abstract

The application relates to the technical field of medicines, and discloses a novel biomarker for early warning of recurrence of chordoma and application, wherein the biomarker is Notch1 intracellular segment O-GlcNAc glycosylation modification. Notch1 intracellular segment O-GlcNAc glycosylation modification is significantly up-regulated in recurrent chordoma, and chordoma patients with high Notch1 intracellular segment O-GlcNAc glycosylation level are more likely to early relapse than chordoma patients with low Notch1 intracellular segment O-GlcNAc glycosylation level; and inhibition of Notch1 and O-GlcNAc glycosylation modification can inhibit tumor growth. Therefore, Notch1 intracellular segment O-GlcNAc glycosylation can be used as a novel biomarker for early warning of recurrence of chordoma, and targeting Notch1 intracellular segment O-GlcNAc glycosylation is a novel treatment strategy for recurrent chordoma.

Description

Technical Field

[0001] This invention relates to the field of pharmaceutical technology, and more specifically, to a novel biomarker for early warning of chordoma recurrence and its application. Background Technology

[0002] Chordoma is a rare primary malignant bone tumor with an annual incidence of approximately one in a million. Although histologically considered a low-grade tumor, chordoma has a recurrence rate exceeding 60%, with a significant proportion of patients experiencing multiple recurrences, making it a leading cause of death. Currently, treatment for recurrent chordoma primarily involves surgery, but repeat surgery is extremely difficult and risky. Furthermore, chordoma is insensitive to radiotherapy and chemotherapy, and there are no effective biological therapies. Therefore, the treatment of recurrent chordoma faces significant challenges. Identifying early warning diagnostic biomarkers for chordoma recurrence, deeply exploring the mechanisms of chordoma recurrence, revealing key factors of recurrence, and developing new therapeutic targets are crucial for improving patient prognosis. Summary of the Invention

[0003] To overcome the aforementioned deficiencies in existing technologies, this invention provides a novel biomarker for early warning of chordoma recurrence and its application. Notch1 intracellular O-GlcNAc glycosylation modification is significantly upregulated in recurrent chordoma, and chordoma patients with high levels of Notch1 intracellular O-GlcNAc glycosylation are more prone to early recurrence compared to patients with low levels of Notch1 intracellular O-GlcNAc glycosylation. Furthermore, inhibiting Notch1 and O-GlcNAc glycosylation modification inhibits tumor growth. Therefore, Notch1 intracellular O-GlcNAc glycosylation can serve as a novel biomarker for early warning of chordoma recurrence, and targeting Notch1 intracellular O-GlcNAc glycosylation is a novel treatment strategy for recurrent chordoma.

[0004] The above-mentioned technical objective of the present invention is achieved through the following technical solution: a novel biomarker for early warning of chordoma recurrence, wherein the biomarker is the intracellular O-GlcNAc glycosylation modification of Notch1.

[0005] Application of a novel biomarker for early warning of chordoma recurrence in early warning of chordoma recurrence.

[0006] Application of a novel biomarker for early warning of chordoma recurrence in the treatment of recurrent chordoma.

[0007] In summary, this invention offers the following advantages: Notch1 intracellular O-GlcNAc glycosylation modification is significantly upregulated in recurrent chordoma, and chordoma patients with high levels of Notch1 intracellular O-GlcNAc glycosylation are more prone to early recurrence compared to patients with low levels of Notch1 intracellular O-GlcNAc glycosylation; furthermore, inhibiting Notch1 and O-GlcNAc glycosylation modification inhibits tumor growth. Therefore, Notch1 intracellular O-GlcNAc glycosylation can serve as a novel biomarker for predicting chordoma recurrence, and targeting Notch1 intracellular O-GlcNAc glycosylation is a novel therapeutic strategy for recurrent chordoma. Attached Figure Description

[0008] Figure 1 This is a comparison of the results of the intracellular O-GlcNAc glycosylation level of Notch1 in primary and recurrent chordoma in this embodiment of the invention;

[0009] Figure 2 This is a relapse-free survival curve analysis diagram of patients with cord tumors grouped according to the level of Notch1 intracellular O-GlcNAc glycosylation in an embodiment of the present invention;

[0010] Figure 3 A is a comparison diagram of the chordoma cell sac formation ability in an embodiment of the present invention;

[0011] Figure 3 B is a comparison diagram of CD15 expression in chordoma cell lines detected by flow cytometry in an embodiment of the present invention.

[0012] Figure 3 C, 3D, and 3E are comparative diagrams of promoter analysis of Notch1 intracellular segment binding in chromosome immunoprecipitation sequencing in embodiments of the present invention.

[0013] Figure 4 This is a flowchart of the animal experiment implementation and a comparison chart of subcutaneous tumor quality in the embodiments of the present invention. Detailed Implementation

[0014] The following is in conjunction with the appendix Figure 1-4 The present invention will be described in further detail below.

[0015] Example: The following experimental materials were used in the experiment: human chordoma specimens, human chordoma cell lines MUG-chor1 and UM-chor1, BALB / c nude mice, carbon dioxide incubator (MCO-18AC), clean bench (JL-CJ-HW1B), Bio-Rad vertical electrophoresis apparatus, Leica automated tissue dehydrator, Leica modular tissue embedding system, Leica fully automated rotary microtome for research, Leica slide spreader / baking machine, and laser confocal microscopy (Leica DMI8).

[0016] Primary and recurrent tumor specimens were collected from 10 pairs of chordoma patients. The Duolink PLA staining kit (Sigma), Notch1 intracellular domain antibody, and O-GlcNAc antibody were used to detect Notch1 intracellular domain-specific O-GlcNAc glycosylation modification. The results are as follows: Figure 1 As shown in the figure, in the same patient-paired tissue, the level of Notch1 intracellular O-GlcNAc glycosylation was significantly higher in recurrent tumors than in the corresponding primary tumors.

[0017] Clinical specimens, clinical information, and follow-up data from 87 patients with chordoma were collected. Notch1 intracellular segment-specific O-GlcNAc glycosylation modification was detected in tumor specimens using the Duolink PLA staining kit, Notch1 intracellular segment antibody, and O-GlcNAc antibody. Patients were divided into high and low groups based on the Notch1 intracellular segment O-GlcNAc glycosylation level. Kaplan-Meier analysis was used to analyze recurrence-free survival in chordoma patients. The results were presented as a linear graph. Figure 2 As shown, by Figure 2 It is known that chordoma patients with high levels of Notch1 intracellular O-GlcNAc glycosylation are more prone to early recurrence.

[0018] We used balloon formation and flow cytometry to observe chordoma stemness in chordoma cells. We found that promoting O-GlcNAc glycosylation modification with PUGNAC and GlcNAc significantly enhanced tumor stemness, while inhibiting O-GlcNAc glycosylation with OSMI-1 and knocking down Notch1 could limit and inhibit tumor stemness reversal (e.g., ...). Figure 3 (As shown in Figure AB). After treating chordoma cells with PUGNAC and GlcNAc, chromosome immunoprecipitation and sequencing techniques revealed that Notch1 cells modified with O-GlcNAc glycosylation exhibited stronger transcriptional activity and tended to bind to the promoters of tumor stemness markers to initiate the transcription of downstream genes (such as...). Figure 3 (as shown in CE).

[0019] In nude mice, 10 mg was injected subcutaneously. 7 Chordoma cells MUG-chor1 were divided into four groups based on different treatments. Results were observed after 12 weeks as follows: Figure 4 As shown, PUGNAC and GlcNAc significantly promoted the growth of subcutaneous tumors, while the use of OSMI-1 to inhibit O-GlcNAc glycosylation and the Notch inhibitor DAPT significantly inhibited tumor growth.

[0020] In conclusion, it can be observed that Notch1 intracellular O-GlcNAc glycosylation is significantly upregulated in recurrent chordomas, and chordoma patients with high levels of Notch1 intracellular O-GlcNAc glycosylation are more prone to early recurrence compared to those with low levels. Furthermore, inhibiting Notch1 and O-GlcNAc glycosylation inhibits tumor growth. Therefore, Notch1 intracellular O-GlcNAc glycosylation can serve as a novel biomarker for predicting chordoma recurrence, and targeting Notch1 intracellular O-GlcNAc glycosylation is a novel therapeutic strategy for recurrent chordoma.

[0021] This specific embodiment is merely an explanation of the present invention and is not intended to limit the invention. After reading this specification, those skilled in the art can make modifications to this embodiment without contributing any inventive step, but such modifications are protected by patent law as long as they are within the scope of the claims of the present invention.

Claims

1. The application of reagents for detecting and warning of chordoma recurrence in the preparation of products for early warning of chordoma recurrence, characterized in that, The biomarker is the intracellular O-GlcNAc glycosylation modification of Notch1.

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